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Sample records for bcr-abl positive cells

  1. Imatinib-dependent tyrosine phosphorylation profiling of Bcr-Abl-positive chronic myeloid leukemia cells

    OpenAIRE

    Preisinger, C.; Schwarz, J. P.; Bleijerveld, O.B.; et al

    2012-01-01

    Bcr-Abl is the major cause and pathogenetic principle of chronic myeloid leukemia (CML). Bcr-Abl results from a chromosomal translocation that fuses the bcr and abl genes, thereby generating a constitutively active tyrosine kinase, which stimulates several signaling networks required for proliferation and survival. peer-reviewed

  2. NK cells are dysfunctional in human chronic myelogenous leukemia before and on imatinib treatment and in BCR-ABL-positive mice.

    Science.gov (United States)

    Chen, C I-U; Koschmieder, S; Kerstiens, L; Schemionek, M; Altvater, B; Pscherer, S; Gerss, J; Maecker, H T; Berdel, W E; Juergens, H; Lee, P P; Rossig, C

    2012-03-01

    Although BCR-ABL+ stem cells in chronic myeloid leukemia (CML) resist elimination by targeted pharmacotherapy in most patients, immunological graft-versus-leukemia effects can cure the disease. Besides cytotoxic T cells, natural killer (NK) cells may have a role in immune control of CML. Here, we explored the functionality of NK cells in CML patients and in a transgenic inducible BCR-ABL mouse model. Compared with controls, NK-cell proportions among lymphocytes were decreased at diagnosis of CML and did not recover during imatinib-induced remission for 10-34 months. Functional experiments revealed limited in vitro expansion of NK cells from CML patients and a reduced degranulation response to K562 target cells both at diagnosis and during imatinib therapy. Consistent with the results in human CML, relative numbers of NK1.1+ NK cells were reduced following induction of BCR-ABL expression in mice, and the defects persisted after BCR-ABL reversion. Moreover, target-induced degranulation by expanded BCR-ABL+ NK cells was compromised. We conclude that CML is associated with quantitative and functional defects within the NK-cell compartment, which is reproduced by induced BCR-ABL expression in mice. Further work will aim at identifying the mechanisms of NK-cell deficiency in CML and at developing strategies to exploit NK cells for immunotherapy. PMID:21904381

  3. Regulation of hTERT by BCR-ABL at multiple levels in K562 cells

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    Chai Juin Hsien

    2011-12-01

    Full Text Available Abstract Background The cytogenetic characteristic of Chronic Myeloid Leukemia (CML is the formation of the Philadelphia chromosome gene product, BCR-ABL. Given that BCR-ABL is the specific target of Gleevec in CML treatment, we investigated the regulation of the catalytic component of telomerase, hTERT, by BCR-ABL at multiple levels in K562 cells. Methods Molecular techniques such as over expression, knockdown, real-time PCR, immunoprecipitation, western blotting, reporter assay, confocal microscopy, telomerase assays and microarray were used to suggest that hTERT expression and activity is modulated by BCR-ABL at multiple levels. Results Our results suggest that BCR-ABL plays an important role in regulating hTERT in K562 (BCR-ABL positive human leukemia cells. When Gleevec inhibited the tyrosine kinase activity of BCR-ABL, phosphorylation of hTERT was downregulated, therefore suggesting a positive correlation between BCR-ABL and hTERT. Gleevec treatment inhibited hTERT at mRNA level and significantly reduced telomerase activity (TA in K562 cells, but not in HL60 or Jurkat cells (BCR-ABL negative cells. We also demonstrated that the transcription factor STAT5a plays a critical role in hTERT gene regulation in K562 cells. Knockdown of STAT5a, but not STAT5b, resulted in a marked downregulation of hTERT mRNA level, TA and hTERT protein level in K562 cells. Furthermore, translocation of hTERT from nucleoli to nucleoplasm was observed in K562 cells induced by Gleevec. Conclusions Our data reveal that BCR-ABL can regulate TA at multiple levels, including transcription, post-translational level, and proper localization. Thus, suppression of cell growth and induction of apoptosis by Gleevec treatment may be partially due to TA inhibition. Additionally, we have identified STAT5a as critical mediator of the hTERT gene expression in BCR-ABL positive CML cells, suggesting that targeting STAT5a may be a promising therapeutic strategy for BCR-ABL positive

