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Sample records for anti-her2 igy antibody-functionalized

  1. Anti-HER2 IgY antibody-functionalized single-walled carbon nanotubes for detection and selective destruction of breast cancer cells

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    Mitra Somenath

    2009-10-01

    Full Text Available Abstract Background Nanocarrier-based antibody targeting is a promising modality in therapeutic and diagnostic oncology. Single-walled carbon nanotubes (SWNTs exhibit two unique optical properties that can be exploited for these applications, strong Raman signal for cancer cell detection and near-infrared (NIR absorbance for selective photothermal ablation of tumors. In the present study, we constructed a HER2 IgY-SWNT complex and demonstrated its dual functionality for both detection and selective destruction of cancer cells in an in vitro model consisting of HER2-expressing SK-BR-3 cells and HER2-negative MCF-7 cells. Methods The complex was constructed by covalently conjugating carboxylated SWNTs with anti-HER2 chicken IgY antibody, which is more specific and sensitive than mammalian IgGs. Raman signals were recorded on Raman spectrometers with a laser excitation at 785 nm. NIR irradiation was performed using a diode laser system, and cells with or without nanotube treatment were irradiated by 808 nm laser at 5 W/cm2 for 2 min. Cell viability was examined by the calcein AM/ethidium homodimer-1 (EthD-1 staining. Results Using a Raman optical microscope, we found the Raman signal collected at single-cell level from the complex-treated SK-BR-3 cells was significantly greater than that from various control cells. NIR irradiation selectively destroyed the complex-targeted breast cancer cells without harming receptor-free cells. The cell death was effectuated without the need of internalization of SWNTs by the cancer cells, a finding that has not been reported previously. Conclusion We have demonstrated that the HER2 IgY-SWNT complex specifically targeted HER2-expressing SK-BR-3 cells but not receptor-negative MCF-7 cells. The complex can be potentially used for both detection and selective photothermal ablation of receptor-positive breast cancer cells without the need of internalization by the cells. Thus, the unique intrinsic properties of SWNTs

  2. Anti-HER2 IgY antibody-functionalized single-walled carbon nanotubes for detection and selective destruction of breast cancer cells

    International Nuclear Information System (INIS)

    Xiao, Yan; Savla, Ronak; Wagner, Paul D; Srivastava, Sudhir; He, Huixin; Gao, Xiugong; Taratula, Oleh; Treado, Stephen; Urbas, Aaron; Holbrook, R David; Cavicchi, Richard E; Avedisian, C Thomas; Mitra, Somenath

    2009-01-01

    Nanocarrier-based antibody targeting is a promising modality in therapeutic and diagnostic oncology. Single-walled carbon nanotubes (SWNTs) exhibit two unique optical properties that can be exploited for these applications, strong Raman signal for cancer cell detection and near-infrared (NIR) absorbance for selective photothermal ablation of tumors. In the present study, we constructed a HER2 IgY-SWNT complex and demonstrated its dual functionality for both detection and selective destruction of cancer cells in an in vitro model consisting of HER2-expressing SK-BR-3 cells and HER2-negative MCF-7 cells. The complex was constructed by covalently conjugating carboxylated SWNTs with anti-HER2 chicken IgY antibody, which is more specific and sensitive than mammalian IgGs. Raman signals were recorded on Raman spectrometers with a laser excitation at 785 nm. NIR irradiation was performed using a diode laser system, and cells with or without nanotube treatment were irradiated by 808 nm laser at 5 W/cm 2 for 2 min. Cell viability was examined by the calcein AM/ethidium homodimer-1 (EthD-1) staining. Using a Raman optical microscope, we found the Raman signal collected at single-cell level from the complex-treated SK-BR-3 cells was significantly greater than that from various control cells. NIR irradiation selectively destroyed the complex-targeted breast cancer cells without harming receptor-free cells. The cell death was effectuated without the need of internalization of SWNTs by the cancer cells, a finding that has not been reported previously. We have demonstrated that the HER2 IgY-SWNT complex specifically targeted HER2-expressing SK-BR-3 cells but not receptor-negative MCF-7 cells. The complex can be potentially used for both detection and selective photothermal ablation of receptor-positive breast cancer cells without the need of internalization by the cells. Thus, the unique intrinsic properties of SWNTs combined with high specificity and sensitivity of IgY

  3. Developing Anti-HER2 Vaccines: Breast Cancer Experience.

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    Al-Awadhi, Aydah; Murray, James Lee; Ibrahim, Nuhad K

    2018-04-25

    Breast cancer accounts for more than one million new cases annually and is the leading cause of death in women globally. HER2 overexpression induces cellular and humoral immune responses against the HER2 protein and is associated with higher tumour proliferation rates. Trastuzumab-based therapies are effectively and widely used as standard of care in HER2-amplified/overexpressed breast cancer patients; one cited mechanism of action is the induction of passive immunity and antibody-dependent cellular cytotoxicity against malignant breast cancer cells. These findings drove the efforts to generate antigen-specific immunotherapy to trigger the patient's immune system to target HER2-overexpressing tumour cells, which led to the development of various vaccines against the HER2 antigen. This manuscript discusses the various anti-HER2 vaccine formulations and strategies and their potential role in the metastatic and adjuvant settings. This article is protected by copyright. All rights reserved. © 2018 UICC.

  4. Targeting breast carcinoma with radioiodinated anti-HER2 Nanobody

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    Pruszynski, Marek; Koumarianou, Eftychia; Vaidyanathan, Ganesan; Revets, Hilde; Devoogdt, Nick; Lahoutte, Tony; Zalutsky, Michael R.

    2013-01-01

    Introduction: With a molecular weight an order of magnitude lower than antibodies but possessing comparable affinities, Nanobodies (Nbs) are attractive as targeting agents for cancer diagnosis and therapy. An anti-HER2 Nb could be utilized to determine HER2 status in breast cancer patients prior to trastuzumab treatment. This provided motivation for the generation of HER2-specific 5F7GGC Nb, its radioiodination and evaluation for targeting HER2 expressing tumors. Methods: 5F7GGC Nb was radioiodinated with 125 I using Iodogen and with 131 I using the residualizing agent N ε -(3-[ 131 I]iodobenzoyl)-Lys 5 -N α -maleimido-Gly 1 -GEEEK ([ 131 I]IB-Mal-D-GEEEK) used previously successfully with intact antibodies. Paired-label internalization assays using BT474M1 cells and tissue distribution experiments in athymic mice bearing BT474M1 xenografts were performed to compare the two labeled Nb preparations. Results: The radiochemical yields for Iodogen and [ 131 I]IB-Mal-D-GEEEK labeling were 83.6 ± 5.0% (n = 10) and 59.6 ± 9.4% (n = 15), respectively. The immunoreactivity of labeled proteins was preserved as confirmed by in vitro and in vivo binding to tumor cells. Biodistribution studies showed that Nb radiolabeled using [ 131 I]IB-Mal-D-GEEEK, compared with the directly labeled Nb, had a higher tumor uptake (4.65 ± 0.61% ID/g vs. 2.92 ± 0.24% ID/g at 8 h), faster blood clearance, lower accumulation in non-target organs except kidneys, and as a result, higher concomitant tumor-to-blood and tumor-to-tissue ratios. Conclusions: Taken together, these results demonstrate that 5F7GGC anti-HER2 Nb labeled with residualizing [ 131 I]IB-Mal-D-GEEEK had better tumor targeting properties compared to the directly labeled Nb suggesting the potential utility of this Nb conjugate for SPECT ( 129 I) and PET imaging ( 124 I) of patients with HER2-expressing tumors.

  5. Novel anti-HER2 monoclonal antibodies: synergy and antagonism with tumor necrosis factor-α

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    Ceran Ceyhan

    2012-10-01

    Full Text Available Abstract Background One-third of breast cancers display amplifications of the ERBB2 gene encoding the HER2 kinase receptor. Trastuzumab, a humanized antibody directed against an epitope on subdomain IV of the extracellular domain of HER2 is used for therapy of HER2-overexpressing mammary tumors. However, many tumors are either natively resistant or acquire resistance against Trastuzumab. Antibodies directed to different epitopes on the extracellular domain of HER2 are promising candidates for replacement or combinatorial therapy. For example, Pertuzumab that binds to subdomain II of HER2 extracellular domain and inhibits receptor dimerization is under clinical trial. Alternative antibodies directed to novel HER2 epitopes may serve as additional tools for breast cancer therapy. Our aim was to generate novel anti-HER2 monoclonal antibodies inhibiting the growth of breast cancer cells, either alone or in combination with tumor necrosis factor-α (TNF-α. Methods Mice were immunized against SK-BR-3 cells and recombinant HER2 extracellular domain protein to produce monoclonal antibodies. Anti-HER2 antibodies were characterized with breast cancer cell lines using immunofluorescence, flow cytometry, immunoprecipitation, western blot techniques. Antibody epitopes were localized using plasmids encoding recombinant HER2 protein variants. Antibodies, either alone or in combination with TNF-α, were tested for their effects on breast cancer cell proliferation. Results We produced five new anti-HER2 monoclonal antibodies, all directed against conformational epitope or epitopes restricted to the native form of the extracellular domain. When tested alone, some antibodies inhibited modestly but significantly the growth of SK-BR-3, BT-474 and MDA-MB-361 cells displaying ERBB2 amplification. They had no detectable effect on MCF-7 and T47D cells lacking ERBB2 amplification. When tested in combination with TNF-α, antibodies acted synergistically on SK-BR-3 cells

  6. Development of an Anti-HER2 Monoclonal Antibody H2Mab-139 Against Colon Cancer.

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    Kaneko, Mika K; Yamada, Shinji; Itai, Shunsuke; Kato, Yukinari

    2018-02-01

    Human epidermal growth factor receptor 2 (HER2) expression has been reported in several cancers, such as breast, gastric, lung, pancreatic, and colorectal cancers. HER2 is overexpressed in those cancers and is associated with poor clinical outcomes. Trastuzumab, a humanized anti-HER2 antibody, provides significant survival benefits for patients with HER2-overexpressing breast cancers and gastric cancers. In this study, we developed a novel anti-HER2 monoclonal antibody (mAb), H 2 Mab-139 (IgG 1 , kappa) and investigated it against colon cancers using flow cytometry, western blot, and immunohistochemical analyses. Flow cytometry analysis revealed that H 2 Mab-139 reacted with colon cancer cell lines, such as Caco-2, HCT-116, HCT-15, HT-29, LS 174T, COLO 201, COLO 205, HCT-8, SW1116, and DLD-1. Although H 2 Mab-139 strongly reacted with LN229/HER2 cells on the western blot, we did not observe a specific signal for HER2 in colon cancer cell lines. Immunohistochemical analyses revealed sensitive and specific reactions of H 2 Mab-139 against colon cancers, indicating that H 2 Mab-139 is useful in detecting HER2 overexpression in colon cancers using flow cytometry and immunohistochemical analyses.

  7. A Single Domain–Based Anti-Her2 Antibody Has Potent Antitumor Activities

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    Xiaoqiong Wu

    2018-04-01

    Full Text Available Human epidermal growth factor receptor 2 (HER2 is overexpressed in approximately 20% to 30% of breast cancers and various other types of cancers, which plays a vital role in the cancer progression. Monoclonal antibodies targeting Her2 are now used in the clinic to treat Her2 overexpression cancer patients. However, relapse or resistance is frequent with the current therapies. To generate a new treatment avenue against Her2, we immunized and selected a specific anti-Her2 single domain antibody C3 for further studies. The C3-Fc antibody drove antibody-dependent cell-mediated cytotoxicity against Her2-positive tumor cells in vitro and resulted in potent antitumor growth in vivo. These data suggest that the C3-Fc antibody may provide an alternative avenue for Her2-positive cancer therapy.

  8. Optimization of an anti-HER2 nanobody expression using the Taguchi method.

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    Farasat, Alireza; Rahbarizadeh, Fatemeh; Ahmadvand, Davoud; Yazdian, Fatemeh

    2017-09-14

    Despite being widely used in immunotherapy of cancer, whole antibodies are limited by several disadvantages. This has led to the advent of novel biomolecules such as nanobodies. Taguchi method is a statistical experimental design to study the effect of multiple variables in biological processes. In an effort to overexpress a recombinant anti-human epidermal growth factor receptor type 2 (HER2) nanobody, we performed a detailed study to find optimal condition of temperature, induction, culture media, vector, and host strain, using Taguchi methodology. A total of 16 various experiments were designed. Total protein of the formulated cultures were assessed by Bradford test and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, followed by size exclusion high performance liquid chromatography to quantify the relative concentration of the nanobody in different expression settings. Western blotting was performed to confirm the expression of the anti-HER2 nanobody. When, individually, optimum parameters determined by Taguchi were applied, including SHuffle strain cultured in LB medium, induced with 0.4 mM isopropyl-β-D-thio-galactoside for 18 h at 24°C, production yield further increased by about 9% (25.4 mg/L), compared to the highest expression setting. Flow cytometry and enzyme-linked immunosorbent assay result indicated improved protein binding in optimized conditions. Overall, our findings provide a basis for further investigations on economical production of recombinant nanobodies to improve production yield and activity.

  9. Current therapeutic strategies of anti-HER2 treatment in advanced breast cancer patients

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    Joanna Huszno

    2016-03-01

    Full Text Available The HER2/neu ( ERBB2 oncogene is amplified and/or overexpressed in approximately 20% of breast cancers, and is a strong prognostic factor for relapse and poor overall survival, particularly in node-positive patients. It is also an important predictor for response to trastuzumab, which has established efficacy against breast cancer with overexpression or amplification of the HER2 oncogene. Treatment with the anti-HER2 humanized monoclonal antibody – trastuzumab significantly improves progression-free and overall survival among patients with HER2-positive breast cancer. However, in most patients with HER2-positive metastatic breast cancer, the disease progresses occurred, what cause the need for new targeted therapies for advanced disease. In clinical trials, there are tested new drugs to improve the results of treatment for this group of patients. This paper presents new drugs introduced into clinical practice for treatment of advanced breast cancer, whose molecular target are receptors of the HER2 family. In addition, new therapeutic strategies and drugs that are currently in clinical researches are discussed.

  10. Survival benefit of anti-HER2 therapy after whole-brain radiotherapy in HER2-positive breast cancer patients with brain metastasis.

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    Zhang, Qian; Chen, Jian; Yu, Xiaoli; Cai, Gang; Yang, Zhaozhi; Cao, Lu; Hu, Chaosu; Guo, Xiaomao; Sun, Jing; Chen, Jiayi

    2016-09-01

    We aimed to assess the survival benefit of epidermal growth factor receptor 2 (HER2)-positive breast cancer patients with brain metastasis (BM) after whole-brain radiotherapy (WBRT) in combination with systemic treatments, especially anti-HER2 therapy. This retrospective study analyzed the overall survival (OS) of 60 HER2-positive breast cancer patients with BM after WBRT in combination with systemic treatments. Among them, 42 patients received chemotherapy while 18 patients did not receive after WBRT. With regard to anti-HER2 therapy, after WBRT, 17 patients received anti-HER2 treatment without prior adjuvant trastuzumab-based therapy, 7 patients received anti-HER2 treatment with prior adjuvant trastuzumab-based therapy, and 36 patients did not receive further anti-HER2 treatment. All patients were followed up regularly until January 23, 2013. The median OS of patients with BM was 12 months. Patients who received anti-HER2 therapy and chemotherapy after WBRT had significantly better survival compared with patients who did not receive further treatment. Patients who received anti-HER2 treatment after WBRT but did not receive adjuvant trastuzumab-based therapy for early breast cancer had better OS, followed by patients who received anti-HER2 agent both in adjuvant treatment and after WBRT and patients who did not receive anti-HER2 treatment. Multivariate analysis showed that Karnofsky Performance Status, control of extracranial metastases, chemotherapy after WBRT, and anti-HER2 therapy combined with WBRT were all independent predictors for OS. Both chemotherapy and anti-HER2 therapy after WBRT could improve OS. Moreover, patients without prior exposure to adjuvant anti-HER2 treatment may have survival benefit superior to those of patients with prior exposure.

  11. Is the skin a sanctuary for breast cancer cells during treatment with anti-HER2 antibodies?

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    Graziano, Vincenzo; Scognamiglio, Maria Teresa; Zilli, Marinella; Giampietro, Jamara; Vici, Patrizia; Natoli, Clara; Grassadonia, Antonino

    2015-01-01

    The occurrence of skin metastases is a common event in patients affected by advanced breast cancer, usually associated with systemic disease progression. Here we describe 2 cases of diffuse cutaneous metastases from HER2-overexpressing breast cancer occurring despite a dramatic response in liver and bone, respectively, during treatment with anti-HER2 antibodies Trastuzumab and Pertuzumab. We discuss the reasons for this discrepancy and suggest a possible implication of impaired immune response in the skin. Future research should provide strategies to overcome the induction of immune privilege in the skin in order to avoid discontinuation of effective treatments.

  12. H2Mab-77 is a Sensitive and Specific Anti-HER2 Monoclonal Antibody Against Breast Cancer.

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    Itai, Shunsuke; Fujii, Yuki; Kaneko, Mika K; Yamada, Shinji; Nakamura, Takuro; Yanaka, Miyuki; Saidoh, Noriko; Chang, Yao-Wen; Handa, Saori; Takahashi, Maki; Suzuki, Hiroyoshi; Harada, Hiroyuki; Kato, Yukinari

    2017-08-01

    Human epidermal growth factor receptor 2 (HER2) plays a critical role in the progression of breast cancers, and HER2 overexpression is associated with poor clinical outcomes. Trastuzumab is an anti-HER2 humanized antibody that leads to significant survival benefits in patients with HER2-positive metastatic breast cancers. In this study, we developed novel anti-HER2 monoclonal antibodies (mAbs) and characterized their efficacy in flow cytometry, Western blot, and immunohistochemical analyses. Initially, we expressed the full length or ectodomain of HER2 in LN229 glioblastoma cells and then immunized mice with ectodomain of HER2 or LN229/HER2, and performed the first screening by enzyme-linked immunosorbent assays using ectodomain of HER2. Subsequently, we selected mAbs according to their efficacy in flow cytometry (second screening), Western blot (third screening), and immunohistochemical analyses (fourth screening). Among 100 mAb clones, only three mAbs reacted with HER2 in Western blot, and clone H 2 Mab-77 (IgG 1 , kappa) was selected. Finally, immunohistochemical analyses with H 2 Mab-77 showed sensitive and specific reactions against breast cancer cells, warranting the use of H 2 Mab-77 to detect HER2 in pathological analyses of breast cancers.

  13. Improved tumor targeting of anti-HER2 nanobody through N-succinimidyl 4-guanidinomethyl-3-iodobenzoate radiolabeling.

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    Pruszynski, Marek; Koumarianou, Eftychia; Vaidyanathan, Ganesan; Revets, Hilde; Devoogdt, Nick; Lahoutte, Tony; Lyerly, H Kim; Zalutsky, Michael R

    2014-04-01

    Nanobodies are approximately 15-kDa proteins based on the smallest functional fragments of naturally occurring heavy chain-only antibodies and represent an attractive platform for the development of molecularly targeted agents for cancer diagnosis and therapy. Because the human epidermal growth factor receptor type 2 (HER2) is overexpressed in breast and ovarian carcinoma, as well as in other malignancies, HER2-specific Nanobodies may be valuable radiodiagnostics and therapeutics for these diseases. The aim of the present study was to evaluate the tumor-targeting potential of anti-HER2 5F7GGC Nanobody after radioiodination with the residualizing agent N-succinimidyl 4-guanidinomethyl 3-(125/131)I-iodobenzoate (*I-SGMIB). The 5F7GGC Nanobody was radiolabeled using *I-SGMIB and, for comparison, with N(ε)-(3-*I-iodobenzoyl)-Lys(5)-N(α)-maleimido-Gly(1)-GEEEK (*I-IB-Mal-d-GEEEK), another residualizing agent, and by direct radioiodination using IODO-GEN ((125)I-Nanobody). The 3 labeled Nanobodies were evaluated in affinity measurements, and paired-label internalization assays were performed on HER2-expressing BT474M1 breast carcinoma cells and in paired-label tissue distribution measurements in mice bearing subcutaneous BT474M1 xenografts. *I-SGMIB-Nanobody was produced in 50.4% ± 3.6% radiochemical yield and exhibited a dissociation constant of 1.5 ± 0.5 nM. Internalization assays demonstrated that intracellular retention of radioactivity was up to 1.5-fold higher for *I-SGMIB-Nanobody than for coincubated (125)I-Nanobody or *I-IB-Mal-d-GEEEK-Nanobody. Peak tumor uptake for *I-SGMIB-Nanobody was 24.50% ± 9.89% injected dose/g at 2 h, 2- to 4-fold higher than observed with other labeling methods, and was reduced by 90% with trastuzumab blocking, confirming the HER2 specificity of localization. Moreover, normal-organ clearance was fastest for *I-SGMIB-Nanobody, such that tumor-to-normal-organ ratios greater than 50:1 were reached by 24 h in all tissues except lungs

  14. Arg9 facilitates the translocation and downstream signal inhibition of an anti-HER2 single chain antibody

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    Hu Yi

    2012-07-01

    Full Text Available Abstract Background HER2 plays a critical role in the pathogenesis of many cancers and is linked to poor prognosis or cancer metastases. Monoclonal antibodies, such as Herceptin against HER2-overexpressing cancers, have showed satisfactory clinical therapeutic effect. However, they have difficulty to surmount obstacles to enter cells or blood–brain barrier. Results In this study, a cell-penetrating peptide Arg9 was linked to the C-terminus of anti-HER2 single chain antibody (MIL5scFv. Flow cytometry, confocal microscopy and electron microscopy analysis all revealed that Arg9 peptide facilitated the penetration of MIL5scFv into HER2-negative cell line NIH3T3 and orientate in mitochondria. More interestingly, Western blot assay showed the potential enhanced bioactivity of MIL5scFv-Arg9 in HER2+ cell line SKOV3, indicating that Arg9 could help large molecules (e.g. antibody to penetrate into cells and therefore enhance its anti-neoplastic function. Conclusions Our work represented an attractive by preliminary strategy to enhance the therapeutic effect of existing antibodies by entering cells easier, or more desirable, surmounting the physical barriers, especially in hard-to-reach cancers such as brain metastases cases.

  15. Comparative Evaluation of Anti-HER2 Affibody Molecules Labeled with 64Cu Using NOTA and NODAGA

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    Vladimir Tolmachev

    2017-01-01

    Full Text Available Imaging using affibody molecules enables discrimination between breast cancer metastases with high and low expression of HER2, making appropriate therapy selection possible. This study aimed to evaluate if the longer half-life of 64Cu (T1/2 = 12.7 h would make 64Cu a superior nuclide compared to 68Ga for PET imaging of HER2 expression using affibody molecules. The synthetic ZHER2:S1 affibody molecule was conjugated with the chelators NOTA or NODAGA and labeled with 64Cu. The tumor-targeting properties of 64Cu-NOTA-ZHER2:S1 and 64Cu-NODAGA-ZHER2:S1 were evaluated and compared with the targeting properties of 68Ga-NODAGA-ZHER2:S1 in mice. Both 64Cu-NOTA-ZHER2:S1 and 64Cu-NODAGA-ZHER2:S1 demonstrated specific targeting of HER2-expressing xenografts. At 2 h after injection of 64Cu-NOTA-ZHER2:S1, 64Cu-NODAGA-ZHER2:S1, and 68Ga-NODAGA-ZHER2:S1, tumor uptakes did not differ significantly. Renal uptake of 64Cu-labeled conjugates was dramatically reduced at 6 and 24 h after injection. Notably, radioactivity uptake concomitantly increased in blood, lung, liver, spleen, and intestines, which resulted in decreased tumor-to-organ ratios compared to 2 h postinjection. Organ uptake was lower for 64Cu-NODAGA-ZHER2:S1. The most probable explanation for this biodistribution pattern was the release and redistribution of renal radiometabolites. In conclusion, monoamide derivatives of NOTA and NODAGA may be suboptimal chelators for radiocopper labeling of anti-HER2 affibody molecules and, possibly, other scaffold proteins with high renal uptake.

  16. Evaluation of an Anti-HER2 Nanobody Labeled with 225Ac for Targeted α-Particle Therapy of Cancer.

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    Pruszynski, Marek; D'Huyvetter, Matthias; Bruchertseifer, Frank; Morgenstern, Alfred; Lahoutte, Tony

    2018-04-02

    Human epidermal growth factor receptor type 2 (HER2) is overexpressed in numerous carcinomas. Nanobodies (Nbs) are the smallest antibody-derived fragments with beneficial characteristics for molecular imaging and radionuclide therapy. Therefore, HER2-targeting nanobodies could offer a valuable platform for radioimmunotherapy, especially when labeled with α-particle emitters, which provide highly lethal and localized radiation to targeted cells with minimal exposure to surrounding healthy tissues. In this study, the anti-HER2 2Rs15d-nanobody was conjugated with 2-(4-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid ( p-SCN-Bn-DOTA) and radiolabeled with an α-emitter 225 Ac with a high yield (>90%) and a radiochemical purity above 95%. The 225 Ac-DOTA-Nb binding affinity was 4.12 ± 0.47 nM with an immunoreactive fraction above 80%. Binding to low HER2-expressing MDA-MB-231 cells was negligible, whereas HER2-overexpressing SKOV-3 cells could be blocked with an excess of unlabeled nanobody, confirming the specificity of binding. Noncompeting binding to HER2 was observed in the presence of an excess of trastuzumab. The cell-associated fraction of 225 Ac-DOTA-Nb was 34.72 ± 16.66% over 24 h. In vitro, the radioconjugate was toxic in an HER2-mediated and dose-dependent manner, resulting in IC 50 values of 10.2 and 322.1 kBq/mL for 225 Ac-DOTA-Nb and the 225 Ac-DOTA control, respectively, on SKOV-3 cells, and 282.2 kBq/mL for 225 Ac-DOTA-Nb on MDA-MB-231 cells. Ex vivo biodistribution studies, performed in mice bearing subcutaneous HER2-overexpressing and low HER2-expressing tumors, showed a fast uptake in SKOV-3 tumors compared to MDA-MB-231 (4.01 ± 1.58% ID/g vs 0.49 ± 0.20% ID/g after 2 h), resulting also in high tumor-to-normal tissue ratios. In addition, coinjection of 225 Ac-DOTA-Nb with Gelofusine reduced kidney retention by 70%. This study shows that 225 Ac-DOTA-Nb is a promising new radioconjugate for targeted α-particle therapy

  17. Restoring Lost Anti-HER-2 Th1 Immunity in Breast Cancer: A Crucial Role for Th1 Cytokines in Therapy and Prevention.

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    Nocera, Nadia F; Lee, M Catherine; De La Cruz, Lucy M; Rosemblit, Cinthia; Czerniecki, Brian J

    2016-01-01

    The ErbB/B2 (HER-2/neu) oncogene family plays a critical role in the development and metastatic spread of several tumor types including breast, ovarian and gastric cancer. In breast cancer, HER-2/neu is expressed in early disease development in a large percentage of DCIS lesions and its expression is associated with an increased risk of invasion and recurrence. Targeting HER-2 with antibodies such as trastuzumab or pertuzumab has improved survival, but patients with more extensive disease may develop resistance to therapy. Interestingly, response to HER-2 targeted therapies correlates with presence of immune response genes in the breast. Th1 cell production of the cytokines interferon gamma (IFNγ) and TNFα can enhance MHC class I expression, PD-L1 expression, augment apoptosis and tumor senescence, and enhances growth inhibition of many anti-breast cancer agents, including anti-estrogens and HER-2 targeted therapies. Recently, we have identified that a loss of anti-HER-2 CD4 Th1 in peripheral blood occurs during breast tumorigenesis and is dramatically diminished, even in Stage I breast cancers. The loss of anti-HER-2 Th1 response is specific and not readily reversed by standard therapies. In fact, this loss of anti-HER-2 Th1 response in peripheral blood correlates with lack of complete response to neoadjuvant therapy and diminished disease-free survival. This defect can be restored with HER-2 vaccinations in both DCIS and IBC. Correcting the anti-HER-2 Th1 response may have significant impact in improving response to HER-2 targeted therapies. Development of immune monitoring systems for anti-HER-2 Th1 to identify patients at risk for recurrence could be critical to improving outcomes, since the anti-HER-2 Th1 response can be restored by vaccination. Correction of the cellular immune response against HER-2 may prevent recurrence in high-risk patients with DCIS and IBC at risk of developing new or recurrent breast cancer.

  18. Upregulation of ER signaling as an adaptive mechanism of cell survival in HER2-positive breast tumors treated with anti-HER2 therapy

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    Giuliano, Mario; Hu, Huizhong; Wang, Yen-Chao; Fu, Xiaoyong; Nardone, Agostina; Herrera, Sabrina; Mao, Sufeng; Contreras, Alejandro; Gutierrez, Carolina; Wang, Tao; Hilsenbeck, Susan G.; De Angelis, Carmine; Wang, Nicholas J.; Heiser, Laura M.; Gray, Joe W.; Lopez-Tarruella, Sara; Pavlick, Anne C.; Trivedi, Meghana V.; Chamness, Gary C.; Chang, Jenny C.; Osborne, C. Kent; Rimawi, Mothaffar F.; Schiff, Rachel

    2015-01-01

    Purpose To investigate the direct effect and therapeutic consequences of epidermal growth factor receptor 2 (HER2)-targeting therapy on expression of estrogen receptor (ER) and Bcl2 in preclinical models and clinical tumor samples. Experimental design Archived xenograft tumors from two preclinical models (UACC812 and MCF7/HER2-18) treated with ER and HER2-targeting therapies, and also HER2+ clinical breast cancer specimens collected in a lapatinib neoadjuvant trial (baseline and week 2 post treatment), were used. Expression levels of ER and Bcl2 were evaluated by immunohistochemistry and western blot. The effects of Bcl2 and ER inhibition, by ABT-737 and fulvestrant respectively, were tested in parental versus lapatinib-resistant UACC812 cells in vitro. Results Expression of ER and Bcl2 was significantly increased in xenograft tumors with acquired resistance to anti-HER2 therapy, compared with untreated tumors, in both preclinical models (UACC812: ER p=0.0014; Bcl2 p<0.001. MCF7/HER2-18: ER p=0.0007; Bcl2 p=0.0306). In the neoadjuvant clinical study, lapatinib treatment for two weeks was associated with parallel upregulation of ER and Bcl2 (Spearman’s coefficient: 0.70; p=0.0002). Importantly, 18% of tumors originally ER-negative (ER−) converted to ER+ upon anti-HER2 therapy. In ER−/HER2+ MCF7/HER2-18 xenografts, ER re-expression was primarily observed in tumors responding to potent combination of anti-HER2 drugs. Estrogen deprivation added to this anti-HER2 regimen significantly delayed tumor progression (p=0.018). In the UACC812 cells, fulvestrant, but not ABT-737, was able to completely inhibit anti-HER2-resistant growth (p<0.0001). Conclusion HER2 inhibition can enhance or restore ER expression with parallel Bcl2 upregulation, representing an ER-dependent survival mechanism potentially leading to anti-HER2 resistance. PMID:26015514

  19. Assessment of the systemic distribution of a bioconjugated anti-Her2 magnetic nanoparticle in a breast cancer model by means of magnetic resonance imaging

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    Huerta-Núñez, L. F. E., E-mail: lidi-huerta@hotmail.com [Universidad del Ejercito y FAM/EMGS-Laboratorio Multidisciplinario de Investigación (Mexico); Villanueva-Lopez, G. Cleva, E-mail: villanuevacleva3@gmail.com [Instituto Politécnico Nacional-Escuela Superior de Medicina-Sección Investigación y Posgrado (Mexico); Morales-Guadarrama, A., E-mail: amorales@ci3m.mx [Centro Nacional de Investigacion en Imagenologia e Instrumentacion Medica-Universidad Autónoma (Mexico); Soto, S., E-mail: cuadrosdobles@hotmail.com; López, J., E-mail: jaimelocr@hotmail.com; Silva, J. G., E-mail: gabrielsilva173@gmail.com [Universidad del Ejercito y FAM/EMGS-Laboratorio Multidisciplinario de Investigación (Mexico); Perez-Vielma, N., E-mail: nadiampv@gmail.com [Instituto Politécnico Nacional - Centro Interdisciplinario de Ciencias de la Salud Unidad Santo Tomás (CICS-UST) (Mexico); Sacristán, E., E-mail: esacristan@ci3m.mx [Centro Nacional de Investigacion en Imagenologia e Instrumentacion Medica-Universidad Autónoma (Mexico); Gudiño-Zayas, Marco E., E-mail: gudino@unam.mx [UNAM, Departamento de Medicina Experimental, Facultad de Medicina (Mexico); González, C. A., E-mail: cgonzalezd@ipn.mx [Universidad del Ejercito y FAM/EMGS-Laboratorio Multidisciplinario de Investigación (Mexico)

    2016-09-15

    The aim of this study was to determine the systemic distribution of magnetic nanoparticles of 100 nm diameter (MNPs) coupled to a specific monoclonal antibody anti-Her2 in an experimental breast cancer (BC) model. The study was performed in two groups of Sprague–Dawley rats: control (n = 6) and BC chemically induced (n = 3). Bioconjugated “anti-Her2-MNPs” were intravenously administered, and magnetic resonance imaging (MRI) monitored its systemic distribution at seven times after administration. Non-heme iron presence associated with the location of the bioconjugated anti-Her2-MNPs in splenic, hepatic, cardiac and tumor tissues was detected by Perl’s Prussian blue (PPB) stain. Optical density measurements were used to semiquantitatively determine the iron presence in tissues on the basis of a grayscale values integration of T1 and T2 MRI sequence images. The results indicated a delayed systemic distribution of MNPs in cancer compared to healthy conditions with a maximum concentration of MNPs in cancer tissue at 24 h post-infusion.

  20. Development and Characterization of a Humanized Anti-HER2 Antibody HuA21 with Potent Anti-Tumor Properties in Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Ruilin Li

    2016-04-01

    Full Text Available Human epidermal growth factor receptor 2 (HER2 is one of the most studied tumor-associated antigens for cancer immunotherapy. An engineered anti-HER-2 chimeric A21 antibody (chA21 is a chimeric antibody targeted to subdomain I of the HER2 extracellular domain. Here, we report the anti-tumor activity of the novel engineered monoclonal antibody humanized chA21 (HuA21 that targets HER2 on the basis of chA21, and we describe the underlying mechanisms. Our results reveal that HuA21 markedly inhibits the proliferation and migration of HER2-overexpressing breast cancer cells and causes enhanced antibody-dependent cell-mediated cytotoxicity potency against HER2-overexpressing tumor cells. In particular, HuA21, but not trastuzumab (Tra, markedly suppresses growth and enhances the internalization of the antibody in Tra-resistant BT-474 breast cancer cells. These characteristics are highly associated with the intrinsic ability of HuA21 to down-regulate HER2 activation and inhibit the extracellular signal-regulated kinase 1/2 (ERK1/2 and protein kinase B (Akt signaling pathways. Furthermore, the combination of HuA21 with Tra synergistically enhances the anti-tumor effects in vitro and in vivo and inhibits HER2 activation and the ERK1/2 and Akt signaling pathways. Altogether, our results suggest that HuA21 may represent a unique anti-HER2 antibody with potential as a therapeutic candidate alone or in combination with other anti-HER2 reagents in cancer therapy.

  1. Development of a Targeted anti-HER2 scFv Chimeric Peptide for Gene Delivery into HER2-Positive Breast Cancer Cells.

    Science.gov (United States)

    Cheraghi, Roya; Nazari, Mahboobeh; Alipour, Mohsen; Majidi, Asia; Hosseinkhani, Saman

    2016-12-30

    Chimeric polymers are known as suitable carriers for gene delivery. Certain properties are critical for a polymer to be used as a gene delivery vector. A new polymer was designed for the targeted delivery of genes into breast cancer cell lines, based on MPG peptide. It is composed of different functional domains, including HIV gp41, nuclear localization sequence of SV40 T-antigen, two C-terminus repeats of histone H1, and the scFv of anti-HER2 antibody. The results demonstrated that the vector can effectively condense plasmid DNA into nanoparticles with an average size of 250nm. Moreover, fusion of the scFv portion to the carrier brought about the specific recognition of HER2. Overall, the transfection efficiency of the vector demonstrated that it could deliver the desired gene into BT-474 HER2-positive breast cancer cells. Copyright © 2016 Elsevier B.V. All rights reserved.

  2. Overview of the trastuzumab (Herceptin) anti-HER2 monoclonal antibody clinical program in HER2-overexpressing metastatic breast cancer. Herceptin Multinational Investigator Study Group.

    Science.gov (United States)

    Shak, S

    1999-08-01

    The recombinant humanized anti-HER2 monoclonal antibody trastuzumab (Herceptin; Genentech, San Francisco, CA) was evaluated in human clinical trials for treatment of women with metastatic breast cancer who have tumors that overexpress HER2. The trastuzumab clinical program consisted of a series of phase I, phase II, and phase III clinical trials. Clinical experience with this novel biologic has been obtained in more than 1,000 women with HER2-overexpressing metastatic breast cancer. Two pivotal trials were performed to evaluate trastuzumab efficacy and safety: (1) trastuzumab in combination with chemotherapy as first-line therapy and (2) trastuzumab as a single agent in second- and third-line chemotherapy. Preliminary results of the pivotal clinical trials that have been presented at national meetings are summarized below. The data suggest that trastuzumab will be an important new treatment option for women with HER2-overexpressing metastatic breast cancer.

  3. Synthesis and Characterization of Anti-HER2 Antibody Conjugated CdSe/CdZnS Quantum Dots for Fluorescence Imaging of Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Takashi Jin

    2009-11-01

    Full Text Available The early detection of HER2 (human epidermal growth factor receptor 2 status in breast cancer patients is very important for the effective implementation of anti-HER2 antibody therapy. Recently, HER2 detections using antibody conjugated quantum dots (QDs have attracted much attention. QDs are a new class of fluorescent materials that have superior properties such as high brightness, high resistance to photo-bleaching, and multi-colored emission by a single-light source excitation. In this study, we synthesized three types of anti-HER2 antibody conjugated QDs (HER2Ab-QDs using different coupling agents (EDC/sulfo-NHS, iminothiolane/sulfo-SMCC, and sulfo-SMCC. As water-soluble QDs for the conjugation of antibody, we used glutathione coated CdSe/CdZnS QDs (GSH-QDs with fluorescence quantum yields of 0.23~0.39 in aqueous solution. Dispersibility, hydrodynamic size, and apparent molecular weights of the GSH-QDs and HER2Ab-QDs were characterized by using dynamic light scattering, fluorescence correlation spectroscopy, atomic force microscope, and size-exclusion HPLC. Fluorescence imaging of HER2 overexpressing cells (KPL-4 human breast cancer cell line was performed by using HER2Ab-QDs as fluorescent probes. We found that the HER2Ab-QD prepared by using SMCC coupling with partially reduced antibody is a most effective probe for the detection of HER2 expression in KPL-4 cells. We have also studied the size dependency of HER2Ab-QDs (with green, orange, and red emission on the fluorescence image of KPL-4 cells.

  4. Polymer–lipid hybrid anti-HER2 nanoparticles for targeted salinomycin delivery to HER2-positive breast cancer stem cells and cancer cells

    Directory of Open Access Journals (Sweden)

    Li J

    2017-09-01

    Full Text Available Jun Li,1,* Wenqing Xu,2,* Xiaoli Yuan,3,* Huaiwen Chen,3 Hao Song,1,4 Bingquan Wang,5 Jun Han5 1College of Pharmacy, Liaocheng University, Liaocheng, Shandong, 2Railway Police College, Zhengzhou, 3Department of Cadre Health Care, Nanjing General Hospital of Nanjing Military Command, Nanjing, Jiangsu, 4Centre for Stem Cell & Regenerative Medicine, Liaocheng People’s Hospital, 5Institute of Biopharmaceutical Research, Liaocheng University, Liaocheng, Shandong, China *These authors contributed equally to this work Purpose: Breast cancer stem cells (CSCs are responsible for the initiation, recurrence, and metastasis of breast cancer. Sufficient evidence has established that breast cancer cells can spontaneously turn into breast CSCs. Thus, it is essential to simultaneously target breast CSCs and cancer cells to maximize the efficacy of breast cancer therapy. HER2 has been found to be overexpressed in both breast CSCs and cancer cells. We developed salinomycin-loaded polymer–lipid hybrid anti-HER2 nanoparticles (Sali-NP-HER2 to target both HER2-positive breast CSCs and cancer cells.Methods: The antitumor activity of Sali-NP-HER2 constructed by conjugating anti-HER2 antibodies to polymer–lipid salinomycin nanoparticles was evaluated in vitro and in vivo.Results: Sali-NP-HER2 efficiently bound to HER2-positive breast CSCs and cancer cells, resulting in enhanced cytotoxic effects compared with non-targeted nanoparticles or salinomycin. In mice bearing breast cancer xenografts, administration of Sali-NP-HER2 exhibited superior efficacy in inhibiting tumor growth. Sali-NP-HER2 reduced the breast tumorsphere formation rate and the proportion of breast CSCs more effectively than non-targeted nanoparticles or salinomycin alone.Conclusion: Sali-NP-HER2 represents a promising approach in treating HER2-positive breast cancer by targeting both breast CSCs and cancer cells. Keywords: nanoparticles, breast cancer, cancer stem cells, salinomycin, HER2

  5. HER2-positive breast cancer, how far away from the cure?-on the current situation of anti-HER2 therapy in breast cancer treatment and survival of patients.

    Science.gov (United States)

    Liao, Ning

    2016-06-01

    With the diagnosis and treatment of tumor enter into the area of precision medical, based on selected targeted molecular typing of patients with individualized diagnosis and treatment play an important role. HER gene encoded epidermal growth factor receptor 2 (HER2) leading to increased early distant metastasis of breast cancer in patients and poor prognosis. However, a number of clinical studies provided evidence-based anti-HER2 targeted therapy and confirmed the benefit of anti-HER2 targeted therapy in patient survival. In recent years, through the tireless efforts of scholars in the field of breast cancer in our country, the whole diagnosis and treatment of breast cancer has accomplished an international standard. But based on a variety of factors, the anti-HER2 targeted therapy between China and the developed countries, and between different areas in China still exists certain gaps, is now a problem need to be solved. This article will analyzing the diagnostic and treatment on HER2-positive breast cancer in the United States and China, exploring reasons and looking for answers to narrow down the gap in the treatment of HER2-positive breast cancer between China and the United States. Improve the anti-HER2 targeted therapy in our country, let the patients get maximum benefit from anti-HER2 targeted therapy.

  6. Anti-HER2 CD4(+) T-helper type 1 response is a novel immune correlate to pathologic response following neoadjuvant therapy in HER2-positive breast cancer.

    Science.gov (United States)

    Datta, Jashodeep; Berk, Erik; Xu, Shuwen; Fitzpatrick, Elizabeth; Rosemblit, Cinthia; Lowenfeld, Lea; Goodman, Noah; Lewis, David A; Zhang, Paul J; Fisher, Carla; Roses, Robert E; DeMichele, Angela; Czerniecki, Brian J

    2015-05-23

    A progressive loss of circulating anti-human epidermal growth factor receptor-2/neu (HER2) CD4(+) T-helper type 1 (Th1) immune responses is observed in HER2(pos)-invasive breast cancer (IBC) patients relative to healthy controls. Pathologic complete response (pCR) following neoadjuvant trastuzumab and chemotherapy (T + C) is associated with decreased recurrence and improved prognosis. We examined differences in anti-HER2 Th1 responses between pCR and non-pCR patients to identify modifiable immune correlates to pathologic response following neoadjuvant T + C. Anti-HER2 Th1 responses in 87 HER2(pos)-IBC patients were examined using peripheral blood mononuclear cells pulsed with 6 HER2-derived class II peptides via IFN-γ ELISPOT. Th1 response metrics were anti-HER2 responsivity, repertoire (number of reactive peptides), and cumulative response across 6 peptides (spot-forming cells [SFC]/10(6) cells). Anti-HER2 Th1 responses of non-pCR patients (n = 4) receiving adjuvant HER2-pulsed type 1-polarized dendritic cell (DC1) vaccination were analyzed pre- and post-immunization. Depressed anti-HER2 Th1 responses observed in treatment-naïve HER2(pos)-IBC patients (n = 22) did not improve globally in T + C-treated HER2(pos)-IBC patients (n = 65). Compared with adjuvant T + C receipt, neoadjuvant T + C - utilized in 61.5 % - was associated with higher anti-HER2 Th1 repertoire (p = 0.048). While pCR (n = 16) and non-pCR (n = 24) patients did not differ substantially in demographic/clinical characteristics, pCR patients demonstrated dramatically higher anti-HER2 Th1 responsivity (94 % vs. 33 %, p = 0.0002), repertoire (3.3 vs. 0.3 peptides, p vs. 22.4 SFC/10(6), p non-pCR patients. After controlling for potential confounders, anti-HER2 Th1 responsivity remained independently associated with pathologic response (odds ratio 8.82, p = 0.016). This IFN-γ(+) immune disparity was mediated by anti-HER2 CD4(+)T-bet(+)IFN-γ(+) (i.e., Th1) - not CD4(+)GATA-3(+)IFN-γ(+) (i.e., Th2

  7. Anti-HER2 antibody and ScFvEGFR-conjugated antifouling magnetic iron oxide nanoparticles for targeting and magnetic resonance imaging of breast cancer

    Directory of Open Access Journals (Sweden)

    Chen H

    2013-10-01

    Full Text Available Hongwei Chen,1,* Liya Wang,1,2,* Qiqi Yu,1,2 Weiping Qian,3 Diana Tiwari,1 Hong Yi,4 Andrew Y Wang,5 Jing Huang,1,2 Lily Yang,3 Hui Mao1,2 1Department of Radiology and Imaging Sciences, 2Center for Systems Imaging, 3Department of Surgery, Emory University School of Medicine, 4Robert Apkarian Electron Microscopy Core, Emory University, Atlanta, GA, 5Ocean NanoTech LLC, Springdale, AK, USA *These authors contributed equally to this work Abstract: Antifouling magnetic iron oxide nanoparticles (IONPs coated with block copolymer poly(ethylene oxide-block-poly(γ-methacryloxypropyltrimethoxysilane (PEO-b-PγMPS were investigated for improving cell targeting by reducing nonspecific uptake. Conjugation of a HER2 antibody, Herceptin®, or a single chain fragment (ScFv of antibody against epidermal growth factor receptor (ScFvEGFR to PEO-b-PγMPS-coated IONPs resulted in HER2-targeted or EGFR-targeted IONPs (anti-HER2-IONPs or ScFvEGFR-IONPs. The anti-HER2-IONPs bound specifically to SK-BR-3, a HER2-overexpressing breast cancer cell line, but not to MDA-MB-231, a HER2-underexpressing cell line. On the other hand, the ScFvEGFR-IONPs showed strong reactivity with MDA-MB-231, an EGFR-positive human breast cancer cell line, but not with MDA-MB-453, an EGFR-negative human breast cancer cell line. Transmission electron microscopy revealed internalization of the receptor-targeted nanoparticles by the targeted cancer cells. In addition, both antibody-conjugated and non-antibody-conjugated IONPs showed reduced nonspecific uptake by RAW264.7 mouse macrophages in vitro. The developed IONPs showed a long blood circulation time (serum half-life 11.6 hours in mice and low accumulation in both the liver and spleen. At 24 hours after systemic administration of ScFvEGFR-IONPs into mice bearing EGFR-positive breast cancer 4T1 mouse mammary tumors, magnetic resonance imaging revealed signal reduction in the tumor as a result of the accumulation of the targeted IONPs

  8. Evaluation of anti-HER2 scFv-conjugated PLGA–PEG nanoparticles on 3D tumor spheroids of BT474 and HCT116 cancer cells

    International Nuclear Information System (INIS)

    Le, Thi Thuy Duong; Pham, Thu Hong; Ngo, Thi Hong Giang; Le, Quang Huan; Nguyen, Trong Nghia; Hoang, Thi My Nhung

    2016-01-01

    Three-dimensional culture cells (spheroids) are one of the multicellular culture models that can be applied to anticancer chemotherapeutic development. Multicellular spheroids more closely mimic in vivo tumor-like patterns of physiologic environment and morphology. In previous research, we designed docetaxel-loaded pegylated poly(D, L-lactide-co-glycolide) nanoparticles conjugated with anti-HER2 single chain antibodies (scFv–Doc–PLGA–PEG) and evaluated them in 2D cell culture. In this study, we continuously evaluate the cellular uptake and cytotoxic effect of scFv–Doc–PLGA–PEG on a 3D tumor spheroid model of BT474 (HER2-overexpressing) and HCT116 (HER2-underexpressing) cancer cells. The results showed that the nanoparticle formulation conjugated with scFv had a significant internalization effect on the spheroids of HER2-overexpressing cancer cells as compared to the spheroids of HER2-underexpressing cancer cells. Therefore, cytotoxic effects of targeted nanoparticles decreased the size and increased necrotic score of HER2-overexpressing tumor spheroids. Thus, these scFv–Doc–PLGA–PEG nanoparticles have potential for active targeting for HER2-overexpressing cancer therapy. In addition, BT474 and HCT116 spheroids can be used as a tumor model for evaluation of targeting therapies. (paper)

  9. Radiobromination of humanized anti-HER2 monoclonal antibody trastuzumab using N-succinimidyl 5-bromo-3-pyridinecarboxylate, a potential label for immunoPET

    Energy Technology Data Exchange (ETDEWEB)

    Mume, Eskender [Organic Chemistry, Department of Chemistry, Uppsala University, S-751 24 Uppsala (Sweden); Orlova, Anna [Affibody AB, S-161 02 Bromma (Sweden); Malmstroem, Per-Uno [Division of Urology, Department of Surgical Sciences, Uppsala University, S-751 85 Uppsala (Sweden); Lundqvist, Hans [Unit of Biomedical Radiation Sciences, Rudbeck Laboratory, Uppsala University, S-751 85 Uppsala (Sweden); Sjoeberg, Stefan [Organic Chemistry, Department of Chemistry, Uppsala University, S-751 24 Uppsala (Sweden); Tolmachev, Vladimir [Unit of Biomedical Radiation Sciences, Rudbeck Laboratory, Uppsala University, S-751 85 Uppsala (Sweden)]. E-mail: vladimir.tolmachev@bms.uu.se

    2005-08-01

    Combining the specificity of radioimmunoscintigraphy and the high sensitivity of PET in an in vivo detection technique could improve the quality of nuclear diagnostics. Positron-emitting nuclide {sup 76}Br (T {sub 1/2}=16.2 h) might be a possible candidate for labeling monoclonal antibodies (mAbs) and their fragments, provided that the appropriate labeling chemistry has been established. For internalizing antibodies, such as the humanized anti-HER2 monoclonal antibody, trastuzumab, radiobromine label should be residualizing, i.e., ensuring that radiocatabolites are trapped intracellularly after the proteolytic degradation of antibody. This study evaluated the chemistry of indirect radiobromination of trastuzumab using N-succinimidyl 5-(tributylstannyl)-3-pyridinecarboxylate. Literature data indicated that the use of this method provided residualizing properties for iodine and astatine labels on some antibodies. An optimized 'one-pot' procedure produced an overall labeling efficiency of 45.5{+-}1.2% over 15 min. The bromine label was stable under physiological and denaturing conditions. The labeled trastuzumab retained its capacity to bind specifically to HER2-expressing SKOV-3 ovarian carcinoma cells in vitro (immunoreactivity more than 75%). However, in vitro cell test did not demonstrate that the radiobromination of trastuzumab using N-succinimidyl 5-bromo-3-pyridinecarboxylate improves cellular retention of radioactivity in comparison with the use of N-succinimidyl 4-bromobenzoate.

  10. A novel dendritic cell-based immunization approach for the induction of durable Th1-polarized anti-HER-2/neu responses in women with early breast cancer

    Science.gov (United States)

    Koski, Gary K.; Koldovsky, Ursula; Xu, Shuwen; Mick, Rosemarie; Sharma, Anupama; Fitzpatrick, Elizabeth; Weinstein, Susan; Nisenbaum, Harvey; Levine, Bruce L; Fox, Kevin; Zhang, Paul; Czerniecki, Brian J

    2011-01-01

    Twenty-seven subjects with HER-2/neu over-expressing ductal carcinoma in situ of the breast were enrolled in a neoadjuvant immunization trial for safety and immunogenicity of DC1-polarized dendritic cells (DC1) pulsed with six HER-2/neu promiscuous MHC class II-binding peptides, plus two additional HLA-A2.1 class I-binding peptides. DC1 were generated with IFN-γ plus a special clinical-grade bacterial endotoxin (LPS) and administered directly into groin lymph nodes four times at weekly intervals prior to scheduled surgical resection of DCIS. Subjects were monitored for the induction of new or enhanced anti-peptide reactivity by IFN-γ ELIspot and ELISA assays performed on Th cells obtained from peripheral blood or excised sentinel lymph nodes. Responses by CTL against HLA-A2.1-binding peptides were measured using peptide-pulsed T2 target cells or HER-2/neu-expressing or non-expressing tumor cell lines. DC1 showed surface phenotype indistinct from “gold standard” inflammatory cocktail-activated DC, but displayed a number of distinguishing functional characteristics including the secretion of soluble factors and enhanced “killer DC” capacity against tumor cells in vitro. Post-immunization, we observed sensitization of Th cells to at least 1 class II peptide in 22 of 25 (88%, 95% exact CI 68.8 – 97.5%) evaluable subjects, while eleven of 13 (84.6%, 95% exact CI 64 – 99.8%) HLA-A2.1 subjects were successfully sensitized to class I peptides. Perhaps most importantly, anti-HER-2/neu peptide responses were observed up to 52 months post-immunization. These data show even in the presence of early breast cancer such DC1 are potent inducers of durable type I-polarized immunity, suggesting potential clinical value for development of cancer immunotherapy. PMID:22130160

  11. Anti-HER2 immunoliposomes for selective delivery of electron paramagnetic resonance imaging probes to HER2-overexpressing breast tumor cells

    Science.gov (United States)

    Burks, Scott R.; Macedo, Luciana F.; Barth, Eugene D.; Tkaczuk, Katherine H.; Martin, Stuart S.; Rosen, Gerald M.; Halpern, Howard J.; Brodie, Angela M.

    2014-01-01

    Electron paramagnetic resonance (EPR) imaging is an emerging modality that can detect and localize paramagnetic molecular probes (so-called spin probes) in vivo. We previously demonstrated that nitroxide spin probes can be encapsulated in liposomes at concentrations exceeding 100 mM, at which nitroxides exhibit a concentration-dependent quenching of their EPR signal that is analogous to the self-quenching of fluorescent molecules. Therefore, intact liposomes encapsulating high concentrations of nitroxides exhibit greatly attenuated EPR spectral signals, and endocytosis of such liposomes represents a cell-activated contrast-generating mechanism. After endocytosis, the encapsulated nitroxide is liberated and becomes greatly diluted in the intracellular milieu. This dequenches the nitroxides to generate a robust intracellular EPR signal. It is therefore possible to deliver a high concentration of nitroxides to cells while minimizing background signal from unendocytosed liposomes. We report here that intracellular EPR signal can be selectively generated in a specific cell type by exploiting its expression of Human Epidermal Growth Factor Receptor 2 (HER2). When targeted by anti-HER2 immunoliposomes encapsulating quenched nitroxides, Hc7 cells, which are novel HER2-overexpressing cells derived from the MCF7 breast tumor cell line, endocytose the liposomes copiously, in contrast to the parent MCF7 cells or control CV1 cells, which do not express HER2. HER2-dependent liposomal delivery enables Hc7 cells to accumulate 750 μM nitroxide intracellularly. Through the use of phantom models, we verify that this concentration of nitroxides is more than sufficient for EPR imaging, thus laying the foundation for using EPR imaging to visualize HER2-overexpressing Hc7 tumors in animals. PMID:20066490

  12. Optimal specific radioactivity of anti-HER2 Affibody molecules enables discrimination between xenografts with high and low HER2 expression levels

    Energy Technology Data Exchange (ETDEWEB)

    Tolmachev, Vladimir [Uppsala University, Biomedical Radiation Sciences, Rudbeck Laboratory, Uppsala (Sweden); Uppsala University, Department of Medical Sciences, Nuclear Medicine, Uppsala (Sweden); Waallberg, Helena [Royal Institute of Technology, School of Biotechnology, Stockholm (Sweden); Sandstroem, Mattias [Uppsala University Hospital, Section of Hospital Physics, Department of Oncology, Uppsala (Sweden); Hansson, Monika; Wennborg, Anders [Affibody AB, Stockholm (Sweden); Orlova, Anna [Uppsala University, Biomedical Radiation Sciences, Rudbeck Laboratory, Uppsala (Sweden)

    2011-03-15

    Overexpression of the HER2 receptor is a biomarker for predicting those patients who may benefit from trastuzumab therapy. Radiolabelled Affibody molecules can be used to visualize HER2 expression in tumour xenografts with high sensitivity. However, previous studies demonstrated that the difference in uptake in xenografts with high and low HER2 expression levels is not proportional to the difference in expression levels. We hypothesized that discrimination between tumours with high and low HER2 expression may be improved by increasing the injected dose (reducing the specific activity) of the tracer. The influence of injected dose of anti-HER2 {sup 111}In-DOTA-Z{sub HER2} {sub 342} Affibody molecule on uptake in SKOV-3 (high HER2 expression) and LS174T (low expression) xenografts was investigated. The optimal range of injected doses enabling discrimination between xenografts with high and low expression was determined. To verify this, tumour uptake was measured in mice carrying both SKOV-3 and LS174T xenografts after injection of either 1 or 15 {mu}g {sup 111}In-DOTA-Z{sub HER2:342}. An increase in the injected dose caused a linear decrease in the radioactivity accumulation in the LS174T xenografts (low HER2 expression). For SKOV-3 xenografts, the dependence of the tumour uptake on the injected dose was less dramatic. The injection of 10-30 {mu}g {sup 111}In-DOTA-Z{sub HER2:342} per mouse led to the largest difference in uptake between the two types of tumour. Experiments in mice bearing two xenografts confirmed that the optimized injected dose enabled better discrimination of expression levels. Careful optimization of the injected dose of Affibody molecules is required for maximum discrimination between xenografts with high and low levels of HER2 expression. This information has potential relevance for clinical imaging applications. (orig.)

  13. tabAnti-HER2 (erbB-2) oncogene effects of phenolic compounds directly isolated from commercial Extra-Virgin Olive Oil (EVOO)

    International Nuclear Information System (INIS)

    Menendez, Javier A; Vazquez-Martin, Alejandro; Garcia-Villalba, Rocio; Carrasco-Pancorbo, Alegria; Oliveras-Ferraros, Cristina; Fernandez-Gutierrez, Alberto; Segura-Carretero, Antonio

    2008-01-01

    The effects of the olive oil-rich Mediterranean diet on breast cancer risk might be underestimated when HER2 (ERBB2) oncogene-positive and HER2-negative breast carcinomas are considered together. We here investigated the anti-HER2 effects of phenolic fractions directly extracted from Extra Virgin Olive Oil (EVOO) in cultured human breast cancer cell lines. Solid phase extraction followed by semi-preparative high-performance liquid chromatography (HPLC) was used to isolate phenolic fractions from commercial EVOO. Analytical capillary electrophoresis coupled to mass spectrometry was performed to check for the composition and to confirm the identity of the isolated fractions. EVOO polyphenolic fractions were tested on their tumoricidal ability against HER2-negative and HER2-positive breast cancer in vitro models using MTT, crystal violet staining, and Cell Death ELISA assays. The effects of EVOO polyphenolic fractions on the expression and activation status of HER2 oncoprotein were evaluated using HER2-specific ELISAs and immunoblotting procedures, respectively. Among the fractions mainly containing the single phenols hydroxytyrosol and tyrosol, the polyphenol acid elenolic acid, the lignans (+)-pinoresinol and 1-(+)-acetoxypinoresinol, and the secoiridoids deacetoxy oleuropein aglycone, ligstroside aglycone, and oleuropein aglycone, all the major EVOO polyphenols (i.e. secoiridoids and lignans) were found to induce strong tumoricidal effects within a micromolar range by selectively triggering high levels of apoptotic cell death in HER2-overexpressors. Small interfering RNA-induced depletion of HER2 protein and lapatinib-induced blockade of HER2 tyrosine kinase activity both significantly prevented EVOO polyphenols-induced cytotoxicity. EVOO polyphenols drastically depleted HER2 protein and reduced HER2 tyrosine autophosphorylation in a dose- and time-dependent manner. EVOO polyphenols-induced HER2 downregulation occurred regardless the molecular mechanism

  14. tabAnti-HER2 (erbB-2 oncogene effects of phenolic compounds directly isolated from commercial Extra-Virgin Olive Oil (EVOO

    Directory of Open Access Journals (Sweden)

    Carrasco-Pancorbo Alegria

    2008-12-01

    Full Text Available Abstract Background The effects of the olive oil-rich Mediterranean diet on breast cancer risk might be underestimated when HER2 (ERBB2 oncogene-positive and HER2-negative breast carcinomas are considered together. We here investigated the anti-HER2 effects of phenolic fractions directly extracted from Extra Virgin Olive Oil (EVOO in cultured human breast cancer cell lines. Methods Solid phase extraction followed by semi-preparative high-performance liquid chromatography (HPLC was used to isolate phenolic fractions from commercial EVOO. Analytical capillary electrophoresis coupled to mass spectrometry was performed to check for the composition and to confirm the identity of the isolated fractions. EVOO polyphenolic fractions were tested on their tumoricidal ability against HER2-negative and HER2-positive breast cancer in vitro models using MTT, crystal violet staining, and Cell Death ELISA assays. The effects of EVOO polyphenolic fractions on the expression and activation status of HER2 oncoprotein were evaluated using HER2-specific ELISAs and immunoblotting procedures, respectively. Results Among the fractions mainly containing the single phenols hydroxytyrosol and tyrosol, the polyphenol acid elenolic acid, the lignans (+-pinoresinol and 1-(+-acetoxypinoresinol, and the secoiridoids deacetoxy oleuropein aglycone, ligstroside aglycone, and oleuropein aglycone, all the major EVOO polyphenols (i.e. secoiridoids and lignans were found to induce strong tumoricidal effects within a micromolar range by selectively triggering high levels of apoptotic cell death in HER2-overexpressors. Small interfering RNA-induced depletion of HER2 protein and lapatinib-induced blockade of HER2 tyrosine kinase activity both significantly prevented EVOO polyphenols-induced cytotoxicity. EVOO polyphenols drastically depleted HER2 protein and reduced HER2 tyrosine autophosphorylation in a dose- and time-dependent manner. EVOO polyphenols-induced HER2 downregulation

  15. Preclinical Evaluation of 18F-Labeled Anti-HER2 Nanobody Conjugates for Imaging HER2 Receptor Expression by Immuno-PET.

    Science.gov (United States)

    Vaidyanathan, Ganesan; McDougald, Darryl; Choi, Jaeyeon; Koumarianou, Eftychia; Weitzel, Douglas; Osada, Takuya; Lyerly, H Kim; Zalutsky, Michael R

    2016-06-01

    The human growth factor receptor type 2 (HER2) is overexpressed in breast as well as other types of cancer. Immuno-PET, a noninvasive imaging procedure that could assess HER2 status in both primary and metastatic lesions simultaneously, could be a valuable tool for optimizing application of HER2-targeted therapies in individual patients. Herein, we have evaluated the tumor-targeting potential of the 5F7 anti-HER2 Nanobody (single-domain antibody fragment; ∼13 kDa) after (18)F labeling by 2 methods. The 5F7 Nanobody was labeled with (18)F using the novel residualizing label N-succinimidyl 3-((4-(4-(18)F-fluorobutyl)-1H-1,2,3-triazol-1-yl)methyl)-5-(guanidinomethyl)benzoate ((18)F-SFBTMGMB; (18)F-RL-I) and also via the most commonly used (18)F protein-labeling prosthetic agent N-succinimidyl 3-(18)F-fluorobenzoate ((18)F-SFB). For comparison, 5F7 Nanobody was also labeled using the residualizing radioiodination agent N-succinimidyl 4-guanidinomethyl-3-(125)I-iodobenzoate ((125)I-SGMIB). Paired-label ((18)F/(125)I) internalization assays and biodistribution studies were performed on HER2-expressing BT474M1 breast carcinoma cells and in mice with BT474M1 subcutaneous xenografts, respectively. Small-animal PET/CT imaging of 5F7 Nanobody labeled using (18)F-RL-I also was performed. Internalization assays indicated that intracellularly retained radioactivity for (18)F-RL-I-5F7 was similar to that for coincubated (125)I-SGMIB-5F7, whereas that for (18)F-SFB-5F7 was lower than coincubated (125)I-SGMIB-5F7 and decreased with time. BT474M1 tumor uptake of (18)F-RL-I-5F7 was 28.97 ± 3.88 percentage injected dose per gram of tissue (%ID/g) at 1 h and 36.28 ± 14.10 %ID/g at 2 h, reduced by more than 90% on blocking with trastuzumab, indicating HER2 specificity of uptake, and was also 26%-28% higher (P < 0.05) than that of (18)F-SFB-5F7. At 2 h, the tumor-to-blood ratio for (18)F-RL-I-5F7 (47.4 ± 13.1) was significantly higher (P < 0.05) than for (18)F-SFB-5F7 (25.4 ± 10

  16. Characterization of an engineered human purine nucleoside phosphorylase fused to an anti-her2/neu single chain Fv for use in ADEPT

    Directory of Open Access Journals (Sweden)

    Wu Anna M

    2009-12-01

    Full Text Available Abstract Background Antibody Directed Enzyme Prodrug Therapy (ADEPT can be used to generate cytotoxic agents at the tumor site. To date non-human enzymes have mainly been utilized in ADEPT. However, these non-human enzymes are immunogenic limiting the number of times that ADEPT can be administered. To overcome the problem of immunogenicity, a fully human enzyme, capable of converting a non-toxic prodrug to cytotoxic drug was developed and joined to a human tumor specific scFv yielding a fully human targeting agent. Methods A double mutant of human purine nucleoside phosphorylase (hDM was developed which unlike the human enzyme can cleave adenosine-based prodrugs. For tumor-specific targeting, hDM was fused to the human anti-HER2/neu single chain Fv (scFv, C6 MH3B1. Enzymatic activity of hDM with its natural substrates and prodrugs was determined using spectrophotomeric approaches. A cell proliferation assay was used to assess the cytotoxicity generated following conversion of prodrug to drug as a result of enzymatic activity of hDM. Affinity of the targeting scFv, C6 MH3B1 fused to hDM to Her2/neu was confirmed using affinity chromatography, surface plasmon resonance, and flow-cytometry. Results In vitro hDM-C6 MH3B1 binds specifically to HER2/neu expressing tumor cells and localizes hDM to tumor cells, where the enzymatic activity of hDM-C6 MH3B1, but not the wild type enzyme, results in phosphorolysis of the prodrug, 2-fluoro-2'-deoxyadenosine to the cytotoxic drug 2-fluoroadenine (F-Ade causing inhibition of tumor cell proliferation. Significantly, the toxic small drug diffuses through the cell membrane of HER2/neu expressing cells as well as cells that lack the expression of HER2/neu, causing a bystander effect. F-Ade is toxic to cells irrespective of their growth rate; therefore, both the slowly dividing tumor cells and the non-dividing neighboring stromal cells that support tumor growth should be killed. Analysis of potential novel MHCII

  17. Prospective Biomarker Analysis of the Randomized CHER-LOB Study Evaluating the Dual Anti-HER2 Treatment With Trastuzumab and Lapatinib Plus Chemotherapy as Neoadjuvant Therapy for HER2-Positive Breast Cancer.

    Science.gov (United States)

    Guarneri, Valentina; Dieci, Maria Vittoria; Frassoldati, Antonio; Maiorana, Antonino; Ficarra, Guido; Bettelli, Stefania; Tagliafico, Enrico; Bicciato, Silvio; Generali, Daniele Giulio; Cagossi, Katia; Bisagni, Giancarlo; Sarti, Samanta; Musolino, Antonino; Ellis, Catherine; Crescenzo, Rocco; Conte, PierFranco

    2015-09-01

    The CHER-LOB randomized phase II study showed that the combination of lapatinib and trastuzumab plus chemotherapy increases the pathologic complete remission (pCR) rate compared with chemotherapy plus either trastuzumab or lapatinib. A biomarker program was prospectively planned to identify potential predictors of sensitivity to different treatments and to evaluate treatment effect on tumor biomarkers. Overall, 121 breast cancer patients positive for human epidermal growth factor 2 (HER2) were randomly assigned to neoadjuvant chemotherapy plus trastuzumab, lapatinib, or both trastuzumab and lapatinib. Pre- and post-treatment samples were centrally evaluated for HER2, p95-HER2, phosphorylated AKT (pAKT), phosphatase and tensin homolog, Ki67, apoptosis, and PIK3CA mutations. Fresh-frozen tissue samples were collected for genomic analyses. A mutation in PIK3CA exon 20 or 9 was documented in 20% of cases. Overall, the pCR rates were similar in PIK3CA wild-type and PIK3CA-mutated patients (33.3% vs. 22.7%; p = .323). For patients receiving trastuzumab plus lapatinib, the probability of pCR was higher in PIK3CA wild-type tumors (48.4% vs. 12.5%; p = .06). Ki67, pAKT, and apoptosis measured on the residual disease were significantly reduced from baseline. The degree of Ki67 inhibition was significantly higher in patients receiving the dual anti-HER2 blockade. The integrated analysis of gene expression and copy number data demonstrated that a 50-gene signature specifically predicted the lapatinib-induced pCR. PIK3CA mutations seem to identify patients who are less likely to benefit from dual anti-HER2 inhibition. p95-HER2 and markers of phosphoinositide 3-kinase pathway deregulation are not confirmed as markers of different sensitivity to trastuzumab or lapatinib. HER2 is currently the only validated marker to select breast cancer patients for anti-HER2 treatment; however, it is becoming evident that HER2-positive breast cancer is a heterogeneous disease. In addition, more

  18. Influence of Alternative Tubulin Inhibitors on the Potency of a Epirubicin-Immunochemotherapeutic Synthesized with an Ultra Violet Light-Activated Intermediate: Influence of incorporating an internal/integral disulfide bond structure and Alternative Tubulin/Microtubule Inhibitors on the Cytotoxic Anti-Neoplastic Potency of Epirubicin-(C3-amide)-Anti-HER2/neu Synthesized Utilizing a UV-Photoactivated Anthracycline Intermediate.

    Science.gov (United States)

    Coyne, C P; Jones, Toni; Bear, Ryan

    2012-11-01

    Immunochemotherapeutics, epirubicin-(C 3 - amide )-SS-[anti-HER2/ neu ] with an internal disulfide bond, and epirubicin-(C 3 - amide )-[anti-HER2/ neu ] were synthesized utilizing succinimidyl 2-[(4,4'-azipentanamido) ethyl]-1,3'-dithioproprionate or succinimidyl 4,4-azipentanoate respectively. Western blot analysis was used to determine the presence of any immunoglobulin fragmentation or IgG-IgG polymerization. Retained HER2/ neu binding characteristics of epirubicin-(C 3 - amide )-[anti-HER2/ neu ] and epirubicin-(C 3 - amide )-SS-[anti-HER2/ neu ] were validated by cell-ELISA using a mammary adenocarcinoma (SKBr-3) population that highly over-expresses trophic HER2/ neu receptor complexes. Cytotoxic anti-neoplastic potency of epirubicin-(C 3 - amide )-[anti-HER2/ neu ] and epirubicin-(C 3 - amide )-SS-[anti-HER2/ neu ] between epirubicin-equivalent concentrations of 10 -10 M and 10 -6 M was determined by measuring the vitality/proliferation of chemotherapeutic-resistant mammary adenocarcinoma (SKBr-3 cell type). Cytotoxic anti-neoplastic potency of benzimidazoles (albendazole, flubendazole, membendazole) and griseofulvin were assessed between 0-to-2 μg/ml and 0-to-100 μg/ml respectively while mebendazole and griseofulvin were analyzed at fixed concentrations of 0.35 μg/ml and 35 g/ml respectively in dual combination with gradient concentrations of epirubicin-(C 3 - amide )-[anti-HER2/ neu ] and epirubicin-(C 3 - amide )-SS-[anti-HER2/ neu ]. Cytotoxic anti-neoplastic potency for epirubicin-(C 3 - amide )-[anti-HER2/ neu ] and epirubicin-(C 3 - amide )-SS-[anti-HER2/ neu ] against chemotherapeutic-resistant mammary adenocarcinoma (SKBr-3) was nearly identical at epirubicin-equivalent concentrations of 10 -10 M and 10 -6 M. The benzimadazoles also possessed cytotoxic anti-neoplastic activity with flubendazole and albendazole being the most and least potent respectively. Similarly, griseofulvin had cytotoxic anti-neoplastic activity and was more potent than

  19. Radiobromination of anti-HER2/neu/ErbB-2 monoclonal antibody using the p-isothiocyanatobenzene derivative of the [76Br]undecahydro-bromo-7,8-dicarba-nido-undecaborate(1-) ion

    International Nuclear Information System (INIS)

    Winberg, Karl Johan; Persson, Mikael; Malmstroem, Per-Uno; Sjoeberg, Stefan; Tolmachev, Vladimir

    2004-01-01

    The monoclonal humanized anti-HER2 antibody trastuzumab was radiolabeled with the positron emitter 76 Br (T ((1)/(2)) =16.2 h). Indirect labeling was performed using the p-isothiocyanatobenzene derivative of the [ 76 Br]undecahydro-bromo-7,8-dicarba-nido-undecaborate(1-) ( 76 Br-NBI) as a precursor molecule. 76 Br-NBI was prepared by bromination of the 7-(p-isothiocyanato-phenyl)dodecahydro-7,8-dicarba-nido-undecaborate(1-) ion (NBI) with a yield of 93-95% using Chloramine-T (CAT) as an oxidant. Coupling of radiobrominated NBI to antibody was performed without intermediate purification, in an ''one pot'' reaction. An overall labeling yield of 55.7 ± 4.8% (mean ± maximum error) was achieved when 300 μg of antibody was labeled. The label was stable in vitro in physiological and denaturing conditions. In a cell binding test, trastuzumab remained immunoreactive after labeling

  20. My IGY in Antarctica

    Science.gov (United States)

    Bentley, Charles

    2012-01-01

    Dr Charles Bentley is the A.P. Crary Professor Emeritus of Geophysics, Department of Geology and Geophysics, University of Wisconsin-Madison. Dr. Bentley joined the Arctic Institute of North America in 1956 to participate in International Geophysical Year (IGY)-related activities in the Antarctic. He wintered over consecutively in 1957 and 1958 at Byrd Station, a station in the interior of West Antarctica that housed 24 men each winter - 12 Navy support people and 12 civilian scientists/technicians. During the austral summers, he also participated in over-snow traverses, first as co-leader, then leader (the other coleader went home after the first year). These traverses consisted of six men and three vehicles, and lasted several months. These traverses covered more than 1609 kilometers (1000 miles) of largely unmapped and unphotographed terrain. During these traverses, connections to Byrd Station were by radio (daily, when the transmission conditions were good enough) and roughly every 2 weeks by resupply flight.

  1. Efficacy of Anti-HER2 Agents in Combination With Adjuvant or Neoadjuvant Chemotherapy for Early and Locally Advanced HER2-Positive Breast Cancer Patients: A Network Meta-Analysis

    Directory of Open Access Journals (Sweden)

    Márcio Debiasi

    2018-05-01

    Full Text Available BackgroundSeveral (neoadjuvant treatments for patients with HER2-positive breast cancer have been compared in different randomized clinical trials. Since it is not feasible to conduct adequate pairwise comparative trials of all these therapeutic options, network meta-analysis offers an opportunity for more detailed inference for evidence-based therapy.MethodsPhase II/III randomized clinical trials comparing two or more different (neoadjuvant treatments for HER2-positive breast cancer patients were included. Relative treatment effects were pooled in two separate network meta-analyses for overall survival (OS and disease-free survival (DFS.Results17 clinical trials met our eligibility criteria. Two different networks of trials were created based on the availability of the outcomes: OS network (15 trials: 37,837 patients; and DFS network (17 trials: 40,992 patients. Two studies—the ExteNET and the NeoSphere trials—were included only in this DFS network because OS data have not yet been reported. The concept of the dual anti-HER2 blockade proved to be the best option in terms of OS and DFS. Chemotherapy (CT plus trastuzumab (T and lapatinib (L and CT + T + Pertuzumab (P are probably the best treatment options in terms of OS, with 62.47% and 22.06%, respectively. In the DFS network, CT + T + Neratinib (N was the best treatment option with 50.55%, followed by CT + T + P (26.59% and CT + T + L (20.62%.ConclusionThis network meta-analysis suggests that dual anti-HER2 blockade with trastuzumab plus either lapatinib or pertuzumab are probably the best treatment options in the (neoadjuvant setting for HER2-positive breast cancer patients in terms of OS gain. Mature OS results are still expected for the Aphinity trial and for the sequential use of trastuzumab followed by neratinib, the treatment that showed the best performance in terms of DFS in our analysis.

  2. Radiobromination of anti-HER2/neu/ErbB-2 monoclonal antibody using the p-isothiocyanatobenzene derivative of the [{sup 76}Br]undecahydro-bromo-7,8-dicarba-nido-undecaborate(1-) ion

    Energy Technology Data Exchange (ETDEWEB)

    Winberg, Karl Johan; Persson, Mikael; Malmstroem, Per-Uno; Sjoeberg, Stefan; Tolmachev, Vladimir E-mail: Vladimir.Tolmachev@bms.uu.se

    2004-05-01

    The monoclonal humanized anti-HER2 antibody trastuzumab was radiolabeled with the positron emitter {sup 76}Br (T{sub ((1)/(2))} =16.2 h). Indirect labeling was performed using the p-isothiocyanatobenzene derivative of the [{sup 76}Br]undecahydro-bromo-7,8-dicarba-nido-undecaborate(1-) ({sup 76}Br-NBI) as a precursor molecule. {sup 76}Br-NBI was prepared by bromination of the 7-(p-isothiocyanato-phenyl)dodecahydro-7,8-dicarba-nido-undecaborate(1-) ion (NBI) with a yield of 93-95% using Chloramine-T (CAT) as an oxidant. Coupling of radiobrominated NBI to antibody was performed without intermediate purification, in an ''one pot'' reaction. An overall labeling yield of 55.7 {+-} 4.8% (mean {+-} maximum error) was achieved when 300 {mu}g of antibody was labeled. The label was stable in vitro in physiological and denaturing conditions. In a cell binding test, trastuzumab remained immunoreactive after labeling.

  3. Comparison between anti-CEA and anti-HER2 212Pb-labeled mAbs during α-RIT of small volume peritoneal carcinomatosis - Role of activity distribution on therapeutic efficacy and toxicity?

    International Nuclear Information System (INIS)

    Elgqvist, J.; Boudousq, V.; Bobyk, L.; Busson, M.; Lozza, C.; Navarro-Teulon, I.; Pouget, J.P.; Maquaire, P.; Torgue, J.

    2015-01-01

    Full text of publication follows. Objectives: we investigated the role of internalizing/non-internalizing monoclonal antibodies (mAbs) on the final outcome (efficacy/toxicity) of mice treated with alpha radioimmunotherapy (α- RIT) using 212 Pb-labeled mAbs. The relationship between distribution of radioactivity at the tissue level and biological parameters was also assessed. Methods: nude mice bearing 2-3 mm peritoneal nodules obtained by xenograft of A-431 tumor cells, expressing low and high level of HER2 and CEA receptors, respectively, were intraperitoneally (i.p.) injected with increasing activities (370-1480 kBq; 37 MBq/mg) of either 35A7 (non-internalising anti-CEA), Trastuzumab (internalizing anti-HER2) or PX (non-specific) 212 Pb-labeled mAbs. Control groups were injected with corresponding amount of unlabeled mAbs or with NaCl. Tumor growth was followed by bioluminescence and median survival (MS) of control and treated mice was determined. 212 Pb-35A7 and 212 Pb-Trastuzumab biodistribution was used to determine the cumulative uptake of radioactivity (UOR) in organs and tumors. Mean absorbed doses were calculated using the MIRD formalism. Haematological, liver and kidney toxicities were also assessed. Distribution of radioactivity at the tissue level was determined by digital micro-autoradiography and the relationship with biological markers of tissue damage was investigated using immunohistochemistry. Results: a mild and transient haematological toxicity in groups treated with the highest amount of activity was observed. MS of the groups treated either with internalizing or non-internalizing 212 Pb-labeled mAbs was significantly improved compared to those treated with non-specific 212 Pb-PX or those only given unlabeled mAbs or just NaCl. MS ranged from 42 days to 94 days using various activity levels of anti-CEA 212 Pb-35A7 while MS was not reached over the follow-up period of 130 days for mice treated with anti-HER2 212 Pb-Trastuzumab. However, UOR and

  4. Comparative evaluation of synthetic anti-HER2 Affibody molecules site-specifically labelled with {sup 111}In using N-terminal DOTA, NOTA and NODAGA chelators in mice bearing prostate cancer xenografts

    Energy Technology Data Exchange (ETDEWEB)

    Malmberg, Jennie; Varasteh, Zohreh; Orlova, Anna [Uppsala University, Preclinical PET Platform, Department of Medicinal Chemistry, Uppsala (Sweden); Perols, Anna; Braun, Alexis; Eriksson Karlstroem, Amelie [AlbaNova University Centre, Division of Molecular Biotechnology, School of Biotechnology, KTH Royal Institute of Technology, Stockholm (Sweden); Altai, Mohamed; Tolmachev, Vladimir [Uppsala University, Unit of Biomedical Radiation Sciences, Rudbeck Laboratory, Uppsala (Sweden); Sandstroem, Mattias [Uppsala University Hospital, Section of Medical Physics, Department of Oncology, Uppsala (Sweden); Garske, Ulrike [Uppsala University Hospital, Department of Medical Sciences, Section of Nuclear Medicine, Uppsala (Sweden)

    2012-03-15

    In disseminated prostate cancer, expression of human epidermal growth factor receptor type 2 (HER2) is one of the pathways to androgen independence. Radionuclide molecular imaging of HER2 expression in disseminated prostate cancer might identify patients for HER2-targeted therapy. Affibody molecules are small (7 kDa) targeting proteins with high potential as tracers for radionuclide imaging. The goal of this study was to develop an optimal Affibody-based tracer for visualization of HER2 expression in prostate cancer. A synthetic variant of the anti-HER2 Z{sub HER2:342} Affibody molecule, Z{sub HER2:S1}, was N-terminally conjugated with the chelators DOTA, NOTA and NODAGA. The conjugated proteins were biophysically characterized by electrospray ionization mass spectroscopy (ESI-MS), circular dichroism (CD) spectroscopy and surface plasmon resonance (SPR)-based biosensor analysis. After labelling with {sup 111}In, the biodistribution was assessed in normal mice and the two most promising conjugates were further evaluated for tumour targeting in mice bearing DU-145 prostate cancer xenografts. The HER2-binding equilibrium dissociation constants were 130, 140 and 90 pM for DOTA-Z{sub HER2:S1}, NOTA-Z{sub HER2:S1} and NODAGA-Z{sub HER2:S1}, respectively. A comparative study of {sup 111}In-labelled DOTA-Z{sub HER2:S1}, NOTA-Z{sub HER2:S1} and NODAGA-Z{sub HER2:S1} in normal mice demonstrated a substantial influence of the chelators on the biodistribution properties of the conjugates. {sup 111}In-NODAGA-Z{sub HER2:S1} had the most rapid clearance from blood and healthy tissues. {sup 111}In-NOTA-Z{sub HER2:S1} showed high hepatic uptake and was excluded from further evaluation. {sup 111}In-DOTA-Z{sub HER2:S1} and {sup 111}In-NODAGA-Z{sub HER2:S1} demonstrated specific uptake in DU-145 prostate cancer xenografts in nude mice. The tumour uptake of {sup 111}In-NODAGA-Z{sub HER2:S1}, 5.6 {+-} 0.4%ID/g, was significantly lower than the uptake of {sup 111}In-DOTA-Z{sub HER2:S1

  5. Avian IgY antibodies: characteristics and applications in immunodiagnostic

    Directory of Open Access Journals (Sweden)

    Lívia Silveira Munhoz

    2014-01-01

    Full Text Available Immunoglobulin Y (IgY is the major antibody isotype in birds, reptiles, amphibia, and lungfish, playing a similar biological role as mammal IgG. Due to its phylogenetic distance, immune diversification and presence in the egg yolk, IgY provide a number of advantages in immunodiagnostic compared to IgG from mammals. Moreover, IgY production is in agreement with international efforts to reduce, refine and if possible, to replace animals in experimentation, contributing substantially in favor of animal welfare. This article presents an overview about structural and functional features, production and applications of IgY in immunodiagnostic, as well as the advantages of chicken antibodies use.

  6. PRODUKSI ANTIBODI KUNING TELUR (IGY ANTI STREPTOCOCCUS MUTANS SEBAGAI ANTI KARIES GIGI

    Directory of Open Access Journals (Sweden)

    Okti Nadia Poetri

    2006-12-01

    Full Text Available The aim of this study was to explore IgY anti Streptococcus mutan production and the ability of Igy Streptococcus mutans blocking adhesion process. The eggs was collected from Single Comb Brown Leghorn which have been immunized by S. mutan. Agar gel precipitation test was done to detect IgY anti S. mutans in serum and egg. Egg which Countain IgY anti S. mutans was collected. IgY anti S. mutans extracted from egg yolk by mean s PEG-Amonium sulfat and purified using fast protein liquid chromatography. The purity of Igy anti S. mutans was determined by UV spectropometer. Biological activities of Igy anti S. mutans to inhibit adhession process was learned by anti adhesion test. We use two dose of IgY, which is 100 ug and 500 ug. Igy anti S. mutans formen in serum five weeks after the first immunization while it formed in egg nine weeks after the first immunization. Igy anti S. mutanss still present in serum andegg until twelve weeks from the first immunization. Igy anti S. mutanss could decrease the amount of bacteria which attach the epithelial cell surface. The amount of sticky bacteria on epithelial cell (without IgY are 40 cell bacteria/epithelial cell. After blocked by IgY anti S. mutanss the amount of bacteria turn into 30 cell bacteria/epithelial cell (for dose of 100 ug IgY and 28 cell bacteria/epitheelial cell (for dose of 500 ug IgY. This research concluded that hens were capable producing IgY anti S. mutanss in egg yolk and it can be used to solve dental caries problem which caused by S. mutanss.