  4. Regulation of hTERT by BCR-ABL at multiple levels in K562 cells

    International Nuclear Information System (INIS)

    The cytogenetic characteristic of Chronic Myeloid Leukemia (CML) is the formation of the Philadelphia chromosome gene product, BCR-ABL. Given that BCR-ABL is the specific target of Gleevec in CML treatment, we investigated the regulation of the catalytic component of telomerase, hTERT, by BCR-ABL at multiple levels in K562 cells. Molecular techniques such as over expression, knockdown, real-time PCR, immunoprecipitation, western blotting, reporter assay, confocal microscopy, telomerase assays and microarray were used to suggest that hTERT expression and activity is modulated by BCR-ABL at multiple levels. Our results suggest that BCR-ABL plays an important role in regulating hTERT in K562 (BCR-ABL positive human leukemia) cells. When Gleevec inhibited the tyrosine kinase activity of BCR-ABL, phosphorylation of hTERT was downregulated, therefore suggesting a positive correlation between BCR-ABL and hTERT. Gleevec treatment inhibited hTERT at mRNA level and significantly reduced telomerase activity (TA) in K562 cells, but not in HL60 or Jurkat cells (BCR-ABL negative cells). We also demonstrated that the transcription factor STAT5a plays a critical role in hTERT gene regulation in K562 cells. Knockdown of STAT5a, but not STAT5b, resulted in a marked downregulation of hTERT mRNA level, TA and hTERT protein level in K562 cells. Furthermore, translocation of hTERT from nucleoli to nucleoplasm was observed in K562 cells induced by Gleevec. Our data reveal that BCR-ABL can regulate TA at multiple levels, including transcription, post-translational level, and proper localization. Thus, suppression of cell growth and induction of apoptosis by Gleevec treatment may be partially due to TA inhibition. Additionally, we have identified STAT5a as critical mediator of the hTERT gene expression in BCR-ABL positive CML cells, suggesting that targeting STAT5a may be a promising therapeutic strategy for BCR-ABL positive CML patients

  5. miR-146b-5p within BCR-ABL1-Positive Microvesicles Promotes Leukemic Transformation of Hematopoietic Cells.

    Science.gov (United States)

    Zhang, Hong-Mei; Li, Qing; Zhu, Xiaojian; Liu, Wei; Hu, Hui; Liu, Teng; Cheng, Fanjun; You, Yong; Zhong, Zhaodong; Zou, Ping; Li, Qiubai; Chen, Zhichao; Guo, An-Yuan

    2016-05-15

    Evidence is accumulating that extracellular microvesicles (MV) facilitate progression and relapse in cancer. Using a model in which MVs derived from K562 chronic myelogenous leukemia (CML) cells transform normal hematopoietic transplants into leukemia-like cells, we defined the underlying mechanisms of this process through gene-expression studies and network analyses of transcription factors (TF) and miRNAs. We found that antitumor miRNAs were increased and several defense pathways were initiated during the early phases of oncogenic transformation. Later, oncomiRs and genes involved in cell cycle, DNA repair, and energy metabolism pathways were upregulated. Regulatory network analyses revealed that a number of TFs and miRNAs were responsible for the pathway dysregulation and the oncogenic transformation. In particular, we found that miR-146b-5p, which was highly expressed in MVs, coordinated the regulation of cancer-related genes to promote cell-transforming processes. Notably, treatment of recipient cells with MV derived from K562 cells expressing mimics of miR-146b-5p revealed that it accelerated the transformation process in large part by silencing the tumor-suppressor NUMB High levels of miR-146b-5p also enhanced reactive oxygen species levels and genome instability of recipient cells. Taken together, our finding showed how upregulation of oncogenic miRNAs in MVs promote hematopoetic cells to a leukemic state, as well as a demonstration for TF and miRNA coregulatory analysis in exploring the dysregulation of cancers and discovering key factors. Cancer Res; 76(10); 2901-11. ©2016 AACR. PMID:27013199

  6. INHIBITION OF APOPTOSIS BY bcr-abl FUSION GENE IN K562 CELLS

    Institute of Scientific and Technical Information of China (English)

    WANG Chun-hong; SUN Bing-zhong; YUAN Yue-chuan

    1999-01-01

    Objective: To investigate the effect of bcr-abl fusion gene on CML cell apoptosis. Methods: Apoptosis of exvivo cultured K562 cells were observed after exposure to synthetic 18 mer antisense oligodeoxynucleotide complementary to the bcr-abl junction (b3a2). Results: Apoptosis of K562 cells was significantly increased associated with inhibition of bcr-abl expression. Conclusion: bcr-abl fusion gene formation due to chromosome translocation may be the major mechanism of CML via inhibition of apoptosis.

  7. Environment-mediated drug resistance in Bcr/Abl-positive acute lymphoblastic leukemia

    OpenAIRE

    Feldhahn, Niklas; Arutyunyan, Anna; Stoddart, Sonia; ZHANG Bin; Schmidhuber, Sabine; Yi, Sun-ju; Kim, Yong-Mi; Groffen, John; Heisterkamp, Nora

    2012-01-01

    Although cure rates for acute lymphoblastic leukemia (ALL) have increased, development of resistance to drugs and patient relapse are common. The environment in which the leukemia cells are present during the drug treatment is known to provide significant survival benefit. Here, we have modeled this process by culturing murine Bcr/Abl-positive acute lymphoblastic leukemia cells in the presence of stroma while treating them with a moderate dose of two unrelated drugs, the farnesyltransferase i...

  8. An asymptomatic 61-year-old man with BCR-ABL-positive bone marrow following autologous transplantation for multiple myeloma

    OpenAIRE

    Roper, Nitin; DeAngelo, Daniel; Kuo, Frank; Cin, Paola Dal; Ghobrial, Irene; Aster, Jon C.

    2010-01-01

    A 61-year-old man treated with an autologous transplant for multiple myeloma was incidentally found to have a high level of BCR-ABL fusion gene-positive cells in his bone marrow. We describe the clinical decision-making process that led us to initiate therapy with imatinib, despite the absence of any clinical evidence of chronic myelogenous leukemia or other BCR-ABL associated hematologic malignancy.

  9. Role of p21 RAS in p210 bcr-abl transformation of murine myeloid cells.

    Science.gov (United States)

    Mandanas, R A; Leibowitz, D S; Gharehbaghi, K; Tauchi, T; Burgess, G S; Miyazawa, K; Jayaram, H N; Boswell, H S

    1993-09-15

    The p21 RAS product has been implicated as part of the downstream signaling of certain nonreceptor tyrosine kinase oncogenes and several growth factor receptor-ligand interactions. We have reported that the chronic myelogenous leukemia oncogene p210 bcr-abl transforms a growth-factor-dependent myeloid cell line NFS/N1.H7 to interleukin-3 (IL-3) independence. In these p210 bcr-abl-transformed cells (H7 bcr-abl.A54) and in two other murine myeloid cell lines transformed to IL-3 independence by p210 bcr-abl, endogenous p21 RAS is activated as determined by an elevated ratio of associated guanosine triphosphate (GTP)/guanosine diphosphate (GDP), assayed by thin-layer chromatography of the nucleotides eluted from p21 RAS after immunoprecipitation with the Y13-259 antibody. Treatment of p210 bcr-abl-transformed cells with a specific tyrosine kinase inhibitor herbimycin A resulted in diminished tyrosine phosphorylation of p210 bcr-abl and associated proteins, without major reduction in expression of the p210 bcr-abl protein itself. Inhibition of p210 bcr-abl-dependent tyrosine phosphorylation resulted in a reduction of active p21RAS-GTP complexes in the transformed cells, in diminished expression of the nuclear early response genes c-jun and c-fos, and in lower cellular proliferation rate. To further implicate p21 RAS in these functional events downstream of p210 bcr-abl tyrosine phosphorylation, we targeted G-protein function directly by limiting the availability of GTP with the inosine monophosphate dehydrogenase inhibitor, tiazofurin (TR). In p210 bcr-abl-transformed cells treated for 4 hours with TR, in which the levels of GTP were reduced by 50%, but GDP, guanosine monophosphate, and adenosine triphosphate (ATP) were unaffected, p210 bcr-abl tyrosine phosphorylation was at control levels. However, expression of c-fos and c-jun nuclear proto-oncogenes were strongly inhibited and p21 RAS activity was downregulated. These findings show that p210 bcr-abl transduces

  10. A non-radioactive assay for precise determination of intracellular levels of imatinib and its main metabolite in Bcr-Abl positive cells

    Czech Academy of Sciences Publication Activity Database

    Mlejnek, P.; Novák, Ondřej; Doležel, P.