  7. IgY antibodies in human nutrition for disease prevention.

    Science.gov (United States)

    Müller, Sandra; Schubert, Andreas; Zajac, Julia; Dyck, Terry; Oelkrug, Christopher

    2015-10-20

    Oral administration of preformed specific antibodies is an attractive approach against infections of the digestive system in humans and animals in times of increasing antibiotic resistances. Previous studies showed a positive effect of egg yolk IgY antibodies on bacterial intoxications in animals and humans. Immunization of chickens with specific antigens offers the possibility to create various forms of antibodies. Research shows that orally applied IgY's isolated from egg yolks can passively cure or prevent diseases of the digestive system. The use of these alternative therapeutic drugs provides further advantages: (1) The production of IgY's is a non-invasive alternative to current methods; (2) The keeping of chickens is inexpensive; (3) The animals are easy to handle; (4) It avoids repetitive bleeding of laboratory animals; (5) It is also very cost effective regarding the high IgY concentration within the egg yolk. Novel targets of these antigen specific antibodies are Helicobacter pylori and also molecules involved in signaling pathways in gastric cancer. Furthermore, also dental caries causing bacteria like Streptococcus mutans or opportunistic Pseudomonas aeruginosa in cystic fibrosis patients are possible targets. Therefore, IgY's included in food for human consumption may be able to prevent or cure human diseases.

  8. Chronobiological studies of chicken IgY: monitoring of infradian, circadian and ultradian rhythms of IgY in blood and yolk of chickens.

    Science.gov (United States)

    He, Jin-Xin; Thirumalai, Diraviyam; Schade, Rüdiger; Zhang, Xiao-Ying

    2014-08-15

    IgY is the functional equivalent of mammalian IgG found in birds, reptiles and amphibians. Many of its biological aspects have been explored with different approaches. In order to evaluate the rhythmicity of serum and yolk IgY, four chickens were examined and reared under the same conditions. To monitor biological oscillations of IgY in yolk and serum, the eggs and blood samples were collected over a 60 day period and the rhythm of yolk and serum IgY was determined by direct-ELISA. Results indicated that, there is a significant circaseptan rhythm in yolk IgY and circaquattran rhythm in serum IgY. The serum IgY concentration reached a peak in the morning, decreased to a minimum during the daytime and increased again at night revealing a significant circadian rhythm was superimposed by an ultradian rhythm. These data are suited to address the controversies concerning the IgY concentration in egg yolk and blood of laying hens. In addition, this study raised new questions, if the different rhythms in yolk and serum are concerned. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. Evaluation of the protective efficacy of immunoglobulin Y (IgY ...

    African Journals Online (AJOL)

    user

    2012-09-06

    Sep 6, 2012 ... IgY, which is transferred from serum to the yolk for ... The lethal potencies of the venom (LD50) were determined by i.p. injection of Swiss outbreed mice ... The protein concentration was measured by Biuret method and the. IgY-preparation ... Mean log10 antibody titer of the IgY- extracted by. Ammonium.

  10. Anti-Pseudomonas aeruginosa IgY antibodies augment bacterial clearance in a murine pneumonia model

    DEFF Research Database (Denmark)

    Thomsen, K.; Christophersen, L.; Bjarnsholt, T.

    2016-01-01

    Background: Oral prophylactic therapy by gargling with pathogen-specific egg yolk immunoglobulins (IgY) may reduce the initial airway colonization with Pseudomonas aeruginosa in cystic fibrosis (CF) patients. IgY antibodies impart passive immunization and we investigated the effects of anti......-P. aeruginosa IgY antibodies on bacterial eradication in a murine pneumonia model. Methods: P. aeruginosa pneumonia was established in Balb/c mice and the effects of prophylactic IgY administration on lung bacteriology, clinical parameters and subsequent inflammation were compared to controls. Results......: Prophylactic administration of IgY antibodies targeting P. aeruginosa significantly reduced the bacterial burden by 2-log 24 h post-infection compared to controls and was accompanied by significantly reduced clinical symptom scores and successive inflammatory cytokine profile indicative of diminished lung...

  11. PRODUKSI IGY SPESIFIK STAPHYLOCOCCUS AUREUS DARI ISOLAT ASAL KASUS STAPHYLOCOCCOSIS PADA KELINCI

    Directory of Open Access Journals (Sweden)

    Triwardhani Cahyaningsih

    2017-04-01

    Full Text Available This experiment aims to produce immunoglobulin Y (Ig-Y Staphylococcus aureus isolates origin staphylococcosis case in rabbits. Vaccination is done several times using Staphylococcus aureus active in 109 cfu/ml. Injecting the chicken is done four weeks in a row, the first week is done by intravenous injection of S. aureus antigens, the second week of the injection sub cutan S. aureus emulsified with Freund's complete adjuvant, followed by the third and fourth week of the injection of S. aureus antigen emulsified with incomplete Freund's adjuvant. Egg samples were taken after four weeks of the last injection for identification, purification, and determination of specific IgY against S. aureus in egg yolk. IgY specificity qualitatively tests performed by the AGP test (To Gel Precipitation. IgY extraction is done using PEG - Ammonium sulfate, purified IgY concentration calculated by the method of Bradford. The results showed that IgY began to be detected in egg yolk at week six after immunization, with the average levels of IgY is 1.7 mg/ml.

  12. Quantitative immunochemistry using IgG and IgY against plant ...

    African Journals Online (AJOL)

    Quantitative immunochemistry using IgG and IgY against plant hormones. ... both of animal origin, using chemical couplings to their respective functional groups. ... enzyme-linked immunosorbent assay; ELISA) to detect and quantify these ...

  13. Production and purification of IgY antibodies from chicken egg yolk

    Directory of Open Access Journals (Sweden)

    Wala Ahmad Amro

    2018-06-01

    Full Text Available The isolation of polyclonal antibodies from the serum of immunized mammals has significantly contributed to scientific research and diagnosis. The fact that recent technologies allow the production of antibodies in the yolk of eggs laid by hens, has led to the development of an alternative method for antibody generation that is less stressful to animals. As hens are kept under almost all their natural conditions, antibodies are isolated from the collected eggs; this technology is expected to become an interesting alternative to the conventionally serum-based techniques that eventually require to sacrifice the animal.Here we present a modified protocol for the isolation of IgY antibodies from immunized chickens and provide comparison between two chicken breeds in relative to IgY yield per egg. Our results show the possibility of generating large quantities of highly pure IgY from chicken eggs and also show large differences in the yield of IgY production between the two studied breeds. The results of this work indicate that IgY technology can be used for the production of primary antibodies for immunological work and disease diagnosis. Keywords: IgY, Chicken egg yolk, Gel filtration chromatography, Salmonella typhimurium

  14. Therapeutic potential of combined anti-IL-1β IgY and anti-TNF-α IgY in guinea pigs with allergic rhinitis induced by ovalbumin.

    Science.gov (United States)

    Guo-Zhu, Hu; Xi-Ling, Zhu; Zhu, Wen; Li-Hua, Wu; Dan, He; Xiao-Mu, Wu; Wen-Yun, Zhou; Wei-Xu, Hu

    2015-03-01

    We have previously demonstrated that anti-IL-1β immunoglobulin yolk(IgY) inhibits pathological responses in allergic asthma guinea pigs induced by ovalbumin(OVA). This study aims to determine whether the combined blockade of IL-1β and TNF-α can more effectively inhibit allergic inflammation in allergic rhinitis(AR) guinea pigs induced by OVA. Healthy guinea pigs treated with saline were used as the healthy control. The AR guinea pigs induced by OVA were randomly divided into (1) the AR model group containing negative control animals treated with intranasal saline; (2) the 0.1% non-specific IgY treatment group treated with non-specific IgY; (3) the 0.1% anti-TNF-α IgY treatment group treated with 0.1% anti-TNF-α IgY; (4) the 0.1% anti-IL-1β IgY treatment group treated with 0.1% anti-IL-1β IgY; (5) the 0.1% combined anti-IL-1β IgY and anti-TNF-α IgY treatment group treated with 0.1% combined anti-IL-1β IgY and anti-TNF-α IgY; and (6) the fluticasone propionate treatment group treated with fluticasone propionate. Cytokines were measured using an enzyme-linked immunosorbent assay. The results showed that IL-1β, IL-5, IL-9, IL-13, IL-18, IL-22, IL-33, TNF-α, TGF-β1 and OVA-specific IgE levels in the peripheral blood (PB) and nasal lavage fluid (NLF) significantly decreased at 2h, 4h or 8h in the 0.1% combined anti-IL-1β IgY and anti-TNF-α IgY treatment group compared to the AR model group and the 0.1% non-specific IgY treatment group (P<0.05). The data suggest that blockade of IL-1β and TNF-α by intranasal instillation of combined anti-IL-1β IgY and anti-TNF-α IgY could be a potential alternative strategy for preventing and treating allergic rhinitis. Copyright © 2014. Published by Elsevier B.V.

  15. Targeted radionuclide therapy with A 177Lu-labeled anti-HER2 nanobody.

    Science.gov (United States)

    D'Huyvetter, Matthias; Vincke, Cécile; Xavier, Catarina; Aerts, An; Impens, Nathalie; Baatout, Sarah; De Raeve, Hendrik; Muyldermans, Serge; Caveliers, Vicky; Devoogdt, Nick; Lahoutte, Tony

    2014-01-01

    RIT has become an attractive strategy in cancer treatment, but still faces important drawbacks due to poor tumor penetration and undesirable pharmacokinetics of the targeting vehicles. Smaller radiolabeled antibody fragments and peptides feature highly specific target accumulation, resulting in low accumulation in healthy tissue, except for the kidneys. Nanobodies are the smallest (MWnanobodies is predominantly dictated by the number of polar residues in the C-terminal amino acid tag. Three nanobodies were produced with different C-terminal amino-acid tag sequences (Myc-His-tagged, His-tagged, and untagged). Dynamic planar imaging of Wistar rats with 111In-DTPA-nanobodies revealed that untagged nanobodies showed a 70% drop in kidney accumulation compared to Myc-His-tagged nanobodies at 50 min p.i.. In addition, coinfusion of untagged nanobodies with the plasma expander Gelofusin led to a final reduction of 90%. Similar findings were obtained with different 177Lu-DTPA-2Rs15d nanobody constructs in HER2pos tumor xenografted mice at 1 h p.i.. Kidney accumulation decreased 88% when comparing Myc-His-tagged to untagged 2Rs15d nanobody, and 95% with a coinfusion of Gelofusin, without affecting the tumor targeting capacity. Consequently, we identified a generic method to reduce kidney retention of radiolabeled nanobodies. Dosimetry calculations of Gelofusin-coinfused, untagged 177Lu-DTPA-2Rs15d revealed a dose of 0.90 Gy/MBq that was delivered to both tumor and kidneys and extremely low doses to healthy tissues. In a comparative study, 177Lu-DTPA-Trastuzumab supplied 6 times more radiation to the tumor than untagged 177Lu-DTPA-2Rs15d, but concomitantly also a 155, 34, 80, 26 and 4180 fold higher radioactivity burden to lung, liver, spleen, bone and blood. Most importantly, nanobody-based targeted radionuclide therapy in mice bearing small estiblashed HER2pos tumors led to an almost complete blockade of tumor growth and a significant difference in event-free survival between the treated and the control groups (Pnanobody's C-terminus, as it has a predominant effect on kidney retention. Moreover, we show successful nanobody-based targeted radionuclide therapy in a xenograft model and highlight the potential of radiolabeled nanobodies as a valuable adjuvant therapy candidate for treatment of minimal residual and metastatic disease.

  16. Dual-Specificity Anti-HER-2/neu Antisense DNA Agents for Breast Cancer Therapy

    National Research Council Canada - National Science Library

    Stein, Stanley

    2001-01-01

    .... To achieve high avidity and specificity, we designed chimeric antisense molecules consisting of a short active DNA fused to a short "anchor" 2'-0-methyl RNA complementary to non-contiguous single...

  17. Interplay between Natural Killer Cells and Anti-HER2 Antibodies: Perspectives for Breast Cancer Immunotherapy

    Directory of Open Access Journals (Sweden)

    Aura Muntasell

    2017-11-01

    Full Text Available Overexpression of the human epidermal growth factor receptor 2 (HER2 defines a subgroup of breast tumors with aggressive behavior. The addition of HER2-targeted antibodies (i.e., trastuzumab, pertuzumab to chemotherapy significantly improves relapse-free and overall survival in patients with early-stage and advanced disease. Nonetheless, considerable proportions of patients develop resistance to treatment, highlighting the need for additional and co-adjuvant therapeutic strategies. HER2-specific antibodies can trigger natural killer (NK cell-mediated antibody-dependent cellular cytotoxicity and indirectly enhance the development of tumor-specific T cell immunity; both mechanisms contributing to their antitumor efficacy in preclinical models. Antibody-dependent NK cell activation results in the release of cytotoxic granules as well as the secretion of pro-inflammatory cytokines (i.e., IFNγ and TNFα and chemokines. Hence, NK cell tumor suppressive functions include direct cytolytic killing of tumor cells as well as the regulation of subsequent antitumor adaptive immunity. Albeit tumors with gene expression signatures associated to the presence of cytotoxic lymphocyte infiltrates benefit from trastuzumab-based treatment, NK cell-related biomarkers of response/resistance to HER2-specific therapeutic antibodies in breast cancer patients remain elusive. Several variables, including (i the configuration of the patient NK cell repertoire; (ii tumor molecular features (i.e., estrogen receptor expression; (iii concomitant therapeutic regimens (i.e., chemotherapeutic agents, tyrosine kinase inhibitors; and (iv evasion mechanisms developed by progressive breast tumors, have been shown to quantitatively and qualitatively influence antibody-triggered NK cell responses. In this review, we discuss possible interventions for restoring/enhancing the therapeutic activity of HER2 therapeutic antibodies by harnessing NK cell antitumor potential through combinatorial approaches, including immune checkpoint blocking/stimulatory antibodies, cytokines and toll-like receptor agonists.

  18. HER2 specific delivery of methotrexate by dendrimer conjugated anti-HER2 mAb

    International Nuclear Information System (INIS)

    Shukla, Rameshwer; Thomas, Thommey P; Desai, Ankur M; Kotlyar, Alina; Park, Steve J; Baker, James R Jr

    2008-01-01

    Herceptin, a humanized monoclonal antibody that binds to human growth factor receptor-2 (HER2), was covalently attached to a fifth-generation (G5) polyamidoamine dendrimer containing the cytotoxic drug methotrexate. The specific binding and internalization of this conjugate labeled with FITC was clearly demonstrated in cell lines overexpressing HER2 by flow cytometry as well as confocal microscopic analysis. In addition, binding and uptake of antibody conjugated dendrimers was completely blocked by excess non-conjugated herceptin. The dendrimer conjugate was also shown to inhibit the dihydrofolate reductase with similar activity to methotrexate. Co-localization experiments with lysotracker red indicate that antibody conjugate, although internalized efficiently into cells, has an unusually long residence time in the lysosome. Somewhat lower cytotoxicity of the conjugate in comparison to free methotrexate was attributed to the slow release of methotrexate from the conjugate and its long retention in the lysosomal pocket

  19. Development of paclitaxel-loaded liposomal systems with anti-her2 ...

    African Journals Online (AJOL)

    Methods: Seven different liposomal systems containing PTX, or MabHer2 or a combination of .... particle distribution via the polydispersity index ... at a constant voltage of 150 V for about 3h. ... using nano-drop spectrophotometer (BioSpec-.

  20. The complementarity-determining region sequences in IgY antivenom hypervariable regions

    Directory of Open Access Journals (Sweden)

    David Gitirana da Rocha

    2017-08-01

    Full Text Available The data presented in this article are related to the research article entitled "Development of IgY antibodies against anti-snake toxins endowed with highly lethal neutralizing activity" (da Rocha et al., 2017 [1]. Complementarity-determining region (CDR sequences are variable antibody (Ab sequences that respond with specificity, duration and strength to identify and bind to antigen (Ag epitopes. B lymphocytes isolated from hens immunized with Bitis arietans (Ba and anti-Crotalus durissus terrificus (Cdt venoms and expressing high specificity, affinity and toxicity neutralizing antibody titers were used as DNA sources. The VLF1, CDR1, CDR2, VLR1 and CDR3 sequences were validated by BLASTp, and values corresponding to IgY VL and VH anti-Ba or anti-Cdt venoms were identified, registered [Gallus gallus IgY Fv Light chain (GU815099/Gallus gallus IgY Fv Heavy chain (GU815098] and used for molecular modeling of IgY scFv anti-Ba. The resulting CDR1, CDR2 and CDR3 sequences were combined to construct the three - dimensional structure of the Ab paratope.

  1. Geomagnetism and Aeronomy activities in Italy during IGY, 1957/58

    Directory of Open Access Journals (Sweden)

    Lucilla Alfonsi

    2009-06-01

    Full Text Available In 2007 several events were organized to celebrate the fiftieth anniversary of the International Geophysical Year
    (IGY, 1957-1958. The celebrations will last until 2009 and are taking place within different contexts: the International
    Polar Year (IPY, the International Heliophysical Year (IHY, the electronic Geophysical Year (eGY
    and the International Year of Planet Earth (IYPE.
    IGY offered a very appropriate and timely occasion to undertake a series of coordinated observations of various
    geophysical phenomena all over the globe. Italy took part in the broad international effort stimulated by IGY. In
    fact, Italy participated in observations and studies in many of the proposed scientific areas, in particular Geomagnetism
    and Aeronomy. The Istituto Nazionale di Geofisica (ING started the installation of observatories,
    and updated and ensured continuous recording of geophysical observations. Geomagnetism, ionospheric
    physics, seismology, and other geophysical disciplines, were advanced. Although much of the work was undertaken
    in Italy, some attention was also devoted to other areas of the world, in particular Antarctica, where Italy
    participated in seismological observations. This paper gives a summary of the Geomagnetism and Ionospheric
    Physics activities within IGY. Furthermore, we highlight the importance of this historical event and its outcomes
    for the improvement of geophysical observations and the post-IGY growth of scientific investigations in Italy.

  2. IGI (the Italian Grid initiative) and its impact on the Astrophysics community

    Science.gov (United States)

    Pasian, F.; Vuerli, C.; Taffoni, G.

    IGI - the Association for the Italian Grid Infrastructure - has been established as a consortium of 14 different national institutions to provide long term sustainability to the Italian Grid. Its formal predecessor, the Grid.it project, has come to a close in 2006; to extend the benefits of this project, IGI has taken over and acts as the national coordinator for the different sectors of the Italian e-Infrastructure present in EGEE. IGI plans to support activities in a vast range of scientificdisciplines - e.g. Physics, Astrophysics, Biology, Health, Chemistry, Geophysics, Economy, Finance - and any possible extensions to other sectors such as Civil Protection, e-Learning, dissemination in Universities and secondary schools. Among these, the Astrophysics community is active as a user, by porting applications of various kinds, but also as a resource provider in terms of computing power and storage, and as middleware developer.

  3. Produksi IgY Antivirus Avian Influenza H5N1 dan Prospek Pemanfaatannya dalam Pengebalan Pasif

    Directory of Open Access Journals (Sweden)

    I Wayan Teguh Wibawan

    2009-09-01

    Full Text Available Immunoglobulin Y (IgY in yolk has been shown in several studies to prevent both bacterial and viralinfections. This research was conducted to find evidence that IgY specific against avian influenza virus(AIV of H5N1 subtype can be produced in a large quantity in egg yolk. Laying hens were vaccinated withAI killed-vaccine (IPB-Shigeta. The IgY was purified using affinity chromatograpy technique, and anti-H5activity was measured using a standard haemaglutination inhibition (HI and agar gel immunodifusion.The concentration of IgY was calculated, and the protein pattern was detected using polyacrilamid gel(AGID electrophoresis (PAGE. Anti H5 antibody as high as 27 – 29 HI units was detected and produce aspecific line of precipitation in AGID. The concentration of IgY was 7.89 mg/ml. Purified specific IgY consistof 6 main protein bands with molecular weights ranging from 35 to 225 kD. These proteins were sensitiveto heat treatment (75oC for 30 minutes, to acid condition (pH2 as well as the pepsin and trypsin. Theseresults indicated the possibility of using specific IgY for passive immunisation to prevent AIV infection oras immunotherapeutic applications for AI treatment in humans.

  4. The potency of chicken egg yolk immunoglobulin (IgY specific as immunotherapy to Mycobacterium tuberculosis infection

    Directory of Open Access Journals (Sweden)

    Sri Agus Sudjarwo

    2017-01-01

    Full Text Available The aim of this study was to characterize of chicken egg yolk immunoglobulins (IgYs specific as immunotherapy to Mycobacterium tuberculosis complex (MTBC infection. Lohmann laying hens were immunized intramuscularly with antigenic of MTBC. Egg yolk was separated from egg white, and IgY antibody was then purified by multiple polyethylene glycols 6000 extraction and ammonium sulfate purification steps. The IgY anti-MTBC concentration in egg yolk increased at 2 weeks and it reached a maximum at 4 weeks after immunization. After 6 weeks, the levels of IgY anti-MTBC decreased gradually. The antibody of MTBC was detected and produces a specific line of precipitation in agar gel precipitation test beginning the week 2 after the first immunization. Analysis of results obtained with ELISA showed a significant increase in the MTBC specific antibodies after 2 weeks and reached a plateau at 4 weeks from the booster immunization. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed the IgY preparation to be pure and dissociated into protein bands with molecular weights of 112, 78, 69, 49, and 28 kDa and Western blot analysis shown the presence of anti-MTBC IgY in egg yolks, with molecular weights of approximately 78 kDa. These results suggested that egg yolk could be a practical strategy in large-scale production of specific anti-MTBC IgY for immunotherapy of TBC.

  5. The ability of IgY to recognize surface proteins of Streptococcus mutans

    Directory of Open Access Journals (Sweden)

    Basri A. Gani

    2009-12-01

    Full Text Available Background: Streptococcus mutans are gram positive bacteria classified into viridians group, and have a role in pathogenesis of dental caries. It’s adhesion to the tooth surface is mediated by cell surface proteins, which interact with specific receptor located in tooth pellicle. Glucan binding protein, Glukosyltransferase, and antigen I/II are basic proteins of S. mutans, which have a role in initiating the interaction. A previous study showed that chicken’s IgY can interfere the interaction. Purpose: The objective of this study was to assess the ability of IgY in recognizing the surface molecule of Streptococcus mutans expressed by various serotypes (c, d, e, f and a strain derived from IPB, Bogor. Method: Western blot was used as a method to determine such capability. Result: The result showed that IgY has a potency to recognize antigen I/II, but not the other proteins on the cell surface of all bacteria tested. Conclusion: The ability of IgY to bind the surface protein, antigen I/II, indicates that this avian antibody could be used as a candidate for anti-adhesion in preventing dental caries.

  6. Evidence of IgY subclass diversification in snakes: evolutionary implications.

    Science.gov (United States)

    Wang, Tao; Sun, Yi; Shao, Wenwei; Cheng, Gang; Li, Lingxiao; Cao, Zubing; Yang, Zhi; Zou, Huiying; Zhang, Wei; Han, Binyue; Hu, Yang; Ren, Liming; Hu, Xiaoxiang; Guo, Ying; Fei, Jing; Hammarström, Lennart; Li, Ning; Zhao, Yaofeng

    2012-10-01

    Mammalian IgG and IgE are thought to have evolved from IgY of nonmammalian tetrapods; however, no diversification of IgY subclasses has been reported in reptiles or birds, which are phylogenetically close to mammals. To our knowledge, we report the first evidence of the presence of multiple IgY-encoding (υ) genes in snakes. Two υ genes were identified in the snake Elaphe taeniura, and three υ genes were identified in the Burmese python (Python molurus bivittatus). Although four of the υ genes displayed a conventional four-H chain C region exon structure, one of the υ genes in the Burmese python lacked the H chain C region 2 exon, thus exhibiting a structure similar to that of the mammalian γ genes. We developed mouse mAbs specific for the IgY1 and IgY2 of E. taeniura and showed that both were expressed in serum; each had two isoforms: one full-length and one truncated at the C terminus. The truncation was not caused by alternative splicing or transcriptional termination. We also identified the μ and δ genes, but no α gene, in both snakes. This study provides valuable clues for our understanding of Ig gene evolution in tetrapods.

  7. IgY Technology: Extraction of Chicken Antibodies from Egg Yolk by Polyethylene Glycol (PEG) Precipitation

    Science.gov (United States)

    Pauly, Diana; Chacana, Pablo A.; Calzado, Esteban G.; Brembs, Björn; Schade, Rüdiger

    2011-01-01

    Hens can be immunized by means of i.m. vaccination (Musculus pectoralis, left and right, injection volume 0.5-1.0 ml) or by means of Gene-Gun plasmid-immunization. Dependent on the immunogenicity of the antigen, high antibody-titres (up to 1:100,000 - 1:1,000,000) can be achieved after only one or 3 - 4 boost immunizations. Normally, a hen lays eggs continuously for about 72 weeks, thereafter the laying capacity decreases. This protocol describes the extraction of total IgY from egg yolk by means of a precipitation procedure (PEG. Polson et al. 1980). The method involves two important steps. The first one is the removal of lipids and the second is the precipitation of total IgY from the supernatant of step one. After dialysis against a buffer (normally PBS) the IgY-extract can be stored at -20°C for more than a year. The purity of the extract is around 80 %, the total IgY per egg varies from 40-80 mg, dependent on the age of the laying hen. The total IgY content increases with the age of the hen from around 40 mg/egg up to 80 mg/egg (concerning PEG precipitation). The laying capacity of a hen per year is around 325 eggs. That means a total potential harvest of 20 g total IgY/year based on a mean IgY content of 60 mg total IgY/egg (see Table 1). PMID:21559009

  8. [Clinical study on the effect of anti-gingivitis IgY toothpaste in control of gingivitis and dental plaque].

    Science.gov (United States)

    Zhang, Wei; Feng, Xi-Ping; Tao, Dan-Ying; Chen, Jian-Fen

    2016-08-01

    To observe the effect of anti-gingivitis IgY toothpaste in control of gingivitis and plaque. The study was a double-blind, randomized, parallel-controlled clinical trail with a total of 100 subjects who were divided into two groups, experimental group and control group. The subjects in experimental group used anti-gingivitis IgY toothpaste to brush twice daily for 3 minutes, and the subjects in control group used none anti-gingivitis IgY toothpaste. The examiner recorded GI, PI and BOP index of all subjects at the baseline, 6-weeks and 12-weeks. SPSS21.0 software package was used for statistical analysis. Twelve weeks later, there were significant differences in GI and BOP between the two groups. Yet no significant difference was found in PI. Anti-gingivitis IgY toothpaste is effective in control of gingivitis.

  9. Aktivitas Biologis Imunoglobulin Yolk Anti Parvovirus Setelah Perlakuan Suhu (BIOLOGY ACTIVITIES OF IgY PARVOVIRUS AFTER HEAT TREATMENT

    Directory of Open Access Journals (Sweden)

    I Gusti Ayu Agung Suartini

    2016-02-01

    Full Text Available This study aims to determine the effect of temperature on the biological activity of various crude and precipitate specific Immunoglobulin (IgY Canine parvovirus (CPV. Hiperimun serum conducted on Isa Brown chickens injected with antigen CPV. Crude yolk Ig preparations derived from chicken serum without purification while the yolk Ig preparations precipitates obtained by the chicken serum was precipitated with ammonium sulfate and dialyzed. Both types of Ig yolk given treatment temperature 50ºC, 60ºC, 70ºC, and 80ºC for 15 minutes. To test Gel Precipitation Test (AGPT is performed to determine whether there is a specific IgY CPV in the serum of chickens. Biological activity of both types of Ig detected with Barriers Haemagglutination test (HI. The design used in this study is completely randomized design factorial. The results of this study indicate that the temperature was highly significant on the activities of IgY crude and precipitates. Activities IgY crude and precipitate down to the treatment temperature of 50ºC, 60ºC, 70ºC, and 80ºC. Geometric Mean Titer crude IgY respectively - were 26.67, 26, 25.33, and <2º Unit HI while IgY precipitates are respectively 26.33, 25.67, 24, and <2º Unit HI. Based on the results of this study concluded that the biological activity of crude IgY better than IgY precipitates after treatment of a wide range of temperatures.

  10. Protective effect of an egg yolk-derived immunoglobulin (IgY) against Prevotella intermedia-mediated gingivitis.

    Science.gov (United States)

    Hou, Y-Y; Zhen, Y-H; Wang, D; Zhu, J; Sun, D-X; Liu, X-T; Wang, H-X; Liu, Y; Long, Y-Y; Shu, X-H

    2014-04-01

    To investigate the effects of an egg yolk-derived immunoglobulin (IgY) specific to Prevotella intermedia in vitro and in vivo. An IgY specific to P. intermedia was produced by immunizing hens with formaldehyde-inactivated P. intermedia and showed high titres when subjected to an ELISA. The obtained IgY inhibited the growth of P. intermedia in a dose-dependent manner at concentrations from 1 to 20 mg ml(-1) in Center for Disease Control and Prevention liquid medium. Forty rats were challenged with P. intermedia on gingivae and then randomly divided into four groups, which were syringed respectively with phosphate-buffered saline, 1 mg ml(-1) of tinidazole, 20 mg ml(-1) of nonspecific IgY and 20 mg ml(-1) of the IgY specific to P. intermedia at a dosage of 300 μl per day. Gingival index (GI), plaque index (PI), bleeding on probing (BOP), counts of white blood cell (WBC) and histopathological slide of the gums were measured after treatment for 15 days. The gingivitis rats treated with the IgY specific to P. intermedia showed significantly decreased GI, PI, BOP and WBC (P intermedia-mediated gingivitis. A new immunoglobulin specific to P. intermedia was developed from egg yolk. This specific IgY can dose-dependently inhibit the growth of P. intermedia and protect rats from gingivitis induced by P. intermedia. The new IgY has potential for the treatment of P. intermedia-mediated gingivitis. © 2013 The Society for Applied Microbiology.

  11. Chicken egg yolk antibodies (IgY) as non-antibiotic production enhancers for use in swine production: a review

    OpenAIRE

    Li, Xiaoyu; Wang, Lili; Zhen, Yuhong; Li, Shuying; Xu, Yongping

    2015-01-01

    In recent years, the use of in-feed antibiotics for growth and disease prevention in livestock production has been under severe scrutiny. The use and misuse of in-feed antibiotics has led to problems with drug residues in animal products and increased bacterial resistance. Chicken egg yolk antibodies (IgY) have attracted considerable attention as an alternative to antibiotics to maintain swine health and performance. Oral administration of IgY possesses many advantages over mammalian IgG such...

  12. Protective effects of chicken egg yolk antibody (IgY) against experimental Vibrio splendidus infection in the sea cucumber (Apostichopus japonicus).

    Science.gov (United States)

    Li, Xiaoyu; Jing, Kailin; Wang, Xitao; Li, Yuan; Zhang, Meixia; Li, Zhen; Xu, Le; Wang, Lili; Xu, Yongping

    2016-01-01

    Vibrio splendidus is one of the most harmful pathogens associated with skin ulceration syndrome in the sea cucumber (Apostichopus japonicus) due to its high virulence and frequency of appearance. The objective of this study was to determine the effectiveness of chicken egg yolk antibody (IgY) against V. splendidus infection in the sea cucumber. Whole V. splendidus cells were used as an immunogen to immunize 20 White Leghorn hens (25 weeks old). IgY was produced from egg yolks obtained from these immunized hens using water dilution, two-step salt precipitation and ultrafiltration. The purity of the IgY produced was approximately 83%. Enzyme-linked Immunosorbent Assay indicated a high specificity for IgY with a maximum antibody titer of 320,000. The growth of V. splendidus in liquid medium was significantly inhibited by IgY in a dose-dependent manner at concentrations ranging from 1 to 10 mg/mL. The protective effects of IgY were evaluated in sea cucumber by intraperitoneally injecting anti-V. splendidus IgY antibodies (10 mg/mL) or immersing the sea cucumber in aqueous IgY (1 g/L) after an intraperitoneal injection of V. splendidus. Intraperitoneal injection resulted in an 80% survival while immersion resulted in a 75% survival during the 11-day experimental period. The survival rates were significantly higher than the positive control and the non-specific IgY group (P sea cucumber treated with specific IgY than those treated with non-specific IgY. The phagocytosis of coelomocytes for V. splendidus in the presence of specific IgY was significantly (P sea cucumbers against V. splendidus infection. Copyright © 2015 Elsevier Ltd. All rights reserved.

  13. Effect of Chicken Egg Yolk Antibodies (IgY) against Diarrhea in Domesticated Animals: A Systematic Review and Meta-Analysis

    OpenAIRE

    Diraviyam, Thirumalai; Zhao, Bin; Wang, Yuan; Schade, Ruediger; Michael, Antonysamy; Zhang, Xiaoying

    2014-01-01

    Background IgY antibodies are serum immunoglobulin in birds, reptiles and amphibians, and are transferred from serum to egg yolk to confer passive immunity to their embryos and offspring. Currently, the oral passive immunization using chicken IgY has been focused as an alternative to antibiotics for the treatment and control of diarrhea in animals and humans. This systematic review was focused to determine the effect of IgY in controlling and preventing diarrhea in domesticated animals includ...

  14. Research on the Earth's Interior Conducted by Russia after IGY: The Geotraverse Project and "Intermargins"

    Directory of Open Access Journals (Sweden)

    A G Rodnikov

    2009-04-01

    Full Text Available Fifty years have passed since the International Geophysical Year (IGY of 1957.58, one of the most important and noble initiatives in the history of science and in the history of humanity in general. IGY became the model for subsequent international scientific initiatives in various fields of solid Earth research, including the Upper Mantle Project (1961.71, the Geodynamic Project (1971.80, the Geotraverse Project (1987.2003, and the "InterMARGINS" Project (2003. The Russian investigations as part of the Geotraverse Project and "InterMARGINS" were aimed at research into the deep structure of the continental margins of East Eurasia, which are characterized by high seismicity, volcanism, and natural cataclysms hazardous to people living there.

  15. IgY antibodies protect against human Rotavirus induced diarrhea in the neonatal gnotobiotic piglet disease model.

    Directory of Open Access Journals (Sweden)

    Celina G Vega

    Full Text Available Group A Rotaviruses are the most common cause of severe, dehydrating diarrhea in children worldwide. The aim of the present work was to evaluate protection against rotavirus (RV diarrhea conferred by the prophylactic administration of specific IgY antibodies (Ab to gnotobiotic piglets experimentally inoculated with virulent Wa G1P[8] human rotavirus (HRV. Chicken egg yolk IgY Ab generated from Wa HRV hyperimmunized hens specifically recognized (ELISA and neutralized Wa HRV in vitro. Supplementation of the RV Ab free cow milk diet with Wa HRV-specific egg yolk IgY Ab at a final ELISA Ab titer of 4096 (virus neutralization -VN- titer = 256 for 9 days conferred full protection against Wa HRV associated diarrhea and significantly reduced virus shedding. This protection was dose-dependent. The oral administration of semi-purified passive IgY Abs from chickens did not affect the isotype profile of the pig Ab secreting cell (ASC responses to Wa HRV infection, but it was associated with significantly fewer numbers of HRV-specific IgA ASC in the duodenum. We further analyzed the pigś immune responses to the passive IgY treatment. The oral administration of IgY Abs induced IgG Ab responses to chicken IgY in serum and local IgA and IgG Ab responses to IgY in the intestinal contents of neonatal piglets in a dose dependent manner. To our knowledge, this is the first study to show that IgY Abs administered orally as a milk supplement passively protect neonatal pigs against an enteric viral pathogen (HRV. Piglets are an animal model with a gastrointestinal physiology and an immune system that closely mimic human infants. This strategy can be scaled-up to inexpensively produce large amounts of polyclonal IgY Abs from egg yolks to be applied as a preventive and therapeutic passive Ab treatment to control RV diarrhea.

  16. Development and immunochemical evaluation of a novel chicken IgY antibody specific for KLK6

    Directory of Open Access Journals (Sweden)

    Sotiropoulou Georgia

    2012-12-01

    Full Text Available Abstract Background Human kallikrein-related peptidase 6 (KLK6 has been implicated in various types of cancer and in neurodegenerative and demyelinating diseases including multiple sclerosis. Further, anti-KLK6 antibodies attenuated disease manifestations in the mouse model of multiple sclerosis. Availability of specific antibodies against KLK6 is fundamental to the development of improved diagnostic and/or immunotherapeutic applications. Here, we exploited the enhanced immunogenicity of mammalian proteins in avian species to generate a polyclonal antibody against KLK6. Results Chicken were immunized with recombinant KLK6 and antibodies Y (IgYs were purified from egg yolk with a simple procedure and evaluated for KLK6 detection by ELISA and Western blot using recombinant proteins and human cell lysates and supernatants. The anti-KLK6 Y polyclonal exhibited high affinity for KLK6 with a detection limit of 30 fmol. On the other hand, the widely used rabbit polyclonal antibody that was raised against the same recombinant KLK6 had a detection limit of 300 fmol. Moreover, the IgYs did not display any crossreactivity with recombinant KLKs or endogenous KLKs and other cellular proteins. Conclusions Based on its high specificity and sensitivity the developed anti-KLK6 IgY is expected to aid the development of improved diagnostic tools for the detection of KLK6 in biological and clinical samples.

  17. Production of IgY by layers injected with Salmonella typhimurium

    Directory of Open Access Journals (Sweden)

    Hee, H. L.

    2017-06-01

    Full Text Available In the present study, production of IgY and performance of layers immunised with Salmonella typhimurium were evaluated. Thirty 23-wk old layers (Lohmann Brown with average body weight of 1.6 � 0.03 kg. were randomly and equally divided into two groups. Both groups were placed in a specific pathogen free room equipped with an exhaust fan. The layers were placed in single battery cages with slightly tilted floor which allowed eggs to roll straight into the collecting tray. Lyophilized, attenuated whole cells of S. typhimurium strain [serotype: Somatic (O, antigen 1, 4, (5, 12, flagella (H N12 phase I: 1; phase II: 1, 2 (Group B] that were emulsified with Freund�s complete adjuvant were used to immunise the layers intramuscularly 3 times at 2-wk interval (treatment group. Layers in the control group were injected with a solution of sterilized phosphate-buffered saline (PBS emulsified with an equal volume of Freund�s complete adjuvant. The experimental period lasted for 14 wks. Specific antibodies were detected by enzyme-linked immunosorbent assay. The results showed that the birds lost weight during the injection period, but the weights slowly increased in the following weeks. However, the weight of layers in the treated group remained lower (P<0.05 when compared to the period before injection. The loss in body weight of layers was the result of reduced feed intake, in particular during the injection weeks. Layers in the control group had a higher egg production, by about 10.1% than layers in the treatment group throughout the experimental period. The egg production for layers in treatment group showed a significant decrease (P<0.05 during the immunization period by 28.8%. The egg production significantly increased (P<0.05 to 71.9% after this period, but the production did not reach the level before immunisation. The weight of eggs of treated layers were significantly lower (P<0.05 than the control layers during the injection weeks. The egg

  18. Moving Beyond IGY: An Electronic Geophysical Year (eGY) Concept

    Science.gov (United States)

    Baker, D. N.; Barton, C. E.; Rodger, A. S.; Thompson, B. J.; Fraser, B.; Papitashvili, V.

    2003-12-01

    During the International Geophysical Year (1957-1958), member countries established many new geophysical observatories pursuing the major IGY objectives - to collect geophysical data as widely as possible and to provide free access to these data for all scientists around the globe. Today, geophysics has attained a rather good understanding within traditional regions, i.e., the atmosphere, ionosphere, magnetosphere, and other such geospheres. At the same time, it has become clear that much of the new and important science is coming from the studies of interfaces and coupling between geospheres. Thus, if geophysical data are made `'transparently'' available to a much wider range of scientists and students than to those who do the observations, then new and exciting discoveries can be expected. An International Association of Geomagnetic and Aeronomy (IAGA) task force, recognizing that a key achievement of the IGY was the establishment of a worldwide system of data centers and physical observatories, proposes that for the 50th anniversary of IGY, the worldwide scientific community should endorse and promote an electronic Geophysical Year (eGY) initiative. The proposed eGY concept would both commemorate the IGY in 2007-2008 and provide a forward impetus to geophysics in 21st century, similar to that provided by the IGY fifty years ago. The IAGA task force strongly advocates: (1) Securing permission and release of existing data; (2) Creating access to information; and (3) Conversion of relevant analog data to digital form. The eGY concept embraces all available and upcoming geophysical data (e.g., atmospheric, ionospheric, geomagnetic, gravity, etc.) through the establishment of a series of virtual geophysical observatories now being `'deployed'' in cyberspace. The eGY concept is modern, global, and timely; it is attractive, pragmatic, and affordable. The eGY is based on the existing and continually developing computing/networking technologies (e.g., XML, Semantic Web

  19. Chicken IgY Fc expressed by Eimeria mitis enhances the immunogenicity of E. mitis.

    Science.gov (United States)

    Qin, Mei; Tang, Xinming; Yin, Guangwen; Liu, Xianyong; Suo, Jingxia; Tao, Geru; Ei-Ashram, Saeed; Li, Yuan; Suo, Xun

    2016-03-21

    Eimeria species are obligate intracellular apicomplexan parasites, causing great economic losses in the poultry industry. Currently wild-and attenuated- type anticoccidial vaccines are used to control coccidiosis. However, their use in fast growing broilers is limited by vaccination side effects caused by medium and/or low immunogenic Eimeria spp. There is, therefore, a need for a vaccine with high immunogenicity for broilers. The avian yolk sac IgY Fc is the avian counterpart of the mammalian IgG Fc, which enhances immunogenicity of Fc-fusion proteins. Here, we developed a stable transgenic Eimeria mitis expressing IgY Fc (Emi.chFc) and investigated whether the avian IgY Fc fragment enhances the immunogenicity of E. mitis. Two-week-old broilers were immunized with either Emi.chFc or wild type Eimeria and challenged with wild type E. mitis to analyze the protective properties of transgenic Emi.chFc. Chickens immunized with Emi.chFc had significantly lower oocyst output, in comparison with PBS, mock control (transgenic E. mitis expressing HA1 from H9N2 avian influenza virus) and wildtype E. mitis immunized groups after challenge, indicating that IgY Fc enhanced the immunogenicity of E. mitis. Our findings suggest that IgY Fc-expressing Eimeria may be a better coccidiosis vaccine, and transgenic Eimeria expressing Fc-fused exogenous antigens may be used as a novel vaccine-delivery vehicle against a wide variety of pathogens.

  20. I.G.Y. Ascaplots annals of the international geophysical year, v.20

    CERN Document Server

    Stoffregen, W

    Annals of the International Geophysical Year, Volume 20, Part II: I.G.Y. Ascaplots is a four-chapter text that provides the data on half-hourly auroral all-sky camera plots from 115 stations for the period 1958-1959. This period cover two winters in the northern hemisphere characterized by high auroral activity. This part also presents the list of stations, as well as the maps of the northern and southern distribution of all-sky cameras, with some modifications and additions to the earlier list. Data from the added Japanese station in the Antarctic are received and are included with the data

  1. IgY Technology: Extraction of Chicken Antibodies from Egg Yolk by Polyethylene Glycol (PEG) Precipitation

    OpenAIRE

    Pauly, Diana; Chacana, Pablo A.; Calzado, Esteban G.; Brembs, Bj?rn; Schade, R?diger

    2011-01-01

    Hens can be immunized by means of i.m. vaccination (Musculus pectoralis, left and right, injection volume 0.5-1.0 ml) or by means of Gene-Gun plasmid-immunization. Dependent on the immunogenicity of the antigen, high antibody-titres (up to 1:100,000 - 1:1,000,000) can be achieved after only one or 3 - 4 boost immunizations. Normally, a hen lays eggs continuously for about 72 weeks, thereafter the laying capacity decreases. This protocol describes the extraction of total IgY from egg yolk b...

  2. Effect of passive immunization by anti-gingipain IgY on periodontal health of dogs

    Directory of Open Access Journals (Sweden)

    Rahman A.K.M. Shofiqur

    2011-09-01

    Full Text Available Anti-gingipain IgY (IgY-GP, known as hyperimmune γ-livetin from egg yolk, inhibits the enzyme activity, growth and adherence of Porphyromonas gingivalis to gingival epithelial cells. Our objective was to evaluate the efficacy of IgY-GP on periodontal health of dogs. IgY-GP was prepared from the egg yolk of hens immunized with the gingipain from Porphyromonas gingivalis ATCC 33277. Two in vivo trial models were conducted on 15 adult dogs with periodontitis by giving IgY-GP-supplemented dog feed for 8 weeks and direct application of the IgY in dental ointment to the periodontal pockets at weekly interval for 4 weeks. Clinical parameters including gingivitis, periodontitis, oral health index, bleeding on probe (BOP, pocket depth (PD, and dental calculus removal pattern for selected premolar teeth were recorded at baseline, 4 and 8 weeks post treatment.IgY-GP showed strong cross-reactivity with gingipain from Porphyromonas gulae and inhibited the enzyme activity in vitro. In the dog trials, IgY-GP resulted in significant improvement of oral health parameters including gingivitis and periodontitis scores, BOP, dental calculus removal. No adverse events during and after antibody applications were noted. Oral immunotherapy by using IgY-GP is a new promising alternative to conventional preventive and therapeutic methods to improve oral health status in dogs.

  3. Effect of chicken egg yolk antibodies (IgY against diarrhea in domesticated animals: a systematic review and meta-analysis.

    Directory of Open Access Journals (Sweden)

    Thirumalai Diraviyam

    Full Text Available BACKGROUND: IgY antibodies are serum immunoglobulin in birds, reptiles and amphibians, and are transferred from serum to egg yolk to confer passive immunity to their embryos and offspring. Currently, the oral passive immunization using chicken IgY has been focused as an alternative to antibiotics for the treatment and control of diarrhea in animals and humans. This systematic review was focused to determine the effect of IgY in controlling and preventing diarrhea in domesticated animals including Piglets, Mice, Poultry and Calves. METHODS AND RESULTS: Previous research reports focused on treatment effect of Chicken IgY against diarrhea were retrieved from different electronic data bases (MEDLINE, EMBASE, SPRINGER-LINK, WILEY, AGRICOLA, MEDWELL Journals, Scientific Publish, Chinese articles from Core periodicals in 2012. A total of 61 studies in 4 different animal classes met the inclusion criteria. Data on study characteristics and outcome measures were extracted. The pooled relative risk (RR of 49 studies of different animals [Piglets--22; Mice--14; Poultry--7 and Calves--6] in meta-analyses revealed that, IgY significantly reduced the risk of diarrhea in treatment group when compare to the placebo. However, the 95% confidence intervals of the majority of studies in animal class piglets and calves embrace RR of one. The same results were obtained in sub group analyses (treatment regiment--prophylactic or therapeutic; pathogen type--bacterial or viral. Perhaps, this inconsistency in the effect of IgY at the individual study level and overall effect measures could be influenced by the methodological heterogeneity. CONCLUSION: The present systematic review (SR and meta-analysis demonstrated the beneficial effect of IgY. This supports the opinion that IgY is useful for prophylaxis and treatment. However, more intensive studies using the gold standard animal experiments with the focus to use IgY alone or in combination with other alternative

  4. Effect of Chicken Egg Yolk Antibodies (IgY) against Diarrhea in Domesticated Animals: A Systematic Review and Meta-Analysis

    Science.gov (United States)

    Diraviyam, Thirumalai; Zhao, Bin; Wang, Yuan; Schade, Ruediger; Michael, Antonysamy; Zhang, Xiaoying

    2014-01-01

    Background IgY antibodies are serum immunoglobulin in birds, reptiles and amphibians, and are transferred from serum to egg yolk to confer passive immunity to their embryos and offspring. Currently, the oral passive immunization using chicken IgY has been focused as an alternative to antibiotics for the treatment and control of diarrhea in animals and humans. This systematic review was focused to determine the effect of IgY in controlling and preventing diarrhea in domesticated animals including Piglets, Mice, Poultry and Calves. Methods and Results Previous research reports focused on treatment effect of Chicken IgY against diarrhea were retrieved from different electronic data bases (MEDLINE, EMBASE, SPRINGER-LINK, WILEY, AGRICOLA, MEDWELL Journals, Scientific Publish, Chinese articles from Core periodicals in 2012). A total of 61 studies in 4 different animal classes met the inclusion criteria. Data on study characteristics and outcome measures were extracted. The pooled relative risk (RR) of 49 studies of different animals [Piglets – 22; Mice – 14; Poultry – 7 and Calves – 6] in meta-analyses revealed that, IgY significantly reduced the risk of diarrhea in treatment group when compare to the placebo. However, the 95% confidence intervals of the majority of studies in animal class piglets and calves embrace RR of one. The same results were obtained in sub group analyses (treatment regiment – prophylactic or therapeutic; pathogen type – bacterial or viral). Perhaps, this inconsistency in the effect of IgY at the individual study level and overall effect measures could be influenced by the methodological heterogeneity. Conclusion The present systematic review (SR) and meta-analysis demonstrated the beneficial effect of IgY. This supports the opinion that IgY is useful for prophylaxis and treatment. However, more intensive studies using the gold standard animal experiments with the focus to use IgY alone or in combination with other alternative

  5. Long-term hazard of recurrence in HER2+ breast cancer patients untreated with anti-HER2 therapy

    DEFF Research Database (Denmark)

    Strasser-Weippl, Kathrin; Horick, Nora; Smith, Ian E

    2015-01-01

    INTRODUCTION: Worldwide, many patients with HER2+ (human epidermal growth factor receptor 2-positive) early breast cancer (BC) do not receive adjuvant trastuzumab. Hazards of recurrence of these patients with respect to hormone receptor status of the primary tumor have not been described. METHODS...

  6. Targeted delivery of doxorubicin-utilizing chitosan nanoparticles surface-functionalized with anti-Her2 trastuzumab

    Directory of Open Access Journals (Sweden)

    Yousefpour P

    2011-09-01

    Full Text Available Parisa Yousefpour1, Fatemeh Atyabi2, Ebrahim Vasheghani-Farahani3, Ali-Akbar Mousavi Movahedi1, Rassoul Dinarvand21Department of Biotechnology, Faculty of Science, University of Tehran, 2Nanotechnology Research Centre, Faculty of Pharmacy, Tehran University of Medical Sciences, 3Biotechnology Group, Department of Chemical Engineering, Faculty of Engineering, Tarbiat Modares University, Tehran, IranBackground: Targeting drugs to their sites of action to overcome the systemic side effects associated with most antineoplastic agents is still a major challenge in pharmaceutical research. In this study, the monoclonal antibody, trastuzumab, was used as a targeting agent in nanoparticles carrying the antitumor drug, doxorubicin, specifically to its site of action.Methods: Chitosan-doxorubicin conjugation was carried out using succinic anhydride as a crosslinker. Trastuzumab was conjugated to self-assembled chitosan-doxorubin conjugate (CS-DOX nanoparticles (particle size, 200 nm via thiolation of lysine residues and subsequent linking of the resulted thiols to chitosan. Conjugation was confirmed by gel permeation chromatography, differential scanning calorimetry, Fourier transform infrared spectroscopy, and 1H nuclear magnetic resonance spectroscopy studies. Dynamic light scattering, transmission electron microscopy, and zeta potential determination were used to characterize the nanoparticles.Results: CS-DOX conjugated nanoparticles had a spherical shape and smooth surface with a narrow size distribution and core-shell structure. Increasing the ratio of doxorubicin to chitosan in the conjugation reaction gave rise to a higher doxorubicin content but lower conjugation efficiency. Trastuzumab-decorated nanoparticles (CS-DOX-mAb contained 47 µg/mg doxorubicin and 33.5 µg/mg trastuzumab. Binding of trastuzumab to the nanoparticles was further probed thermodynamically by isothermal titration calorimetry. Fluorescence microscopy demonstrated enhanced and selective uptake of CS-DOX-mAb by Her2+ cancer cells compared with nontargeted CS-DOX nanoparticles and free drug.Conclusion: Antibody-conjugated nanoparticles were shown to discriminate between Her2+ and Her2- cells, and thus have the potential to be used in active targeted drug delivery, with reduction of drug side effects in Her2+ breast and ovarian cancers.Keywords: chitosan, doxorubicin, self-assembled nanoparticles, active targeting, trastuzumab

  7. Antiparasitic effects induced by polyclonal IgY antibodies anti-phospholipase A2 from Bothrops pauloensis venom.

    Science.gov (United States)

    Borges, Isabela Pacheco; Silva, Mariana Ferreira; Santiago, Fernanda Maria; de Faria, Lucas Silva; Júnior, Álvaro Ferreira; da Silva, Rafaela José; Costa, Mônica Soares; de Freitas, Vitor; Yoneyama, Kelly Aparecida Geraldo; Ferro, Eloísa Amália Vieira; Lopes, Daiana Silva; Rodrigues, Renata Santos; de Melo Rodrigues, Veridiana

    2018-06-01

    Activities of phospholipases (PLAs) have been linked to pathogenesis in various microorganisms, and implicated in cell invasion and so the interest in these enzymes as potential targets that could contribute to the control of parasite survival and proliferation. Chicken eggs immunized with BnSP-7, a Lys49 phospholipase A 2 (PLA 2 ) homologue from Bothrops pauloensis snake venom, represent an excellent source of polyclonal antibodies with potential inhibitory activity on parasite PLA s. Herein, we report the production, characterization and anti-parasitic effect of IgY antibodies from egg yolks of hens immunized with BnSP-7. Produced antibodies presented increasing avidity and affinity for antigenic toxin epitopes throughout immunization, attaining a plateau after 4weeks. Pooled egg yolks-purified anti-BnSP-7 IgY antibodies were able to specifically recognize different PLA 2 s from Bothrops pauloensis and Bothrops jararacussu venom. Antibodies also neutralized BnSP-7 cytotoxic activity in C2C12 cells. Also, the antibodies recognized targets in Leishmania (Leishmania) amazonensis and Toxoplasma gondii extracts by ELISA and immunofluorescence assays. Anti-BnSP-7 IgY antibodies were cytotoxic to T. gondii tachyzoite and L. (L.) amazonensis promastigotes, and were able to decrease proliferation of both parasites treated before infection. These data suggest that the anti-BnSP-7 IgY is an important tool for discovering new parasite targets and blocking parasitic effects. Copyright © 2018 Elsevier B.V. All rights reserved.

  8. General applicability of chicken egg yolk antibodies: the performance of IgY immunoglobulins raised against the hypoxia-inducible factor 1alpha

    OpenAIRE

    Camenisch, G; Tini, M; Chilov, D; Kvietikova, I; Srinivas, V; Caro, J; Spielmann, P; Wenger, R H; Gassmann, M

    1999-01-01

    Avian embryos and neonates acquire passive immunity by transferring maternal immunoglobulins from serum to egg yolk. Despite being a convenient source of antibodies, egg yolk immunoglobulins (IgY) from immunized hens have so far received scant attention in research. Here we report the generation and rapid isolation of IgY from the egg yolk of hens immunized against the alpha subunit of the human hypoxia-inducible factor 1 (HIF-1alpha). Anti-HIF-1alpha IgY antibodies were affinity purified and...

  9. Solar Cycle variations of ƒoF2 from IGY to 1990

    Directory of Open Access Journals (Sweden)

    M. K. Goel

    Full Text Available Noontime monthly median values of F2-layer critical frequency foF2 (m for some ionospheric stations representing low- and mid-latitudes are examined for their dependence on solar activity for the years 1957 (IGY to 1990. This is the period for which ionospheric data in digital form is available in two CD-ROMs at the World Data Center, Boulder. It is observed that at mid-latitudes, foF2 (m shows nearly a linear relationship with R12 (the 12-month running average of the Zurich sunspot number, though this relation is nonlinear for low-latitudes. These results indicate some departures from the existing information often used in theoretical and applied areas of space research.Key words. Ionosphere (equatorial ionosphere; mid-latitude ionosphere; modelling and forecasting

  10. Grid: From EGEE to EGI and from INFN-Grid to IGI

    International Nuclear Information System (INIS)

    Giselli, A.; Mazzuccato, M.

    2009-01-01

    In the last fifteen years the approach of the computational Grid has changed the way to use computing resources. Grid computing has raised interest worldwide in academia, industry, and government with fast development cycles. Great efforts, huge funding and resources have been made available through national, regional and international initiatives aiming at providing Grid infrastructures, Grid core technologies, Grid middle ware and Grid applications. The Grid software layers reflect the architecture of the services developed so far by the most important European and international projects. In this paper Grid e-Infrastructure story is given, detailing European, Italian and international projects such as EGEE, INFN-Grid and NAREGI. In addition the sustainability issue in the long-term perspective is described providing plans by European and Italian communities with EGI and IGI.

  11. Eficacia experimental de anticuerpos IgY producidos en huevos, contra el veneno de la serpiente peruana Bothrops atrox Experimental efficacy of IgY antibodies produced in eggs against the venom of the Peruvian snake Bothrops atrox

    Directory of Open Access Journals (Sweden)

    Julio C. Mendoza

    2012-03-01

    Full Text Available Objetivos. Desarrollar un protocolo de inmunización para producir inmunoglobulinas IgY de origen aviar contra el veneno de la serpiente peruana Bothrops atrox y evaluar la capacidad neutralizante. Materiales y métodos. Se inmunizaron seis gallinas de postura de la raza hy line brown con 500 μg/dosis de veneno de B. atrox en un periodo de dos meses. Cada semana, los huevos fueron colectados para el aislamiento de inmunoglobulinas IgY a partir de la yema, usando dos pasos consecutivos con αcido caprνlico y sulfato de amonio. La detecciσn de anticuerpos se realizσ por inmunodifusiσn doble mientras que el tνtulo y reactividad cruzada se determinaron por las técnicas de ELISA y Western blot. El cálculo de DL50 y de la DE50 del antiveneno IgY producido se realizó utilizando el método de Probits. Resultados. La masa de anticuerpos aislados fue de 8,5 ± 1,35 mg de IgY/mL de yema. Asimismo, la DE50 del antiveneno aviar fue calculada en 575 μL de antiveneno/mg de veneno. Adicionalmente, los ensayos de reactividad cruzada mostraron que el veneno de B. atrox comparte mas epνtopes comunes con el veneno de B. brazili (47% que con otros veneno del mismo género, en tanto que los venenos de Lachesis muta (19% y Crotalus durissus (12% mostraron una baja reactividad cruzada. Conclusiones. Se ha obtenido IgY purificada contra el veneno de B. atrox con capacidad neutralizante y se ha demostrado su utilidad como herramienta inmunoanalítica para evaluar la reactividad cruzada con venenos de otras especies.Objectives. To develop an immunization protocol in order to produce avian IgY immunoglobulins against Bothrops atrox Peruvian snake venom and to evaluate its neutralizing capacity. Materials and methods. Six Hy Line Brown hens were immunized each two weeks using 500μg/doses of B. atrox venom in a period of two months. Each week, eggs were collected for IgY isolation from yolk using two consecutive steps with caprilic acid and ammonium sulfate

  12. Production, Characterization and Use of Monoclonal Antibodies Recognizing IgY Epitopes Shared by Chicken, Turkey, Pheasant, Peafowl and Sparrow

    Directory of Open Access Journals (Sweden)

    Ajda Biček

    2004-01-01

    Full Text Available Chicken antibodies are not only a part of immune defense but are more and more popular commercial products in form of chicken polyclonal, monoclonal or recombinant antibodies. We produced and characterized mouse monoclonal antibodies (mAbs that recognize epitopes located on heavy or light chain of chicken immunoglobulin Y (chIgY shared also by some other Phasianidae birds. The use of mAbs 1F5 and 2F10 that recognize heavy chain on chIgY common epitopes was demonstrated on immunoglobulins of turkey, pheasant and peafowl. Chicken IgY light chain specific mAb 3E10 revealed the presence of common epitopes on immunoglobulins of turkey, pheasant and sparrow. Monoclonal antibody clone 1F5/3G2 was used to prepare horseradish peroxidase (HRP conjugate and immunoadsorbent column. Conjugated mAbs were demonstrated to be excellent secondary antibodies for diagnostics of certain infections in different avian species. Since they do not react with mammalian immunoglobulins using our mAbs as secondary antibodies in human serodiagnostics would minimize background staining that appears when using mouse detection system. In dot immunobinding assay (DIBA and immunoblot assay they recognized specific IgY antibodies against Mycoplasma synoviae, Mycoplasma gallisepticum and Newcastle disease virus in sera of infected or vaccinated birds. Immunoadsorption as a method for removal of IgY from samples in which Mycoplasma synoviae specific IgY was predominant immunoglobulin class enabled more exact demonstration of specific IgA and IgM antibodies. Herein we are presenting effective mAbs useful in diagnostics of avian and mammalian infections as well as in final steps of detection and purification of chicken antibodies and their subunits produced in vivo or in vitro as polyclonal, monoclonal or recombinant antibodies.

  13. Growth inhibition of Staphylococcus aureus and escherichia coli strains by neutralizing IgY antibodies from ostrich egg yolk

    Directory of Open Access Journals (Sweden)

    Fernando Luiz Tobias

    2012-06-01

    Full Text Available Ostrich raising around the world have some key factors and farming profit depend largely on information and ability of farmers to rear these animals. Non fertilized eggs from ostriches are discharged in the reproduction season. Staphylococcus aureus and Escherichia coli are microorganisms involved in animal and human diseases. In order to optimize the use of sub products of ostrich raising, non fertilized eggs of four selected birds were utilized for development of polyclonal IgY antibodies. The birds were immunized (200ug/animal with purified recombinant staphylococcal enterotoxin C (recSEC and synthetic recRAP, both derived from S. aureus, and recBFPA and recEspB involved in E. coli pathogenicity, diluted in FCA injected in the braquial muscle. Two subsequent immunization steps with 21 days intervals were repeated in 0,85% saline in FIA. Blood and eggs samples were collected before and after immunization steps. Egg yolk immunoglobulins were purified by precipitation with 19% sodium sulfate and 20% ammonium sulphate methodologies. Purified IgY 50µL aliquots were incubated in 850µL BHI broth containing 50µL inoculums of five strains of S. aureus and five strains of E.coli during four hours at 37ºC. Growth inhibition was evaluated followed by photometry reading (DO550nm. Egg yolk IgY preparation from hiperimmunized birds contained antibodies that inhibited significantly (p<0,05 growth of strains tested. Potential use of ostrich IgY polyclonal antibodies as a diagnostic and therapeutic tool is proposed for diseased animals.

  14. OBTENCIÓN DE ANTICUERPOS POLICLONALES IgY ANTIPARVOVIRUS CANINO A PARTIR DE YEMA DE HUEVO DE GALLINA

    Directory of Open Access Journals (Sweden)

    Pinto, J.

    2005-06-01

    Full Text Available Inmunizando un grupo de diez gallinas de postura raza Lohmann de 16 semanas de edad con parvovirus canino (CPV cepa vacunal, se obtuvieron anticuerpos IgY policlonales contra CPV en yema de huevo.En la extracción de los anticuerpos de la yema se requirieron dos pasos, el primero para la remoción de lípidos y el segundo para la precipitación de proteínas. Para la remoción de lípidos se usó el método PBS:Cloroformo y para la precipitación de los anticuerpos solubles (Inmunoglobulinas-IgY se usó el método de salting-out con sulfato de amonio ((NH42SO4. La evaluación del proceso se efectuó empleando el método comercial estándar “EGGstract IgY Purification System®” de PROMEGA.La metodología empleada permitió la obtención de anticuerpos IgY policlonales contra CPV a partir de yema de huevo de gallina en concentraciones altas por mililitro de yema, con una pureza aceptable y títulos altos; los resultados fueron comparables con el método comercial.

  15. Homogeneous immunoassay for human IgG using oriented hen egg IgY immobilized on gold sol nanoparticles

    International Nuclear Information System (INIS)

    Yeritsyan, H.E.; Gasparyan, V.K.

    2012-01-01

    Homogeneous immunoassays using (red) gold nanoparticles represent an attractive detection scheme because of the option of photometric readout. We have applied oriented immobilization of hen egg immunoglobulin Y (IgY) on gold nanoparticles when developing a homogeneous immunoassay for human IgG. In oriented immobilization, as opposed to random immobilization, the antigen binding capabilities of the antibodies are retained. It is shown that such immunoassay has significantly better sensitivity in comparison with methods based on conventional immobilization of affinity-purified antibodies. It is also shown that hen egg IgY is better suited than rabbit antibodies, because much more antibody can be immobilized on gold nanoparticles without any destabilization, probably because of the more acidic nature of these antibodies. In addition, hen egg IgY can be supplied in higher quantity and can be prepared more easily than IgG from rabbits. Bleeding and slaughtering of animals is not needed. The assay presented here has a wide detection range (30-500 ng. mL -1 ) and a limit of detection as low as 30 ng. mL -1 of human IgG. (author)

  16. Extensive diversification of IgD-, IgY-, and truncated IgY(δFc)-encoding genes in the red-eared turtle (Trachemys scripta elegans).

    Science.gov (United States)

    Li, Lingxiao; Wang, Tao; Sun, Yi; Cheng, Gang; Yang, Hui; Wei, Zhiguo; Wang, Ping; Hu, Xiaoxiang; Ren, Liming; Meng, Qingyong; Zhang, Ran; Guo, Ying; Hammarström, Lennart; Li, Ning; Zhao, Yaofeng

    2012-10-15

    IgY(ΔFc), containing only CH1 and CH2 domains, is expressed in the serum of some birds and reptiles, such as ducks and turtles. The duck IgY(ΔFc) is produced by the same υ gene that expresses the intact IgY form (CH1-4) using different transcriptional termination sites. In this study, we show that intact IgY and IgY(ΔFc) are encoded by distinct genes in the red-eared turtle (Trachemys scripta elegans). At least eight IgY and five IgY(ΔFc) transcripts were found in a single turtle. Together with Southern blotting, our data suggest that multiple genes encoding both IgY forms are present in the turtle genome. Both of the IgY forms were detected in the serum using rabbit polyclonal Abs. In addition, we show that multiple copies of the turtle δ gene are present in the genome and that alternative splicing is extensively involved in the generation of both the secretory and membrane-bound forms of the IgD H chain transcripts. Although a single μ gene was identified, the α gene was not identified in this species.

  17. Quality of electricity service: Evaluation of nuisance index (IGI) of industrial customers; Qualite du service electrique:evaluation de l`indice de gene individuel des clients industriels

    Energy Technology Data Exchange (ETDEWEB)

    Naggar, R. [Hydro-Quebec, Montreal, PQ (Canada)

    1996-08-01

    The effects of power interruption on individual industrial customers by computing an individual nuisance index (IGI) is one of the tools planned by Hydro-Quebec to measure quality of service to its customers. When fully functioning, IGI will represent a combined value of loss of sales, overtime, lost materials and other direct costs, each IGI tailor-made for a particular company. Data for computing the index will be obtained from Hydro-Quebec`s own customer classification database, plus a commercial technical database (DTC) containing data required for the assessment of the nuisance, and a survey carried out by Hydro-Quebec involving some 1600 industrial customers. As of this date, the DTC is not yet available. A statistical analysis of survey responses was substituted to provide default values based on available parameters. Hydro-Quebec is confident that this new approach to evaluating service quality will open new horizons in quality assurance. 3 refs., 5 tabs.

  18. Anti-Pseudomonas aeruginosa IgY Antibodies Induce Specific Bacterial Aggregation and Internalization in Human Polymorphonuclear Neutrophils

    DEFF Research Database (Denmark)

    Thomsen, K.; Christophersen, L.; Bjarnsholt, T.

    2015-01-01

    with P. aeruginosa by augmenting the phagocytic competence of PMNs may postpone the deteriorating chronic biofilm infection. Anti-P. aeruginosa IgY antibodies significantly increase the PMN-mediated respiratory burst and subsequent bacterial killing of P. aeruginosa in vitro. The mode of action...... is attributed to IgY-facilitated formation of immobilized bacteria in aggregates, as visualized by fluorescence microscopy and the induction of increased bacterial hydrophobicity. Thus, the present study demonstrates that avian egg yolk immunoglobulins (IgY) targeting P. aeruginosa modify bacterial fitness...

  19. Egg yolk IgY: protection against rotavirus induced diarrhea and modulatory effect on the systemic and mucosal antibody responses in newborn calves.

    Science.gov (United States)

    Vega, C; Bok, M; Chacana, P; Saif, L; Fernandez, F; Parreño, V

    2011-08-15

    Bovine rotavirus (BRV) is an important cause of diarrhea in newborn calves. Local passive immunity is the most efficient protective strategy to control the disease. IgY technology (the use of chicken egg yolk immunoglobulins) is an economic and practical alternative to prevent BRV diarrhea in dairy calves. The aim of this study was to evaluate the protection and immunomodulation induced by the oral administration of egg yolk enriched in BRV specific IgY to experimentally BRV infected calves. All calves in groups Gp 1, 2 and 3 received control colostrum (CC; BRV virus neutralization Ab titer - VN=65,536; ELISA BRV IgG(1)=16,384) prior to gut closure. After gut closure, calves received milk supplemented with 6% BRV-immune egg yolk [(Gp 1) VN=2048; ELISA IgY Ab titer=4096] or non-immune control egg yolk [(Gp 2) VNcontrols (Gp 3 and 4, respectively). Calves were inoculated with 10(5.85)focus forming units (FFU) of virulent BRV IND at 2 days of age. Control calves (Gp 3 and 4) and calves fed control IgY (Gp 2) were infected and developed severe diarrhea. Around 80% calves in Gp 1 (IgY 4096) were infected, but they showed 80% (4/5) protection against BRV diarrhea. Bovine RV-specific IgY Ab were detected in the feces of calves in Gp 1, indicating that avian antibodies (Abs) remained intact after passage through the gastrointestinal tract. At post infection day 21, the duodenum was the major site of BRV specific antibody secreting cells (ASC) in all experimental groups. Mucosal ASC responses of all isotypes were significantly higher in the IgY treated groups, independently of the specificity of the treatment, indicating that egg yolk components modulated the immune response against BRV infection at the mucosal level. These results indicate that supplementing newborn calves' diets for the first 14 days of life with egg yolk enriched in BRV-specific IgY represents a promising strategy to prevent BRV diarrhea. Moreover a strong active ASC immune response is induced in the

  20. Production and purification of avian antibodies (IgYs from inclusion bodies of a recombinant protein central in NAD+ metabolism

    Directory of Open Access Journals (Sweden)

    Paula A. Moreno-González

    2013-08-01

    Full Text Available The use of hens for the production of polyclonal antibodies reduces animal intervention and moreover yields a higher quantity of antibodies than other animal models.  The phylogenetic distance between bird and mammal antigens, often leads to more specific avian antibodies than their mammalian counterparts.Since a large amount of antigen is required for avian antibody production, the use of recombinant proteins for this procedure has been growing faster over the last years. Nevertheless, recombinant protein production through heterologous systems frequently prompts the protein to precipitate, forming insoluble aggregates of limited utility (inclusion bodies. A methodology for the production of avian polyclonal antibodies, using recombinant protein from inclusion bodies is presented in this article.In order to produce the antigen, a recombinant Nicotinamide mononucleotide adenylyltransferase from Giardia intestinalis (His-GiNMNAT was expressed in Escherichia coli.  The protein was purified through solubilization from inclusion bodies prior to its renaturalization.  Antibodies were purified from egg yolk of immunized hens by water dilution, followed by ammonium sulfate precipitation and thiophilic affinity chromatography.The purified antibodies were tested against His-GiNMNAT protein in Western blot essays. From one egg yolk, 14.4 mg of highly pure IgY were obtained; this antibody was able to detect 15ng of His-GiNMNAT.  IgY specificity was improved by means of antigen affinity purification, allowing its use for parasite protein recognition.

  1. Updates on antibody functions in Mycobacterium tuberculosis infection and their relevance for developing a vaccine against tuberculosis.

    Science.gov (United States)

    Achkar, Jacqueline M; Prados-Rosales, Rafael

    2018-04-12

    A more effective vaccine to control tuberculosis (TB), a major global public health problem, is urgently needed. Current vaccine candidates focus predominantly on eliciting cell-mediated immunity but other arms of the immune system also contribute to protection against TB. We review here recent studies that enhance our current knowledge of antibody-mediated functions against Mycobacterium tuberculosis. These findings, which contribute to the increasing evidence that antibodies have a protective role against TB, include demonstrations that firstly distinct human antibody Fc glycosylation patterns, found in latent M. tuberculosis infection but not in active TB, influence the efficacy of the host to control M. tuberculosis infection, secondly antibody isotype influences human antibody functions, and thirdly that antibodies targeting M. tuberculosis surface antigens are protective. We discuss these findings in the context of TB vaccine development and highlight the need for further research on antibody-mediated immunity in M. tuberculosis infection. Copyright © 2018 Elsevier Ltd. All rights reserved.

  2. Anti-Interleukin-1 Beta/Tumor Necrosis Factor-Alpha IgY Antibodies Reduce Pathological Allergic Responses in Guinea Pigs with Allergic Rhinitis

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    Hu Wei-xu

    2016-01-01

    Full Text Available This study aims to determine whether the combined blockade of IL-1β and TNF-α can alleviate the pathological allergic inflammatory reaction in the nasal mucosa and lung tissues in allergic rhinitis (AR guinea pigs. Healthy guinea pigs treated with saline were used as the healthy controls. The AR guinea pigs were randomly divided into (1 the AR model group treated with intranasal saline; (2 the 0.1% nonspecific IgY treatment group; (3 the 0.1% anti-TNF-α IgY treatment group; (4 the 0.1% anti-IL-1β IgY treatment group; (5 the 0.1% combined anti-IL-1β and TNF-α IgY treatment group; and (6 the fluticasone propionate treatment group. The inflammatory cells were evaluated using Wright’s staining. Histopathology was examined using hematoxylin-eosin staining. The results showed that the number of eosinophils was significantly decreased in the peripheral blood, nasal lavage fluid, and bronchoalveolar lavage fluid (P<0.05, and eosinophil, neutrophil, and lymphocyte infiltration and edema were significantly reduced or absent in the nasal mucosa and lung tissues (P<0.05 in the combined 0.1% anti-IL-1β- and TNF-α IgY-treated guinea pigs. The data suggest that topical blockade of IL-1β and TNF-α could reduce pathological allergic inflammation in the nasal mucosa and lung tissues in AR guinea pigs.

  3. Anti-Interleukin-1 Beta/Tumor Necrosis Factor-Alpha IgY Antibodies Reduce Pathological Allergic Responses in Guinea Pigs with Allergic Rhinitis.

    Science.gov (United States)

    Wei-Xu, Hu; Wen-Yun, Zhou; Xi-Ling, Zhu; Zhu, Wen; Li-Hua, Wu; Xiao-Mu, Wu; Hui-Ping, Wei; Wen-Ding, Wang; Dan, He; Qin, Xiang; Guo-Zhu, Hu

    2016-01-01

    This study aims to determine whether the combined blockade of IL-1β and TNF-α can alleviate the pathological allergic inflammatory reaction in the nasal mucosa and lung tissues in allergic rhinitis (AR) guinea pigs. Healthy guinea pigs treated with saline were used as the healthy controls. The AR guinea pigs were randomly divided into (1) the AR model group treated with intranasal saline; (2) the 0.1% nonspecific IgY treatment group; (3) the 0.1% anti-TNF-α IgY treatment group; (4) the 0.1% anti-IL-1β IgY treatment group; (5) the 0.1% combined anti-IL-1β and TNF-α IgY treatment group; and (6) the fluticasone propionate treatment group. The inflammatory cells were evaluated using Wright's staining. Histopathology was examined using hematoxylin-eosin staining. The results showed that the number of eosinophils was significantly decreased in the peripheral blood, nasal lavage fluid, and bronchoalveolar lavage fluid (P guinea pigs. The data suggest that topical blockade of IL-1β and TNF-α could reduce pathological allergic inflammation in the nasal mucosa and lung tissues in AR guinea pigs.

  4. Dengue virus specific IgY provides protection following lethal dengue virus challenge and is neutralizing in the absence of inducing antibody dependent enhancement.

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    Ashley L Fink

    2017-07-01

    Full Text Available Dengue hemorrhagic fever (DHF and dengue shock syndrome (DSS are severe disease manifestations that can occur following sequential infection with different dengue virus serotypes (DENV1-4. At present, there are no licensed therapies to treat DENV-induced disease. DHF and DSS are thought to be mediated by serotype cross-reactive antibodies that facilitate antibody-dependent enhancement (ADE by binding to viral antigens and then Fcγ receptors (FcγR on target myeloid cells. Using genetically engineered DENV-specific antibodies, it has been shown that the interaction between the Fc portion of serotype cross-reactive antibodies and FcγR is required to induce ADE. Additionally, it was demonstrated that these antibodies were as neutralizing as their non-modified variants, were incapable of inducing ADE, and were therapeutic following a lethal, antibody-enhanced infection. Therefore, we hypothesized that avian IgY, which do not interact with mammalian FcγR, would provide a novel therapy for DENV-induced disease. We demonstrate here that goose-derived anti-DENV2 IgY neutralized DENV2 and did not induce ADE in vitro. Anti-DENV2 IgY was also protective in vivo when administered 24 hours following a lethal DENV2 infection. We were also able to demonstrate via epitope mapping that both full-length and alternatively spliced anti-DENV2 IgY recognized different epitopes, including epitopes that have not been previously identified. These observations provide evidence for the potential therapeutic applications of goose-derived anti-DENV2 IgY.

  5. Dengue virus specific IgY provides protection following lethal dengue virus challenge and is neutralizing in the absence of inducing antibody dependent enhancement.

    Science.gov (United States)

    Fink, Ashley L; Williams, Katherine L; Harris, Eva; Alvine, Travis D; Henderson, Thomas; Schiltz, James; Nilles, Matthew L; Bradley, David S

    2017-07-01

    Dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) are severe disease manifestations that can occur following sequential infection with different dengue virus serotypes (DENV1-4). At present, there are no licensed therapies to treat DENV-induced disease. DHF and DSS are thought to be mediated by serotype cross-reactive antibodies that facilitate antibody-dependent enhancement (ADE) by binding to viral antigens and then Fcγ receptors (FcγR) on target myeloid cells. Using genetically engineered DENV-specific antibodies, it has been shown that the interaction between the Fc portion of serotype cross-reactive antibodies and FcγR is required to induce ADE. Additionally, it was demonstrated that these antibodies were as neutralizing as their non-modified variants, were incapable of inducing ADE, and were therapeutic following a lethal, antibody-enhanced infection. Therefore, we hypothesized that avian IgY, which do not interact with mammalian FcγR, would provide a novel therapy for DENV-induced disease. We demonstrate here that goose-derived anti-DENV2 IgY neutralized DENV2 and did not induce ADE in vitro. Anti-DENV2 IgY was also protective in vivo when administered 24 hours following a lethal DENV2 infection. We were also able to demonstrate via epitope mapping that both full-length and alternatively spliced anti-DENV2 IgY recognized different epitopes, including epitopes that have not been previously identified. These observations provide evidence for the potential therapeutic applications of goose-derived anti-DENV2 IgY.

  6. Chicken Egg Yolk Antibodies (IgY) for Prophylaxis and Treatment of Rotavirus Diarrhea in Human and Animal Neonates: A Concise Review

    Science.gov (United States)

    Thu, Hlaing Myat; Myat, Theingi Win; Win, Mo Mo; Thant, Kyaw Zin; Rahman, Shofiqur; Umeda, Kouji; Nguyen, Sa Van; Icatlo, Faustino C.; Higo-Moriguchi, Kyoko; Taniguchi, Koki; Tsuji, Takao; Oguma, Keiji; Kim, Sang Jong; Bae, Hyun Suk

    2017-01-01

    The rotavirus-induced diarrhea of human and animal neonates is a major public health concern worldwide. Until recently, no effective therapy is available to specifically inactivate the rotavirion particles within the gut. Passive immunotherapy by oral administration of chicken egg yolk antibody (IgY) has emerged of late as a fresh alternative strategy to control infectious diseases of the alimentary tract and has been applied in the treatment of diarrhea due to rotavirus infection. The purpose of this concise review is to evaluate evidence on the properties and performance of anti-rotavirus immunoglobulin Y (IgY) for prevention and treatment of rotavirus diarrhea in human and animal neonates. A survey of relevant anti-rotavirus IgY basic studies and clinical trials among neonatal animals (since 1994-2015) and humans (since 1982-2015) have been reviewed and briefly summarized. Our analysis of a number of rotavirus investigations involving animal and human clinical trials revealed that anti-rotavirus IgY significantly reduced the severity of clinical manifestation of diarrhea among IgY-treated subjects relative to a corresponding control or placebo group. The accumulated information as a whole depicts oral IgY to be a safe and efficacious option for treatment of rotavirus diarrhea in neonates. There is however a clear need for more randomized, placebo controlled and double-blind trials with bigger sample size to further solidify and confirm claims of efficacy and safety in controlling diarrhea caused by rotavirus infection especially among human infants with health issues such as low birth weights or compromised immunity in whom it is most needed. PMID:28316465

  7. PURIFICACIÓN DE IgY CONTRA LA SUBUNIDAD NR3 DEL RECEPTOR NMDA DE CEREBRO DE RATA

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    Gina Méndez C

    2008-04-01

    Full Text Available Objetivo. Obtener anticuerpos tipo IgY contra péptidos sintéticos de las subunidades NR3A y NR3B del receptor NMDA de ratas, para reconocer y seguir la expresión de estas subunidades en extractos de cerebro de rata de diferentes edades. Materiales y métodos. Se diseñaron dos péptidos empleando los sistemas de la base de datos Entrez y el programa ClustalW-PBIL de alineamientos múltiples contra las subunidades NR3A y NR3B del receptor NMDA; una vez sintetizados por el método SSPS-fmoc fueron utilizados para inocular gallinas (Gallus gallus, variedad Hy Line Brown de 16 semanas de edad; al cabo de 57 días postinoculación se purificó IgY específica y se enfrentaron a extractos de cerebro de rata postnatal y adulta. Resultados. Se detectaron las subunidades NR3A y NR3B y se relacionó su expresión con la edad del animal; siendo mayor la expresión de la subunidad NR3A en extracto de cerebro de rata postnatal. No se encontró diferencia marcada en la expresión de la subunidad NR3B en las edades mencionadas. Conclusiones. Esta es la primera investigación que emplea proteína nativa para el reconocimiento de la subunidad NR3 del receptor NMDA, lo cual muestra la especificidad de los anticuerpos generados y contribuye con el entendimiento de las funciones de este receptor y su relación con la regulación de la memoria espacial.

  8. Hen egg yolk antibodies (IgY, production and use for passive immunization against bacterial enteric infections in chicken: a review

    Directory of Open Access Journals (Sweden)

    Chalghoumi R.

    2009-01-01

    Full Text Available Enteric infections caused by Salmonella remain a major public health burden worldwide. Poultry, particularly chickens, are known to be the main reservoir for this zoonotic pathogen. Therefore, the prevention and monitoring of Salmonella infection during the live phase may greatly reduce the contamination of poultry meat during slaughter and processing. With the ban on sub-therapeutic antibiotic usage in Europe and the increasingly strictness of the European legislation on food hygiene, passive immunization by oral administration of pathogen-specific hen egg yolk antibody (IgY may be a useful and attractive alternative. This review offers summarized information about IgY production and the use of these antibodies for passive immunization, particularly in poultry.

  9. A trial with IgY chicken antibodies to eradicate faecal carriage of Klebsiella pneumoniae and Escherichia coli producing extended-spectrum beta-lactamases

    OpenAIRE

    Jonsson, Anna-Karin; Larsson, Anders; Tängdén, Thomas; Melhus, Åsa; Lannergård, Anders

    2015-01-01

    Background: Extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae is an emerging therapeutic challenge, especially in the treatment of urinary tract infections. Following an outbreak of CTX-M-15 Klebsiella pneumoniae in Uppsala, Sweden, an orphan drug trial on IgY chicken antibodies was undertaken in an attempt to eradicate faecal carriage of ESBL-producing K. pneumoniae and Escherichia coli.Methods: Hens were immunised with epitopes from freeze-dried, whole-cell bacteria (ESBL...

  10. Chicken Egg Yolk Antibodies (IgY) for Prophylaxis and Treatment of Rotavirus Diarrhea in Human and Animal Neonates: A Concise Review

    OpenAIRE

    Thu, Hlaing Myat; Myat, Theingi Win; Win, Mo Mo; Thant, Kyaw Zin; Rahman, Shofiqur; Umeda, Kouji; Nguyen, Sa Van; Icatlo, Faustino C.; Higo-Moriguchi, Kyoko; Taniguchi, Koki; Tsuji, Takao; Oguma, Keiji; Kim, Sang Jong; Bae, Hyun Suk; Choi, Hyuk Joon

    2017-01-01

    The rotavirus-induced diarrhea of human and animal neonates is a major public health concern worldwide. Until recently, no effective therapy is available to specifically inactivate the rotavirion particles within the gut. Passive immunotherapy by oral administration of chicken egg yolk antibody (IgY) has emerged of late as a fresh alternative strategy to control infectious diseases of the alimentary tract and has been applied in the treatment of diarrhea due to rotavirus infection. The purpos...

  11. In vivo tracking of genetically engineered, anti-HER2/neu directed natural killer cells to HER2/neu positive mammary tumors with magnetic resonance imaging

    International Nuclear Information System (INIS)

    Daldrup-Link, Heike E.; Meier, Reinhardt; Metz, Stephan; Settles, Marcus; Rummeny, Ernst J.; Rudelius, Martina; Piontek, Guido; Schlegel, Juergen; Piert, Morand; Uherek, Christoph; Wels, Winfried

    2005-01-01

    The purpose of this study is to optimize labeling of the human natural killer (NK) cell line NK-92 with iron-oxide-based contrast agents and to monitor the in vivo distribution of genetically engineered NK-92 cells, which are directed against HER2/neu receptors, to HER2/neu positive mammary tumors with magnetic resonance (MR) imaging. Parental NK-92 cells and genetically modified HER2/neu specific NK-92-scFv(FRP5)-zeta cells, expressing a chimeric antigen receptor specific to the tumor-associated ErbB2 (HER2/neu) antigen, were labeled with ferumoxides and ferucarbotran using simple incubation, lipofection and electroporation techniques. Labeling efficiency was evaluated by MR imaging, Prussian blue stains and spectrometry. Subsequently, ferucarbotran-labeled NK-92-scFv(FRP5)-zeta (n=3) or parental NK-92 cells were intravenously injected into the tail vein of six mice with HER2/neu-positive NIH 3T3 mammary tumors, implanted in the mammary fat pad. The accumulation of the cells in the tumors was monitored by MR imaging before and 12 and 24 h after cell injection (p.i.). MR data were correlated with histopathology. Both the parental NK-92 and the genetically modified NK-92-scFv(FRP5)-zeta cells could be labeled with ferucarbotran and ferumoxides by lipofection and electroporation, but not by simple incubation. The intracellular cytoplasmatic iron-oxide uptake was significantly higher after labeling with ferucarbotran than ferumoxides (P 6 NK-92-scFv(FRP5)-zeta cells into tumor-bearing mice, MR showed a progressive signal decline in HER2/neu-positive mammary tumors at 12 and 24 h (p.i.). Conversely, injection of 5 x 10 6 parental NK-92 control cells, not directed against HER2/neu receptors, did not cause significant signal intensity changes of the tumors. Histopathology confirmed an accumulation of the former, but not the latter cells in tumor tissue. The human natural killer cell line NK-92 can be efficiently labeled with clinically applicable iron-oxide contrast agents, and the accumulation of these labeled cells in murine tumors can be monitored in vivo with MR imaging. This MR cell tracking technique may be applied to monitor NK-cell based immunotherapies in patients in order to assess the presence and extent of NK-cell tumor accumulations and, thus, to determine therapy response early and non-invasively. (orig.)