    2011-01-01

    Roč. 83, č. 5 (2011), s. 1466-1471. ISSN 0039-9140 R&D Projects: GA ČR GA301/08/1649 Institutional research plan: CEZ:AV0Z50380511 Keywords : K562 cells * P-glycoprotein * Multidrug resistance * N-desmethyl imatinib * CGP 74588 Subject RIV: EF - Botanics Impact factor: 3.794, year: 2011

  11. Evaluation of Morpholino Antisense Oligos’ Role on BCR-ABL Gene Silencing in the K562 Cell Line

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    Bahman Delalat

    2010-01-01

    Full Text Available Objective: Chronic myeloid leukemia (CML develops when a hematopoietic stem cellacquires the BCR/ABL fusion gene. This causes these transformed hematopoietic cellsto have a greater than normal proliferation rate. Scientists attempt to improve the CMLtreatment process by silencing the BCR/ABL oncogene. In this work, we used morpholinoantisense oligos to silence the BCR/ABL oncogene.Materials and Methods: In this study, the K562 was used as a BCR/ABL fusion-genepositive cell line and the Jurkat cell line as a control. We explored the inhibiting capacityof morpholino antisense oligos in the the expression of the BCR/ABL oncogene andstudied their p210 BCR/ABL suppression, inhibition of cell proliferation and stimulation ofapoptosis in the K562 cells after 24 and 48 hours. Endo-Porter was used for delivery ofmorpholino antisense oligos into cell cytosols. Meanwhile, flow cytometric analysis wasperformed in order to determine the appropriate concentration of morpholino antisenseoligos.Results: Prolonged exposure of the K562 cell line to the morpholino antisense oligostargeted against the BCR-ABL gene showed proliferation inhibition as its main feature.After western blotting, we found that complete silencing of BCR/ABL was achieved, butflow cytometric analysis showed no broad apoptosis.Conclusion: The results indicate that the Morpholino antisense oligo is able to inhibitp210 BCR/ABL; however, it cannot induce broad apoptosis due to co-silencing of BCR.

  12. Curcumin synergistically augments bcr/abl phosphorethieate antisense oligonucleotides to inhibit growth of chronic myelogenous leukemia cells

    Institute of Scientific and Technical Information of China (English)

    Kun-zhong ZHANG; Jian-hua XU; Xiu-wang HUANG; Li-xian WU; Yu SU; Yuan-zhong CHEN

    2007-01-01

    Aim: To investigate the growth inhibition effect of the combination of bcr/abl phosphorothioate antisense oligonucleotides (PS-ASODN) and curcumin (cur), and the possible mechanisms of cur on the chronic myelogenous leukemia cell line K562. Methods: The K562 cell line was used as a P210bcr/abl-positive cell model in vitro and was exposed to different concentrations of PS-ASODN (0-20 μmol/L), cur (0-20 μmol/L), or a combination of both. Growth inhibition and apoptosis of K562 cells were assessed by MTT assay and AO/EB fluorescent staining, respec-tively. The expression levels of P210bct/abl, NF-κB and heat shock protein 90 (Hsp90) were assessed by Western blot. Results: Exposure to cur (5-20 μmol/L) and PS-ASODN (5-20 μmol/L) resulted in a synergistic inhibitory effect on cell growth.Growth inhibition was associated with the inhibition of the proliferation and in-duction of apoptosis. Western blot analysis showed that the drugs synergisti-cally downregulated the level of P210bcr/abl and NF-κB. Cur downregulated Hsp90,whereas no synergism was observed when cur was combined with PS-ASODN.Conclusion: PS-ASODN and cur exhibited a synergistic inhibitory effect on the cell growth of K562. The synergistic growth inhibition was mediated through different mechanisms that involved the inhibition of P210bcr/abl.

  13. Susceptibility of Ph-positive all to TKI therapy associated with Bcr-Abl rearrangement patterns: a retrospective analysis.

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    Yu Jing

    Full Text Available BACKGROUND: Tyrosine kinase inhibitors (TKIs have demonstrated success in the treatment of acute lymphoblastic leukemia (ALL in patients that express BCR-ABL rearrangements (Philadelphia chromosome [Ph]. The current study aimed to assess the efficacy of TKIs and prognostic factors in the treatment of adults with Ph+-ALL. METHODS: In this multicenter retrospective study, the relationship between Ph+-ALL and treatment outcomes among Chinese patients receiving TKI-containing induction/consolidation chemotherapy was examined. A total of 86 Ph+-ALL patients were included and followed for 3.85 (0.43-9.30 years. Overall survival (OS and event-free survival (EFS were analyzed. RESULTS: A total of 86 Ph+-ALL patients (40 females and 46 males; median age: 34.0 years were enrolled, including those with BCR/ABL transcripts 190 (n = 52, 210 (n = 25, and 230 (n = 2; BCR/ABL isoform determination was not available for 7 patients. Mortality was influenced by variable BCR/ABL transcripts and TKI administration, and BCR/ABL transcripts, hematopoietic stem cell transplantation (HSCT, and TKI administration were associated with the occurrence of events. The OS rate in the TKI administration group during steady state was significantly higher compared with those patients who did not receive TKI administration (P = 0.008, the EFS rate in the TKI administration group during steady state was significantly higher compared with those patients who did not receive TKIs (P = 0.012, and also higher than those with TKI salvage administration (P = 0.004. BCR/ABL transcripts 210 showed preferable OS and EFS compared with BCR/ABL transcripts 190 and 230 (P<0.05 for each. CONCLUSIONS: The susceptibility of Ph+-ALL to TKI associated with the patterns of BCR-ABL rearrangement is demonstrated for the first time, thus adding another risk-stratifying molecular prognostic tool for the management of patients with Ph+-ALL.

  14. Autophagy induction by Bcr-Abl-expressing cells facilitates their recovery from a targeted or nontargeted treatment.

    LENUS (Irish Health Repository)

    Crowley, Lisa C

    2012-01-31

    Although Imatinib has transformed the treatment of chronic myeloid leukemia (CML), it is not curative due to the persistence of resistant cells that can regenerate the disease. We have examined how Bcr-Abl-expressing cells respond to two mechanistically different therapeutic agents, etoposide and Imatinib. We also examined Bcr-Abl expression at low and high levels as elevated expression has been associated with treatment failure. Cells expressing low levels of Bcr-Abl undergo apoptosis in response to the DNA-targeting agent (etoposide), whereas high-Bcr-Abl-expressing cells primarily induce autophagy. Autophagic populations engage a delayed nonapoptotic death; however, sufficient cells evade this and repopulate following the withdrawal of the drug. Non-Bcr-Abl-expressing 32D or Ba\\/F3 cells induce both apoptosis and autophagy in response to etoposide and can recover. Imatinib treatment induces both apoptosis and autophagy in all Bcr-Abl-expressing cells and populations rapidly recover. Inhibition of autophagy with ATG7 and Beclin1 siRNA significantly reduced the recovery of Imatinib-treated K562 cells, indicating the importance of autophagy for the recovery of treated cells. Combination regimes incorporating agents that disrupt Imatinib-induced autophagy would remain primarily targeted and may improve response to the treatment in CML.

  15. Cytoprotective effect of imatinib mesylate in non-BCR-ABL-expressing cells along with autophagosome formation

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    Ohtomo, Tadashi [Department of Biochemistry, Tokyo Medical University, Tokyo (Japan); Miyazawa, Keisuke, E-mail: miyazawa@tokyo-med.ac.jp [Department of Biochemistry, Tokyo Medical University, Tokyo (Japan); Naito, Munekazu [Department of Anatomy, Tokyo Medical University, Tokyo (Japan); Moriya, Shota [Department of Biochemistry, Tokyo Medical University, Tokyo (Japan); Kuroda, Masahiko [Department of Molecular Pathology, Tokyo Medical University, Tokyo (Japan); Itoh, Masahiro [Department of Anatomy, Tokyo Medical University, Tokyo (Japan); Tomoda, Akio [Department of Biochemistry, Tokyo Medical University, Tokyo (Japan)