  12. In vivo tracking of genetically engineered, anti-HER2/neu directed natural killer cells to HER2/neu positive mammary tumors with magnetic resonance imaging

    Energy Technology Data Exchange (ETDEWEB)

    Daldrup-Link, Heike E. [UCSF Medical Center, Department of Radiology, San Francisco, CA (United States); Meier, Reinhardt; Metz, Stephan; Settles, Marcus; Rummeny, Ernst J. [Technical University Munich, Department of Radiology, Munich (Germany); Rudelius, Martina; Piontek, Guido; Schlegel, Juergen [Technical University Munich, Institute of Pathology, Division of Neuropathology, Munich (Germany); Piert, Morand [Technical University Munich, Department of Nuclear Medicine, Munich (Germany); Uherek, Christoph; Wels, Winfried [University of Frankfurt, Georg Speyer House, Frankfurt (Germany)

    2005-01-01

    The purpose of this study is to optimize labeling of the human natural killer (NK) cell line NK-92 with iron-oxide-based contrast agents and to monitor the in vivo distribution of genetically engineered NK-92 cells, which are directed against HER2/neu receptors, to HER2/neu positive mammary tumors with magnetic resonance (MR) imaging. Parental NK-92 cells and genetically modified HER2/neu specific NK-92-scFv(FRP5)-zeta cells, expressing a chimeric antigen receptor specific to the tumor-associated ErbB2 (HER2/neu) antigen, were labeled with ferumoxides and ferucarbotran using simple incubation, lipofection and electroporation techniques. Labeling efficiency was evaluated by MR imaging, Prussian blue stains and spectrometry. Subsequently, ferucarbotran-labeled NK-92-scFv(FRP5)-zeta (n=3) or parental NK-92 cells were intravenously injected into the tail vein of six mice with HER2/neu-positive NIH 3T3 mammary tumors, implanted in the mammary fat pad. The accumulation of the cells in the tumors was monitored by MR imaging before and 12 and 24 h after cell injection (p.i.). MR data were correlated with histopathology. Both the parental NK-92 and the genetically modified NK-92-scFv(FRP5)-zeta cells could be labeled with ferucarbotran and ferumoxides by lipofection and electroporation, but not by simple incubation. The intracellular cytoplasmatic iron-oxide uptake was significantly higher after labeling with ferucarbotran than ferumoxides (P<0.05). After intravenous injection of 5 x 10{sup 6} NK-92-scFv(FRP5)-zeta cells into tumor-bearing mice, MR showed a progressive signal decline in HER2/neu-positive mammary tumors at 12 and 24 h (p.i.). Conversely, injection of 5 x 10{sup 6} parental NK-92 control cells, not directed against HER2/neu receptors, did not cause significant signal intensity changes of the tumors. Histopathology confirmed an accumulation of the former, but not the latter cells in tumor tissue. The human natural killer cell line NK-92 can be efficiently labeled with clinically applicable iron-oxide contrast agents, and the accumulation of these labeled cells in murine tumors can be monitored in vivo with MR imaging. This MR cell tracking technique may be applied to monitor NK-cell based immunotherapies in patients in order to assess the presence and extent of NK-cell tumor accumulations and, thus, to determine therapy response early and non-invasively. (orig.)

  13. The HER2 CISH pharmDx(™) Kit in the assessment of breast cancer patients for anti-HER2 treatment.

    Science.gov (United States)

    Foged, Niels T; Brügmann, Anja; Jørgensen, Jan T

    2013-04-01

    Testing for amplification of the human EGF receptor 2 (HER2) gene by in situ hybridization is a central principle for the identification of breast cancer patients likely to respond to treatments directed toward HER2. However, its application in clinical routine has been somewhat restricted by the typical use of a visualization system based on fluorescence (FISH), which requires skilled, work-intensive, high-magnification quantitative microscopy. The US FDA has recently approved the HER2 CISH pharmDx™ Kit, which is characterized by employing a chromogenic visualization system that allows quantification of the HER2 gene and centromere 17 reference signals by relatively low-magnification brightfield microscopy. It is indicated as an aid in the assessment of patients for whom Herceptin(®) (trastuzumab) treatment is being considered.

  14. Limiting the protein corona: A successful strategy for in vivo active targeting of anti-HER2 nanobody-functionalized nanostars.

    Science.gov (United States)

    D'Hollander, Antoine; Jans, Hilde; Velde, Greetje Vande; Verstraete, Charlotte; Massa, Sam; Devoogdt, Nick; Stakenborg, Tim; Muyldermans, Serge; Lagae, Liesbet; Himmelreich, Uwe

    2017-04-01

    Gold nanoparticles hold great promise as anti-cancer theranostic agents against cancer by actively targeting the tumor cells. As this potential has been supported numerously during in vitro experiments, the effective application is hampered by our limited understanding and control of the interactions within complex in vivo biological systems. When these nanoparticles are exposed to a biological environment, their surfaces become covered with proteins and biomolecules, referred to as the protein corona, reducing the active targeting capabilities. We demonstrate a chemical strategy to overcome this issue by reducing the protein corona's thickness by blocking the active groups of the self-assembled monolayer on gold nanostars. An optimal blocking agent, 2-mercapto ethanol, has been selected based on charge and length of the carbon chain. By using a nanobody as a biological ligand of the human epidermal growth factor 2 receptor (HER2), the active targeting is demonstrated in vitro and in vivo in an experimental tumor model by using darkfield microscopy and photoacoustic imaging. In this study, we have established gold nanostars as a conceivable theranostic agent with a specificity for HER2-positive tumors. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. Determining true HER2 gene status in breast cancers with polysomy by using alternative chromosome 17 reference genes: implications for anti-HER2 targeted therapy.

    Science.gov (United States)

    Tse, Chun Hing; Hwang, Harry C; Goldstein, Lynn C; Kandalaft, Patricia L; Wiley, Jesse C; Kussick, Steven J; Gown, Allen M

    2011-11-01

    The ratio of human epidermal growth factor receptor 2 (HER2) to CEP17 by fluorescent in situ hybridization (FISH) with the centromeric probe CEP17 is used to determine HER2 gene status in breast cancer. Increases in CEP17 copy number have been interpreted as representing polysomy 17. However, pangenomic studies have demonstrated that polysomy 17 is rare. This study tests the hypothesis that the use of alternative chromosome 17 reference genes might more accurately assess true HER2 gene status. In all, 171 patients with breast cancer who had HER2 FISH that had increased mean CEP17 copy numbers (> 2.6) were selected for additional chromosome 17 studies that used probes for Smith-Magenis syndrome (SMS), retinoic acid receptor alpha (RARA), and tumor protein p53 (TP53) genes. A eusomic copy number exhibited in one or more of these loci was used to calculate a revised HER2-to-chromosome-17 ratio by using the eusomic gene locus as the reference. Of 132 cases classified as nonamplified on the basis of their HER2:CEP17 ratios, 58 (43.9%) were scored as amplified by using alternative chromosome 17 reference gene probes, and 13 (92.9%) of 14 cases scored as equivocal were reclassified as amplified. Among the cases with mean HER2 copy number of 4 to 6, 41 (47.7%) of 86 had their HER2 gene status upgraded from nonamplified to amplified, and four (4.7%) of 86 were upgraded from equivocal to amplified. Our results support the findings of recent pangenomic studies that true polysomy 17 is uncommon. Additional FISH studies that use probes to the SMS, RARA, and TP53 genes are an effective way to determine the true HER2 amplification status in patients with polysomy 17 and they have important potential implications for guiding HER2-targeted therapy in breast cancer.

  16. Imaging biomarkers to predict response to anti-HER2 (ErbB2) therapy in preclinical models of breast cancer

    Science.gov (United States)

    Shah, Chirayu; Miller, Todd W.; Wyatt, Shelby K.; McKinley, Eliot T.; Olivares, Maria Graciela; Sanchez, Violeta; Nolting, Donald D.; Buck, Jason R.; Zhao, Ping; Ansari, M. Sib; Baldwin, Ronald M.; Gore, John C.; Schiff, Rachel; Arteaga, Carlos L.; Manning, H. Charles

    2010-01-01

    Purpose To evaluate non-invasive imaging methods as predictive biomarkers of response to trastuzumab in mouse models of HER2-overexpressing breast cancer. The correlation between tumor regression and molecular imaging of apoptosis, glucose metabolism, and cellular proliferation was evaluated longitudinally in responding and non-responding tumor-bearing cohorts. Experimental Design Mammary tumors from MMTV/HER2 transgenic female mice were transplanted into syngeneic female mice. BT474 human breast carcinoma cell line xenografts were grown in athymic nude mice. Tumor cell apoptosis (NIR700-Annexin-V accumulation), glucose metabolism ([18F]FDG-PET), and proliferation ([18F]FLT-PET) were evaluated throughout a bi-weekly trastuzumab regimen. Imaging metrics were validated by direct measurement of tumor size and immunohistochemical (IHC) analysis of cleaved caspase-3, phosphorylated AKT (p-AKT) and Ki67. Results NIR700-Annexin-V accumulated significantly in trastuzumab-treated MMTV/HER2 and BT474 tumors that ultimately regressed, but not in non-responding or vehicle-treated tumors. Uptake of [18F]FDG was not affected by trastuzumab treatment in MMTV/HER2 or BT474 tumors. [18F]FLT PET imaging predicted trastuzumab response in BT474 tumors but not in MMTV/HER2 tumors, which exhibited modest uptake of [18F]FLT. Close agreement was observed between imaging metrics and IHC analysis. Conclusions Molecular imaging of apoptosis accurately predicts trastuzumab-induced regression of HER2(+) tumors and may warrant clinical exploration to predict early response to neoadjuvant trastuzumab. Trastuzumab does not appear to alter glucose metabolism substantially enough to afford [18F]FDG-PET significant predictive value in this setting. Although promising in one preclinical model, further studies are required to determine the overall value of [18F]FLT-PET as a biomarker of response to trastuzumab in HER2+ breast cancer. PMID:19584166

  17. Detection of Clonorchis sinensis circulating antigen in sera from Chinese patients by immunomagnetic bead ELISA based on IgY.

    Directory of Open Access Journals (Sweden)

    Ge Nie

    Full Text Available BACKGROUND: Clonorchiasis, caused by Clonorchis sinensis, is widely distributed in Southeast Asia including China. Clonorchiasis is included in control programs of neglected tropical diseases by World Health Organization (WHO because it is one of the major health problems in most endemic areas. Diagnosis of clonorchiasis plays a key role in the control programs. However, so far, there is no satisfactory method for clonorchiasis because of low sensitivity, poor practicality and high false positivity of available diagnostic tools. METHODOLOGY/PRINCIPAL FINDINGS: We developed an immunomagnetic bead enzyme-linked immunosorbent assay (ELISA based on IgY (egg yolk immunoglobulin against cysteine proteinase of C. sinensis for detection of circulating antigen in serum samples of patients infected with C. sinensis. The polyclonal IgY, coated with magnetic beads, was used as a capture antibody and a monoclonal IgG labeled with horseradish peroxidase as a detection antibody in the IgY-based immunomagnetic bead ELISA system (IgY-IMB-ELISA. The results showed that the sensitivity of IgY-IMB-ELISA was 93.3% (14 of 15 in cases of heavy infection (5000 to 9999 eggs per gram feces, i.e, EPG 5000-9999, 86.7% (13 of 15 in cases of moderate infection (EPG 1000-4999 and 75.0% (9 of 12 in cases of light infection (EPG <1000 of clonorchiasis. Together 36 of total 42 (85.7% serum samples of human clonorchiasis gave a positive reaction. There was a significant correlation between ELISA optical density and egg counts (EPG with a correlation coefficient of 0.83 in total 42 patients. There were no positive results in patients with trichinosis (n = 10 or cysticercosis (n = 10. Cross-reactivity was 6.7% (2 of 30 with schistosomiasis japonica and 10.0% (3 of 30 with paragonimiasis, respectively. No positive reaction was found in 20 healthy persons. CONCLUSIONS: Our findings suggest that IgY-IMB-ELISA appears to be a sensitive and specific assay for detection of circulating

  18. Detection of Clonorchis sinensis circulating antigen in sera from Chinese patients by immunomagnetic bead ELISA based on IgY.

    Science.gov (United States)

    Nie, Ge; Wang, Ting; Lu, Shengjun; Liu, Wenqi; Li, Yonglong; Lei, Jiahui

    2014-01-01

    Clonorchiasis, caused by Clonorchis sinensis, is widely distributed in Southeast Asia including China. Clonorchiasis is included in control programs of neglected tropical diseases by World Health Organization (WHO) because it is one of the major health problems in most endemic areas. Diagnosis of clonorchiasis plays a key role in the control programs. However, so far, there is no satisfactory method for clonorchiasis because of low sensitivity, poor practicality and high false positivity of available diagnostic tools. We developed an immunomagnetic bead enzyme-linked immunosorbent assay (ELISA) based on IgY (egg yolk immunoglobulin) against cysteine proteinase of C. sinensis for detection of circulating antigen in serum samples of patients infected with C. sinensis. The polyclonal IgY, coated with magnetic beads, was used as a capture antibody and a monoclonal IgG labeled with horseradish peroxidase as a detection antibody in the IgY-based immunomagnetic bead ELISA system (IgY-IMB-ELISA). The results showed that the sensitivity of IgY-IMB-ELISA was 93.3% (14 of 15) in cases of heavy infection (5000 to 9999 eggs per gram feces, i.e, EPG 5000-9999), 86.7% (13 of 15) in cases of moderate infection (EPG 1000-4999) and 75.0% (9 of 12) in cases of light infection (EPG <1000) of clonorchiasis. Together 36 of total 42 (85.7%) serum samples of human clonorchiasis gave a positive reaction. There was a significant correlation between ELISA optical density and egg counts (EPG) with a correlation coefficient of 0.83 in total 42 patients. There were no positive results in patients with trichinosis (n = 10) or cysticercosis (n = 10). Cross-reactivity was 6.7% (2 of 30) with schistosomiasis japonica and 10.0% (3 of 30) with paragonimiasis, respectively. No positive reaction was found in 20 healthy persons. Our findings suggest that IgY-IMB-ELISA appears to be a sensitive and specific assay for detection of circulating antigen in human clonorchiasis.

  19. Hypersensitive detection and quantitation of BoNT/A by IgY antibody against substrate linear-peptide.

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    Tao Li

    Full Text Available Botulinum neurotoxin A (BoNT/A, the most acutely poisonous substance to humans known, cleave its SNAP-25 substrate with high specificity. Based on the endopeptidase activity, different methods have been developed to detect BoNT/A, but most lack ideal reproducibility or sensitivity, or suffer from long-term or unwanted interferences. In this study, we developed a simple method to detect and quantitate trace amounts of botulinum neurotoxin A using the IgY antibody against a linear-peptide substrate. The effects of reaction buffer, time, and temperature were analyzed and optimized. When the optimized assay was used to detect BoNT/A, the limit of detection of the assay was 0.01 mouse LD50 (0.04 pg, and the limit of quantitation was 0.12 mouse LD50/ml (0.48 pg. The findings also showed favorable specificity of detecting BoNT/A. When used to detect BoNT/A in milk or human serum, the proposed assay exhibited good quantitative accuracy (88% < recovery < 111%; inter- and intra-assay CVs < 18%. This method of detection took less than 3 h to complete, indicating that it can be a valuable method of detecting BoNT/A in food or clinical diagnosis.

  20. Production and characterisation of a novel chicken IgY antibody raised against C-terminal peptide from human thymidine kinase 1.

    Science.gov (United States)

    Wu, Chuanjing; Yang, Rongjiang; Zhou, Ji; Bao, Shing; Zou, Li; Zhang, Pinggan; Mao, Yongrong; Wu, Jianping; He, Qimin

    2003-06-01

    Egg yolk is a good source of highly specific antibodies against mammalian antigens because of the phylogenetic distance between birds and mammals. Chicken egg yolk immunoglobulins (IgY) were generated to a synthetic 31-amino acid peptide from the C-terminal of human HeLa thymidine kinase 1 (TK1) enzyme. The anti-TK1 IgY antibody was purified using affinity chromatography against the 31-amino acid peptide. The purified antibody inhibited the catalytic activity of the TK1 enzyme in the CEM TK1(+) cells and recognized the 25-kDa subunit and tetrameric form of TK1, which has a pI value of 8.3. No immunoreaction was observed in CEM TK1(-) cells. Western blot of the serum TK1 (S-TK1) also showed that only a single band was found in the serum of patients with malignancies. No band was seen in healthy serum. Furthermore, dot blots and enhanced chemiluminescence (ECL) detection of S-TK1 performed on sera of preoperative patients with gastric cancer (GC) (n=31) and healthy controls (n=62) showed that the levels of S-TK1 in the sera of cancer patients were significantly different (Pstage cancer patients (four breast carcinomas, three hepatocarcinomas and four thyroid carcinomas) indicated that a strong staining of TK1 enzyme was found in the cytoplasm of malignant cells. No staining or weak staining was seen in normal tissues. We suggest that screening for TK1 using anti-TK1 IgY may be potentially useful for serological and immunohistochemical detection of TK1 as an early prognosis and for monitoring patients undergoing treatment.

  1. Detection of Shiga toxin-producing Escherichia coli by sandwich enzyme-linked immunosorbent assay using chicken egg yolk IgY antibodies

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    Yanil R Parma

    2012-06-01

    Full Text Available Enterohemorrhagic Escherichia coli (EHEC, a subset of Shiga toxin producing E. coli (STEC is associated with a spectrum of diseases that includes diarrhea, hemorrhagic colitis and a life-threatening hemolytic uremic syndrome (HUS. Regardless of serotype, Shiga toxins (Stx1 and/or Stx2 are uniformly expressed by all EHEC, and so exploitable targets for laboratory diagnosis of these pathogens. In this study, a sandwich ELISA for determination of Shiga toxin (Stx was developed using anti-Stx2 B subunit antibodies and its performance was compared with that of the Vero cell assay and a commercial immunoassay kit. Chicken IgY was used as capture antibody and a HRP-conjugated rabbit IgG as the detection antibody. The anti-Stx2B IgY was harvested from eggs laid by hens immunized with a recombinant protein fragment. Several parameters were tested in order to optimize the sandwich ELISA assay, including concentration of antibodies, type and concentration of blocking agent, and incubation temperatures. Supernatants from 42 STEC strains of different serotypes and stx variants, including stx2EDL933, stx2vha, stx2vhb, stx2g, stx1EDL933 and stx1d were tested. All Stx variants were detected by the sandwich ELISA, with a detection limit of 400 ng /ml Stx2. Twenty three strains negative for stx genes, including different bacteria species, showed no activity in Vero cell assay and produced negative results in ELISA, except for 2 strains. Our results show that anti-Stx2B IgY sandwich ELISA could be used in routine diagnosis as a rapid, specific and economic method for detection of Shiga toxin-producing E. coli.

  2. Detection of Shiga toxin-producing Escherichia coli by sandwich enzyme-linked immunosorbent assay using chicken egg yolk IgY antibodies

    Science.gov (United States)

    Parma, Y. R.; Chacana, P. A.; Lucchesi, P. M. A.; Rogé, A.; Granobles Velandia, C. V.; Krüger, A.; Parma, A. E.; Fernández-Miyakawa, M. E.

    2012-01-01

    Enterohemorrhagic Escherichia coli (EHEC), a subset of Shiga toxin producing E. coli (STEC) is associated with a spectrum of diseases that includes diarrhea, hemorrhagic colitis and a life-threatening hemolytic-uremic syndrome (HUS). Regardless of serotype, Shiga toxins (Stx1 and/or Stx2) are uniformly expressed by all EHEC, and so exploitable targets for laboratory diagnosis of these pathogens. In this study, a sandwich ELISA for determination of Shiga toxin (Stx) was developed using anti-Stx2B subunit antibodies and its performance was compared with that of the Vero cell assay and a commercial immunoassay kit. Chicken IgY was used as capture antibody and a HRP-conjugated rabbit IgG as the detection antibody. The anti-Stx2B IgY was harvested from eggs laid by hens immunized with a recombinant protein fragment. Several parameters were tested in order to optimize the sandwich ELISA assay, including concentration of antibodies, type and concentration of blocking agent, and incubation temperatures. Supernatants from 42 STEC strains of different serotypes and stx variants, including stx2EDL933, stx2vha, stx2vhb, stx2g, stx1EDL933, and stx1d were tested. All Stx variants were detected by the sandwich ELISA, with a detection limit of 115 ng/ml Stx2. Twenty three strains negative for stx genes, including different bacteria species, showed no activity in Vero cell assay and produced negative results in ELISA, except for two strains. Our results show that anti-Stx2B IgY sandwich ELISA could be used in routine diagnosis as a rapid, specific and economic method for detection of Shiga toxin-producing E. coli. PMID:22919675

  3. PRODUCTION AND PURIFICATION OF IgY ANTIBODIES AS A NOVEL TOOL TO PURIFY THE NR1 SUBUNIT OF NMDA RECEPTO

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    Edgar Antonio Reyes Montaño

    2011-12-01

    Full Text Available Producing polyclonal antibodies (IgY inchickens has advantages over those obtainedin other animal models, since theyhave been used as a tool for studyingdifferent proteins (NMDA glutamate receptorin our case, specifically the NR1subunit. We produced specific antibodiesagainst expression products by thealternative splicing of the gene encodingNMDA receptor NR1 subunit in adult ratbrain. Three peptides corresponding tothe splicing sites (N1, C1 and C2’ cassetteswere designed, synthesised and usedindividually as antigens in hens. Specificimmunoglobulins were purified fromyolks. The antibodies were then used forpurifying the NMDA receptor NR1 subunitusing affinity chromatography couplingthe three antibodies to the support.R

  4. Hen egg yolk antibodies (IgY), production and use for passive immunization against bacterial enteric infections in chicken : a review

    OpenAIRE

    Chalghoumi, Raja; Beckers, Yves; Portetelle, Daniel; Thewis, André

    2009-01-01

    Les anticorps du jaune d’oeuf de poule (IgY), production et utilisation en immunisation passive contre les infections entériques bactériennes : une revue. Les infections entériques causées par Salmonella constituent un problème majeur de santé publique à travers le monde. Il est bien connu que la volaille, en particulier le poulet de chair, constitue le principal réservoir pour ce pathogène zoonotique. Par conséquent, la prévention et la surveillance de Salmonella au cours de la phase d’éleva...

  5. Therapeutic potential of anti-IL-1β IgY in guinea pigs with allergic asthma induced by ovalbumin.

    Science.gov (United States)

    Wei-xu, Hu; Qin, Xiang; Zhu, Wen; Yuan-yi, Chen; Li-feng, Zeng; Zhi-yong, Liu; Dan, He; Xiao-mu, Wu; Guo-zhu, Hu

    2014-03-01

    Interleukin-1 beta (IL-1β) plays pivotal roles in the progression of allergic airway inflammation. This study aims to determine whether the blockade of IL-1β can inhibit airway inflammation in guinea pigs with allergic asthma induced by the inhalation of aerosolized ovalbumin (OVA). Healthy guinea pigs treated with saline were used as normal controls (group C). The guinea pigs with allergic asthma induced by the inhalation of aerosolized OVA were randomly divided into three groups: (1) the M group containing negative control animals treated with saline; (2) the Z1 group containing animals treated by the inhalation of atomized 0.1% anti-IL-1β immunoglobulin yolk (IgY); and (3) the Z2 group containing positive control animals that were treated with budesonide. The inflammatory cells in the peripheral blood (PB) and bronchoalveolar lavage fluid (BALF) were evaluated using methylene blue and eosin staining. Cytokine concentrations were measured using an enzyme-linked immunosorbent assay. Pulmonary sections were examined using hematoxylin-eosin staining. Allergic inflammation and damage to the pulmonary tissues were decreased in the Z1 group compared to the M group. Eosinophils and neutrophils in the PB and BALF were significantly decreased in the Z1 group compared to the M group (Pguinea pigs with allergic asthma. The inhibitory activity may be due to the decrease in the numbers of eosinophils and neutrophils and the reduced levels of inflammatory cytokines and IgE in the PB and BALF. Copyright © 2013 Elsevier Ltd. All rights reserved.

  6. Three IgH isotypes, IgM, IgA and IgY are expressed in Gentoo penguin and zebra finch.

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    Binyue Han

    Full Text Available Previous studies on a limited number of birds suggested that the IgD-encoding gene was absent in birds. However, one of our recent studies showed that the gene was definitely expressed in the ostrich and emu. Interestingly, we also identified subclass diversification of IgM and IgY in these two birds. To better understand immunoglobulin genes in birds, in this study, we analyzed the immunoglobulin heavy chain genes in the zebra finch (Taeniopygia guttata and Gentoo penguin (Pygoscelis papua, belonging respectively to the order Passeriformes, the most successful bird order in terms of species diversity and numbers, and Sphenisciformes, a relatively primitive avian order. Similar to the results obtained in chickens and ducks, only three genes encoding immunoglobulin heavy chain isotypes, IgM, IgA and IgY, were identified in both species. Besides, we detected a transcript encoding a short membrane-bound IgA lacking the last two CH exons in the Gentoo penguin. We did not find any evidence supporting the presence of IgD gene or subclass diversification of IgM/IgY in penguin or zebra finch. The obtained data in our study provide more insights into the immunoglobulin heavy chain genes in birds and may help to better understand the evolution of immunoglobulin genes in tetrapods.

  7. Three IgH isotypes, IgM, IgA and IgY are expressed in Gentoo penguin and zebra finch

    Science.gov (United States)

    Han, Haitang; Zhao, Yaofeng; Pan, Qingjie; Ren, Liming

    2017-01-01

    Previous studies on a limited number of birds suggested that the IgD-encoding gene was absent in birds. However, one of our recent studies showed that the gene was definitely expressed in the ostrich and emu. Interestingly, we also identified subclass diversification of IgM and IgY in these two birds. To better understand immunoglobulin genes in birds, in this study, we analyzed the immunoglobulin heavy chain genes in the zebra finch (Taeniopygia guttata) and Gentoo penguin (Pygoscelis papua), belonging respectively to the order Passeriformes, the most successful bird order in terms of species diversity and numbers, and Sphenisciformes, a relatively primitive avian order. Similar to the results obtained in chickens and ducks, only three genes encoding immunoglobulin heavy chain isotypes, IgM, IgA and IgY, were identified in both species. Besides, we detected a transcript encoding a short membrane-bound IgA lacking the last two CH exons in the Gentoo penguin. We did not find any evidence supporting the presence of IgD gene or subclass diversification of IgM/IgY in penguin or zebra finch. The obtained data in our study provide more insights into the immunoglobulin heavy chain genes in birds and may help to better understand the evolution of immunoglobulin genes in tetrapods. PMID:28403146

  8. Quantification of antibody (IgY) titers in hen eggs following immunization and their use in detecting cell surface molecules on nitrocellulose membranes.

    Science.gov (United States)

    Ruan, Guang-Ping; Ma, Li; Meng, Xiao-Jing; Meng, Min-Jie; Wang, Xiao-Ning; Lin, Ying; Wu, Zheng-Qiang; He, Xiao-Wei; Wang, Ju-Fang; Zhu, Yong

    2007-01-01

    HLA-A*0201 alpha chain and beta2m were expressed from a prokaryotic system, and after refolding and purification, the alpha chain and beta2m were used to immunize eight laying hens. The titer of egg yolk antibody against alpha chain increased from 10(2) to 10(5.3) The titer of egg yolk antibody against beta2m increased from 10(1) to 10(4.7). The extent of titer increase is similar between the two antigens. An average of 135 mg purified polyclonal antibody (IgY) can be easily obtained from one egg yolk. The use of egg collection rather than serum collection is compatible with modern animal protection regulations. An average of 28 eggs were obtained from a laying hen every month, with a total amount of 3780 mg immunoglobulin extracted from one immunized hen every month, which would be equivalent to 630 mL of serum or 1260 mL of blood per month. Chickens are an optimal host for the production of polyclonal antibodies with high titer and high yield. Purified IgY was labeled with horseradish peroxidase and reacted with PBMC on nitrocellulose membranes indicating that the antibody can bind to the native conformation of class I HLA molecule on PBMC.

  9. Three IgH isotypes, IgM, IgA and IgY are expressed in Gentoo penguin and zebra finch.

    Science.gov (United States)

    Han, Binyue; Li, Yan; Han, Haitang; Zhao, Yaofeng; Pan, Qingjie; Ren, Liming

    2017-01-01

    Previous studies on a limited number of birds suggested that the IgD-encoding gene was absent in birds. However, one of our recent studies showed that the gene was definitely expressed in the ostrich and emu. Interestingly, we also identified subclass diversification of IgM and IgY in these two birds. To better understand immunoglobulin genes in birds, in this study, we analyzed the immunoglobulin heavy chain genes in the zebra finch (Taeniopygia guttata) and Gentoo penguin (Pygoscelis papua), belonging respectively to the order Passeriformes, the most successful bird order in terms of species diversity and numbers, and Sphenisciformes, a relatively primitive avian order. Similar to the results obtained in chickens and ducks, only three genes encoding immunoglobulin heavy chain isotypes, IgM, IgA and IgY, were identified in both species. Besides, we detected a transcript encoding a short membrane-bound IgA lacking the last two CH exons in the Gentoo penguin. We did not find any evidence supporting the presence of IgD gene or subclass diversification of IgM/IgY in penguin or zebra finch. The obtained data in our study provide more insights into the immunoglobulin heavy chain genes in birds and may help to better understand the evolution of immunoglobulin genes in tetrapods.

  10. Obtención, purificación y caracterización de anticuerpos policlonales IgY desarrollados en gallina, dirigidos contra aislamientos colombianos de Giardia duodenalis.

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    Dabeiba Adriana García

    2005-12-01

    Full Text Available Introducción. El desarrollo de anticuerpos policlonales requiere de animales de laboratorio, generalmente, el conejo, que deben sangrarse para su obtención. Como alternativa, se han utilizado las gallinas. Objetivo. Desarrollar anticuerpos policlonales IgY anti-Giardia duodenalis y evaluar diferentes métodos para su purificación a partir de yema de huevo. Materiales y métodos. Se inmunizaron tres gallinas intramuscularmente con trofozoítos del parásito: las inmunizaciones se realizaron a los 0, 15, 30, 45, 60, 90 y 120 días. Se recolectaron huevos en cada etapa de la inmunización y se purificó la IgY por deslipidación (D y precipitación (P mediante cinco protocolos diferentes: M1: (P: sulfato de amonio/D: dextrán sulfato-cloruro de calcio, M2: (D: dextrán sulfato-cloruro de calcio/P: sulfato de amonio, M3: (D: cloroformo/ P: sulfato de amonio 50%, M4: (D: solución A/P:solución B y M5: (D: cloroformo/P: sulfato de amonio 30%. Se realizó evaluación inmunoquímica de la IgY anti-Giardia duodenalis mediante inmunodifusión, contrainmunoelectroforesis e inmunoelectrotransferencia (Western blot; la pureza de IgY por SDS-PAGE en presencia y ausencia de reductor y la concentración de la inmunoglobulina (mg/mL por espectrofotometría y densitometría. Resultados. La IgY anti-G. duodenalis mediante evaluación inmunoquímica presentó títulos hasta de 1:32. La inmunoglobulina en ausencia de reductor mostró una banda de 180 kd y en su presencia bandas de 30 y 68 kd, características de sus cadenas liviana y pesada, respectivamente. Las mayores concentraciones de inmunoglobulina se recuperaron con el método dos (M2, 4,6 mg de IgY por mL de yema de partida. Conclusiones. Por su facilidad y economía de producción en gallinas, los anticuerpos policlonales IgY anti-Giardia así obtenidos podrán utilizarse para desarrollar inmunoensayos que detecten el parásito en eluídos de heces.

  11. CARACTERIZACIÓN DE IGYS ANTI-LECTINA DE Salvia bogotensis Y SU APLICACIÓN EN ESTUDIOS CITOQUÍMICOS PARA LA DETECCIÓN DEL ANTÍGENO TN

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    Nohora Vega

    2010-02-01

    Full Text Available Las inmunoglobulinas aisladas de la yema de huevo (IgY son muy utilizadas actualmente en diversos campos de las ciencias biológicas, dadas sus ventajas frente a las IgG séricas de mamíferos. En un trabajo previo establecimos las condiciones de obtención de IgY dirigidas contra la lectinas de Salvia bogotensis; su utilización en estudios inmunocitoquímicos requiere conocer sus principales características moleculares y las condiciones para la interacción IgY-lectina. La lectina de Salvia bogotensis (SBoL reconoce específicamente el antígeno Tn, marcador tumoral en muchos tipos de cáncer, pero se requieren herramientas adicionales para evidenciar esta interacción a nivel celular. Dada la disponibilidad de IgY anti-lectina de S. bogotensis, se realizó este trabajo con el objeto de caracterizar molecularmente estas IgY y evaluar su utilización en estudios inmunocitoquímicos para la detección del antígeno Tn en células tumorales. A las IgY purificadas se les determinó su punto isoeléctrico, peso molecular y contenido de carbohidratos. Para establecer la especificidad de interacción IgY-SBoL se obtuvieron lectinas homólogas y heterólogas y se ensayaron por ELLSA. La detección del antígeno Tn en las líneas celulares MCF-7 y HeLa con la lectina y las IgYs marcadas con biotina o peroxidasa se realizó por CELISA e inmunocitoquímica. Los resultados mostraron que los anticuerpos IgY anti-SBoL son una herramienta de una alta sensibilidad para los ensayos de reconocimiento específico del antígeno Tn.

  12. Development of IgY antibodies in chickens and IgG in rabbits immunized against proteins of Pythium insidiosum isolated from horses in the state of Rio de Janeiro Desenvolvimento de Anticorpos IgY em galinhas e IgG em coelhos imunizados contra proteínas de Pythium insidiosum isolado de equinos no Estado do Rio de Janeiro, Brasil

    Directory of Open Access Journals (Sweden)

    Maria Fabíola Nunes Rangel

    2010-01-01

    Full Text Available Pythiosis is caused by Pythium insidiosum and the occurrence of disease in horses was described in the North and Northwest State of Rio de Janeiro, Brazil. The disease was described in cattle, sheep, humans, and horses in different states and regions across the country. This paper describes the development of IgY and IgG polyclonal antibodies, in chicken and rabbits, respectively against proteins extracted from kunkers and hyphae of P. insidiosum from affected horses. The proteins were recognized by chicken, rabbit and horse antibodies by immunodiffusion and Western blot against majority bands of 27 and 43 KDa, and titrated by ELISA. The antibodies IgY developed by the first time against Brazilian strains of P. insidiosum may represent a valuable tool in the detection of antigens of the pathogen and contribute to further studies aimed at immunotherapy and knowledge about this disease in endemic areas in Rio de Janeiro and in Brazil.Pitiose é causada por Pythium insidiosum e a doença foi descrita em equinos no Norte e Noroeste do Estado do Rio de Janeiro, Brasil. A doença foi descrita em bovinos, ovelhas, humanos e cavalos em diferentes estados e regiões do país. Este trabalho descreve o desenvolvimento de anticorpos policlonais, IgY e IgG, em galinha e coelho, respectivamente, contra proteínas extraídas de kunkers e hifas de P. insidiosum de cavalos doentes. As proteínas foram reconhecidas por anticorpos de galinha, coelho e cavalos contra as bandas majoritárias de 27 e 34 KDa em imunodifusão e Western blot tituladas por ELISA. Os anticorpos IgY desenvolvidos pela primeira vez contra cepas brasileiras de P. insidiosum podem representar um valioso instrumento na detecção de antígenos de patógenos e contribuem para novos estudos baseados na imunoterapia e no entendimento sobre esta doença em áreas endêmicas no Rio de Janeiro e em todo o país.

  13. A trial with IgY chicken antibodies to eradicate faecal carriage of Klebsiella pneumoniae and Escherichia coli producing extended-spectrum beta-lactamases

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    Anna-Karin Jonsson

    2015-11-01

    Full Text Available Background: Extended-spectrum beta-lactamase (ESBL-producing Enterobacteriaceae is an emerging therapeutic challenge, especially in the treatment of urinary tract infections. Following an outbreak of CTX-M-15 Klebsiella pneumoniae in Uppsala, Sweden, an orphan drug trial on IgY chicken antibodies was undertaken in an attempt to eradicate faecal carriage of ESBL-producing K. pneumoniae and Escherichia coli. Methods: Hens were immunised with epitopes from freeze-dried, whole-cell bacteria (ESBL-producing K. pneumoniae and E. coli and recombinant proteins of two K. pneumoniae fimbriae subunits (fimH and mrkD. The egg yolks were processed according to good manufacturing practice and the product was stored at−20°C until used. Using an internal database from the outbreak and the regular laboratory database, faecal carriers were identified and recruited from May 2005 to December 2013. The participants were randomised in a placebo-controlled 1:1 manner. Results: From 749 eligible patients, 327 (44% had deceased, and only 91 (12% were recruited and signed the informed consent. In the initial screening performed using the polymerase chain reaction, 24 participants were ESBL positive and subsequently randomised and treated with either the study drug or a placebo. The study was powered for 124 participants. Because of a very high dropout rate, the study was prematurely terminated. From the outbreak cohort (n=247, only eight patients were screened, and only one was positive with the outbreak strain in faeces. Conclusions: The present study design, using IgY chicken antibodies for the eradication of ESBL-producing K. pneumonia and E. coli, was ineffective in reaching its goal due to high mortality and other factors resulting in a low inclusion rate. Spontaneous eradication of ESBL-producing bacteria was frequently observed in recruited participants, which is consistent with previous reports.

  14. A trial with IgY chicken antibodies to eradicate faecal carriage of Klebsiella pneumoniae and Escherichia coli producing extended-spectrum beta-lactamases

    Science.gov (United States)

    Jonsson, Anna-Karin; Larsson, Anders; Tängdén, Thomas; Melhus, Åsa; Lannergård, Anders

    2015-01-01

    Background Extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae is an emerging therapeutic challenge, especially in the treatment of urinary tract infections. Following an outbreak of CTX-M-15 Klebsiella pneumoniae in Uppsala, Sweden, an orphan drug trial on IgY chicken antibodies was undertaken in an attempt to eradicate faecal carriage of ESBL-producing K. pneumoniae and Escherichia coli. Methods Hens were immunised with epitopes from freeze-dried, whole-cell bacteria (ESBL-producing K. pneumoniae and E. coli) and recombinant proteins of two K. pneumoniae fimbriae subunits (fimH and mrkD). The egg yolks were processed according to good manufacturing practice and the product was stored at−20°C until used. Using an internal database from the outbreak and the regular laboratory database, faecal carriers were identified and recruited from May 2005 to December 2013. The participants were randomised in a placebo-controlled 1:1 manner. Results From 749 eligible patients, 327 (44%) had deceased, and only 91 (12%) were recruited and signed the informed consent. In the initial screening performed using the polymerase chain reaction, 24 participants were ESBL positive and subsequently randomised and treated with either the study drug or a placebo. The study was powered for 124 participants. Because of a very high dropout rate, the study was prematurely terminated. From the outbreak cohort (n=247), only eight patients were screened, and only one was positive with the outbreak strain in faeces. Conclusions The present study design, using IgY chicken antibodies for the eradication of ESBL-producing K. pneumonia and E. coli, was ineffective in reaching its goal due to high mortality and other factors resulting in a low inclusion rate. Spontaneous eradication of ESBL-producing bacteria was frequently observed in recruited participants, which is consistent with previous reports. PMID:26560861

  15. Chicken IgY Fc linked to Bordetella avium ompA and Taishan Pinus massoniana pollen polysaccharide adjuvant enhances macrophage function and specific immune responses

    Directory of Open Access Journals (Sweden)

    Zhu Ruiliang

    2016-11-01

    Full Text Available Fc-fusion technologies, in which immunoglobulin Fc is genetically fused to an antigenic protein, have been developed to confer antibody-like properties to proteins and peptides. Mammalian IgG Fc fusion exhibits improved antigen-induced immune responses by providing aggregates with high avidity for the IgG Fc receptor and salvaging the antigenic portion from endosomal degradation. However, whether the linked chicken IgY Fc fragment shares similar characteristics to mammalian IgG Fc remains unclear. In this study, we linked the chicken IgY Fc gene to the outer membrane protein A (ompA of Borderella avium through overlapping PCR. The fusion gene was cloned into the pPIC9 plasmid to construct the recombinant Pichia pastoris transformant expressing the ompA–Fc fusion protein. The effects of the linked Fc on macrophage vitality, activity, efficiency of antigen processing, and immune responses induced by the fused ompA were investigated. Furthermore, the effect of Taishan Pinus massoniana pollen polysaccharide (TPPPS, an immunomodulator, on chicken macrophage activation was evaluated. TPPPS was also used as an adjuvant to investigate its immunomodulatory effect on immunoresponses induced by the fused ompA–Fc in chickens. The pinocytosis, phagocytosis, secretion of nitric oxide and TNF-α, and MHC-II molecular expression of the macrophages treated with the fused ompA–Fc were significantly higher than those of the macrophages treated with ompA alone. The addition of TPPPS to the fused ompA–Fc further enhanced macrophage functions. The fused ompA–Fc elicited higher antigen-specific immune responses and protective efficacy compared with ompA alone. Moreover, the fused ompA–Fc conferred higher serum antibody titers, serum IL-2 and IL-4 concentrations, CD4+ and CD8+ T-lymphocyte counts, lymphocyte transformation rate, and protection rate compared with ompA alone. Notably, the prepared TPPPS adjuvant ompA–Fc vaccines induced high immune

  16. A dual amplified electrochemical immunosensor for ofloxacin: Polypyrrole film-Au nanocluster as the matrix and multi-enzyme-antibody functionalized gold nanorod as the label

    International Nuclear Information System (INIS)

    Zang, Shuai; Liu, Yingju; Lin, Mouhong; Kang, Jianli; Sun, Yuanming; Lei, Hongtao

    2013-01-01

    Graphical abstract: Schematic representation of the OFL electrochemical immunosensor using Au nanoclusters/PPy/GCE as the substrate and multi-HRP-GNR-Ab2 bioconjugates as the label. Highlights: ► Gold nanorod was used to load HRP and Ab 2 to form multi-HRP-GNR-Ab 2 . ► A sensitive immunosensor for ofloxacin was constructed using the homemade antibody. ► A dual signal amplified strategy was based on the PPy-Au and multi-HRP-GNR-Ab 2 . -- Abstract: In this work, an electrochemical immunosensor, basing on a dual signal amplified strategy by employing a biocompatible polypyrrole film-Au nanocluster matrix as a sensor platform and multi-enzyme-antibody functionalized gold nanorod as an electrochemical detection label, is established for sensitive detection of ofloxacin (OFL). Firstly, polypyrrole film and Au nanoclusters were progressively fabricated onto the surface of a glassy carbon electrode via electropolymerization and electrochemical deposition, respectively. Such PPy-Au nanocomposite modified electrode was used to immobilize OFL-OVA, blocked with the blocking reagent, and then associated with the corresponding antibody. Secondly, gold nanorod (GNR) was synthesized to load horseradish peroxidase (HRP) and horseradish peroxidase-secondary antibody (HRP-Ab 2 ), and the resulting nanostructure (multi-HRP-GNR-Ab 2 ) was applied as the detection label. The fabrication process of the ordered multilayer structure and immunosensor were characterized by scanning electron microscopy (SEM) and electrochemical measurements, respectively. Finally, based on a competitive immunoassay, i.e., the association ability with the corresponding antibody between the captured antigen and free OFL in the solution, the fabricated immunosensor exhibited a sensitive response to OFL in the range from 0.08 to 410 ng/mL with a detection limit of 0.03 ng/mL. The current immunosensor exhibited good sensitivity, selectivity and long-term stability. This amplification strategy shows excellent

  17. Systematic Review of the Side Effects Associated With Anti-HER2-Targeted Therapies Used in the Treatment of Breast Cancer, on Behalf of the EORTC Quality of Life Group

    NARCIS (Netherlands)

    Sodergren, Samantha C.; Copson, Ellen; White, Alice; Efficace, Fabio; Sprangers, Mirjam; Fitzsimmons, Deborah; Bottomley, Andrew; Johnson, Colin D.

    2016-01-01

    Targeted therapies (TTs), notably trastuzumab, have improved outcomes for breast cancer characterised by overexpression of human epidermal growth factor receptors including HER2. Compared with chemotherapy treatments, TTs are more specific in their targets and are delivered over longer periods of

  18. Systematic Review of the Side Effects Associated With Anti-HER2-Targeted Therapies Used in the Treatment of Breast Cancer, on Behalf of the EORTC Quality of Life Group.

    Science.gov (United States)

    Sodergren, Samantha C; Copson, Ellen; White, Alice; Efficace, Fabio; Sprangers, Mirjam; Fitzsimmons, Deborah; Bottomley, Andrew; Johnson, Colin D

    2016-06-01

    Targeted therapies (TTs), notably trastuzumab, have improved outcomes for breast cancer characterised by overexpression of human epidermal growth factor receptors including HER2. Compared with chemotherapy treatments, TTs are more specific in their targets and are delivered over longer periods of time, thus presenting different side-effect profiles. The objective of this paper is to systematically review and describe the side effects associated with TTs used in the adjuvant and metastatic settings for HER2+ breast cancer. The MEDLINE, EMBASE, CINAHL, Web of Science and Cochrane Library databases were searched from January 2007 to March 2015 to identify clinical trials and prospective studies reporting toxicities associated with TTs (mainly trastuzumab and lapatinib) used without other therapies in the treatment of HER2-positive breast cancer. Two independent reviewers selected papers based on their titles and abstracts. All papers selected by either reviewer were included. A third reviewer extracted and tabulated the relevant data using a data extraction form. We identified 5478 papers, of which 299 were reviewed and 18 trials identified involving 6980 patients. A total of 66 side effects were identified, including 46 "patient-based" symptoms and 20 "medically defined" outcomes. Side effects were more common for patients treated with therapies other than trastuzumab or with dual-HER2 regimens and for patients with metastatic disease. Diarrhoea and skin rash were the most prevalent symptoms, experienced by 29 % and 22 % of patients overall, respectively. There were 119 (2 %) cardiac events reported, and these were not exclusive to trastuzumab-treated patients. The majority of side effects (n = 52) were experienced by 1 % or less of patients and were predominantly of grade 1/2 toxicity. This systematic review provides a detailed analysis of side effects of HER2+ therapies in a large number of patients included in trials, enabling an accurate estimate of prevalence and a complete understanding of the patients' experience. This will help clinicians and patients in treatment planning.

  19. Changes in 2-fluoro-2-deoxy-D-glucose incorporation, hexokinase activity and lactate production by breast cancer cells responding to treatment with the anti-HER-2 antibody trastuzumab

    Energy Technology Data Exchange (ETDEWEB)

    Cheyne, Richard W. [School of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD (United Kingdom); Trembleau, Laurent; McLaughlin, Abbie [School of Natural and Computing Sciences, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD (United Kingdom); Smith, Tim A.D., E-mail: t.smith@abdn.ac.u [School of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD (United Kingdom)

    2011-04-15

    Introduction: Changes in 2-[{sup 18}F]-fluoro-2-deoxy-D-glucose (FDG) incorporation by tumors, detected using positron emission tomography, during response to chemotherapy are utilized clinically in patient management. Here, the effect of treatment with growth-inhibitory doses of the anti-human epidermal growth factor receptor-2 antibody trastuzumab (Herceptin) on the incorporation of FDG by breast tumor cells was measured along with hexokinase (HK) and glucose transport to determine the potential of FDG-positron emission tomography in predicting response to these biological anti-cancer therapies and their modulatory effects on the steps involved in FDG incorporation. Methods: The sensitivity to trastuzumab of three breast tumor cell lines, SKBr3, MDA-MB-453 and MDA-MB-468, expressing human epidermal growth factor receptor-2 at high, medium and low levels, respectively, was determined using MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay over a 6-day period, and a clonogenic assay was carried out after 7- and 10-day exposures. FDG incorporation by cells treated with growth-inhibitory doses of trastuzumab was carried out after 4 h and 2, 4 and 6 days of treatment. Glucose transport (rate of uptake of the non-metabolizable analogue [{sup 3}H]O-methyl-D-glucose), HK activity and lactate production were measured on cells treated with inhibitory doses of trastuzumab for 6 days. Results: The IC{sub 50} doses for SKBr3 and MDA-MB-453 and the IC{sub 20} dose for MDA-MB-468 after 6 days of treatment with trastuzumab were 0.25, 1 and 170 {mu}g/ml, respectively. FDG incorporation by SKBr3 and MDA-MB-453 cells was found to be decreased using IC{sub 50} doses of trastuzumab for 6 days. At the IC{sub 50} doses, FDG incorporation was also decreased at 4 days and, in the case of MDA-MB-453, even after 4 h of treatment. Decreased FDG incorporation corresponded with decreased HK activity in these cells. Lactate production, previously suggested to be a potential measure of response, was found to be significantly decreased by SKBr3 and MDA-MB-453 cells responding to trastuzumab. Conclusion: FDG incorporation at the tumor cell level is modulated by treatment with growth-inhibitory doses of trastuzumab due to modulation of HK activity. Changes in lactate production may also be a useful determinant of response to trastuzumab.

  20. Safety and efficacy of neratinib (HKI-272) plus vinorelbine in the treatment of patients with ErbB2-positive metastatic breast cancer pretreated with anti-HER2 therapy.

    Science.gov (United States)

    Awada, A; Dirix, L; Manso Sanchez, L; Xu, B; Luu, T; Diéras, V; Hershman, D L; Agrapart, V; Ananthakrishnan, R; Staroslawska, E

    2013-01-01

    Neratinib (HKI-272) is a potent irreversible pan-ErbB tyrosine kinase inhibitor with clinical activity in patients with ErbB2/HER2-positive breast cancer. Phase I of this open-label, phase I/II study investigated the maximum tolerated dose (MTD) of oral neratinib (160 or 240 mg/day) plus vinorelbine (25 mg/m2; days 1 and 8 of each 21-day cycle) in patients with solid tumors. Phase II assessed the safety, clinical activity, and pharmacokinetics of the combination in patients with HER2-positive metastatic breast cancer; the primary efficacy end point was objective response (OR). In phase I (n=12), neratinib (240 mg) plus vinorelbine (25 mg/m2) was established as the MTD. In phase II, 79 patients with HER2-positive metastatic breast cancer were treated at the MTD. The most common treatment-related adverse events were diarrhea (96%), neutropenia (54%), and nausea (50%). Three patients discontinued treatment due to diarrhea. No clinically important skin side-effects were observed. The OR rate in assessable phase II patients was 41% (no prior lapatinib) and 8% (prior lapatinib). There was no evidence of pharmacokinetic interaction between neratinib and vinorelbine. Neratinib plus vinorelbine showed promising antitumor activity and no unexpected toxic effects in HER2-positive metastatic breast cancer patients. Trial registration ClinicalTrials.gov #NCT00706030.

  1. Utility of a Novel Three-Dimensional and Dynamic (3DD Cell Culture System for PK/PD Studies: Evaluation of a Triple Combination Therapy at Overcoming Anti-HER2 Treatment Resistance in Breast Cancer

    Directory of Open Access Journals (Sweden)

    Anusha Ande

    2018-05-01

    Full Text Available Background: Emergence of Human epidermal growth factor receptor 2 (HER2 therapy resistance in HER2-positive (HER2+ breast cancer (BC poses a major clinical challenge. Mechanisms of resistance include the over-activation of the PI3K/mTOR and Src pathways. This work aims to investigate a novel combination therapy that employs paclitaxel (PAC, a mitotic inhibitor, with everolimus (EVE, an mTOR inhibitor, and dasatinib (DAS, an Src kinase inhibitor, as a modality to overcome resistance.Methods: Static (two dimensional, 2D and three-dimensional dynamic (3DD cell culture studies were conducted using JIMT-1 cells, a HER2+ BC cell line refractory to HER2 therapies. Cell viability and caspase-3 expression were examined after JIMT-1 cell exposure to agents as monotherapy or in combination using a 2D setting. A pharmacokinetic/pharmacodynamic (PK/PD combination study with PAC+DAS+EVE was conducted over 3 weeks in a 3DD setting. PAC was administered into the system via a 3 h infusion followed by the addition of a continuous infusion of EVE+DAS 24 h post-PAC dosing. Cell counts and caspase-3 expression were quantified every 2 days. A semi-mechanistic PK/PD model was developed using the 2D data and scaled up to capture the 3DD data. The final model integrated active caspase-3 as a biomarker to bridge between drug exposures and cancer cell dynamics. Model fittings were performed using Monolix software.Results: The triple combination significantly induced caspase-3 activity in the 2D cell culture setting. In the 3DD cell culture setting, sequential dosing of PAC then EVE+DAS showed a 5-fold increase in caspase-3 activity and 8.5-fold decrease in the total cell number compared to the control. The semi-mechanistic PK/PD models fit the data well, capturing the time-course profiles of drug concentrations, caspase-3 expression, and cell counts in the 2D and 3DD settings.Conclusion: A novel, sequential triple combination therapeutic regimen was successfully evaluated in both 2D and 3DD in vitro cell culture systems. The efficacy of this combination at inhibiting the cellular proliferation and re-growth of HER2/mTOR resistant cell line, JIMT-1, is demonstrated. A biomarker-linked PK/PD model successfully captured all time-course data. The latter can be used as a modeling platform for a direct translation from 3DD in vitro settings to the clinic.

  2. Obtención de anticuerpos policlonales IgY antiparvovirus canino y evaluación en un sistema de aglutinación con látex

    Directory of Open Access Journals (Sweden)

    Jannet González-Figueredo

    2015-12-01

    Full Text Available La parvovirosis canina es una de las principales infecciones que provoca gastroenteritis, fundamentalmente en cachorros, con altos índices de morbiletalidad. Los diagnosticadores más utilizados se basan en la detección de partículas virales excretadas durante la fase aguda de la enfermedad. Algunos requieren equipos especializados, lo que aumenta los costos y el tiempo de diagnóstico. Por esto, la simplificación de estos métodos con alta sensibilidad y especificidad es prioritaria. En Cuba solo se logra un diagnóstico presuntivo sin la completa confirmación de la enfermedad, principalmente por la escasez o ausencia de un medio diagnóstico en toda la red de consultorios y clínicas veterinarias, rápido, eficaz y ajustable a nuestras condiciones. Este trabajo tuvo como objetivo obtener anticuerpos policlonales IgY a partir de yema de huevo de gallina y evaluarlos mediante un sistema de látex-aglutinación, para su posible uso como terapia y principalmente en el diagnóstico. Se inmunizaron por vía intramuscular dos gallinas de raza Leghorn con una cepa atenuada de parvovirus canino (PVC tipo 2. Se aplicaron 8 inoculaciones por ave cada 15 días. Se cosecharon los huevos y se purificaron los anticuerpos por el método de sulfato de dextrana/sulfato de sodio. Se determinó el título de IgY anti-PVC por el método de Inhibición de la Hemoaglutinación (IH y con antígeno de PVC acoplado a partículas de látex de poliestireno. Por IH se obtuvo un título de IgY anti-PVC de 1:1024. Las mezclas de anticuerpos con títulos alto y medio de ambas gallinas aglutinaron con los reactivos preparados con 230 y 460 µg/mL de antígeno. La mayor intensidad de la reacción y la mejor detectabilidad correspondieron a los reactivos elaborados con los anticuerpos de mayor título.

  3. Elisa de captura com IgY para quantificação de acetato de lupeol em Vernonia scorpioides lam. Pers (asteraceae Capture IgY-elisa to quantify lupeol acetate in Vernonia scorpioides lam. Pers (asteraceae

    Directory of Open Access Journals (Sweden)

    Ronald Bastos Freire

    2004-08-01

    Full Text Available O presente experimento descreve, pela primeira vez, a elaboração de um ensaio imunoenzimático (ELISA de captura com anticorpos IgY para a detecção e quantificação de acetato de lupeol (LAc em Vernonia scorpioides Lam. Pers (Asteraceae. Anticorpos anti-LAc, obtidos partir do soro das aves e gemas de seus ovos, após imunização com o conjugado acetato de lupeol-cBSA, foram concentrados em coluna contendo LAc-BSA acoplado a sepharose. A eficácia da metodologia de detecção imunológica foi de 97%, com índices de sensibilidade e especificidade de 99% e 95%, respectivamente com limites de detecção do ensaio entre 0,02µg.g-1 (inferior e 10 µg.g-1 (superior. A robustez do método foi atestada pela sua elevada reprodutibilidade (entre 94,75% e 96,81%, e pelo baixo coeficiente de variação interna (4,22 ± 1,03% nas condições de execução descritas.For the first time, a yolk immunoglobulins-based immunenzymatic assay (capture IgY-ELISA was carried out to detect Lupeol acetate (LAc from Vernonia scorpioides Lam. Pers (Asteraceae. Antibodies (IgY against lupeol acetate (anti-LAc antibodies were raised in White Leghorn hens immunized with LAc conjugated to the bovine serum albumin (LAc-BSA. The anti-LAc antibodies were recovered by cleanup columns containing LAc-BSA coupled to sepharose. The capture IgY-ELISA efficacy was of 97% when the predictive indices of sensitivity and specificity were 99.0% and 95%, respectively. The lowest and highest detection limits were of 0.02µg g-1 and 10µg.g-1 of plant extract, respectively. The strength of this method was attested by its high reproducibility (between 94.75% and 96.81%, and a low internal variation (4.22 ± 1.03%, under the described conditions.

  4. Modification of Antibody Function by Mutagenesis.

    Science.gov (United States)

    Dasch, James R; Dasch, Amy L

    2017-09-01

    The ability to "fine-tune" recombinant antibodies by mutagenesis separates recombinant antibodies from hybridoma-derived antibodies because the latter are locked with respect to their properties. Recombinant antibodies can be modified to suit the application: Changes in isotype, format (e.g., scFv, Fab, bispecific antibodies), and specificity can be made once the heavy- and light-chain sequences are available. After immunoglobulin heavy and light chains for a particular antibody have been cloned, the binding site-namely, the complementarity determining regions (CDR)-can be manipulated by mutagenesis to obtain antibody variants with improved properties. The method described here is relatively simple, uses commercially available reagents, and is effective. Using the pComb3H vector, a commercial mutagenesis kit, PfuTurbo polymerase (Agilent), and two mutagenic primers, a library of phage with mutagenized heavy and light CDR3 can be obtained. © 2017 Cold Spring Harbor Laboratory Press.

  5. Feeding different levels of vitamin E and selenium has no effect on serum immunoglobulin Y (IgY production by layers vaccinated against Escherichia coli and avian encephalomyelitis virus Alimentação com diferentes níveis de vitamina E e selênio não influencia a produção de imunoglobulina Y (IgY no soro de poedeiras leves vacinadas contra Escherichia coli e encefalomielite aviária

    Directory of Open Access Journals (Sweden)

    Giselle Kindlein

    2007-10-01

    Full Text Available The effects of vitamin E and selenium (Se supplementation on the immunity of hens vaccinated against a mixture of six swine-pathogenic Escherichia coli (EC and avian encephalomyelitis virus (AEV were studied. Antibody production (AbP was evaluated in ninety 49 to 57-week-old H&N Nick Chick hens fed diets containing 14IU Vitamin E kg-1 (basal diet, 27, 59, 111, or 111IU vitamin E kg-1 + 0.56ppm Se supplementation. At 51 wks of age, half of the hens were vaccinated against EC, and all birds were vaccinated against AEV. At 53-weeks of age, the birds received a second dose of EC vaccine. Blood samples were collected weekly and serum was analyzed by ELISA for anti-EC IgY and was expressed as optical density (OD. Vaccinated hens had higher serum OD than the non-vaccinated hens (P£0.05. Vaccinated hens fed 27 and 59IU of vitamin E/kg had a higher (POs efeitos da suplementação de vitamina E e Selênio (Se na imunidade de galinhas vacinadas contra uma mistura de 6 sorotipos patogênicos de Escherichia coli (EC e o vírus da encefalomielite aviária (VEA foram estudados. A produção de anticorpos foi avaliada em galinhas H&N Nick Chick durante a 49a e 57a semanas de vida. As aves foram alimentadas com dietas contendo 14UI de Vitamina E kg-1 (dieta basal, 27, 59, 111 e 111UI de Vitamina E kg-1 + 0,56ppm Se suplementar. Às 51 semanas de idade, metade das galinhas foi vacinada contra EC, e todas as aves foram vacinadas contra VEA. Às 53 semanas, as aves receberam a segunda vacina contra EC. Amostras de sangue foram coletadas semanalmente e o soro foi analisado por ELISA para anti-EC IgY e expresso como densidade óptica (DO. Galinhas vacinadas tiveram maior DO do que as não-vacinadas (P<0,05. Aves vacinadas que receberam 27 e 59 UI de vitamina E/kg tiveram maior DO do soro (P<0,05 do que as alimentadas com 111 UI + Se. Os antígenos utilizados mostraram não ser modelos satisfatórios para estudar a influência de micronutrientes na resposta imune de

  6. Evaluation of Chicken IgY Generated Against Canine Parvovirus Viral-Like Particles and Development of Enzyme-Linked Immunosorbent Assay and Immunochromatographic Assay for Canine Parvovirus Detection.

    Science.gov (United States)

    He, Jinxin; Wang, Yuan; Sun, Shiqi; Zhang, Xiaoying

    2015-11-01

    Immunoglobulin Y (IgY) antibodies were generated against canine parvovirus virus-like particles (CPV-VLPs) antigen using chickens. Anti-CPV-VLPs-IgY was extracted from hen egg yolk and used for developing enzyme-linked immunosorbent assay (ELISA) and immunochromatographic assay (ICA) for the detection of CPV in dog feces. The cutoff negative values for anti-CPV-VLPs-IgY were determined using negative fecal samples (already confirmed by polymerase chain reaction [PCR]). In both ELISA and ICA, there was no cross-reaction with other diarrheal pathogens. Thirty-four fecal samples were collected from dogs with diarrhea, of which 26.47% were confirmed as CPV-positive samples by PCR, while 29.41% and 32.35% of the samples were found to be positive by ELISA and ICA, respectively. The developed ELISA and ICA exhibited 97.06% and 94.12% conformity with PCR. Higher sensitivity and specificity were observed for IgY-based ELISA and ICA. Thus, they could be suitable for routine use in the diagnosis of CPV in dogs.

  7. Site-Specific Antibody Functionalization Using Tetrazine-Styrene Cycloaddition.

    Science.gov (United States)

    Umlauf, Benjamin J; Mix, Kalie A; Grosskopf, Vanessa A; Raines, Ronald T; Shusta, Eric V

    2018-05-03

    Biologics, such as antibody-drug conjugates, are becoming mainstream therapeutics. Consequently, methods to functionalize biologics without disrupting their native properties are essential for identifying, characterizing, and translating candidate biologics from the bench to clinical practice. Here, we present a method for site-specific, carboxy-terminal modification of single-chain antibody fragments (scFvs). ScFvs displayed on the surface of yeast were isolated and functionalized by combining intein-mediated expressed protein ligation (EPL) with inverse electron-demand Diels-Alder (IEDDA) cycloaddition using a styrene-tetrazine pair. The high thiol concentration required to trigger EPL can hinder the subsequent chemoselective ligation reactions; therefore, the EPL reaction was used to append styrene to the scFv, limiting tetrazine exposure to damaging thiols. Subsequently, the styrene-functionalized scFv was reacted with tetrazine-conjugated compounds in an IEDDA cycloaddition to generate functionalized scFvs that retain their native binding activity. Rapid functionalization of yeast surface-derived scFv in a site-directed manner could find utility in many downstream laboratory and preclinical applications.

  8. Detecting Lyme disease using antibody-functionalized carbon nanotubes

    Science.gov (United States)

    Dailey, Jennifer; Lerner, Mitchell; Goldsmith, Brett; Brisson, Dustin; Johnson, A. T. Charlie

    2011-03-01

    We combine antibodies for Lyme flagellar protein with carbon nanotube transistors to create an electronic sensor capable of definitive detection of Lyme disease. Over 35,000 cases of Lyme disease are reported in the United States each year, of which more than 23 percent are originally misdiagnosed. Rational design of the coupling of the biological system to the electronic system gives us a flexible sensor platform which we can apply to several biological systems. By coupling these antibodies to carbon nanotubes in particular, we allow for fast, sensitive, highly selective, electronic detection. Unlike antibody or biomarker detection, bacterial protein detection leads to positive identification of both early and late stage bacterial infections, and is easily expandable to environmental monitoring.

  9. IGY: A New Model for LGBTQ Youth Groups?

    Science.gov (United States)

    Brooks, Orren

    2017-01-01

    This study explores the experiences of homosexual teenagers who are members of the Israeli lesbian, gay, bisexual, transgender, and queer youth organization called Israel Gay Youth. Through an interview-based study with members and counselors, this study reveals how although various therapeutic practices take place during the weekly meetings, all…

  10. (IgY- antibodies) prepared against Walterinnesia aegyptia snake ...

    African Journals Online (AJOL)

    Four groups of eight chickens were immunized intramuscularly with Walterinnesia aegyptia snake venoms mixed with Freund's complete adjuvant during the period from 1st October 2009 to 1st October 2011 at the Center of Excellence in Biotechnology Research, King Saud University, Saudi Arabia. Three weeks later, the ...

  11. Antibody-functionalized porous silicon nanoparticles for vectorization of hydrophobic drugs.

    Science.gov (United States)

    Secret, Emilie; Smith, Kevin; Dubljevic, Valentina; Moore, Eli; Macardle, Peter; Delalat, Bahman; Rogers, Mary-Louise; Johns, Terrance G; Durand, Jean-Olivier; Cunin, Frédérique; Voelcker, Nicolas H

    2013-05-01

    We describe the preparation of biodegradable porous silicon nanoparticles (pSiNP) functionalized with cancer cell targeting antibodies and loaded with the hydrophobic anti-cancer drug camptothecin. Orientated immobilization of the antibody on the pSiNP is achieved using novel semicarbazide based bioconjugate chemistry. To demonstrate the generality of this targeting approach, the three antibodies MLR2, mAb528 and Rituximab are used, which target neuroblastoma, glioblastoma and B lymphoma cells, respectively. Successful targeting is demonstrated by means of flow cytometry and immunocytochemistry both with cell lines and primary cells. Cell viability assays after incubation with pSiNPs show selective killing of cells expressing the receptor corresponding to the antibody attached on the pSiNP. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Antibody-Functionalized Carbon Nanotube Transistors as Biosensors for the Detection of Prostate Cancer

    Science.gov (United States)

    2013-09-01

    NaOH . Note: Although the elution of the His-tagged protein from the magnetic beads was not 100% efficient, we obtained significant amounts of protein...ice. Cells were resuspended (2.5 mL per gram cells) in ice-cold PBS (Biowhittaker, without magnesium or calcium) containing Roche complete protease...NaH2PO4, 0.15 M NaCl, pH adjusted to 7.35 with NaOH ) for 2 h. Upon completion, the chipswerewashedwithwater and placed in a solution of 11.3mM NiCl2

  13. A33 antibody-functionalized exosomes for targeted delivery of doxorubicin against colorectal Cancer.

    Science.gov (United States)

    Li, Yan; Gao, Yuan; Gong, Chunai; Wang, Zhuo; Xia, Qingming; Gu, Fenfen; Hu, Chuling; Zhang, Lijuan; Guo, Huiling; Gao, Shen

    2018-06-20

    Exosomes have emerged as a promising drug carrier with low immunogenicity, high biocompatibility and delivery efficiency. Here in, we isolated exosomes from A33-positive LIM1215 cells (A33-Exo) and loaded them with doxorubicin (Dox). Furthermore, we coated surface-carboxyl superparamagnetic iron oxide nanoparticles (US) with A33 antibodies (A33Ab-US), expecting that these A33 antibodies on the surface of the nanoparticles could bind to A33-positive exosomes and form a complex (A33Ab-US-Exo/Dox) to target A33-positive colon cancer cells. The results showed that A33Ab-US-Exo/Dox had good binding affinity and antiproliferative effect in LIM1215 cells, as shown by increased uptake of the complex. In vivo study showed that A33Ab-US-Exo/Dox had an excellent tumor targeting ability, and was able to inhibit tumor growth and prolong the survival of the mice with reduced cardiotoxicity. In summary, exosomes functionalized by targeting ligands through coating with high-density antibodies may prove to be a novel delivery system for targeted drugs against human cancers. Copyright © 2018 Elsevier Inc. All rights reserved.

  14. Antibody-Functionalized Carbon Nanotube Transistors as Biosensors for the Detection of Prostate Cancer

    Science.gov (United States)

    2010-07-01

    the attachment by AFM and electronic measurement (Fig. 1.2). We have also begun experiments to quantify the sensor response to solutions of OPN...to facilitate IMAC purification. Each of the sc Fv were expressed, purified, and binding activity ch aracterized via surface plasm on resonance (SPR

  15. Integrated geographic information systems (IGIS) analysis and definition of the tectonic framework of northern Mexico

    Science.gov (United States)

    Martinez Pina, Carlos Manuel

    Crustal rupture structures reactivated in the course of the tectonic history of northern Mexico are the surface expressions of planes of weakness, in the form of simple or composite rectilinear features or slightly curved, defined as lineaments. Unless otherwise defined as strike-slip faults, lineaments are part of parallel and sub-parallel oblique convergent or oblique divergent tectonic zones cross cutting the Sierra Madre Occidental and northern Mexico, in a NW trend. These shear zones are the response to the oblique subduction of the Farallon plate beneath North America. Kinematic analysis of five selected sites in northern Mexico, three basins and two compressional shear zones, proved possible a combination of shear mechanism diagram and models from analogue materials, with satellite imagery and geographic information systems, as an aid to define strike-slip fault motion. This was done using a reverse engineering process by comparing geometries. One of the sites assessed, involving the Parras Basin, Coahuila Block (CB), San Marcos fault, a postulated PBF-1 fault, allowed for palinpastic reconstruction of the CB that corroborated the results of the vector motion defined, in addition to an extension of ˜25% in a northwest southeast direction. A GIS-based compilation and georeferenced regional structural studies by several researchers were used as ground control areas (GCA); their interpolation and interpretation, resulted in a tectonic framework map of northern Mexico. In addition, shaded relief models overlaid by the lineaments / fault layer allowed structural analyses of basins related to these major structures. Two important results were obtained from this study: the Tepehuanes-San Luis-fault (TSL) and the Guadalupe fault, named herein, displaces the Villa de Reyes graben, and the Aguascalientes graben, respectively, to the SE, confirming their left lateral vector motion; afterwards TSL was displaced south by the right lateral strike slip Taxco-San Miguel de Allende fault. The second result refers to the hypothesis that the Mesa Central was brought to its present location by a subduction zone located to the north. This subduction zone coincides with several researchers who postulated the idea. The compressional zones refer to segments of the Sinforosa and a postulated Aquinquari fault located in the stratotectonic Guerrero Terrane regarded as a highly mineralized zone. Negative anomalies near -200 milligals are strongly suggestive of a cratonic block identified in western Chihuahua, it being named the Western Chihuahua Cratonic Block (WCCB). In the southwestern portion of the North American craton the age provinces are well documented, but the block versus mobile belt idea has not been put forth or emphasized. The present study combines data of several types, sedimentological, structural, igneous geochemistry, and geochronologic data to evaluate this behavior in SW NA, and the proposed block is tested against these data. The presence of the WCCB is supported by a wide variety of data. Basins, troughs, aulacogens, bimodal volcanism, and other rift and rift shoulder features, characterize the spatially constrained mobile belts. Mobile belts surrounding the WCCB contain geologic records of the events going back to 1.4 Ga, with different aspects being dominant over geologic time. Mobile belts will participate in compression,(subduction), extension (rifting), and transform (lateral) faulting. The WCCB may have been derived from closely, adjacent, North American craton by mobile belt action. This study has shown that integration of data is essential, because allows detection of differences in hypotheses for the same event in the same area. This integration capability is what makes integrated geographic information systems a powerful tool, not only for their synergy, but because they can be combined with specific techniques that provide data before going to conduct fieldwork. Whether the issue of defining the tectonic framework of northern Mexico can be resolved or not, depends on the viability of integrating volumes of data from research, hypotheses, or maps, and put together under the same geographic frame.

  16. Optimized protocol of chicken antibody (IgY) purification providing electrophoretically homogenous preparations

    Czech Academy of Sciences Publication Activity Database

    Hodek, P.; Trefil, P.; Šimůnek, Jiří; Hudeček, J.; Stiborová, M.

    2013-01-01

    Roč. 8, č. 1 (2013), 113-124 ISSN 1452-3981 Grant - others:GA ČR(CZ) GAP303/12/G163 Program:GA Institutional support: RVO:67985904 Keywords : yolk immunoglobulin * purification protocol * bio-compatible chemicals Subject RIV: CG - Electrochemistry Impact factor: 1.956, year: 2013

  17. Quantitative immunochemistry using IgG and IgY against plant ...

    African Journals Online (AJOL)

    Viviane

    2Department of Sciences, College of Teacher Education – UERJ, Rua Dr. Francisco Portela ... Janeiro (Universidade Federal Rural do Rio de Janeiro – UFRRJ), BR 465 km 07, 23890-000, Seropédica - RJ – Brazil. .... The clot that was formed.

  18. Significance of United Nations Conference on Environment and Development (UNCED). Chikyu summit no igi

    Energy Technology Data Exchange (ETDEWEB)

    Shimizu, T [Ministry of Construction, Tokyo (Japan)

    1992-10-01

    On the significance of UNCED, the background, the results of the conference and the reflection to the construction administration were described. The causes of global environmental problems such as the global warming, the ozone layer destruction and the decrease of forests can mainly be classified into 3. These are the problem of socio-economic activities of advanced countries such as mass production, mass consumption and mass dumping, the problem originating from the poverty in developing countries and the problem originating from the interests between advanced countries and developing countries. The main results in the UNCED were as follows: the signature to the two conventions as to the climate change and the biological diversity, the statement as to the principle for forests, the Rio Declaration which should be said to be the world constitution concerning the environment and development, and the adoption of the Agenda 21 of the draft which summarized the policies and measures. The Agenda 21 includes the fund, the technology transfer and the organization problems. In order to reflect these problems to the construction administration, it is necessary to promote the energy policy, the establishment of efficient traffic system, the effective land utilization, the appropriate management of river spaces, the urban policy and the recycling of resources.

  19. Lower antibody functionality in middle-aged adults compared to adolescents after primary meningococcal vaccination : Role of IgM

    NARCIS (Netherlands)

    van der Heiden, Marieke; van Ravenhorst, Mariette B; Bogaard, Marjan; Boots, Annemieke M H; Berbers, Guy A M; Buisman, Anne-Marie

    Introduction: Successful vaccination of elderly persons is often hampered by immunological ageing, leaving part of the elderly population vulnerable for infectious diseases. As an alternative, timely vaccinations might be administered at middle-age, before reaching old age. Studies evaluating the

  20. Clonal relationships between thyroid-stimulating hormone receptor-stimulating antibodies illustrate the effect of hypermutation on antibody function

    DEFF Research Database (Denmark)

    Padoa, Carolyn J; Larsen, Sanne L; Hampe, Christiane S

    2009-01-01

    relationship and derivation from a single precursor B-cell clone. The IGHV-region genes of the two mAbs underwent high degrees of somatic hypermutation by sharing numerous mutations before diverging, while the IGLV genes evolved separately. Interestingly, the mutations were present in both the complementarity......, in experimentally immunized mice, multiple pathogenic antibodies to TSHR can arise from a single clone by a series of somatic mutations in the V-region genes and may give an insight into how such antibodies develop spontaneously in autoimmune Graves' disease....

  1. Elaboration of Stable and Antibody Functionalized Positively Charged Colloids by Polyelectrolyte Complexation between Chitosan and Hyaluronic Acid

    Directory of Open Access Journals (Sweden)

    Ramona C. Polexe

    2013-07-01

    Full Text Available In this study, we describe the elaboration of multifunctional positively charged polyelectrolyte complex (PEC nanoparticles, designed to be stable at physiological salt concentration and pH, for effective targeted delivery. These nanoparticles were obtained by charge neutralization between chitosan (CS as polycation and hyaluronic acid (HA as polyanion. We showed that the course of the complexation process and the physico-chemical properties of the resulting colloids were impacted by (i internal parameters such as the Degree of Acetylation (DA, i.e., the molar ration of acetyl glucosamine residues and molar mass of CS, the HA molar mass and (ii external parameters like the charge mixing ratio and the polymer concentrations. As a result, nonstoichiometric colloidal PECs were obtained in water or PBS (pH 7.4 and remained stable over one month. The polymer interactions were characterized by thermal analysis (DSC and TGA and the morphology was studied by scanning electron microscopy. A model antibody, anti-ovalbumine (OVA immunoglobulin A (IgA was sorbed on the particle surface in water and PBS quantitatively in 4 h. The CS-HA/IgA nanoparticles average size was between 425–665 nm with a positive zeta potential. These results pointed out that CS-HA can be effective carriers for use in targeted drug delivery.

  2. Selection of a novel anti-nicotine vaccine: influence of antigen design on antibody function in mice.

    Directory of Open Access Journals (Sweden)

    David C Pryde

    Full Text Available Anti-nicotine vaccines may aid smoking cessation via the induction of anti-nicotine antibodies (Ab which reduce nicotine entering the brain, and hence the associated reward. Ab function depends on both the quantity (titer and the quality (affinity of the Ab. Anti-nicotine vaccines tested previously in clinical studies had poor efficacy despite high Ab titer, and this may be due to inadequate function if Ab of low affinity were induced. In this study, we designed and synthesized a series of novel nicotine-like haptens which were all linked to diphtheria toxoid (DT as carrier, but which differed in the site of attachment of linker to nicotine, the nature of linker used, and the handle used to attach the hapten to DT. The resulting hapten conjugates were evaluated in a mouse model, using CpG (a TLR9 agonist and aluminum hydroxide (Al(OH3 as adjuvants, whereby Ab titers, affinity and function were evaluated using a radiolabeled nicotine challenge model. A series of additional linkers varying in length, rigidity and polarity were used with a single hapten to generate additional DT-conjugates, which were also tested in mice. Conjugates made with different haptens resulted in various titers of anti-nicotine Ab. Several haptens gave similarly high Ab titers, but among these, Ab affinity and hence function varied considerably. Linker also influenced Ab titer, affinity and function. These results demonstrate that immune responses induced in mice by nicotine-conjugate antigens are greatly influenced by hapten design including site of attachment of linker to nicotine, the nature of linker used, and the handle used to attach the hapten to DT. While both Ab titer and affinity contributed to function, affinity was more sensitive to antigen differences.

  3. Antibody functionalized graphene biosensor for label-free electrochemical immunosensing of fibrinogen, an indicator of trauma induced coagulopathy.

    Science.gov (United States)

    Saleem, Waqas; Salinas, Carlos; Watkins, Brian; Garvey, Gavin; Sharma, Anjal C; Ghosh, Ritwik

    2016-12-15

    An antibody, specific to fibrinogen, has been covalently attached to graphene and deposited onto screen printed electrodes using a chitosan hydrogel binder to prepare an inexpensive electrochemical fibrinogen biosensor. Fourier Transform Infrared (FT-IR) spectroscopy has been utilized to confirm the presence of the antibody on the graphene scaffold. Electrochemical Impedance Spectroscopy (EIS) has been utilized to demonstrate that the biosensor responds in a selective manner to fibrinogen in aqueous media even in the presence of plasminogen, a potentially interfering molecule in the coagulopathy cascade. Furthermore, the biosensor was shown to reliably sense fibrinogen in the presence of high background serum albumin levels. Finally, we demonstrated detection of clinically relevant fibrinogen concentrations (938-44,542μg/dL) from human serum and human whole blood samples using this biosensor. This biosensor can potentially be used in a point-of-care device to detect the onset of coagulopathy and monitor response following therapeutic intervention in trauma patients. Thus this biosensor may improve the clinical management of patients with trauma-induced coagulopathy. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. A novel activating chicken IgY FcR is related to leukocyte receptor complex (LRC)

    NARCIS (Netherlands)

    Viertlboeck, B.C.; Schmitt, R.; Hanczaruk, M.A.; Crooijmans, R.P.M.A.; Groenen, M.A.M.; Gobel, T.W.

    2009-01-01

    FcRs have multifaceted roles in the immune system. Chicken FcRs were demonstrated on macrophages decades ago; however, only recently the chicken Ig-like receptor AB1, encoded in the leukocyte receptor complex, was molecularly identified as a high-affinity FcR. The present study was initiated to

  5. Insights into the chicken IgY with emphasis on the generation and applications of chicken recombinant monoclonal antibodies.

    Science.gov (United States)

    Lee, Warren; Syed Atif, Ali; Tan, Soo Choon; Leow, Chiuan Herng

    2017-08-01

    The advantages of chicken (Gallus gallus domesticus) antibodies as immunodiagnostic and immunotherapeutic biomolecules has only been recently recognized. Even so, chicken antibodies remain less-well characterized than their mammalian counterparts. This review aims at providing a current overview of the structure, function, development and generation of chicken antibodies. Additionally, brief but comprehensive insights into current knowledge pertaining to the immunogenetic framework and diversity-generation of the chicken immunoglobulin repertoire which have contributed to the establishment of recombinant chicken mAb-generating methods are discussed. Focus is provided on the current methods used to generate antibodies from chickens with added emphasis on the generation of recombinant chicken mAbs and its derivative formats. The advantages and limitations of established protocols for the generation of chicken mAbs are highlighted. The various applications of recombinant chicken mAbs and its derivative formats in immunodiagnostics and immunotherapy are further detailed. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Production, Characterization and Use of Monoclonal Antibodies Recognizing IgY Epitopes Shared by Chicken, Turkey, Pheasant, Peafowl and Sparrow

    OpenAIRE

    Narat, Mojca; Biček, Ajda; Vadnjal, Robert; Benčina, Dušan

    2004-01-01

    Chicken antibodies are not only a part of immune defense but are more and more popular commercial products in form of chicken polyclonal, monoclonal or recombinant antibodies. We produced and characterized mouse monoclonal antibodies (mAbs) that recognize epitopes located on heavy or light chain of chicken immunoglobulin Y (chIgY) shared also by some other Phasianidae birds. The use of mAbs 1F5 and 2F10 that recognize heavy chain on chIgY common epitopes was demonstrated on immunoglobulins of...

  7. Roles played by coral reef ecosystems in mitigating global warming. Ondanka taisaku ni okeru sangosho no igi

    Energy Technology Data Exchange (ETDEWEB)

    Komiyama, H [The University of Tokyo, Tokyo (Japan). Faculty of Engineering

    1993-05-01

    This Paper describes briefly a question of whether growing coral reefs could be an effective means to fix CO2 in sea water in order to mitigate global warming: Growing coral reefs could be a promising method from the following three views: Photosynthesis performed by diverse ecosystems that constitute coral reefs is thought to excel calcification (accompanying CO2 discharge from sea to the atmosphere); corals all over the world would fix CO2 at an annual rate of several hundred million tons; and no much energies are consumed to fix CO2. However, these favorable views depend upon another condition that a time period for the CO2 fixation is not too short (a measure for the time being is 100 years or longer). Elucidating where organic matters generated by the photosynthesis would go is an important question to meet this requirement. The paper indicates that measuring nitrogen and phosphor balances in addition to carbon balance is effective for measuring ratio of a net photosynthesis rate to the calcification rate. 4 refs., 1 fig., 4 tabs.

  8. Microscale depletion of high abundance proteins in human biofluids using IgY14 immunoaffinity resin: analysis of human plasma and cerebrospinal fluid.

    Science.gov (United States)

    Hyung, Seok-Won; Piehowski, Paul D; Moore, Ronald J; Orton, Daniel J; Schepmoes, Athena A; Clauss, Therese R; Chu, Rosalie K; Fillmore, Thomas L; Brewer, Heather; Liu, Tao; Zhao, Rui; Smith, Richard D

    2014-11-01

    Removal of highly abundant proteins in plasma is often carried out using immunoaffinity depletion to extend the dynamic range of measurements to lower abundance species. While commercial depletion columns are available for this purpose, they generally are not applicable to limited sample quantities (depletion media can be prohibitive for larger-scale studies. Modern LC-MS instrumentation provides the sensitivity necessary to scale-down depletion methods with minimal sacrifice to proteome coverage, which makes smaller volume depletion columns desirable for maximizing sample recovery when samples are limited, as well as for reducing the expense of large-scale studies. We characterized the performance of a 346 μL column volume microscale depletion system, using four different flow rates to determine the most effective depletion conditions for ∼6-μL injections of human plasma proteins and then evaluated depletion reproducibility at the optimum flow rate condition. Depletion of plasma using a commercial 10-mL depletion column served as the control. Results showed depletion efficiency of the microscale column increased as flow rate decreased, and that our microdepletion was reproducible. In an initial application, a 600-μL sample of human cerebrospinal fluid (CSF) pooled from multiple sclerosis patients was depleted and then analyzed using reversed phase liquid chromatography-mass spectrometry to demonstrate the utility of the system for this important biofluid where sample quantities are more commonly limited.

  9. Detection in chick embryo of fetoproteins not recognized by the dam's immune system and of soluble alloantigens. Presumptive teratogenic and abortogenic capacity of their specific IgY

    Directory of Open Access Journals (Sweden)

    Rodríguez-Burgos Antonio

    2003-06-01

    Full Text Available Abstract Background The aim of this work was to detect antigens, non-self to the dam, potentially present in chick embryo prior to organogenesis with a view to establishing the consequences of their neutralization on chick development. To this end, hens were immunized with the extract from embryos incubated for 53 h. Their eggs were either used to isolate immunoglobulins for dot and blot tests or incubated for variable lengths of time. Results Immunoblot tests, using adsorbed primary and secondary antibodies against paternal serum, revealed the presence of at least four antigens of 32, 34, 70 and 200 kDa that can be classified as soluble alloantigens. The same antibodies against chick embryo extracts (between 53 h and 9 showed at least five aged antigens of 34, 52, 90, 200 and 250 kDa, not detected in cock serum, that can thus be considered as soluble, foreign to the immunized hens and transitory antigens. The abnormalities observed included arrested development and fetal death, as well as minor functional damage in the few chicks that were born alive. The ratio of abnormal to normal embryos was 2.85 in the experimental group and 0.43 in the control group. With regard to congenital anomalies it must be said that of the 81 eggs incubated only four chicks were born alive, and of these, only one had a healthy birth and subsequent growth. The other three showed a transitory ataxia and one of them presented adult lumbar scoliosis and asymmetric pelvis. Conclusions The problem of recurrent spontaneous abortions is revisited in the light of these results. Some recent data suggest that soluble alloantigens may be candidates for a new etiological entity in recurrent spontaneous abortions. They can also be the cause of some congenital anomalies. The soluble, foreign, transitory antigens may have a similar effect although there is no supportive data in the literature.

  10. Refinement of the production of antigen-specific hen egg yolk antibodies (IgY intended for passive dietary immunization in animals. A review

    Directory of Open Access Journals (Sweden)

    Marcq, C.