    2010-01-01

    Treatment with imatinib mesylate (IM) results in an increased viable cell number of non-BCR-ABL-expressing cell lines by inhibiting spontaneous apoptosis. Electron microscopy revealed an increase of autophagosomes in response to IM. IM attenuated the cytotoxic effect of cytosine arabinoside, as well as inhibiting cell death with serum-deprived culture. Cytoprotection with autophagosome formation by IM was observed in various leukemia and cancer cell lines as well as normal murine embryonic fibroblasts (MEFs). Complete inhibition of autophagy by knockdown of atg5 in the Tet-off atg5{sup -/-} MEF system attenuated the cytoprotective effect of IM, indicating that the effect is partially dependent on autophagy. However, cytoprotection by IM was not mediated through suppression of ROS production via mitophagy, ER stress via ribophagy, or proapoptotic function of ABL kinase. Although the target tyrosine kinase(s) of IM remains unclear, our data provide novel therapeutic possibilities of using IM for cytoprotection.

  16. The Role of MEK Kinase 1 in Bcr-Abl-Induced Self-Renewal Activity of Embryonic Stem Cells

    OpenAIRE

    NAKAMURA, Yukinori; Yujiri, Toshiaki; Tanizawa, Yukio

    2006-01-01

    BCR-ABL oncogene, the moecular hallmark of chronic mylogenous leukemia, arises in a promitive hematopoietic stem cell that has the capacity for both differentiation and self-renewal. Its product, Bcr-Abl protein, has been shown to sctivate signal transducers and activators of transcription 3 (STAT3) and to promote self-renewal in embryonic stem (ES)cells, even in the absencs of leukemia in hibitory factor (LIF). MEK kinase 1 (MEKK1) is a 196-kDa mitogen-activated protein kinase (MAPK) kinase ...

  17. Perturbation of energy metabolism by fatty-acid derivative AIC-47 and imatinib in BCR-ABL-harboring leukemic cells.

    Science.gov (United States)

    Shinohara, Haruka; Kumazaki, Minami; Minami, Yosuke; Ito, Yuko; Sugito, Nobuhiko; Kuranaga, Yuki; Taniguchi, Kohei; Yamada, Nami; Otsuki, Yoshinori; Naoe, Tomoki; Akao, Yukihiro

    2016-02-01

    In Ph-positive leukemia, imatinib brought marked clinical improvement; however, further improvement is needed to prevent relapse. Cancer cells efficiently use limited energy sources, and drugs targeting cellular metabolism improve the efficacy of therapy. In this study, we characterized the effects of novel anti-cancer fatty-acid derivative AIC-47 and imatinib, focusing on cancer-specific energy metabolism in chronic myeloid leukemia cells. AIC-47 and imatinib in combination exhibited a significant synergic cytotoxicity. Imatinib inhibited only the phosphorylation of BCR-ABL; whereas AIC-47 suppressed the expression of the protein itself. Both AIC-47 and imatinib modulated the expression of pyruvate kinase M (PKM) isoforms from PKM2 to PKM1 through the down-regulation of polypyrimidine tract-binding protein 1 (PTBP1). PTBP1 functions as alternative splicing repressor of PKM1, resulting in expression of PKM2, which is an inactive form of pyruvate kinase for the last step of glycolysis. Although inactivation of BCR-ABL by imatinib strongly suppressed glycolysis, compensatory fatty-acid oxidation (FAO) activation supported glucose-independent cell survival by up-regulating CPT1C, the rate-limiting FAO enzyme. In contrast, AIC-47 inhibited the expression of CPT1C and directly fatty-acid metabolism. These findings were also observed in the CD34(+) fraction of Ph-positive acute lymphoblastic leukemia cells. These results suggest that AIC-47 in combination with imatinib strengthened the attack on cancer energy metabolism, in terms of both glycolysis and compensatory activation of FAO. PMID:26607903

  18. Chibby drives β catenin cytoplasmic accumulation leading to activation of the unfolded protein response in BCR-ABL1+ cells.