    2013-01-01

    Full Text Available Antibodies have become essential tools in recent decades, with a wide range of applications in the laboratory and in human and veterinary medicine. The use of laying hens, instead of mammals, to obtain the necessary antibodies from the eggs is a major advance in terms of animal welfare because it makes blood sampling obsolete. However, the advantages of this technology are numerous, in addition to the animal welfare aspect. With a carefully designed immunization protocol, it is possible to enhance both the hen's immune response and its welfare during the process. This review puts forward recommendations how to do this and discusses recent approaches on improving the technology.

  11. Detection of Shiga toxin-producing Escherichia coli by sandwich enzyme-linked immunosorbent assay using chicken egg yolk IgY antibodies

    OpenAIRE

    Parma, Y. R.; Chacana, P. A.; Lucchesi, P. M. A.; Rogé, A.; Granobles Velandia, C. V.; Krüger, A.; Parma, A. E.; Fernández-Miyakawa, M. E.

    2012-01-01

    Enterohemorrhagic Escherichia coli (EHEC), a subset of Shiga toxin producing E. coli (STEC) is associated with a spectrum of diseases that includes diarrhea, hemorrhagic colitis and a life-threatening hemolytic-uremic syndrome (HUS). Regardless of serotype, Shiga toxins (Stx1 and/or Stx2) are uniformly expressed by all EHEC, and so exploitable targets for laboratory diagnosis of these pathogens. In this study, a sandwich ELISA for determination of Shiga toxin (Stx) was developed using anti-St...

  12. Detection of Shiga toxin-producing Escherichia coli by sandwich enzyme-linked immunosorbent assay using chicken egg yolk IgY antibodies

    OpenAIRE

    Yanil R Parma; Pablo A Chacana; Paula María Alejandra Lucchesi; Ariel eRoge; Claudia V Granobles Velandia; Alejandra eKrüger; Alejandra eKrüger; Alberto E. Parma; Mariano Enrique Fernandez Miyakawa

    2012-01-01

    Enterohemorrhagic Escherichia coli (EHEC), a subset of Shiga toxin producing E. coli (STEC) is associated with a spectrum of diseases that includes diarrhea, hemorrhagic colitis and a life-threatening hemolytic uremic syndrome (HUS). Regardless of serotype, Shiga toxins (Stx1 and/or Stx2) are uniformly expressed by all EHEC, and so exploitable targets for laboratory diagnosis of these pathogens. In this study, a sandwich ELISA for determination of Shiga toxin (Stx) was developed using anti-St...

  13. Hen egg yolk antibodies (IgY), production and use for passive immunization against bacterial enteric infections in chicken: a review

    OpenAIRE

    Chalghoumi R.; Beckers Y.; Portetelle D.; Théwis A.

    2009-01-01

    Enteric infections caused by Salmonella remain a major public health burden worldwide. Poultry, particularly chickens, are known to be the main reservoir for this zoonotic pathogen. Therefore, the prevention and monitoring of Salmonella infection during the live phase may greatly reduce the contamination of poultry meat during slaughter and processing. With the ban on sub-therapeutic antibiotic usage in Europe and the increasingly strictness of the European legislation on food hygiene, passiv...

  14. (18)F-nanobody for PET imaging of HER2 overexpressing tumors.

    Science.gov (United States)

    Xavier, Catarina; Blykers, Anneleen; Vaneycken, Ilse; D'Huyvetter, Matthias; Heemskerk, Jan; Lahoutte, Tony; Devoogdt, Nick; Caveliers, Vicky

    2016-04-01

    Radiolabeled nanobodies are exciting new probes for molecular imaging due to high affinity, high specificity and fast washout from the blood. Here we present the labeling of an anti-HER2 nanobody with (18)F and its validation for in vivo assessment of HER2 overexpression. The GMP grade anti-HER2 nanobody was labeled with the prosthetic group, N-succinimidyl-4-[(18)F]fluorobenzoate ([(18)F]-SFB), and its biodistribution, tumor targeting and specificity were evaluated in mouse and rat tumor models. [(18)F]FB-anti-HER2 nanobody was prepared with a 5-15% global yield (decay corrected) and a specific activity of 24.7 ± 8.2 MBq/nmol. In vivo studies demonstrated a high specific uptake for HER2 positive xenografts (5.94 ± 1.17 and 3.74 ± 0.52%IA/g, 1 and 3h p.i.) with high tumor-to-blood and tumor-to-muscle ratios generating high contrast PET imaging. The probe presented fast clearance through the kidneys (4%IA/g at 3h p.i.). [(18)F]FB-anti-HER2 nanobody is able to image HER2 expressing tumors when co-administered with the anti-HER2 therapeutic antibody trastuzumab (Herceptin), indicating the possibility of using the tracer in patients undergoing Herceptin therapy. The GMP grade anti-HER2 nanobody was labeled with (18)F. This new PET probe for imaging HER2 overexpression in tumors has ample potential for clinical translation. Copyright © 2016 Elsevier Inc. All rights reserved.

  15. 18F-nanobody for PET imaging of HER2 overexpressing tumors

    International Nuclear Information System (INIS)

    Xavier, Catarina; Blykers, Anneleen; Vaneycken, Ilse; D'Huyvetter, Matthias; Heemskerk, Jan; Lahoutte, Tony; Devoogdt, Nick; Caveliers, Vicky

    2016-01-01

    Introduction: Radiolabeled nanobodies are exciting new probes for molecular imaging due to high affinity, high specificity and fast washout from the blood. Here we present the labeling of an anti-HER2 nanobody with 18 F and its validation for in vivo assessment of HER2 overexpression. Methods: The GMP grade anti-HER2 nanobody was labeled with the prosthetic group, N-succinimidyl-4-[ 18 F]fluorobenzoate ([ 18 F]-SFB), and its biodistribution, tumor targeting and specificity were evaluated in mouse and rat tumor models. Results: [ 18 F]FB-anti-HER2 nanobody was prepared with a 5–15% global yield (decay corrected) and a specific activity of 24.7 ± 8.2 MBq/nmol. In vivo studies demonstrated a high specific uptake for HER2 positive xenografts (5.94 ± 1.17 and 3.74 ± 0.52%IA/g, 1 and 3 h p.i.) with high tumor-to-blood and tumor-to-muscle ratios generating high contrast PET imaging. The probe presented fast clearance through the kidneys (4%IA/g at 3 h p.i.). [ 18 F]FB-anti-HER2 nanobody is able to image HER2 expressing tumors when co-administered with the anti-HER2 therapeutic antibody trastuzumab (Herceptin), indicating the possibility of using the tracer in patients undergoing Herceptin therapy. Conclusions: The GMP grade anti-HER2 nanobody was labeled with 18 F. This new PET probe for imaging HER2 overexpression in tumors has ample potential for clinical translation.

  16. An anti-CD34 antibody-functionalized clinical-grade POSS-PCU nanocomposite polymer for cardiovascular stent coating applications: a preliminary assessment of endothelial progenitor cell capture and hemocompatibility.

    Directory of Open Access Journals (Sweden)

    Aaron Tan

    Full Text Available In situ endothelialization of cardiovascular implants has emerged in recent years as an attractive means of targeting the persistent problems of thrombosis and intimal hyperplasia. This study aimed to investigate the efficacy of immobilizing anti-CD34 antibodies onto a POSS-PCU nanocomposite polymer surface to sequester endothelial progenitor cells (EPCs from human blood, and to characterize the surface properties and hemocompatibility of this surface. Amine-functionalized fumed silica was used to covalently conjugate anti-CD34 to the polymer surface. Water contact angle, fluorescence microscopy, and scanning electron microscopy were used for surface characterization. Peripheral blood mononuclear cells (PBMCs were seeded on modified and pristine POSS-PCU polymer films. After 7 days, adhered cells were immunostained for the expression of EPC and endothelial cell markers, and assessed for the formation of EPC colonies. Hemocompatibility was assessed by thromboelastography, and platelet activation and adhesion assays. The number of EPC colonies formed on anti-CD34-coated POSS-PCU surfaces was not significantly higher than that of POSS-PCU (5.0±1.0 vs. 1.7±0.6, p>0.05. However, antibody conjugation significantly improved hemocompatibility, as seen from the prolonged reaction and clotting times, decreased angle and maximum amplitude (p<0.05, as well as decreased platelet adhesion (76.8±7.8 vs. 8.4±0.7, p<0.05 and activation. Here, we demonstrate that POSS-PCU surface immobilized anti-CD34 antibodies selectively captured CD34+ cells from peripheral blood, although only a minority of these were EPCs. Nevertheless, antibody conjugation significantly improves the hemocompatibility of POSS-PCU, and should therefore continue to be explored in combination with other strategies to improve the specificity of EPC capture to promote in situ endothelialization.

  17. Kinetika Immunoglobulin Kuning Telur Antiparvovirus Anjing Pada Anjing (KINETICS OF ANTICANINE PARVOVIRUS YOLK IMMUNOGLOBULIN IN DOGS

    Directory of Open Access Journals (Sweden)

    I Gusti Ayu Agung Suartini

    2016-07-01

    Full Text Available Kinetic study on Anti CPV IgY has been performed on six dogs aged 5-10 months. The IgY was injectedintravenously at dose of 21.4mg /10kg body weight. IgY levels in the blood were determined by ELISA. Aresearch was conducted to find out the kinetics of Anti CPV IgY in dogs blood. The kinetics of IgY wascalculated by using regression analysis to determine the association on the levels of IgY in serum againsttime at injection. The results showed that kinetic parameters were calculated based on first order kinetics.The constant elimination rate of IgY was at the range between 0.007 to 0.015 / h. IgY concentration in thedogs blood was from 0.746 to 0.992 mg / mL. The half-life of IgY was from 1.65 to 4.01 / d. Volumedistribution of IgY was between 21.47 to 28,55 / mL. Total IgY in the dog bodies (AUC was from 42,60 to142,00 mg / mL.h. The duration of the IgY in the dog’s body was 3.08 to 8.51 days. Clearance time of IgY was0.15 to 0.50 mL / h. In conclusion the kinetics of anti CPV IgY in dog’s body follow one compartment andfirst order model, which are only distributed in the blood with the half-life at 2.5 days, and IgY has lesspossibility to accumulate in the body compared to the IgG.

  18. Immunohistochemical detection of Her-2/neu overexpression in ...

    African Journals Online (AJOL)

    Materials and Methods: Immunohistochemical staining for Her-2/neu was performed on 10% formalin-fixed, paraffinembedded primary carcinoma of the breast from 83 patients, between 2003 and 2007 using anti-Her-2/neu rabbit polyclonal antibody (DakoCytomation, CA, USA) and reactivity detected by an avidin-biotin ...

  19. Validation of the 4B5 rabbit monoclonal antibody in determining Her2/neu status in breast cancer

    NARCIS (Netherlands)

    van der Vegt, Bert; de Bock, G.H.; Bart, J.; Zwartjes, N.G.; Wesseling, J.

    HER2 overexpression in breast cancer is associated with worse clinical outcome. To select patients for anti-Her2-based therapy immunohistochemistry is commonly performed as a first step to assess Her2 status. However, interobserver and interlaboratory variability can significantly compromise

  20. Combination therapies for the treatment of HER2-positive breast cancer: current and future prospects.

    Science.gov (United States)

    Brandão, Mariana; Pondé, Noam F; Poggio, Francesca; Kotecki, Nuria; Salis, Mauren; Lambertini, Matteo; de Azambuja, Evandro

    2018-05-24

    HER2-positive disease is an aggressive subtype of breast cancer that has been revolutionized by anti-HER2 directed therapies. Multiple drugs have been developed and are currently in clinical use, including trastuzumab, lapatinib, pertuzumab, T-DM1, and neratinib, alone or combined in 'dual HER2-blockade' regimens. Areas covered: A comprehensive literature review was performed regarding the current state and the future of combination regimens containing anti-HER2 agents, focusing on their efficacy, toxicity, and cost-effectiveness. Expert commentary: The combination of trastuzumab/pertuzumab is approved in all disease settings, while trastuzumab/neratinib is approved in the adjuvant setting and trastuzumab/lapatinib in metastatic disease. Meanwhile, as breast cancer biology and resistance mechanisms become clearer, combinations with drugs like PI3K/Akt/mTOR inhibitors, CDK4/6 inhibitors, anti-PD(L)1 antibodies, endocrine therapy, and new anti-HER2 agents (panHER and HER2 tyrosine kinase inhibitors, bispecific antibodies, anti-HER3 antibodies, and antibody-drug conjugates) are being extensively tested in clinical trials. More specific strategies for the 'triple-positive' (estrogen receptor-positive/HER2-positive) disease are also being explored. However, there is an urgent need for the development of predictive biomarkers for a better tailoring of anti-HER2 directed therapy. This is the only way to further improve clinical outcomes and quality of life and to decrease costs and toxicities of unnecessary treatments.

  1. A gene expression signature of Retinoblastoma loss-of-function predicts resistance to neoadjuvant chemotherapy in ER-positive/HER2-positive breast cancer patients.

    Science.gov (United States)

    Risi, Emanuela; Grilli, Andrea; Migliaccio, Ilenia; Biagioni, Chiara; McCartney, Amelia; Guarducci, Cristina; Bonechi, Martina; Benelli, Matteo; Vitale, Stefania; Biganzoli, Laura; Bicciato, Silvio; Di Leo, Angelo; Malorni, Luca

    2018-07-01

    HER2-positive (HER2+) breast cancers show heterogeneous response to chemotherapy, with the ER-positive (ER+) subgroup deriving less benefit. Loss of retinoblastoma tumor suppressor gene (RB1) function has been suggested as a cardinal feature of breast cancers that are more sensitive to chemotherapy and conversely resistant to CDK4/6 inhibitors. We performed a retrospective analysis exploring RBsig, a gene signature of RB loss, as a potential predictive marker of response to neoadjuvant chemotherapy in ER+/HER2+ breast cancer patients. We selected clinical trials of neoadjuvant chemotherapy ± anti-HER2 therapy in HER2+ breast cancer patients with available information on gene expression data, hormone receptor status, and pathological complete response (pCR) rates. RBsig expression was computed in silico and correlated with pCR. Ten studies fulfilled the inclusion criteria and were included in the analysis (514 patients). Overall, of 211 ER+/HER2+ breast cancer patients, 49 achieved pCR (23%). The pCR rate following chemotherapy ± anti-HER2 drugs in patients with RBsig low expression was significantly lower compared to patients with RBsig high expression (16% vs. 30%, respectively; Fisher's exact test p = 0.015). The area under the ROC curve (AUC) was 0.62 (p = 0.005). In the 303 ER-negative (ER-)/HER2+ patients treated with chemotherapy ± anti-HER2 drugs, the pCR rate was 43%. No correlation was found between RBsig expression and pCR rate in this group. Low expression of RBsig identifies a subset of ER+/HER2+ patients with low pCR rates following neoadjuvant chemotherapy ± anti-HER2 therapy. These patients may potentially be spared chemotherapy in favor of anti-HER2, endocrine therapy, and CDK 4/6 inhibitor combinations.

  2. Specific egg yolk immunoglobulin as a new preventive approach for Shiga-toxin-mediated diseases.

    Directory of Open Access Journals (Sweden)

    Paola Neri

    Full Text Available Shiga toxins (Stxs are involved in the development of severe systemic complications associated with enterohemorrhagic Escherichia coli (EHEC infection. Various neutralizing agents against Stxs are under investigation for management of EHEC infection. In this study, we immunized chickens with formalin-inactivated Stx-1 or Stx-2, and obtained immunoglobulin Y (IgY from the egg yolk. Anti-Stx-1 IgY and anti-Stx-2 IgY recognized the corresponding Stx A subunit and polymeric but not monomeric B subunit. Anti-Stx-1 IgY and anti-Stx-2 IgY suppressed the cytotoxicity of Stx-1 and Stx-2 to HeLa 229 cells, without cross-suppressive activity. The suppressive activity of these IgY was abrogated by pre-incubation with the corresponding recombinant B subunit, which suggests that the antibodies directed to the polymeric B subunits were predominantly involved in the suppression. In vivo, the intraperitoneal or intravenous administration of these IgY rescued mice from death caused by intraperitoneal injection of the corresponding toxin at a lethal dose. Moreover, oral administration of anti-Stx-2 IgY reduced the mortality of mice infected intestinally with EHEC O157:H7. Our results therefore suggest that anti-Stx IgY antibodies may be considered as preventive agents for Stx-mediated diseases in EHEC infection.

  3. Durable Clinical Benefit of Pertuzumab in a Young Patient with BRCA2 Mutation and HER2-Overexpressing Breast Cancer Involving the Brain

    Directory of Open Access Journals (Sweden)

    Anna Koumarianou

    2016-01-01

    Full Text Available Patients with HER2-positive breast cancer and brain metastases have limited treatment options, and, as a result of their poor performance status and worse prognosis, they are underrepresented in clinical trials. Not surprisingly, these patients may not be fit enough to receive any active treatment and are offered supportive therapy. BRCA2 mutations are reported to be rarely associated with HER2-overexpressing advanced breast cancer and even more rarely with brain metastases at diagnosis. We report on a BRCA2-positive breast cancer patient with metastatic disease in multiple sites, including the brain, and poor performance status who exhibited an extraordinary clinical and imaging response to the novel anti-HER2 therapy pertuzumab after multiple lines of therapy including anti-HER2 targeting. To our knowledge, the clinicopathologic and therapeutic characteristics of this patient point to a unique case and an urgent need for further investigation of pertuzumab in patients with brain metastases.

  4. Adjuverende medicinsk behandling tilpatienter med lokoregionalt recidiv af brystkræft

    DEFF Research Database (Denmark)

    Farooq, Farah Choudary; Kamby, Claus

    2016-01-01

    randomized trials of adjuvant medical treatment with chemotherapy, anti-hormonal and anti HER2 treatments for patients radically treated for ILRR. Certain groups of patients may benefit from adjuvant systemic treatment. However larger randomized trials are needed.......There is 10-30% risk of developing isolated locoregional recurrence (ILRR) after mastectomy for primary breast cancer. Currently, there is no standard treatment for ILRR and therefore patients with ILRR cause an oncological task. This review investigates existing literature concerning relevant...

  5. Mass spectrometry characterization for N-glycosylation of immunoglobulin Y from hen egg yolk.

    Science.gov (United States)

    Sheng, Long; He, Zhenjiao; Liu, Yaping; Ma, Meihu; Cai, Zhaoxia

    2018-03-01

    Immunoglobulin Y (IgY) is a new therapeutic antibody that exists in hen egg yolk. It is a glycoprotein, not much is known about its N-glycan structures, site occupancy and site-specific N-glycosylation. In this study, purified protein from hen egg yolk was identified as IgY based on SDS-PAGE and MALDI-TOF/TOF MS. N-glycan was released from IgY using peptide-N4-(N-acetyl-beta-glucosaminyl) asparagine-amidase treatment, and the molecular weight of IgY was calculated using the difference between the molecular weight of IgY and deglycosylated IgY. Two potential N-Glycosylation sites (ASN 308 and ASN 409 ) were detected on IgY by nanoLC-ESI MS. Sugar chains were separated using normal phase liquid chromatography after fluorescence labeling, and 17 N-glycan structures were confirmed using ESI-MS. The sugar chain pattern contained high-mannose oligosaccharide, hybrid oligosaccharide and complex oligosaccharide. These results could lead to other important information regarding IgY glycosylation. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Quantification and imaging of HER2 protein using nanocrystals conjugated with single-domain antibodies

    International Nuclear Information System (INIS)

    Glukhov, S; Berestovoy, M; Nabiev, I; Sukhanova, A; Chames, P; Baty, D

    2017-01-01

    This study dealt with quantification and imaging of human epidermal growth factor receptor 2 (HER2), an important prognostic marker for cancer diagnosis and treatment, using specific quantum-dot-based conjugates. Fluorescent inorganic nanocrystals or quantum dots (QDs) are extremely highly resistant to photobleaching and have a high emission quantum yield and a continuous range of emission spectra, from the ultraviolet to the infrared regions. Ultrasmall nanoprobes consisting of highly affine anti-HER2 single-domain antibodies (sdAbs or 'nanobodies') conjugated with QDs in a strictly oriented manner have been designed. QDs with a fluorescence peak maxima at wavelengths of 562 nm, 569 nm, 570 nm or in the near-infrared region were used. Here, we present our results of ISA quantification of HER2 protein, in situ imaging of HER2 protein on the surface of HER2-positive SK-BR-3 cells in immunohistochemical experiments, and counting of stained with anti-HER2 conjugates HER2-positive SK-BR-3 cells in their mixture with unstained cells of the same culture in flow cytometry experiments. The data demonstrate that the anti-HER2 QD–sdAb conjugates obtained are highly specific and sensitive and could be used in numerous applications for advanced integrated diagnosis. (paper)

  7. Quantification and imaging of HER2 protein using nanocrystals conjugated with single-domain antibodies

    Science.gov (United States)

    Glukhov, S.; Berestovoy, M.; Chames, P.; Baty, D.; Nabiev, I.; Sukhanova, A.

    2017-01-01

    This study dealt with quantification and imaging of human epidermal growth factor receptor 2 (HER2), an important prognostic marker for cancer diagnosis and treatment, using specific quantum-dot-based conjugates. Fluorescent inorganic nanocrystals or quantum dots (QDs) are extremely highly resistant to photobleaching and have a high emission quantum yield and a continuous range of emission spectra, from the ultraviolet to the infrared regions. Ultrasmall nanoprobes consisting of highly affine anti-HER2 single-domain antibodies (sdAbs or "nanobodies") conjugated with QDs in a strictly oriented manner have been designed. QDs with a fluorescence peak maxima at wavelengths of 562 nm, 569 nm, 570 nm or in the near-infrared region were used. Here, we present our results of ISA quantification of HER2 protein, in situ imaging of HER2 protein on the surface of HER2-positive SK-BR-3 cells in immunohistochemical experiments, and counting of stained with anti-HER2 conjugates HER2-positive SK-BR-3 cells in their mixture with unstained cells of the same culture in flow cytometry experiments. The data demonstrate that the anti-HER2 QD-sdAb conjugates obtained are highly specific and sensitive and could be used in numerous applications for advanced integrated diagnosis.

  8. Glycoengineered Monoclonal Antibodies with Homogeneous Glycan (M3, G0, G2, and A2 Using a Chemoenzymatic Approach Have Different Affinities for FcγRIIIa and Variable Antibody-Dependent Cellular Cytotoxicity Activities.

    Directory of Open Access Journals (Sweden)

    Masaki Kurogochi

    Full Text Available Many therapeutic antibodies have been developed, and IgG antibodies have been extensively generated in various cell expression systems. IgG antibodies contain N-glycans at the constant region of the heavy chain (Fc domain, and their N-glycosylation patterns differ during various processes or among cell expression systems. The Fc N-glycan can modulate the effector functions of IgG antibodies, such as antibody-dependent cellular cytotoxicity (ADCC and complement dependent cytotoxicity (CDC. To control Fc N-glycans, we performed a rearrangement of Fc N-glycans from a heterogeneous N-glycosylation pattern to homogeneous N-glycans using chemoenzymatic approaches with two types of endo-β-N-acetyl glucosaminidases (ENG'ases, one that works as a hydrolase to cleave all heterogeneous N-glycans, another that is used as a glycosynthase to generate homogeneous N-glycans. As starting materials, we used an anti-Her2 antibody produced in transgenic silkworm cocoon, which consists of non-fucosylated pauci-mannose type (Man2-3GlcNAc2, high-mannose type (Man4-9GlcNAc2, and complex type (Man3GlcNAc3-4 N-glycans. As a result of the cleavage of several ENG'ases (endoS, endoM, endoD, endoH, and endoLL, the heterogeneous glycans on antibodies were fully transformed into homogeneous-GlcNAc by a combination of endoS, endoD, and endoLL. Next, the desired N-glycans (M3; Man3GlcNAc1, G0; GlcNAc2Man3GlcNAc1, G2; Gal2GlcNAc2Man3GlcNAc1, A2; NeuAc2Gal2GlcNAc2Man3GlcNAc1 were transferred from the corresponding oxazolines to the GlcNAc residue on the intact anti-Her2 antibody with an ENG'ase mutant (endoS-D233Q, and the glycoengineered anti-Her2 antibody was obtained. The binding assay of anti-Her2 antibody with homogenous N-glycans with FcγRIIIa-V158 showed that the glycoform influenced the affinity for FcγRIIIa-V158. In addition, the ADCC assay for the glycoengineered anti-Her2 antibody (mAb-M3, mAb-G0, mAb-G2, and mAb-A2 was performed using SKBR-3 and BT-474 as target

  9. Glycoengineered Monoclonal Antibodies with Homogeneous Glycan (M3, G0, G2, and A2) Using a Chemoenzymatic Approach Have Different Affinities for FcγRIIIa and Variable Antibody-Dependent Cellular Cytotoxicity Activities.

    Science.gov (United States)

    Kurogochi, Masaki; Mori, Masako; Osumi, Kenji; Tojino, Mami; Sugawara, Shu-Ichi; Takashima, Shou; Hirose, Yuriko; Tsukimura, Wataru; Mizuno, Mamoru; Amano, Junko; Matsuda, Akio; Tomita, Masahiro; Takayanagi, Atsushi; Shoda, Shin-Ichiro; Shirai, Takashi

    2015-01-01

    Many therapeutic antibodies have been developed, and IgG antibodies have been extensively generated in various cell expression systems. IgG antibodies contain N-glycans at the constant region of the heavy chain (Fc domain), and their N-glycosylation patterns differ during various processes or among cell expression systems. The Fc N-glycan can modulate the effector functions of IgG antibodies, such as antibody-dependent cellular cytotoxicity (ADCC) and complement dependent cytotoxicity (CDC). To control Fc N-glycans, we performed a rearrangement of Fc N-glycans from a heterogeneous N-glycosylation pattern to homogeneous N-glycans using chemoenzymatic approaches with two types of endo-β-N-acetyl glucosaminidases (ENG'ases), one that works as a hydrolase to cleave all heterogeneous N-glycans, another that is used as a glycosynthase to generate homogeneous N-glycans. As starting materials, we used an anti-Her2 antibody produced in transgenic silkworm cocoon, which consists of non-fucosylated pauci-mannose type (Man2-3GlcNAc2), high-mannose type (Man4-9GlcNAc2), and complex type (Man3GlcNAc3-4) N-glycans. As a result of the cleavage of several ENG'ases (endoS, endoM, endoD, endoH, and endoLL), the heterogeneous glycans on antibodies were fully transformed into homogeneous-GlcNAc by a combination of endoS, endoD, and endoLL. Next, the desired N-glycans (M3; Man3GlcNAc1, G0; GlcNAc2Man3GlcNAc1, G2; Gal2GlcNAc2Man3GlcNAc1, A2; NeuAc2Gal2GlcNAc2Man3GlcNAc1) were transferred from the corresponding oxazolines to the GlcNAc residue on the intact anti-Her2 antibody with an ENG'ase mutant (endoS-D233Q), and the glycoengineered anti-Her2 antibody was obtained. The binding assay of anti-Her2 antibody with homogenous N-glycans with FcγRIIIa-V158 showed that the glycoform influenced the affinity for FcγRIIIa-V158. In addition, the ADCC assay for the glycoengineered anti-Her2 antibody (mAb-M3, mAb-G0, mAb-G2, and mAb-A2) was performed using SKBR-3 and BT-474 as target cells, and

  10. The impact of N-glycosylation on conformation and stability of immunoglobulin Y from egg yolk.

    Science.gov (United States)

    Sheng, Long; He, Zhenjiao; Chen, Jiahui; Liu, Yaofa; Ma, Meihu; Cai, Zhaoxia

    2017-03-01

    Immunoglobulin Y (IgY) is a new therapeutic antibody, and its applications in industry are very broad. To provide insight into the effects of N-glycosylation on IgY, its conformation and stability were studied. In this research, IgY was extracted from egg yolk and then digested by peptide-N4-(N-acetyl-beta-glucosaminyl) asparagine-amidase. SDS-PAGE and infrared absorption spectrum showed that carbohydrates were distinctly reduced after enzymolysis. The circular dichroism spectrum indicated that the IgY molecule became more flexible and disordered after removal of N-glycan. The fluorescence intensity revealed that Trp residues were buried in a more hydrophobic environment after disposal of N-glycan. Storage stability decreased with the removal of oligosaccharide chains based on size-exclusion chromatography analysis. Deglycosylated IgY exhibited less resistance to guanidine hydrochloride-induced unfolding. After deglycosylation, IgY was more sensitive to pepsin. Therefore, N-glycosylation played an important role in the maintenance of the structure and stability of IgY. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Protection Efficacy of Multivalent Egg Yolk Immunoglobulin against Eimeria Tenella Infection in Chickens

    Directory of Open Access Journals (Sweden)

    JJ Xu

    2013-09-01

    Full Text Available Background: To control avian coccidiosis with drug-independent strategy effec­tively and safely, multivalent hyperimmune egg yolk immunoglobulin (IgY was prepared and its ability to protect against Eimeria tenella infection was evaluated.Methods: Hens were orally immunized with live oocysts of 5 species of Eimeria for six times, antibody titers in serum and yolk were monitored by indirect enzyme-linked immunosorbent assay. The specific IgY was isolated, purified and lyophi­lized. IgY powder was orally administrated as dietary supplement in newly hatched chicks at various dosages. Birds were orally challenged with 10000 sporulated oo­cysts of E. tenella at 10 days of age, weighed and killed at 8 days post challenge, and the protective effect was assessed.Results: The averge yeid of IgY was 9.2 mg/ml yolk, the antibody titer of IgY reached to 1:163840 per mg with the purity up to 98%. Chickens fed IgY resulted in reduced mortality, increased body weight gain (BWG, reduced oocyst shedding, reduced caecal lesion score and increased anti-coccidial index. In terms of BWG and caecal lesion, IgY significantly enhanced the resistance of bird at ≥ 0.05% of IgY in the diet when compared with the challenged control group (P0.05.Conclusion: Supplementing newly hatched chicks with Eimeria-specific IgY represents a promising strategy to prevent avian coccidiosis.

  12. Using Emotional Intelligence in Personalized Adaptation

    NARCIS (Netherlands)

    Damjanovic, Violeta; Kravcik, Milos

    2008-01-01

    Damjanovic, V. & Kravcik, M. (2007). Using Emotional Intelligence in Personalized Adaptation. In M. D. Lytras & A. Naeve (Eds.), Ubiquitous and Pervasive Knowledge and Learning Management (pp. 158-197). IGI Publishing.

  13. Using Emotional Intelligence in Personalized Adaptation

    NARCIS (Netherlands)

    Damjanovic, Violeta; Kravcik, Milos

    2007-01-01

    Damjanovic, V. & Kravcik, M. (2007). Using Emotional Intelligence in Personalized Adaptation. In V. Sugumaran (Ed.), Intelligent Information Technologies: Concepts, Methodologies, Tools, and Applications (pp. 1716-1742). IGI Publishing.

  14. POTENSI NETRALISASI IMUNOGLOBULIN Y ANTITETANUS YANG DIISOLASI DARI TELUR AYAM (THE POTENCY NETRALIZATION OF ANTI TETANUS IMMUNOGLOBULIN Y THAT WERE ISOLATED FROM CHICKEN EGGS

    Directory of Open Access Journals (Sweden)

    I Nyoman Suartha

    2007-06-01

    Full Text Available The porpuse of study was to explore the potential use of? anti tetanus IgY from eggs yolk as a substitute for anti tetanus serum raised in ?horses. The eggs were collected from chickens which have previously been immunized with tetanus toxoid. Neutralization potency test of anti tetanus IgY determined by ?Spearman-Karber method.? The highest mean titer of anti tetanus of egg yolk was 80.16 ? 33.55 IU/ml and the lowest was 1.69 ? 0.63 IU/ml. The concentration? of purified IgY was 1.644 ? 0.424 mg/ml. Spearman-Karber value of potency of anti tetanus IgY are 35 IU/ml. ?This research concluded that Chickens was capable of produced of anti tetanus in eggs yolk with value of potency are 35 IU/ml.

  15. Effect of inoculation route on the production of antibodies and histological characteristics of the spleen in laying hens

    Directory of Open Access Journals (Sweden)

    SF Eto

    2012-03-01

    Full Text Available Recent studies have reported the use of IgY antibody in the prevention or treatment of diseases in animals. IgY can be obtained in large amounts from the yolk of chicken eggs through a low-cost process. This study evaluated the effect of different routes of inoculation on antibody production and spleen morphological characteristics of laying hens (White Leghorn inoculated with sheep red blood cells. The analysis of the results showed that the intramuscular route is the most efficient for total antibody production in the primary immune response, while the intravenous route is the most efficient in producing IgY antibodies in the secondary immune response. No histological changes were observed in the spleen of laying hens. This study could be useful for developing protocols of antigen inoculation in laying hens for IgY antibody production.

  16. Using immunoglobulin Y as an alternative antibody for the detection of hepatitis A virus in frozen liver sections

    Directory of Open Access Journals (Sweden)

    Gentil Arthur Bentes

    2015-06-01

    Full Text Available An increasing amount of research has been conducted on immunoglobulin Y (IgY because the use of IgY offers several advantages with respect to diagnostic testing, including its easy accessibility, low cost and translatability to large-scale production, in addition to the fact that it can be ethically produced. In a previous work, immunoglobulin was produced and purified from egg yolks (IgY reactive to hepatitis A virus (HAV antigens. In the present work, this anti-HAV-specific IgY was used in an indirect immunofluorescence assay to detect viral antigens in liver biopsies that were obtained from experimentally infected cynomolgus monkeys. Fields that were positive for HAV antigen were detected in liver sections using confocal microscopy. In conclusion, egg yolks from immunised hens may be a reliable source for antibody production, which can be employed for immunological studies.

  17. Ten-Competence: Life-Long Competence Development and Learning

    NARCIS (Netherlands)

    Koper, Rob; Specht, Marcus

    2006-01-01

    Koper, R., & Specht, M. (2008). Ten-Competence: Life-Long Competence Development and Learning. In M-A. Cicilia (Ed.), Competencies in Organizational e-learning: concepts and tools (pp. 234-252). Hershey: IGI-Global.

  18. Recensione / Recension / Book review

    Directory of Open Access Journals (Sweden)

    Bravo Fabio

    2010-03-01

    Full Text Available Book review of:Sette R., Cases on Technologies for Teaching Criminology and Victimology: Methodologies and practices, IGI Global – Information Science Reference, Hershey PA, 2010

  19. 77 FR 5028 - Withdrawal of Approval of New Animal Drug Applications

    Science.gov (United States)

    2012-02-01

    .... DATES: Withdrawal of approval is effective February 13, 2012. FOR FURTHER INFORMATION CONTACT: John...- 3115. NADA 100-689 DIFIL Syrup (diethylcarbamazine Evsco Pharmaceuticals, an Affiliate of citrate). IGI...

  20. NASA's Origins and the Dawn of the Space Age. No. 10; Monographs in Aerospace History

    Science.gov (United States)

    Portree, David S. F.

    1998-01-01

    The twenty page narrative describes historical circumstances around Sputnik, the International Geophysical Year (IGY) and the formation of NASA from NACA in 1957-1958. Appendices include reproductions of relevant historical documents.

  1. The updated network meta-analysis of neoadjuvant therapy for HER2-positive breast cancer.

    Science.gov (United States)

    Nakashoji, Ayako; Hayashida, Tetsu; Yokoe, Takamichi; Maeda, Hinako; Toyota, Tomoka; Kikuchi, Masayuki; Watanuki, Rurina; Nagayama, Aiko; Seki, Tomoko; Takahashi, Maiko; Abe, Takayuki; Kitagawa, Yuko

    2018-01-01

    We previously described a systematic assessment of the neoadjuvant therapies for human epidermal growth factor receptor-2 (HER2) positive breast cancer, using network meta-analysis. Accumulation of new clinical data has compelled us to update the analysis. Randomized trials comparing different anti-HER2 regimens in the neoadjuvant setting were included, and odds ratio for pathologic complete response (pCR) in seven treatment arms were assessed by pooling effect sizes. Direct and indirect comparisons using a Bayesian statistical model were performed. All statistical tests were two-sided. A database search identified 993 articles with 13 studies meeting the eligibility criteria, including three new studies with lapatinib (lpnb). In an indirect comparison, dual anti-HER2 agents with CT achieved a better pCR rate than other arms. The credibility intervals of CT + tzmb + lpnb arm were largely reduced compared to our former report, which we added sufficient clinical evidence by this update. Values of surface under the cumulative ranking (SUCRA) suggested that CT + tzmb + pzmb had the highest probability of being the best treatment arm for pCR, widening the difference between the top two dual-HER2 blockade arms compared to our former report. The overall consistency with our first report enhanced the credibility of the results. Network meta-analysis using new clinical data firmly establish that combining two anti-HER2 agents with CT is most effective against HER2-positive breast cancer in the neoadjuvant setting. New pzmb related trials are required to fully determine the best neoadjuvant dual-HER2 blockade regimen. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. A virosomal formulated Her-2/neu multi-peptide vaccine induces Her-2/neu-specific immune responses in patients with metastatic breast cancer: a phase I study.

    Science.gov (United States)

    Wiedermann, Ursula; Wiltschke, C; Jasinska, J; Kundi, M; Zurbriggen, R; Garner-Spitzer, E; Bartsch, R; Steger, G; Pehamberger, H; Scheiner, O; Zielinski, C C

    2010-02-01

    We have previously shown in mice that vaccination with three Her-2-peptides representing B-cell epitopes of the extracellular domain of Her-2/neu induces Her-2/neu-specific IgG antibodies with strong anti-tumor activity in vitro and in vivo. We have now finalized a phase I clinical trial with an anti-Her-2/neu vaccine-construct of immunopotentiating reconstituted influenza virosomes with the three peptides in patients with metastatic breast cancer (MBC). Ten MBC patients with low protein overexpression of Her-2/neu of MBC (+ or ++ upon immunohistochemistry, FISH negative) and positive hormone receptor status were enrolled in a single center phase I study. The virosomal formulated vaccine, consisting of 10 microg/peptide, was intramuscularly applied three times on days 1, 28, and 56. The primary endpoint of the study, which lasted 12 weeks, was safety, the secondary endpoint immunogenicity. Local erythema at the injection site was the only vaccine-related side effect occurring in four patients. In 8 of 10 patients an increase in peptide-specific antibody titer measured by ELISA was found. Importantly, the induced antibodies were also directed against the native Her-2/neu protein. Cellular immune responses, as measured by in vitro production of IL-2, IFN-c, and TNF-a of PBMCs showed a marked increase after vaccination in the majority of vaccinees. Notably, the number of CD4+CD25+Foxp3+T regulatory cells, which were significantly increased compared to healthy controls prior to vaccination, was markedly reduced following vaccination. In all, the immunological responses after vaccination indicated that the patients in stage IV of disease were immunocompetent and susceptible to vaccination. The Her-2/neu multipeptide vaccine was safe, well tolerated and effective in overcoming immunological tolerance to Her-2/neu. The induction of anti-Her-2-specific antibodies could result in clinical benefit comparable to passive anti-Her-2 antibody therapy.

  3. Suppressors for Human Epidermal Growth Factor Receptor 2/4 (HER2/4): A New Family of Anti-Toxoplasmic Agents in ARPE-19 Cells.

    Science.gov (United States)

    Kim, Yeong Hoon; Bhatt, Lokraj; Ahn, Hye-Jin; Yang, Zhaoshou; Lee, Won-Kyu; Nam, Ho-Woo

    2017-10-01

    The effects of tyrosine kinase inhibitors (TKIs) were evaluated on growth inhibition of intracellular Toxoplasma gondii in host ARPE-19 cells. The number of tachyzoites per parasitophorous vacuolar membrane (PVM) was counted after treatment with TKIs. T. gondii protein expression was assessed by western blot. Immunofluorescence assay was performed using Programmed Cell Death 4 (PDCD4) and T. gondii GRA3 antibodies. The TKIs were divided into 3 groups; non-epidermal growth factor receptor (non-EGFR), anti-human EGFR 2 (anti-HER2), and anti-HER2/4 TKIs, respectively. Group I TKIs (nintedanib, AZD9291, and sunitinib) were unable to inhibit proliferation without destroying host cells. Group II TKIs (lapatinib, gefitinib, erlotinib, and AG1478) inhibited proliferation up to 98% equivalent to control pyrimethamine (5 μM) at 20 μM and higher, without affecting host cells. Group III TKIs (neratinib, dacomitinib, afatinib, and pelitinib) inhibited proliferation up to 98% equivalent to pyrimethamine at 1-5 μM, but host cells were destroyed at 10-20 μM. In Group I, TgHSP90 and SAG1 inhibitions were weak, and GRA3 expression was moderately inhibited. In Group II, TgHSP90 and SAG1 expressions seemed to be slightly enhanced, while GRA3 showed none to mild inhibition; however, AG1478 inhibited all proteins moderately. Protein expression was blocked in Group III, comparable to pyrimethamine. PDCD4 and GRA3 were well localized inside the nuclei in Group I, mildly disrupted in Group II, and were completely disrupted in Group III. This study suggests the possibility of a vital T. gondii TK having potential HER2/4 properties, thus anti-HER2/4 TKIs may inhibit intracellular parasite proliferation with minimal adverse effects on host cells.

  4. Neoadjuvant therapy for early-stage breast cancer: the clinical utility of pertuzumab

    International Nuclear Information System (INIS)

    Gollamudi, Jahnavi; Parvani, Jenny G; Schiemann, William P; Vinayak, Shaveta

    2016-01-01

    Approximately 20% of breast cancer patients harbor tumors that overexpress human epidermal growth factor receptor 2 (HER2; also known as ErbB2), a receptor tyrosine kinase that belongs to the epidermal growth factor receptor family of receptor tyrosine kinases. HER2 amplification and hyperactivation drive the growth and survival of breast cancers through the aberrant activation of proto-oncogenic signaling systems, particularly the Ras/MAP kinase and PI3K/AKT pathways. Although HER2-positive (HER2 + ) breast cancer was originally considered to be a highly aggressive form of the disease, the clinical landscape of HER2 + breast cancers has literally been transformed by the approval of anti-HER2 agents for adjuvant and neoadjuvant settings. Indeed, pertuzumab is a novel monoclonal antibody that functions as an anti-HER2 agent by targeting the extracellular dimerization domain of the HER2 receptor; it is also the first drug to receive an accelerated approval by the US Food and Drug Administration for use in neoadjuvant settings in early-stage HER2 + breast cancer. Here, we review the molecular and cellular factors that contribute to the pathophysiology of HER2 in breast cancer, as well as summarize the landmark preclinical and clinical findings underlying the approval and use of pertuzumab in the neoadjuvant setting. Finally, the molecular mechanisms operant in mediating resistance to anti-HER2 agents, and perhaps to pertuzumab as well, will be discussed, as will the anticipated clinical impact and future directions of pertuzumab in breast cancer patients

  5. Generation and characterization of chicken egg yolk antibodies against propionibacterium acnes for the prevention of acne vulgaris

    Directory of Open Access Journals (Sweden)

    Karthika Selvan

    2012-01-01

    Full Text Available Introduction: Antigen-specific antibody has been widely used for immunological analysis in the field of diagnosis as well as in pure scientific research, where the IgY antibodies can be raised against P acnes antigen. Material and Methods: To produce IgY against Propionibacterium acnes, laying hens were immunized with P acnes (MTCC No: 1951 and subsequent booster injections were given. The antibodies produced were purified from the egg yolk of immunized chicken using the polyethylene glycol and ammonium sulfate precipitation method and, further, by Diethylaminoethyl (DEAE cellulose ion-exchange column chromatography. The protein fraction of IgY was isolated from the egg yolk. The separation was rapid, and the success of each step was viewed on Sodium Dodecyl Sulphate-polyacrylamide gel electrophoresis (SDS-PAGE. The reactivity of anti-P acnes was evaluated by the Enzyme-linked immunosorbent assay (ELISA test and the dot-immunoassay. Results: With ELISA, the highest titter of 1:10000 was observed on the 150 th day after vaccination. The results of dot-immunoassay suggested that anti-P acnes IgY developed a brown color as positive reaction, which showed the antigen-antibody binding even after a maximum dilution of 1/500. These results suggest that anti-acne IgY was produced and had strong specific antibody reactivity. Conclusion: The findings indicate that anti-acne IgY is worth utilizing as a preventive agent for acne vulgaris.

  6. Production, characterization and applications for Toxoplasma gondii-specific polyclonal chicken egg yolk immunoglobulins.

    Directory of Open Access Journals (Sweden)

    Álvaro Ferreira Júnior

    Full Text Available Toxoplasma gondii may cause abortions, ocular and neurological disorders in warm-blood hosts. Immunized mammals are a wide source of hyperimmune sera used in different approaches, including diagnosis and the study of host-parasite interactions. Unfortunately, mammalian antibodies present limitations for its production, such as the necessity for animal bleeding, low yield, interference with rheumatoid factor, complement activation and affinity to Fc mammalian receptors. IgY antibodies avoid those limitations; therefore they could be an alternative to be applied in T. gondii model.In this study we immunized hens with soluble tachyzoite antigens of T. gondii (STAg and purified egg yolk antibodies (IgY by an inexpensive and simple method, with high yield and purity degree. IgY anti-STAg antibodies presented high avidity and were able to recognize a broad range of parasite antigens, although some marked differences were observed in reactivity profile between antibodies produced in immunized hens and mice. Interestingly, IgY antibodies against Neospora caninum and Eimeria spp. did not react to STAg. We also show that IgY antibodies were suitable to detect T. gondii forms in paraffin-embedded sections and culture cell monolayers.Due to its cost-effectiveness, high production yield and varied range of possible applications, polyclonal IgY antibodies are useful tools for studies involving T. gondii.

  7. The Ability of Immunoglobulin Yolk Recognized the Antigen in the Tissue of Ascaridia galli

    Directory of Open Access Journals (Sweden)

    Darmawi

    2012-12-01

    Full Text Available Antigen-antibody reaction is an important tool for the analysis of localization of target molecules, including antigenic protein within worm tissues. The purpose of the present research was to demonstrate the ability of immunoglobulin yolk (IgY anti-excretory/secretory recognized the antigen in the tissue of Ascaridia galli by mean of immunohistochemistry method. The excretory/secretory protein was procured from A. galli and concentrated by mean of vivaspin 30,000 MWCO. IgY was produced by egg yolks of immunized chickens with excretory/secretory, and purified using fast protein liquid chromatography (FPLC method. A. galli adult worms were cut in transversal and longitudinal section of the center and anterior region. Slides were incubated with both primary IgY for overnight at 4 oC and secondary antibody rabbit anti-chicken IgY HRP-conjugate for one hour at room temperature. The slides were stained with 3-amino, 9-ethylcarbazole (AEC chromogen, counterstained with Lillie Mayer Haematoxylin, and mounted in glyserin aqueous mount. Antigen-antibody reaction was investigated under a microscope. The result showed that antigen was appeared in the tissues such as cuticle, epicuticle, buccal cavity, and eggs inside the uterine of A. galli. This research concluded that IgY stimulated by the excretory/secretory was able to recognized the antigen scattered in the tissues of A. galli so the IgY could be applied for immunodiagnostic.

  8. Green turtles (Chelonia mydas) have novel asymmetrical antibodies

    Science.gov (United States)

    Work, Thierry M.; Dagenais, Julie; Breeden, Renee; Schneemann, Anette; Sung, Joyce; Hew, Brian; Balazs, George H.; Berestecky, John M.

    2015-01-01

    Igs in vertebrates comprise equally sized H and L chains, with exceptions such as H chain–only Abs in camels or natural Ag receptors in sharks. In Reptilia, Igs are known as IgYs. Using immunoassays with isotype-specific mAbs, in this study we show that green turtles (Chelonia mydas) have a 5.7S 120-kDa IgY comprising two equally sized H/L chains with truncated Fc and a 7S 200-kDa IgY comprised of two differently sized H chains bound to L chains and apparently often noncovalently associated with an antigenically related 90-kDa moiety. Both the 200- and 90-kDa 7S molecules are made in response to specific Ag, although the 90-kDa molecule appears more prominent after chronic Ag stimulation. Despite no molecular evidence of a hinge, electron microscopy reveals marked flexibility of Fab arms of 7S and 5.7S IgY. Both IgY can be captured with protein G or melon gel, but less so with protein A. Thus, turtle IgY share some characteristics with mammalian IgG. However, the asymmetrical structure of some turtle Ig and the discovery of an Ig class indicative of chronic antigenic stimulation represent striking advances in our understanding of immunology.

  9. Terapia combinada con Trastuzumab en el tratamiento del Cáncer de mama: eficacia y efectos adversos

    OpenAIRE

    Hernández Martín, María Sol

    2015-01-01

    Introducción: El cáncer de mama es el más frecuente entre la población femenina. Entre un 25-30% de los cánceres de mama son HER2-positivo. Este tipo de cáncer, se ha relacionado con una mayor agresividad clínica e histológica, mayor riesgo de recurrencia y muerte asociada al cáncer de mama. Trastuzumab, es un anticuerpo monoclonal humanizado contra este receptor. Actualmente se dispone de cuatro agentes anti-HER2 autorizados: Trastuzumab, Pertuzumab, Lapatinib y Trastuzumab...

  10. Energetics and Structure Prediction of the Network of Homo- and Hetero-Oligomers Formed by the Transmembrane Domains of the ErbReceptor Family of Proteins

    Science.gov (United States)

    2006-06-01

    amino acid residue motif, Small-x-x-Large-G/A, consist- ing of a small residue (Gly, Ala , Ser, Thr, or Pro) in the zero position, a large aliphatic...residue ( Ala , Val, Leu, or Ile) in position 3, followed by Gly or Ala in position four.15 This motif was identified in a large number of receptor tyrosine...M. A., Codony-Servat, J., Albanell, J., Rojo, F., Arribas , J. & Baselga, J. (2001). Trastuzumab (her- ceptin), a humanized anti-Her2 receptor

  11. Evaluation of Human Epidermal Growth Factor Receptor 2 (HER2) Gene Status in Human Breast Cancer Formalin-Fixed Paraffin-Embedded (FFPE) Tissue Specimens by Fluorescence In Situ Hybridization (FISH).

    Science.gov (United States)

    Hwang, Harry C; Gown, Allen M

    2016-01-01

    Current standard of care requires that HER2 gene testing be performed on all newly diagnosed invasive breast cancers in order to determine eligibility for anti-HER2 antibody therapy and should be performed in accordance with current ASCO-CAP guidelines (Hammond et al., J Clin Oncol 29(15):e458, 2011; Wolff et al., J Clin Oncol 31(31):3997-4013, 2013). Here we describe a HER2 FISH methodology to evaluate HER2 gene status in FFPE breast tumor specimens.

  12. Protective Effect of Sucralfate on Activity of IgY against Recombinant VacA of Helicobacter pylori%硫糖铝对抗幽门螺杆菌重组VacA卵黄抗体的保护作用

    Institute of Scientific and Technical Information of China (English)

    郭丽媛; 杨致邦; 田一玲; 黄伟; 韩飞; 范贵荣; 向瑜

    2009-01-01

    目的 观察硫糖铝对抗幽门螺杆菌重组细胞空泡毒素抗原(VacA)卵黄抗体(IgY)的保护作用,为制备能耐受胃酸和蛋白酶的IgY制剂奠定基础.方法 诱导重组菌DH5α-vacA-pQE30,大量表达并纯化VacA重组蛋白,免疫洛曼母鸡,制备VacA IgY,并进行纯化.在不同pH值的IgY液中加入不同浓度的硫糖铝和不同浓度的胃蛋白酶,将含不同浓度硫糖铝的IgY反复冻融7次,将含30%糖铝的IgY室温放置1 d~4周,ELISA法测定各实验组IgY的相对抗体活性(AT/Ac).结果 在pH 1.5、2.0和3.0的条件下,含30%~60%硫糖铝的IgY的相对抗体活性高于不加硫糖铝的IgY;在pH 1.5条件下,60%硫糖铝可使IgY的相对抗体活性保持68.7%;在pH 2.0和3.0条件下,50%和40%以上的硫糖铝几乎可完全保持IgY的相对抗体活性.在pH值为2.0和3.0时,正常胃蛋白酶浓度(0.02 mg/ml)下,30%以上的硫糖铝均可有效保护IgY的相对抗体活性.30%以上的硫糖铝可增强IgY的抗冻融能力.室温放置4周后,含30%硫糖铝的IgY仍能保持80%以上的相对抗体活性.结论 30%以上的硫糖铝可增强VacA IgY对低pH和胃蛋白酶的耐受能力及抗冻融能力,是较理想的IgY保护剂.

  13. 硫糖铝在体外对抗幽门螺杆菌HpaA IgY的保护作用评价%Evaluation on Protective Effects of Sucralfate on Anti-H.pylori HpaA IgY in Vitro

    Institute of Scientific and Technical Information of China (English)

    邓西川; 黄伟; 戴维; 胡晓莉; 沈旭东; 杨致邦

    2011-01-01

    目的 评价硫糖铝对HpaAIgY的保护作用,为制备IgY口服制剂奠定基础.方法 大量诱导工程菌DH5α-HpaA-pQE30,制备纯化的重组蛋白HpaA.以HpaA为抗原接种洛曼母鸡,制备纯化的IgY.在不同pH值,舍不同浓度胃蛋白酶的HpaAIgY液中加入不同浓度的硫糖铝;将IgY液反复冻融;以ELISA法测定HpaAIgY的剩余抗体活性(AR/AC).结果 在pH1.5条件下,60%硫糖铝可使IgY抗体剩余活性维持65.8%;在pH 2.0条件下,舍30%硫糖铝的IgY可使抗体活性维持85.4%,含50%硫糖铝的IgY可完全维持抗体活性.在pH 3.0条件下,含30%硫糖铝的IgY可使抗体活性维持87.3%,而含40%以上硫糖铝的IgY几乎可完全维持抗体活性.在胃蛋白酶浓度0.02mg/mL条件下,当含10%、30%、50%硫糖铝IgY的剩余活性,在pH值2.0时,分别为62.8%、71.6%、82.5%;在pH值3.0时,分别为75.6%、87.4%、94.5%;反复冻融后,含10%硫糖铝的IgY可使抗体活性提高到70.6%,含30%硫糖铝可使IgY的抗体活性提高到82.7%,50%硫糖铝可提高到89.3%.结论 30%以上的硫糖铝可增强VacAIgY对低pH和胃蛋白酶的耐受能力,抗冻融能力,是较理想的IgY保护剂.

  14. Development and Structure/Property Relationship of New Electron Accepting Polymers Based on Thieno[2',3':4,5]pyrido[2,3- <i>g> ]thieno[3,2- c ]quinoline-4,10-dione for All-Polymer Solar Cells

    Energy Technology Data Exchange (ETDEWEB)

    Jung, In Hwan; Zhao, Donglin; Jang, Jaeyoung; Chen, Wei; Landry, Erik S.; Lu, Luyao; Talapin, Dmitri V.; Yu, Luping

    2015-09-08

    Several electron accepting polymers having weak accepting–strong accepting (WA-SA) and strong accepting–strong accepting (SA-SA) monomer alternation were synthesized for studies of structure/property relationship in all-polymer solar cells. Two kinds of cyclic amide monomers, 4,10-bis(2-butyloctyl)-thieno[2',3':5,6]pyrido[3,4-g]thieno-[3,2-c]isoquinoline-5,11-dione (TPTI) and 5,11-bis(2-butyloctyl)-thieno[2',3':4,5]pyrido[2,3-g]thieno[3,2-c]quinoline-4,10-dione (TPTQ), were synthesized as weak accepting monomers (WA). Difluorinated TPTQ (FTPTQ) and well-known perylene diimide (PDI) monomers were synthesized as strong electron accepting monomers (SA). By using 1-chloronaphthalene (CN) as a cosolvent, the morphology of the polymer blended films can be finely tuned to achieve better ordering toward face-on mode and favorable phase separation between electron donor and acceptor, resulting in significant enhancement of short circuit current (Jsc) and fill factor (FF). The fluorination in the TPTQ unit reduced the dipole moment of the D–A complex and gave a negative effect on a polymer system. PFP showed worse electron accepting property with lower electron mobility than PQP. It is reasoned that the internal polarization plays an important role in the design of electron accepting polymers. As a result, PQP having TPTQ monomer exhibited the best photovoltaic performance with power conversion efficiency (PCE) of 3.52% (Voc = 0.71 V, Jsc = 8.57 mA/cm2, FF = 0.58) at a weight ratio of PTB7-Th:PQP = 1:1, under AM 1.5G.

  15. Development and Structure/Property Relationship of New Electron Accepting Polymers Based on Thieno[2',3':4,5]pyrido[2,3- <i>g> ]thieno[3,2- c ]quinoline-4,10-dione for All-Polymer Solar Cells

    Energy Technology Data Exchange (ETDEWEB)

    Jung, In Hwan; Zhao, Donglin; Jang, Jaeyoung; Chen, Wei; Landry, Erik S.; Lu, Luyao; Talapin, Dmitri V.; Yu, Luping

    2015-09-08

    Several electron accepting polymers having weak accepting–strong accepting (WA-SA) and strong accepting–strong accepting (SA-SA) monomer alternation were synthesized for studies of structure/property relationship in all-polymer solar cells. Two kinds of cyclic amide monomers, 4,10-bis(2-butyloctyl)-thieno[2',3':5,6]pyrido[3,4-g]thieno-[3,2-c]isoquinoline-5,11-dione (TPTI) and 5,11-bis(2-butyloctyl)-thieno[2',3':4,5]pyrido[2,3-g]thieno[3,2-c]quinoline-4,10-dione (TPTQ), were synthesized as weak accepting monomers (WA). Difluorinated TPTQ (FTPTQ) and well-known perylene diimide (PDI) monomers were synthesized as strong electron accepting monomers (SA). By using 1-chloronaphthalene (CN) as a cosolvent, the morphology of the polymer blended films can be finely tuned to achieve better ordering toward face-on mode and favorable phase separation between electron donor and acceptor, resulting in significant enhancement of short circuit current (Jsc) and fill factor (FF). The fluorination in the TPTQ unit reduced the dipole moment of the D–A complex and gave a negative effect on a polymer system. PFP showed worse electron accepting property with lower electron mobility than PQP. It is reasoned that the internal polarization plays an important role in the design of electron accepting polymers. As a result, PQP having TPTQ monomer exhibited the best photovoltaic performance with power conversion efficiency (PCE) of 3.52% (Voc = 0.71 V, Jsc = 8.57 mA/cm2, FF = 0.58) at a weight ratio of PTB7-Th:PQP = 1:1, under AM 1.5G.

  16. Production and Purification Immunoglobulin against E. coli in Egg Yolk

    Directory of Open Access Journals (Sweden)

    Mohammadreza Nassiri

    2016-08-01

    Full Text Available Introduction Chicken is the only avian species in which polyclonal antibodies, like IgG is transported from the hen to the egg yolk in a similar manner as the transport of mammalian IgG from the mother to the fetus. Immunoglobulin Y in the chicken is transported to the egg and accumulates in the egg yolk in large quantities. IgY is an egg yolk antibody that has been used widely for treatment and prevention of infections in humans and animal. IgY is used for passive protection of the pathogen infections such as Escherichia coli, bovine and human rotavirus, bovine coronavirus, salmonella, staphylococcus and Pseudomonas. IgY is a promising candidate as an alternative to antibiotics. Eschericha coli strains of serotype O157: H7 belongs to a family of pathogenic E. coli called enterohemorrhagic E. coli (EHEC strains responsible for hemorrhagic colitis, bloody or non-bloody diarrhea, and hemolytic uremic syndrome in humans. This strain of E. coli pathogenises by adhering to host intestinal epithelium and forming bacterial colonies. The purpose of this study was to produce and purify immunoglobulin Y against E. coli O157:H7 and develop specific polyclonal anti E. coli antibody in the egg yolk. Materials and Methods Sixteen-week-old laying hens (Mashhad, Iran were kept in individual cages with food and water ad libitum. Immunization of hens was performed by intramuscularly injecting killed E. coli O157: H7 with an equal volume of Freund’s complete adjuvant into two sides of chest area (Sigma, USA for the first immunization. Two booster immunizations followed up using complete and incomplete Freund’s adjuvants in two weeks interval. Freund’s adjuvant without antigen was injected to the control group. Two weeks after the last injection, the eggs were collected daily for eight weeks, marked and stored at 4 ºC. In order to IgY purification, eggs were collected. Purification of IgY from egg yolk was based on Polson and using PEG6000. Finally, the

  17. Thermoinactivation Mechanism of Glucose Isomerase

    Science.gov (United States)

    Lim, Leng Hong; Saville, Bradley A.

    In this article, the mechanisms of thermoinactivation of glucose isomerase (GI) from Streptomyces rubiginosus (in soluble and immobilized forms) were investigated, particularly the contributions of thiol oxidation of the enzyme's cysteine residue and a "Maillard-like" reaction between the enzyme and sugars in high fructose corn syrup (HFCS). Soluble GI (SGI) was successfully immobilized on silica gel (13.5 μm particle size), with an activity yield between 20 and 40%. The immobilized GI (IGI) has high enzyme retention on the support during the glucose isomerization process. In batch reactors, SGI (half-life =145 h) was more stable than IGI (half-life=27 h) at 60°C in HFCS, whereas at 80°C, IGI (half-life=12 h) was more stable than SGI (half-life=5.2 h). IGI was subject to thiol oxidation at 60°C, which contributed to the enzyme's deactivation. IGI was subject to thiol oxidation at 80°C, but this did not contribute to the deactivation of the enzyme. SGI did not undergo thiol oxidation at 60°C, but at 80°C SGI underwent severe precipitation and thiol oxidation, which caused the enzyme to deactivate. Experimental results show that immobilization suppresses the destablizing effect of thiol oxidation on GI. A "Maillard-like" reaction between SGI and the sugars also caused SGI thermoinactivation at 60, 70, and 80°C, but had minimal effect on IGI. At 60 and 80°C, IGI had higher thermostability in continuous reactors than in batch reactors, possibily because of reduced contact with deleterious compounds in HFCS.

  18. Clearance and Biodistribution of Liposomally Encapsulated Nitroxides: A Model for Targeted Delivery of Electron Paramagnetic Resonance Imaging Probes to Tumors

    Science.gov (United States)

    Burks, Scott R.; Legenzov, Eric A.; Rosen, Gerald M.

    2011-01-01

    Electron paramagnetic resonance (EPR) imaging using nitroxides as molecular probes is potentially a powerful tool for the detection and physiological characterization of micrometastatic lesions. Encapsulating nitroxides in anti-HER2 immunoliposomes at high concentrations to take advantage of the “self-quenching” phenomenon of nitroxides allows generation of robust EPR signals in HER2-overexpressing breast tumor cells with minimal background from indifferent tissues or circulating liposomes. We investigated the in vivo pharmacological properties of nitroxides encapsulated in sterically stabilized liposomes designed for long circulation times. We show that circulation times of nitroxides can be extended from hours to days; this increases the proportion of liposomes in circulation to enhance tumor targeting. Furthermore, nitroxides encapsulated in sterically stabilized anti-HER2 immunoliposomes can be delivered to HER2-overexpressing tumors at micromolar concentrations, which should be imageable by EPR. Lastly, after in vivo administration, liposomally encapsulated nitroxide signal also appears in the liver, spleen, and kidneys. Although these organs are spatially distinct and would not hinder tumor imaging in our model, understanding nitroxide signal retention in these organs is essential for further improvements in EPR imaging contrast between tumors and other tissues. These results lay the foundation to use liposomally delivered nitroxides and EPR imaging to visualize tumor cells in vivo. PMID:21737567

  19. Targeted imaging of ovarian cancer cells using viral nanoparticles doped with indocyanine green

    Science.gov (United States)

    Guerrero, Yadir; Bahmani, Baharak; Jung, Bonsu; Vullev, Valentine; Kundra, Vikas; Anvari, Bahman

    2013-03-01

    Our group has constructed a new type of viral nanoparticles (VNPs) from genome-depleted plant infecting brome mosaic virus (BMV) that encapsulates the FDA-approved near infrared (NIR) indocyanine green (ICG)[1]. We refer to these VNPs as optical viral ghosts (OVGs) since the constructs lack the genomic content of wild-type BMV. One of our areas of interest is the application of OVGs for real-time intraoperative NIR fluorescence imaging of small peritoneal ovarian tumor nodules. We target human epidermal growth factor receptor-2 (HER-2) expression in ovarian cancer as a biomarker associated with ovarian cancer, since its over-expression is linked to the disease's progression to death. We functionalize the OVGs with anti-HER-2 monoclonal antibodies using reductive amination methods. We used fluorescence imaging to visualize the SKOV-3 cells (high HER-2 expression) after incubation with free ICG, OVGs, and functionalized OVGs. Our results suggest the possibility of using anti-HER2 conjugated OVGs in conjunction with cytoreductive surgery to detect small tumor nodules (<5cm) which currently are not excised during surgery.

  20. Establishment of H2Mab-119, an Anti-Human Epidermal Growth Factor Receptor 2 Monoclonal Antibody, Against Pancreatic Cancer.

    Science.gov (United States)

    Yamada, Shinji; Itai, Shunsuke; Nakamura, Takuro; Chang, Yao-Wen; Harada, Hiroyuki; Suzuki, Hiroyoshi; Kaneko, Mika K; Kato, Yukinari

    2017-12-01

    Human epidermal growth factor receptor 2 (HER2) is overexpressed in breast cancer and is associated with poor clinical outcomes. In addition, HER2 expression has been reported in other cancers, such as gastric, colorectal, lung, and pancreatic cancers. An anti-HER2 humanized antibody, trastuzumab, leads to significant survival benefits in patients with HER2-overexpressing breast cancers and gastric cancers. Herein, we established a novel anti-HER2 monoclonal antibody (mAb), H 2 Mab-119 (IgG 1 , kappa), and characterized its efficacy against pancreatic cancers using flow cytometry, Western blot, and immunohistochemical analyses. H 2 Mab-119 reacted with pancreatic cancer cell lines, such as KLM-1, Capan-2, and MIA PaCa-2, but did not react with PANC-1 in flow cytometry analysis. Western blot analysis also revealed a moderate signal for KLM-1 and a weak signal for MIA PaCa-2, although H 2 Mab-119 reacted strongly with LN229/HER2 cells. Finally, immunohistochemical analyses with H 2 Mab-119 revealed sensitive and specific reactions against breast and colon cancers but did not react with pancreatic cancers, indicating that H 2 Mab-119 is useful for detecting HER2 overexpression in pancreatic cancers using flow cytometry and Western blot analyses.

  1. Advantages and disadvantages of technologies for HER2 testing in breast cancer specimens.

    Science.gov (United States)

    Furrer, Daniela; Sanschagrin, François; Jacob, Simon; Diorio, Caroline

    2015-11-01

    Human epidermal growth factor receptor 2 (HER2) plays a central role as a prognostic and predictive marker in breast cancer specimens. Reliable HER2 evaluation is central to determine the eligibility of patients with breast cancer to targeted anti-HER2 therapies such as trastuzumab and lapatinib. Presently, several methods exist for the determination of HER2 status at different levels (protein, RNA, and DNA level). In this review, we discuss the main advantages and disadvantages of the techniques developed so far for the evaluation of HER2 status in breast cancer specimens. Each technique has its own advantages and disadvantages. It is therefore not surprising that no consensus has been reached so far on which technique is the best for the determination of HER2 status. Currently, emphasis must be put on standardization of procedures, internal and external quality control assessment, and competency evaluation of already existing methods to ensure accurate, reliable, and clinically meaningful test results. Development of new robust and accurate diagnostic assays should also be encouraged. In addition, large clinical trials are warranted to identify the technique that most reliably predicts a positive response to anti-HER2 drugs. Copyright© by the American Society for Clinical Pathology.

  2. NIR-to-visible upconversion nanoparticles for fluorescent labeling and targeted delivery of siRNA

    International Nuclear Information System (INIS)

    Jiang Shan; Zhang Yong; Lim, Kian Meng; Sim, Eugene K W; Ye Lei

    2009-01-01

    Near-infrared (NIR)-to-visible upconversion fluorescent nanoparticles were synthesized and used for imaging and targeted delivery of small interfering RNA (siRNA) to cancer cells. Silica-coated NaYF 4 upconversion nanoparticles (UCNs) co-doped with lanthanide ions (Yb/Er) were synthesized. Folic acid and anti-Her2 antibody conjugated UCNs were used to fluorescently label the folate receptors of HT-29 cells and Her2 receptors of SK-BR-3 cells, respectively. The intracellular uptake of the folic acid and antibody conjugated UCNs was visualized using a confocal fluorescence microscope equipped with an NIR laser. siRNA was attached to anti-Her2 antibody conjugated UCNs and the delivery of these nanoparticles to SK-BR-3 cells was studied. Meanwhile, a luciferase assay was established to confirm the gene silencing effect of siRNA. Upconversion nanoparticles can serve as a fluorescent probe and delivery system for simultaneous imaging and delivery of biological molecules.

  3. NIR-to-visible upconversion nanoparticles for fluorescent labeling and targeted delivery of siRNA

    Science.gov (United States)

    Jiang, Shan; Zhang, Yong; Lim, Kian Meng; Sim, Eugene K. W.; Ye, Lei

    2009-04-01

    Near-infrared (NIR)-to-visible upconversion fluorescent nanoparticles were synthesized and used for imaging and targeted delivery of small interfering RNA (siRNA) to cancer cells. Silica-coated NaYF4 upconversion nanoparticles (UCNs) co-doped with lanthanide ions (Yb/Er) were synthesized. Folic acid and anti-Her2 antibody conjugated UCNs were used to fluorescently label the folate receptors of HT-29 cells and Her2 receptors of SK-BR-3 cells, respectively. The intracellular uptake of the folic acid and antibody conjugated UCNs was visualized using a confocal fluorescence microscope equipped with an NIR laser. siRNA was attached to anti-Her2 antibody conjugated UCNs and the delivery of these nanoparticles to SK-BR-3 cells was studied. Meanwhile, a luciferase assay was established to confirm the gene silencing effect of siRNA. Upconversion nanoparticles can serve as a fluorescent probe and delivery system for simultaneous imaging and delivery of biological molecules.

  4. Evolving landscape of human epidermal growth factor receptor 2-positive breast cancer treatment and the future of biosimilars.

    Science.gov (United States)

    Jackisch, Christian; Lammers, Philip; Jacobs, Ira

    2017-04-01

    Human epidermal growth factor receptor 2-positive (HER2+) breast cancer comprises approximately 15%-20% of all breast cancers and is associated with a poor prognosis. The introduction of anti-HER2 therapy has significantly improved clinical outcomes for patients with HER2+ breast cancer, and multiple HER2-directed agents (ie, trastuzumab, pertuzumab, lapatinib, and ado-trastuzumab emtansine [T-DM1]) are approved for clinical use in various settings. The treatment landscape for patients with HER2+ breast cancer is continuing to evolve. While novel agents and therapeutic strategies are emerging, biologic therapies, particularly trastuzumab, are likely to remain a mainstay of treatment. However, access issues create barriers to the use of biologics, and there is evidence for underuse of trastuzumab worldwide. A biosimilar is a biologic product that is highly similar to a licensed biologic in terms of product safety and effectiveness. Biosimilars of trastuzumab are in development and may soon become available. The introduction of biosimilars may improve access to anti-HER2 therapies by providing additional treatment options and lower-cost alternatives. Because HER2-targeted drugs may be administered for extended periods of time and in combination with other systemic therapies, biosimilars have the potential to result in significant savings for healthcare systems. Herein we review current and emerging treatment options for, and discuss the possible role of biosimilars in, treating patients with HER2+ breast cancer. Copyright © 2017 Authors, Pfizer Inc. Published by Elsevier Ltd.. All rights reserved.

  5. Emerging treatments for HER2-positive early-stage breast cancer: focus on neratinib.

    Science.gov (United States)

    Kourie, Hampig Raphael; El Rassy, Elie; Clatot, Florian; de Azambuja, Evandro; Lambertini, Matteo

    2017-01-01

    Over the last decades, a better understanding of breast cancer heterogeneity provided tools for a biologically based personalization of anticancer treatments. In particular, the overexpression of the human epidermal growth factor receptor 2 (HER2) by tumor cells provided a specific target in these HER2-positive tumors. The development of the monoclonal antibody trastuzumab, and its approval in 1998 for the treatment of patients with metastatic disease, radically changed the natural history of this aggressive subtype of breast cancer. These findings provided strong support for the continuous research in targeting the HER2 pathway and implementing the development of new anti-HER2 targeted agents. Besides trastuzumab, a series of other anti-HER2 agents have been developed and are currently being explored for the treatment of breast cancer patients, including those diagnosed with early-stage disease. Among these agents, neratinib, an oral tyrosine kinase inhibitor that irreversibly inhibits HER1, HER2, and HER4 at the intracellular level, has shown promising results, including when administered to patients previously exposed to trastuzumab-based treatment. This article aims to review the available data on the role of the HER2 pathway in breast cancer and on the different targeted agents that have been studied or are currently under development for the treatment of patients with early-stage HER2-positive disease with a particular focus on neratinib.

  6. Effects of Fab' fragments of specific egg yolk antibody (IgY-Fab') against Shewanella putrefaciens on the preservation of refrigerated turbot.

    Science.gov (United States)

    Zhang, Qian; Lin, Hong; Sui, Jianxin; Wang, Jingxue; Cao, Limin

    2015-01-01

    In our previous studies the specific egg yolk antibody (IgY) against Shewanella putrefaciens (one of the specific spoilage organisms for marine products during aerobic chilling storage) demonstrated significant activity to prolong the shelf life of refrigerated fish. The exploitation of the antigen-binding fragment plus the hinge region (IgY-Fab') is now considered a promising method for improving the efficiency of such natural antimicrobial agents. The antimicrobial activity of IgY-Fab' against S. putrefaciens was investigated using refrigerated turbot as samples. By microbial, chemical and sensory tests, it was shown to be able to effectively inhibit bacterial growth and prolong the shelf life of samples, with an efficiency evaluated significantly higher than that of whole IgY with the same molarity. The interaction between IgY agents and S. putrefaciens cells was also investigated, and the IgY-Fab' showed a much greater ability to damage cell membranes than the whole IgY. Compared to whole IgY with the same molarity, IgY-Fab' demonstrated higher and more durable antimicrobial efficiency. Such a result was assumed to be closely related to its structural properties (such as the much lower molecular weight), which may enhance its ability to influence physiological activities of antigen bacteria, especially the property or/and structure of cell membranes. © 2014 Society of Chemical Industry.

  7. Invitro Assessment of Bacteriostatic Potency of Egg Yolk Immunoglobulin against Escherichia coli

    Directory of Open Access Journals (Sweden)

    Vikrama Chakravarthi. P1

    Full Text Available The present study was carried out in commercial layer chickens to assess the bacteriostatic potency of egg yolk immunoglobulin IgY against food poisoning pathogen. The O antigen of food poisoning pathogen Escherichia coli was prepared and used to immunize commercial layer chickens. The eggs which contain anti-E.Coli IgY was collected on 30 th day of first injection and stored at 4 0 C. The antibacterial IgY was separated by water dilution method (10 times diluted with distilled water, pH 5.0 - 5.5, incubated at 4 0 C for 6 hrs and purified by 60 % ammonium sulphate. The recovery of IgY was in range of 57-62 %. The pathogens in Tryptic soya broth (approx. 6X108/ ml were cultured with anti-E.coli IgY @ 20 mg /ml and inhibitory effect was measured in UV spectrophotometer at 550 nm. The resultant growth curve indicated that the application of polyclonal antibodies (Ig Y on meat could be used to prevent the E.coli food poisoning. [Veterinary World 2010; 3(10.000: 460-462

  8. Egg Yolk Antibodies, State of the Art and Future Prospects.

    Science.gov (United States)

    Schade, Rüdiger; Hlinak, Andreas

    1996-01-01

    Immunization of chickens and extraction of antibodies from egg yolk belongs to the alternative methods since the animals suffering is reduced by non-invasive antibody-sampling. Also, the number of animals needed to produce a certain amount of antibody can be reduced since chickens produce a significant higher antibody quantity than rabbits. Despite its several advantages this technology (IgY-technology) is rather scarcely used. Traditional behavior as well as limited or no information at all may hamper a broader acceptance at present. However, significant arguments exist in chicken housing, the choice of appropriate IgY-extraction methods and a lack of information regarding the use of IgY-antibodies. This paper intends to give a short introduction in the IgY-technology, to briefly discuss the state of the art and to inform on recent developments and discussions in this field. The suitability of IgY for special fields of application (as a result of the structural differences between IgY and IgG) is emphasized (e.g. assays combining IgG and IgY, immunization of chickens against highly conserved anti-genes). In addition, it is stressed that the IgY-technology as an alternative method can particularly integrate requirements of animal protection (reduce, replace, refine), science (characteristics of avian immune system and resulting properties of IgY) and economy (amount of IgY produced from one chicken).

  9. The interaction between calreticulin and immunoglobulin G and immunoglobulin Y

    DEFF Research Database (Denmark)

    Møllegaard, Karen Mai; Duus, Karen; Træholt, Sofie Dietz

    2011-01-01

    accumulating in support of calreticulin as a polypeptide binding chaperone. In contrast to mammalian immunoglobulin G (IgG), which has complex type N-glycans, chicken immunoglobulin Y (IgY) possesses a monoglucosylated high mannose N-linked glycan, which is a ligand for calreticulin. Here, we have used solid...... and solution-phase assays to analyze the in vitro binding of calreticulin, purified from human placenta, to human IgG and chicken IgY in order to compare the interactions. In addition, peptides from the respective immunoglobulins were included to further probe the binding specificity of calreticulin....... The experiments demonstrate the ability of calreticulin to bind to denatured forms of both IgG and IgY regardless of the glycosylation state of the proteins. Furthermore, calreticulin exhibits binding to peptides (glycosylated and non-glycosylated) derived from trypsin digestion of both immunoglobulins...

  10. Effect of processing on the detectability of peanut protein by ELISA.

    Science.gov (United States)

    Iqbal, Amjad; Ateeq, Nadia

    2013-12-01

    Chicken IgY was used for the detection and quantification of peanut proteins by indirect competitive ELISA. The method was optimized by using a checker board approach to determine the optimal concentration of coating antigen, primary antibody and secondary antibody. Peanut protein could be detected in foods down to levels of 10 ppm. The effect of physical (heat treatment at 80 °C and 100 °C) and chemical (acid, alkali and reducing sugar) treatments on the IgY binding of peanut proteins was investigated. The optimized assay was relatively sensitive for the roasted peanut proteins. However, the binding ability of chicken IgYs to peanut proteins was found to be significantly altered by denaturation and hydrolysis of proteins. It was also observed that the effect of Millard chemistry on the detectability of peanut protein was less pronounced at high temperatures than at low temperatures. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. Analytical evidences of the use of iron-gall ink as a pigment on miniature paintings

    Science.gov (United States)

    Aceto, Maurizio; Calà, Elisa

    2017-12-01

    Iron-gall ink (IGI) has been used by scribes for writing since at least the 4th century CE. Another typical use of this ink was for drawing: many Old Masters created beautiful sketches in brown-black hues. Despite its widespread use to draw lines, it seems like IGI was hardly used for painting as well. In fact, the number of identification on manuscripts is very low at present. This could be partially due to a lack of reliable diagnostic information. In this work we tried to better define the possibility of identifying IGI as a pigment on illuminate manuscripts, evaluating the pros and cons of three different techniques: UV-visible diffuse reflectance spectrophotometry with optic fibres (FORS), Raman spectroscopy and XRF spectrometry. With concern to in situ non-invasive analysis, Raman spectroscopy has the best diagnostic power but FORS seems to provide the better compromise between selectivity and ease of application. Moreover, new analytical evidences was given on the particular use of IGI by ancient illuminators: a non-invasive and micro-invasive diagnostic survey on Western manuscripts datable in the range 6-16th centuries was carried out showing that, apart from its widespread use as an ink for writing and drawing, IGI was largely used as a pigment too. The large number of identification obtained allows us to hypothesise that this pigment was used all through medieval Europe up to at least the Renaissance, where its use is already documented in drawing. The occurrence of IGI in miniature paintings older than 6th century or more recent than 16th century cannot be excluded, as is its use beyond Europe; further measurements could instead widen the time range and the geographic area. Nevertheless, the present study allows shedding a new light on the use of this colourant all along the period of medieval and Renaissance miniature painting art.

  12. Comparative study on immunoglobulin Y transfer from breeding hens to egg yolk and progeny chicks in different breeds of poultry

    Directory of Open Access Journals (Sweden)

    Ritu Agrawal

    2016-04-01

    Full Text Available Aim: This study was undertaken to compare the immunoglobulin Y (IgY level and its efficacy in laying hens of four different breeds of poultry (viz.,Vanraja, Gramapriya, BlackRock, and KalingaBrown and its relative transfer in egg yolk and chick. Materials and Methods: This study was conducted in 48 apparently healthy laying hens vaccinated with Salmonella inactivated polyvalent vaccine, eggs and progeny chicks; 12 each from four different breeds of poultry,viz.,Vanraja, Gramapriya, BlackRock, and KalingaBrown. The methodology included measurement of egg and yolk weight, total protein and IgY in egg yolk, total serum protein and IgY in breeding hens, and progeny chicks and extent of IgY transfer from hens to yolk then to chicks. Further, Salmonella-specific antibodies in breeding hens, egg yolk and progeny chicks were assessed using O and H antigen by tube agglutination test. Results: The egg weight differed nonsignificantly (p>0.05 among breeds, however, breed wise significant variation (p0.05 difference among breed was found in total protein of egg yolk and chick. The IgY concentration in hens, egg yolk and chick was found to be in the range of 5.35±0.63- 5.83±0.65, 2.3±0.1-2.6±0.2, and 1.3±0.11-1.7±0.16 mg/ml, respectively which is uniform and independent of total protein concentration at all the three levels. Significant breed variations were not observed in maternal IgY transfer from breeding hens to chicks and were 25.62±1.42-36.06±4.34% of total IgY in parent flock. Moderate to higher rate of seroprevalence with peak titers of 1:640 against Salmonella-specific antibodies was observed in only 41.6% of breeding hens. Conclusion: No significant difference in the rate of transfer of IgY was observed in four breeds studied (viz.,Vanraja, Gramapriya, BlackRock, and KalingaBrown and moderate seropositivity was detected for Salmonella-specific antibodies in progeny chicks.

  13. Prophylactic and therapeutic efficacy of avian antibodies against influenza virus H5N1 and H1N1 in mice.

    Directory of Open Access Journals (Sweden)

    Huan H Nguyen

    Full Text Available BACKGROUND: Pandemic influenza poses a serious threat to global health and the world economy. While vaccines are currently under development, passive immunization could offer an alternative strategy to prevent and treat influenza virus infection. Attempts to develop monoclonal antibodies (mAbs have been made. However, passive immunization based on mAbs may require a cocktail of mAbs with broader specificity in order to provide full protection since mAbs are generally specific for single epitopes. Chicken immunoglobulins (IgY found in egg yolk have been used mainly for treatment of infectious diseases of the gastrointestinal tract. Because the recent epidemic of highly pathogenic avian influenza virus (HPAIV strain H5N1 has resulted in serious economic losses to the poultry industry, many countries including Vietnam have introduced mass vaccination of poultry with H5N1 virus vaccines. We reasoned that IgY from consumable eggs available in supermarkets in Vietnam could provide protection against infections with HPAIV H5N1. METHODS AND FINDINGS: We found that H5N1-specific IgY that are prepared from eggs available in supermarkets in Vietnam by a rapid and simple water dilution method cross-protect against infections with HPAIV H5N1 and related H5N2 strains in mice. When administered intranasally before or after lethal infection, the IgY prevent the infection or significantly reduce viral replication resulting in complete recovery from the disease, respectively. We further generated H1N1 virus-specific IgY by immunization of hens with inactivated H1N1 A/PR/8/34 as a model virus for the current pandemic H1N1/09 and found that such H1N1-specific IgY protect mice from lethal influenza virus infection. CONCLUSIONS: The findings suggest that readily available H5N1-specific IgY offer an enormous source of valuable biological material to combat a potential H5N1 pandemic. In addition, our study provides a proof-of-concept for the approach using virus

  14. Cyber Situation Awareness through Instance-Based Learning: Modeling the Security Analyst in a Cyber-Attack Scenario

    Science.gov (United States)

    2012-01-01

    Chocolate Avenue Hershey PA 17033 Tel: 717-533-8845 Fax: 717-533-8661 E-mail: cust@igi-global.com Web site: http://www.igi-global.com Copyright © 2011...program and obtain control on the machine (event 21st out of 25). During the course of this simple scenario, a security analyst is able to observe...G. A. (1989). Recognition-primed deci- sions. In Rouse, W. B. (Ed.), Advances in man- machine system research (Vol. 5, pp. 47–92). Greenwich, CT

  15. From the Editor

    OpenAIRE

    Ugur DEMIRAY

    2011-01-01

    Dear readers of TOJDE,TOJDE appears on your screen now as Volume 12, Number: 3/2. This issue covers as a Special Issue on “Usage of Second Life Applications in Generally Education and Especially in Distance Education” field. Almost 9 months ago, I was planning to publish a book by agreement with IGI publishing, USA, on “Usage of Second Life Applications in Generally Education and Especially in Distance Education” field. And also recently, I had signed a draft contract with IGI Publishing too....

  16. Annals of the International Geophysical Year solar radio emission during the International Geophysical Year

    CERN Document Server

    Smerd, S F

    1969-01-01

    Annals of the International Geophysical Year, Volume 34: Solar Radio Emission During the International Geophysical Year covers the significant solar radio emission events observed during the International Geophysical Year (IGY). This book is composed of six chapters, and begins with a summary of tabulated quantities describing solar radio emission during the IGY. The tabulated figures illustrate the method of recording the position of radio sources on the sun, the use of symbols in describing the structure of bursts observed at single frequencies, and the different types used in a spectral

  17. Human Epidermal Growth Factor Receptor 2 Overexpression in Micropapillary and Other Variants of Urothelial Carcinoma.