    Science.gov (United States)

    Mancini, Manuela; Leo, Elisa; Takemaru, Ken-Ichi; Campi, Virginia; Borsi, Enrica; Castagnetti, Fausto; Gugliotta, Gabriele; Santucci, Maria Alessandra; Martinelli, Giovanni

    2013-09-01

    Chronic myeloid leukemia (CML) is a myeloproliferative disease caused by the constitutive tyrosine kinase (TK) activity of the BCR-ABL fusion protein. However, the phenotype of leukemic stem cells (LSC) is sustained by β catenin rather than by the BCR-ABL TK. β catenin activity in CML is contingent upon its stabilization proceeding from the BCR-ABL-induced phosphorylation at critical residues for interaction with the Adenomatous polyposis coli (APC)/Axin/glycogen synthase kinase 3 (GSK3) destruction complex or GSK3 inactivating mutations. Here we studied the impact of β catenin antagonist Chibby (CBY) on β catenin signaling in BCR-ABL1+ cells. CBY is a small conserved protein which interacts with β catenin and impairs β catenin-mediated transcriptional activation through two distinct molecular mechanisms: 1) competition with T cell factor (TCF) or lymphoid enhancer factor (LEF) for β catenin binding; and 2) nuclear export of β catenin via interaction with 14-3-3. We found that its enforced expression in K562 cell line promoted β catenin cytoplasmic translocation resulting in inhibition of target gene transcription. Moreover, cytoplasmic accumulation of β catenin activated the endoplasmic reticulum (ER) stress-associated pathway known as unfolded protein response (UPR). CBY-driven cytoplasmic accumulation of β catenin is also a component of BCR-ABL1+ cell response to the TK inhibitor Imatinib (IM). It evoked the UPR activation leading to the induction of BCL2-interacting mediator of cell death (BIM) by UPR sensors. BIM, in turn, contributed to the execution phase of apoptosis in the activation of ER resident caspase 12 and mobilization of Ca(2+) stores. PMID:23707389

  19. Inhibitory Effects of Omacetaxine on Leukemic Stem Cells and BCR-ABL-Induced Chronic Myeloid Leukemia and Acute Lymphoblastic Leukemia in Mice

    OpenAIRE

    Chen, Yaoyu; Hu, Yiguo; Michaels, Shawnya; Segal, David; Brown, Dennis; Li, Shaoguang

    2009-01-01

    Omacetaxine mepesuccinate (formerly homoharringtonine) is a molecule with a mechanism of action that is different from tyrosine kinase inhibitors and its activity in chronic myeloid leukemia (CML) seems to be independent of BCR-ABL mutation status. Using BCR-ABL-expressing myelogenous and lymphoid cell lines and mouse models of CML and B cell acute lymphoblastic leukemia (B-ALL) induced by wild type BCR-ABL or T315I mutant-BCR-ABL, we evaluated the inhibitory effects of omacetaxine on CML and...

  20. Location of CTP-OD1-HA and CTP-OD2-HA fusion peptide in K562 cells and its interaction with BCR-ABL protein

    Directory of Open Access Journals (Sweden)

    Heng XIAO

    2013-11-01

    Full Text Available Objective To investigate the intracellular location of cytoplasmic transduction peptide-oligomerization domain 1-hemagglutinin (CTP-OD1-HA and cytoplasmic transduction peptide-oligomerization domain 2-hemagglutinin (CTP-OD2-HA fusion peptide and interaction with BCR-ABL protein in chronic myelocytic leukemia (CML K562 cells. Methods The prepared CTP-OD1-HA and CTP-OD2-HA fusion peptide were transduced into K562 cells, and the cytoplasmic transduction peptideoligomerization domain-hemagglutinin (CTP-OD-HA peptide was used as positive control, and cytoplasmic transduction peptidehemagglutinin (CTP-HA and PBS were used as negative control. The intracellular location of the fusion peptide was demonstrated by immunofluorescence and Western blotting. The interactions between the fusion peptides and BCR-ABL protein were detected by His-pull down and CoIP test. Results Immunofluorescence assay showed that both fusion peptides CTP-OD1-HA and CTP-OD2-HA could penetrate the cell membrane, and they were mainly localized in cytoplasm. Western blotting also confirmed the existence of the two fusion peptides in the cytoplasm. His-pull down showed that CTP-OD1-HA and CTP-OD2-HA could directly bind BCRABL protein outside the cells, and CoIP experiment showed that both fusion peptides could interact with BCR-ABL protein and form complex in the K562 cells. The negative control CTP-HA fusion peptide and PBS showed no such effects. Conclusion CTPOD1-HA and CTP-OD2-HA fusion peptides can successfully target into CML K562 cells and locate in cytoplasm, and it interacts with BCR-ABL protein. DOI: 10.11855/j.issn.0577-7402.2013.11.006