    Science.gov (United States)

    Behzatoğlu, Kemal; Yörükoğlu, Kutsal; Demir, Hale; Bal, Nebil

    2016-06-21

    Human epidermal growth factor receptor 2 (HER2) protein overexpression or gene amplification has been shown in urothelial bladder cancer. This could be helpful when using targeted anti-HER2 therapy on these tumors. To evaluate HER2 immunohistochemical expression in conventional urothelial carcinoma (UC), in situ UC, and UC variants primarily in micropapillary urothelial carcinoma (MPUC). The study evaluated 60 MPUC cases; 25 invasive, 20 low-grade noninvasive, and 10 high-grade noninvasive UC cases; 8 in situ UC cases; and 69 UC variant cases. The immunohistochemistry staining was scored according to recommendations of the American Society of Clinical Oncology/College of American Pathologists 2013 HER2 test guideline established for breast cancer and only 3+ staining was considered HER2 overexpression. HER2 overexpression was determined by 3+ staining. 34 of 60 MPUC cases (56%) showed HER2 overexpression (3+ staining). We observed 3+ staining HER2 overexpression in nine of 25 conventional invasive UC cases (36%), four of eight in situ UC cases (50%), and three of six lipid cell variant cases (50%). 3+ staining HER2 overexpression was not seen in eight glandular, six small cell, and five sarcomatoid variant cases. HER2 overexpression was negative in the 20 low-grade noninvasive UC cases but positive in two of the 10 high-grade noninvasive UC cases (20%). We observed HER2 overexpression most commonly in MPUC cases. We also found HER2 overexpression in conventional invasive and in situ UC cases. Pure in situ UC and conventional invasive UC, especially MPUC, could be candidate tumors for treatment with anti-HER2 antibody (trastuzumab therapy). Targeted therapy has a limited place in treatment of bladder cancer. In this study, human epidermal growth factor receptor 2 (HER2) overexpression in bladder carcinomas was evaluated in a large number of cases. Anti-HER2 therapy could be used in bladder cancers, as in breast and gastric cancers. Copyright © 2016 European

  18. Novel paradigm for immunotherapy of ovarian cancer by engaging prophylactic immunity against hepatitis B virus.

    Science.gov (United States)

    Malecki, Marek; Putzer, Emily; Quach, Caroline; Dodivenaka, Chaitanya; Tombokan, Xenia

    2016-12-01

    Only eight women out of one hundred diagnosed with ovarian epithelial cancers, which progressed to the clinical stage IV, survive 10 years. First line therapies: surgery, chemotherapy, and radiation therapy inflict very serious iatrogenic consequences. Passive immunotherapy of ovarian cancers offers only low efficacy. Prophylactic and therapeutic vaccines for ovarian cancers are not available. Interestingly, prophylactic vaccines for Hepatitis B Viruses (HBV) are very effective. The specific aim of this work was to design, synthesize, and administer biomolecules, which would engage prophylactic, vaccination-induced immunity for HBV towards killing of ovarian cancer cells with high specificity and efficacy. Tissue biopsies, ascites, and blood were acquired from the patients, whose identities were entirely concealed in accordance with the Declaration of Helsinki, pursuant to the Institutional Review Board approval, and with the Patients' informed consent. By biomolecular engineering, we have created a novel family of biomolecules: antibody × vaccine engineered constructs (AVEC: anti-HER-2 × HBsAg). We have collected the blood from the volunteers, and measured the titers of anti-HBV antibodies resulting from the FDA approved and CDC scheduled HBV vaccinations. We have acquired tumor biopsies, ascites, and blood from patients suffering from the advanced ovarian cancers. We have established cultures of HER-2 over-expressing epithelial ovarian cancers: OV-90, TOC-112D, SKOV-3, as well as human ovary surface epithelial (HOSE) and human artery endothelial (HAE) cells. Treatment of the HER-2+ ovarian cancer cells with AVEC: anti-HER-2 × HBsAg, accompanied by administration of blood drawn from patients with high titers of the anti-HBV antibodies, resulted in much higher therapeutic efficacy as compared to treatment with the naked anti-HER-2 antibodies alone and/or with the relevant isotype antibodies. This treatment had practically no effect upon the HOSE and HAE

  19. Neoadjuvant therapy for early-stage breast cancer: the clinical utility of pertuzumab

    Directory of Open Access Journals (Sweden)

    Gollamudi J

    2016-02-01

    Full Text Available Jahnavi Gollamudi,1,* Jenny G Parvani,2,* William P Schiemann,3 Shaveta Vinayak3,4 1Department of Internal Medicine, 2Department of Biomedical Engineering, 3Case Comprehensive Cancer Center, Case Western Reserve University, 4Department of Hematology and Oncology, University Hospitals Case Medical Center, Cleveland, OH, USA *These authors contributed equally to this work Abstract: Approximately 20% of breast cancer patients harbor tumors that overexpress human epidermal growth factor receptor 2 (HER2; also known as ErbB2, a receptor tyrosine kinase that belongs to the epidermal growth factor receptor family of receptor tyrosine kinases. HER2 amplification and hyperactivation drive the growth and survival of breast cancers through the aberrant activation of proto-oncogenic signaling systems, particularly the Ras/MAP kinase and PI3K/AKT pathways. Although HER2-positive (HER2+ breast cancer was originally considered to be a highly aggressive form of the disease, the clinical landscape of HER2+ breast cancers has literally been transformed by the approval of anti-HER2 agents for adjuvant and neoadjuvant settings. Indeed, pertuzumab is a novel monoclonal antibody that functions as an anti-HER2 agent by targeting the extracellular dimerization domain of the HER2 receptor; it is also the first drug to receive an accelerated approval by the US Food and Drug Administration for use in neoadjuvant settings in early-stage HER2+ breast cancer. Here, we review the molecular and cellular factors that contribute to the pathophysiology of HER2 in breast cancer, as well as summarize the landmark preclinical and clinical findings underlying the approval and use of pertuzumab in the neoadjuvant setting. Finally, the molecular mechanisms operant in mediating resistance to anti-HER2 agents, and perhaps to pertuzumab as well, will be discussed, as will the anticipated clinical impact and future directions of pertuzumab in breast cancer patients. Keywords: breast cancer

  20. Radiation binary targeted therapy for HER-2 positive breast cancers: assumptions, theoretical assessment and future directions

    Energy Technology Data Exchange (ETDEWEB)

    Mundy, Daniel W [School of Nuclear Engineering, Purdue University, 400 Central Drive, West Lafayette, IN 47909 (United States); Harb, Wael [Horizon Oncology, The Care Group, Unity Medical Center, Lafayette, IN 47901 (United States); Jevremovic, Tatjana [School of Nuclear Engineering, Purdue University, 400 Central Drive, West Lafayette, IN 47909 (United States)

    2006-03-21

    A novel radiation targeted therapy is investigated for HER-2 positive breast cancers. The proposed concept combines two known approaches, but never used together for the treatment of advanced, relapsed or metastasized HER-2 positive breast cancers. The proposed radiation binary targeted concept is based on the anti HER-2 monoclonal antibodies (MABs) that would be used as vehicles to transport the nontoxic agent to cancer cells. The anti HER-2 MABs have been successful in targeting HER-2 positive breast cancers with high affinity. The proposed concept would utilize a neutral nontoxic boron-10 predicting that anti HER-2 MABs would assure its selective delivery to cancer cells. MABs against HER-2 have been a widely researched strategy in the clinical setting. The most promising antibody is Trastuzumab (Herceptin (registered) ). Targeting HER-2 with the MAB Trastuzumab has been proven to be a successful strategy in inducing tumour regression and improving patient survival. Unfortunately, these tumours become resistant and afflicted women succumb to breast cancer. In the proposed concept, when the tumour region is loaded with boron-10 it is irradiated with neutrons (treatment used for head and neck cancers, melanoma and glioblastoma for over 40 years in Japan and Europe). The irradiation process takes less than an hour producing minimal side effects. This paper summarizes our recent theoretical assessments of radiation binary targeted therapy for HER-2 positive breast cancers on: the effective drug delivery mechanism, the numerical model to evaluate the targeted radiation delivery and the survey study to find the neutron facility in the world that might be capable of producing the radiation effect as needed. A novel method of drug delivery utilizing Trastuzumab is described, followed by the description of a computational Monte Carlo based breast model used to determine radiation dose distributions. The total flux and neutron energy spectra of five currently available

  1. Lapatinib or Trastuzumab Plus Taxane Therapy for Human Epidermal Growth Factor Receptor 2-Positive Advanced Breast Cancer: Final Results of NCIC CTG MA.31.

    Science.gov (United States)

    Gelmon, Karen A; Boyle, Frances M; Kaufman, Bella; Huntsman, David G; Manikhas, Alexey; Di Leo, Angelo; Martin, Miguel; Schwartzberg, Lee S; Lemieux, Julie; Aparicio, Samuel; Shepherd, Lois E; Dent, Susan; Ellard, Susan L; Tonkin, Katia; Pritchard, Kathleen I; Whelan, Timothy J; Nomikos, Dora; Nusch, Arnd; Coleman, Robert E; Mukai, Hirofumi; Tjulandin, Sergei; Khasanov, Rustem; Rizel, Shulamith; Connor, Anne P; Santillana, Sergio L; Chapman, Judith-Anne W; Parulekar, Wendy R

    2015-05-10

    The efficacy of lapatinib versus trastuzumab combined with taxanes in the first-line setting of human epidermal growth factor receptor 2 (HER2) -positive metastatic breast cancer (BC) is unknown. The MA.31 trial compared a combination of first-line anti-HER2 therapy (lapatinib or trastuzumab) and taxane therapy for 24 weeks, followed by the same anti-HER2 monotherapy until progression. Stratification was by prior (neo)adjuvant anti-HER2 therapy, prior (neo)adjuvant taxane, planned taxane, and liver metastases. The primary end point was intention-to-treat (ITT) progression-free survival (PFS), defined as time from random assignment to progression by RECIST (version 1.0) criteria, or death for patients with locally assessed HER2-positive tumors. The primary test statistic was a stratified log-rank test for noninferiority. PFS was also assessed for patients with centrally confirmed HER2-positive tumors. From July 17, 2008, to December 1, 2011, 652 patients were accrued from 21 countries, resulting in 537 patients with centrally confirmed HER2-positive tumors. Median follow-up was 21.5 months. Median ITT PFS was 9.0 months with lapatinib and 11.3 months with trastuzumab. By ITT analysis, PFS was inferior for lapatinib compared with trastuzumab, with a stratified hazard ratio (HR) of 1.37 (95% CI, 1.13 to 1.65; P = .001). In patients with centrally confirmed HER2-positive tumors, median PFS was 9.1 months with lapatinib and 13.6 months with trastuzumab (HR, 1.48; 95% CI, 1.20 to 1.83; P < .001). More grade 3 or 4 diarrhea and rash were observed with lapatinib (P < .001). PFS results were supported by the secondary end point of overall survival, with an ITT HR of 1.28 (95% CI, 0.95 to 1.72; P = .11); in patients with centrally confirmed HER2-positive tumors, the HR was 1.47 (95% CI, 1.03 to 2.09; P = .03). As first-line therapy for HER2-positive metastatic BC, lapatinib combined with taxane was associated with shorter PFS and more toxicity compared with trastuzumab

  2. A single-domain antibody-linked Fab bispecific antibody Her2-S-Fab has potent cytotoxicity against Her2-expressing tumor cells.

    Science.gov (United States)

    Li, Aifen; Xing, Jieyu; Li, Li; Zhou, Changhua; Dong, Bin; He, Ping; Li, Qing; Wang, Zhong

    2016-12-01

    Her2, which is frequently overexpressed in breast cancer, is one of the most studied tumor-associated antigens for cancer therapy. Anti-HER2 monoclonal antibody, trastuzumab, has achieved significant clinical benefits in metastatic breast cancer. In this study, we describe a novel bispecific antibody Her2-S-Fab targeting Her2 by linking a single domain anti-CD16 VHH to the trastuzumab Fab. The Her2-S-Fab antibody can be efficiently expressed and purified from Escherichia coli, and drive potent cancer cell killing in HER2-overexpressing cancer cells. In xenograft model, the Her2-S-Fab suppresses tumor growth in the presence of human immune cells. Our results suggest that the bispecific Her2-S-Fab may provide a valid alternative to Her2 positive cancer therapy.

  3. Breast and gastrointestinal cancer updates from ASCO 2015.

    Science.gov (United States)

    Dawood, Shaheenah

    2015-01-01

    This review focuses on the updates presented at the ASCO 2015 symposium in breast and gastrointestinal malignancies. Some were practice changing while others gave us an exciting glimpse into what's to come in the very near future. Immunotherapy was the buzz word this year with data presented on every tumor site. Data on the efficacy of anti PD-1 agents in colorectal, hepatocellular and gastric cancer were presented. In breast cancer we saw data on a new and exciting therapeutic target in the form of androgen receptor among triple receptor negative breast tumors presented. Positive results of the PALOMA 3 trial were presented that has given women with hormone receptor positive metastatic breast cancer another therapeutic option. Furthermore data on strategies to further improve anti her2 therapy, optimizing of chemotherapy in the early and advanced stage and various strategies to improve endocrine therapy among patients with breast cancer were presented.

  4. New agents for the management of resistant metastatic breast cancer.

    Science.gov (United States)

    Anampa, Jesus; Sparano, Joseph A

    2017-12-01

    Metastatic breast cancer (MBC) is an incurable disease and treatment is directed towards symptom palliation and survival prolongation. Treatment selection in patients is based on tumor biology, age, comorbidities, performance status, tumor burden, and prior treatment history. Areas covered: This present review summarizes the recent treatment strategies in the management of MBC, highlighting regimens after first-line therapy. Topics discussed include new strategies for endocrine therapy, anti-HER2 therapy, and promising strategies for the management of triple negative breast cancer. Expert opinion: MBC is a heterogeneous entity and despite recent advances, there is significant room for improvement of treatment beyond first-line therapies. Combination regimens that can maximize clinical efficacy while minimizing toxicities are required. Current investigation approaches in advanced stages of clinical development include immunoconjugates, immune checkpoint blockade, novel cyclin-dependent-kinase inhibitors, and PARP inhibitors for MBC associated with germline BRCA mutations. We recommend that every patient with MBC should be evaluated for clinical trial options.

  5. [HER2 gene amplification assay: is CISH an alternative to FISH?].

    Science.gov (United States)

    Denoux, Yves; Arnould, Laurent; Fiche, Maryse; Lannes, B; Couturier, Jérôme; Vincent-Salomon, Anne; Penault-Llorca, Frédérique; Antoine, M; Balaton, A; Baranzelli, M C; Becette, V; Bellocq, J P; Bibeau, F; Ettore, F; Fridman, V; Gnassia, J P; Jacquemier, J; MacGrogan, G; Mathieu, M C; Migeon, C; Rigaud, C; Roger, P; Sigal-Zafrani, B; Simony-Lafontaine, J; Trassard, M; Treilleux, I; Verriele, V; Voigt, J J

    2003-12-01

    The HER2 proto-oncogene encodes a transmembrane protein, which is considered to function as a growth factor receptor. Overexpression of this protein found by immunohistochemistry in about 20% of infiltrating breast carcinomas, has a predictive value of response to treatment by trastuzumab, an anti-HER2 humanized monoclonal antibody. Search for HER2 gene amplification is necessary to adapt the immunohistochemical technique quality and also in the cases of delicate analysis or weak overexpression. It is usually carried out by Fluorescence In Situ Hybridization (FISH). A more recent hybridization technique, named CISH because of its chromogenic revelation is an alternative method, which gives highly correlated results with FISH. We present details of this technique, which may be more familiar for the pathologists than FISH, because reading analysis is similar to that of immunohistochemical staining.

  6. Self-assembling complexes of quantum dots and scFv antibodies for cancer cell targeting and imaging.

    Directory of Open Access Journals (Sweden)

    Tatiana A Zdobnova

    Full Text Available Semiconductor quantum dots represent a novel class of fluorophores with unique physical and chemical properties which could enable a remarkable broadening of the current applications of fluorescent imaging and optical diagnostics. Complexes of quantum dots and antibodies are promising visualising agents for fluorescent detection of selective biomarkers overexpressed in tumor tissues. Here we describe the construction of self-assembling fluorescent complexes of quantum dots and anti-HER1 or anti-HER2/neu scFv antibodies and their interactions with cultured tumor cells. A binding strategy based on a very specific non-covalent interaction between two proteins, barnase and barstar, was used to connect quantum dots and the targeting antibodies. Such a strategy allows combining the targeting and visualization functions simply by varying the corresponding modules of the fluorescent complex.

  7. HER-2-positive metastatic breast cancer: new possibilities for therapy

    Directory of Open Access Journals (Sweden)

    E. V. Artamonova

    2013-01-01

    Full Text Available This article is devoted to modern approaches in HER-2-positive metastatic breast cancer therapy. Recently treatment algorithm for this type of cancer included trastuzumab plus cytostatic in first line, continuation of trastuzumab with another chemotherapy regimen in second line, further switch to lapatinib and eventual return to trastuzumab after progression. Nowadays our options are broader owing to new anti-HER-2 agents which are pertruzumab and T-DM1. Now the most effective therapy regimen in first line is double HER-2 blockade (trastuzumab + pertuzumab in combination with docetaxel. Benefit of new agent T-DM1 versus combination of lapatinib and capecitabin is proved in patients progressed on trastuzumab and taxanes. T-DM1 also showed high efficacy as salvage therapy in intensively pretreated patients with meta- static HER-2-positive breast cancer who progressed on taxanes, trastuzumab and lapatinib.

  8. Identification of early breast cancer patient cohorts who may benefit from lapatinib therapy

    DEFF Research Database (Denmark)

    Strasser-Weippl, Kathrin; Horick, Nora; Smith, Ian E

    2016-01-01

    with HER2+ early breast cancer not treated with trastuzumab. We performed subgroup analyses and number-needed-to-treat (NNT) calculations using patient and tumour associated predictors. Hormone receptor negative (HR-) patients on lapatinib had a significantly prolonged disease-free survival (DFS) compared...... 5 years) was between 5.9 (node positive patients trastuzumab for HR unselected patients (e.g. 15.6 for DFS at 4 years in HERA). In a subgroup analysis of the adjuvant TEACH trial, we show...... that anti-HER2 monotherapy with a TKI is beneficial as adjuvant therapy in a subgroup of patients. NNT in HER2+ HR- patients are in range with those reported from up-front adjuvant trastuzumab trials....

  9. Radiobromination of monoclonal antibody using potassium [{sup 76}Br] (4 isothiocyanatobenzyl-ammonio)-bromo-decahydro-closo-dodecaborate (Bromo-DABI)

    Energy Technology Data Exchange (ETDEWEB)

    Bruskin, Alexander; Sivaev, Igor; Persson, Mikael; Lundqvist, Hans; Carlsson, Joergen; Sjoeberg, Stefan; Tolmachev, Vladimir E-mail: Vladimir.Tolmachev@bms.uu.se

    2004-02-01

    The use of charged linkers in attaching radiohalogens to tumor-seeking biomolecules may improve intracellular retention of the radioactive label after internalization and degradation of targeting proteins. Derivatives of polyhedral boron clusters, such as closo-dodecaborate (2-) anion, might be possible charged linkers. In this study, a bifunctional derivative of closo-dodecaborate, (4-isothiocyanatobenzyl-ammonio)-undecahydro-closo-dodecaborate (DABI) was labeled with positron-emitting nuclide {sup 76}Br (T = 16.2 h) and coupled to anti-HER2/neu humanized antibody Trastuzumab. The overall labeling yield at optimized conditions was 80.7 {+-} 0.6%. The label was proven to be stable in vitro in physiological and a set of denaturing conditions. The labeled antibody retained its capacity to bind to HER-2/neu antigen expressing cells. The results of the study demonstrated feasibility for using derivatives of closo-dodecaborate in indirect labeling of antibodies for radioimmunoPET.

  10. Optimal Synthetic Glycosylation of a Therapeutic Antibody.

    Science.gov (United States)

    Parsons, Thomas B; Struwe, Weston B; Gault, Joseph; Yamamoto, Keisuke; Taylor, Thomas A; Raj, Ritu; Wals, Kim; Mohammed, Shabaz; Robinson, Carol V; Benesch, Justin L P; Davis, Benjamin G

    2016-02-12

    Glycosylation patterns in antibodies critically determine biological and physical properties but their precise control is a significant challenge in biology and biotechnology. We describe herein the optimization of an endoglycosidase-catalyzed glycosylation of the best-selling biotherapeutic Herceptin, an anti-HER2 antibody. Precise MS analysis of the intact four-chain Ab heteromultimer reveals nonspecific, non-enzymatic reactions (glycation), which are not detected under standard denaturing conditions. This competing reaction, which has hitherto been underestimated as a source of side products, can now be minimized. Optimization allowed access to the purest natural form of Herceptin to date (≥90 %). Moreover, through the use of a small library of sugars containing non-natural functional groups, Ab variants containing defined numbers of selectively addressable chemical tags (reaction handles at Sia C1) in specific positions (for attachment of cargo molecules or "glycorandomization") were readily generated.

  11. Targeted nanodiamonds as phenotype-specific photoacoustic contrast agents for breast cancer.

    Science.gov (United States)

    Zhang, Ti; Cui, Huizhong; Fang, Chia-Yi; Cheng, Kun; Yang, Xinmai; Chang, Huan-Cheng; Forrest, M Laird

    2015-03-01

    The aim is to develop irradiated nanodiamonds (INDs) as a molecularly targeted contrast agent for high-resolution and phenotype-specific detection of breast cancer with photoacoustic (PA) imaging. The surface of acid treated radiation-damaged nanodiamonds was grafted with PEG to improve its stability and circulation time in blood, followed by conjugation to an anti-HER2 peptide with a final nanoparticle size of approximately 92 nm. Immunocompetent mice bearing orthotopic HER2-positive or negative tumors were administered INDs and PA imaged using an 820-nm near-infrared laser. PA images demonstrated that INDs accumulate in tumors and completely delineated the entire tumor within 10 h. HER2 targeting significantly enhanced imaging of HER2-positive tumors. Pathological examination demonstrated INDs are nontoxic. PA technology is adaptable to low-cost bedside medicine, and with new contrast agents described herein, PA can achieve high-resolution (sub-mm) and phenotype-specific monitoring of cancer growth.

  12. Liposomes derivatized with multimeric copies of KCCYSL peptide as targeting agents for HER-2-overexpressing tumor cells

    Directory of Open Access Journals (Sweden)

    Ringhieri P

    2017-01-01

    Full Text Available Paola Ringhieri,1 Silvia Mannucci,2 Giamaica Conti,2 Elena Nicolato,2 Giulio Fracasso,3 Pasquina Marzola,4 Giancarlo Morelli,1 Antonella Accardo1 1Department of Pharmacy and Interuniversity Research Centre on Bioactive Peptides (CIRPeB, University of Naples “Federico II”, Napoli, 2Department of Neurological Biomedical and Movement Sciences, 3Section of Immunology, Department of Medicine, 4Department of Informatics, University of Verona, Verona, Italy Abstract: Mixed liposomes, obtained by coaggregation of 1,2-dioleoyl-sn-glycero-3-phosphocholine and of the synthetic monomer containing a gadolinium complex ([C18]2DTPA[Gd] have been prepared. Liposomes externally decorated with KCCYSL (P6.1 peptide sequence in its monomeric, dimeric, and tetrameric forms are studied as target-selective delivery systems toward cancer cells overexpressing human epidermal growth factor receptor-2 (HER-2 receptors. Derivatization of liposomal surface with targeting peptides is achieved using the postmodification method: the alkyne-peptide derivative Pra-KCCYSL reacts, through click chemistry procedures, with a synthetic surfactant modified with 1, 2, or 4 azido moieties previously inserted in liposome formulation. Preliminary in vitro data on MDA-MB-231 and BT-474 cells indicated that liposomes functionalized with P6.1 peptide in its tetrameric form had better binding to and uptake into BT-474 cells compared to liposomes decorated with monomeric or dimeric versions of the P6.1 peptide. BT-474 cells treated with liposomes functionalized with the tetrameric form of P6.1 showed high degree of liposome uptake, which was comparable with the uptake of anti-HER-2 antibodies such as Herceptin. Moreover, magnetic MRI experiments have demonstrated the potential of liposomes to act as MRI contrast agents. Keywords: anti-HER2 liposomes, target peptide, KCCYSL peptide, breast cancer, click chemistry, branched peptides 

  13. Phase I clinical trial of HER2-specific immunotherapy with concomitant HER2 kinase inhibtion

    Directory of Open Access Journals (Sweden)

    Hamilton Erika

    2012-02-01

    Full Text Available Abstract Background Patients with HER2-overexpressing metastatic breast cancer, despite initially benefiting from the monoclonal antibody trastuzumab and the EGFR/HER2 tyrosine kinase inhibitor lapatinib, will eventually have progressive disease. HER2-based vaccines induce polyclonal antibody responses against HER2 that demonstrate enhanced anti-tumor activity when combined with lapatinib in murine models. We wished to test the clinical safety, immunogenicity, and activity of a HER2-based cancer vaccine, when combined with lapatinib. Methods We immunized women (n = 12 with metastatic, trastuzumab-refractory, HER2-overexpressing breast cancer with dHER2, a recombinant protein consisting of extracellular domain (ECD and a portion of the intracellular domain (ICD of HER2 combined with the adjuvant AS15, containing MPL, QS21, CpG and liposome. Lapatinib (1250 mg/day was administered concurrently. Peripheral blood antibody and T cell responses were measured. Results This regimen was well tolerated, with no cardiotoxicity. Anti-HER2-specific antibody was induced in all patients whereas HER2-specific T cells were detected in one patient. Preliminary analyses of patient serum demonstrated downstream signaling inhibition in HER2 expressing tumor cells. The median time to progression was 55 days, with the majority of patients progressing prior to induction of peak anti-HER2 immune responses; however, 300-day overall survival was 92% (95% CI: 77-100%. Conclusions dHER2 combined with lapatinib was safe and immunogenic with promising long term survival in those with HER2-overexpressing breast cancers refractory to trastuzumab. Further studies to define the anticancer activity of the antibodies induced by HER2 vaccines along with lapatinib are underway. Trial registry ClinicalTrials.gov NCT00952692

  14. A prospective, non-randomized phase II trial of Trastuzumab and Capecitabine in patients with HER2 expressing metastasized pancreatic cancer

    Directory of Open Access Journals (Sweden)

    Endlicher Esther

    2009-01-01

    Full Text Available Abstract Background Pancreatic cancer is the fourth most common cause of cancer related death in Western countries. Advantages in surgical techniques, radiation and chemotherapy had almost no impact on the long term survival of affected patients. Therefore, the need for better treatment strategies is urgent. HER2, a receptor tyrosine kinase of the EGFR family, involved in signal transduction pathways leading to cell growth and differentiation is overexpressed in a number of cancers, including breast and pancreatic cancer. While in breast cancer HER2 has already been successfully used as a treatment target, there are only limited data evaluating the effects of inhibiting HER2 tyrosine kinases in patients with pancreatic cancer. Methods Here we report the design of a prospective, non-randomized multi-centered Phase II clinical study evaluating the effects of the Fluoropyrimidine-carbamate Capecitabine (Xeloda ® and the monoclonal anti-HER2 antibody Trastuzumab (Herceptin® in patients with non-resectable, HER2 overexpressing pancreatic cancer. Patients eligible for the study will receive Trastuzumab infusions on day 1, 8 and 15 concomitant to the oral intake of Capecitabine from day 1 to day 14 of each three week cylce. Cycles will be repeated until tumor progression. A total of 37 patients will be enrolled with an interim analysis after 23 patients. Discussion Primary end point of the study is to determine the progression free survival after 12 weeks of bimodal treatment with the chemotherapeutic agent Capecitabine and the anti-HER2 antibody Trastuzumab. Secondary end points include patient's survival, toxicity analysis, quality of life, the correlation of HER2 overexpression and clinical response to Trastuzumab treatment and, finally, the correlation of CA19-9 plasma levels and progression free intervals.

  15. Breast Cancer Immunotherapy

    Institute of Scientific and Technical Information of China (English)

    Juhua Zhou; Yin Zhong

    2004-01-01

    Breast cancer is a leading cause of cancer-related deaths in women worldwide. Although tumorectomy,radiotherapy, chemotherapy and hormone replacement therapy have been used for the treatment of breast cancer, there is no effective therapy for patients with invasive and metastatic breast cancer. Immunotherapy may be proved effective in treating patients with advanced breast cancer. Breast cancer immunotherapy includes antibody based immunotherapy, cancer vaccine immunotherapy, adoptive T cell transfer immunotherapy and T cell receptor gene transfer immunotherapy. Antibody based immunotherapy such as the monoclonal antibody against HER-2/neu (trastuzumab) is successfully used in the treatment of breast cancer patients with over-expressed HER-2/neu, however, HER-2/neu is over-expressed only in 25-30% of breast cancer patients. Cancer vaccine immunotherapy is a promising method to treat cancer patients. Cancer vaccines can be used to induce specific anti-tumor immunity in breast cancer patients, but cannot induce objective tumor regression. Adoptive T cell transfer immunotherapy is an effective method in the treatment of melanoma patients. Recent advances in anti-tumor T cell generation ex vivo and limited clinical trial data have made the feasibility of adoptive T cell transfer immunotherapy in the treatment of breast cancer patients. T cell receptor gene transfer can redirect the specificity of T cells. Chimeric receptor, scFv(anti-HER-2/neu)/zeta receptor, was successfully used to redirect cytotoxic T lymphocyte hybridoma cells to obtain anti-HER-2/neu positive tumor cells, suggesting the feasibility of treatment of breast cancer patients with T cell receptor gene transfer immunotherapy. Clinical trials will approve that immunotherapy is an effective method to cure breast cancer disease in the near future.

  16. Breast Cancer Immunotherapy

    Institute of Scientific and Technical Information of China (English)

    JuhuaZhou; YinZhong

    2004-01-01

    Breast cancer is a leading cause of cancer-related deaths in women worldwide. Although tumorectomy, radiotherapy, chemotherapy and hormone replacement therapy have been used for the treatment of breast cancer, there is no effective therapy for patients with invasive and metastatic breast cancer. Immunotherapy may be proved effective in treating patients with advanced breast cancer. Breast cancer immunotherapy includes antibody based immunotherapy, cancer vaccine immunotherapy, adoptive T cell transfer immunotherapy and T cell receptor gene transfer immunotherapy. Antibody based immunotherapy such as the monoclonal antibody against HER-2/neu (trastuzumab) is successfully used in the treatment of breast cancer patients with over-expressed HER-2/neu, however, HER-2/neu is over-expressed only in 25-30% of breast cancer patients. Cancer vaccine immunotherapy is a promising method to treat cancer patients. Cancer vaccines can be used to induce specific anti-tumor immunity in breast cancer patients, but cannot induce objective tumor regression. Adoptive T cell transfer immunotherapy is an effective method in the treatment of melanoma patients. Recent advances in anti-tumor T cell generation ex vivo and limited clinical trial data have made the feasibility of adoptive T cell transfer immunotherapy in the treatment of breast cancer patients. T cell receptor gene transfer can redirect the specificity of T cells. Chimeric receptor, scFv(anti-HER-2/neu)/zeta receptor, was successfully used to redirect cytotoxic T lymphocyte hybridoma cells to obtain anti-HER-2/neu positive tumor cells, suggesting the feasibility of treatment of breast cancer patients with T cell receptor gene transfer immunotherapy. Clinical trials will approve that immunotherapy is an effective method to cure breast cancer disease in the near future. Cellular & Molecular Immunology.

  17. Emerging treatments for HER2-positive early-stage breast cancer: focus on neratinib

    Directory of Open Access Journals (Sweden)

    Kourie HR

    2017-07-01

    Full Text Available Hampig Raphael Kourie,1 Elie El Rassy,1 Florian Clatot,2,3 Evandro de Azambuja,4 Matteo Lambertini3,4 1Department of Oncology, Faculty of Medicine, Saint Joseph University, Beirut, Lebanon; 2Department of Medical Oncology and IRON/U1245, Centre Henri Becquerel, Rouen, France; 3Breast Cancer Translational Research Laboratory, 4Department of Medicine, Institut Jules Bordet, Université Libre de Bruxelles, Brussels, Belgium Abstract: Over the last decades, a better understanding of breast cancer heterogeneity provided tools for a biologically based personalization of anticancer treatments. In particular, the overexpression of the human epidermal growth factor receptor 2 (HER2 by tumor cells provided a specific target in these HER2-positive tumors. The development of the monoclonal antibody trastuzumab, and its approval in 1998 for the treatment of patients with metastatic disease, radically changed the natural history of this aggressive subtype of breast cancer. These findings provided strong support for the continuous research in targeting the HER2 pathway and implementing the development of new anti-HER2 targeted agents. Besides trastuzumab, a series of other anti-HER2 agents have been developed and are currently being explored for the treatment of breast cancer patients, including those diagnosed with early-stage disease. Among these agents, neratinib, an oral tyrosine kinase inhibitor that irreversibly inhibits HER1, HER2, and HER4 at the intracellular level, has shown promising results, including when administered to patients previously exposed to trastuzumab-based treatment. This article aims to review the available data on the role of the HER2 pathway in breast cancer and on the different targeted agents that have been studied or are currently under development for the treatment of patients with early-stage HER2-positive disease with a particular focus on neratinib. Keywords: breast cancer, HER2-positive, neratinib, early-stage, targeted

  18. Near-infrared quantum dots for HER2 localization and imaging of cancer cells.

    Science.gov (United States)

    Rizvi, Sarwat B; Rouhi, Sepideh; Taniguchi, Shohei; Yang, Shi Yu; Green, Mark; Keshtgar, Mo; Seifalian, Alexander M

    2014-01-01

    Quantum dots are fluorescent nanoparticles with unique photophysical properties that allow them to be used as diagnostic, therapeutic, and theranostic agents, particularly in medical and surgical oncology. Near-infrared-emitting quantum dots can be visualized in deep tissues because the biological window is transparent to these wavelengths. Their small sizes and free surface reactive groups that can be conjugated to biomolecules make them ideal probes for in vivo cancer localization, targeted chemotherapy, and image-guided cancer surgery. The human epidermal growth factor receptor 2 gene (HER2/neu) is overexpressed in 25%-30% of breast cancers. The current methods of detection for HER2 status, including immunohistochemistry and fluorescence in situ hybridization, are used ex vivo and cannot be used in vivo. In this paper, we demonstrate the application of near-infrared-emitting quantum dots for HER2 localization in fixed and live cancer cells as a first step prior to their in vivo application. Near-infrared-emitting quantum dots were characterized and their in vitro toxicity was established using three cancer cell lines, ie, HepG2, SK-BR-3 (HER2-overexpressing), and MCF7 (HER2-underexpressing). Mouse antihuman anti-HER2 monoclonal antibody was conjugated to the near-infrared-emitting quantum dots. In vitro toxicity studies showed biocompatibility of SK-BR-3 and MCF7 cell lines with near-infrared-emitting quantum dots at a concentration of 60 μg/mL after one hour and 24 hours of exposure. Near-infrared-emitting quantum dot antiHER2-antibody bioconjugates successfully localized HER2 receptors on SK-BR-3 cells. Near-infrared-emitting quantum dot bioconjugates can be used for rapid localization of HER2 receptors and can potentially be used for targeted therapy as well as image-guided surgery.

  19. Validated biomarkers: The key to precision treatment in patients with breast cancer.

    Science.gov (United States)

    Duffy, Michael J; O'Donovan, Norma; McDermott, Enda; Crown, John

    2016-10-01

    Recent DNA sequencing and gene expression studies have shown that at a molecular level, almost every case of breast cancer is unique and different from other breast cancers. For optimum management therefore, every patient should receive treatment that is guided by the molecular composition of their tumor, i.e., precision treatment. While such a scenario is still some distance into the future, biomarkers are beginning to play an important role in preparing the way for precision treatment. In particular, biomarkers are increasingly being used for predicting patient outcome and informing as to the most appropriate type of systemic therapy to be administered. Mandatory biomarkers for every newly diagnosed case of breast cancer are estrogen receptors and progesterone receptors in selecting patients for endocrine treatment and HER2 for identifying patients likely to benefit from anti-HER2 therapy. Amongst the best validated prognostic biomarker tests are uPA/PAI-1, MammaPrint and Oncotype DX. Although currently, there are no biomarkers available for predicting response to specific forms of chemotherapy, uPA/PAI-1 and Oncotype DX can aid the identification of lymph node-negative patients that are most likely to benefit from adjuvant chemotherapy, in general. In order to accelerate progress towards precision treatment for women with breast cancer, we need additional predictive biomarkers, especially for enhancing the positive predictive value for endocrine and anti-HER2 therapies, as well as biomarkers for predicting response to specific forms of chemotherapy. The ultimate biomarker test for achieving the goal of precision treatment for patients with breast cancer will likely require a combination of gene sequencing and transcriptomic analysis of every patient's tumor. Copyright © 2016 Elsevier Ltd. All rights reserved.

  20. A sandwich-type electrochemical immunosensor based on in situ silver deposition for determination of serum level of HER2 in breast cancer patients.

    Science.gov (United States)

    Shamsipur, Mojtaba; Emami, Mahdi; Farzin, Leila; Saber, Reza

    2018-04-30

    The sensitive quantification of Human Epidermal growth factor Receptor 2 (HER2), as a key prognostic tumor marker, plays a critical role in screening, early diagnosis and management of breast cancer. This paper describes a sandwich-type immunoassay with silver signal enhancement strategy for highly sensitive detection of HER2. For this purpose, the target capturing step was designed by functionalization of 3-aminopropyltrimethoxysilane coated magnetite nanoparticles with antibody (antiHER2/APTMS-Fe 3 O 4 ), as a platform bioconjugate (PB), and immobilized at a bare GCE. Then, in the presence of label-free immunosensor, the PB was covered by magnetic gold nanoparticles self-assembled with thiolated antibodies (antiHER2/Hyd@AuNPs-APTMS-Fe 3 O 4 ) containing chemically reduced silver ions, as a label bioconjugate (LB). Under optimum conditions, a linear relationship between the differential pulse voltammetric (DPV) stripping signal of silver and the logarithm of HER2 concentrations was obtained in the range of 5.0 × 10 -4 -50.0ngmL -1 (R 2 = 0.9906) with a detection limit of 2.0 × 10 -5 ngmL -1 . The effectiveness of this protocol was evaluated experimentally through employing of designed immunosensor for detection of the serum level of tumor marker. The good consistency of the results with those obtained by the enzyme-linked immunosorbent assay (ELISA) conventional method (p-value of < 0.05) showed that this immunosensor can be applied for the testing of HER2 in clinical samples of breast cancer patients. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Passive Immune-Protection of Litopenaeus vannamei against Vibrio harveyi and Vibrio parahaemolyticus Infections with Anti-Vibrio Egg Yolk (IgY-Encapsulated Feed

    Directory of Open Access Journals (Sweden)

    Xiaojian Gao

    2016-05-01

    Full Text Available Vibrio spp. are major causes of mortality in white shrimp (Litopenaeus vannamei which is lacking adaptive immunity. Passive immunization with a specific egg yolk antibody (IgY is a potential method for the protection of shrimp against vibriosis. In this study, immune effects of the specific egg yolk powders (IgY against both V. harveyi and V. parahaemolyticus on white shrimp were evaluated. The egg yolk powders against V. harveyi and V. parahaemolyticus for passive immunization of white shrimp were prepared, while a tube agglutination assay and an indirect enzyme-linked immunosorbent assay (ELISA were used for detection of IgY titer. Anti-Vibrio egg yolk was encapsulated by β-cyclodextrin, which could keep the activity of the antibody in the gastrointestinal tract of shrimp. The results showed that the anti-Vibrio egg powders had an inhibiting effect on V. harveyi and V. parahaemolyticus in vitro. Lower mortality of infected zoeae, mysis, and postlarva was observed in groups fed with anti-Vibrio egg powders, compared with those fed with normal egg powders. The bacterial load in postlarva fed with specific egg powders in seeding ponds was significantly lower than those fed with normal egg powders in seeding ponds. These results show that passive immunization by oral administration with specific egg yolk powders (IgY may provide a valuable protection of vibrio infections in white shrimp.

  2. An immunoenzymatic assay for the diagnosis of hepatitis A utilising immunoglobulin Y

    Directory of Open Access Journals (Sweden)

    Alexandre dos Santos da Silva

    2012-11-01

    Full Text Available The detection of anti-hepatitis A virus (HAV antibody levels by diagnostic kits in the convalescent period of disease generally use immunoglobulin G (IgG, which is expensive. An alternative to IgG is immunoglobulin Y (IgY, an immunoglobulin antibody encountered in birds and reptiles. The aim of this study was to develop a competitive immunoenzymatic assay to measure total anti-HAV antibody levels using anti-HAV IgY as the capture and conjugated immunoglobulins. For this purpose, anti-HAV IgY was conjugated to horseradish peroxidase (HRP and the optimal dilution of HRP-conjugated antibodies was evaluated to establish the competitive immuneenzymatic assay. The results obtained from our "in-house" assay were plotted on a receiver operator curve, which showed a sensitivity of 95% and a specificity of 98.8%, demonstrating that a competitive anti-HAV IgY immunoenzymatic assay developed "in house" could be used as an alternative to commercial assays that utilise IgG.

  3. Elisa development for detection of glyphosat resistant gm soybean

    Directory of Open Access Journals (Sweden)

    Владислав Геннадійович Спиридонов

    2015-11-01

    Full Text Available During research we have utilized recombinant enzyme 5-enolpyruvylshikimate-3-phosphate synthase (CP4 EPSPS, conferring resistance to glyphosate for GM soybean, for the hen immunization and obtaining specific yolk antibodies IgY. Stages of ELISA development that can detect at least 0,1 % of GM-soybean resistant to glyphosate were present

  4. Successive immunoglobulin and cytokine expression in the small intestine of juvenile chicken

    NARCIS (Netherlands)

    Lammers, A.; Wieland, W.H.; Kruijt, L.; Jansma, A.; Straetemans, T.; Schots, A.; Hartog, den C.G.; Parmentier, H.K.

    2010-01-01

    The intestinal mucosa is of major importance for immune development. To further study the ontogeny of avian mucosal immunity, mRNA levels of IgM, IgY and IgA, the polymeric immunoglobulin receptor (pIgR) and a number of cytokines were determined at different ages in jejunum and ileum of

  5. Development of hen antihepatitis B antigen IgY-based conjugate for ELISA assay

    Directory of Open Access Journals (Sweden)

    Najat Muayed Nafea

    2015-01-01

    Conclusions: This study showed that laying hens can be used as an alternative source for production of polyclonal antibodies against HBsAg and anti-HBs IgY could be labeled with HRP enzyme and could subsequently be used successfully as secondary antibody in ELISA for detection of HBsAg in the patients sera.

  6. Optimizing Two-Color Semiconductor Nanocrystal Immunoassays in Single Well Microtiter Plate Formats

    Directory of Open Access Journals (Sweden)

    W. Russ Algar

    2011-08-01

    Full Text Available The simultaneous detection of two analytes, chicken IgY (IgG and Staphylococcal enterotoxin B (SEB, in the single well of a 96-well plate is demonstrated using luminescent semiconductor quantum dot nanocrystal (NC tracers. The NC-labeled antibodies were prepared via sulfhydryl-reactive chemistry using a facile protocol that took

  7. Scale-Independent Relational Query Processing

    Science.gov (United States)

    2013-10-04

    source options are also available, including Postgresql, MySQL , and SQLite. These mod- ern relational databases are generally very complex software systems...and Their Application to Data Stream Management. IGI Global, 2010. [68] George Reese. Database Programming with JDBC and Java , Second Edition. Ed. by

  8. Game-Based Life-Long Learning

    NARCIS (Netherlands)

    Kelle, Sebastian; Sigurðarson, Steinn; Westera, Wim; Specht, Marcus

    2010-01-01

    Kelle, S., Sigurðarson, S., Westera, W., & Specht, M. (2011). Game-Based Life-Long Learning. In G. D. Magoulas (Ed.), E-Infrastructures and Technologies for Lifelong Learning: Next Generation Environments (pp. 337-349). Hershey, PA: IGI Global.

  9. Generation of Anti-Boa Immunoglobulin Antibodies for Serodiagnostic Applications, and Their Use to Detect Anti-Reptarenavirus Antibodies in Boa Constrictor.

    Directory of Open Access Journals (Sweden)

    Yegor Korzyukov

    Full Text Available Immunoglobulins (Igs, the key effectors of the adaptive immune system, mediate the specific recognition of foreign structures, i.e. antigens. In mammals, IgM production commonly precedes the production of IgG in the response to an infection. The reptilian counterpart of IgG is IgY, but the exact kinetics of the reptilian immune response are less well known. Boid inclusion body disease (BIBD, an often fatal disease of captive boas and pythons has been linked to reptarenavirus infection, and BIBD is believed to be immunosuppressive. However, so far, the study of the serological response towards reptarenaviruses in BIBD has been hampered by the lack of reagents. Thus we set up a purification protocol for boa constrictor IgY and IgM, which should also be applicable for other snake species. We used centrifugal filter units, poly ethylene glycol precipitation and gel permeation chromatography to purify and separate the IgM and IgY fractions from boa constrictor serum, which we further used to immunise rabbits. We affinity purified IgM and IgY specific reagents from the produced antiserum, and labelled the reagents with horseradish peroxidase. Finally, using the sera of snakes with known exposure to reptarenaviruses we demonstrated that the newly generated reagents can be utilised for serodiagnostic purposes, such as immunoblotting and immunofluorescent staining. To our knowledge, this is the first report to show reptarenavirus-specific antibodies in boa constrictors.

  10. Measuring the Style of Innovative Thinking among Engineering Students

    Science.gov (United States)

    Passig, David; Cohen, Lizi

    2014-01-01

    Background: Many tools have been developed to measure the ability of workers to innovate. However, all of them are based on self-reporting questionnaires, which raises questions about their validity Purpose: The aim was to develop and validate a tool, called Ideas Generation Implementation (IGI), to objectively measure the style and potential of…

  11. Guiding histological assessment of uterine lesions using 3D in vitro ultrasonography and stereotaxis

    DEFF Research Database (Denmark)

    Vandermeulen, Liselore; Cornelis, Ann; Rasmussen, Christina Kjærgaard

    2017-01-01

    , the uterus was re-evaluated by 3D in vitro ultrasonography and in vitro gel instillation sonography (iGIS). The lesion of interest was pinpointed by inserting an intramuscular injection needle using a free-hand 2D-ultrasound guided technique to focus the macroscopic and the microscopic examination...

  12. Looking Forward to the electronic Geophysical Year

    Science.gov (United States)

    Kamide, Y.; Baker, D. N.; Thompson, B.; Barton, C.; Kihn, E.

    2004-12-01

    During the International Geophysical Year (1957-1958), member countries established many new capabilities pursuing the major IGY objectives of collecting geophysical data as widely as possible and providing free access to these data for all scientists around the globe. A key achievement of the IGY was the establishment of a worldwide system of data centers and physical observatories. The worldwide scientific community has now endorsed and is promoting an electronic Geophysical Year (eGY) initiative. The proposed eGY concept would both commemorate the 50th anniversary of the IGY in 2007-2008 and would provide a forward impetus to geophysics in the 21st century, similar to that provide by the IGY fifty years ago. The eGY concept advocates the establishment of a series of virtual geophysical observatories now being deployed in cyberspace. We discuss plans to aggregate measurements into a readily accessible database along with analysis, visualization, and display tools that will make information available and useful to the scientific community, to the user community, and to the general public. We are examining the possibilities for near-realtime acquisition of data and utilization of forecast tools in order to provide users with advanced space weather capabilities. This program will provide powerful tools for education and public outreach concerning the connected Sun-Earth System.

  13. An innovative biocatalyst for production of ethanol from xylose in a continuous bioreactor.

    Science.gov (United States)

    Silva, C R; Zangirolami, T C; Rodrigues, J P; Matugi, K; Giordano, R C; Giordano, R L C

    2012-01-05

    The use of the hemicellulose fraction of biomass may be important for the feasibility of the production of second generation bioethanol. Wild strains of Saccharomyces cerevisiae are widely used in industry for production of 1st generation ethanol, and the robustness of this yeast is an important advantage in large scale applications. Isomerization of xylose to xylulose is an essential step in this process. This reaction is catalyzed by glucose isomerase (GI). A new biocatalyst is presented here for the simultaneous isomerization and fermentation (SIF) of xylose. GI from Streptomyces rubiginosus was immobilized in chitosan, through crosslinking with glutaraldehyde, and the support containing the immobilized GI (IGI-Ch) was co-immobilized with S. cerevisiae, in calcium alginate gel. The immobilization experiments led to high immobilized protein loads (30-68 mg × g(support)(-1)), high yields (circa of 100%) and high recovered enzyme activity (>90%). The IGI-Ch derivative with maximum activity presented 1700 IU × g(catalyst)(-1), almost twice the activity of a commercial immobilized GI, GENSWEET(®) IGI-HF. At typical operational conditions for xylose SIF operation (pH 5, 30-35 °C, presence of nutrients and ethanol concentrations in the medium up to 70 L(-1)), both derivatives, IGI-Ch and GENSWEET(®) IGI-HF retained app. 90% of the initial activity after 120 h, while soluble GI was almost completely inactive at pH 5, 30 °C. The isomerization xylose/xylulose, catalyzed by IGI-Ch, reached the equilibrium in batch experiments after 4h, with 12,000 IU × L(-1) (7 g(der) × L(-1)), at pH 5 and 30 °C, in the presence of fermentation nutrients. After co-immobilization of IGI-Ch with yeast in alginate gel, this biocatalyst succeeded in producing 12 g × L(-1) of ethanol, 9.5 g × L(-1) of xylitol, 2.5 g × L(-1) of glycerol and 1.9 g × L(-1) of acetate after consumption of 50 g × L(-1) of xylose, in 48 h, using 32.5 × 10(3) IU × L(-1) and 20 g(yeast) × L(-1), at 35

  14. 抗肠道病毒71型特异性卵黄抗体的制备及鉴定%Preparation and identification of specific chicken egg yolk immunoglobulins against enterovirus 71

    Institute of Scientific and Technical Information of China (English)

    朱思思; 赵肃清; 李月明; 温俊林

    2011-01-01

    0bjsctive iPreparhg specificity chicken egg yolk antibody (IgY ) against enteiovius 71 (EV 71) and detecting its biobgical activities,this is significantfor prevention and treatnentof EV 71 infection .M ethods:Fourhensweie inmurmed Ihree tineswiih tie purified him an EV71 virus as an antigen .The inmunogbbulh Y (IgY ) antibody was extracted and purified from tie egg yolk by water combine pofyeihylene gfycol (water-peg) m eihod .The yield and molecularm ass of IgY were analysed by Bradford m eihod and SDS-PAGE electrophore- sis.ELBA and A gar double in munodiffusbn were used for detect 1he titer of IgY ,while tie inmunoreactivily of antHiV71 IgY was tested using W estem bbt.Results :The yield and purily of IgY were 7 .76 m g/m land 94.86% respective^ .Results of ELBA hdicated 1hat1he specific an- trEV71 IgY antibody appealed in tie egg yolk 10 days afterprinary inmuniation and reached 1he highest titer 1 = 20 480 after 40 days,while 1he titer of A gar double in m unodiffusion was 1 = 16 .W estem bbt analysis revealed lhattie purified IgY antibody owned good inmunoreactivily and a specifically combination capacily of EV71 virus.Conclusbn :Egg yolk is a large scale source of antidodys specifically against EV 71 virus .The antrEV 71 IgY extracted and purified by W aterPEG m etiod possess good production ,purtly ,and bbbgical activities.%目的:制备特异性抗肠道病毒71型(Enterovirus 71,EV71)的卵黄抗体并检测其生物学性能,这对EV71感染手足口病的预防和治疗具有重大的意义.方法:用纯化并灭活后EV71作为抗原免疫健康产蛋鸡.采用本实验室水提综合聚乙二醇法提取纯化鸡卵黄抗体;用Bradford法测定卵黄抗体提取液的蛋白含量;还原性SDS-PAGE凝胶电泳分析测定提取蛋白的纯度及相对分子质量;分别用ELISA、双向免疫琼脂扩散检测纯化特异性卵黄抗体抗EV71效价;Western blot分析特异性卵黄抗体的免疫反应性.结果:卵黄中卵黄抗体含量达7.76 mg

  15. Leveraging the International Polar Year Legacy: Providing Historical Perspective for IPY Education, Outreach and Communication Efforts

    Science.gov (United States)

    Tsukernik, M.; McCaffrey, M. S.

    2006-12-01

    As the International Polar Year 2007-2008 (IPY) is fast approaching, it is important to look back and learn from the previous experience. Over 125 years ago, when an Austrian explorer and naval officer Lt. Karl Weyprecht called for an international yearlong intensive effort to study the Polar Regions, he probably never imagined that his model for international collaboration would become so widely popular. Frustrated by the lack of coordinated, international collaboration in research activities, Weyprecht proposed an intensive burst of research activity over the course of at least a year. The first IPY began in 1882 with 12 nations establishing 13 stations in the Arctic and 2 in the Southern Hemisphere. The initial yearlong plan did not go beyond data collection. However, the idea lived in the minds of scientists worldwide and the second IPY followed the first one 50 years later. By 1932, technology evolved significantly, and on top of ground-based meteorological and geophysical measurements, data collection also included radiosonde and acoustic atmospheric measurements. Occurring during a global economic depression, and between world wars, the second IPY faced many challenges. However, 40 permanent stations were established, some of which are still active. Scientific exploration also reached remote frontiers from Antarctica to the Earth's ionosphere. Less than a decade after the WWII, the idea of the next IPY started to circulate in scientific circles. The world was focused on space exploration and the word "polar" seemed too narrow for the gigantic projects planned for the 1957. That is why the initial idea of the third IPY evolved into the International Geophysical Year (IGY), although polar regions were still a major focus. The success of the IGY is almost overwhelming the first Earth orbiting satellites, a traverse of Antarctica, a discovery of the Radiation Belt, a series of science education films about IGY activities and research themes are just a few

  16. [Isolation of cariogenic Streptococcus mutans and the inhibitory effect of egg yolk antibody on caries].

    Science.gov (United States)

    Li, X L; Zhang, Z; Li, Z X; Deng, N J; Zeng, B; Chen, Y M

    2017-04-09

    Objective: To isolate the cariogenic Streptococcus mutans (Sm) strains and study the therapeutical effect of egg yolk antibody (IgY) of the Sm on dental caries development. Methods: Sm strains were isolated from the children's dental plaque samples. Morphological, biochemical and molecular biological methods were applied to identify the serotype, acid producing and adhesion abilities of isolated Sm strains. After inactivation one of the Sm strains was used as antigen to immune laying hens to collect and extract the specific anti-Sm IgY. The rats were infected with Sm (serotype e). After 16 weeks of infection, all the rats were found developing dental caries. The rats were then randomly divided into two groups. The rats in experimental group were supplied with diet containing anti-Sm IgY while the rats in control group with normal IgY. All rats were sacrificed after another 8 weeks' observation. The degree of caries for each rat was assessed using Keyes' method. Results: We isolated 7 Sm strains from the children's dental plaque samples in the present study. The numbers of serotype c, e, f, k were 3, 2, 0 and 2, respectively. All strains showed similar morphological and biochemical characters as standard UA159 Sm strain, and possessed strong capabilities of acid production and adherence. Interestingly, even the same serotypec strains, such as No.3 and No.7 strains, demonstrated significant difference on acid producing and adherence capabilities. After 16 weeks infection with serotype e strain, the rats' mandibular teeth were apparently decayed, and treatment with specific anti-Sm IgY obviously attenuated the development of caries in the experiment group rats (16.4±2.0) compared with that in the control group rats (30.2±9.3) ( Pcariogenic Sm strains of different serotypes were isolated, which possesses similar morphology and biochemical characters. Although belonging to the same serotype strains they always show significant difference in acid-producing and

  17. The Prospect of Immunoglobulin Y for Therapy of Canine parvovirus Infection in Dogs

    Directory of Open Access Journals (Sweden)

    I Gusti Ayu Agung Suartini

    2015-06-01

    Full Text Available Canine parvovirus (CPV is a highly infectious virus. The virus causes death in dogs worldwide. The mortality rate due to infection of CPV in dog reaches 91%. Prevention of CPV infection in puppies has been done by vaccination which is effectively proven. Protective mechanisms of maternal antibodies contribute to the failure of vaccination. Highly stable characteristics of parvovirus enable the virus still exist in the environment. Various therapies are performed only to suppress the clinical symptoms but can not reduce puppy mortalities. This review discusses CPV alternative therapy and the advantages using immunoglobulin Y (IgY specific antibodies isolated from chicken egg yolk. Immunoglobulin Y will neutralize the virus, so it can not infect host cells. Intravenous IgY therapy has shown to suppress the spread of CPV infection and prevent death.

  18. A novel colloidal gold labeled antigen for the detection of Deoxynivalenol using an immunochromatographic assay method

    Science.gov (United States)

    Jin, Yu; Liu, Renrong; Zhu, Lixin; Chen, Zhenzhen

    2017-11-01

    In this paper, an immunochromatographic assay card was developed for the detection of DON in feed and cereals using a novel colloidal gold labeling method. For the colloidal gold immunochromatographic rapid detection (GICD) card, a monoclonal antibody DON-mAb and a goat anti-chicken IgY were drawn on NC membrane as the test line (T line) and the control line (C line) respectively. A gold labeled DON-CBSA conjugate and a gold labeled chicken IgY were sprayed onto the conjugate pad. The GICD card has cut-off levels of 50ng/mL for DON, which is invulnerable to matrix interference, and applicable to a wide range of samples. The GICD detecting results of feed and grain samples were compared with the results of ELISA testing, which showed good consistency.

  19. Changes of serum NO/iNOS after whole abdominal irradiation in rats

    International Nuclear Information System (INIS)

    Lin Hong; Wu Wei; Zhang Kunhe; Chen Jiang; Wang Yang; Luo Hui; Huang Xiaoqing

    2004-01-01

    Objective: To observe the changes of serum NO/NOS concentrations in rats after whole abdominal irradiation and their relationships with intestinal radiation injury. Methods: Thirty health rats were randomly divided into three groups: normal control group, irradiation control group, and IgY protection group. The sera of rats were collected and the NO and iNOS were assayed. Results: NO and iNOS levels increased evidently in the irradiation control group in contrast with those of the normal control group (P<0.01). NO and iNOS levels increased slightly in the IgY protection group in contrast with the irradiation control group (P<0.01) and were close to the levels of the normal control group. The changes of NO and NOS concentrations were synchronous. Conclusion: NO and iNOS are good indices for detecting intestinal radiation injury. (authors)

  20. Semimechanistic model describing gastric emptying and glucose absorption in healthy subjects and patients with type 2 diabetes

    DEFF Research Database (Denmark)

    Alskär, Oskar; Bagger, Jonatan I; Røge, Rikke M.

    2016-01-01

    The integrated glucose-insulin (IGI) model is a previously published semimechanistic model that describes plasma glucose and insulin concentrations after glucose challenges. The aim of this work was to use knowledge of physiology to improve the IGI model's description of glucose absorption and ga...... model provides a better description and improves the understanding of dynamic glucose tests involving oral glucose....... and gastric emptying after tests with varying glucose doses. The developed model's performance was compared to empirical models. To develop our model, data from oral and intravenous glucose challenges in patients with type 2 diabetes and healthy control subjects were used together with present knowledge...... glucose absorption was superior to linear absorption regardless of the gastric emptying model applied. The semiphysiological model developed performed better than previously published empirical models and allows better understanding of the mechanisms underlying glucose absorption. In conclusion, our new...

  1. The development of the Australian Space Forecast Centre (ASFC)

    Science.gov (United States)

    Wilkinson, Phil; Kennewell, John A.; Cole, David

    2018-05-01

    The Ionospheric Prediction Service (IPS) was formed in 1947 to provide monthly prediction services for high frequency (HF) radio, in particular to support HF communications with the United Kingdom. It was quickly recognized that to be effective such a service also had to provide advice when ionospheric storms prevented HF communications from taking place. With the advent of the International Geophysical Year (IGY), short-term forecasts were also required for research programmes and the task of supplying the Australian input to these was given to Frank Cook, of the IPS, while Jack Turner, also of the IPS, supervised the generation of ionospheric maps to support high latitude HF communications. These two important IGY activities formed the platform on which all future IPS services would be built. This paper reviews the development of the Australian Space Forecast Centre (ASFC), which arose from these early origins.

  2. Functional Imaging of HER2-Positive Metastatic Breast Cancer Using 64Cu-DOTA-Trastuzumab Positron Emission Tomography

    Science.gov (United States)

    Mortimer, Joanne E.; Bading, James R.; Colcher, David M.; Conti, Peter S.; Frankel, Paul H.; Carroll, Mary I.; Tong, Shan; Poku, Erasmus; Miles, Joshua K.; Shively, John E.; Raubitschek, Andrew A.

    2014-01-01

    Women with human epidermal growth factor receptor 2 (HER2)-positive breast cancer are candidates for treatment with the anti-HER2 antibody trastuzumab. Assessment of HER2 status in recurrent disease is usually made by core needle biopsy of a single lesion which may not be representative of the larger tumor mass or other sites of disease. Our long-range goal is to develop positron emission tomography (PET) of radiolabeled trastuzumab for systemically assessing tumor HER2 expression and identifying appropriate use of anti-HER2 therapies. The purpose of this study was to evaluate PET-CT of 64Cu-DOTA-trastuzumab for detecting and measuring tumor uptake of trastuzumab in patients with HER2-positive metastatic breast cancer. Methods Eight women with biopsy-confirmed HER2-positive metastatic breast cancer and no anti-HER2 therapy for ≥ 4 mo underwent complete staging, including 18F-fluorodeoxyglucose (FDG)/PET-CT. For 6 of the 8 patients, 64Cu-DOTA-trastuzumab injection (364-512 MBq, 5 mg trastuzumab) was preceded by trastuzumab infusion (45 mg). PET-CT (PET scan duration 1 h) was performed 21-25 (“Day 1”) and 47-49 (“Day 2”) h after 64Cu-DOTA-trastuzumab injection. Scan fields of view were chosen based on 18F-FDG/PET-CT. Lesions visualized relative to adjacent tissue on PET were considered PET-positive; analysis was limited to lesions identifiable on CT. Radiolabel uptake in prominent lesions was measured as maximum single-voxel standardized uptake value (SUVmax). Results Liver uptake of 64Cu was reduced approximately 75% with the 45 mg trastuzumab pre-dose, without significant effect on tumor uptake. The study included 89 CT-positive lesions; detection sensitivity was 77, 89 and 93% for Day 1, Day 2 and 18F-FDG, respectively. On average, tumor uptake was similar for 64Cu-DOTA-trastuzumab and 18F-FDG [SUVmax (mean, range): Day 1 (8.1, 3.0-22.5, n=48); Day 2 (8.9, 0.9-28.9, n=38); 18F-FDG (9.7, 3.3-25.4, n=56)], but the extent of same-lesion uptake was not

  3. Functional imaging of human epidermal growth factor receptor 2-positive metastatic breast cancer using (64)Cu-DOTA-trastuzumab PET.

    Science.gov (United States)

    Mortimer, Joanne E; Bading, James R; Colcher, David M; Conti, Peter S; Frankel, Paul H; Carroll, Mary I; Tong, Shan; Poku, Erasmus; Miles, Joshua K; Shively, John E; Raubitschek, Andrew A

    2014-01-01

    Women with human epidermal growth factor receptor 2 (HER2)-positive breast cancer are candidates for treatment with the anti-HER2 antibody trastuzumab. Assessment of HER2 status in recurrent disease is usually made by core needle biopsy of a single lesion, which may not represent the larger tumor mass or other sites of disease. Our long-range goal is to develop PET of radiolabeled trastuzumab for systemically assessing tumor HER2 expression and identifying appropriate use of anti-HER2 therapies. The purpose of this study was to evaluate PET/CT of (64)Cu-DOTA-trastuzumab for detecting and measuring tumor uptake of trastuzumab in patients with HER2-positive metastatic breast cancer. Eight women with biopsy-confirmed HER2-positive metastatic breast cancer and no anti-HER2 therapy for 4 mo or longer underwent complete staging, including (18)F-FDG PET/CT. For 6 of the 8 patients, (64)Cu-DOTA-trastuzumab injection (364-512 MBq, 5 mg of trastuzumab) was preceded by trastuzumab infusion (45 mg). PET/CT (PET scan duration 1 h) was performed 21-25 (day 1) and 47-49 (day 2) h after (64)Cu-DOTA-trastuzumab injection. Scan fields of view were chosen on the basis of (18)F-FDG PET/CT. Tumor detection sensitivity and uptake analyses were limited to lesions identifiable on CT; lesions visualized relative to adjacent tissue on PET were considered PET-positive. Radiolabel uptake in prominent lesions was measured as maximum single-voxel standardized uptake value (SUVmax). Liver uptake of (64)Cu was reduced approximately 75% with the 45-mg trastuzumab predose, without significant effect on tumor uptake. The study included 89 CT-positive lesions. Detection sensitivity was 77%, 89%, and 93% for day 1, day 2, and (18)F-FDG, respectively. On average, tumor uptake was similar for (64)Cu-DOTA-trastuzumab and (18)F-FDG (SUVmax and range, 8.1 and 3.0-22.5 for day 1 [n = 48]; 8.9 and 0.9-28.9 for day 2 [n = 38]; 9.7 and 3.3-25.4 for (18)F-FDG [n = 56]), but same-lesion SUVmax was not correlated

  4. Outer membrane vesicles of Gallibacterium anatis induce protective immunity in egg-laying hens.

    Science.gov (United States)

    Pors, Susanne E; Pedersen, Ida J; Skjerning, Ragnhild Bager; Thøfner, Ida C N; Persson, Gry; Bojesen, Anders M

    2016-11-15

    Gallibacterium anatis causes infections in the reproductive tract of egg-laying hens and induce increased mortality and decreased egg production. New prophylactic measures are needed in order to improve animal welfare and production efficiency. Bacterial outer membrane vesicles (OMVs) have previously shown promising results in protection against infections and we hypothesized that OMVs could serve as an immunogen to protect egg-laying hens against G. anatis. To investigate the immunogenic potential of G. anatis OMVs, two in vivo studies in egg-laying hens were made. The trials assessedthe degree of protection provided by immunization with G. anatis OMV against challenge and the IgY responses in serum after immunization and challenge, respectively. A total of 64 egg-laying hens were included in the trials. OMVs for immunization were produced and purified from a high-producing G. anatis ΔtolR mutant. Challenge was done with G. anatis 12656-12 and evaluated by scoring lesions and bacterial re-isolation rates from peritoneum. Finally, levels of OMV-specific IgY in sera were assayed by ELISA. Immunization with OMVs decreased the lesions scores significantly, while the bacterial re-isolation remained unchanged. Furthermore, a high OMV-specific IgY response was induced by immunization and subsequent challenge of the hens. The results strongly indicate that immunization with G. anatis OMVs provides significant protection against G. anatis challenge and induces specific antibody responses with high titers of OMV-specific IgY in serum. The results therefore show great promise for OMV based vaccines aiming at providing protecting against G. anatis in egg-laying hens. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Space Weather and Real-Time Monitoring

    Directory of Open Access Journals (Sweden)

    S Watari

    2009-04-01

    Full Text Available Recent advance of information and communications technology enables to collect a large amount of ground-based and space-based observation data in real-time. The real-time data realize nowcast of space weather. This paper reports a history of space weather by the International Space Environment Service (ISES in association with the International Geophysical Year (IGY and importance of real-time monitoring in space weather.

  6. Trace metals, melanin-based pigmentation and their interaction influence immune parameters in feral pigeons (Columba livia).

    Science.gov (United States)

    Chatelain, M; Gasparini, J; Frantz, A

    2016-04-01

    Understanding the effects of trace metals emitted by anthropogenic activities on wildlife is of great concern in urban ecology; yet, information on how they affect individuals, populations, communities and ecosystems remains scarce. In particular, trace metals may impact survival by altering the immune system response to parasites. Plumage melanin is assumed to influence the effects of trace metals on immunity owing to its ability to bind metal ions in feathers and its synthesis being coded by a pleiotropic gene. We thus hypothesized that trace metal exposure would interact with plumage colouration in shaping immune response. We experimentally investigated the interactive effect between exposure to an environmentally relevant range of zinc and/or lead and melanin-based plumage colouration on components of the immune system in feral pigeons (Columba livia). We found that zinc increased anti-keyhole limpet hemocyanin (KLH) IgY primary response maintenance, buffered the negative effect of lead on anti-KLH IgY secondary response maintenance and tended to increase T-cell mediated phytohaemagglutinin (PHA) skin response. Lead decreased the peak of the anti-KLH IgY secondary response. In addition, pheomelanic pigeons exhibited a higher secondary anti-KLH IgY response than did eumelanic ones. Finally, T-cell mediated PHA skin response decreased with increasing plumage eumelanin level of birds exposed to lead. Neither treatments nor plumage colouration correlated with endoparasite intensity. Overall, our study points out the effects of trace metals on some parameters of birds' immunity, independently from other confounding urbanization factors, and underlines the need to investigate their impacts on other life history traits and their consequences in the ecology and evolution of host-parasite interactions.

  7. 1,5-anhydroglucitol is associated with early-phase insulin secretion in chinese patients with newly diagnosed type 2 diabetes mellitus.

    Science.gov (United States)

    Ma, Xiaojing; Hao, Yaping; Hu, Xiang; Luo, Yuqi; Deng, Zixuan; Zhou, Jian; Bao, Yuqian; Jia, Weiping

    2015-05-01

    The goal of the present study was to explore the correlations of 1,5-anhydroglucitol (l,5-AG), glycated hemoglobin (HbA1c), and glycated albumin (GA) with insulin sensitivity and secretion. In total, 302 patients with newly diagnosed type 2 diabetes mellitus (166 men, 136 women) were enrolled in this study. The homeostasis model assessment for insulin resistance (HOMA-IR) and homeostasis model assessment for β-cell function (HOMA-β) were calculated to determine the basal insulin sensitivity and secretion. The insulinogenic index (IGI) was used to evaluate early-phase insulin secretion. 1,5-AG and GA were assayed via the enzymatic method, and HbA1c was detected by high-pressure liquid chromatography. Among all 302 subjects, the serum 1,5-AG level was 13.1±7.2 μg/mL, and the HbA1c and GA levels [median (interquartile range)] were 6.7% (6.2-7.3%) and 17.7% (16.0-19.5%), respectively. Increased 1,5-AG quartiles were accompanied by trends toward a decreased HOMA-IR and an increased HOMA-β and IGI (for all trends, P1). 1,5-AG was negatively associated with HOMA-IR (r=-0.200, P1) and positively associated with HOMA-β and IGI (r=0.210 and 0.413, respectively; both P1). 1,5-AG was independently related to HOMA-IR and HOMA-β and exhibited an independent positive association with IGI (standardized β=0.242, P1). Additionally, both HbA1c and GA were independently correlated with HOMA-IR and HOMA-β. 1,5-AG is not only correlated with basal insulin sensitivity and secretion, but also closely associated with early-phase insulin secretion in Chinese patients with newly diagnosed type 2 diabetes mellitus.

  8. First Results of Minimum Fisher Regularisation as Unfolding Method for JET NE213 Liquid Scintillator Neutron Spectrometry

    Czech Academy of Sciences Publication Activity Database

    Mlynář, Jan; Adams, J. M.; Bertalot, L.; Conroy, S.

    2005-01-01

    Roč. 74, 1-4 (2005), s. 781-786 ISSN 0920-3796. [Symposium on Fusion Technology - SOFT/23rd./. Benátky, 20.9.2004-24.9.2004] Institutional research plan: CEZ:AV0Z20430508 Keywords : Tokamak * fusion * neutron diagnostic * spectrum unfolding * scintillator regularisation Subject RIV: BL - Plasma and Gas Discharge Physics Impact factor: 0.981, year: 2005 http://soft2004.igi.cnr.it/

  9. A High Level Model of a Conscious Embodied Agent

    Czech Academy of Sciences Publication Activity Database

    Wiedermann, Jiří

    2010-01-01

    Roč. 2, č. 3 (2010), s. 62-78 ISSN 1942-9045 R&D Projects: GA ČR GAP202/10/1333 Institutional research plan: CEZ:AV0Z10300504 Keywords : embodied agent * internal world models * higher cognitive function Subject RIV: IN - Informatics, Computer Science http://www.igi-global.com/article/high-level-model-conscious-embodied/46147

  10. Central plane of the ring current responsible for geomagnetic disturbance in the South-American region

    International Nuclear Information System (INIS)

    Kane, R.P.; Trivedi, N.B.

    1981-01-01

    Using hourly values of H, D, Z from a network of South American stations, operative during the IGY-IGC, the latitude dependence of storm effects was studied. It was found that whereas there were considerable distortions due to conductivity anomalies under the Andes, there was also evidence of latitudinal excursions of overhead current system, not only from storm to storm but even during the course of the same storm

  11. Sulphur mountain: Cosmic ray intensity records

    International Nuclear Information System (INIS)

    Venkatesan, D.; Mathews, T.

    1985-01-01

    This book deals with the comic ray intensity registrations at the Sulphur Mountain Cosmic Ray Laboratory. The time series of intensity form a valuable data-set, for studying cosmic ray intensity variations and their dependence on solar activity. The IGY neutron monitor started operating from July 1, 1957 and continued through 1963. Daily mean values are tabulated for the period and these are also represented in plots. This monitor was set up by the National Research Council of Canada

  12. Vähesest piisab? / Harri Slip

    Index Scriptorium Estoniae

    Slip, Harri

    2011-01-01

    Nuppkuularid hinnaga 70-105 eurot: AKG K370; Beyerdynamic DTX 101iE; Bose IE2; Creative Aurvana In-Ear 2; Denon AH-C560R; Grado iGi; Jamo wEAR In30; Klipsch S4; Philips SHE9750/10; Sennheiser CX 400-II; Shure SE 215; Skullcandy Heavy Metal; Sony MDR-EX510LP Ultimate Ears 600.Ultimate Ears 600

  13. CORAL SNAKE ANTIVENOM PRODUCED IN CHICKENS (Gallus domesticus

    Directory of Open Access Journals (Sweden)

    Irma Aguilar

    2014-01-01

    Full Text Available The production of anti-snake venom from large mammal's blood has been found to be low-yielding and arduous, consequently, antivenom immunoglobulins for treatment are achieved regularly as polyvalent serum. We have standardized an undemanding technique for making purified immunoglobulin IgY antivenom consisting of polyclonal antibodies against coral snake venom in the egg yolk of immunized hens. We have adapted a reported process of antibody purification from egg yolks, and achieved 90% antibody purity. The customized technique consisted of the removal of lipids from distilled water-diluted egg yolks by a freeze–thaw sequence. The specific immunoglobulins were present in the egg yolk for up to 180 days postimmunization. Therefore, by means of small venom quantities, a significant amount of immunoglobulins were found in an adequately purified state (The obtained material contained about 90% pure IgY. The antigen binding of the immunoglobulins was detected by a double immunodiffusion test. Titers of antibodies in the yolk were estimated with a serum protection assay (Median effective dose = ED50 (ED50= 477 mg/kg. Given that breeding hens is economically feasible, egg gathering is noninvasive and the purification of IgY antibodies is quick and easy, chicken immunization is an excellent alternative for the production of polyclonal antibodies. To the best of our knowledge, this is the first coral snake antivenom prepared in birds.

  14. Protective efficacy of immunoglobulins Y prepared against Cerastes cerastes snake venom in the Kingdom of Saudi Arabia.

    Science.gov (United States)

    Moussa, Ihab M; Hessan, Ashgan M; Aleisa, Abdulaziz M; Al-Arfaj, Abdullah A; Salem-Bekhit, Mounier M; AlRejai, Salim A

    2012-08-01

    To prepare and evaluate the protective efficacy of immunoglobulin Y (IgY) prepared against local Saudi Cerastes cerastes snake venom. The study was conducted between October 2009 and October 2011 at the Center of Excellence in Biotechnology Research, King Saud University, Riyadh, Kingdom of Saudi Arabia. The study designed as follow; 4 groups of 8 chickens were immunized intramuscularly with Cerastes cerastes snake venoms mixed with Freund's complete adjuvant. Three weeks later, the injections were repeated with the venoms with incomplete Freund's adjuvant. Three boosters were given with the venoms at 3 weeks intervals. The IgY was extracted by ammonium sulphate-caprylic acid method, the antibody titer were tested by enzyme linked immunosorbant assay, and the protective efficacies of the extracted immunoglobulins were performed. Immunoglobulin Y preparation extracted by ammonium sulphate-caprylic acid method showed lack of low molecular weight bands. The bands representing IgY-antibodies, which have molecular weights ranged from 180-200 KD, appeared sharp and clear. Furthermore, evaluation of the prepared protective value of IgY-antibodies revealed one ml of extracted IgY-antibodies containing 15 mg/ml anti Cerastes cerastes; specific IgY could produce 100% protection against 50 LD50. Laying hens could be used as an alternative source of polyclonal antibodies against Cerastes cerastes snake venoms due to several advantages as compared with mammals.

  15. Generation and Characterization of Polyclonal Antibody Against Part of Immunoglobulin Constant Heavy υ Chain of Goose

    Science.gov (United States)

    Zhao, Panpan; Guo, Yongli; Ma, Bo; Wang, Junwei

    2014-01-01

    Immunoglobulin Y (abbreviated as IgY) is a type of immunoglobulin that is the major antibody in bird, reptile, and lungfish blood. IgY consists of two light (λ) and two heavy (υ) chains. In the present study, polyclonal antibody against IgYFc was generated and evaluated. rIgYCυ3/Cυ4 was expressed in Escherichia coli, purified and utilized to raise polyclonal antibody in rabbit. High affinity antisera were obtained, which successfully detected the antigen at a dilution of 1:204,800 for ELISA assay. The antibody can specifically recognize both rIgYCυ3/Cυ4 and native IgY by Western bolt analysis. Furthermore, the serum of Grus japonensis or immunoglobulin of chicken, duck, turkey, and silkie samples and dynamic changes of serum GoIgY after immunogenicity with GPV-VP3-virus-like particles (GPV-VP3-VLPs) can be detected with the anti-GoIgYFc polyclonal antibody. These results suggested that the antibody is valuable for the investigation of biochemical properties and biological functions of GoIgY. PMID:25171010

  16. Multimodal system for the planning and guidance of bronchoscopy

    Science.gov (United States)

    Higgins, William E.; Cheirsilp, Ronnarit; Zang, Xiaonan; Byrnes, Patrick

    2015-03-01

    Many technical innovations in multimodal radiologic imaging and bronchoscopy have emerged recently in the effort against lung cancer. Modern X-ray computed-tomography (CT) scanners provide three-dimensional (3D) high-resolution chest images, positron emission tomography (PET) scanners give complementary molecular imaging data, and new integrated PET/CT scanners combine the strengths of both modalities. State-of-the-art bronchoscopes permit minimally invasive tissue sampling, with vivid endobronchial video enabling navigation deep into the airway-tree periphery, while complementary endobronchial ultrasound (EBUS) reveals local views of anatomical structures outside the airways. In addition, image-guided intervention (IGI) systems have proven their utility for CT-based planning and guidance of bronchoscopy. Unfortunately, no IGI system exists that integrates all sources effectively through the complete lung-cancer staging work flow. This paper presents a prototype of a computer-based multimodal IGI system that strives to fill this need. The system combines a wide range of automatic and semi-automatic image-processing tools for multimodal data fusion and procedure planning. It also provides a flexible graphical user interface for follow-on guidance of bronchoscopy/EBUS. Human-study results demonstrate the system's potential.

  17. Oral and parenteral immunization of chickens (Gallus gallus) against West Nile virus with recombinant envelope protein

    Science.gov (United States)

    Fassbinder-Orth, C. A.; Hofmeister, Erik K.; Weeks-Levy, C.; Karasov, W.H.

    2009-01-01

    West Nile virus (WNV) causes morbidity and mortality in humans, horses, and in more than 315 bird species in North America. Currently approved WNV vaccines are designed for parenteral administration and, as yet, no effective oral WNV vaccines have been developed. WNV envelope (E) protein is a highly antigenic protein that elicits the majority of virus-neutralizing antibodies during a WNV immune response. Leghorn chickens were given three vaccinations (each 2 wk apart) of E protein orally (20 ??g or 100 ??g/dose), of E protein intramuscularly (IM, 20 ??g/dose), or of adjuvant only (control group) followed by a WNV challenge. Viremias were measured post-WNV infection, and three new enzyme-linked immunosorbent assays were developed for quantifying IgM, IgY, and IgA-mediated immune response of birds following WNV infection. WNV viremia levels were significantly lower in the IM group than in both oral groups and the control group. Total WNV E protein-specific IgY production was significantly greater, and WNV nonstructural 1-specific IgY was significantly less, in the IM group compared to all other treatment groups. The results of this study indicate that IM vaccination of chickens with E protein is protective against WNV infection and results in a significantly different antibody production profile as compared to both orally vaccinated and nonvaccinated birds. ?? 2009 American Association of Avian Pathologists.

  18. Aerosol single-scattering albedo and asymmetry parameter from MFRSR observations during the ARM Aerosol IOP 2003

    Directory of Open Access Journals (Sweden)

    E. I. Kassianov

    2007-06-01

    Full Text Available Multi-filter Rotating Shadowband Radiometers (MFRSRs provide routine measurements of the aerosol optical depth (τ at six wavelengths (0.415, 0.5, 0.615, 0.673, 0.870 and 0.94 μm. The single-scattering albedo (π0 is typically estimated from the MFRSR measurements by assuming the asymmetry parameter (<I>g>. In most instances, however, it is not easy to set an appropriate value of <I>g> due to its strong temporal and spatial variability. Here, we introduce and validate an updated version of our retrieval technique that allows one to estimate simultaneously π0 and <I>g> for different types of aerosol. We use the aerosol and radiative properties obtained during the Atmospheric Radiation Measurement (ARM Program's Aerosol Intensive Operational Period (IOP to validate our retrieval in two ways. First, the MFRSR-retrieved optical properties are compared with those obtained from independent surface, Aerosol Robotic Network (AERONET, and aircraft measurements. The MFRSR-retrieved optical properties are in reasonable agreement with these independent measurements. Second, we perform radiative closure experiments using the MFRSR-retrieved optical properties. The calculated broadband values of the direct and diffuse fluxes are comparable (~5 W/m2 to those obtained from measurements.

  19. Discovery of iron group impurity ion spin states in single crystal Y{sub 2}SiO{sub 5} with strong coupling to whispering gallery photons

    Energy Technology Data Exchange (ETDEWEB)

    Goryachev, Maxim; Farr, Warrick G.; Carmo Carvalho, Natalia do; Creedon, Daniel L.; Le Floch, Jean-Michel [ARC Centre of Excellence for Engineered Quantum Systems, University of Western Australia, 35 Stirling Highway, Crawley WA 6009 (Australia); Probst, Sebastian [Physikalisches Institut, Karlsruhe Institute of Technology, D-76128 Karlsruhe (Germany); Bushev, Pavel [Experimentalphysik, Universität des Saarlandes, D-66123 Saarbrücken (Germany); Tobar, Michael E., E-mail: michael.tobar@uwa.edu.au [ARC Centre of Excellence for Engineered Quantum Systems, School of Physics, University of Western Australia, Crawley 6009 (Australia)

    2015-06-08

    Interaction of Whispering Gallery Modes (WGMs) with dilute spin ensembles in solids is an interesting paradigm of Hybrid Quantum Systems potentially beneficial for Quantum Signal Processing applications. Unexpected ion transitions are measured in single crystal Y{sub 2}SiO{sub 5} using WGM spectroscopy with large Zero Field Splittings at 14.7 GHz, 18.4 GHz, and 25.4 GHz, which also feature considerable anisotropy of the g-tensors as well as two inequivalent lattice sites, indicating spins from Iron Group Ion (IGI) impurities. The comparison of undoped and Rare-Earth doped crystals reveal that the IGIs are introduced during co-doping of Eu{sup 3+} or Er{sup 3+} with concentration at much lower levels of order 100 ppb. The strong coupling regime between an ensemble of IGI spins and WGM photons have been demonstrated at 18.4 GHz and near zero field. This approach together with useful optical properties of these ions opens avenues for “spins-in-solids” Quantum Electrodynamics.

  20. Phase I Study of 68Ga-HER2-Nanobody for PET/CT Assessment of HER2 Expression in Breast Carcinoma.

    Science.gov (United States)

    Keyaerts, Marleen; Xavier, Catarina; Heemskerk, Johannes; Devoogdt, Nick; Everaert, Hendrik; Ackaert, Chloé; Vanhoeij, Marian; Duhoux, Francois P; Gevaert, Thierry; Simon, Philippe; Schallier, Denis; Fontaine, Christel; Vaneycken, Ilse; Vanhove, Christian; De Greve, Jacques; Lamote, Jan; Caveliers, Vicky; Lahoutte, Tony

    2016-01-01

    Human epidermal growth factor receptor 2 (HER2) status is one of the major tumor characteristics in breast cancer to guide therapy. Anti-HER2 treatment has clear survival advantages in HER2-positive breast carcinoma patients. Heterogeneity in HER2 expression between primary tumor and metastasis has repeatedly been described, resulting in the need to reassess HER2 status during the disease course. To avoid repeated biopsy with potential bias due to tumor heterogeneity, Nanobodies directed against HER2 have been developed as probes for molecular imaging. Nanobodies, which are derived from unique heavy-chain-only antibodies, are the smallest antigen-binding antibody fragments and have ideal characteristics for PET imaging. The primary aims were assessment of safety, biodistribution, and dosimetry. The secondary aim was to investigate tumor-targeting potential. In total, 20 women with primary or metastatic breast carcinoma (score of 2+ or 3+ on HER2 immunohistochemical assessment) were included. Anti-HER2-Nanobody was labeled with (68)Ga via a NOTA derivative. Administered activities were 53-174 MBq (average, 107 MBq). PET/CT scans for dosimetry assessment were obtained at 10, 60, and 90 min after administration. Physical evaluation and blood analysis were performed for safety evaluation. Biodistribution was analyzed for 11 organs using MIM software; dosimetry was assessed using OLINDA/EXM. Tumor-targeting potential was assessed in primary and metastatic lesions. No adverse reactions occurred. A fast blood clearance was observed, with only 10% of injected activity remaining in the blood at 1 h after injection. Uptake was seen mainly in the kidneys, liver, and intestines. The effective dose was 0.043 mSv/MBq, resulting in an average of 4.6 mSv per patient. The critical organ was the urinary bladder wall, with a dose of 0.406 mGy/MBq. In patients with metastatic disease, tracer accumulation well above the background level was demonstrated in most identified sites of

  1. NSCLC and HER2: between lights and shadows.

    Science.gov (United States)

    Ricciardi, Giuseppina Rosaria Rita; Russo, Alessandro; Franchina, Tindara; Ferraro, Giuseppa; Zanghì, Mariangela; Picone, Antonio; Scimone, Antonino; Adamo, Vincenzo

    2014-12-01

    The therapeutic landscape of non-small-cell lung cancer (NSCLC) has dramatically changed in the last few years with the introduction of molecularly targeted agents, leading to unprecedented results in lung tumors with a paradigmatic shift from a "one size fits all" approach to an histologic and molecular-based approach. The discovery of epidermal growth factor receptor (EGFR) mutations in NSCLC in 2004 and the marked response to the EGFR tyrosine kinase inhibitor gefitinib, in a small subset of patients harboring these genetic abnormalities, stimulated the study of other kinase mutants involvement in NSCLC. The incredible story of ALK rearranged tumors, with the rapid Food and Drug Administration approval of Crizotinib after only 4 years from the discovery of EML4-ALK translocation in NSCLC, has profoundly influenced the concept of drug development in NSCLC, paving the way to a novel series of molecularly selected studies with specific inhibitors. The identification of these oncogenic drivers has dramatically changed the genetic landscape of NSCLC moving away from the old concept of a large indistinct histological entity to a combination of rare clinically relevant molecular subsets. Recently, a renewed interest has been emerging on the human epidermal growth factor-2 (HER2) pathway. Genetic aberrations of this signaling pathway have been reported over time to be associated in NSCLC with different sensitivity to the EGFR tyrosine kinase inhibitors, to have a possible prognostic role and more recently HER2 amplification has been emerged as a possible mechanism in EGFR-mutated tumors of acquired resistance to the EGFR tyrosine kinase inhibitors. In addition, dysregulation of the HER2 pathway, in particular HER2 mutations (mostly, in-frame exon 20 insertions), may represent a possible novel therapeutic target in NSCLC, paving the way for a new generation of targeted agents in NSCLC. Since anecdotal case reports of clinical activity of anti-HER2 agents in NSCLC

  2. EGFR gene amplification is relatively common and associates with outcome in intestinal adenocarcinoma of the stomach, gastro-oesophageal junction and distal oesophagus

    International Nuclear Information System (INIS)

    Birkman, Eva-Maria; Ålgars, Annika; Lintunen, Minnamaija; Ristamäki, Raija; Sundström, Jari; Carpén, Olli

    2016-01-01

    Approximately 50 % of gastric adenocarcinomas belong to a molecular subgroup characterised by chromosomal instability and a strong association with the intestinal histological subtype. This subgroup typically contains alterations in the receptor tyrosine kinase–RAS pathway, for example EGFR or HER2 gene amplifications leading to protein overexpression. In clinical practice, HER2 overexpressing metastatic gastric cancer is known to respond to treatment with anti-HER2 antibodies. By contrast, anti-EGFR antibodies have not been able to provide survival benefit in clinical trials, which, however, have not included patient selection based on the histological subtype or EGFR gene copy number analysis of the tumours. To examine the role of EGFR as a potential biomarker, we studied the prevalence, clinicopathological associations as well as prognostic role of EGFR and HER2 expression and gene amplification in intestinal adenocarcinomas of the stomach, gastro-oesophageal junction and distal oesophagus. Tissue samples from 220 patients were analysed with EGFR and HER2 immunohistochemistry. Those samples with moderate/strong staining intensity were further analysed with silver in situ hybridization to quantify gene copy numbers. The results were associated with clinical patient characteristics and survival. Moderate/strong EGFR protein expression was found in 72/220 (32.7 %) and EGFR gene amplification in 31/220 (14.1 %) of the tumours, while moderate/strong HER2 protein expression was detected in 31/220 (14.1 %) and HER2 gene amplification in 29/220 (13.2 %) of the tumours. EGFR and HER2 genes were co-amplified in eight tumours (3.6 %). EGFR gene amplification was more common in tumours of distal oesophagus/gastro-oesophageal junction/cardia than in those of gastric corpus (p = 0.013). It was associated with shortened time to cancer recurrence (p = 0.026) and cancer specific survival (p = 0.033). EGFR gene amplification is relatively common in intestinal adenocarcinomas

  3. The Effect of Histone Hyperacetylation on Viability of Basal-Like Breast Cancer Cells MDA-MB-231

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    Aliasghar Rahimian

    2017-06-01

    Full Text Available Background The Basal-Like breast cancer, is always known for lack of expression of estrogen receptor (ER, progesterone receptor (PR and as well, absence of epidermal growth factor receptor 2 (HER2 gene amplification. Improper expression pattern of ER, PR, and Her2, makes Basal-Like breast tumors resistant to the current hormonal and anti HER2 treatments. In recent decades, several studies have been conducted to investigate the regulatory role of chemical modifications of core histones in gene expression. Their results have shown that histone acetylation is involved in regulation of cell survival. Acetylation of core histones is regulated by the epigenetic-modifying enzymes named Histone Deacetylases (HDACs. As a new approach to control the viability of breast tumor cells resistant to the hormonal and anti-HER2 treatments, we have targeted the HDACs. Using Trichostatin A (TSA as a known HDACs inhibitor, we have tried to hyperacetylate the core histones of MDA-MB-231 cells as an in vitro model of Basal-Like breast tumors. Then we have investigated the effect of histone hyperacetylation on viability of MDA-MB-231 cells. Methods MDA-MB-231 cells were cultured in RPMI 1640 medium containing 10% fetal bovine serum (FBS and were incubated at 37°C, in a humidified incubator with 5% CO2 atmosphere. Then cells were treated with different concentrations of TSA including: 50, 100, 200, 400, 800 and 1000 nM or control (1% DMSO. After 24 and 48 hours, viability of cells was evaluated by MTT assay. Results After 24 and 48h exposure to different concentrations of TSA, MDA-MB-231 cells showed a maximum tolerable dose. At higher concentrations, TSA decreased the percentage of cell viability through a time-dose dependent manner. IC50 value for 48h treatment was 600 nM. Conclusions Our results indicate that HDACs inhibition and subsequently hyperacetylation of histones, leads to cytotoxic effects on breast tumor cells resistant to the current treatments. Following

  4. Cardiotoxicity of novel HER2-targeted therapies.

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    Sendur, Mehmet A N; Aksoy, Sercan; Altundag, Kadri

    2013-08-01

    Trastuzumab, an anti-HER2 humanized monoclonal antibody, is the standard treatment for both early and metastatic HER2-positive breast cancer. In addition to other chemotherapeutic agents, trastuzumab significantly improves response rate and survival in HER2-positive early and metastatic breast cancer. Although it is well known that trastuzumab therapy is closely associated with both symptomatic and asymptomatic cardiotoxicity, less is known about novel HER2-targeted therapies. The aim of this review is to discuss the cardiac safety data from recent studies of novel anti-HER2 drugs other than trastuzumab. Novel HER2-targeted therapies showed favorable results in HER2 positive metastatic breast cancer patients. Pubmed database, ASCO and San Antonio Breast Cancer Symposium Meeting abstracts were searched until January 2013 using the following search keywords; 'trastuzumab, trastuzumab cardiotoxicity, HER-2 targeted therapies, lapatinib, pertuzumab, trastuzumab emtansine, afatinib and neratinib'; papers which were considered relevant for the aim of this review were selected by the authors. Lapatinib, pertuzumab, T-DM1, neratinib and afatinib molecules are evaluated in the study. In a comprehensive analysis, 3689 lapatinib treated patients enrolled in 49 trials; asymptomatic cardiac events were reported in 53 patients (1.4%) and symptomatic grade III and IV systolic dysfunction was observed only in 7 patients (0.2%) treated with lapatinib. In phase I-III trials of pertuzumab, cardiac dysfunction was seen in 4.5-14.5% of patients with pertuzumab treatment and cardiac dysfunction was usually grade I and II. Cardiotoxicity of pertuzumab was usually reported with the trastuzumab combination and no additive cardiotoxicity was reported with addition of pertuzumab to trastuzumab. T-DM1 had a better safety profile compared to trastuzumab, no significant cardiotoxicity was observed with T-DM1 in heavily pre-treated patients. In the EMILIA study, only in 1.7% of patients in the T

  5. A Comprehensive Evaluation Of HER-2/NEU In Human Breast Cancer By Immunohistochemistry And Fluorescence In Situ Hybridization

    International Nuclear Information System (INIS)

    Abdel Fattah, H.I.; Seif, A.A.; El Hadidi, E.S.; Mohamed, A.A.; El Shinawi, M.E.; Shabaan, M.A.M.

    2012-01-01

    Background: Accurate diagnostic assessment of human epidermal growth factor receptor-2 (HER2/neu) is essential and a prerequisite for appropriate application of the humanized anti-HER-2 monoclonal antibody trastuzumab (Herceptin) to the treatment of patients with breast cancer. The food drug administration (FDA) approved immunohistochemistry (IHC) and Fluorescence in situ hybridization (FISH) for HER2/neu evaluation. IHC is the most widely applicable diagnostic modality in studying HER-2 status. FISH is used for HER2/neu assessment in cases with an equivocal IHC status (score 2+). Objectives: The aim of this study was to detect the amplification and/or expression of HER2/neu in breast cancer using FISH and IHC techniques and to evaluate these applied techniques for their potential and clinical application, with special consideration of equivocal cases by IHC (2+). Subjects and Methods: Assessment of HER2/neu gene expression was made by FISH analysis using the HER2/CEP dual color probe (Vysis) in paraffin-embedded tissue sections of 32 breast cancer patients who were grouped into stages 1+, 2+ or 3+ based on IHC (CBII monoclonal antibody), 4 were classified as IHC 0, 4 were classified as IHC 1 +, 22 were classified as the borderline group; IHC 2+, and 2 were classified as IHC 3+. ER, PR, CEA and CA15-3 were performed to all cases. Survival data was obtained from 25 patients only. Results: The cut-off suggested for HER2/neu amplification by FISH ratio was > 1.3. No statistical significance was found between HER2/neu -by either FISH or IHC- and the different prognostic parameters. The overall survival for the studied patients -in average 3 years- was 16/25 (64%). A significant statistical association was revealed between breast cancer patients’ survival outcome and HER2/neu amplification (p < 0.05) using chi square test. Conclusion: All breast cancer patients should be assessed for HER2/neu status. IHC is a well established method for assessing HER2/neu status in

  6. Trastuzumab-binding peptide display by Tobacco mosaic virus

    International Nuclear Information System (INIS)

    Frolova, Olga Y.; Petrunia, Igor V.; Komarova, Tatiana V.; Kosorukov, Vyacheslav S.; Sheval, Eugene V.; Gleba, Yuri Y.; Dorokhov, Yuri L.

    2010-01-01

    Human epidermal growth factor receptor-2 (HER2/neu) is a target for the humanized monoclonal antibody trastuzumab. Recently, trastuzumab-binding peptides (TBP) of HER2/neu that inhibit proliferation of breast cancer cells were identified. We have now studied conditions of efficient assembly in vivo of Tobacco mosaic virus (TMV)-based particles displaying TBP on its surface. The system is based on an Agrobacterium-mediated co-delivery of binary vectors encoding TMV RNA and coat protein (CP) with TBP in its C-terminal extension into plant leaves. We show how the fusion of amino acid substituted TBP (sTBP) to CP via a flexible peptide linker can improve the manufacturability of recombinant TMV (rTMV). We also reveal that rTMV particles with exposed sTBP retained trastuzumab-binding capacity but lost an anti-HER2/neu immunogenic scaffold function. Mouse antibodies against rTMV did not recognize HER2/neu on surface of human SK-BR-3 cells.

  7. A novel immuno-gold labeling protocol for nanobody-based detection of HER2 in breast cancer cells using immuno-electron microscopy.

    Science.gov (United States)

    Kijanka, M; van Donselaar, E G; Müller, W H; Dorresteijn, B; Popov-Čeleketić, D; El Khattabi, M; Verrips, C T; van Bergen En Henegouwen, P M P; Post, J A

    2017-07-01

    Immuno-electron microscopy is commonly performed with the use of antibodies. In the last decade the antibody fragment indicated as nanobody (VHH or single domain antibody) has found its way to different applications previously done with conventional antibodies. Nanobodies can be selected to bind with high affinity and specificity to different antigens. They are small (molecular weight ca. 15kDa) and are usually easy to produce in microorganisms. Here we have evaluated the feasibility of a nanobody binding to HER2 for application in immuno-electron microscopy. To obtain highest labeling efficiency combined with optimal specificity, different labeling conditions were analysed, which included nanobody concentration, fixation and blocking conditions. The obtained optimal protocol was applied for post-embedment labeling of Tokuyasu cryosections and for pre-embedment labeling of HER2 for fluorescence microscopy and both transmission and scanning electron microscopy. We show that formaldehyde fixation after incubation with the anti-HER2 nanobody, improves labeling intensity. Among all tested blocking agents the best results were obtained with a mixture of cold water fish gelatine and acetylated bovine serum albumin, which prevented a-specific interactions causing background labeling while preserving specific interactions at the same time. In conclusion, we have developed a nanobody-based protocol for immuno-gold labeling of HER2 for Tokuyasu cryosections in TEM as well as for pre-embedment gold labeling of cells for both TEM and SEM. Copyright © 2017. Published by Elsevier Inc.

  8. Synthesis, characterization and theranostic evaluation of Indium-111 labeled multifunctional superparamagnetic iron oxide nanoparticles

    International Nuclear Information System (INIS)

    Zolata, Hamidreza; Abbasi Davani, Fereydoun; Afarideh, Hossein

    2015-01-01

    Indium-111 labeled, Trastuzumab-Doxorubicin Conjugated, and APTES-PEG coated magnetic nanoparticles were designed for tumor targeting, drug delivery, controlled drug release, and dual-modal tumor imaging. Superparamagnetic iron oxide nanoparticles (SPIONs) were synthesized by thermal decomposition method to obtain narrow size particles. To increase SPIONs circulation time in blood and decrease its cytotoxicity in healthy tissues, SPIONs surface was modified with 3-Aminopropyltriethoxy Silane (APTES) and then were functionalized with N-Hydroxysuccinimide (NHS) ester of Polyethylene Glycol Maleimide (NHS-PEG-Mal) to conjugate with thiolated 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6, 9,-triacetic acid (PCTA) bifunctional chelator (BFC) and Trastuzumab antibody. In order to tumor SPECT/MR imaging, SPIONs were labeled with Indium-111 (T 1/2 = 2.80d). NHS ester of monoethyl malonate (MEM-NHS) was used for conjugation of Doxorubicin (DOX) chemotherapeutic agent onto SPIONs surface. Mono-Ethyl Malonate allows DOX molecules to be attached to SPIONs via pH-sensitive hydrazone bonds which lead to controlled drug release in tumor region. Active and passive tumor targeting were achieved through incorporated anti-HER2 (Trastuzumab) antibody and EPR effect of solid tumors for nanoparticles respectively. In addition to in vitro assessments of modified SPIONs in SKBR3 cell lines, their theranostic effects were evaluated in HER2 + breast tumor bearing BALB/c mice via biodistribution study, dual-modal molecular imaging and tumor diameter measurements

  9. Asymptomatic Changes in Cardiac Function Can Occur in DCIS Patients Following Treatment with HER-2/neu Pulsed Dendritic Cell Vaccines

    Science.gov (United States)

    Bahl, Susan; Roses, Robert; Sharma, Anupama; Koldovsky, Ursula; Xu, Shuwen; Weinstein, Susan; Nisenbaum, Harvey; Fox, Kevin; Pasha, Theresa; Zhang, Paul; Araujo, Louis; Carver, Joseph; Czerniecki, Brian J

    2009-01-01

    Background Targeting HER-2/neu with Trastuzumab has been associated with development of cardiac toxicity. Methods Twenty-seven patients with ductal carcinoma in situ (DCIS) of the breast completed an IRB approved clinical trial of a HER-2/neu targeted dendritic cell based vaccine. Four weekly vaccinations were administered prior to surgical resection. All subjects underwent pre- and post-vaccine cardiac monitoring by MUGA/ECHO scanning allowing for a comparison of cardiac function. Results In 3 of 27 vaccinated patients (11%) transient asymptomatic decrements in ejection fraction of greater than 15% were noted after vaccination. Notably, evidence of circulating anti-HER-2/neu antibody was found prior to vaccination in all three patients, but cardiac toxicity was not noted until induction of cellular mediated immune responses. Conclusions This is the first description of HER-2/neu targeted vaccination associated with an incidence of cardiac changes, and the induction of cellular immune responses combined with antibody may contribute to changes in cardiac function. PMID:19800453

  10. Immunization with a novel chimeric peptide representing B and T cell epitopes from HER2 extracellular domain (HER2 ECD) for breast cancer.

    Science.gov (United States)

    Mahdavi, Manijeh; Keyhanfar, Mehrnaz; Jafarian, Abbas; Mohabatkar, Hassan; Rabbani, Mohammad

    2014-12-01

    Because of direct stimulating immune system against disease, vaccination or active immunotherapy is preferable compared to passive immunotherapy. For this purpose, a newly designed chimeric peptide containing epitopes for both B and T cells from HER2 ECD subdomain III was proposed. To evaluate the effects of the active immunization, a discontinuous B cell epitope peptide was selected based on average antigenicity by bioinformatics analysis. The selected peptide was collinearly synthesized as a chimera with a T helper epitope from the protein sequence of measles virus fusion (208-302) using the GPSL linker. Three mice were immunized with the chimeric peptide. Reactive antibodies with HER2 protein in ELISA and immunofluorescence assays with no cross-reactivity were generated. The 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay indicated that the anti-peptide sera had inhibitory effects on proliferation of SK-BR-3 cells. Hence, the newly designed, discontinuous chimeric peptide representing B and T cell epitopes from subdomain III of HER2-ECD can form the basis for future vaccines design, where these data can be applied for monoclonal antibody production targeting the distinct epitope of HER2 receptor compared to the two broadly used anti-HER2 monoclonal antibodies, Herceptin and pertuzumab.

  11. Herceptin Enhances the Antitumor Effect of Natural Killer Cells on Breast Cancer Cells Expressing Human Epidermal Growth Factor Receptor-2

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    Xiao Tian

    2017-10-01

    Full Text Available Optimal adoptive cell therapy (ACT should contribute to effective cancer treatment. The unique ability of natural killer (NK cells to kill cancer cells independent of major histocompatibility requirement makes them suitable as ACT tools. Herceptin, an antihuman epidermal growth factor receptor-2 (anti-HER2 monoclonal antibody, is used to treat HER2+ breast cancer. However, it has limited effectiveness and possible severe cardiotoxicity. Given that Herceptin may increase the cytotoxicity of lymphocytes, we explored the possible augmentation of NK cell cytotoxicity against HER2+ breast cancer cells by Herceptin. We demonstrated that Herceptin could interact with CD16 on NK cells to expand the cytotoxic NK (specifically, CD56dim cell population. Additionally, Herceptin increased NK cell migration and cytotoxicity against HER2+ breast cancer cells. In a pilot study, Herceptin-treated NK cells shrunk lung nodular metastasis in a woman with HER2+ breast cancer who could not tolerate the cardiotoxic side effects of Herceptin. Our findings support the therapeutic potential of Herceptin-treated NK cells in patients with HER2+ and Herceptin-intolerant breast cancer.

  12. Rabbit antibodies for hormone receptors and HER2 evaluation in breast cancer Anticorpos de coelho para avaliação de receptores hormonais e HER2 em câncer de mama

    Directory of Open Access Journals (Sweden)

    Rafael Malagoli Rocha

    2009-01-01

    Full Text Available BACKGROUND: Novel rabbit monoclonal antibodies (RabMab for estrogen (ER, progesterone (PR receptors and HER2 evaluation by immunohistochemistry have recently been commercially released. We compared the RabMab anti-ER, anti-PR and anti-HER2 to mouse monoclonal antibodies (Mab using tissue microarrays (TMA of breast carcinomas. METHODS: Two TMA containing breast carcinomas were built. Sections were immunostained using anti-ER and anti-PR, Mab and RabMab. The sections stained for ER and PR were evaluated considering positive those tumors in which more than 1% of the tumor cell nuclei stained moderate or strong. For HER2, the immunostained sections were evaluated using the ASCO/CAP guidelines for HER2. Chromogenic in situ hybridization (CISH was used as the gold standard for HER2 evaluation. CISH was evaluated using the Zymed HER2 CISH interpretation guidelines. RESULTS: RabMab against ER have similar staining patterns compared to the 6F11 (Mab, but stronger than 1D5 (Mab from three different suppliers. The RabMab against PR provide stronger and sharper immunohistochemical signals compared to Mab. The detection of HER2 protein overexpression was more prevalent with the polyclonal antibodies and RabMab than with the Mab. These were more specific than the RabMab, which were more sensitive when compared to CISH. CONCLUSION: The novel RabMab against ER and PR showed higher intensity of staining than the Mab. The RabMab against HER2 is more sensitive than Mab, however, Mab presented more specificity than RabMab when compared to CISH for HER2 evaluation of breast carcinomas.OBJETIVOS: Novos anticorpos monoclonais de coelho (RabMab para a avaliação imuno-histoquímica de receptores de estrógeno (RE, progesterona (RP e HER2 foram lançados comercialmente. Comparamos os RabMab anti-RE, anti-RP e anti-HER2 com os anticorpos monoclonais de camundongo (Mab utilizando tissue microarrays (TMA de carcinomas de mama. MÉTODOS: Foram construídos dois TMAs de

  13. Relevance of deep learning to facilitate the diagnosis of HER2 status in breast cancer

    Science.gov (United States)

    Vandenberghe, Michel E.; Scott, Marietta L. J.; Scorer, Paul W.; Söderberg, Magnus; Balcerzak, Denis; Barker, Craig

    2017-04-01

    Tissue biomarker scoring by pathologists is central to defining the appropriate therapy for patients with cancer. Yet, inter-pathologist variability in the interpretation of ambiguous cases can affect diagnostic accuracy. Modern artificial intelligence methods such as deep learning have the potential to supplement pathologist expertise to ensure constant diagnostic accuracy. We developed a computational approach based on deep learning that automatically scores HER2, a biomarker that defines patient eligibility for anti-HER2 targeted therapies in breast cancer. In a cohort of 71 breast tumour resection samples, automated scoring showed a concordance of 83% with a pathologist. The twelve discordant cases were then independently reviewed, leading to a modification of diagnosis from initial pathologist assessment for eight cases. Diagnostic discordance was found to be largely caused by perceptual differences in assessing HER2 expression due to high HER2 staining heterogeneity. This study provides evidence that deep learning aided diagnosis can facilitate clinical decision making in breast cancer by identifying cases at high risk of misdiagnosis.

  14. Radiolabeling of biological vectors by poly-aza-macrocyclic complexes

    International Nuclear Information System (INIS)

    Moreau, M.

    2012-01-01

    This work conducted at the 'Institut de Chimie Moleculaire de l'Universite de Bourgogne' carries at first on the synthesis of bifunctional chelating agents suitable for the chelation of trivalent radio-metals, including indium-111. The greater part of this work was then dedicated to the grafting of a DOTA derivative bifunctional chelating agent on different antibodies or fragments of monoclonal antibodies: trastuzumab (anti-HER2 treatment of breast cancer), cetuximab (anti EGFR, treatment of many cancers, including colorectal cancer) and abciximab (antiplatelet). Particular attention was paid to the characterization of various immuno-conjugates. The critical step of this thesis consisted in the indium-111 radiolabeling of two previously prepared immuno-conjugates: trastuzumab and cetuximab. These steps of radiolabelling allowed us to determine the immunoreactive fraction and affinity of each radiotracer. Thus, we were able to study the in vivo biodistribution of the radiotracers in tumour-bearing mice by SPECT-CT. We also developed an original method for the labeling of a Fab antibody fragment in order to monitor the biodistribution of the antiplatelet agent (abciximab). Finally, we also validated the concept of multimodal imaging through grafting and radiolabeling of a bimodal agent for optical and SPECT imaging on bacterial lipopolysaccharide. Thanks to this work, we gained an expertise in antibodies radiolabeling. The results obtained allow to consider the labeling of antibodies or other biomolecules, and the use of other radionuclides for PET imaging and radioimmunotherapy. (author)

  15. Metastatic gastric cancer – focus on targeted therapies

    Directory of Open Access Journals (Sweden)

    Meza-Junco J

    2012-06-01

    Full Text Available Judith Meza-Junco, Michael B SawyerDepartment of Oncology, Cross Cancer Institute, Edmonton, Alberta, CanadaAbstract: Gastric cancer (GC is currently the second leading cause of cancer death worldwide; unfortunately, most patients will present with locally advanced or metastatic disease. Despite recent progress in diagnosis, surgery, chemotherapy, and radiotherapy, prognosis remains poor. A better understanding of GC biology and signaling pathways is expected to improve GC therapy, and the integration of targeted therapies has recently become possible and appears to be promising. This article focuses on anti-Her-2 therapy, specifically trastuzumab, as well as other epidermal growth factor receptor antagonists such as cetuximab, panitumub, matuzumab, nimotzumab, gefitinib, and erlotinib. Additionally, drugs that target angiogenesis pathways are also under investigation, particulary bevacizumab, ramucirumab, sorafenib, sunitinib, and cediranib. Other targeted agents in preclinical or early clinical development include mTOR inhibitors, anti c-MET, polo-like kinase 1 inhibitors, anti-insulin-like growth factor, anti-heat shock proteins, and small molecules targeting Hedgehog signaling.Keywords: gastric cancer, targeted therapy, antiangiogenesis drugs, anti-EGFR drugs

  16. Tailoring DNA vaccines: designing strategies against HER2 positive cancers

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    Cristina eMarchini

    2013-05-01

    Full Text Available The crucial role of HER2 in epithelial transformation and its selective overexpression on cancer tissues makes it an ideal target for cancer immunotherapies such as passive immunotherapy with Trastuzumab. There are, however, a number of concerns regarding the use of monoclonal antibodies which include resistance, repeated treatments, considerable costs and side effects that make active immunotherapies against HER2 desirable alternative approaches. The efficacy of anti-HER2 DNA vaccination has been widely demonstrated in transgenic cancer-prone mice, which recapitulate several features of human breast cancers. Nonetheless, the rational design of a cancer vaccine able to trigger a long lasting immunity, and thus prevent tumor recurrence in patients, would require the understanding of how tolerance and immunosuppression regulate antitumor immune responses and, at the same time, the identification of the most immunogenic portions of the target protein. We herein retrace the findings that led to our most promising DNA vaccines that, by encoding human/rat chimeric forms of HER2, are able to circumvent peripheral tolerance. Preclinical data obtained with these chimeric DNA vaccines have provided the rationale for their use in an ongoing phase I clinical trial (EudraCT 2011-001104-34.

  17. Selective internalization of self-assembled artificial oil bodies by HER2/neu-positive cells

    International Nuclear Information System (INIS)

    Chiang, Chung-Jen; Lin, Che-Chin; Lin, Li-Jen; Chang, Chih-Hsiang; Chao, Yun-Peng

    2011-01-01

    A novel delivery carrier was developed using artificial oil bodies (AOBs). Plant seed oil bodies (OBs) consist of a triacylglycerol matrix surrounded by a monolayer of phospholipids embedded with the storage protein oleosin (Ole). Ole consists of a central hydrophobic domain with two amphiphatic arms that extrude from the surface of OBs. In this study, a bivalent anti-HER2/neu affibody domain (ZH2) was fused with Ole at the C terminus. After overproduction in Escherichia coli, the fusion protein (Ole-ZH2) was recovered to assemble AOBs. The size of self-assembled AOBs was tailored by varying the oil/Ole-ZH2 ratio and pH to reach a nanoscale. Upon co-incubation with tumor cells, the nanoscale AOBs encapsulated with a hydrophobic fluorescence dye were selectively internalized by HER2/neu-overexpressing cells and displayed biocompatibility with the cells. In addition, the ZH2-mediated endosomal entry of AOBs occurred in a time- and AOB dose-dependent manner. The internalization efficiency was as high as 90%. The internalized AOBs disintegrated at the non-permissive pH (e.g. in acidic endosomes) and the cargo dye was released. Results of in vitro study revealed a sustained and prolonged release profile. Taken together, our findings indicate the potential of AOBs as a delivery carrier.

  18. Neratinib for the treatment of breast cancer.

    Science.gov (United States)

    Prové, Annemie; Dirix, Luc

    2016-10-03

    Neratinib is an orally available, pan-HER inhibitor with clinical activity in patients with HER2-amplified and HER2-mutated breast cancer. Areas Covered. A summary of publically available and relevant clinical data on neratinib. Expert Opinion. Neratinib (N) is clearly distinct from lapatinib (L), a difference based on its broad anti-HER effect, its covalent target binding and its toxicity profile. The main toxicity of neratinib is gastro-intestinal and is essentially limited to diarrhea. Although not directly compared with single agent lapatinib, skin toxicity is much less pronounced with N. The direct clinical comparison of N-capecitabine versus L-capecitabine is the subject of the ongoing NALA-trial. In patients with advanced disease, neratinib has clinically relevant activity in patients with trastuzumab(T)-pretreated and unpretreated disease. In patients having completed one year of adjuvant trastuzumab, an additional year of neratinib further reduces the risk of recurrence of invasive disease. The activity of neratinib in HER2-mutated advanced disease is subject of ongoing clinical trials but preclinical and early clinical results are promising. Neratinib is a usefull drug and a valuable addition to the different anti-HER2-drugs avalaible for patients with HER2-overexpressing and HER2-mutated breast cancer.

  19. Targeted Approaches Applied to Uncommon Diseases: A Case of Salivary Duct Carcinoma Metastatic to the Brain Treated with the Multikinase Inhibitor Neratinib

    Directory of Open Access Journals (Sweden)

    Karl R. Sorenson

    2017-08-01

    Full Text Available Salivary duct carcinoma is a rare malignancy associated with hormone receptor and human epidermal growth factor receptor 2 (HER2 overexpression. Local surgical control is the cornerstone of therapy, but a subset of patients develops metastatic disease portending a poor prognosis and limited management options. Intracranial metastases are an uncommon manifestation and present a therapeutic challenge. We report the case of a 31-year-old male who presented with facial pain and swelling subsequently diagnosed with salivary duct carcinoma. Our patient underwent extensive locoregional resection and analysis of the tumor tissue demonstrated evidence of androgen receptor expression and HER2 overexpression. His course was complicated by metastatic extra- and intracranial recurrence despite combined modality treatment with radiation and chemotherapy followed by anti-HER2 monoclonal antibody therapy and androgen deprivation therapy. After exhausting standard treatment options, he received experimental therapy with a new small-molecule tyrosine kinase inhibitor, neratinib, with evidence of a transient clinical response and no significant adverse effects. This case exemplifies the potential and limitations of targeted therapy, particularly when applied to patients with rare diseases and presentations.

  20. Targeted Approaches Applied to Uncommon Diseases: A Case of Salivary Duct Carcinoma Metastatic to the Brain Treated with the Multikinase Inhibitor Neratinib.

    Science.gov (United States)

    Sorenson, Karl R; Piovezani Ramos, Guilherme; Villasboas Bisneto, Jose Caetano; Price, Katharine

    2017-01-01

    Salivary duct carcinoma is a rare malignancy associated with hormone receptor and human epidermal growth factor receptor 2 (HER2) overexpression. Local surgical control is the cornerstone of therapy, but a subset of patients develops metastatic disease portending a poor prognosis and limited management options. Intracranial metastases are an uncommon manifestation and present a therapeutic challenge. We report the case of a 31-year-old male who presented with facial pain and swelling subsequently diagnosed with salivary duct carcinoma. Our patient underwent extensive locoregional resection and analysis of the tumor tissue demonstrated evidence of androgen receptor expression and HER2 overexpression. His course was complicated by metastatic extra- and intracranial recurrence despite combined modality treatment with radiation and chemotherapy followed by anti-HER2 monoclonal antibody therapy and androgen deprivation therapy. After exhausting standard treatment options, he received experimental therapy with a new small-molecule tyrosine kinase inhibitor, neratinib, with evidence of a transient clinical response and no significant adverse effects. This case exemplifies the potential and limitations of targeted therapy, particularly when applied to patients with rare diseases and presentations.

  1. HER2 mutated breast cancer responds to treatment with single agent neratinib, a second generation HER2/EGFR tyrosine kinase inhibitor

    Science.gov (United States)

    Ben–Baruch, Noa Efrat; Bose, Ron; Kavuri, Shyam M.; Ma, Cynthia X.; Ellis, Matthew J.

    2015-01-01

    Activating mutations in the HER2 tyrosine kinase have been identified in human breast cancers that lack HER2 gene amplification. These patients are not candidates for HER2 targeted drugs under current standards of care, but preclinical data strongly suggest that these patients will benefit from anti-HER2 drugs. In this case report, we describe a young woman with metastatic breast cancer whose tumor was found to carry a HER2 L755S mutation, which is in the kinase domain of HER2. Treatment with the second generation HER2/EGFR tyrosine kinase inhibitor, neratinib, resulted in partial response and dramatic improvement in the patient’s function status. This partial response lasted 11 months and when the patient’s cancer progressed, she was treated with neratinib plus capecitabine and her cancer again responded. This second response parallels the benefit seen with continuing trastuzumab in HER2 amplified breast cancer after disease progression. This case is the first report, to our knowledge, of successful single agent treatment of HER2 mutated breast cancer. Two clinical trials of neratinib for HER2 mutated, metastatic breast cancer are currently enrolling patients. Further, data from The Cancer Genome Atlas project have identified HER2 mutations in a wide range of solid tumors, including bladder, colorectal, and non-small cell lung cancer, suggesting that clinical trials of neratinib or neratinib-based combinations for HER2 mutated solid tumors is warranted. PMID:26358790

  2. Pharmacodynamics, pharmacokinetics and clinical efficacy of neratinib in HER2-positive breast cancer and breast cancer with HER2 mutations.

    Science.gov (United States)

    Kourie, Hampig Raphael; Chaix, Marie; Gombos, Andrea; Aftimos, Phillippe; Awada, Ahmad

    2016-08-01

    Despite the availability of several potent HER2-directed targeted agents, primary and acquired resistance continues to influence patient outcomes in HER2-positive breast cancer. Neratinib is an irreversible pan-HER tyrosine kinase inhibitor in late-phase clinical development. This review article focuses on neratinib in the treatment of HER2-positive breast cancer - early and metastatic stage - and HER2-mutant breast cancer, with particular emphasis on the pharmacokinetics and pharmacodynamics of the drug. The phase III ExteNET trial shows that neratinib improves 2-year invasive disease-free survival after trastuzumab-based adjuvant therapy in early-stage HER2-positive breast cancer, and in particular HER2+/HR+ tumors. Survival data are awaited. The investigational role of neratinib in high-risk patients or conversely in de-escalation dual regimens with other anti-HER2 therapies and without chemotherapy are of interest. Phase II trials show that neratinib has efficacy, either as monotherapy or in combination with other chemotherapeutic or endocrine agents, in patients with HER2-positive metastatic breast cancer and in tumors harboring HER2 mutations. The role of neratinib in therapeutic algorithms of HER2-positive patients, as well as delaying CNS events, awaits the results of ongoing trials such as NALA. Diarrhea, the main toxicity of neratinib, can be effectively managed with early loperamide prophylaxis.

  3. Tyrosine kinase inhibitors for brain metastases in HER2-positive breast cancer.

    Science.gov (United States)

    Duchnowska, Renata; Loibl, Sibylle; Jassem, Jacek

    2018-06-01

    Approximately 30-50% of advanced HER2-positive breast cancer patients will develop central nervous system (CNS) metastases, with an annual risk of around 10%, and a half of them will die from brain progression. An increased risk of brain metastases is also seen in patients with early HER2-positive breast cancer administered curative therapy. Brain metastases in HER2-positive breast cancer patients usually constitute the first site of recurrence. The administration of anti-HER2 monoclonal antibodies, trastuzumab and pertuzumab, considerably delays the onset of symptomatic brain disease: however, the limited penetration of these compounds into the CNS hinders their efficacy. The small-molecule tyrosine kinase inhibitors of epidermal growth factor receptors family have established activity in HER2-positive breast cancer in both advanced disease and neoadjuvant setting. Favorable physico-chemical properties of these compounds allow them for a more efficient penetration through the blood-brain barrier, and hold the promise for more effective prevention and treatment of brain metastases. In this article we review the role of currently available or investigational HER2 tyrosine kinase inhibitors: lapatinib, neratinib, afatinib and tucatinib in the treatment of brain metastases in HER2-positive breast cancer patients. Copyright © 2018 Elsevier Ltd. All rights reserved.

  4. HER2-Mutated Breast Cancer Responds to Treatment With Single-Agent Neratinib, a Second-Generation HER2/EGFR Tyrosine Kinase Inhibitor.

    Science.gov (United States)

    Ben-Baruch, Noa Efrat; Bose, Ron; Kavuri, Shyam M; Ma, Cynthia X; Ellis, Matthew J

    2015-09-01

    Activating mutations in the HER2 tyrosine kinase have been identified in human breast cancers that lack HER2 gene amplification. These patients are not candidates for HER2-targeted drugs under current standards of care, but preclinical data strongly suggest that these patients will benefit from anti-HER2 drugs. This case report describes a young woman with metastatic breast cancer whose tumor was found to carry a HER2 L755S mutation, which is in the kinase domain of HER2. Treatment with the second-generation HER2/EGFR tyrosine kinase inhibitor neratinib resulted in partial response and dramatic improvement in the patient's functional status. This partial response lasted 11 months, and when the patient's cancer progressed, she was treated with neratinib plus capecitabine and her cancer again responded. This second response parallels the benefit seen with continuing trastuzumab in HER2-amplified breast cancer after disease progression. This case represents the first report, to our knowledge, of successful single-agent treatment of HER2-mutated breast cancer. Two clinical trials of neratinib for HER2-mutated metastatic breast cancer are currently enrolling patients. Further, data from The Cancer Genome Atlas project have identified HER2 mutations in a wide range of solid tumors, including bladder, colorectal, and non-small cell lung cancers, suggesting that clinical trials of neratinib or neratinib-based combinations for HER2-mutated solid tumors is warranted. Copyright © 2015 by the National Comprehensive Cancer Network.

  5. Functionalized nanoscale oil bodies for targeted delivery of a hydrophobic drug

    Energy Technology Data Exchange (ETDEWEB)

    Chiang, Chung-Jen; Lin, Che-Chin; Lu, Tzu-Li; Wang, Hesin-Fu, E-mail: cjchiang@mail.cmu.edu.tw [Department of Medical Laboratory Science and Biotechnology, China Medical University, 91 Hsue-Shih Road, Taichung 40402, Taiwan (China)

    2011-10-14

    Effective formulations of hydrophobic drugs for cancer therapies are challenging. To address this issue, we have sought to nanoscale artificial oil bodies (NOBs) as an alternative. NOBs are lipid-based particles which consist of a central oil space surrounded by a monolayer of oleosin (Ole)-embedded phospholipids (PLs). Ole was first fused with the anti-HER2/neu affibody (Ole-ZH2), and the resulting hybrid protein was overproduced in Escherichia coli. ZH2-displayed NOBs were then assembled by sonicating the mixture containing plant oil, PLs, and isolated Ole-ZH2 in one step. To illustrate their usefulness, functionalized NOBs were employed to encapsulate a hydrophobic anticancer drug, Camptothecin (CPT). As a result, these CPT-loaded NOBs remained stable in serum and the release of CPT at the non-permissive condition exhibited a sustained and prolonged profile. Moreover, plain NOBs were biocompatible whereas CPT-loaded NOBs exerted a strong cytotoxic effect on HER2/neu-positive cells in vitro. Administration of xenograft nude mice with CPT-loaded NOBs also led to the regression of solid tumors in an effective way. Overall, the result indicates the potential of NOBs for targeted delivery of hydrophobic drugs.

  6. How to treat brain metastasis in 2012?

    International Nuclear Information System (INIS)

    Braccini, A.L.; Azria, D.; Mazeron, J.J.; Mornex, F.; Jacot, W.; Metellus, P.; Tallet, A.

    2012-01-01

    During the last French radiation oncology society annual congress, the therapeutic options for the management of brain metastases were presented. The indications and limits of surgery, stereotactic radiotherapy and whole brain radiotherapy, as well as their benefit in terms of overall survival, local control and improvement of the functional and neuro-cognitive status were discussed. The prognosis significance of the different phenotypes of breast cancer on the risk for BM as well as their roles in the treatment of brain metastases were also described. Surgery improves overall survival for patients with a single brain metastase and should be considered in the case of symptomatic lesions. The overall survival of patients treated with stereotactic radiotherapy do not differ from that of patients treated with surgery. These treatments should be mainly considered for patients with good performance status, one to three small brain metastases (< 3 cm) and limited extracranial disease. Whole brain radiotherapy is more and more discussed in adjuvant setting due to potential late neuro-cognitive toxicity. This toxicity could be improved with the development of techniques sparing the hippocampus. HER2+ and triple-negative breast cancer patients are at increased risk for brain metastases. Prognosis of these patients differs as the overall survival of HER2+ patients has improved with anti-HER2 therapies. The optimal combination of local and systemic therapies remain to be determined. (authors)

  7. Sanctuary site leptomeningeal metastases in HER-2 positive breast cancer: A review in the era of trastuzumab.

    Science.gov (United States)

    Kordbacheh, T; Law, W Y; Smith, I E

    2016-04-01

    The development of trastuzumab and other targeted systemic therapies has transformed the management of HER-2 positive breast cancers. However, as patients live longer and systemic therapies may not cross the blood brain barrier a rising number of patients are developing leptomeningeal metastases and brain metastases as a sanctuary site of disease. Intrathecal trastuzumab has been reported to treat these. We describe a breast cancer patient with HER-2 positive leptomeningeal disease in the spinal cord successfully treated with intrathecal trastuzumab and methotrexate, alongside systemic anti-HER-2 therapy and radiotherapy. We also review the literature to date on the efficacy and safety of intrathecal trastuzumab, and recent evidence suggesting that intrathecal trastuzumab passes via the blood brain barrier into the serum to achieve intravenous concentrations similar to that seen with systemic therapy alone. Overall, intrathecal trastuzumab appears to be a safe and often effective treatment for leptomeningeal metastases in HER-2 positive breast cancer. Ongoing phase I and II studies are required to determine optimum dosing schedules, validate CSF and CSF-to-serum pharmacokinetics, determine efficacy, and to assess the added benefits or disadvantages of prior radiotherapy and concomitant systemic therapy. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. Management of advanced gastric cancer: An overview of major findings from meta-analysis

    Science.gov (United States)

    Cai, Danxian; Li, Wende; Hui, Jialiang; Liu, Chuan; Zhao, Yanxia; Li, Guoxin

    2016-01-01

    This study aims to provide an overview of different treatment for advanced gastric cancer. In the present study, we systematically reviewed the major findings from relevant meta-analyses. A total of 54 relevant papers were searched via the PubMed, Web of Science, and Google scholar databases. They were classified according to the mainstay treatment modalities such as surgery, chemotherapy and others. Primary outcomes including overall survival, response rate, disease-free survival, recurrence-free survival, progression-free survival, time-to-progression, time-to failure, recurrence and safety were summarized. The recommendations and uncertainties regarding the treatment of advanced gastric cancer were also proposed. It was suggested that laparoscopic gastrectomy was a safe and technical alternative to open gastrectomy. Besides, neoadjuvant chemotherapy and adjuvant chemotherapy were thought to benefit the survival over surgery alone. And it was demonstrated in the study that targeted therapy like anti-angiogenic and anti-HER2 agents but anti-EGFR agent might have a significant survival benefit. PMID:27655725

  9. Use of maintenance endocrine therapy after chemotherapy in metastatic breast cancer.

    Science.gov (United States)

    Sutherland, S; Miles, D; Makris, A

    2016-12-01

    For women with oestrogen receptor+ metastatic breast cancer (MBC), the options for systemic treatment include endocrine therapy (ET) and chemotherapy. For women whose disease is also HER2+, anti-HER2 therapies are also routinely used either with chemotherapy or less commonly with ET. Where chemotherapy is used as initial therapy, treatment is often discontinued due to cumulative toxicity in the absence of disease progression. In this setting, there is the option of introducing ET with the aim of prolonging response and delaying relapse. Literature review revealed four trials addressing the question of whether there is a benefit from introducing ET following chemotherapy for MBC. We also sought evidence for alternative approaches, including concurrent chemotherapy and ET and continuing chemotherapy until disease progression. The evidence for the use of ET after chemotherapy in MBC is limited, and the trials done were small. Furthermore, they were performed at a time when both the chemotherapy regimens and ET were different from those used currently. Despite these limitations, there is probably a modest improvement in time to progression for the sequential use of ET after chemotherapy but with no overall survival benefit. An alternative approach, particularly considering agents with relatively low toxicity, such as orally bioavailable fluoropyrimidines, is to continue chemotherapy until disease progression. Where chemotherapy for MBC is discontinued due to toxicity, in the absence of progression, the use of ET, with its relatively low toxicity, is a reasonable approach with the aim of delaying relapse. Copyright © 2016. Published by Elsevier Ltd.

  10. Trastuzumab emtansine: first global approval.

    Science.gov (United States)

    Ballantyne, Anita; Dhillon, Sohita

    2013-05-01

    Genentech and ImmunoGen are collaborating on the development of trastuzumab emtansine, a HER2 antibody-drug conjugate that comprises Genentech's trastuzumab antibody linked to ImmunoGen's anti-mitotic agent, mertansine (a maytansine derivative; also known as DM1). The conjugate combines two strategies: the anti-HER2 activity of trastuzumab, and the targeted intracellular delivery of mertansine, a tubulin polymerisation inhibitor which interferes with mitosis and promotes apoptosis. The linker in trastuzumab emtansine is a non-reducible thioether linker, N-succinimidyl-4-(N-maleimidomethyl) cyclohexane-1-carboxylate (SMCC, designated MCC after conjugation). Trastuzumab emtansine (Kadcyla™) has been launched in the USA as second-line monotherapy for HER2-positive metastatic breast cancer, and has been filed for approval in the EU and Japan in this indication. Trastuzumab emtansine is in phase III development as first-line combination therapy or monotherapy for metastatic HER2-positive breast cancer, and as third-line monotherapy for metastatic HER2-positive breast cancer. Phase II development is underway for early-stage breast cancer and phase II/III development is underway in patients with HER2-positive gastric cancer. This article summarizes the milestones in the development of trastuzumab emtansine leading to this first approval for the treatment of patients with HER2-positive, metastatic breast cancer who previously received trastuzumab and a taxane, separately or in combination.

  11. Color feature extraction of HER2 Score 2+ overexpression on breast cancer using Image Processing

    Directory of Open Access Journals (Sweden)

    Muhimmah Izzati

    2018-01-01

    Full Text Available One of the major challenges in the development of early diagnosis to assess HER2 status is recognized in the form of Gold Standard. The accuracy, validity and refraction of the Gold Standard HER2 methods are widely used in laboratory (Perez, et al., 2014. Method determining the status of HER2 (human epidermal growth factor receptor 2 is affected by reproductive problems and not reliable in predicting the benefit from anti-HER2 therapy (Nuciforo, et al., 2016. We extracted color features by methods adopting Statistics-based segmentation using a continuous-scale naïve Bayes approach. In this study, there were three parts of the main groups, namely image acquisition, image segmentation, and image testing. The stages of image acquisition consisted of image data collection and color deconvolution. The stages of image segmentation consisted of color features, classifier training, classifier prediction, and skeletonization. The stages of image testing were image testing, expert validation, and expert validation results. Area segmentation of the membrane is false positive and false negative. False positive and false negative from area are called the area of system failure. The failure of the system can be validated by experts that the results of segmentation region is not membrane HER2 (noise and the segmentation of the cytoplasm region. The average from 40 data of HER2 score 2+ membrane images show that 75.13% of the area is successfully recognized by the system.

  12. Immunohistochemistry and alternative FISH testing in breast cancer with HER2 equivocal amplification.

    Science.gov (United States)

    Agersborg, Sally; Mixon, Christopher; Nguyen, Thanh; Aithal, Sramila; Sudarsanam, Sucha; Blocker, Forrest; Weiss, Lawrence; Gasparini, Robert; Jiang, Shiping; Chen, Wayne; Hess, Gregory; Albitar, Maher

    2018-03-22

    While HER2 testing is well established in directing appropriate treatment for breast cancer, a small percentage of cases show equivocal results by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). Alternative probes may be used in equivocal cases. We present a single community-based institution's experience in further evaluating these cases. Between 2014 and 2016, 4255 samples were submitted for HER2 amplification testing by alternative probes, TP53, RAI1, and RARA. Of the patients tested by FISH, 505/3908 (12.9%) also had IHC data. Most (73.9%) FISH equivocal cases remained equivocal after IHC testing. However, 50.5% of equivocal cases were classified as HER2 amplified by alternative probes. Most cases were positive by more than one probe: 78% of positive cases by RAI1 and 73.9% by TP53. There was a significant difference between IHC and FISH alternative testing (p alternative FISH testing. Available data showed that 41% of patients were treated with palbociclib and were positive by alternative FISH. The prevalence of double HER2 equivocal cases and the discrepancy between IHC and alternative FISH testing suggest that FISH alternative testing using both RAI1 and TP53 probes is necessary for conclusive classification. Because almost half of FISH equivocal cases converted to HER2 amplified upon alternative testing, clinical studies to determine the benefit of anti-HER2 therapy in these patients are urgently needed.

  13. Caleosin-based nanoscale oil bodies for targeted delivery of hydrophobic anticancer drugs

    International Nuclear Information System (INIS)

    Chiang, Chung-Jen; Lin, Li-Jen; Chen, Chih-Jung

    2011-01-01

    Nanoscale artificial oil bodies (NOBs) could be assembled from plant oil, phospholipids (PLs), and oleosin (Ole) as previously reported. NOBs have a lipid-based structure that contains a central oil space enclosed by a monolayer of Ole-bound PLs. As an oil structural protein, Ole functions to maintain the integrity of NOBs. Like Ole, caleosin (Cal) is a plant oil-associated protein. In this study, we investigated the feasibility of NOBs assembled by Cal for targeted delivery of drugs. Cal was first fused with anti-HER2/neu affibody (ZH2), and the resulting fusion gene (Cal–ZH2) was then expressed in Escherichia coli. Consequently, NOBs assembled with the fusion protein were selectively internalized by HER2/neu-positive tumor cells. The internalization efficiency could reach as high as 90%. Furthermore, a hydrophobic anticancer drug, Camptothecin (CPT), was encapsulated into Cal-based NOBs. These CPT-loaded NOBs had a size around 200 nm and were resistant to hemolysis. Release of CPT from NOBs at the non-permissive condition followed a sustained and prolonged profile. After administration of the CPT formulation, Cal–ZH2-displayed NOBs exhibited a strong antitumor activity toward HER2/neu-positive cells both in vitro and in vivo. The result indicates the potential of Cal-based NOBs for targeted delivery of hydrophobic drugs.

  14. Brain metastasis in human epidermal growth factor receptor 2-positive breast cancer: from biology to treatment

    Energy Technology Data Exchange (ETDEWEB)

    Koo, Tae Ryool [Dept. of Radiation Oncology, Hallym University Chuncheon Sacred Heart Hospital, Chuncheon (Korea, Republic of); Kim, In Ah [Dept. of Radiation Oncology, Seoul National University Bundang Hospital, Seongnam (Korea, Republic of)

    2016-03-15

    Overexpression of human epidermal growth factor receptor 2 (HER2) is found in about 20% of breast cancer patients. With treatment using trastuzumab, an anti-HER2 monoclonal antibody, systemic control is improved. Nonetheless, the incidence of brain metastasis does not be improved, rather seems to be increased in HER2-positive breast cancer. The mainstay treatment for brain metastases is radiotherapy. According to the number of metastatic lesions and performance status of patients, radiosurgery or whole brain radiotherapy can be performed. The concurrent use of a radiosensitizer further improves intracranial control. Due to its large molecular weight, trastuzumab has a limited ability to cross the blood-brain barrier. However, small tyrosine kinase inhibitors such as lapatinib, has been noted to be a promising agent that can be used as a radiosensitizer to affect HER2-positive breast cancer. This review will outline general management of brain metastases and will focus on preclinical findings regarding the radiosensitizing effect of small molecule HER2 targeting agents.

  15. Circulating myeloid-derived suppressor cells increase in patients undergoing neo-adjuvant chemotherapy for breast cancer.

    Science.gov (United States)

    Wesolowski, Robert; Duggan, Megan C; Stiff, Andrew; Markowitz, Joseph; Trikha, Prashant; Levine, Kala M; Schoenfield, Lynn; Abdel-Rasoul, Mahmoud; Layman, Rachel; Ramaswamy, Bhuvaneswari; Macrae, Erin R; Lustberg, Maryam B; Reinbolt, Raquel E; Mrozek, Ewa; Byrd, John C; Caligiuri, Michael A; Mace, Thomas A; Carson, William E

    2017-11-01

    This study sought to evaluate whether myeloid-derived suppressor cells (MDSC) could be affected by chemotherapy and correlate with pathologic complete response (pCR) in breast cancer patients receiving neo-adjuvant chemotherapy. Peripheral blood levels of granulocytic (G-MDSC) and monocytic (M-MDSC) MDSC were measured by flow cytometry prior to cycle 1 and 2 of doxorubicin and cyclophosphamide and 1st and last administration of paclitaxel or paclitaxel/anti-HER2 therapy. Of 24 patients, 11, 6 and 7 patients were triple negative, HER2+ and hormone receptor+, respectively. 45.8% had pCR. Mean M-MDSC% were types. G-MDSC levels at the last draw were numerically lower in patients with pCR (1.15; 95% CI 0.14-2.16) versus patients with no pCR (2.71; 95% CI 0-5.47). There was no significant rise in G-MDSC from draw 1 to 3 in African American patients, and at draw 3 G-MDSC levels were significantly lower in African Americans versus Caucasians (p < 0.05). It was concluded that G-MDSC% increased during doxorubicin and cyclophosphamide therapy, but did not significantly differ between patients based on pathologic complete response.

  16. [Evaluation and classification of drug therapy for breast cancer with bone-only metastasis].

    Science.gov (United States)

    Meng, X Y; Song, S T

    2017-03-23

    Skeleton is one of the most common metastatic organs for breast cancer, which has a better prognosis than visceral metastases. Bone-only metastasis was defined"non-measurable" in the RECIST (Response Evaluation Criteria in Solid Tumors) criteria, and was excluded by clinical trials. However, patients with bone-only metastasis are also in need of effective treatment to prolong survival. Endocrine therapy is the most important treatment for bone metastatic patients. Tumor response of bone metastases can be determined objectively by bone-window CT. Effective treatment should be continued if the symptoms are relieved or osteogenesis is observed. Osteoblastic change in bone-window CT is a sign of improvement after treatment. Endocrine therapy is proper for ER-positive patients. The patients with initial osteoblastic metastasis should not be treated with salvage chemotherapy or anti-HER2 treatment, only if osteolytic metastasis or visceral metastasis is observed. Bishosphonates are just auxiliary drugs in bone metastasis, which should not be abused.

  17. Molecular targeted therapy for advanced gastric cancer.

    Science.gov (United States)

    Kim, Jong Gwang

    2013-03-01

    Although medical treatment has been shown to improve quality of life and prolong survival, no significant progress has been made in the treatment of advanced gastric cancer (AGC) within the last two decades. Thus, the optimum standard first-line chemotherapy regimen for AGC remains debatable, and most responses to chemotherapy are partial and of short duration; the median survival is approximately 7 to 11 months, and survival at 2 years is exceptionally > 10%. Recently, remarkable progress in tumor biology has led to the development of new agents that target critical aspects of oncogenic pathways. For AGC, many molecular targeting agents have been evaluated in international randomized studies, and trastuzumab, an anti-HER-2 monoclonal antibody, has shown antitumor activity against HER-2-positive AGC. However, this benefit is limited to only ~20% of patients with AGC (patients with HER-2-positive AGC). Therefore, there remains a critical need for both the development of more effective agents and the identification of molecular predictive and prognostic markers to select those patients who will benefit most from specific chemotherapeutic regimens and targeted therapies.

  18. Spectral counting assessment of protein dynamic range in cerebrospinal fluid following depletion with plasma-designed immunoaffinity columns

    Directory of Open Access Journals (Sweden)

    Borg Jacques

    2011-06-01

    Full Text Available Abstract Background In cerebrospinal fluid (CSF, which is a rich source of biomarkers for neurological diseases, identification of biomarkers requires methods that allow reproducible detection of low abundance proteins. It is therefore crucial to decrease dynamic range and improve assessment of protein abundance. Results We applied LC-MS/MS to compare the performance of two CSF enrichment techniques that immunodeplete either albumin alone (IgYHSA or 14 high-abundance proteins (IgY14. In order to estimate dynamic range of proteins identified, we measured protein abundance with APEX spectral counting method. Both immunodepletion methods improved the number of low-abundance proteins detected (3-fold for IgYHSA, 4-fold for IgY14. The 10 most abundant proteins following immunodepletion accounted for 41% (IgY14 and 46% (IgYHSA of CSF protein content, whereas they accounted for 64% in non-depleted samples, thus demonstrating significant enrichment of low-abundance proteins. Defined proteomics experiment metrics showed overall good reproducibility of the two immunodepletion methods and MS analysis. Moreover, offline peptide fractionation in IgYHSA sample allowed a 4-fold increase of proteins identified (520 vs. 131 without fractionation, without hindering reproducibility. Conclusions The novelty of this study was to show the advantages and drawbacks of these methods side-to-side. Taking into account the improved detection and potential loss of non-target proteins following extensive immunodepletion, it is concluded that both depletion methods combined with spectral counting may be of interest before further fractionation, when searching for CSF biomarkers. According to the reliable identification and quantitation obtained with APEX algorithm, it may be considered as a cheap and quick alternative to study sample proteomic content.

  19. PEMANFAATAN TELUR AYAM SEBAGAI PABRIK BIOLOGIS (KAJIAN PUSTAKA

    Directory of Open Access Journals (Sweden)

    I WAYAN TEGUH WIBAWAN

    2012-09-01

    Full Text Available ABSTRAK Penelitian tentang pemanfaatan telur sebagai pabrik biologis dilakukan sangat komprehensif di beberapa institusi perguruan tinggi di Inonesia, yakni Fakultas Kedokteran Hewan (FKH Institut Pertanian Bogor (IPB, FKH Universitas Gadjah Mada (UGM, FKH Universitas Udayana (UNUD, FKH Universitas Syahkuala (Unsyiah, Fakultas Kedokteran Gigi Universitas Indonesia. Jejaring penelitian dibangun dengan baik dan telah menghasilkan beberapa produk, antara lain: telur berkhasiat flu burung H5N1, telur Anti Tetanus Serum, Telur Anti Diare, Telur anti Plaque dan Telur Anti White Spot Syndrome Virus pada udang. Secara ilmiah khasiat IgY spesifik dalam kuning telur sebagai senyawa therapeutic telah diuji dan tinggal memerlukan sentuhan akhir tersendiri untuk dapat disajikan sebagai produk komersial. Peran industri yang relevan sangat dibutuhkan dalam mewujudkan hal ini. THE USE OF EGG AS A BIOLOGICAL PRODUCER FOR SPESIFIC ANTIBODIES ABSTRACTS The use of egg as a biological producer for specific antibodies were done intensively among well known universities in Indonesia, such as Faculty of Veterinary Medicine (FVM of FVM of IPB, FVM of UGM, FVM of UDAYANA and FVM of UNSYIAH as well as Faculty of Dental Medicine Indonesia University. The collaboration and research networking among those institutions run as expected and succeed to produce some important products such as eggs containing IgY anti avian flu as well as anti tetanus, anti diarrhea, anti plaques and eggs anti WSSV in shrimp. The potency of specific IgY as immunotherapeutic substances had been studied and need the commercial touch from the respective industries until this products in market available.

  20. Analysis of immunoglobulin transcripts in the ostrich Struthio camelus, a primitive avian species.

    Directory of Open Access Journals (Sweden)

    Tian Huang

    Full Text Available Previous studies on the immunoglobulin (Ig genes in avian species are limited (mainly to galliformes and anseriformes but have revealed several interesting features, including the absence of the IgD and Igκ encoding genes, inversion of the IgA encoding gene and the use of gene conversion as the primary mechanism to generate an antibody repertoire. To better understand the Ig genes and their evolutionary development in birds, we analyzed the Ig genes in the ostrich (Struthio camelus, which is one of the most primitive birds. Similar to the chicken and duck, the ostrich expressed only three IgH chain isotypes (IgM, IgA and IgY and λ light chains. The IgM and IgY constant domains are similar to their counterparts described in other vertebrates. Although conventional IgM, IgA and IgY cDNAs were identified in the ostrich, we also detected a transcript encoding a short membrane-bound form of IgA (lacking the last two C(H exons that was undetectable at the protein level. No IgD or κ encoding genes were identified. The presence of a single leader peptide in the expressed heavy chain and light chain V regions indicates that gene conversion also plays a major role in the generation of antibody diversity in the ostrich. Because the ostrich is one of the most primitive living aves, this study suggests that the distinct features of the bird Ig genes appeared very early during the divergence of the avian species and are thus shared by most, if not all, avian species.

  1. Antibodies against Venom of the Snake Deinagkistrodon acutus.

    Science.gov (United States)

    Lee, Chi-Hsin; Lee, Yu-Ching; Liang, Meng-Huei; Leu, Sy-Jye; Lin, Liang-Tzung; Chiang, Jen-Ron; Yang, Yi-Yuan

    2016-01-01

    Snake venom protein from Deinagkistrodon acutus (DA protein), one of the major venomous species in Taiwan, causes hemorrhagic symptoms that can lead to death. Although horse-derived antivenin is a major treatment, relatively strong and detrimental side effects are seen occasionally. In our study, yolk immunoglobulin (IgY) was purified from eggs, and DA protein was recognized using Western blotting and an enzyme-linked immunosorbent assay (ELISA), similar to therapeutic horse antivenin. The ELISA also indicated that specific IgY antibodies were elicited after the fifth booster, plateaued, and lasted for at least 3 months. To generate monoclonal single-chain variable fragment (scFv) antibodies, we used phage display technology to construct two libraries with short or long linkers, containing 6.24 × 10(8) and 5.28 × 10(8) transformants, respectively. After four rounds of biopanning, the eluted phage titer increased, and the phage-based ELISA indicated that the specific clones were enriched. Nucleotide sequences of 30 individual clones expressing scFv were analyzed and classified into four groups that all specifically recognized the DA venom protein. Furthermore, based on mass spectrometry, the scFv-bound protein was deduced to be snake venom metalloproteinase proteins. Most importantly, both IgY and mixed scFv inhibited the lethal effect in mice injected with the minimum lethal dosage of the DA protein. We suggest that together, these antibodies could be applied to the development of diagnostic agents or treatments for snakebite envenomation in the future. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  2. The glycosylated IgII extracellular domain of EMMPRIN is implicated in the induction of MMP-2.

    Science.gov (United States)

    Papadimitropoulou, Adriana; Mamalaki, Avgi

    2013-07-01

    EMMPRIN is a widely expressed transmembrane glycoprotein that plays important roles in many physiological and pathological processes, such as tumor invasion and metastasis. It stimulates the production of matrix metalloproteinase (MMPs) by tumor-associated fibroblasts. In the present study, our aim was to (a) to investigate if the IgII loop domain of the extracellular domain (ECD) of EMMPRIN contributes to the MMP production by fibroblasts and (b) to evaluate the significance of glycosylation in this process. For this purpose, we expressed the ECD, IgI, or IgII domains of EMMPRIN, in their glycosylated and non-glycosylated forms, in the heterologous expression systems of P. pastoris and E. coli, respectively. Dermal fibroblasts were treated with purified recombinant domains and proteins from cell extracts and supernatants were analyzed by Western blot and zymography assays. Fibroblasts treated with ECD-, IgI-, and IgII-glycosylated domains of EMMPRIN significantly stimulated the gelatinolytic activity of MMP-2, compared to untreated fibroblasts, whereas no significant effect was observed after treatment with the non-glycosylated ECD, IgI, and IgII domains. Western blot analysis from cell extracts and supernatants revealed that only the glycosylated forms were able to stimulate MMP-2 production and secretion, respectively. Quantitative PCR revealed that this effect was not attributed to transcriptional alterations. This study showed that N-glycosylation was a prerequisite for efficient MMP-2 production, with the IgII loop domain contributing significantly to this process. Perturbation of the function of IgII-EMMPRIN loop could have potential therapeutic value in the inhibition of MMP-2-dependent cancer cell invasion and metastasis.

  3. Use of Candida-specific chicken egg yolk antibodies to inhibit the adhering of Candida to denture base materials: prevention of denture stomatitis.

    Science.gov (United States)

    Kamikawa, Yoshiaki; Fujisaki, Junichi; Nagayama, Tomohiro; Kawasaki, Kiyotsugu; Hirabayashi, Daisuke; Hamada, Tomofumi; Sakamoto, Ryoich; Mukai, Hiroshi; Sugihara, Kazumasa

    2016-09-01

    Polyclonal anti-Candida chicken egg yolk antibodies (anti-IgY) were used to investigate the prevention of adherence of Candida species to denture base material in vitro. Candida is a potential virulence factor that can cause systemic infection and even death in immunocompromised individuals. Because long-term antifungal treatment may lead to the emergence of drug-resistant strains, it is necessary to develop novel preventive measures and treatments for candidiasis. Three types of chicken egg yolk antibodies were used in this study: non-specific antibody (control IgY), Candida albicans-specific antibody (anti-C.a.IgY) and Candida glabrata-specific antibody (anti-C.g.IgY). A mixture of different dilutions of each antibody with a suspension of Candida species and denture base material was incubated for 3 h, and then the colony-forming units of Candida on the denture base material were counted. Compared with control IgY, anti-C.a.IgY and anti-C.g.IgY significantly inhibited the adherence of C. albicans, but anti-C.a.IgY tended to be more potent than anti-C.g.IgY. The adherence of C. glabrata was also inhibited significantly by anti-C.a.IgY and anti-C.g.IgY with almost equivalent potency, indicating that their actions against C. glabrata were comparable. This study revealed the inhibitory effects of anti-C.a.IgY and anti-C.g.IgY against the adherence of C. albicans and C. glabrata to denture base material. This finding indicates the possibility of a beneficial effect of IgYs for the prevention of denture stomatitis and candidiasis in clinical settings. © 2014 John Wiley & Sons A/S and The Gerodontology Association. Published by John Wiley & Sons Ltd.

  4. Development and primary application of ELISA method for detecting residual calf serum protein

    International Nuclear Information System (INIS)

    Li Hua; Yu Jiankun; Hong Chao; Long Ruixiang; Zhai Yougang; Xu Qiongfang; Cui Pingfang; Ding Xuefeng; Xie Zhongping

    2005-01-01

    To develop a new DAS-ELISA (double antibody sandwich ELISA) kit for detecting residual calf serum protein (CSP) in vaccines, calf sera from different district were pooled and used to immunize rabbits and hens respectively. Then, the IgY from yolk and the anti-CSP IgG from rabbit were separated and purified. The purified IgY was used as the coating antibody, and purified rabbit anti-CSP IgG was labeled by HRP. The optimal concentration of IgY was 25-30 μg/mL. The coating buffer was 0.01mol/L PBS(pH7.4) containing 0.4% glutin. The optimal dilutions of HRP-IgG were from 1:2000 to 1:3000. The sensitivity of this ELISA method was higher (up to 2.5μg/mL) than that of current RPHA, the variation coefficient was about 6.3%-9.4%, and the recovery rate was 90.4%-112.8%. Furthermore, there was no cross-reaction with sera of pig, monkey and guinea pig. Twenty lots of vaccines with Al(OH) 3 or without Al(OH) 3 were tested by ELISA and RPHA respectively. The results proved that the adjuvant of Al(OH) 3 had fewer influence on ELISA than on RPHA, the variation of PRHA among different lots of vaccines was more significant than ELISA. The ELISA method is a highly sensitive and useful method to detect CSP in vaccines. (authors)

  5. THE USE OF THE SPECIFIC ANTI-SALMONELLA POLYCLONAL ANTIBODIES ISOLATED FROM HEN EGGS, IN SALMONELLOSIS PROPHYLAXIS

    Directory of Open Access Journals (Sweden)

    CRISTE ADRIANA

    2007-01-01

    Full Text Available The administration of increased doses of antibodies in groupsexperimentally infected with Salmonella gallinarum, in order torecord the efficiency of their administration in salmonellosisprophylaxis was the aim of our research. When a low infectiondose, 1x107 CFU Salmonella gallinarum, was used theadministration of IgY polyclonal antibodies as immunoglobulinextract, or even yolk administration had a protective effect againstgerms invasion. This effect was not recorded when a 10 folds higherdose was administered (1x108 CFU. The prophylactic effect of theadministration of polyclonal antibodies is demonstrated.

  6. Observed travel times for Multiply-reflected ScS waves from a deep-focus earthquake

    Directory of Open Access Journals (Sweden)

    A. F. ESPISOSA

    1966-06-01

    Full Text Available Tlie deep-focus Argentinean earthquake of December 8
    1962, generated multiply reflected ScS phases which were recorded very
    clearly at stations of the IGY and the TJSC&GS standardized worldwide
    networks and at Canadian stations. The data gathered from this earthquake
    for the multiply-reflected ScS and sScS were used to construct the
    travel times and to extend them to shorter epicentral distances. These
    new data brought to light an error in published travel times for the 2(ScS
    phase.

  7. THE USE OF THE SPECIFIC ANTI-SALMONELLA POLYCLONAL ANTIBODIES ISOLATED FROM HEN EGGS, IN SALMONELLOSIS PROPHYLAXIS

    Directory of Open Access Journals (Sweden)

    ADRIANA CRISTE

    2007-05-01

    Full Text Available The administration of increased doses of antibodies in groups experimentallyinfected with Salmonella gallinarum, in order to record the efficiency of theiradministration in salmonellosis prophylaxis was the aim of our research. When alow infection dose, 1x107 CFU Salmonella gallinarum, was used theadministration of IgY polyclonal antibodies as immunoglobulin extract, or evenyolk administration had a protective effect against germs invasion. This effect wasnot recorded when a 10 folds higher dose was administered (1x108 CFU. Theprophylactic effect of the administration of polyclonal antibodies is demonstrated.

  8. Auroral spectrograph data annals of the international geophysical year, v.25

    CERN Document Server

    Carrigan, Anne; Norman, S J

    1964-01-01

    Annals of the International Geophysical Year, Volume 25: Auroral Spectrograph Data is a five-chapter text that contains tabulations of auroral spectrograph data. The patrol spectrograph built by the Perkin-Elmer Corporation for the Aurora and Airglow Program of the IGY is a high-speed, low-dispersion, automatic instrument designed to photograph spectra of aurora occurring along a given magnetic meridian of the sky. Data from each spectral frame were recorded on an IBM punched card. The data recorded on the cards are printed onto the tabulations in this volume. These tabulations are available

  9. Fresno Air Terminal, California. Revised Uniform Summary of Surface Weather Observations (RUSSWO). Parts A-F

    Science.gov (United States)

    1981-01-15

    PSYCHROMETRIC SUMMARY ICS S. T.) [ WET SILS TESPERATURE DEPRESSIOS (F) TOTAL I TOTAL , 0 1.2 3-, -6 7-5 𔄃-u a 10 12112 141S. 117igi.l,-20121-.12.222324 l25...VZ T ! ’Nt CA 72 -79 OCT SlAleof STATION UAXJ TEMs M" PAGL 2 18002-2000O New0ti 1.1 T WET BULB TEMPERATURE DEPRESSIO (F) TOTAL TOTAL *(F) 0 1.2 3-4 1

  10. Whistlers and audio-frequency emissions monthly summaries of whistlers and emissions for the period July 1957 - December 1958

    CERN Document Server

    Morgan, M G

    1965-01-01

    Annals of the International Geophysical Year, Volume 37: Whistlers and Audio-Frequency Emissions presents the principal results obtained in Whistlers-East synoptic program publications. Although whistlers can be observed at any time of day, it is found that they occur primarily at night. The greatest incidence of whistlers during the International Geophysical Year (IGY) period occurred in both hemispheres in the geomagnetic latitude range 50-60ʻ. The day-to-day correlation of whistler activity at geomagnetically conjugate stations was sometimes very low and sometimes remarkably high. This book

  11. Enhancement of anamnestic immunospecific antibody response in orally immunized chickens

    DEFF Research Database (Denmark)

    Mayo, Susan; Carlsson, Hans-Erik; Zagon, Andrea

    2008-01-01

    Production of immunospecific egg yolk antibodies (IgY antibodies) in egg laying hens through oral immunization is an attractive alternative to conventional antibody production in mammals for economic reasons as well as for animal welfare reasons. Oral immunization results in a systemic humoral...... of the immunization in week 18, demonstrating the presence of memory cells following the two initial oral immunizations. Considering that oral immunization results in approximately ten times lower concentrations of immunospecific antibodies in the egg yolk, compared to traditional subcutaneous immunization schemes...

  12. Outer membrane vesicles of Gallibacterium anatis induce protective immunity in egg-laying hens

    DEFF Research Database (Denmark)

    Pors, Susanne Elisabeth; Pedersen, Ida Just; Skjerning, Ragnhild Bager

    2016-01-01

    Gallibacterium anatis causes infections in the reproductive tract of egg-laying hens and induce increased mortality and decreased egg production. New prophylactic measures are needed in order to improve animal welfare and production efficiency. Bacterial outer membrane vesicles (OMVs) have...... ΔtolR mutant. Challenge was done with G. anatis 12656-12 and evaluated by scoring lesions and bacterial re-isolation rates from peritoneum. Finally, levels of OMV-specific IgY in sera were assayed by ELISA. Immunization with OMVs decreased the lesions scores significantly, while the bacterial re...

  13. Repérages bibliographiques

    Directory of Open Access Journals (Sweden)

    Sabrina Mommolin

    2013-04-01

    Full Text Available Gouvernance – Governance AYANSO A., CHATTERJEE D., CHO D. I. (2011, “E-Government readiness index: A methodology and analysis”, Government Information Quarterly, vol. 28, n° 4, pp. 522-532. CHEN Y.-C., CHU P.-Y. (2011, Electronic Governance and Cross-Boundary Collaboration: Innovations and Advancing Tools, Hershey, PA: IGI Global, 422 p. ISBN: 9781609607548 CONCHA G., ASTUDILLO H., PORRÚA M. (et al. (2011, “E-Government procurement observatory, maturity model and early measurements”, Gove...

  14. Acquisition of resistance after continuous infection with Ascaridia galli in chickens

    DEFF Research Database (Denmark)

    Ferdushy, T; Schou, T W; Norup, Liselotte Rothmann

    2014-01-01

    Acquired resistance against Ascaridia galli infection was studied in seventy-two 18-week-old white Leghorn chickens allocated to six groups (G1–G6). In order to understand the population dynamics following trickle-infection (100 eggs per chicken twice weekly), chickens of subgroups of G1 were...... with 500 eggs. G6 was left as uninfected control. Necropsy at week 10 after first inoculation revealed a lower establishment rate, an impaired development and a more posterior localization of the larvae in G4 (trickle-infected-treated-challenged) compared with G5 (treated-challenged). IgY level in serum...

  15. Anti-Pseudomonas aeruginosa IgY antibodies promote bacterial opsonization and augment the phagocytic activity of polymorphonuclear neutrophils

    DEFF Research Database (Denmark)

    Thomsen, Kim; Christophersen, Lars; Jensen, Peter Østrup

    2016-01-01

    Moderation of polymorphonuclear neutrophils (PMNs) as part of a critical defense against invading pathogens may offer a promising therapeutic approach to supplement the antibiotic eradication of Pseudomonas aeruginosa infection in non-chronically infected cystic fibrosis (CF) patients. We have...... observed that egg yolk antibodies (IgY) harvested from White leghorn chickens that target P. aeruginosa opsonize the pathogen and enhance the PMN-mediated respiratory burst and subsequent bacterial killing in vitro. The effects on PMN phagocytic activity were observed in different Pseudomonas aeruginosa...

  16. Molecular Mechanisms of Microbially Facilitated Corrosion

    Science.gov (United States)

    1986-10-23

    destierliaoice Mickels et ill. inl experimne desi 19810c, the efferts ol amipil ’ td grazi gig amiud resoutrce loirtitioli itg gas well dril Ii in i (Morrison...utilized todihlerenti- Gendrean td lakol isms contained all enlarged carliosyl hand tt 1740 cvi 1. This Lijotn co ati osY at group i nch uded Biacillus... Ihq *It mtl’ idi~ o, Chlratvi .44 Alr"It 04’/ ,t44 .4. lh ,,f,, 4 4 l,/. 4 I’m ~ .. 1igI4.ii . -hdC4itS %e.iliti, I I At% S Ir. r,- (Atc-,,n ift 215

  17. Changes in the repertoire of natural antibodies caused by immunization with bacterial antigens

    DEFF Research Database (Denmark)

    Shilova, N V; Navakouski, M J; Huflejt, M

    2011-01-01

    The repertoire of natural anti-glycan antibodies in naïve chickens and in chickens immunized with bacteria Burkholderia mallei, Burkholderia pseudomallei, and Francisella tularensis as well as with peptides from an outer membrane protein of B. pseudomallei was studied. A relatively restricted pat...... pattern of natural antibodies (first of all IgY against bacterial cell wall peptidoglycan fragments, L-Rha, and core N-acetyllactosamine) shrank and, moreover, the level of detectable antibodies decreased as a result of immunization....

  18. Social mapping for supporting sensemaking and collaboration: the case of development informatics research in South Africa

    CSIR Research Space (South Africa)

    Marais, Mario A

    2017-06-01

    Full Text Available of Research on Socio-Technical Design and Social Networking Systems. New York: IGI Global, 2009. [28] R. Heeks, “ICT4D 2.0: The next phase of applying ICT for international development,” Computer (Long. Beach. Calif)., vol. 41, no. 6, pp. 26–31, 2008. [29... issues in communications and media research. New York: Nova Science, 2005, pp. 79–94. [33] J. Murumba, E. Micheni, and A. Njuguna, “Evaluating preparedness for social networks integration into learning: A case study of Inoorero University,” in 2015 IST...

  19. 2 D gel based analysis of biological variability of the human plasma proteome

    DEFF Research Database (Denmark)

    Rentsch, Maria Louise; Jessen, Flemming

    individuals and within an individual changes will also happen over time (e.g. after meal intake). Thus, the aim of the present study was to examine the inter-individual variability of plasma protein levels in humans after meal intake. Five subjects consumed three single meals in a randomised order separated...... by one-week interval. Blood samples were drawn before the meal intake and five times during 24 hours for proteome analysis. Plasma was fractionated by use of IgY-12 spin column depleting the 12 highly abundant proteins and further processed for two-dimensional gel electrophoresis. The plasma proteome...

  20. A virtual radiation belt observatory: Looking forward to the electronic geophysical year

    Science.gov (United States)

    Baker, D. N.; Green, J. C.; Kroehl, H. W.; Kihn, E.; Virbo Team

    During the International Geophysical Year (1957-1958), member countries established many new capabilities pursuing the major IGY objectives of collecting geophysical data as widely as possible and providing free access to these data for all scientists around the globe. A key achievement of the IGY was the establishment of a worldwide system of data centers and physical observatories. The worldwide scientific community has now endorsed and is promoting an electronic Geophysical Year (eGY) initiative. The proposed eGY concept would both commemorate the 50th anniversary of the IGY in 2007-2008 and would provide a forward impetus to geophysics in the 21st century, similar to that provide by the IGY fifty years ago. The eGY concept advocates the establishment of a series of virtual geophysical observatories now being deployed in cyberspace. We are developing the concept of a Virtual Radiation Belt Observatory (ViRBO) that will bring together near-earth particle and field measurements acquired by NASA, NOAA, DoD, DOE, and other spacecraft. We discuss plans to aggregate these measurements into a readily accessible database along with analysis, visualization, and display tools that will make radiation belt information available and useful both to the scientific community and to the user community. We envision that data from the various agencies along with models being developed under the auspices of the National Science Foundation Center for Integrated Space Weather Modeling (CISM) will help us to provide an excellent `climatology' of the radiation belts over the past several decades. In particular, we would plan to use these data to drive physical models of the radiation belts to form a gridded database which would characterize particle and field properties on solar-cycle (11-year) time scales. ViRBO will also provide up-to-date specification of conditions for event analysis and anomaly resolution. We are even examining the possibilities for near-realtime acquisition of

  1. Human IgG is produced in a pro-form that requires clipping of C-terminal lysines for maximal complement activation

    DEFF Research Database (Denmark)

    van den Bremer, E. T. J.; Beurskens, F. J.; Voorhorst, M.

    2015-01-01

    Human IgG is produced with C-terminal lysines that are cleaved off in circulation. The function of this modification was unknown and generally thought not to affect antibody function. We recently reported that efficient C1q binding and complement-dependent cytotoxicity (CDC) requires IgG hexameri...

  2. Double mutation of cell wall proteins CspB and PBP1a increases secretion of the antibody Fab fragment from Corynebacterium glutamicum

    Science.gov (United States)

    2014-01-01

    Background Among other advantages, recombinant antibody-binding fragments (Fabs) hold great clinical and commercial potential, owing to their efficient tissue penetration compared to that of full-length IgGs. Although production of recombinant Fab using microbial expression systems has been reported, yields of active Fab have not been satisfactory. We recently developed the Corynebacterium glutamicum protein expression system (CORYNEX®) and demonstrated improved yield and purity for some applications, although the system has not been applied to Fab production. Results The Fab fragment of human anti-HER2 was successfully secreted by the CORYNEX® system using the conventional C. glutamicum strain YDK010, but the productivity was very low. To improve the secretion efficiency, we investigated the effects of deleting cell wall-related genes. Fab secretion was increased 5.2 times by deletion of pbp1a, encoding one of the penicillin-binding proteins (PBP1a), mediating cell wall peptidoglycan (PG) synthesis. However, this Δpbp1a mutation did not improve Fab secretion in the wild-type ATCC13869 strain. Because YDK010 carries a mutation in the cspB gene encoding a surface (S)-layer protein, we evaluated the effect of ΔcspB mutation on Fab secretion from ATCC13869. The Δpbp1a mutation showed a positive effect on Fab secretion only in combination with the ΔcspB mutation. The ΔcspBΔpbp1a double mutant showed much greater sensitivity to lysozyme than either single mutant or the wild-type strain, suggesting that these mutations reduced cell wall resistance to protein secretion. Conclusion There are at least two crucial permeability barriers to Fab secretion in the cell surface structure of C. glutamicum, the PG layer, and the S-layer. The ΔcspBΔpbp1a double mutant allows efficient Fab production using the CORYNEX® system. PMID:24731213

  3. Critical analysis of the potential for therapeutic targeting of mammalian target of rapamycin (mTOR in gastric cancer

    Directory of Open Access Journals (Sweden)

    Inokuchi M

    2014-04-01

    Full Text Available Mikito Inokuchi,1 Keiji Kato,1 Kazuyuki Kojima,2 Kenichi Sugihara1 1Department of Surgical Oncology, 2Department of Minimally Invasive Surgery, Tokyo Medical and Dental University, Tokyo, Japan Abstract: Multidisciplinary treatment including chemotherapy has become the global standard of care for patients with metastatic gastric cancer (mGC; nonetheless, survival remains poor. Although many molecular-targeted therapies have been developed for various cancers, only anti-HER2 treatment has produced promising results in patients with mGC. Mammalian target of rapamycin (mTOR plays a key role in cell proliferation, antiapoptosis, and metastasis in signaling pathways from the tyrosine kinase receptor, and its activation has been demonstrated in gastric cancer (GC cells. This review discusses the clinical relevance of mTOR in GC and examines its potential as a therapeutic target in patients with mGC. Preclinical studies in animal models suggest that suppression of the mTOR pathway inhibits the proliferation of GC cells and delays tumor progression. The mTOR inhibitor everolimus has been evaluated as second- or third-line treatment in clinical trials. Adverse events were well tolerated although the effectiveness of everolimus alone was limited. Everolimus is now being evaluated in combination with chemotherapy in Phase III clinical studies in this subgroup of patients. Two Phase III studies include exploratory biomarker research designed to evaluate the predictive value of the expression or mutation of molecules related to the Akt/mTOR signaling pathway. These biomarker studies may lead to the realization of targeted therapy for selected patients with mGC in the future. Keywords: gastric cancer, mTOR, everolimus

  4. EGFR and AKT1 overexpression are mutually exclusive and associated with a poor survival in resected gastric adenocarcinomas.

    Science.gov (United States)

    Petrini, Iacopo; Lencioni, Monica; Vasile, Enrico; Fornaro, Lorenzo; Belluomini, Lorenzo; Pasquini, Giulia; Ginocchi, Laura; Caparello, Chiara; Musettini, Gianna; Vivaldi, Caterina; Caponi, Sara; Ricci, Sergio; Proietti, Agenese; Fontanini, Gabriella; Naccarato, Antonio Giuseppe; Nardini, Vincenzo; Santi, Stefano; Falcone, Alfredo

    2018-02-14

    The evaluation of molecular targets in gastric cancer has demonstrated the predictive role of HER2 amplification for trastuzumab treatment in metastatic gastric cancer. Besides HER2, other molecular targets are under evaluation in metastatic gastric tumors. However, very little is known about their role in resected tumors. We evaluated the expression of HER2, EGFR, MET, AKT1 and phospho-mTOR in resected stage II-III adenocarcinomas. Ninety-two patients with resected stomach (63%) or gastro-esophageal adenocarcinomas (27%) were evaluated. Antibodies anti-HER2, EGFR, MET, AKT1 and phospho-mTOR were used for immunostaining of formalin-fixed paraffin-embedded slides. Using FISH, HER2 amplification was evaluated in cases with an intermediate (+2) staining. EGFR overexpression (11%) was a poor prognostic factor for overall survival (3-year OS: 47% vs 77%; Log-Rank p= 0.033). MET overexpression (36%) was associated with a trend for a worse survival (3-year OS: 65% vs 77%; Log-Rank p= 0.084). HER2 amplification/overexpression and mTOR hyper-phosphorylation were observed in 13% and 48% of tumors, respectively. AKT1 overexpression (8%) was not a prognostic factor by itself (p= 0.234). AKT1 and EGFR overexpression was mutually exclusive and patients with EGFR or AKT1 overexpression experienced a poor prognosis (3-year OS: 52% vs. 79%, Log-Rank p= 0.005). EGFR is confirmed a poor prognostic factor in resected gastric cancers. We firstly describe a mutually exclusive overexpression of EGFR and AKT1 with potential prognostic implications, suggesting the relevance of this pathway for the growth of gastric cancers.

  5. Improved Debulking of Peritoneal Tumor Implants by Near-Infrared Fluorescent Nanobody Image Guidance in an Experimental Mouse Model.

    Science.gov (United States)

    Debie, Pieterjan; Vanhoeij, Marian; Poortmans, Natalie; Puttemans, Janik; Gillis, Kris; Devoogdt, Nick; Lahoutte, Tony; Hernot, Sophie

    2017-10-31

    Debulking followed by combination chemotherapy is currently regarded as the most effective treatment for advanced ovarian cancer. Prognosis depends drastically on the degree of debulking. Accordingly, near-infrared (NIR) fluorescence imaging has been proposed to revolutionize cancer surgery by acting as a sensitive, specific, and real-time tool enabling visualization of cancer lesions. We have previously developed a NIR-labeled nanobody that allows fast, specific, and high-contrast imaging of HER2-positive tumors. In this study, we applied this tracer during fluorescence-guided surgery in a mouse model and investigated the effect on surgical efficiency. 0.5 × 10 6 SKOV3.IP1-Luc+ cells were inoculated intraperitoneally in athymic mice and were allowed to grow for 30 days. Two nanomoles of IRDye800CW-anti-HER2 nanobody was injected intravenously. After 1h30, mice were killed, randomized in two groups, and subjected to surgery. In the first animal group (n = 7), lesions were removed by a conventional surgical protocol, followed by excision of remaining fluorescent tissue using a NIR camera. The second group of mice (n = 6) underwent directly fluorescence-guided surgery. Bioluminescence imaging was performed before and after surgery. Resected tissue was categorized as visualized during conventional surgery or not, fluorescent or not, and bioluminescent positive or negative. Fluorescence imaging allowed clear visualization of tumor nodules within the abdomen, up to submillimeter-sized lesions. Fluorescence guidance resulted in significantly reduced residual tumor as compared to conventional surgery. Moreover, sensitivity increased from 59.3 to 99.0 %, and the percentage of false positive lesions detected decreased from 19.6 to 7.1 %. This study demonstrates the advantage of intraoperative fluorescence imaging using nanobody-based tracers on the efficiency of debulking surgery.

  6. Leptin/HER2 crosstalk in breast cancer: in vitro study and preliminary in vivo analysis

    International Nuclear Information System (INIS)

    Fiorio, Elena; Bonetti, Franco; Giordano, Antonio; Cetto, Gian Luigi; Surmacz, Eva; Mercanti, Anna; Terrasi, Marianna; Micciolo, Rocco; Remo, Andrea; Auriemma, Alessandra; Molino, Annamaria; Parolin, Veronica; Di Stefano, Bruno

    2008-01-01

    Obesity in postmenopausal women is associated with increased breast cancer risk, development of more aggressive tumors and resistance to certain anti-breast cancer treatments. Some of these effects might be mediated by obesity hormone leptin, acting independently or modulating other signaling pathways. Here we focused on the link between leptin and HER2. We tested if HER2 and the leptin receptor (ObR) can be coexpressed in breast cancer cell models, whether these two receptors can physically interact, and whether leptin can transactivate HER2. Next, we studied if leptin/ObR can coexist with HER2 in breast cancer tissues, and if presence of these two systems correlates with specific clinicopathological features. Expression of ObR, HER2, phospo-HER2 was assessed by immonoblotting. Physical interactions between ObR and HER2 were probed by immunoprecipitation and fluorescent immunostaining. Expression of leptin and ObR in breast cancer tissues was detected by immunohistochemistry (IHC). Associations among markers studied by IHC were evaluated using Fisher's exact test for count data. HER2 and ObR were coexpressed in all studied breast cancer cell lines. In MCF-7 cells, HER2 physically interacted with ObR and leptin treatment increased HER2 phosphorylation on Tyr 1248. In 59 breast cancers, the presence of leptin was correlated with ObR (the overall association was about 93%). This result was confirmed both in HER2-positive and in HER2-negative subgroups. The expression of leptin or ObR was numerically more frequent in larger (> 10 mm) tumors. Coexpression of HER2 and the leptin/ObR system might contribute to enhanced HER2 activity and reduced sensitivity to anti-HER2 treatments

  7. Native mass spectrometry and ion mobility characterization of trastuzumab emtansine, a lysine-linked antibody drug conjugate.

    Science.gov (United States)

    Marcoux, Julien; Champion, Thierry; Colas, Olivier; Wagner-Rousset, Elsa; Corvaïa, Nathalie; Van Dorsselaer, Alain; Beck, Alain; Cianférani, Sarah

    2015-08-01

    Antibody-drug conjugates (ADCs) are biochemotherapeutics consisting of a cytotoxic chemical drug linked covalently to a monoclonal antibody. Two main classes of ADCs, namely cysteine and lysine conjugates, are currently available on the market or involved in clinical trials. The complex structure and heterogeneity of ADCs makes their biophysical characterization challenging. For cysteine conjugates, hydrophobic interaction chromatography is the gold standard technique for studying drug distribution, the naked antibody content, and the average drug to antibody ratio (DAR). For lysine ADC conjugates on the other hand, which are not amenable to hydrophobic interaction chromatography because of their higher heterogeneity, denaturing mass spectrometry (MS) and UV/Vis spectroscopy are the most powerful approaches. We report here the use of native MS and ion mobility (IM-MS) for the characterization of trastuzumab emtansine (T-DM1, Kadcyla(®)). This lysine conjugate is currently being considered for the treatment of human epidermal growth factor receptor 2 (HER2)-positive breast cancer, and combines the anti-HER2 antibody trastuzumab (Herceptin(®)), with the cytotoxic microtubule-inhibiting maytansine derivative, DM1. We show that native MS combined with high-resolution measurements and/or charge reduction is beneficial in terms of the accurate values it provides of the average DAR and the drug load profiles. The use of spectral deconvolution is discussed in detail. We report furthermore the use of native IM-MS to directly determine DAR distribution profiles and average DAR values, as well as a molecular modeling investigation of positional isomers in T-DM1. © 2015 The Protein Society.

  8. Co-Expansion of Cytokine-Induced Killer Cells and Vγ9Vδ2 T Cells for CAR T-Cell Therapy.

    Directory of Open Access Journals (Sweden)

    Shou-Hui Du

    Full Text Available Gamma delta (γδ T cells and cytokine-induced killer (CIK cells, which are a heterogeneous population of T lymphocytes and natural killer T (NKT cells, have been separately expanded ex vivo and shown to be capable of targeting and mediating cytotoxicity against various tumor cells in a major histocompatibility complex-unrestricted manner. However, the co-expansion and co-administration of these immune cells have not been explored. In this study we describe an efficient method to expand simultaneously both CIK and Vγ9Vδ2 T cells, termed as CIKZ cells, from human peripheral blood mononuclear cells (PBMCs using Zometa, interferon-gamma (IFN-γ, interleukin 2 (IL-2, anti-CD3 antibody and engineered K562 feeder cells expressing CD64, CD137L and CD86. A 21-day culture of PBMCs with this method yielded nearly 20,000-fold expansion of CIKZ cells with γδ T cells making up over 20% of the expanded population. The expanded CIKZ cells exhibited antitumor cytotoxicity and could be modified to express anti-CD19 chimeric antigen receptor (CAR, anti-CEA CAR, and anti-HER2 CAR to enhance their specificity and cytotoxicity against CD19-, CEA-, or HER2-positive tumor cells. The tumor inhibitory activity of anti-CD19 CAR-modified CIKZ cells was further demonstrated in vivo in a Raji tumor mouse model. The findings herein substantiate the feasibility of co-expanding CIK and γδ cells for adoptive cellular immunotherapy applications such as CAR T-cell therapy against cancer.

  9. Preparation and characterization of micro-cell membrane chromatographic column with N-hydroxysuccinimide group-modified silica-based porous layer open tubular capillary.

    Science.gov (United States)

    Xu, Liang; Xu, Bei; Zhao, Zhi-Yu; Yang, Hui-Ping; Tang, Cheng; Dong, Lin-Yi; Liu, Kun; Fu, Li; Wang, Xian-Hua

    2017-09-22

    Cell membrane chromatography (CMC) is an effective tool in screening active compounds from natural products and studying membrane protein interactions. Nevertheless, it always consumes a large amount of cells (e.g. 10 7 -10 8 ) for column preparation. To overcome this, micro-CMC (mCMC), that employs a silica capillary as membrane carrier, was developed. However, both CMC and mCMC suffer from short column life span (e.g. 3days), mainly due to the falling-off of cellular membranes (CMs). This has greatly limited further application of CMC and mCMC, especially when the cells are hard to obtain. To solve this, N-hydroxysuccinimide (NHS)-modified silica-based porous layer open tubular capillary was first prepared for mCMC. The NHS groups can easily react with amino groups on CMs to form a stable covalent bond under a mild condition. So, CMs immobilized on the NHS-modified capillary are less likely to fall off. To verify this, SKBR3/mCMC (Her2 positive) and BALL1/mCMC (CD20 positive) columns were prepared. Two monoclonal antibody drugs, trastuzumab (anti-Her2) and rituximab (anti-CD20), were selected as analytes to characterize the columns. As a result, NHS-modified column for mCMC can afford higher chromatographic retention than non-modified column. Besides, the column life span was significantly improved to more than 16days for SKBR3/mCMC and 14days for BALL1/mCMC, while the compared column showed a sharp decline in retention factor in first 3days. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. CD4+ and CD8+ T cells can act separately in tumour rejection after immunization with murine pneumotropic virus chimeric Her2/neu virus-like particles.

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    Kalle Andreasson

    Full Text Available BACKGROUND: Immunization with murine pneumotropic virus virus-like particles carrying Her2/neu (Her2MPtVLPs prevents tumour outgrowth in mice when given prophylactically, and therapeutically if combined with the adjuvant CpG. We investigated which components of the immune system are involved in tumour rejection, and whether long-term immunological memory can be obtained. METHODOLOGY AND RESULTS: During the effector phase in BALB/c mice, only depletion of CD4+ and CD8+ in combination, with or without NK cells, completely abrogated tumour protection. Depletion of single CD4+, CD8+ or NK cell populations only had minor effects. During the immunization/induction phase, combined depletion of CD4+ and CD8+ cells abolished protection, while depletion of each individual subset had no or negligible effect. When tumour rejection was studied in knock-out mice with a C57Bl/6 background, protection was lost in CD4-/-CD8-/- and CD4-/-, but not in CD8-/- mice. In contrast, when normal C57Bl/6 mice were depleted of different cell types, protection was lost irrespective of whether only CD4+, only CD8+, or CD4+ and CD8+ cells in combination were eradicated. No anti-Her2/neu antibodies were detected but a Her2/neu-specific IFNgamma response was seen. Studies of long-term memory showed that BALB/c mice could be protected against tumour development when immunized together with CpG as long as ten weeks before challenge. CONCLUSION: Her2MPtVLP immunization is efficient in stimulating several compartments of the immune system, and induces an efficient immune response including long-term memory. In addition, when depleting mice of isolated cellular compartments, tumour protection is not as efficiently abolished as when depleting several immune compartments together.

  11. Her-2 neu (Cerb-B2) expression in fine needle aspiration samples of breast carcinoma: A pilot study comparing FISH, CISH and immunocytochemistry.

    Science.gov (United States)

    Kapila, Kusum; Al-Awadhi, S; Francis, Im

    2011-04-01

    Breast cancers with Her-2 neu gene amplification are recognized as important markers for aggressive disease and targets which respond to therapy with trastuzumab. Her-2 neu testing on histological sections is routinely performed to select patients who may benefit from anti- Her-2 neu therapy. Few reports are available which document Her-2 neu status on fine needle aspirates (FNA). This pilot study is to document expression of Her-2 neu (Cerb-B2) on cytospin smears from FNA of patients with breast carcinoma. Twenty samples of FNA already collected for diagnostic purposes from patients with primary breast carcinoma were studied for demonstration of Her-2 neu expression by immunohistochemistry (IHC), Fluorescent in-situ hybridization (FISH) and chromogenic in-situ hybridization (CISH) on cytospin smears from FNA. Their expression was compared with tissue sections where possible. Good correlation was observed between Her-2 neu protein expression and gene amplification in cytospin smears. Three of five (60%) breast carcinomas cases with 2+ and 3+ staining on IHC showed gene amplification by FISH and CISH. Three of 7 (43%) and 5 of 7 (71%) cases negative/1+ staining on IHC did not show gene amplification by FISH and CISH respectively. Tissue sections from 10 cases with 2+ and 3+ staining for Her2neu by IHC showed gene amplification in 8 cases. Demonstration of Her-2 neu by IHC, FISH or CISH in FNA is possible and may play a role in the management of patients with advanced breast cancer or those cases where surgical resection is not advisable.

  12. Her-2 neu (Cerb-B2 expression in fine needle aspiration samples of breast carcinoma: A pilot study comparing FISH, CISH and immunocytochemistry

    Directory of Open Access Journals (Sweden)

    Kusum Kapila

    2011-01-01

    Full Text Available Background: Breast cancers with Her-2 neu gene amplification are recognized as important markers for aggressive disease and targets which respond to therapy with trastuzumab. Her-2 neu testing on histological sections is routinely performed to select patients who may benefit from anti- Her-2 neu therapy. Few reports are available which document Her-2 neu status on fine needle aspirates (FNA. Aim: This pilot study is to document expression of Her-2 neu (Cerb-B2 on cytospin smears from FNA of patients with breast carcinoma. Materials and Methods: Twenty samples of FNA already collected for diagnostic purposes from patients with primary breast carcinoma were studied for demonstration of Her-2 neu expression by immunohistochemistry (IHC, Fluorescent in-situ hybridization (FISH and chromogenic in-situ hybridization (CISH on cytospin smears from FNA. Their expression was compared with tissue sections where possible. Results: Good correlation was observed between Her-2 neu protein expression and gene amplification in cytospin smears. Three of five (60% breast carcinomas cases with 2+ and 3+ staining on IHC showed gene amplification by FISH and CISH. Three of 7 (43% and 5 of 7 (71% cases negative/1+ staining on IHC did not show gene amplification by FISH and CISH respectively. Tissue sections from 10 cases with 2+ and 3+ staining for Her2neu by IHC showed gene amplification in 8 cases. Conclusion: Demonstration of Her-2 neu by IHC, FISH or CISH in FNA is possible and may play a role in the management of patients with advanced breast cancer or those cases where surgical resection is not advisable.

  13. Antitumor activity of the multikinase inhibitor regorafenib in patient-derived xenograft models of gastric cancer.

    Science.gov (United States)

    Huynh, Hung; Ong, Richard; Zopf, Dieter

    2015-10-29

    Unresectable gastric cancer is associated with poor outcomes, with few treatment options available after failure of cytotoxic chemotherapy. Clinical trials of targeted therapies have generally shown no survival benefit in gastric cancer, with the exceptions of the antibodies ramucirumab (anti-VEGFR2) and trastuzumab (anti-HER2/neu). Given the efficacy of the multikinase inhibitor regorafenib in other gastrointestinal tumors, we investigated its potential in gastric cancer. The antitumor activity of oral regorafenib was assessed in eight murine patient-derived gastric cancer xenograft models. Dose-response experiments assessed the efficacy and tolerability of oral regorafenib 5, 10, and 15 mg/kg/day in two models, with 10 mg/kg/day selected for further investigation in all eight models. Tumor weight and volume was monitored during treatment; tumor cell proliferation, angiogenesis, apoptosis, and intracellular signaling were assessed using immunohistochemistry and Western blotting of total tumor lysates at the end of treatment. Regorafenib showed dose-dependent inhibition of tumor growth and was well tolerated, with no significant decreases in bodyweight or evident toxicity. Regorafenib 10 mg/kg/day significantly inhibited tumor growth in all eight models (72 to 96 %; all p Regorafenib reduced tumor angiogenesis 3- to 11-fold versus controls in all models (all p Regorafenib was effective in patient-derived models of gastric cancer of different histological subtypes, with inhibition of tumor growth, angiogenesis, and tumor-cell proliferation observed in almost all models. These findings are consistent with the observed activity of regorafenib in preclinical models of other gastrointestinal tumors, and support further clinical investigation in gastric cancer.

  14. HER2 and β-catenin protein location: importance in the prognosis of breast cancer patients and their correlation when breast cancer cells suffer stressful situations.

    Science.gov (United States)

    Cuello-Carrión, F Darío; Shortrede, Jorge E; Alvarez-Olmedo, Daiana; Cayado-Gutiérrez, Niubys; Castro, Gisela N; Zoppino, Felipe C M; Guerrero, Martín; Martinis, Estefania; Wuilloud, Rodolfo; Gómez, Nidia N; Biaggio, Verónica; Orozco, Javier; Gago, Francisco E; Ciocca, Leonardo A; Fanelli, Mariel A; Ciocca, Daniel R

    2015-02-01

    In human breast cancer, β-catenin localization has been related with disease prognosis. Since HER2-positive patients are an important subgroup, and that in breast cancer cells a direct interaction of β-catenin/HER2 has been reported, in the present study we have explored whether β-catenin location is related with the disease survival. The study was performed in a tumor bank from patients (n = 140) that did not receive specific anti-HER2 therapy. The proteins were detected by immunohistochemistry in serial sections, 47 (33.5%) patients were HER2-positive with a long follow-up. HER2-positive patients that displayed β-catenin at the plasma membrane (completely surrounding the tumour cells) showed a significant better disease-free survival and overall survival than the patients showing the protein on other locations. Then we explored the dynamics of the co-expression of β-catenin and HER2 in human MCF-7 and SKBR3 cells exposed to different stressful situations. In untreated conditions MCF-7 and SKBR3 cells showed very different β-catenin localization. In MCF-7 cells, cadmium administration caused a striking change in β-catenin localization driving it from plasma membrane to cytoplasmic and perinuclear areas and HER2 showed a similar localization patterns. The changes induced by cadmium were compared with heat shock, H2O2 and tamoxifen treatments. In conclusion, this study shows the dynamical associations of HER2 and β-catenin and their changes in subcellular localizations driven by stressful situations. In addition, we report for the first time the correlation between plasma membrane associated β-catenin in HER2-positive breast cancer and survival outcome, and the importance of the protein localization in breast cancer samples.

  15. A Molecularly Targeted Theranostic Probe for Ovarian Cancer

    Science.gov (United States)

    Chen, Wenxue; Bardhan, Rizia; Bartels, Marc; Perez-Torres, Carlos; Pautler, Robia G.; Halas, Naomi J.; Joshi, Amit

    2014-01-01

    Overexpression of the human epidermal growth factor receptor (HER) family has been implicated in ovarian cancer because of its participation in signaling pathway regulating cellular proliferation, differentiation, motility, and survival. Currently, effective diagnostic and therapeutic schemes are lacking for treating ovarian cancer and consequently ovarian cancer has a high mortality rate. While HER2 receptor expression does not usually affect the survival rates of ovarian cancer to the same extent as in breast cancer, it can be employed as a docking site for directed nanotherapies in cases with de novo or acquired chemotherapy resistance. In this study, we have exploited a novel gold nanoshell-based complex (nanocomplex) for targeting, dual modal imaging, and photothermal therapy of HER2 overexpressing and drug resistant ovarian cancer OVCAR3 cells in vitro. The nanocomplexes are engineered to simultaneously provide contrast as fluorescence optical imaging probe and a magnetic resonance imaging (MRI) agent. Both immunofluorescence staining and MRI successfully demonstrate that nanocomplex-anti-HER2 conjugates specifically bind to OVCAR3 cells as opposed to the control, MDA-MB-231 cells, which have low HER2 expression. In addition, nanocomplexes targeted to OVCAR3 cells, when irradiated with near infrared (NIR) laser result in selective destruction of cancer cells through photothermal ablation. We also demonstrate that NIR light therapy and the nanocomplexes by themselves are non-cytotoxic in vitro. To the best of our knowledge, this is the first demonstration of a successful integration of dual modal bioimaging with photothermal cancer therapy for treatment of ovarian cancer. Based on their efficacy in vitro, these nanocomplexes are highly promising for image guided photo-thermal therapy of ovarian cancer as well as other HER2 overexpressing cancers. PMID:20371708

  16. Evaluation of Antigen-Conjugated Fluorescent Beads to Identify Antigen-Specific B Cells.

    Science.gov (United States)

    Correa, Isabel; Ilieva, Kristina M; Crescioli, Silvia; Lombardi, Sara; Figini, Mariangela; Cheung, Anthony; Spicer, James F; Tutt, Andrew N J; Nestle, Frank O; Karagiannis, Panagiotis; Lacy, Katie E; Karagiannis, Sophia N

    2018-01-01

    Selection of single antigen-specific B cells to identify their expressed antibodies is of considerable interest for evaluating human immune responses. Here, we present a method to identify single antibody-expressing cells using antigen-conjugated fluorescent beads. To establish this, we selected Folate Receptor alpha (FRα) as a model antigen and a mouse B cell line, expressing both the soluble and the membrane-bound forms of a human/mouse chimeric antibody (MOv18 IgG1) specific for FRα, as test antibody-expressing cells. Beads were conjugated to FRα using streptavidin/avidin-biotin bridges and used to select single cells expressing the membrane-bound form of anti-FRα. Bead-bound cells were single cell-sorted and processed for single cell RNA retrotranscription and PCR to isolate antibody heavy and light chain variable regions. Variable regions were then cloned and expressed as human IgG1/k antibodies. Like the original clone, engineered antibodies from single cells recognized native FRα. To evaluate whether antigen-coated beads could identify specific antibody-expressing cells in mixed immune cell populations, human peripheral blood mononuclear cells (PBMCs) were spiked with test antibody-expressing cells. Antigen-specific cells could comprise up to 75% of cells selected with antigen-conjugated beads when the frequency of the antigen-positive cells was 1:100 or higher. In PBMC pools, beads conjugated to recombinant antigens FRα and HER2 bound antigen-specific anti-FRα MOv18 and anti-HER2 Trastuzumab antibody-expressing cells, respectively. From melanoma patient-derived B cells selected with melanoma cell line-derived protein-coated fluorescent beads, we generated a monoclonal antibody that recognized melanoma antigen-coated beads. This approach may be further developed to facilitate analysis of B cells and their antibody profiles at the single cell level and to help unravel humoral immune repertoires.

  17. Photothermal effects of multi-walled carbon nanotubes on the viability of BT-474 cancer cells

    International Nuclear Information System (INIS)

    Chou, Hung-Tao; Wang, Tsung-Pao; Lee, Chi-Young; Tai, Nyan-Hwa; Chang, Hwan-You

    2013-01-01

    Functionalized multi-walled carbon nanotubes (f-MWCNTs) were conjugated to an antibody of BT-474 cancer cells (f-MWCNTs-ab), and the photothermal effect of the f-MWCNTs-ab for BT-474 cancer cell destruction was demonstrated. After near-infrared irradiation, the f-MWCNTs-ab were more capable of killing cancer cells and possessed higher cell specificity than f-MWCNTs. Quantitative results showed that the viability of the cancer cells was affected by the concentration of the f-MWCNTs-ab solution, irradiation time, and settling time after irradiation. The membrane impermeable fluorescence dye ethidium bromide was used to detect cell viability after near-infrared irradiation, and the results agreed with those obtained from the Alamar Blue cell viability assay. The EtBr fluorescence results suggest that the cell membrane, attached to f-MWCNTs-ab, was damaged after irradiation, which led to cell death and necrosis. Using confocal microscopy, a few f-MWCNTs-ab were detected in the cell, indicating the endocytosis effect. The results not only explain the improved efficiency of thermotherapy but also indicate that necrosis may result from protein denaturation attributing to the heated f-MWCNTs-ab in the cell. Highlights: ► f-MWCNTs conjugated with anti-HER2 antibody by chemical method. ► Kill breast cancer cells by using low dose f-MWCNTs-ab due to photothermal effect. ► Use EtBr fluorescent to prove that the cell membrane was broken by heated f-MWCNTs. ► Few f-MWCNTs-ab were detected in the cell indicating the endocytosis effect. ► Necrosis may result from protein denaturation due to contact with the heated CNTs.

  18. Tumor-Targeting Salmonella typhimurium A1-R in Combination with Trastuzumab Eradicates HER-2-Positive Cervical Cancer Cells in Patient-Derived Mouse Models.

    Directory of Open Access Journals (Sweden)

    Yukihiko Hiroshima

    Full Text Available We have previously developed mouse models of HER-2-positive cervical cancer. Tumors in nude mice had histological structures similar to the original tumor and were stained by anti-HER-2 antibody in the same pattern as the patient's cancer. We have also previously developed tumor-targeting Salmonella typhimurium A1-R and have demonstrated its efficacy against patient-derived tumor mouse models, both alone and in combination. In the current study, we determined the efficacy of S. typhimurium A1-R in combination with trastuzumab on a patient-cancer nude-mouse model of HER-2 positive cervical cancer. Mice were randomized to 5 groups and treated as follows: (1 no treatment; (2 carboplatinum (30 mg/kg, ip, weekly, 5 weeks; (3 trastuzumab (20 mg/kg, ip, weekly, 5 weeks; (4 S. typhimurium A1-R (5 × 107 CFU/body, ip, weekly, 5 weeks; (5 S. typhimurium A1-R (5 × 107 CFU/body, ip, weekly, 5 weeks + trastuzumab (20 mg/kg, ip, weekly, 5 weeks. All regimens had significant efficacy compared to the untreated mice. The relative tumor volume of S. typhimurium A1-R + trastuzumab-treated mice was smaller compared to trastuzumab alone (p = 0.007 and S. typhimurium A1-R alone (p = 0.039. No significant body weight loss was found compared to the no treatment group except for carboplatinum-treated mice (p = 0.021. Upon histological examination, viable tumor cells were not detected, and replaced by stromal cells in the tumors treated with S. typhimurium A1-R + trastuzumab. The results of the present study suggest that S. typhimurium A1-R and trastuzumab in combination are highly effective against HER-2-expressing cervical cancer.

  19. Evaluating the potential of {sup 188}Re-SOCTA-trastuzumab as a new radioimmunoagent for breast cancer treatment

    Energy Technology Data Exchange (ETDEWEB)

    Luo, T.-Y. [Isotope Application Division, Institute of Nuclear Energy Research, P.O. Box 3-27, Longtan, Taoyuan 325, Taiwan (China)], E-mail: tylo@iner.gov.tw; Tang, I-C.; Wu, Y.-L.; Hsu, K.-L. [Isotope Application Division, Institute of Nuclear Energy Research, P.O. Box 3-27, Longtan, Taoyuan 325, Taiwan (China); Liu, S.-W. [Chemistry Division, Institute of Nuclear Energy Research, Taoyuan 325, Taiwan (China); Kung, H.-C. [Department of Electrical Engineering, Tung Nan University, Taipei 222, Taiwan (China); Lai, P.-S. [Department of Chemistry, National Chung Hsing University, Taichung 402, Taiwan (China); Lin, W.-J. [Isotope Application Division, Institute of Nuclear Energy Research, P.O. Box 3-27, Longtan, Taoyuan 325, Taiwan (China)

    2009-01-15

    Introduction: Radioimmunotherapy, which utilizes monoclonal antibodies and therapeutic radioisotopes against antigen-expressing tumor tissues, is an attractive therapeutic approach for cancer therapy. Trastuzumab (Herceptin) is a humanized anti-HER-2/neu monoclonal antibody for breast cancer treatment. In this paper, we introduce a new radioimmunoagent, {sup 188}Re-trastuzumab, via a bifunctional ligand, succinimidyl 3,6-diaza-5-oxo-3-[2-((triphenylmethyl)thio)ethyl] -8-[(triphenylmethyl)thio]octanoate (SOCTA), and evaluate its potential to be a therapeutic radiopharmaceutical for breast cancer treatment. Methods: Equimolar amounts of SOCTA and trastuzumab were selected to react, and the conjugation ratio of SOCTA-trastuzumab was evaluated by the MALDI-TOF method. The immunoreactivity of SOCTA-trastuzumab was compared with nonconjugated trastuzumab in HER-2/neu overexpressing human breast cancer cell BT-474. Biodistribution experiment and microSPECT/CT images of {sup 188}Re-SOCTA-trastuzumab being administered intravenously to SCID mice bearing xenografted BT-474 breast cancer were investigated to evaluate the tumor-targeting capability. Results: The covalent attachment of SOCTA to trastuzumab (at 1:1 molar ratio) resulted in the averaged conjugation ratio of 0.27{+-}0.06 (n=3). The complex could easily be labeled with {sup 188}Re and achieve 95% radiochemical purity (RCP) after 1 h of reaction at room temperature. The in vitro stability study also revealed that the RCP of {sup 188}Re-SOCTA-trastuzumab was at a value of more than 85% after 48 h of incubation with human serum. The immunoreactivity evaluation showed that SOCTA-trastuzumab and nonconjugated trastuzumab had similar binding capacity (B{sub max}) to HER-2/neu receptor in BT-474 cells. The animal experiments showed that {sup 188}Re-SOCTA-trastuzumab accumulated more intensively in the tumor site as compared to normal tissue. Conclusion: We suggest that {sup 188}Re-SOCTA-trastuzumab could be a potential

  20. SU-E-I-81: Targeting of HER2-Expressing Tumors with Dual PET-MR Imaging Probes

    Energy Technology Data Exchange (ETDEWEB)

    Xu, P; Peng, Y; Sun, M; Yang, X [Suzhou Institute of Biomedical Engineering and Technology Chinese Academy o, Suzhou, Jiangsu (China)

    2015-06-15

    Purpose: The detection of human epidermal growth factor receptor type 2 (HER2) expression in malignant tumors provides important information influencing patient management. Radionuclide in vivo imaging of HER2 may permit the detection of HER2 in both primary tumors and metastases by a single noninvasive procedure. Trastuzumab, effective in about 15 % of women with breast cancer, downregulates signalling through the Akt/PI3K and MAPK pathways.These pathways modulate metabolism which can be monitored by positron emission tomography (PET) and magnetic resonance imaging (MRI). Methods: The relationship between response of HER2 overexpressing tumours and changes in imaging PET or SPECT and MRI will be examined by a integrated bimodal imaging probe.Small (7 kDa) high-affinity anti-HER2 Affibody molecules and KCCYSL targeting peptide may be suitable tracers for visualization of HER2-expressing tumors. Peptide-conjugated iron oxide nanoparticles (Fe3O4 NPs) as MRI imaging and CB-TE2A as PET imaging are integrated into a single synthetic molecule in the HER2 positive cancer. Results: One of targeted contrast bimodal imaging probe agents was synthesized and evaluated to target HER2-expressing tumors in a HER2 positive rat model. We will report the newest results regarding the development of bimodal imaging probes. Conclusion: The preliminary results of the bimodal imaging probe presents high correlation of MRI signal and PET imaging intensity in vivo. This unique feature can hardly be obtained by single model contrast agents. It is envisioned that this bimodal agents can hold great potential for accurate detection of HER2-expressing tumors which are critical for clinical management of the disease.

  1. Targeted Therapy for Breast Cancer Prevention

    Science.gov (United States)

    den Hollander, Petra; Savage, Michelle I.; Brown, Powel H.

    2013-01-01

    With a better understanding of the etiology of breast cancer, molecularly targeted drugs have been developed and are being testing for the treatment and prevention of breast cancer. Targeted drugs that inhibit the estrogen receptor (ER) or estrogen-activated pathways include the selective ER modulators (tamoxifen, raloxifene, and lasofoxifene) and aromatase inhibitors (AIs) (anastrozole, letrozole, and exemestane) have been tested in preclinical and clinical studies. Tamoxifen and raloxifene have been shown to reduce the risk of breast cancer and promising results of AIs in breast cancer trials, suggest that AIs might be even more effective in the prevention of ER-positive breast cancer. However, these agents only prevent ER-positive breast cancer. Therefore, current research is focused on identifying preventive therapies for other forms of breast cancer such as human epidermal growth factor receptor 2 (HER2)-positive and triple-negative breast cancer (TNBC, breast cancer that does express ER, progesterone receptor, or HER2). HER2-positive breast cancers are currently treated with anti-HER2 therapies including trastuzumab and lapatinib, and preclinical and clinical studies are now being conducted to test these drugs for the prevention of HER2-positive breast cancers. Several promising agents currently being tested in cancer prevention trials for the prevention of TNBC include poly(ADP-ribose) polymerase inhibitors, vitamin D, and rexinoids, both of which activate nuclear hormone receptors (the vitamin D and retinoid X receptors). This review discusses currently used breast cancer preventive drugs, and describes the progress of research striving to identify and develop more effective preventive agents for all forms of breast cancer. PMID:24069582

  2. A novel antibody engineering strategy for making monovalent bispecific heterodimeric IgG antibodies by electrostatic steering mechanism.

    Science.gov (United States)

    Liu, Zhi; Leng, Esther C; Gunasekaran, Kannan; Pentony, Martin; Shen, Min; Howard, Monique; Stoops, Janelle; Manchulenko, Kathy; Razinkov, Vladimir; Liu, Hua; Fanslow, William; Hu, Zhonghua; Sun, Nancy; Hasegawa, Haruki; Clark, Rutilio; Foltz, Ian N; Yan, Wei

    2015-03-20

    Producing pure and well behaved bispecific antibodies (bsAbs) on a large scale for preclinical and clinical testing is a challenging task. Here, we describe a new strategy for making monovalent bispecific heterodimeric IgG antibodies in mammalian cells. We applied an electrostatic steering mechanism to engineer antibody light chain-heavy chain (LC-HC) interface residues in such a way that each LC strongly favors its cognate HC when two different HCs and two different LCs are co-expressed in the same cell to assemble a functional bispecific antibody. We produced heterodimeric IgGs from transiently and stably transfected mammalian cells. The engineered heterodimeric IgG molecules maintain the overall IgG structure with correct LC-HC pairings, bind to two different antigens with comparable affinity when compared with their parental antibodies, and retain the functionality of parental antibodies in biological assays. In addition, the bispecific heterodimeric IgG derived from anti-HER2 and anti-EGF receptor (EGFR) antibody was shown to induce a higher level of receptor internalization than the combination of two parental antibodies. Mouse xenograft BxPC-3, Panc-1, and Calu-3 human tumor models showed that the heterodimeric IgGs strongly inhibited tumor growth. The described approach can be used to generate tools from two pre-existent antibodies and explore the potential of bispecific antibodies. The asymmetrically engineered Fc variants for antibody-dependent cellular cytotoxicity enhancement could be embedded in monovalent bispecific heterodimeric IgG to make best-in-class therapeutic antibodies. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. Antitumor activity of pan-HER inhibitors in HER2-positive gastric cancer.

    Science.gov (United States)

    Yoshioka, Takahiro; Shien, Kazuhiko; Namba, Kei; Torigoe, Hidejiro; Sato, Hiroki; Tomida, Shuta; Yamamoto, Hiromasa; Asano, Hiroaki; Soh, Junichi; Tsukuda, Kazunori; Nagasaka, Takeshi; Fujiwara, Toshiyoshi; Toyooka, Shinichi

    2018-04-01

    Molecularly targeted therapy has enabled outstanding advances in cancer treatment. Whereas various anti-human epidermal growth factor receptor 2 (HER2) drugs have been developed, trastuzumab is still the only anti-HER2 drug presently available for gastric cancer. In this study, we propose novel treatment options for patients with HER2-positive gastric cancer. First, we determined the molecular profiles of 12 gastric cancer cell lines, and examined the antitumor effect of the pan-HER inhibitors afatinib and neratinib in those cell lines. Additionally, we analyzed HER2 alteration in 123 primary gastric cancers resected from Japanese patients to clarify possible candidates with the potential to respond to these drugs. In the drug sensitivity analysis, both afatinib and neratinib produced an antitumor effect in most of the HER2-amplified cell lines. However, some cells were not sensitive to the drugs. When the molecular profiles of the cells were compared based on the drug sensitivities, we found that cancer cells with lower mRNA expression levels of IGFBP7, a tumor suppressor gene that inhibits the activation of insulin-like growth factor-1 receptor (IGF-1R), were less sensitive to pan-HER inhibitors. A combination therapy consisting of pan-HER inhibitors and an IGF-1R inhibitor, picropodophyllin, showed a notable synergistic effect. Among 123 clinical samples, we found 19 cases of HER2 amplification and three cases of oncogenic mutations. In conclusion, afatinib and neratinib are promising therapeutic options for the treatment of HER2-amplified gastric cancer. In addition to HER2 amplification, IGFBP7 might be a biomarker of sensitivity to these drugs, and IGF-1R-targeting therapy can overcome drug insensitiveness in HER2-amplified gastric cancer. © 2018 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

  4. Neratinib overcomes trastuzumab resistance in HER2 amplified breast cancer.

    Science.gov (United States)

    Canonici, Alexandra; Gijsen, Merel; Mullooly, Maeve; Bennett, Ruth; Bouguern, Noujoude; Pedersen, Kasper; O'Brien, Neil A; Roxanis, Ioannis; Li, Ji-Liang; Bridge, Esther; Finn, Richard; Siamon, Dennis; McGowan, Patricia; Duffy, Michael J; O'Donovan, Norma; Crown, John; Kong, Anthony

    2013-10-01

    Trastuzumab has been shown to improve the survival outcomes of HER2 positive breast cancer patients. However, a significant proportion of HER2-positive patients are either inherently resistant or develop resistance to trastuzumab. We assessed the effects of neratinib, an irreversible panHER inhibitor, in a panel of 36 breast cancer cell lines. We further assessed its effects with or without trastuzumab in several sensitive and resistant breast cancer cells as well as a BT474 xenograft model. We confirmed that neratinib was significantly more active in HER2-amplified than HER2 non-amplified cell lines. Neratinib decreased the activation of the 4 HER receptors and inhibited downstream pathways. However, HER3 and Akt were reactivated at 24 hours, which was prevented by the combination of trastuzumab and neratinib. Neratinib also decreased pHER2 and pHER3 in acquired trastuzumab resistant cells. Neratinib in combination with trastuzumab had a greater growth inhibitory effect than either drug alone in 4 HER2 positive cell lines. Furthermore, trastuzumab in combination with neratinib was growth inhibitory in SKBR3 and BT474 cells which had acquired resistance to trastuzumab as well as in a BT474 xenograft model. Innately trastuzumab resistant cell lines showed sensitivity to neratinib, but the combination did not enhance response compared to neratinib alone. Levels of HER2 and phospho-HER2 showed a direct correlation with sensitivity to neratinib. Our data indicate that neratinib is an effective anti-HER2 therapy and counteracted both innate and acquired trastuzumab resistance in HER2 positive breast cancer. Our results suggest that combined treatment with trastuzumab and neratinib is likely to be more effective than either treatment alone for both trastuzumab-sensitive breast cancer as well as HER2-positive tumors with acquired resistance to trastuzumab.

  5. In vitro and in vivo studies of the combination of IGF1R inhibitor figitumumab (CP-751,871) with HER2 inhibitors trastuzumab and neratinib.

    Science.gov (United States)

    Chakraborty, Ashok K; Zerillo, Cynthia; DiGiovanna, Michael P

    2015-08-01

    The insulin-like growth factor I receptor (IGF1R) has been linked to resistance to HER2-directed therapy with trastuzumab (Herceptin). We examined the anti-tumor activity of figitumumab (CP-751,871), a human monoclonal antibody that blocks IGF1R ligand binding, alone and in combination with the therapeutic anti-HER2 antibody trastuzumab and the pan-HER family tyrosine kinase inhibitor neratinib, using in vitro and in vivo breast cancer model systems. In vitro assays of proliferation, apoptosis, and signaling, and in vivo anti-tumor experiments were conducted in HER2-overexpressing (BT474) and HER2-normal (MCF7) models. We find single-agent activity of the HER2-targeting drugs but not figitumumab in the BT474 model, while the reverse is true in the MCF7 model. However, in both models, combining figitumumab with HER2-targeting drugs shows synergistic anti-proliferative and apoptosis-inducing effects, and optimum inhibition of downstream signaling. In murine xenograft models, synergistic anti-tumor effects were observed in the HER2-normal MCF7 model for the combination of figitumumab with trastuzumab, and, in the HER2-overexpressing BT474 model, enhanced anti-tumor effects were observed for the combination of figitumumab with either trastuzumab or neratinib. Analysis of tumor extracts from the in vivo experiments showed evidence of the most optimal inhibition of downstream signaling for the drug combinations over the single-agent therapies. These results suggest promise for such combinations in treating patients with breast cancer, and that, unlike the case for single-agent therapy, the therapeutic effects of such combinations may be independent of expression levels of the individual receptors or the single-agent activity profile.

  6. Bispecific antibody complex pre-targeting and targeted delivery of polymer drug conjugates for imaging and therapy in dual human mammary cancer xenografts. Targeted polymer drug conjugates for cancer diagnosis and therapy

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    Khaw, Ban-An; Gada, Keyur S.; Patil, Vishwesh; Panwar, Rajiv; Mandapati, Savitri [Northeastern University, Department of Pharmaceutical Sciences, Bouve College of Health Sciences, School of Pharmacy, Boston, MA (United States); Hatefi, Arash [Rutgers University, Department of Pharmaceutics, New Brunswick, NJ (United States); Majewski, Stan [West Virginia University, Department of Radiology, Morgantown, WV (United States); Weisenberger, Andrew [Thomas Jefferson National Accelerator Facility, Jefferson Lab, Newport News, VA (United States)

    2014-08-15

    Doxorubicin, a frontline chemotherapeutic agent, limited by its cardiotoxicity and other tissue toxicities, was conjugated to N-terminal DTPA-modified polyglutamic acid (D-Dox-PGA) to produce polymer pro-drug conjugates. D-Dox-PGA or Tc-99 m labeled DTPA-succinyl-polylysine polymers (DSPL) were targeted to HER2-positive human mammary carcinoma (BT-474) in a double xenografted SCID mouse model also hosting HER2-negative human mammary carcinoma (BT-20). After pretargeting with bispecific anti-HER2-affibody-anti-DTPA-Fab complexes (BAAC), anti-DTPA-Fab or only phosphate buffered saline, D-Dox-PGA or Tc-99 m DSPL were administered. Positive therapeutic control mice were injected with Dox alone at maximum tolerated dose (MTD). Only BT-474 lesions were visualized by gamma imaging with Tc-99 m-DSPL; BT-20 lesions were not. Therapeutic efficacy was equivalent in mice pretargeted with BAAC/targeted with D-Dox-PGA to mice treated only with doxorubicin. There was no total body weight (TBW) loss at three times the doxorubicin equivalent MTD with D-Dox-PGA, whereas mice treated with doxorubicin lost 10 % of TBW at 2 weeks and 16 % after the second MTD injection leading to death of all mice. Our cancer imaging and pretargeted therapeutic approaches are highly target specific, delivering very high specific activity reagents that may result in the development of a novel theranostic application. HER/2 neu specific affibody-anti-DTPA-Fab bispecific antibody pretargeting of HER2 positive human mammary xenografts enabled exquisite targeting of polymers loaded with radioisotopes for molecular imaging and doxorubicin for effective therapy without the associating non-tumor normal tissue toxicities. (orig.)

  7. The Challenges of Recombinant Endostatin in Clinical Application: Focus on the Different Expression Systems and Molecular Bioengineering

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    Abbas Mohajeri

    2017-04-01

    Full Text Available Angiogenesis plays an essential role in rapid growing and metastasis of the tumors. Inhibition of angiogenesis is a putative strategy for cancer therapy. Endostatin (Es is an attractive anti-angiogenesis protein with some clinical application challenges including; short half-life, instability in serum and requirement to high dosage. Therefore, production of recombinant endostatin (rEs is necessary in large scale. The production of rEs is difficult because of its structural properties and is high-cost. Therefore, this review focused on the different expression systems that involved in rEs production including; mammalian, baculovirus, yeast, and Escherichia coli (E. coli expression systems. The evaluating of the results of different expression systems declared that none of the mentioned systems can be considered to be generally superior to the other. Meanwhile with considering the advantages and disadvantage of E. coli expression system compared with other systems beside the molecular properties of Es, E. coli expression system can be a preferred expression system for expressing of the Es in large scale. Also, the molecular bioengineering and sustained release formulations that lead to improving of its stability and bioactivity will be discussed. Point mutation (P125A of Es, addition of RGD moiety or an additional zinc biding site to N-terminal of Es , fusing of Es to anti-HER2 IgG or heavy-chain of IgG, and finally loading of the endostar by PLGA and PEG- PLGA nanoparticles and gold nano-shell particles are the effective bioengineering methods to overcome to clinical changes of endostatin.

  8. Ekspresi HER2/neu(c-ErbB2 pada Kanker Kolorektal

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    Nita Afriani

    2014-12-01

    Full Text Available Kanker kolorektal merupakan keganasan pada kolon dan rektum, penyebab morbiditas danmortalitas ketiga terbanyak diantara keganasan lainnya. Penelitian ekspresi HER2 pada kankerkolorektal memiliki rentang yang cukup jauh yaitu 0-83% dan belum pernah dipublikasi dilndonesia. Penelitian ini bertujuan untuk menilai ekspresi HER2 pada kanker kolorektal. Studicross-secfiona/ dilakukan terhadap 51 sediaan blok parafin kanker kolorektal rentang tahunJanuari 2011-Desember 2012 di Departemen Patologi Anatorni RS Cipto MangunkusumoJakarta. Penilaian karakteristik sampel diambil dari rekam medis dan penilaian histopatologidari sediaan HE pasien kanker kolorektal. Pulasan imunohistokimia HER2 menggunakanantibodipoliklonalantiHER2(DAKO. Rata-rata usia penderita adalah 57.8t13.54tahun,58.8%penderita adalah laki-laki dan 41.2o/o perempuan. Semua sediaan adalah adenokarsinomakolorektal. Ekspresi HER2 positif ditemukan sebanyak 5(9.8% kasus.AbstractColorectal cancer is a neoplasia of colon and rectum ,and also the 3rd leading cause ofmorbidity and mortality among men and women in the world. HER2 expression has controversialresults ranging from 0-83% and never been pubiished in lndonesia. This research aims tostudy HER2 expression in colorectal cancer. Cross-sectional study was conducted in 51 casesof colorectal cancer from January 2011 until December 2012 in Pathology Department ofCipto Mangunkusumo Hospital. Patient characteristic was reviewed from medical record andhistopathologic was evaluated from HE slides. lmunohistochemistry staining used anti HER2polyclonal antibody frorn DAKO. Mean age was 57.8113.54 years, 58.8% were males and41.2o4 were females. Positive HER2 was 5 cases (9.8%.

  9. Concordance Between FISH Analysis of Her-2/Neu Gene in Breast Duct Carcinoma and Corresponding Axillary Nodal Metastases: Egyptian National Cancer Institute Experience.

    Science.gov (United States)

    Badawy, Omnia M; Hassan, Hannan; ELBakey, Heba A; Mosaad, Maha

    2018-05-10

    Breast cancer is a major health problem in Egypt. Her-2/Neu gene is routinely assessed for all breast cancer patients primarily by immunohistochemistry. At National Cancer Institute (NCI), Cairo University, Flourescence In Situ hybridization (FISH) analysis of Her-2/Neu gene is carried out for Her-2/Neu score 2 and for some cases of score 3 (particularly those assessed outside NCI). The test is performed essentially on the primary tumor. However, some situations require testing on corresponding lymph node metastases. There is a debate about the concordance between Her-2/Neu status in the primary tumor and synchronous lymph node metastases in various studies. The aim of this study was to test for the concordance between Her-2/Neu status in the primary breast tumor and corresponding axillary nodal metastases. This is a retrospective study in which FISH analysis of Her-2/Neu was carried out simultaneously on archived material of 50 cases previously diagnosed as invasive duct carcinoma and the corresponding nodal metastases from the Pathology Department, NCI. There was complete concordance between Her-2 status in the primary tumor and the corresponding axillary lymph node metastatic deposits in which Her-2 was amplified in 44% of the studied cohort of Egyptian patients. Her-2/Neu gene assessed by FISH analysis on synchronous lymph node metastases is strongly correlated with the primary tumor. Hence, it is justified to carry out the Her-2/Neu test on synchronous lymph nodes to decide on whether to carry out anti-Her-2/Neu target therapy. Further studies on other metastatic sites is recommended.

  10. Evaluation of Antigen-Conjugated Fluorescent Beads to Identify Antigen-Specific B Cells

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    Isabel Correa

    2018-03-01

    Full Text Available Selection of single antigen-specific B cells to identify their expressed antibodies is of considerable interest for evaluating human immune responses. Here, we present a method to identify single antibody-expressing cells using antigen-conjugated fluorescent beads. To establish this, we selected Folate Receptor alpha (FRα as a model antigen and a mouse B cell line, expressing both the soluble and the membrane-bound forms of a human/mouse chimeric antibody (MOv18 IgG1 specific for FRα, as test antibody-expressing cells. Beads were conjugated to FRα using streptavidin/avidin-biotin bridges and used to select single cells expressing the membrane-bound form of anti-FRα. Bead-bound cells were single cell-sorted and processed for single cell RNA retrotranscription and PCR to isolate antibody heavy and light chain variable regions. Variable regions were then cloned and expressed as human IgG1/k antibodies. Like the original clone, engineered antibodies from single cells recognized native FRα. To evaluate whether antigen-coated beads could identify specific antibody-expressing cells in mixed immune cell populations, human peripheral blood mononuclear cells (PBMCs were spiked with test antibody-expressing cells. Antigen-specific cells could comprise up to 75% of cells selected with antigen-conjugated beads when the frequency of the antigen-positive cells was 1:100 or higher. In PBMC pools, beads conjugated to recombinant antigens FRα and HER2 bound antigen-specific anti-FRα MOv18 and anti-HER2 Trastuzumab antibody-expressing cells, respectively. From melanoma patient-derived B cells selected with melanoma cell line-derived protein-coated fluorescent beads, we generated a monoclonal antibody that recognized melanoma antigen-coated beads. This approach may be further developed to facilitate analysis of B cells and their antibody profiles at the single cell level and to help unravel humoral immune repertoires.

  11. Improved decision making for prioritizing tumor targeting antibodies in human xenografts: Utility of fluorescence imaging to verify tumor target expression, antibody binding and optimization of dosage and application schedule.

    Science.gov (United States)

    Dobosz, Michael; Haupt, Ute; Scheuer, Werner

    2017-01-01

    Preclinical efficacy studies of antibodies targeting a tumor-associated antigen are only justified when the expression of the relevant antigen has been demonstrated. Conventionally, antigen expression level is examined by immunohistochemistry of formalin-fixed paraffin-embedded tumor tissue section. This method represents the diagnostic "gold standard" for tumor target evaluation, but is affected by a number of factors, such as epitope masking and insufficient antigen retrieval. As a consequence, variances and discrepancies in histological staining results can occur, which may influence decision-making and therapeutic outcome. To overcome these problems, we have used different fluorescence-labeled therapeutic antibodies targeting human epidermal growth factor receptor (HER) family members and insulin-like growth factor-1 receptor (IGF1R) in combination with fluorescence imaging modalities to determine tumor antigen expression, drug-target interaction, and biodistribution and tumor saturation kinetics in non-small cell lung cancer xenografts. For this, whole-body fluorescence intensities of labeled antibodies, applied as a single compound or antibody mixture, were measured in Calu-1 and Calu-3 tumor-bearing mice, then ex vivo multispectral tumor tissue analysis at microscopic resolution was performed. With the aid of this simple and fast imaging method, we were able to analyze the tumor cell receptor status of HER1-3 and IGF1R, monitor the antibody-target interaction and evaluate the receptor binding sites of anti-HER2-targeting antibodies. Based on this, the most suitable tumor model, best therapeutic antibody, and optimal treatment dosage and application schedule was selected. Predictions drawn from obtained imaging data were in excellent concordance with outcome of conducted preclinical efficacy studies. Our results clearly demonstrate the great potential of combined in vivo and ex vivo fluorescence imaging for the preclinical development and characterization of

  12. Targeted Delivery of Deoxycytidine Kinase to Her2-Positive Cells Enhances the Efficacy of the Nucleoside Analog Fludarabine.

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    Sujatha P Koduvayur

    Full Text Available Cytotoxic drugs, such as nucleoside analogs and toxins, commonly suffer from off-target effects. One approach to mitigate this problem is to deliver the cytotoxic drug selectively to the intended site. While for toxins this can be achieved by conjugating the cell-killing moiety to a targeting moiety, it is not an option for nucleoside analogs, which rely on intracellular enzymes to convert them to their active triphosphorylated form. To overcome this limitation, and achieve site-targeted activation of nucleoside analogs, we fused the coding region of a prodrug-activating enzyme, deoxycytidine kinase (dCK, to affinity reagents that bind to the Her2 cell surface protein. We evaluated dCK fusions to an anti-Her2 affibody and Designed Ankyrin Repeat Protein (DARPin for their ability to kill cancer cells by promoting the activation of the nucleoside analog fludarabine. Cell staining and flow cytometry experiments with three Her2 positive cancer cell lines (BT-474-JB, JIMT-1 and SK-OV-3 indicate dCK fusions binding and cellular internalization. In contrast, these reagents bind only weakly to the Her2 negative cell line, MCF-7. Cell proliferation assays indicate that SK-OV-3 and BT-474-JB cell lines exhibit significantly reduced proliferation rates when treated with targeting-module fused dCK and fludarabine, compared to fludarabine alone. These findings demonstrate that we have succeeded in delivering active dCK into the Her2-positive cells, thereby increasing the activation of fludarabine, which ultimately reduces the dose of nucleoside analog needed for cell killing. This strategy may help establish the therapeutic index required to differentiate between healthy tissues and cancer cells.

  13. Integrating molecular mechanisms and clinical evidence in the management of trastuzumab resistant or refractory HER-2⁺ metastatic breast cancer.

    Science.gov (United States)

    Wong, Hilda; Leung, Roland; Kwong, Ava; Chiu, Joanne; Liang, Raymond; Swanton, Charles; Yau, Thomas

    2011-01-01

    Human epidermal growth factor receptor (HER)-2(+) breast cancer is a distinct molecular and clinical entity, the prognosis of which is improved by trastuzumab. However, primary resistance to trastuzumab is observed in >50% of patients with HER-2(+) advanced breast cancer, and the majority of patients who initially respond to treatment eventually develop disease progression. To facilitate crosstrial comparisons and the understanding of resistance mechanisms, we propose a unifying definition of trastuzumab resistance as progression at first radiological reassessment at 8-12 weeks or within 3 months after first-line trastuzumab in the metastatic setting or new recurrences diagnosed during or within 12 months after adjuvant trastuzumab. In contrast, we define trastuzumab-refractory breast cancer as disease progression after two or more lines of trastuzumab-containing regimens that initially achieved disease response or stabilization at first radiological assessment. We review mechanisms of trastuzumab resistance mediated by p95HER-2 overexpression, phosphoinositide 3-kinase pathway activation, and signaling pathway activation driven by HER-3, epidermal growth factor receptor, and insulin-like growth factor 1 receptor. We distinguish in vitro from in vivo evidence, highlighting that most data describing trastuzumab resistance are derived from preclinical studies or small retrospective patient cohorts, and discuss targeted therapeutic approaches to overcome resistance. Prospective analysis through clinical trials with robust tissue collection procedures, prior to and following acquisition of resistance, integrated with next-generation tumor genome sequencing technologies, is identified as a priority area for development. The identification of predictive biomarkers is of paramount importance to optimize health economic costs and enhance stratification of anti-HER-2 targeted therapies.

  14. The sensitivity and efficacy method of PIK3CA exon 9 E545A as a high diagnostic accuracy in breast cancer

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    Desriani

    2018-06-01

    Full Text Available The phosphatidylinositol 3-kinases (PIK3s are lipid kinases. Mutation in the exon 9 and exon 20 determined as a predictive factor in anti-HER-2 therapy. In some countries, such as Singapore, China, and Peru, PIK3CA exon 9 E545A was reported to produce the highest rate of mutation. In this research, we developed and optimized PIK3CA exon 9 E545A detection methods with intercalating dye SYBR Green I based on the Tm Shift approach by using prepared recombinant plasmid pGEMT-easy PIK3CA exon 9 and PIK3CA exon 9 E545A. Recombinant plasmid was used due to the limited number of samples. Methods: Recombinant plasmid was prepared based on manufactured procedures, and this process was then followed by Tm prediction with Poland software, Tm Shift SYBR Green I development, and its characterization (reproducibility, repeatability, sensitivity, qPCR efficiency, and qPCR amplification, respectively. Result: A method for PIK3CA E545A detection based on TM shift SYBR Green I has been successfully developed. The melting temperature for PIK3CA exon 9 was 78.1 ± 0.1 °C, while that for PIK3CA exon E545A was 80.20 °C. The Tm of mutant was the same as that predicted using Polland Software. The reproducibility of the methods was high, with the coefficient values for inter and intra assays were below 10% with a high sensitivity at 1%, while R2 0.99 and PCR efficiency was 97.75%. Conclusion: The results presented here demonstrate that the PIK3CA exon 9 E545A detection method has a good sensitivity and efficacy assay, which proves that the method has a high diagnostic accuracy in breast cancer. Keywords: SYBR Green I, PIK3CA E545A, Breast cancer, Real time PCR, Recombinant plasmid

  15. Photothermal effects of multi-walled carbon nanotubes on the viability of BT-474 cancer cells

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    Chou, Hung-Tao [Department of Materials Science and Engineering, National Tsing Hua University, No. 101, Sec. 2 Kuang-Fu Rd., Hsin-chu 30013, Taiwan (China); Wang, Tsung-Pao [Department of Medical Science, National Tsing Hua University, No. 101, Sec. 2 Kuang-Fu Rd., Hsin-chu 30013, Taiwan (China); Lee, Chi-Young [Department of Materials Science and Engineering, National Tsing Hua University, No. 101, Sec. 2 Kuang-Fu Rd., Hsin-chu 30013, Taiwan (China); Tai, Nyan-Hwa, E-mail: nhtai@mx.nthu.edu.tw [Department of Materials Science and Engineering, National Tsing Hua University, No. 101, Sec. 2 Kuang-Fu Rd., Hsin-chu 30013, Taiwan (China); Chang, Hwan-You, E-mail: hychang@mx.nthu.edu.tw [Department of Medical Science, National Tsing Hua University, No. 101, Sec. 2 Kuang-Fu Rd., Hsin-chu 30013, Taiwan (China)

    2013-03-01

    Functionalized multi-walled carbon nanotubes (f-MWCNTs) were conjugated to an antibody of BT-474 cancer cells (f-MWCNTs-ab), and the photothermal effect of the f-MWCNTs-ab for BT-474 cancer cell destruction was demonstrated. After near-infrared irradiation, the f-MWCNTs-ab were more capable of killing cancer cells and possessed higher cell specificity than f-MWCNTs. Quantitative results showed that the viability of the cancer cells was affected by the concentration of the f-MWCNTs-ab solution, irradiation time, and settling time after irradiation. The membrane impermeable fluorescence dye ethidium bromide was used to detect cell viability after near-infrared irradiation, and the results agreed with those obtained from the Alamar Blue cell viability assay. The EtBr fluorescence results suggest that the cell membrane, attached to f-MWCNTs-ab, was damaged after irradiation, which led to cell death and necrosis. Using confocal microscopy, a few f-MWCNTs-ab were detected in the cell, indicating the endocytosis effect. The results not only explain the improved efficiency of thermotherapy but also indicate that necrosis may result from protein denaturation attributing to the heated f-MWCNTs-ab in the cell. Highlights: Black-Right-Pointing-Pointer f-MWCNTs conjugated with anti-HER2 antibody by chemical method. Black-Right-Pointing-Pointer Kill breast cancer cells by using low dose f-MWCNTs-ab due to photothermal effect. Black-Right-Pointing-Pointer Use EtBr fluorescent to prove that the cell membrane was broken by heated f-MWCNTs. Black-Right-Pointing-Pointer Few f-MWCNTs-ab were detected in the cell indicating the endocytosis effect. Black-Right-Pointing-Pointer Necrosis may result from protein denaturation due to contact with the heated CNTs.

  16. HER2 signaling pathway activation and response of breast cancer cells to HER2-targeting agents is dependent strongly on the 3D microenvironment

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    Weigelt, Britta; Lo, Alvin T; Park, Catherine C; Gray, Joe W; Bissell, Mina J

    2009-07-27

    Development of effective and durable breast cancer treatment strategies requires a mechanistic understanding of the influence of the microenvironment on response. Previous work has shown that cellular signaling pathways and cell morphology are dramatically influenced by three-dimensional (3D) cultures as opposed to traditional two-dimensional (2D) monolayers. Here, we compared 2D and 3D culture models to determine the impact of 3D architecture and extracellular matrix (ECM) on HER2 signaling and on the response of HER2-amplified breast cancer cell lines to the HER2-targeting agents Trastuzumab, Pertuzumab and Lapatinib. We show that the response of the HER2-amplified AU565, SKBR3 and HCC1569 cells to these anti-HER2 agents was highly dependent on whether the cells were cultured in 2D monolayer or 3D laminin-rich ECM gels. Inhibition of {beta}1 integrin, a major cell-ECM receptor subunit, significantly increased the sensitivity of the HER2-amplified breast cancer cell lines to the humanized monoclonal antibodies Trastuzumab and Pertuzumab when grown in a 3D environment. Finally, in the absence of inhibitors, 3D cultures had substantial impact on HER2 downstream signaling and induced a switch between PI3K-AKT- and RAS-MAPKpathway activation in all cell lines studied, including cells lacking HER2 amplification and overexpression. Our data provide direct evidence that breast cancer cells are able to rapidly adapt to different environments and signaling cues by activating alternative pathways that regulate proliferation and cell survival, events that may play a significant role in the acquisition of resistance to targeted therapies.

  17. Position for Site-Specific Attachment of a DOTA Chelator to Synthetic Affibody Molecules Has a Different Influence on the Targeting Properties of 68Ga-Compared to 111In-Labeled Conjugates

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    Hadis Honarvar

    2014-12-01

    Full Text Available Affibody molecules, small (7 kDa scaffold proteins, are a promising class of probes for radionuclide molecular imaging. Radiolabeling of Affibody molecules with the positron-emitting nuclide 68Ga would permit the use of positron emission tomography (PET, providing better resolution, sensitivity, and quantification accuracy than single-photon emission computed tomography (SPECT. The synthetic anti-HER2 ZHER2:S1 Affibody molecule was conjugated with DOTA at the N-terminus, in the middle of helix 3, or at the C-terminus. The biodistribution of 68Ga- and 111In-labeled Affibody molecules was directly compared in NMRI nu/nu mice bearing SKOV3 xenografts. The position of the chelator strongly influenced the biodistribution of the tracers, and the influence was more pronounced for 68Ga-labeled Affibody molecules than for the 111In-labeled counterparts. The best 68Ga-labeled variant was 68Ga-[DOTA-A1]-ZHER2:S1 which provided a tumor uptake of 13 ± 1 %ID/g and a tumor to blood ratio of 39 ± 12 at 2 hours after injection. 111In-[DOTA-A1]-ZHER2:S1 and 111In-[DOTA-K58]-ZHER2:S1 were equally good at this time point, providing a tumor uptake of 15 to 16 %ID/g and a tumor to blood ratio in the range of 60 to 80. In conclusion, the selection of the best position for a chelator in Affibody molecules can be used for optimization of their imaging properties. This may be important for the development of Affibody-based and other protein-based imaging probes.

  18. Sensitivity of HER-2/neu antibodies in archival tissue samples: potential source of error in immunohistochemical studies of oncogene expression.

    Science.gov (United States)

    Press, M F; Hung, G; Godolphin, W; Slamon, D J

    1994-05-15

    HER-2/neu oncogene amplification and overexpression of breast cancer tissue has been correlated with poor prognosis in women with both node-positive and node-negative disease. However, several studies have not confirmed this association. Review of these studies reveals the presence of considerable methodological variability including differences in study size, follow-up time, techniques and reagents. The majority of papers with clinical follow-up information are immunohistochemical studies using archival, paraffin-embedded breast cancers, and a variety of HER-2/neu antibodies have been used in these studies. Very little information, however, is available about the ability of the antibodies to detect overexpression following tissue processing for paraffin-embedding. Therefore, a series of antibodies, reported in the literature or commercially available, were evaluated to assess their sensitivity and specificity as immunohistochemical reagents. Paraffin-embedded samples of 187 breast cancers, previously characterized as frozen specimens for HER-2/neu amplification by Southern blot and for overexpression by Northern blot, Western blot, and immunohistochemistry, were used. Two multitumor paraffin-embedded tissue blocks were prepared from the previously analyzed breast cancers as a panel of cases to test a series of previously studied and/or commercially available anti-HER-2/neu antibodies. Immunohistochemical staining results obtained with 7 polyclonal and 21 monoclonal antibodies in sections from paraffin-embedded blocks of these breast cancers were compared. The ability of these antibodies to detect overexpression was extremely variable, providing an important explantation for the variable overexpression rate reported in the literature.

  19. Genetic Predictors of Response to Systemic Therapy in Esophagogastric Cancer.

    Science.gov (United States)

    Janjigian, Yelena Y; Sanchez-Vega, Francisco; Jonsson, Philip; Chatila, Walid K; Hechtman, Jaclyn F; Ku, Geoffrey Y; Riches, Jamie C; Tuvy, Yaelle; Kundra, Ritika; Bouvier, Nancy; Vakiani, Efsevia; Gao, Jianjiong; Heins, Zachary J; Gross, Benjamin E; Kelsen, David P; Zhang, Liying; Strong, Vivian E; Schattner, Mark; Gerdes, Hans; Coit, Daniel G; Bains, Manjit; Stadler, Zsofia K; Rusch, Valerie W; Jones, David R; Molena, Daniela; Shia, Jinru; Robson, Mark E; Capanu, Marinela; Middha, Sumit; Zehir, Ahmet; Hyman, David M; Scaltriti, Maurizio; Ladanyi, Marc; Rosen, Neal; Ilson, David H; Berger, Michael F; Tang, Laura; Taylor, Barry S; Solit, David B; Schultz, Nikolaus

    2018-01-01

    The incidence of esophagogastric cancer is rapidly rising, but only a minority of patients derive durable benefit from current therapies. Chemotherapy as well as anti-HER2 and PD-1 antibodies are standard treatments. To identify predictive biomarkers of drug sensitivity and mechanisms of resistance, we implemented prospective tumor sequencing of patients with metastatic esophagogastric cancer. There was no association between homologous recombination deficiency defects and response to platinum-based chemotherapy. Patients with microsatellite instability-high tumors were intrinsically resistant to chemotherapy but more likely to achieve durable responses to immunotherapy. The single Epstein-Barr virus-positive patient achieved a durable, complete response to immunotherapy. The level of ERBB2 amplification as determined by sequencing was predictive of trastuzumab benefit. Selection for a tumor subclone lacking ERBB2 amplification, deletion of ERBB2 exon 16, and comutations in the receptor tyrosine kinase, RAS, and PI3K pathways were associated with intrinsic and/or acquired trastuzumab resistance. Prospective genomic profiling can identify patients most likely to derive durable benefit to immunotherapy and trastuzumab and guide strategies to overcome drug resistance. Significance: Clinical application of multiplex sequencing can identify biomarkers of treatment response to contemporary systemic therapies in metastatic esophagogastric cancer. This large prospective analysis sheds light on the biological complexity and the dynamic nature of therapeutic resistance in metastatic esophagogastric cancers. Cancer Discov; 8(1); 49-58. ©2017 AACR. See related commentary by Sundar and Tan, p. 14 See related article by Pectasides et al., p. 37 This article is highlighted in the In This Issue feature, p. 1 . ©2017 American Association for Cancer Research.

  20. Dual-Ligand Modified Polymer-Lipid Hybrid Nanoparticles for Docetaxel Targeting Delivery to Her2/neu Overexpressed Human Breast Cancer Cells.

    Science.gov (United States)

    Yang, Zhe; Tang, Wenxin; Luo, Xingen; Zhang, Xiaofang; Zhang, Chao; Li, Hao; Gao, Di; Luo, Huiyan; Jiang, Qing; Liu, Jie

    2015-08-01

    In this study, a dual-ligand polymer-lipid hybrid nanoparticle drug delivery vehicle comprised of an anti-HER2/neu peptide (AHNP) mimic with a modified HIV-1 Tat (mTAT) was established for the targeted treatment of Her2/neu-overexpressing cells. The resultant dual-ligand hybrid nanoparticles (NPs) consisted of a poly(lactide-co-glycolide) core, a near 90% surface coverage of the lipid monolayer, and a 5.7 nm hydrated polyethylene glycol shell. Ligand density optimization study revealed that cellular uptake efficiency of the hybrid NPs could be manipulated by controlling the surface-ligand densities. Furthermore, the cell uptake kinetics and mechanism studies showed that the dual-ligand modifications of hybrid NPs altered the cellular uptake pathway from caveolae-mediated endocytosis (CvME) to the multiple endocytic pathways, which would significantly enhance the NP internalization. Upon the systemic investigation of the cellular uptake behavior of dual-ligand hybrid NPs, docetaxel (DTX), a hydrophobic anticancer drug, was successfully encapsulated into dual-ligand hybrid NPs with high drug loading for Her2/neu-overexpressing SK-BR-3 breast cancer cell treatment. The DTX-loaded dual-ligand hybrid NPs showed a decreased burst release and a more gradual sustained drug release property. Because of the synergistic effect of dual-ligand modification, DTX-loaded dual-ligand hybrid NPs exerted substantially better therapeutic potency against SK-BR-3 cancer cells than other NP formulations and free DTX drugs. These results demonstrate that the dual-ligand hybrid NPs could be a promising vehicle for targeted drug delivery to treat breast cancer.

  1. Dual-Labeled Near-Infrared/99mTc Imaging Probes Using PAMAM-Coated Silica Nanoparticles for the Imaging of HER2-Expressing Cancer Cells

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    Haruka Yamaguchi

    2016-07-01

    Full Text Available We sought to develop dual-modality imaging probes using functionalized silica nanoparticles to target human epidermal growth factor receptor 2 (HER2-overexpressing breast cancer cells and achieve efficient target imaging of HER2-expressing tumors. Polyamidoamine-based functionalized silica nanoparticles (PCSNs for multimodal imaging were synthesized with near-infrared (NIR fluorescence (indocyanine green (ICG and technetium-99m (99mTc radioactivity. Anti-HER2 antibodies were bound to the labeled PCSNs. These dual-imaging probes were tested to image HER2-overexpressing breast carcinoma cells. In vivo imaging was also examined in breast tumor xenograft models in mice. SK-BR3 (HER2 positive cells were imaged with stronger NIR fluorescent signals than that in MDA-MB231 (HER2 negative cells. The increased radioactivity of the SK-BR3 cells was also confirmed by phosphor imaging. NIR images showed strong fluorescent signals in the SK-BR3 tumor model compared to muscle tissues and the MDA-MB231 tumor model. Automatic well counting results showed increased radioactivity in the SK-BR3 xenograft tumors. We developed functionalized silica nanoparticles loaded with 99mTc and ICG for the targeting and imaging of HER2-expressing cells. The dual-imaging probes efficiently imaged HER2-overexpressing cells. Although further studies are needed to produce efficient isotope labeling, the results suggest that the multifunctional silica nanoparticles are a promising vehicle for imaging specific components of the cell membrane in a dual-modality manner.

  2. Nanobody-functionalized PEG-b-PCL polymersomes and their targeting study.

    Science.gov (United States)

    Zou, Tao; Dembele, Fatimata; Beugnet, Anne; Sengmanivong, Lucie; Trepout, Sylvain; Marco, Sergio; de Marco, Ario; Li, Min-Hui

    2015-11-20

    We prepared and characterized polymersomes functionalized with nanobodies (VHHs) on the basis of biocompatible, biodegradable and FDA-approved poly(ethylene glycol)-block-poly(ϵ-caprolactone) (PEG-b-PCL). Fluorescein isothiocyanate (FITC) and N-beta-maleimidopropyl-oxysuccinimide ester were allowed reacting with H2N-PEG-b-PCL to produce FITC and maleimide (Mal) functionalized copolymers, Mal-PEG-b-PCL and FITC-PEG-b-PCL. A mixture of MeO-PEG-b-PCL, Mal-PEG-b-PCL and FITC-PEG-b-PCL was used to prepare polymersomes by thin film hydration and nanoprecipitation methods. Morphological studies by cryogenic transmission electron microscopy (Cryo-TEM) showed that the nanoparticles exhibited predominantly vesicular structures (polymersomes). Their mean diameters measured by dynamic light scattering were around 150 nm and the zeta-potentials around -1 mV at pH 7.4. The nanoparticles were functionalized with either anti-HER2 (VHH1) or anti-GFP (VHH2) nanobodies using maleimide-cysteine chemistry. Their particle size and zeta-potential increased slightly after nanobody-functionalization. The specific binding of VHH-functionalized polymersomes and control nanoparticles towards HER2 positive breast cancer cells was analyzed by flow cytometry and confocal microscopy. The collected results represent the first report which experimentally demonstrates that VHH1-functionalized PEO-b-PCL polymersomes can target specifically breast cancer cells expressing HER2 receptors. The detailed morphological and cell-binding studies described herein pave the way for future in vivo studies to evaluate the feasibility to use such nanoparticles for targeted drug delivery. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Evaluation of Antigen-Conjugated Fluorescent Beads to Identify Antigen-Specific B Cells

    Science.gov (United States)

    Correa, Isabel; Ilieva, Kristina M.; Crescioli, Silvia; Lombardi, Sara; Figini, Mariangela; Cheung, Anthony; Spicer, James F.; Tutt, Andrew N. J.; Nestle, Frank O.; Karagiannis, Panagiotis; Lacy, Katie E.; Karagiannis, Sophia N.

    2018-01-01

    Selection of single antigen-specific B cells to identify their expressed antibodies is of considerable interest for evaluating human immune responses. Here, we present a method to identify single antibody-expressing cells using antigen-conjugated fluorescent beads. To establish this, we selected Folate Receptor alpha (FRα) as a model antigen and a mouse B cell line, expressing both the soluble and the membrane-bound forms of a human/mouse chimeric antibody (MOv18 IgG1) specific for FRα, as test antibody-expressing cells. Beads were conjugated to FRα using streptavidin/avidin-biotin bridges and used to select single cells expressing the membrane-bound form of anti-FRα. Bead-bound cells were single cell-sorted and processed for single cell RNA retrotranscription and PCR to isolate antibody heavy and light chain variable regions. Variable regions were then cloned and expressed as human IgG1/k antibodies. Like the original clone, engineered antibodies from single cells recognized native FRα. To evaluate whether antigen-coated beads could identify specific antibody-expressing cells in mixed immune cell populations, human peripheral blood mononuclear cells (PBMCs) were spiked with test antibody-expressing cells. Antigen-specific cells could comprise up to 75% of cells selected with antigen-conjugated beads when the frequency of the antigen-positive cells was 1:100 or higher. In PBMC pools, beads conjugated to recombinant antigens FRα and HER2 bound antigen-specific anti-FRα MOv18 and anti-HER2 Trastuzumab antibody-expressing cells, respectively. From melanoma patient-derived B cells selected with melanoma cell line-derived protein-coated fluorescent beads, we generated a monoclonal antibody that recognized melanoma antigen-coated beads. This approach may be further developed to facilitate analysis of B cells and their antibody profiles at the single cell level and to help unravel humoral immune repertoires. PMID:29628923

  4. A novel IgA-like immunoglobulin in the reptile Eublepharis macularius.

    Science.gov (United States)

    Deza, Francisco Gambón; Espinel, Christian Sánchez; Beneitez, Julio Valdueza

    2007-01-01

    The appearance of antibody genes over evolution coincided with the origin of the vertebrates. Reptiles are of great interest in evolution since they are the link between the amphibians, birds, and mammals. This work describes the presence of a gene in the reptile leopard gecko (Eublepharis macularius) where phylogenetic studies suggest that it is the gene orthologue of immunoglobulin A (IgA) and immunoglobulin X (IgX) in Xenopus. Messenger RNA samples taken from different tissues showed expression of this antibody in intestinal tissue. Data on the structure deduced from the sequence of nucleotides showed an antibody with four domains in the constant region. There is a sequence of 20 amino acids in the C terminus similar to the secretory tail of immunoglobulin M (IgM) and IgA. A detailed analysis of the sequence of amino acids displayed a paradox, i.e., domains CH1 and CH2 showed a clear homology with domains CH1 and CH2 of immunoglobulin Y (IgY) while domains CH3 and CH4 were homologous with domains CH3 and CH4 of IgM. This homology pattern is also seen in Xenopus IgX and bird IgA. The most logical explanation for this phenomenon is that a recombination between the IgM and IgY gave rise to the IgA.

  5. Single Chain Antibody Fragment against Venom from the Snake Daboia russelii formosensis

    Directory of Open Access Journals (Sweden)

    Chi-Hsin Lee

    2017-10-01

    Full Text Available Russell’s vipers containing hemotoxic and neurotoxic venom commonly cause snake envenomation. Horse-derived antivenom is a specific antidote, but its production is expensive and has side effects. Developing a cost-effective and more tolerable therapeutic strategy is favorable. In this study, using glutaraldehyde-attenuated Daboia russelii formosensis (DRF venom proteins to immunize chickens, polyclonal yolk-immunoglobulin (IgY antibodies were generated and showed a specific binding affinity. Phage display technology was used to generate two antibody libraries of single-chain variable fragments (scFvs containing 3.4 × 107 and 5.5 × 107 transformants, respectively. Phage-based ELISA indicated that specific clones were enriched after bio-panning. The nucleotide sequences of scFv-expressing clones were analyzed and classified into six groups in the short linker and four groups in the long linker. These scFv antibodies specifically bound to DRF proteins, but not other venom proteins. Mass spectrometric data suggested that these scFv antibodies may recognize phospholipase A2 RV-4 or RV-7. In vivo studies showed that anti-DRF IgY exhibited complete protective effects and mixed scFv antibodies increased the survival rate and time of mice challenged with a lethal dose of DRF proteins. These antibodies can be potentially applied in a rapid diagnostic method or for treatment in the future.

  6. Identification and Extraction of Chicken Egg Yolk Immunoglobulin from Egg by Polyethylene Glycol (PEG Precipitation

    Directory of Open Access Journals (Sweden)

    Mohammad Mehdi Soltan Dallal

    2016-01-01

    Full Text Available Background: Staphylococcus aureus strains exhibiting multiple antibiotic resistances are isolatedfrom most communities and hospital infections. Treatment of patients with these infections hasbeen difficult. The aim of this study was to detect and extract, the egg yolk immunoglobulin Y asa potential source of anti- S. aureus antibody.Methods: Specific IgY was produced by immunizing hens with formalin-killed S. aureus. Thespecificity of serum`s antibody was confirmed by ELISA method. The antibodies were extractedfrom egg yolk by polyethylene glycol (PEG precipitation. Proteins were analysed by sodiumdodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE.Results: Chicken egg yolk antibodies (IgY were raised against S. aureus in the serum afterinjections. Up to 104 dilution specific antibodies were determined in serum.Conclusion: The results of the ELISA indicates the specificity of the immunoglobulin Y to thetarget antigen. In order to find a viable alternative to antibiotic treatments, more research must bedone on the ability of these antibodies to inhibit the growth of S. aureus.

  7. Single Chain Antibody Fragment against Venom from the Snake Daboia russelii formosensis.

    Science.gov (United States)

    Lee, Chi-Hsin; Lee, Yu-Ching; Lee, Yueh-Lun; Leu, Sy-Jye; Lin, Liang-Tzung; Chen, Chi-Ching; Chiang, Jen-Ron; Mwale, Pharaoh Fellow; Tsai, Bor-Yu; Hung, Ching-Sheng; Yang, Yi-Yuan

    2017-10-27

    Russell's vipers containing hemotoxic and neurotoxic venom commonly cause snake envenomation. Horse-derived antivenom is a specific antidote, but its production is expensive and has side effects. Developing a cost-effective and more tolerable therapeutic strategy is favorable. In this study, using glutaraldehyde-attenuated Daboia russelii formosensis (DRF) venom proteins to immunize chickens, polyclonal yolk-immunoglobulin (IgY) antibodies were generated and showed a specific binding affinity. Phage display technology was used to generate two antibody libraries of single-chain variable fragments (scFvs) containing 3.4 × 10⁷ and 5.5 × 10⁷ transformants, respectively. Phage-based ELISA indicated that specific clones were enriched after bio-panning. The nucleotide sequences of scFv-expressing clones were analyzed and classified into six groups in the short linker and four groups in the long linker. These scFv antibodies specifically bound to DRF proteins, but not other venom proteins. Mass spectrometric data suggested that these scFv antibodies may recognize phospholipase A2 RV-4 or RV-7. In vivo studies showed that anti-DRF IgY exhibited complete protective effects and mixed scFv antibodies increased the survival rate and time of mice challenged with a lethal dose of DRF proteins. These antibodies can be potentially applied in a rapid diagnostic method or for treatment in the future.

  8. Differential effects of B7-1 blockade in the rat experimental autoimmune encephalomyelitis model

    DEFF Research Database (Denmark)

    Gallon, L; Chandraker, A; Issazadeh-Navikas, Shohreh

    1997-01-01

    that CD28-B7 blockade by systemic administration of CTLA4Ig prevents actively induced EAE. Since CTLA4Ig binds to both B7-1 and B7-2, we used a mutant form of CTLA4Ig (CTLA4IgY100F) that binds only B7-1, to study the role of B7-1 blockade in this model. Such a reagent avoids the potential of signaling...... treated with systemic CTLA4gY100F did not. More importantly, systemic administration of CTLA4IgY100F abrogated the protective effect of ex vivo treated APCs. These data suggest an important regulatory role for B7-1, perhaps through binding to CTLA4, in this model of EAE. Understanding the role......Blocking the CD28-B7 T cell costimulatory activation pathway protects animals from developing experimental autoimmune encephalomyelitis (EAE). In the mouse EAE model, selective blockade of B7-1 by specific mAbs has been shown to protect animals from EAE. In the Lewis rat model, we have shown...

  9. Epidemic infectious gastrointestinal illness aboard U.S. Navy ships deployed to the Middle East during peacetime operations – 2000–2001

    Directory of Open Access Journals (Sweden)

    Bresee Joseph S

    2006-02-01

    Full Text Available Abstract Background Infectious gastrointestinal illness (IGI outbreaks have been reported in U.S. Navy ships and could potentially have an adverse mission impact. Studies to date have been anecdotal. Methods We conducted a retrospective analysis of weekly reported disease and non-battle injury health data collected in 2000 – 2001 from 44 U.S. Navy ships while sailing in the 5th Fleet (Persian Gulf and nearby seas. Results During this period, 11 possible IGI outbreaks were identified. Overall, we found 3.3 outbreaks per 100 ship-weeks, a mean outbreak duration of 4.4 weeks, and a mean cumulative ship population attack rate of 3.6%. Morbidity, represented by days lost due to personnel being placed on sick-in-quarters status, was higher during outbreak weeks compared to non-outbreak weeks (p = 0.002. No clear seasonal distribution was identified. Conclusion Explosive outbreaks due to viruses and bacteria with the potential of incapacitating large proportions of the crew raise serious concerns of mission impact and military readiness.

  10. Reply to Nicholson's comment on "Consistent calculation of aquatic gross production from oxygen triple isotope measurements" by Kaiser (2011

    Directory of Open Access Journals (Sweden)

    J. Kaiser

    2012-08-01

    Full Text Available The comment by Nicholson (2011a questions the "consistency" of the "definition" of the "biological end-member" used by Kaiser (2011a in the calculation of oxygen gross production. "Biological end-member" refers to the relative oxygen isotope ratio difference between photosynthetic oxygen and Air-O2 (abbreviated 17δP and 18δP for 17O/16O and 18O/16O, respectively. The comment claims that this leads to an overestimate of the discrepancy between previous studies and that the resulting gross production rates are "30% too high". Nicholson recognises the improved accuracy of Kaiser's direct calculation ("dual-delta" method compared to previous approximate approaches based on 17O excess (17Δ and its simplicity compared to previous iterative calculation methods. Although he correctly points out that differences in the normalised gross production rate (<i>g> are largely due to different input parameters used in Kaiser's "base case" and previous studies, he does not acknowledge Kaiser's observation that iterative and dual-delta calculation methods give exactly the same <i>g> for the same input parameters (disregarding kinetic isotope fractionation during air-sea exchange. The comment is based on misunderstandings with respect to the "base case" 17δP and 18δP values. Since direct measurements of 17δP and 18δPdo not exist or have been lost, Kaiser constructed the "base case" in a way that was consistent and compatible with literature data. Nicholson showed that an alternative reconstruction of 17δP gives <i>g> values closer to previous studies. However, unlike Nicholson, we refrain from interpreting either reconstruction as a benchmark for the accuracy of <i>g>. A number of publications over the last 12 months

  11. Optimizing ultrasound molecular imaging of secreted frizzled related protein 2 expression in angiosarcoma.

    Directory of Open Access Journals (Sweden)

    James K Tsuruta

    Full Text Available Secreted frizzled related protein 2 (SFRP2 is a tumor endothelial marker expressed in angiosarcoma. Previously, we showed ultrasound molecular imaging with SFRP2-targeted contrast increased average video pixel intensity (VI of angiosarcoma vessels by 2.2 ± 0.6 VI versus streptavidin contrast. We hypothesized that redesigning our contrast agents would increase imaging performance. Improved molecular imaging reagents were created by combining NeutrAvidin™-functionalized microbubbles with biotinylated SFRP2 or IgY control antibodies. When angiosarcoma tumors in nude mice reached 8 mm, time-intensity, antibody loading, and microbubble dose experiments optimized molecular imaging. 10 minutes after injection, the control-subtracted time-intensity curve (TIC for SFRP2-targeted contrast reached a maximum, after subtracting the contribution of free-flowing contrast. SFRP2 antibody-targeted VI was greater when contrast was formulated with 10-fold molar excess of maleimide-activated NeutrAvidin™ versus 3-fold (4.5 ± 0.18 vs. 0.32 ± 0.15, VI ± SEM, 5 x 106 dose, p < 0.001. Tumor vasculature returned greater average video pixel intensity using 5 x 107 versus 5 x 106 microbubbles (21.2 ± 2.5 vs. 4.5 ± 0.18, p = 0.0011. Specificity for tumor vasculature was confirmed by low VI for SFRP2-targeted, and control contrast in peri-tumoral vasculature (3.2 ± 0.52 vs. 1.6 ± 0.71, p = 0.92. After optimization, average video pixel intensity of tumor vasculature was 14.2 ± 3.0 VI units higher with SFRP2-targeted contrast versus IgY-targeted control (22.1 ± 2.5 vs. 7.9 ± 1.6, p < 0.001. After log decompression, 14.2 ΔVI was equal to ~70% higher signal, in arbitray acoustic units (AU, for SFRP2 versus IgY. This provided ~18- fold higher acoustic signal enhancement than provided previously by 2.2 ΔVI. Basing our targeted contrast on NeutrAvidin™-functionalized microbubbles, using IgY antibodies for our control contrast, and optimizing our imaging protocol

  12. A hybrid biomechanical intensity based deformable image registration of lung 4DCT

    International Nuclear Information System (INIS)

    Samavati, Navid; Velec, Michael; Brock, Kristy

    2015-01-01

    Deformable image registration (DIR) has been extensively studied over the past two decades due to its essential role in many image-guided interventions (IGI). IGI demands a highly accurate registration that maintains its accuracy across the entire region of interest. This work evaluates the improvement in accuracy and consistency by refining the results of Morfeus, a biomechanical model-based DIR algorithm.A hybrid DIR algorithm is proposed based on, a biomechanical model–based DIR algorithm and a refinement step based on a B-spline intensity-based algorithm. Inhale and exhale reconstructions of four-dimensional computed tomography (4DCT) lung images from 31 patients were initially registered using the biomechanical DIR by modeling contact surface between the lungs and the chest cavity. The resulting deformations were then refined using the intensity-based algorithm to reduce any residual uncertainties. Important parameters in the intensity-based algorithm, including grid spacing, number of pyramids, and regularization coefficient, were optimized on 10 randomly-chosen patients (out of 31). Target registration error (TRE) was calculated by measuring the Euclidean distance of common anatomical points on both images after registration. For each patient a minimum of 30 points/lung were used.Grid spacing of 8 mm, 5 levels of grid pyramids, and regularization coefficient of 3.0 were found to provide optimal results on 10 randomly chosen patients. Overall the entire patient population (n = 31), the hybrid method resulted in mean ± SD (90th%) TRE of 1.5 ± 1.4 (2.9) mm compared to 3.1 ± 1.9 (5.6) using biomechanical DIR and 2.6 ± 2.5 (6.1) using intensity-based DIR alone.The proposed hybrid biomechanical modeling intensity based algorithm is a promising DIR technique which could be used in various IGI procedures. The current investigation shows the efficacy of this approach for the registration of 4DCT images of the lungs with average accuracy of 1.5

  13. Gut-Associated Lymphoid Tissue: A Key Tissue Inside the Mucosal Immune System of Hens Immunized with Escherichia coli F4.

    Science.gov (United States)

    Peralta, Maria F; Magnoli, Alejandra; Alustiza, Fabrisio; Nilson, Armando; Miazzo, Raúl; Vivas, Adriana

    2017-01-01

    Immunoglobulin Y (IgY) is the predominant antibody found in hen's ( Gallus domesticus ) egg yolk. This antibody, developed against several microorganisms in hen egg yolk, has been successfully used as an alternative to immunoglobulins from mammals for use in immunodiagnostics and immunotherapy. Enteropathogenic Escherichia coli (E.coli) F 4 is the main etiological agent associated with swine neonatal diarrhea, and it causes notable economic losses in swine production. The aim of the present study was to evaluate the relationship between humoral immune response and the activation of gut-associated lymphoid tissue (GALT) in laying hens intramuscularly immunized with E. coli F 4 . Adult laying Shaver hens were immunized with a bacterin based on an inactivated lysate E. coli F 4 strain that was originally isolated from neonatal piglet diarrhea, following a recommended schedule. The percentage of B lymphocytes in blood and spleen homogenates was determined by flow cytometry. Villi histomorphometry and the size of germinal centers (GC) activated in GALT and the spleen were measured in histological samples either stained with hematoxylin/eosin or through immunofluorescence. Antibody and isotype-specific antibodies in serum and egg yolk were measured using indirect enzyme-linked immunosorbent assay (ELISA). Secretory and serum immunoglobulin A (IgA) were measured by ELISA tests. Laying hen with intramuscular immunization with E. coli F 4 lysate, activated both mucosal and systemic protection. Mucosal protection was provided through B lymphocytes, and most of them were activated on Peyer's patches and esophageal tonsils, in GALT. Furthermore, increased B lymphocyte number in the lamina propria of the gut, and increased intraepithelial plasmatic cell number, produced high levels of mucosal IgA. Activated B lymphocytes interacted with absorptive cells, immune cells, and microbiota in the gut, producing signals that were translated into a powerful physical defense by producing

  14. Antiviral Biologic Produced in DNA Vaccine/Goose Platform Protects Hamsters Against Hantavirus Pulmonary Syndrome When Administered Post-exposure.

    Directory of Open Access Journals (Sweden)

    Nicole Haese

    Full Text Available Andes virus (ANDV and ANDV-like viruses are responsible for most hantavirus pulmonary syndrome (HPS cases in South America. Recent studies in Chile indicate that passive transfer of convalescent human plasma shows promise as a possible treatment for HPS. Unfortunately, availability of convalescent plasma from survivors of this lethal disease is very limited. We are interested in exploring the concept of using DNA vaccine technology to produce antiviral biologics, including polyclonal neutralizing antibodies for use in humans. Geese produce IgY and an alternatively spliced form, IgYΔFc, that can be purified at high concentrations from egg yolks. IgY lacks the properties of mammalian Fc that make antibodies produced in horses, sheep, and rabbits reactogenic in humans. Geese were vaccinated with an ANDV DNA vaccine encoding the virus envelope glycoproteins. All geese developed high-titer neutralizing antibodies after the second vaccination, and maintained high-levels of neutralizing antibodies as measured by a pseudovirion neutralization assay (PsVNA for over 1 year. A booster vaccination resulted in extraordinarily high levels of neutralizing antibodies (i.e., PsVNA80 titers >100,000. Analysis of IgY and IgYΔFc by epitope mapping show these antibodies to be highly reactive to specific amino acid sequences of ANDV envelope glycoproteins. We examined the protective efficacy of the goose-derived antibody in the hamster model of lethal HPS. α-ANDV immune sera, or IgY/IgYΔFc purified from eggs, were passively transferred to hamsters subcutaneously starting 5 days after an IM challenge with ANDV (25 LD50. Both immune sera, and egg-derived purified IgY/IgYΔFc, protected 8 of 8 and 7 of 8 hamsters, respectively. In contrast, all hamsters receiving IgY/IgYΔFc purified from normal geese (n=8, or no-treatment (n=8, developed lethal HPS. These findings demonstrate that the DNA vaccine/goose platform can be used to produce a candidate antiviral

  15. Effect of an artificial Ascaridia galli infection on egg production, immune response, and liver lipid reserve of free-range laying hens.

    Science.gov (United States)

    Sharma, N; Hunt, P W; Hine, B C; McNally, J; Sharma, N K; Iqbal, Z; Normant, C; Andronicos, N M; Swick, R A; Ruhnke, I

    2018-02-01

    This study was conducted to determine the effect of Ascaridia galli infection on free-range laying hens. Lohmann Brown laying hens (n = 200) at 17 wk of age were allocated to 4 treatment groups (n = 50 per group), each with 5 replicate pens of 10 hens. Hens in 3 treatment groups were orally inoculated with different doses of embryonated A. galli eggs: low (250 eggs), medium (1,000 eggs), and high (2,500 eggs) levels, whereas hens of the control group were not infected. Infection levels were monitored using excreta egg counts and mature A. galli worm counts in the intestine. Anti A. galli antibody titers (IgY) in the serum were measured prior to infection, and at 6, 11, 15, and 20 wk post infection (PI) and in egg yolk at 11 and 20 wk PI. Parameters evaluated included feed intake, egg production, egg weight, egg mass, FCR, liver weight, liver fat, and intra epithelial immune cell infiltration. The results showed no difference in feed intake, body weight, or FCR among any treatment groups (P > 0.05). Egg production was lower in the low infection group compared to other groups at 20 wk of age (P < 0.01). Serum IgY was higher in the infected groups' hens at 20 wk PI compared to control group hens (P < 0.01). Yolk IgY increased significantly over time and was higher in infected hens compared to hens of the control group at 11 and 20 wk PI (P < 0.001). No differences were observed in liver lipid content or intraepithelial lymphocytes infiltration among treatment groups. Ascaridia galli eggs in the coprodeum content and adult A. galli worm count were higher in infected hens compared to hens of the control group (P < 0.01). In conclusion, the effects of artificial infection with A. galli on the parameters investigated were minor, and egg yolk antibody may be a more reliable indicator of A. galli infection than serum antibody or excreta egg count. © 2017 Poultry Science Association Inc.

  16. Detection of multiple tumor markers using ultra-long carbon nanotube devices

    Science.gov (United States)

    So, Hye-Mi; Park, Dong-Won; Kim, Beom Soo; Kong, Ki-Jeong; Buh, Gyoung-Ho; Chang, Hyunju; Lee, Jeong-O.; Kong, Jing

    2008-03-01

    For the simultaneous detection of multiple tumor markers, we have fabricated ultra-long carbon nanotube sensors that can detect carcinoembryonic antigen (CEA) and prostate specific antigen (PSA), simultaneously. Ultra-long carbon nanotubes, several millimeters long, were grown by ethanol CVD, and fabricated as FET sensors by using conventional photolithography. To functionalize each segment of a single ultra-long nanotube device with multiple-tumor markers, we first functionalize the entire device with CDI-Tween 20 linking molecules, and then immobilized CEA and PSA antibodies using the microfluidic channel. The electrical conductance from CEA-antibody functionalized and PSA-antibody functionalized segment of a ultra-long carbon nanotube device was monitored simultaneously with Ag/AgCl reference electrode as a liquid gate. We will discuss the advantages of long-nanotube device in detail.

  17. Circulating antibody to myelin basic protein in relapsing-remitting multiple sclerosis

    International Nuclear Information System (INIS)

    Biggins, J.A.; Taylor, A.; Caspary, E.A.

    1978-01-01

    Sera from multiple sclerosis patients with relapsing-remitting disease and normal subjects were tested for antibody to myelin basic protein by a sensitive radioimmunoassay. The results showed a marginally decreased titre in multiple sclerosis superimposed on a seasonal variation. There was no correlation with the clinical state of the patients. Results are discussed briefly in relation to humoral antibody function in multiple sclerosis and experimental autoimmune encephalitis. (author)

  18. Validation of cosmogenic nuclide production rate scaling factors through direct measurement

    Energy Technology Data Exchange (ETDEWEB)

    Graham, I.J. E-mail: i.graham@gns.cri.nz; Barry, B.J.; Ditchburn, R.G.; Whitehead, N.E

    2000-10-01

    {sup 7}Be produced in water targets by nuclear interactions of cosmic rays has been measured to determine cosmogenic nuclide production rates as a function of altitude (sea level to 2 km) and geomagnetic latitude (20-79 deg. S). Relative intensities of low energy cosmic ray neutrons have at the same time been measured using neutron monitors based on IGY/NM-64 designed to efficiently thermalise ca. 2-30 MeV neutrons. The research is on-going and we present here preliminary data from the past two years. Water target and neutron flux results are in general agreement, and are consistent with the altitude-dependent scaling factors of Lal [Earth Planet. Sci. Lett. 104 (1991) 4241]. Significant differences between the sea level, latitude-dependent neutron flux data and Lal's predictions are possibly related to the response function of the detector.

  19. Early Japanese contributions to space weather research (1945–1960

    Directory of Open Access Journals (Sweden)

    A. Nishida

    2010-04-01

    Full Text Available Major contributions by Japanese scientists in the period of 1945 to 1960 are reviewed. This was the period when the foundation of the space weather research was laid by ground-based observations and theoretical research. Important contributions were made on such subjects as equatorial ionosphere in quiet times, tidal wind system in the ionosphere, formation of the F2 layer, VLF propagation above the ionosphere, and precursory phenomena (type IV radio outburst and polar cap absorption to storms. At the IGY (1957, 1958, research efforts were intensified and new programs in space and Antarctica were initiated. Japanese scientists in this discipline held a tight network for communication and collaboration that has been kept to this day.

  20. Validation of cosmogenic nuclide production rate scaling factors through direct measurement

    CERN Document Server

    Graham, I J; Ditchburn, R G; Whitehead, N E

    2000-01-01

    sup 7 Be produced in water targets by nuclear interactions of cosmic rays has been measured to determine cosmogenic nuclide production rates as a function of altitude (sea level to 2 km) and geomagnetic latitude (20-79 deg. S). Relative intensities of low energy cosmic ray neutrons have at the same time been measured using neutron monitors based on IGY/NM-64 designed to efficiently thermalise ca. 2-30 MeV neutrons. The research is on-going and we present here preliminary data from the past two years. Water target and neutron flux results are in general agreement, and are consistent with the altitude-dependent scaling factors of Lal [Earth Planet. Sci. Lett. 104 (1991) 4241]. Significant differences between the sea level, latitude-dependent neutron flux data and Lal's predictions are possibly related to the response function of the detector.

  1. Development of the control algorithm of processes of intensive hygrothermal impact on capillary and porous materials in the conditions of the vacuum

    Directory of Open Access Journals (Sweden)

    Larina Ludmila

    2017-01-01

    Full Text Available Objective of this research is creation of an algorithm of a control system of the modes of the intensive hygrothermal influence (IGI in the conditions of a vacuum when performing the corresponding operations: moistening; the subsequent, if necessary, cyclic drying from within preparation of top of footwear; damp thermal treatment on universal installation with adjustable parameters of a working environment. For assessment of quality of the intensified hygrothermal impact on preparations of top of footwear the integrated criteria of efficiency of processes were used. Ensuring automatic control of parameters of processes of IGV on preparations of top of footwear in universal vacuum installation will allow to control quality of preparations upon transition from performance of one operation to another according to standard manufacturing techniques of footwear.

  2. Molecular characterization and expression of the M6 gene of grass carp hemorrhage virus (GCHV), an aquareovirus.

    Science.gov (United States)

    Qiu, T; Lu, R H; Zhang, J; Zhu, Z Y

    2001-07-01

    The complete nucleotide sequence of M6 gene of grass carp hemorrhage virus (GCHV) was determined. It is 2039 nucleotides in length and contains a single large open reading frame that could encode a protein of 648 amino acids with predicted molecular mass of 68.7 kDa. Amino acid sequence comparison revealed that the protein encoded by GCHV M6 is closely related to the protein mu1 of mammalian reovirus. The M6 gene, encoding the major outer-capsid protein, was expressed using the pET fusion protein vector in Escherichia coli and detected by Western blotting using chicken anti-GCHV immunoglobulin (IgY). The result indicates that the protein encoded by M6 may share a putative Asn-42-Pro-43 proteolytic cleavage site with mu1.

  3. Short Trips and a Traverse

    Science.gov (United States)

    Cameron, Richard

    2012-01-01

    Dr. Cameron joined the Arctic Institute of North America in 1956 to participate in IGY-related activities in Antarctica. He served as Chief Glaciologist at Wilkes Station, on the coast of East Antarctica. This was a joint Navy-civilian operation consisting of 17 Navy personnel and 10 scientists. Specifically, his glaciological team consisted of two colleagues with whom he had worked before - Olav Loken in Norway in the summer of 1953, and John Molholm in Greenland in the summer of 1954. This team spent much of its time at a remote station established 80 kilometers (50 miles) inland, where they conducted both meteorological and glaciological studies. One of the glaciological studies entailed digging a 35-meter (approx.115-foot) vertical pit to study snow densification and stratigraphy. The assignment for the Navy Seabees was to first establish a joint US-NZ base at Cape Hallett and then go along the coast of East Antarctica and set up Wilkes Station.

  4. Oblique Multi-Camera Systems - Orientation and Dense Matching Issues

    Science.gov (United States)

    Rupnik, E.; Nex, F.; Remondino, F.

    2014-03-01

    The use of oblique imagery has become a standard for many civil and mapping applications, thanks to the development of airborne digital multi-camera systems, as proposed by many companies (Blomoblique, IGI, Leica, Midas, Pictometry, Vexcel/Microsoft, VisionMap, etc.). The indisputable virtue of oblique photography lies in its simplicity of interpretation and understanding for inexperienced users allowing their use of oblique images in very different applications, such as building detection and reconstruction, building structural damage classification, road land updating and administration services, etc. The paper reports an overview of the actual oblique commercial systems and presents a workflow for the automated orientation and dense matching of large image blocks. Perspectives, potentialities, pitfalls and suggestions for achieving satisfactory results are given. Tests performed on two datasets acquired with two multi-camera systems over urban areas are also reported.

  5. Proposing co-design of personas as a method to heighten validity and engage users

    DEFF Research Database (Denmark)

    Albrechtsen, Charlotte; Pedersen, Majbrit; Pedersen, Nicholai Friis

    2016-01-01

    This paper proposes co-designing personas with users as a strategy to overcome a challenge inherent in the design of personas or fictitious users: On one hand, personas should appear realistic and believable as individuals, and on the other hand, personas should represent a broader range of users....... By involving empirical users in all parts of the process of persona design, the risk of creating personas that are too stereotypical is minimized, as the participating users enrich the data on which the personas are based with up-to-date and firsthand contextual knowledge. Advantages of co-designing personas...... with users is illustrated by a case from higher education in which personas were co-designed with students as part of a project aiming at designing a smartphone application for Master's thesis students. © 2016, IGI Global....

  6. Towards freeform curved blazed gratings using diamond machining

    Science.gov (United States)

    Bourgenot, C.; Robertson, D. J.; Stelter, D.; Eikenberry, S.

    2016-07-01

    Concave blazed gratings greatly simplify the architecture of spectrographs by reducing the number of optical components. The production of these gratings using diamond-machining offers practically no limits in the design of the grating substrate shape, with the possibility of making large sag freeform surfaces unlike the alternative and traditional method of holography and ion etching. In this paper, we report on the technological challenges and progress in the making of these curved blazed gratings using an ultra-high precision 5 axes Moore-Nanotech machine. We describe their implementation in an integral field unit prototype called IGIS (Integrated Grating Imaging Spectrograph) where freeform curved gratings are used as pupil mirrors. The goal is to develop the technologies for the production of the next generation of low-cost, compact, high performance integral field unit spectrometers.

  7. OBLIQUE MULTI-CAMERA SYSTEMS – ORIENTATION AND DENSE MATCHING ISSUES

    Directory of Open Access Journals (Sweden)

    E. Rupnik

    2014-03-01

    Full Text Available The use of oblique imagery has become a standard for many civil and mapping applications, thanks to the development of airborne digital multi-camera systems, as proposed by many companies (Blomoblique, IGI, Leica, Midas, Pictometry, Vexcel/Microsoft, VisionMap, etc.. The indisputable virtue of oblique photography lies in its simplicity of interpretation and understanding for inexperienced users allowing their use of oblique images in very different applications, such as building detection and reconstruction, building structural damage classification, road land updating and administration services, etc. The paper reports an overview of the actual oblique commercial systems and presents a workflow for the automated orientation and dense matching of large image blocks. Perspectives, potentialities, pitfalls and suggestions for achieving satisfactory results are given. Tests performed on two datasets acquired with two multi-camera systems over urban areas are also reported.

  8. Oscillation and nonoscillation of solutions to even order self-adjoint differential equations

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    Ondrej Dosly

    2003-11-01

    Full Text Available We establish oscillation and nonoscilation criteria for the linear differential equation $$ (-1^nig(t^alpha y^{(n}ig^{(n}- frac{gamma_{n,alpha}}{t^{2n-alpha}}y=q(ty,quad alpha otin {1, 3, dots , 2n-1}, $$ where $$ gamma_{n,alpha}=frac{1}{4^n}prod_{k=1}^n(2k-1-alpha^2 $$ and $q$ is a real-valued continuous function. It is proved, using these criteria, that the equation $$ (-1^nig(t^alpha y^{(n}ig^{(n} -ig(frac{gamma_{n,alpha}}{t^{2n-alpha}} + frac{gamma}{t^{2n-alpha}lg^2 t}igy = 0 $$ is nonoscillatory if and only if $$ gamma leq ilde gamma_{n,alpha}:= frac{1}{4^n}prod_{k=1}^n(2k-1-alpha^2 sum_{k=1}^nfrac{1}{(2k-1-alpha^2}. $$

  9. A Small-Unit Amphibious Operation Combat Model.

    Science.gov (United States)

    1983-03-01

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  10. Evaluation of fasting state-/oral glucose tolerance test-derived measures of insulin release for the detection of genetically impaired β-cell function.

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    Silke A Herzberg-Schäfer

    Full Text Available BACKGROUND: To date, fasting state- and different oral glucose tolerance test (OGTT-derived measures are used to estimate insulin release with reasonable effort in large human cohorts required, e.g., for genetic studies. Here, we evaluated twelve common (or recently introduced fasting state-/OGTT-derived indices for their suitability to detect genetically determined β-cell dysfunction. METHODOLOGY/PRINCIPAL FINDINGS: A cohort of 1364 White European individuals at increased risk for type 2 diabetes was characterized by OGTT with glucose, insulin, and C-peptide measurements and genotyped for single nucleotide polymorphisms (SNPs known to affect glucose- and incretin-stimulated insulin secretion. One fasting state- and eleven OGTT-derived indices were calculated and statistically evaluated. After adjustment for confounding variables, all tested SNPs were significantly associated with at least two insulin secretion measures (p≤0.05. The indices were ranked according to their associations' statistical power, and the ranks an index obtained for its associations with all the tested SNPs (or a subset were summed up resulting in a final ranking. This approach revealed area under the curve (AUC(Insulin(0-30/AUC(Glucose(0-30 as the best-ranked index to detect SNP-dependent differences in insulin release. Moreover, AUC(Insulin(0-30/AUC(Glucose(0-30, corrected insulin response (CIR, AUC(C-Peptide(0-30/AUC(Glucose(0-30, AUC(C-Peptide(0-120/AUC(Glucose(0-120, two different formulas for the incremental insulin response from 0-30 min, i.e., the insulinogenic indices (IGI(2 and IGI(1, and insulin 30 min were significantly higher-ranked than homeostasis model assessment of β-cell function (HOMA-B; p<0.05. AUC(C-Peptide(0-120/AUC(Glucose(0-120 was best-ranked for the detection of SNPs involved in incretin-stimulated insulin secretion. In all analyses, HOMA-β displayed the highest rank sums and, thus, scored last. CONCLUSIONS/SIGNIFICANCE: With AUC(Insulin(0

  11. Immune responses of a native and an invasive bird to Buggy Creek Virus (Togaviridae: Alphavirus and its arthropod vector, the swallow bug (Oeciacus vicarius.

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    Carol A Fassbinder-Orth

    Full Text Available Invasive species often display different patterns of parasite burden and virulence compared to their native counterparts. These differences may be the result of variability in host-parasite co-evolutionary relationships, the occurrence of novel host-parasite encounters, or possibly innate differences in physiological responses to infection between invasive and native hosts. Here we examine the adaptive, humoral immune responses of a resistant, native bird and a susceptible, invasive bird to an arbovirus (Buggy Creek virus; Togaviridae: Alphavirus and its ectoparasitic arthropod vector (the swallow bug; Oeciacus vicarius. Swallow bugs parasitize the native, colonially nesting cliff swallow (Petrochelidon pyrrhonota and the introduced house sparrow (Passer domesticus that occupies nests in cliff swallow colonies. We measured levels of BCRV-specific and swallow bug-specific IgY levels before nesting (prior to swallow bug exposure and after nesting (after swallow bug exposure in house sparrows and cliff swallows in western Nebraska. Levels of BCRV-specific IgY increased significantly following nesting in the house sparrow but not in the cliff swallow. Additionally, house sparrows displayed consistently higher levels of swallow bug-specific antibodies both before and after nesting compared to cliff swallows. The higher levels of BCRV and swallow bug specific antibodies detected in house sparrows may be reflective of significant differences in both antiviral and anti-ectoparasite immune responses that exist between these two avian species. To our knowledge, this is the first study to compare the macro- and microparasite-specific immune responses of an invasive and a native avian host exposed to the same parasites.

  12. Neuroinvasion of the highly pathogenic influenza virus H7N1 is caused by disruption of the blood brain barrier in an avian model.

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    Aida J Chaves

    Full Text Available Influenza A virus (IAV causes central nervous system (CNS lesions in avian and mammalian species, including humans. However, the mechanism used by IAV to invade the brain has not been determined. In the current work, we used chickens infected with a highly pathogenic avian influenza (HPAI virus as a model to elucidate the mechanism of entry of IAV into the brain. The permeability of the BBB was evaluated in fifteen-day-old H7N1-infected and non-infected chickens using three different methods: (i detecting Evans blue (EB extravasation into the brain, (ii determining the leakage of the serum protein immunoglobulin Y (IgY into the brain and (iii assessing the stability of the tight-junction (TJ proteins zonula occludens-1 and claudin-1 in the chicken brain at 6, 12, 18, 24, 36 and 48 hours post-inoculation (hpi. The onset of the induced viremia was evaluated by quantitative real time RT-PCR (RT-qPCR at the same time points. Viral RNA was detected from 18 hpi onward in blood samples, whereas IAV antigen was detected at 24 hpi in brain tissue samples. EB and IgY extravasation and loss of integrity of the TJs associated with the presence of viral antigen was first observed at 36 and 48 hpi in the telencephalic pallium and cerebellum. Our data suggest that the mechanism of entry of the H7N1 HPAI into the brain includes infection of the endothelial cells at early stages (24 hpi with subsequent disruption of the TJs of the BBB and leakage of virus and serum proteins into the adjacent neuroparenchyma.

  13. Produção e purificação de imunoglobulinas Y policlonais anti-Leptospira spp.

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    Tatiane C.F. Tavares

    2013-09-01

    Full Text Available Objetivou-se verificar se galinhas imunizadas com uma solução de Leptospira interrogans inativadas e proteínas de membrana externa do sorovar Hardjo, poderiam produzir anticorpos policlonais específicos anti-leptospiras, detectáveis em testes ELISA. Foram imunizados oito galinhas com 25 semanas de idade, da raça White Leghorn, sendo três imunizadas com uma suspensão de leptospiras inativadas, três com uma solução de proteínas de membrana externa extraída do sorovar Hardjo e duas controle. Coletas de sangue foram realizadas quinzenalmente e de ovos diariamente. A IgY foi purificada a partir da gema dos ovos utilizando para a delipidação o método de diluição em água ácida e a precipitação com sulfato de amônio. Nos testes ELISA realizados para verificar a especificidade da IgY, foi demonstrada a produção de anticorpos anti-Leptospira, tanto no soro quanto nas gemas purificadas. O pico de produção de anticorpos específicos ocorreu na 5º semana após a primeira imunização. Ficou demonstrada a possibilidade da indução da produção de anticorpos específicos em galinhas imunizadas com leptospiras do sorovar Hardjo inativadas, bem como, com proteínas de membrana externa (PME extraidas desse sorovar. As galinhas imunizadas com uma suspensão de leptospiras inativadas ou com PME de Leptospira interrogans do sorovar Hardjo produziram anticorpos reativos a PME Hardjo detectáves por teste ELISA.

  14. Retrospective

    Science.gov (United States)

    Brooks, David A.

    Charting a course toward an uncertain future is always a risky business, especially among shoals of fiscal restraint or national tragedy, and the prudent navigator is well advised to remember where he's been as he looks ahead. The ocean and space sciences are poised for grand joint adventures, but shrinking budgets and the lingering Challenger numbness are restrictive lee shores that must be considered when laying plans. To sharpen the focus on future choices, it may be helpful to glance in the geophysical rearview mirror and remember some of the challenges and opportunities of a different era.A quarter century is a long time, but many images from 25 years ago can still be recalled in crisp detail, like photographs in a scrapbook. In 1961, results from the International Geophysical Year (IGY) filled the pages of the Transactions of the American Geophysical Union, and the U.S. program of space exploration finally was underway with conviction. The Indian Ocean Expedition, conceived during the IGY, ushered in a new era of international oceanography. The TIROS III satellite beamed to earth fuzzy pictures of tropical storms and revealed the intricate writhings of the Gulf Stream. Forecasters and fluid dynamicists suddenly saw new horizons, and geophysical turbulence became a major topic at the IUGG Symposium in Marseilles, France. Papers with prescient themes were presented at the AGU Ocean Section meeting: June Pattullo (then at Oregon State College, Corvallis) on heat storage in the Pacific; Ferris Webster (then at Woods Hole Oceanographic Institution, Woods Hole, Mass.) on Gulf Stream meanders. Polar oceanography was well represented in AGU journals: Kenneth Hunkins (at what was then called the Lamont Geological Observatory, Palisades, N.Y.) described the Alpha Rise, discovered from a drifting Arctic ice island, and Edward Thiel (then at the University of Minnesota, Minneapolis) and his co-workers discussed open ocean tides, gravimetrically measured from Antarctic

  15. Food availability and maternal immunization affect transfer and persistence of maternal antibodies in nestling pigeons.

    Directory of Open Access Journals (Sweden)

    Ahmad Ismail

    Full Text Available The ability of mothers to transfer antibodies (Abs to their young and the temporal persistence of maternal Abs in offspring constitute important life-history traits that can impact the evolution of host-parasite interactions. Here, we examined the effects of food availability and parental immunization on the transfer and persistence of maternal antibodies in nestling pigeons (Columba livia. This species can transmit maternal Abs to offspring before hatching through the egg yolk and potentially after hatching through crop milk. However, the role of this postnatal substance in immunity remains elusive. We used a full cross-fostering design to disentangle the effects of food limitation and parental immunization both before and after hatching on the levels and persistence of maternal Abs in chicks. Parents were immunized via injection with keyhole limpet hemocyanin antigens. Using an immunoassay that specifically detected the IgY antibodies that are known to be transmitted via the yolk, we found that the levels of anti-KLH Abs in newly hatched chicks were positively correlated with the levels of anti-KLH Abs in the blood of their biological mothers. However, this correlation was not present between chicks and their foster parents, suggesting limited IgY transfer via crop milk to the chick's bloodstream. Interestingly, biological mothers subjected to food limitation during egg laying transferred significantly fewer specific maternal Abs, which suggests that the transfer of antibodies might be costly for them. In addition, the persistence of maternal Abs in a chick's bloodstream was not affected by food limitation or the foster parents' anti-KLH Ab levels; it was only affected by the initial level of maternal anti-KLH Abs that were present in newly hatched chicks. These results suggest that the maternal transfer of Abs could be costly but that their persistence in an offspring's bloodstream may not necessarily be affected by environmental conditions.

  16. Association of NEFA composition with insulin sensitivity and beta cell function in the Prospective Metabolism and Islet Cell Evaluation (PROMISE) cohort.

    Science.gov (United States)

    Johnston, Luke W; Harris, Stewart B; Retnakaran, Ravi; Giacca, Adria; Liu, Zhen; Bazinet, Richard P; Hanley, Anthony J

    2018-04-01

    Our aim was to determine the longitudinal associations of individual NEFA with the pathogenesis of diabetes, specifically with differences in insulin sensitivity and beta cell function over 6 years in a cohort of individuals who are at risk for diabetes. In the Prospective Metabolism and Islet Cell Evaluation (PROMISE) longitudinal cohort, 477 participants had serum NEFA measured at the baseline visit and completed an OGTT at three time points over 6 years. Outcome variables were calculated using the OGTT values. At each visit, insulin sensitivity was assessed using the HOMA2 of insulin sensitivity (HOMA2-%S) and the Matsuda index, while beta cell function was assessed using the insulinogenic index over HOMA-IR (IGI/IR) and the insulin secretion-sensitivity index-2 (ISSI-2). Generalised estimating equations were used, adjusting for time, waist, sex, ethnicity, baseline age, alanine aminotransferase (ALT) and physical activity. NEFA were analysed as both concentrations (nmol/ml) and proportions (mol%) of the total fraction. Participants' (73% female, 70% with European ancestry) insulin sensitivity and beta cell function declined by 14-21% over 6 years of follow-up. In unadjusted models, several NEFA (e.g. 18:1 n-7, 22:4 n-6) were associated with lower insulin sensitivity, however, nearly all of these associations were attenuated in fully adjusted models. In adjusted models, total NEFA, 16:0, 18:1 n-9 and 18:2 n-6 (as concentrations) were associated with 3.7-8.0% lower IGI/IR and ISSI-2, while only 20:5 n-3 (as mol%) was associated with 7.7% higher HOMA2-%S. Total NEFA concentration was a strong predictor of lower beta cell function over 6 years. Our results suggest that the association with beta cell function is due to the absolute size of the serum NEFA fraction, rather than the specific fatty acid composition.

  17. High Serum Pepsinogen I and beta Helicobacter pylori Infection Are Risk Factors for Aspirin-Induced Gastroduodenal Injury.

    Science.gov (United States)

    Shan, Jing; Lei, Hongjun; Shi, Wei; Sun, Xiaobin; Tang, Yu; Ren, Chunrong

    2018-01-01

    Whether gastric hyperchlorhydria and Helicobacter pylori infection contribute to aspirin-induced gastroduodenal injury still lacks evidence. Because serum pepsinogens (PGs) and gastrin-17 (G17) can reflect gastric acid secretion, this study intended to elucidate whether serum PGs, serum G17, and H. pylori infection are associated with aspirin-induced gastrointestinal injury. A total of 60 patients taking low-dose aspirin for more than 1 month were enrolled in this study. Serum PG I, PG II, and G17 were determined using ELISA. A 14C-urea breath test was used for the detection of an H. pylori infection. The modified Lanza score was used to evaluate the degree of gastroduodenal injury under endoscopy. The median serum PG I level was significantly higher in the intensive gastroduodenal injury (IGI) group compared to that in the mild gastroduodenal injury group (155.0 vs. 116.6 ng/mL, p = 0.006). The H. pylori infection rate was significantly higher in the IGI group (73 vs. 40%, p = 0.037). Receiver operator characteristic curves analysis revealed that the cutoff value of PG I was 123 ng/mL, with 80% sensitivity and 61.4% specificity. H. pylori infection combined with PG I at >123 ng/mL had an OR (95% CI) of 15.8 (2.4 ± 104.5) for the prediction of aspirin-induced gastroduodenal injury. Key Messages: Serum PG I and H. pylori infection could be used to identify potential high-risk aspirin-induced gastroduodenal injury patients. © 2017 S. Karger AG, Basel.

  18. Immunologic responses in corn snakes (Pantherophis guttatus) after experimentally induced infection with ferlaviruses.

    Science.gov (United States)

    Neul, Annkatrin; Schrödl, Wieland; Marschang, Rachel E; Bjick, Tina; Truyen, Uwe; von Buttlar, Heiner; Pees, Michael

    2017-04-01

    OBJECTIVE To measure immunologic responses of snakes after experimentally induced infection with ferlaviruses. ANIMALS 42 adult corn snakes (Pantherophis guttatus) of both sexes. PROCEDURES Snakes were inoculated intratracheally with genogroup A (n = 12), B (12), or C (12) ferlavirus (infected groups) or cell-culture supernatant (6; control group) on day 0. Three snakes from each infected group were euthanized on days 4, 16, 28, and 49, and 3 snakes from the control group were euthanized on day 49. Blood samples were collected from live snakes on days -6 (baseline), 4, 16, 28, and 49. Hematologic tests were performed and humoral responses assessed via hemagglutination-inhibition assays and ELISAs. Following euthanasia, gross pathological and histologic evaluations and virus detection were performed. RESULTS Severity of clinical signs of and immunologic responses to ferlavirus infection differed among snake groups. Hematologic values, particularly WBC and monocyte counts, increased between days 4 and 16 after infection. A humoral response was identified between days 16 and 28. Serum IgM concentrations increased from baseline earlier than IgY concentrations, but the IgY relative increase was higher at the end of the study. The hemagglutination-inhibition assay revealed that the strongest reactions in all infected groups were against the strain with which they had been infected. Snakes infected with genogroup A ferlavirus had the strongest immune response, whereas those infected with genogroup B had the weakest responses. CONCLUSIONS AND CLINICAL RELEVANCE Results of this experimental study suggested that the ferlavirus strain with the highest virulence induced the weakest immune response in snakes.

  19. Controlling feeding behavior by chemical or gene-directed targeting in the brain: What’s so spatial about our methods?

    Directory of Open Access Journals (Sweden)

    Arshad M Khan

    2013-12-01

    Full Text Available Intracranial chemical injection (ICI methods have been used to identify the locations in the brain where feeding behavior can be controlled acutely. Scientists conducting ICI studies often document their injection site locations, thereby leaving kernels of valuable location data for others to use to further characterize feeding control circuits. Unfortunately, this rich dataset has not yet been formally contextualized with other published neuroanatomical data. In particular, axonal tracing studies have delineated several neural circuits originating in the same areas where ICI injection feeding-control sites have been documented, but it remains unclear whether these circuits participate in feeding control. However, comparing injection sites with other types of location data requires careful anatomical registration between the datasets. Here, a conceptual framework is presented for how such anatomical registration efforts can be performed. For example, by using a simple atlas alignment tool, a hypothalamic locus sensitive to the orexigenic effects of neuropeptide Y (NPY can be aligned accurately with the locations of neurons labeled by anterograde tracers or those known to express NPY receptors or feeding-related peptides. This approach can also be applied to those intracranial gene-directed injection (IGI methods (e.g., site-specific recombinase methods, RNA expression or interference, optogenetics and pharmacosynthetics that involve viral injections to targeted neuronal populations. Spatial alignment efforts can be accelerated if location data from ICI/IGI methods are mapped to stereotaxic brain atlases to allow powerful neuroinformatics tools to overlay different types of data in the same reference space. Atlas-based mapping will be critical for community-based sharing of location data for feeding control circuits, and will accelerate our understanding of structure-function relationships in the brain for mammalian models of obesity and

  20. The Antarctic Master Directory - a fundamental data management element for the International Polar Year 2007-2008

    Science.gov (United States)

    Scharfen, G.; Bauer, R.

    2004-12-01

    A successful International Polar Year (IPY) in 2007-2008 will extend the scientific spirit of international collaboration and exploration first undertaken in earlier IPYs and the 1957/58 International Geophysical Year (IGY) to the current era of advanced collection and analysis technology. The IGY not only led to a number of important scientific achievements; it also established an enduring data system - the World Data Centers - which continues today. Effective utilization of the vast arrays of data which will result from the coming IPY will challenge data managers and scientists alike. Coordinating the collection, assembly, archival and international exchange of disparate and voluminous data sets requires advance planning and the involvement of the relevant science agencies and data managers to utilize and extend existing capabilities. The IPY Planning Group has identified key objectives indicating that data management is an essential part of the IPY planning process including: - Ensure data collected under the IPY are made available in an open and timely manner - Intensify the recovery of relevant historical data and ensure that these also are made openly available - Develop and embrace new technological and logistical capabilities The Scientific Committee on Antarctic Research (SCAR) and Committee of Managers of National Antarctic Programmes (COMNAP) have established the Joint Committee on Antarctic Data Management (JCADM) to develop the Antarctic Master Directory (AMD) to enable scientists to find and access the data sets collected more than 22 countries in the Antarctic. Incorporating concepts developed as part of the AMD and extending them to cover the scope of the IPY is an important part of a successful IPY data management program. This paper identifies major aspects of the AMD and how it can serve the IPY.

  1. Natural History of Impaired Glucose Tolerance in Japanese Americans: Change in Visceral Adiposity is Associated with Remission from Impaired Glucose Tolerance to Normal Glucose Tolerance.

    Science.gov (United States)

    Onishi, Yukiko; Hayashi, Tomoshige; Sato, Kyoko K; Leonetti, Donna L; Kahn, Steven E; Fujimoto, Wilfred Y; Boyko, Edward J

    2018-05-30

    To describe the roles of intra-abdominal fat and its change in the remission of impaired glucose tolerance (IGT) to normal glucose tolerance (NGT). We followed 157 Japanese Americans with IGT at baseline for 10-11 years without external intervention. We measured intra-abdominal and abdominal subcutaneous fat area (IAFA and ASFA) by computed tomography at baseline and at 5-6 years of follow-up. Change in IAFA and ASFA (ΔIAFA and ΔASFA) were calculated by subtracting baseline fat area from 5-6 year follow-up fat area. Glucose and insulin at fasting and during a 75-g oral glucose tolerance test, insulinogenic index (IGI [Δinsulin/Δglucose (30-0 min)]) and homeostasis model assessment for insulin resistance (HOMA-IR) were measured at baseline. Fourty-four subjects remitted to NGT. Among those with lower IAFA (≤median 91.31 cm 2 ) and the lowest tertile of ΔIAFA, 45% remitted, while with higher IAFA (>91.31 cm 2 ) and the highest tertile of ΔIAFA, only 12.5% remitted. ΔIAFA was significantly associated with remission to NGT (multiple-adjusted odd ratio [1-SD decrease] 1.93, 95% CI 1.10-3.36) independent of IAFA, ASFA, ΔASFA, IGI, HOMA-IR, age, sex, and family history of diabetes. In the natural history of IGT, change in intra-abdominal fat was associated with remission to NGT. Copyright © 2018. Published by Elsevier B.V.

  2. Oral peptide specific egg antibody to intestinal sodium-dependent phosphate co-transporter-2b is effective at altering phosphate transport in vitro and in vivo.

    Science.gov (United States)

    Bobeck, Elizabeth A; Hellestad, Erica M; Sand, Jordan M; Piccione, Michelle L; Bishop, Jeff W; Helvig, Christian; Petkovich, Martin; Cook, Mark E

    2015-06-01

    Hyperimmunized hens are an effective means of generating large quantities of antigen specific egg antibodies that have use as oral supplements. In this study, we attempted to create a peptide specific antibody that produced outcomes similar to those of the human pharmaceutical, sevelamer HCl, used in the treatment of hyperphosphatemia (a sequela of chronic renal disease). Egg antibodies were generated against 8 different human intestinal sodium-dependent phosphate cotransporter 2b (NaPi2b) peptides, and hNaPi2b peptide egg antibodies were screened for their ability to inhibit phosphate transport in human intestinal Caco-2 cell line. Antibody produced against human peptide sequence TSPSLCWT (anti-h16) was specific for its peptide sequence, and significantly reduced phosphate transport in human Caco-2 cells to 25.3±11.5% of control nonspecific antibody, when compared to nicotinamide, a known inhibitor of phosphate transport (P≤0.05). Antibody was then produced against the mouse-specific peptide h16 counterpart (mouse sequence TSPSYCWT, anti-m16) for further analysis in a murine model. When anti-m16 was fed to mice (1% of diet as dried egg yolk powder), egg yolk immunoglobulin (IgY) was detected using immunohistochemical staining in mouse ileum, and egg anti-m16 IgY colocalized with a commercial goat anti-NaPi2b antibody. The effectiveness of anti-m16 egg antibody in reducing serum phosphate, when compared to sevelamer HCl, was determined in a mouse feeding study. Serum phosphate was reduced 18% (Pegg yolk powder) and 30% (Pegg immunoglobulin. The methods described and the findings reported show that oral egg antibodies are useful and easy to prepare reagents for the study and possible treatment of select diseases. © 2015 Poultry Science Association Inc.

  3. In vitro study of protein release from AFCo1 and implications in mucosal immunisation

    Directory of Open Access Journals (Sweden)

    José Raúl Dopico

    2012-04-01

    Full Text Available Los anticuerpos aviares (IgY presentan algunas ventajas con relación a los anticuerpos IgG de mamíferos, debido a su fácil obtención y purificación y su bajo costo de producción. El objetivo de este trabajo fue estudiar la factibilidad de acoplar anticuerpos IgY a partículas de poliestireno y evaluar su desempeño en ensayos de látex-aglutinación en lámina. Para este propósito se utilizó como modelo la detección del antígeno de superficie de la hepatitis B (HBsAg. Gallinas Leghorn se inmunizaron con el ingrediente farmacéutico activo que se utiliza en la elaboración de la vacuna recombinante cubana Heberbiovac-HB. Los anticuerpos obtenidos se purificaron y emplearon en la preparación de reactivos de látex-aglutinación (0,8 µm para la detección del HBsAg, los cuales se evaluaron con 50 muestras de suero frente a un reactivo comercial similar. Los resultados se evaluaron de satisfactorios. Se demostró la factibilidad de acoplar anticuerpos aviares a partículas de látex y se abre una perspectiva al empleo de este tipo de ensayo para el diagnóstico rápido de diversas enfermedades.

  4. The CU 2-D-MAX-DOAS instrument – Part 2: Raman scattering probability measurements and retrieval of aerosol optical properties

    Energy Technology Data Exchange (ETDEWEB)

    Ortega, Ivan; Coburn, Sean; Berg, Larry K.; Lantz, Kathy; Michalsky, Joseph; Ferrare, Richard A.; Hair, Johnathan W.; Hostetler, Chris A.; Volkamer, Rainer

    2016-01-01

    The multiannual global mean of aerosol optical depth at 550 nm (AOD550) over land is ~0.19, and that over oceans is ~0.13. About 45 % of the Earth surface shows AOD550 smaller than 0.1. There is a need for measurement techniques that are optimized to measure aerosol optical properties under low AOD conditions. We present an inherently calibrated retrieval (i.e., no need for radiance calibration) to simultaneously measure AOD and the aerosol phase function parameter, <i>g>, based on measurements of azimuth distributions of the Raman scattering probability (RSP), the near-absolute rotational Raman scattering (RRS) intensity. We employ radiative transfer model simulations to show that for solar azimuth RSP measurements at solar elevation and solar zenith angle (SZA) smaller than 80°, RSP is insensitive to the vertical distribution of aerosols and maximally sensitive to changes in AOD and <i>g> under near-molecular scattering conditions. The University of Colorado two-dimensional Multi-AXis Differential Optical Absorption Spectroscopy (CU 2-D-MAX-DOAS) instrument was deployed as part of the Two Column Aerosol Project (TCAP) at Cape Cod, MA, during the summer of 2012 to measure direct sun spectra and RSP from scattered light spectra at solar relative azimuth angles (SRAAs) between 5 and 170°. During two case study days with (1) high aerosol load (17 July, 0.3 < AOD430 < 0.6) and (2) near-molecular scattering conditions (22 July, AOD430 < 0.13) we compare RSP-based retrievals of AOD430 and <i>g> with data from a co-located CIMEL sun photometer, Multi-Filter Rotating Shadowband Radiometer (MFRSR), and an airborne High Spectral Resolution Lidar (HSRL-2). The average difference (relative to DOAS) for AOD430 is +0.012 ± 0.023 (CIMEL), -0.012 ± 0.024 (MFRSR), -0.011 ± 0.014 (HSRL-2), and +0.023 ± 0.013 (CIMELAOD - MFRSRAOD) and yields the following

  5. Optimized preparation and preliminary evaluation of [64Cu]-DOTA-trastuzumab for targeting ErbB2/Neu expression

    International Nuclear Information System (INIS)

    Behrooz Alirezapour; Mohammad Javad Rasaee

    2013-01-01

    , spleen, kidney and other tissues demonstrates a similar pattern to the other radiolabeled anti-HER2 immunoconjugates. 64 Cu-DOTA-trastuzumab is a potential compound for molecular imaging of PET for diagnosis and treatment studies and follow-up of HER2 expression in oncology. (author)

  6. HER2 induced EMT and tumorigenicity in breast epithelial progenitor cells is inhibited by coexpression of EGFR.

    Science.gov (United States)

    Ingthorsson, S; Andersen, K; Hilmarsdottir, B; Maelandsmo, G M; Magnusson, M K; Gudjonsson, T

    2016-08-11

    The members of the epidermal growth factor receptor (EGFR) kinase family are important players in breast morphogenesis and cancer. EGFR2/HER2 and EGFR expression have a prognostic value in certain subtypes of breast cancer such as HER2-amplified, basal-like and luminal type B. Many clinically approved small molecular inhibitors and monoclonal antibodies have been designed to target HER2, EGFR or both. There is, however, still limited knowledge on how the two receptors are expressed in normal breast epithelium, what effects they have on cellular differentiation and how they participate in neoplastic transformation. D492 is a breast epithelial cell line with stem cell properties that can undergo epithelial to mesenchyme transition (EMT), generate luminal- and myoepithelial cells and form complex branching structures in three-dimensional (3D) culture. Here, we show that overexpression of HER2 in D492 (D492(HER2)) resulted in EMT, loss of contact growth inhibition and increased oncogenic potential in vivo. HER2 overexpression, furthermore, inhibited endogenous EGFR expression. Re-introducing EGFR in D492(HER2) (D492(HER2/EGFR)) partially reversed the mesenchymal state of the cells, as an epithelial phenotype reappeared both in 3D cultures and in vivo. The D492(HER2/EGFR) xenografts grow slower than the D492(HER2) tumors, while overexpression of EGFR alone (D492(EGFR)) was not oncogenic in vivo. Consistent with the EGFR-mediated epithelial phenotype, overexpression of EGFR drove the cells toward a myoepithelial phenotype in 3D culture. The effect of two clinically approved anti-HER2 and EGFR therapies, trastuzumab and cetuximab, was tested alone and in combination on D492(HER2) xenografts. While trastuzumab had a growth inhibitory effect compared with untreated control, the effect of cetuximab was limited. When administered in combination, the growth inhibitory effect of trastuzumab was less pronounced. Collectively, our data indicate that in HER2-overexpressing D492

  7. Distinct distribution and prognostic significance of molecular subtypes of breast cancer in Chinese women: a population-based cohort study

    Directory of Open Access Journals (Sweden)

    Cai Qiuyin

    2011-07-01

    detection and anti-HER2 therapy to potentially benefit a high proportion of breast cancer patients in China.

  8. A Qualitative Study Comparing the Assay Performance Characteristics Between the 2007 and the 2013 American Society for Clinical Oncology and College of American Pathologists HER2 Scoring Methods in Mucinous Epithelial Ovarian Cancer

    Science.gov (United States)

    Chen, Chi-Kuan; Lee, Ming-Yung; Lin, Wea-Lung; Wang, Yu-Ting; Han, Chih-Ping; Yu, Cheng-Ping; Chao, Wan-Ru

    2014-01-01

    Abstract The remarkable success of trastuzumab and other newly developed anti-HER2 (human epidermal growth factor receptor 2) therapies in breast, gastric, or gastroesophageal junction cancer patients has supported us to investigate the HER2 status and its possible therapeutic implication in mucinous epithelial ovarian cancer (EOC). However, there is currently no standardization of HER2 scoring criteria in mucinous EOC. In this study, we aimed to compare both the assay performance characteristics of the 2007 and the 2013 American Society for Clinical Oncology and College of American Pathologists scoring methods. Forty-nine tissue microarray samples of mucinous EOC from Asian women were analyzed by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) tests using the 2007 and the 2013 criteria, respectively. The overall concordance between IHC and FISH by the 2007 criteria was 97.92 % (kappa = 0.921), and that by the 2013 criteria was 100% (kappa = 1.000). The percentage of Her2 FISH-amplified cases showed an increasing trend significantly through their corresponding HER2 IHC ordinals by the 2007 and the 2013 criteria, respectively (P < 0.001, P < 0.001). After excluding equivocal cases, the specificity (100%) and positive predictive value (100%) were unchanged under either the 2007 or the 2013 criteria. The sensitivity (100%), negative predictive value (NPV) (100%), and accuracy (100%) of HER2 IHC were higher under the 2013 criteria than those (sensitivity 87.5%, NPV 97.6%, and accuracy 97.9%) under the 2007 criteria. Of the total 49 cases, the number (n = 4) of HER2 IHC equivocal results under the 2013 criteria was 4-fold higher than that (n = 1) under the 2007 criteria (8.16% vs 2.04%). Conclusively, if first tested by IHC, the 2013 criteria caused more equivocal HER2 IHC cases to be referred to Her2 FISH testing than the 2007 criteria. That decreased the false-negative rate of HER2 status and increased the detection

  9. Amplification and high-level expression of heat shock protein 90 marks aggressive phenotypes of human epidermal growth factor receptor 2 negative breast cancer.

    Science.gov (United States)

    Cheng, Qing; Chang, Jeffrey T; Geradts, Joseph; Neckers, Leonard M; Haystead, Timothy; Spector, Neil L; Lyerly, H Kim

    2012-04-17

    Although human epidermal growth factor receptor 2 (HER2) positive or estrogen receptor (ER) positive breast cancers are treated with clinically validated anti-HER2 or anti-estrogen therapies, intrinsic and acquired resistance to these therapies appears in a substantial proportion of breast cancer patients and new therapies are needed. Identification of additional molecular factors, especially those characterized by aggressive behavior and poor prognosis, could prioritize interventional opportunities to improve the diagnosis and treatment of breast cancer. We compiled a collection of 4,010 breast tumor gene expression data derived from 23 datasets that have been posted on the National Center for Biotechnology Information (NCBI) Gene Expression Omnibus (GEO) database. We performed a genome-scale survival analysis using Cox-regression survival analyses, and validated using Kaplan-Meier Estimates survival and Cox Proportional-Hazards Regression survival analyses. We conducted a genome-scale analysis of chromosome alteration using 481 breast cancer samples obtained from The Cancer Genome Atlas (TCGA), from which combined expression and copy number data were available. We assessed the correlation between somatic copy number alterations and gene expression using analysis of variance (ANOVA). Increased expression of each of the heat shock protein (HSP) 90 isoforms, as well as HSP transcriptional factor 1 (HSF1), was correlated with poor prognosis in different subtypes of breast cancer. High-level expression of HSP90AA1 and HSP90AB1, two cytoplasmic HSP90 isoforms, was driven by chromosome coding region amplifications and were independent factors that led to death from breast cancer among patients with triple-negative (TNBC) and HER2-/ER+ subtypes, respectively. Furthermore, amplification of HSF1 was correlated with higher HSP90AA1 and HSP90AB1 mRNA expression among the breast cancer cells without amplifications of these two genes. A collection of HSP90AA1, HSP90AB1 and HSF

  10. The vitamin E analog, alpha-tocopheryloxyacetic acid enhances the anti-tumor activity of trastuzumab against HER2/neu-expressing breast cancer

    Directory of Open Access Journals (Sweden)

    Penichet Manuel L

    2011-11-01

    Full Text Available Abstract Background HER2/neu is an oncogene that facilitates neoplastic transformation due to its ability to transduce growth signals in a ligand-independent manner, is over-expressed in 20-30% of human breast cancers correlating with aggressive disease and has been successfully targeted with trastuzumab (Herceptin®. Because trastuzumab alone achieves only a 15-30% response rate, it is now commonly combined with conventional chemotherapeutic drugs. While the combination of trastuzumab plus chemotherapy has greatly improved response rates and increased survival, these conventional chemotherapy drugs are frequently associated with gastrointestinal and cardiac toxicity, bone marrow and immune suppression. These drawbacks necessitate the development of new, less toxic drugs that can be combined with trastuzumab. Recently, we reported that orally administered alpha-tocopheryloxyacetic acid (α-TEA, a novel ether derivative of alpha-tocopherol, dramatically suppressed primary tumor growth and reduced the incidence of lung metastases both in a transplanted and a spontaneous mouse model of breast cancer without discernable toxicity. Methods In this study we examined the effect of α-TEA plus HER2/neu-specific antibody treatment on HER2/neu-expressing breast cancer cells in vitro and in a HER2/neu positive human xenograft tumor model in vivo. Results We show in vitro that α-TEA plus anti-HER2/neu antibody has an increased cytotoxic effect against murine mammary tumor cells and human breast cancer cells and that the anti-tumor effect of α-TEA is independent of HER2/neu status. More importantly, in a human breast cancer xenograft model, the combination of α-TEA plus trastuzumab resulted in faster tumor regression and more tumor-free animals than trastuzumab alone. Conclusion Due to the cancer cell selectivity of α-TEA, and because α-TEA kills both HER2/neu positive and HER2/neu negative breast cancer cells, it has the potential to be effective and

  11. Regional Nodal Irradiation After Breast Conserving Surgery for Early HER2-Positive Breast Cancer: Results of a Subanalysis From the ALTTO Trial.

    Science.gov (United States)

    Gingras, Isabelle; Holmes, Eileen; De Azambuja, Evandro; Nguyen, David H A; Izquierdo, Miguel; Anne Zujewski, Jo; Inbar, Moshe; Naume, Bjorn; Tomasello, Gianluca; Gralow, Julie R; Wolff, Antonio C; Harris, Lyndsay; Gnant, Michael; Moreno-Aspitia, Alvaro; Piccart, Martine J; Azim, Hatem A

    2017-08-01

    Two randomized trials recently demonstrated that regional nodal irradiation (RNI) could reduce the risk of recurrence in early breast cancer; however, these trials were conducted in the pretrastuzumab era. Whether these results are applicable to human epidermal growth factor receptor 2 (HER2)-positive breast cancer patients treated with anti-HER2-targeted therapy is unknown. This retrospective analysis was performed on patients with node-positive breast cancer who were enrolled in the Adjuvant Lapatinib and/or Trastuzumab Treatment Optimization phase III adjuvant trial and subjected to BCS. The primary objective of the present study was to examine the effect of RNI on disease-free survival (DFS). A multivariable cox regression analysis adjusted for number of positive lymph nodes, tumor size, grade, age, hormone receptors status, presence of macrometastatis, treatment arm, and chemotherapy timing was carried out to investigate the relationship between RNI and DFS. One thousand six hundred sixty-four HER2-positive breast cancer patients were included, of whom 878 (52.8%) had received RNI to the axillary, supraclavicular, and/or internal mammary lymph nodes. Patients in the RNI group had higher nodal burden and more frequently had tumors larger than 2 cm. At a median follow-up of 4.5 years, DFS was 84.3% in the RNI group and 88.3% in the non-RNI group. No differences in regional recurrence (0.9 % vs 0.6 %) or in overall survival (93.6% vs 95.3%) were observed between the two groups. After adjustment in multivariable analysis, there was no statistically significant association between RNI and DFS (hazard ratio = 0.96, 95% confidence interval = 0.71 to 1.29). Our analysis did not demonstrate a DFS benefit of RNI in HER2-positive, node-positive patients treated with adjuvant HER2-targeted therapy. The benefit of RNI in HER2-positive breast cancer needs further testing within randomized clinical trials. © The Author 2017. Published by Oxford University Press. All

  12. Homogeneous plate based antibody internalization assay using pH sensor fluorescent dye.

    Science.gov (United States)

    Nath, Nidhi; Godat, Becky; Zimprich, Chad; Dwight, Stephen J; Corona, Cesear; McDougall, Mark; Urh, Marjeta

    2016-04-01

    Receptor-mediated antibody internalization is a key mechanism underlying several anti-cancer antibody therapeutics. Delivering highly toxic drugs to cancer cells, as in the case of antibody drug conjugates (ADCs), efficient removal of surface receptors from cancer cells and changing the pharmacokinetics profile of the antibody drugs are some of key ways that internalization impacts the therapeutic efficacy of the antibodies. Over the years, several techniques have been used to study antibody internalization including radiolabels, fluorescent microscopy, flow cytometry and cellular toxicity assays. While these methods allow analysis of internalization, they have limitations including a multistep process and limited throughput and are generally endpoint assays. Here, we present a new homogeneous method that enables time and concentration dependent measurements of antibody internalization. The method uses a new hydrophilic and bright pH sensor dye (pHAb dye), which is not fluorescent at neutral pH but becomes highly fluorescent at acidic pH. For receptor mediated antibody internalization studies, antibodies against receptors are conjugated with the pHAb dye and incubated with the cells expressing the receptors. Upon binding to the receptor, the dyes conjugated to the antibody are not fluorescent because of the neutral pH of the media, but upon internalization and trafficking into endosomal and lysosomal vesicles the pH drops and dyes become fluorescent. The enabling attributes of the pHAb dyes are the hydrophilic nature to minimize antibody aggregation and bright fluorescence at acidic pH which allows development of simple plate based assays using a fluorescent reader. Using two different therapeutic antibodies--Trastuzumab (anti-HER2) and Cetuximab (anti-EGFR)--we show labeling with pHAb dye using amine and thiol chemistries and impact of chemistry and dye to antibody ration on internalization. We finally present two new approaches using the pHAb dye, which will be

  13. Chemotherapy for isolated locoregional recurrence of breast cancer (CALOR): a randomised trial.

    Science.gov (United States)

    Aebi, Stefan; Gelber, Shari; Anderson, Stewart J; Láng, István; Robidoux, André; Martín, Miguel; Nortier, Johan W R; Paterson, Alexander H G; Rimawi, Mothaffar F; Cañada, José Manuel Baena; Thürlimann, Beat; Murray, Elizabeth; Mamounas, Eleftherios P; Geyer, Charles E; Price, Karen N; Coates, Alan S; Gelber, Richard D; Rastogi, Priya; Wolmark, Norman; Wapnir, Irene L

    2014-02-01

    Patients with isolated locoregional recurrences (ILRR) of breast cancer have a high risk of distant metastasis and death from breast cancer. We aimed to establish whether adjuvant chemotherapy improves the outcome of such patients. The CALOR trial was a pragmatic, open-label, randomised trial that accrued patients with histologically proven and completely excised ILRR after unilateral breast cancer who had undergone a mastectomy or lumpectomy with clear surgical margins. Eligible patients were enrolled from hospitals worldwide and were centrally randomised (1:1) to chemotherapy (type selected by the investigator; multidrug for at least four courses recommended) or no chemotherapy, using permuted blocks, and stratified by previous chemotherapy, oestrogen-receptor and progesterone-receptor status, and location of ILRR. Patients with oestrogen-receptor-positive ILRR received adjuvant endocrine therapy, radiation therapy was mandated for patients with microscopically involved surgical margins, and anti-HER2 therapy was optional. The primary endpoint was disease-free survival. All analyses were by intention to treat. This study is registered with ClinicalTrials.gov, number NCT00074152. From Aug 22, 2003, to Jan 31, 2010, 85 patients were randomly assigned to receive chemotherapy and 77 were assigned to no chemotherapy. At a median follow-up of 4·9 years (IQR 3·6-6 ·0), 24 (28%) patients had disease-free survival events in the chemotherapy group compared with 34 (44%) in the no chemotherapy group. 5-year disease-free survival was 69% (95% CI 56-79) with chemotherapy versus 57% (44-67) without chemotherapy (hazard ratio 0·59 [95% CI 0·35-0·99]; p=0·046). Adjuvant chemotherapy was significantly more effective for women with oestrogen-receptor-negative ILRR (pinteraction=0·046), but analyses of disease-free survival according to the oestrogen-receptor status of the primary tumour were not statistically significant (pinteraction=0·43). Of the 81 patients who

  14. Generation of IgE-based immunotherapies against HER-2 overexpressing tumours

    International Nuclear Information System (INIS)

    Knittelfelder, R.

    2010-01-01

    In combination with chemotherapy or radiation, passive immunotherapy with monoclonal antibodies is state of the art in cancer therapy. For this purpose, two properties of antibodies are harnessed: i) via the Fab fragment they bind a specific tumour antigen and ii) via the Fc portion they recruit effector cells and activate the complement system. One of these antibodies is trastuzumab (Herceptin), a growth-inhibitory humanized monoclonal IgG1 antibody recognizing the tumour antigen HER-2, which is overexpressed in 30% of human breast cancers. Interestingly, all antibodies applied for passive immunotherapy are so far exclusively of the IgG subclass. In contrast, antibodies of the IgE subclass are best-known for their detrimental function in type I hypersensitivity. It is little-known that IgE has anti-tumour capacity which could be exploited for immunotherapy of cancer. Thus, the aim of this doctoral thesis was to examine alternative strategies for cancer treatment based on IgE antibodies, and to compare their efficacy with that of IgG. The oral immunization route is well suited for the induction of a Th2 immunity including high affine IgE responses to administered antigens. Therefore, the establishment of an IgE dependent food allergy model in mice is described, which we applied also for our cancer studies. When mice were fed with different concentrations of ovalbumin under concomitant anti-acid medication, an antigen-specific IgE induction in a Th2 environment could be achieved. This oral vaccination regimen was also used for feeding HER-2 mimotopes, i.e. epitope-mimics of the anti-HER-2 IgG antibody trastuzumab. Indeed, these mimotopes induced IgE antibodies recognizing the tumour antigen which were able to bind HER-2 overexpressing breast cancer cells and led to tumour cell lysis. Complementary to this active immunotherapeutic approach a trastuzumab-like IgE antibody for passive immunotherapy was constructed. We could show that this trastuzumab IgE exhibited the

  15. (67/68)Ga-labeling agent that liberates (67/68)Ga-NOTA-methionine by lysosomal proteolysis of parental low molecular weight polypeptides to reduce renal radioactivity levels.

    Science.gov (United States)

    Uehara, Tomoya; Rokugawa, Takemi; Kinoshita, Mai; Nemoto, Souki; Fransisco Lazaro, Guerra Gomez; Hanaoka, Hirofumi; Arano, Yasushi

    2014-11-19

    The renal localization of gallium-67 or gallium-68 ((67/68)Ga)-labeled low molecular weight (LMW) probes such as peptides and antibody fragments constitutes a problem in targeted imaging. Wu et al. previously showed that (67)Ga-labeled S-2-(4-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid (SCN-Bz-NOTA)-conjugated methionine ((67)Ga-NOTA-Met) was rapidly excreted from the kidney in urine following lysosomal proteolysis of the parental (67)Ga-NOTA-Bz-SCN-disulfide-stabilized Fv fragment (Bioconjugate Chem., (1997) 8, 365-369). In the present study, a new (67/68)Ga-labeling reagent for LMW probes that liberates (67/68)Ga-NOTA-Met was designed, synthesized, and evaluated using longer-lived (67)Ga in order to reduce renal radioactivity levels. We employed a methionine-isoleucine (MI) dipeptide bond as the cleavable linkage. The amine residue of MI was coupled with SCN-Bz-NOTA for (67)Ga-labeling, while the carboxylic acid residue of MI was derivatized to maleimide for antibody conjugation in order to synthesize NOTA-MI-Mal. A Fab fragment of the anti-Her2 antibody was thiolated with iminothiolane, and NOTA-MI-Mal was conjugated with the antibody fragment by maleimide-thiol chemistry. The Fab fragment was also conjugated with SCN-Bz-NOTA (NOTA-Fab) for comparison. (67)Ga-NOTA-MI-Fab was obtained at radiochemical yields of over 95% and was stable in murine serum for 24 h. In the biodistribution study using normal mice, (67)Ga-NOTA-MI-Fab registered significantly lower renal radioactivity levels from 1 to 6 h postinjection than those of (67)Ga-NOTA-Fab. An analysis of urine samples obtained 6 h after the injection of (67)Ga-NOTA-MI-Fab showed that the majority of radioactivity was excreted as (67)Ga-NOTA-Met. In the biodistribution study using tumor-bearing mice, the tumor to kidney ratios of (67)Ga-NOTA-MI-Fab were 4 times higher (6 h postinjection) than those of (67)Ga-NOTA-Fab. Although further studies including the structure of radiometabolites and

  16. Targeting and killing of glioblastoma with activated T cells armed with bispecific antibodies

    International Nuclear Information System (INIS)

    Zitron, Ian M; Thakur, Archana; Norkina, Oxana; Barger, Geoffrey R; Lum, Lawrence G; Mittal, Sandeep

    2013-01-01

    Since most glioblastomas express both wild-type EGFR and EGFRvIII as well as HER2/neu, they are excellent targets for activated T cells (ATC) armed with bispecific antibodies (BiAbs) that target EGFR and HER2. ATC were generated from PBMC activated for 14 days with anti-CD3 monoclonal antibody in the presence of interleukin-2 and armed with chemically heteroconjugated anti-CD3×anti-HER2/neu (HER2Bi) and/or anti-CD3×anti-EGFR (EGFRBi). HER2Bi- and/or EGFRBi-armed ATC were examined for in vitro cytotoxicity using MTT and 51 Cr-release assays against malignant glioma lines (U87MG, U118MG, and U251MG) and primary glioblastoma lines. EGFRBi-armed ATC killed up to 85% of U87, U118, and U251 targets at effector:target ratios (E:T) ranging from 1:1 to 25:1. Engagement of tumor by EGFRBi-armed ATC induced Th1 and Th2 cytokine secretion by armed ATC. HER2Bi-armed ATC exhibited comparable cytotoxicity against U118 and U251, but did not kill HER2-negative U87 cells. HER2Bi- or EGFRBi-armed ATC exhibited 50—80% cytotoxicity against four primary glioblastoma lines as well as a temozolomide (TMZ)-resistant variant of U251. Both CD133– and CD133+ subpopulations were killed by armed ATC. Targeting both HER2Bi and EGFRBi simultaneously showed enhanced efficacy than arming with a single BiAb. Armed ATC maintained effectiveness after irradiation and in the presence of TMZ at a therapeutic concentration and were capable of killing multiple targets. High-grade gliomas are suitable for specific targeting by armed ATC. These data, together with additional animal studies, may provide the preclinical support for the use of armed ATC as a valuable addition to current treatment regimens

  17. In vitro cytotoxicity of {sup 211}At-labeled trastuzumab in human breast cancer cell lines: effect of specific activity and HER2 receptor heterogeneity on survival fraction

    Energy Technology Data Exchange (ETDEWEB)

    Akabani, Gamal [Department of Radiology, Duke University Medical Center, P.O. Box 3808, Durham, NC 27710 (United States); Carlin, Sean [Department of Radiology, Duke University Medical Center, P.O. Box 3808, Durham, NC 27710 (United States); Welsh, Phil [Department of Radiology, Duke University Medical Center, P.O. Box 3808, Durham, NC 27710 (United States); Zalutsky, Michael R. [Department of Radiology, Duke University Medical Center, P.O. Box 3808, Durham, NC 27710 (United States)]. E-mail: zalut001@mc.duke.edu

    2006-04-15

    Introduction: Radioimmunotherapy with anti-HER2 monoclonal antibodies (mAbs) such as trastuzumab is a promising strategy for treating HER2-positive breast and ovarian carcinoma patients. The objective of this study was to determine the cytotoxic effectiveness of trastuzumab labeled with the 7.2-h half-life {alpha}-particle emitter {sup 211}At. Methods: Experiments were performed on SKBr-3, BT-474 and the transfected MCF7/HER2-18 human breast carcinoma cell lines. Intrinsic radiosensitivity was determined after exposure to external beam irradiation. The cytotoxicity of {sup 211}At-labeled trastuzumab was measured by clonogenic assays. The distribution of HER2 receptor expression on the cell lines was measured using fluorescence-activated cell sorting. A pharmacokinetic (PK)/microdosimetric model was established to assess the effects of specific activity (SA), HER2 receptor expression and absorbed dose on survival fraction (SF). Results: With external beam irradiation, the 2-Gy SF for BT-474, SKBr-3 and MCF7/HER2-18 cells was 0.78, 0.53 and 0.64 Gy, respectively. Heterogeneous HER2 expression was observed, with a subpopulation of cells lacking measurable receptor (14.5%, SKBr-3; 0.34%, MCF-7/HER2; 1.73%, BT-474). When plotted as a function of activity concentration, SF curves were biphasic and inversely proportional to SA; however, when the model was applied and absorbed doses calculated, the SF curve was monoexponential independent of SA. Thus, the PK model was able to demonstrate the effects of competition between cold and labeled mAb. These studies showed that the relative biological effectiveness of {sup 211}At-labeled trastuzaumab was about 10 times higher than that of external beam therapy. Conclusion: These in vitro studies showed that {sup 211}At-labeled trastuzumab mAb is an effective cytotoxic agent for the treatment of HER2-positive tumor cells. The SA of the labeled mAb and the homogeneity of HER2 receptor expression are important variables influencing

  18. Can breast cancer patients with HER2 dual-equivocal tumours be managed as HER2-negative disease?

    Science.gov (United States)

    Tong, Yiwei; Chen, Xiaosong; Fei, Xiaochun; Lin, Lin; Wu, Jiayi; Huang, Ou; He, Jianrong; Zhu, Li; Chen, Weiguo; Li, Yafen; Shen, Kunwei

    2018-01-01

    Increasing human epidermal growth factor receptor 2 (HER2) immunohistochemistry (IHC)/fluorescence in situ hybridisation (FISH) dual-equivocal breast tumours are reported after the 2013 American Society of Clinical Oncology/College of American Pathologists (ASCO/CAP) guideline update. The aim of this study is to investigate the clinico-pathologic characteristics, treatment patterns and disease outcome of these patients with HER2 dual-equivocal tumours. Patients with HER2 IHC 2+ and available FISH results were retrospectively analysed from the Comprehensive Breast Health Center, Shanghai Ruijin Hospital. The 2013 ASCO/CAP guideline was applied to define HER2-positive, dual-equivocal and -negative groups. Patient characteristics, systemic treatment patterns and survival were compared among these groups. Reverse transcriptase-polymerase chain reaction-based assays were applied to test HER2 mRNA expression level. Among 691 patients included, 133 (19.25%) were HER2 positive, 25 (3.62%) were HER2 dual-equivocal and 533 (77.13%) were HER2 negative. Univariate and multivariate analyses stated that HER2 dual-equivocal tumours shared more similarity with HER2-negative tumours, whereas HER2-positive tumours had rather different clinico-pathologic features. HER2 dual-equivocal tumours had similar HER2 mRNA levels compared with HER2-negative tumours (P = 0.26), which were much less compared with HER2-positive breast cancer. Besides, adjuvant systemic treatment patterns were comparable between HER2-negative and dual-equivocal tumours, and none of HER2 dual-equivocal patients received anti-HER2 treatment. There was no survival difference among these three groups (P = 0.43). HER2 dual-equivocal tumours share more similarity with HER2-negative disease in terms of clinico-pathologic features, HER2 mRNA levels, adjuvant systemic treatment patterns and disease outcome, which deserves further clinical evaluation. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Tyrosine kinase inhibitors as modulators of trastuzumab-mediated antibody-dependent cell-mediated cytotoxicity in breast cancer cell lines.

    Science.gov (United States)

    Collins, Denis M; Gately, Kathy; Hughes, Clare; Edwards, Connla; Davies, Anthony; Madden, Stephen F; O'Byrne, Kenneth J; O'Donovan, Norma; Crown, John

    2017-09-01

    Trastuzumab is an anti-HER2 monoclonal antibody (mAb) therapy capable of antibody-dependent cell-mediated cytotoxicity (ADCC) and used in the treatment of HER2+ breast cancer. Through interactions with FcƴR+ immune cell subsets, trastuzumab functions as a passive immunotherapy. The EGFR/HER2-targeting tyrosine kinase inhibitor (TKI) lapatinib and the next generation TKIs afatinib and neratinib, can alter HER2 levels, potentially modulating the ADCC response to trastuzumab. Using LDH-release assays, we investigated the impact of antigen modulation, assay duration and peripheral blood mononuclear cell (PBMC) activity on trastuzumab-mediated ADCC in breast cancer models of maximal (SKBR3) and minimal (MCF-7) target antigen expression to determine if modulating the ADCC response to trastuzumab using TKIs may be a viable approach for enhancing tumor immune reactivity. HER2 levels were determined in lapatinib, afatinib and neratinib-treated SKBR3 and MCF-7 using high content analysis (HCA). Trastuzumab-mediated ADCC was assessed following treatment with TKIs utilising a colorimetric LDH release-based protocol at 4 and 12h timepoints. PBMC activity was assessed against non-MHC-restricted K562 cells. A flow cytometry-based method (CFSE/7-AAD) was also used to measure trastuzumab-mediated ADCC in medium-treated SKBR3 and MCF-7. HER2 antigen levels were significantly altered by the three TKIs in both cell line models. The TKIs significantly reduced LDH levels directly in SKBR3 cells but not MCF-7. Lapatinib and neratinib augment trastuzumab-related ADCC in SKBR3 but the effect was not consistent with antigen expression levels and was dependent on volunteer PBMC activity (vs. K562). A 12h assay timepoint produced more consistent results. Trastuzumab-mediated ADCC (PBMC:target cell ratio of 10:1) was measured at 7.6±4.7% (T12) by LDH assay and 19±3.2 % (T12) using the flow cytometry-based method in the antigen-low model MCF-7. In the presence of effector cells with high

  20. De-Escalation Strategies in Human Epidermal Growth Factor Receptor 2 (HER2)-Positive Early Breast Cancer (BC): Final Analysis of the West German Study Group Adjuvant Dynamic Marker-Adjusted Personalized Therapy Trial Optimizing Risk Assessment and Therapy Response Prediction in Early BC HER2- and Hormone Receptor-Positive Phase II Randomized Trial-Efficacy, Safety, and Predictive Markers for 12 Weeks of Neoadjuvant Trastuzumab Emtansine With or Without Endocrine Therapy (ET) Versus Trastuzumab Plus ET.

    Science.gov (United States)

    Harbeck, Nadia; Gluz, Oleg; Christgen, Matthias; Kates, Ronald Ernest; Braun, Michael; Küemmel, Sherko; Schumacher, Claudia; Potenberg, Jochem; Kraemer, Stefan; Kleine-Tebbe, Anke; Augustin, Doris; Aktas, Bahriye; Forstbauer, Helmut; Tio, Joke; von Schumann, Raquel; Liedtke, Cornelia; Grischke, Eva-Maria; Schumacher, Johannes; Wuerstlein, Rachel; Kreipe, Hans Heinrich; Nitz, Ulrike Anneliese

    2017-09-10

    Purpose Human epidermal growth factor receptor 2 (HER2)-positive/hormone receptor (HR)-positive breast cancer is a distinct subgroup associated with lower chemotherapy sensitivity and slightly better outcome than HER2-positive/HR-negative disease. Little is known about the efficacy of the combination of endocrine therapy (ET) with trastuzumab or with the potent antibody-cytotoxic, anti-HER2 compound trastuzumab emtansine (T-DM1) with or without ET for this subgroup. The West German Study Group trial, ADAPT (Adjuvant Dynamic Marker-Adjusted Personalized Therapy Trial Optimizing Risk Assessment and Therapy Response Prediction in Early Breast Cancer) compares pathologic complete response (pCR) rates of T-DM1 versus trastuzumab with ET in early HER2-positive/HR-positive breast cancer. Patients and Methods In this prospective, neoadjuvant, phase II trial, 375 patients with early breast cancer with HER2-positive and HR-positive status (n = 463 screened) were randomly assigned to 12 weeks of T-DM1 with or without ET or to trastuzumab with ET. The primary end point was pCR (ypT0/is/ypN0). Early response was assessed in 3-week post-therapeutic core biopsies (proliferation decrease ≥ 30% Ki-67 or cellularity response). Secondary end points included safety and predictive impact of early response on pCR. Adjuvant therapy followed national standards. Results Baseline characteristics were well balanced among the arms. More than 90% of patients completed the therapy per protocol. pCR was observed in 41.0% of patients treated with T-DM1, 41.5% of patients treated with T-DM1 and ET, and 15.1% with trastuzumab and ET ( P < .001). Early responders (67% of patients with assessable response) achieved pCR in 35.7% compared with 19.8% in nonresponders (odds ratio, 2.2; 95% CI, 1.24 to 4.19). T-DM1 was associated with a significantly higher prevalence of grade 1 to 2 toxicities, especially thrombocytopenia, nausea, and elevation of liver enzymes. Overall toxicity was low; seventeen

  1. Actionable gene-based classification toward precision medicine in gastric cancer

    Directory of Open Access Journals (Sweden)

    Hiroshi Ichikawa

    2017-10-01

    Full Text Available Abstract Background Intertumoral heterogeneity represents a significant hurdle to identifying optimized targeted therapies in gastric cancer (GC. To realize precision medicine for GC patients, an actionable gene alteration-based molecular classification that directly associates GCs with targeted therapies is needed. Methods A total of 207 Japanese patients with GC were included in this study. Formalin-fixed, paraffin-embedded (FFPE tumor tissues were obtained from surgical or biopsy specimens and were subjected to DNA extraction. We generated comprehensive genomic profiling data using a 435-gene panel including 69 actionable genes paired with US Food and Drug Administration-approved targeted therapies, and the evaluation of Epstein-Barr virus (EBV infection and microsatellite instability (MSI status. Results Comprehensive genomic sequencing detected at least one alteration of 435 cancer-related genes in 194 GCs (93.7% and of 69 actionable genes in 141 GCs (68.1%. We classified the 207 GCs into four The Cancer Genome Atlas (TCGA subtypes using the genomic profiling data; EBV (N = 9, MSI (N = 17, chromosomal instability (N = 119, and genomically stable subtype (N = 62. Actionable gene alterations were not specific and were widely observed throughout all TCGA subtypes. To discover a novel classification which more precisely selects candidates for targeted therapies, 207 GCs were classified using hypermutated phenotype and the mutation profile of 69 actionable genes. We identified a hypermutated group (N = 32, while the others (N = 175 were sub-divided into six clusters including five with actionable gene alterations: ERBB2 (N = 25, CDKN2A, and CDKN2B (N = 10, KRAS (N = 10, BRCA2 (N = 9, and ATM cluster (N = 12. The clinical utility of this classification was demonstrated by a case of unresectable GC with a remarkable response to anti-HER2 therapy in the ERBB2 cluster. Conclusions This actionable gene

  2. The extracellular domain of Her2 in serum as a biomarker of breast cancer.

    Science.gov (United States)

    Perrier, Alexandre; Gligorov, Joseph; Lefèvre, Guillaume; Boissan, Mathieu

    2018-02-28

    Breast cancer is a major health problem worldwide. In ~15% of breast cancers, the epidermal growth factor receptor HER2, a transmembrane protein, is overexpressed. This HER2 overexpression is associated with an aggressive form of the disease and a poor clinical prognosis. The extracellular domain (ECD) of HER2 is released into the blood by a proteolytic mechanism known as "ECD shedding". This proteolytic shedding leaves a constitutively active truncated receptor in the membrane that is 10-100-fold more oncogenic than the full-length receptor and promotes the growth and survival of cancer cells. Shedding of the HER2 ECD is increased during metastasis: whereas 15% of primary breast cancer patients have elevated levels of serum HER2 ECD (sHER2 ECD), the levels reach 45% in patients with metastatic disease. Thus, sHER2 ECD has been proposed as a promising biomarker for cancer recurrence and for monitoring the disease status of patients overexpressing HER2. Nevertheless, in 2016, the American Society of Clinical Oncology advises clinicians not to use soluble HER2 levels to guide their choice of adjuvant therapy for patients with HER2-positive breast cancer, because the evidence was considered not strong enough. Currently, biomarkers such as carcinoembryonic antigen and cancer antigen 15-3 are widely used to monitor metastatic breast cancer disease even if the level of evidence of clinical impact of this monitoring is poor. In this article, we review the evidence that sHER2 ECD might be used in some situations as a biomarker for breast cancer. Although this serum biomarker will not replace the direct measurement of tumor HER2 status for diagnosis of early-stage tumors; it might be especially useful in metastatic disease for prognosis, as an indicator of cancer progression and of therapy response, particularly to anti-HER2 therapies. Owing to these data, sHER2 ECD should be considered as a promising biomarker to detect cancer recurrence and metastasis.

  3. Development and preclinical characterisation of 99mTc-labelled Affibody molecules with reduced renal uptake

    International Nuclear Information System (INIS)

    Ekblad, Torun; Karlstroem, Amelie Eriksson; Tran, Thuy; Orlova, Anna; Feldwisch, Joachim; Widstroem, Charles; Abrahmsen, Lars; Wennborg, Anders; Tolmachev, Vladimir

    2008-01-01

    Affibody molecules are low molecular weight proteins (7 kDa), which can be selected to bind to tumour-associated target proteins with subnanomolar affinity. Because of rapid tumour localisation and clearance from nonspecific compartments, Affibody molecules are promising tracers for molecular imaging. Earlier, 99m Tc-labelled Affibody molecules demonstrated specific targeting of tumour xenografts. However, the biodistribution was suboptimal either because of hepatobiliary excretion or high renal uptake of the radioactivity. The goal of this study was to optimise the biodistribution of Affibody molecules by chelator engineering. Anti-HER2 Z HER2:342 Affibody molecules, carrying the mercaptoacetyl-glutamyl-seryl-glutamyl (maESE), mercaptoacetyl-glutamyl-glutamyl-seryl (maEES) and mercaptoacetyl-seryl-glutamyl-glutamyl (maSEE) chelators, were prepared by peptide synthesis and labelled with 99m Tc. The tumour-targeting capacity of these conjugates was compared with each other and with the best previously available conjugate, 99m Tc-maEEE-Z HER2:342, in nude mice bearing SKOV-3 xenografts. The tumour-targeting capacity of the most promising conjugate, 99m Tc-maESE-Z HER2:342, was compared with radioiodinated Z HER2:342 . All novel conjugates demonstrated successful tumour targeting and a low degree of hepatobiliary excretion. The renal uptakes of serine-containing conjugates, 33±5, 68±21 and 71±10%IA/g, for 99m Tc-maESE-Z HER2:342 , 99m Tc-maEES-Z HER2:342 and 99m Tc-maSEE-Z HER2:342 , respectively, were significantly reduced in comparison with 99m Tc-maEEE-Z HER2:342 (102 ± 13%IA/g). For 99m Tc-maESE-Z HER2:342 , a tumour uptake of 9.6±1.8%IA/g and a tumour-to-blood ratio of 58±6 were reached at 4 h p.i. A combination of serine and glutamic acid residues in the chelator sequence confers increased renal excretion and relatively low renal uptake of 99m Tc-labelled Affibody molecules. In combination with preserved targeting capacity, this improved imaging of targets

  4. Development of the designed ankyrin repeat protein (DARPin) G3 for HER2 molecular imaging

    Energy Technology Data Exchange (ETDEWEB)

    Goldstein, Robert; Livanos, Maria; Bhavsar, Gaurav; Rashid, Mohammed; Miranda, Enrique; Tolner, Berend; Meyer, Tim; Chester, Kerry [UCL Cancer Institute, London (United Kingdom); Sosabowski, Jane; Leyton, Julius; Mather, Stephen [Queen Mary University of London, Centre for Molecular Oncology, Barts Cancer Institute, London (United Kingdom); Vigor, Kim [Clare Hall Laboratories, Biotherapeutics Development Unit, Cancer Research UK, South Mimms (United Kingdom); Nagy-Davidescu, Gabriela; Plueckthun, Andreas [Universitaet Zuerich, Biochemisches Institut, Zuerich (Switzerland); Yeung, Jenny [UCL Cancer Institute, London (United Kingdom); UCL Institute of Child Health, London (United Kingdom)

    2014-11-13

    Human epidermal growth factor receptor-2 (HER2) overexpression is a predictor of response to anti-HER2 therapy in breast and gastric cancer. Currently, HER2 status is assessed by tumour biopsy, but this may not be representative of the larger tumour mass or other metastatic sites, risking misclassification and selection of suboptimal therapy. The designed ankyrin repeat protein (DARPin) G3 binds HER2 with high affinity at an epitope that does not overlap with trastuzumab and is biologically inert. We hypothesized that radiolabelled DARPin G3 would be capable of selectively imaging HER2-positive tumours, and aimed to identify a suitable format for clinical application. G3 DARPins tagged with hexahistidine (His{sub 6}) or with histidine glutamate (HE){sub 3} and untagged G3 DARPins were manufactured using a GMP-compatible Pichia pastoris protocol and radiolabelled with {sup 125}I, or with {sup 111}In via DOTA linked to a C-terminal cysteine. BALB/c mice were injected with radiolabelled G3 and tissue biodistribution was evaluated by gamma counting. The lead construct ((HE){sub 3}-G3) was assessed in mice bearing HER2-positive human breast tumour (BT474) xenografts. For both isotopes, (HE){sub 3}-G3 had significantly lower liver uptake than His{sub 6}-G3 and untagged G3 counterparts in non-tumour-bearing mice, and there was no significantly different liver uptake between His{sub 6}-G3 and untagged G3. (HE){sub 3}-G3 was taken forward for evaluation in mice bearing HER2-positive tumour xenografts. The results demonstrated that radioactivity from {sup 111}In-(HE){sub 3}-G3 was better maintained in tumours and cleared faster from serum than radioactivity from {sup 125}I-(HE){sub 3}-G3, achieving superior tumour-to-blood ratios (343.7 ± 161.3 vs. 22.0 ± 11.3 at 24 h, respectively). On microSPECT/CT, {sup 111}In-labelled and {sup 125}I-labelled (HE){sub 3}-G3 could image HER2-positive tumours at 4 h after administration, but there was less normal tissue uptake of

  5. Development of the designed ankyrin repeat protein (DARPin) G3 for HER2 molecular imaging

    International Nuclear Information System (INIS)

    Goldstein, Robert; Livanos, Maria; Bhavsar, Gaurav; Rashid, Mohammed; Miranda, Enrique; Tolner, Berend; Meyer, Tim; Chester, Kerry; Sosabowski, Jane; Leyton, Julius; Mather, Stephen; Vigor, Kim; Nagy-Davidescu, Gabriela; Plueckthun, Andreas; Yeung, Jenny

    2015-01-01

    Human epidermal growth factor receptor-2 (HER2) overexpression is a predictor of response to anti-HER2 therapy in breast and gastric cancer. Currently, HER2 status is assessed by tumour biopsy, but this may not be representative of the larger tumour mass or other metastatic sites, risking misclassification and selection of suboptimal therapy. The designed ankyrin repeat protein (DARPin) G3 binds HER2 with high affinity at an epitope that does not overlap with trastuzumab and is biologically inert. We hypothesized that radiolabelled DARPin G3 would be capable of selectively imaging HER2-positive tumours, and aimed to identify a suitable format for clinical application. G3 DARPins tagged with hexahistidine (His 6 ) or with histidine glutamate (HE) 3 and untagged G3 DARPins were manufactured using a GMP-compatible Pichia pastoris protocol and radiolabelled with 125 I, or with 111 In via DOTA linked to a C-terminal cysteine. BALB/c mice were injected with radiolabelled G3 and tissue biodistribution was evaluated by gamma counting. The lead construct ((HE) 3 -G3) was assessed in mice bearing HER2-positive human breast tumour (BT474) xenografts. For both isotopes, (HE) 3 -G3 had significantly lower liver uptake than His 6 -G3 and untagged G3 counterparts in non-tumour-bearing mice, and there was no significantly different liver uptake between His 6 -G3 and untagged G3. (HE) 3 -G3 was taken forward for evaluation in mice bearing HER2-positive tumour xenografts. The results demonstrated that radioactivity from 111 In-(HE) 3 -G3 was better maintained in tumours and cleared faster from serum than radioactivity from 125 I-(HE) 3 -G3, achieving superior tumour-to-