  1. MEK kinase 1 is essential for Bcr-Abl-induced STAT3 and self-renewal activity in embryonic stem cells.

    Science.gov (United States)

    Nakamura, Yukinori; Yujiri, Toshiaki; Nawata, Ryouhei; Tagami, Kozo; Tanizawa, Yukio

    2005-11-17

    BCR-ABL oncogene, the molecular hallmark of chronic myelogenous leukemia, arises in a primitive hematopoietic stem cell that has the capacity for both differentiation and self-renewal. Its product, Bcr-Abl protein, has been shown to activate signal transducers and activators of transcription 3 (STAT3) and to promote self-renewal in embryonic stem (ES) cells, even in the absence of leukemia inhibitory factor (LIF). MEK kinase 1 (MEKK1) is a 196-kDa mitogen-activated protein kinase (MAPK) kinase kinase involved in Bcr-Abl signal transduction. To investigate the role of MEKK1 in Bcr-Abl-induced transformation of stem cells, p210 Bcr-Abl was stably transfected into wild-type (WT(p210)) and MEKK1-/- (MEKK1-/-(p210)) ES cells. Bcr-Abl enhanced MEKK1 expression in ES transfectants, as it does in other Bcr-Abl-transformed cells. In the absence of LIF, WT(p210) cells showed constitutive STAT3 activation and formed rounded, compact colonies having strong alkaline phosphatase activity, a characteristic phenotype of undifferentiated ES cells. MEKK1-/-(p210) cells, by contrast, showed less STAT3 activity than WT(p210) cells and formed large, flattened colonies having weak alkaline phosphatase activity, a phenotype of differentiated ES cells. These results indicate that MEKK1 plays a key role in Bcr-Abl-induced STAT3 activation and in ES cells' capacity for LIF-independent self-renewal, and may thus be involved in Bcr-Abl-mediated leukemogenesis in stem cells. PMID:16044153

  2. Additive antileukemia effects by GFI1B- and BCR-ABL-specific siRNA in advanced phase chronic myeloid leukemic cells.

    Science.gov (United States)

    Koldehoff, M; Zakrzewski, J L; Beelen, D W; Elmaagacli, A H

    2013-07-01

    Previous studies demonstrated selective inhibition of the BCR-ABL (breakpoint cluster region-Abelson murine leukemia oncogene) tyrosine kinase by RNA interference in leukemic cells. In this study, we evaluated the effect of BCR-ABL small interfering RNA (siRNA) and GFI1B siRNA silencing on chronic myeloid leukemia (CML) cells in myeloid blast crises. The GFI1B gene was mapped to chromosome 9 and is, therefore, located downstream of the BCR-ABL translocation in CML cells. Co-transfection of BCR-ABL siRNA and GFI1B siRNA dramatically decreased cell viability and significantly induced apoptosis and inhibited proliferation in K562 cells (P<0.0001) and primary advanced phase CML cells (P<0.0001) versus controls. Furthermore, combining of BCR-ABL siRNA and GFI1B siRNA significantly modified the expression of several relevant genes including Myc, MDR1, MRP1 and tyrosyl-phosphoproteins in primary CML cells. Our data suggest that silencing of both BCR-ABL siRNA and GFI1B siRNA is associated with an additive antileukemic effect against K562 cells and primary advanced CML cells, further validating these genes as attractive therapeutic targets. PMID:23788109

  3. Stat5 Exerts Distinct, Vital Functions in the Cytoplasm and Nucleus of Bcr-Abl+ K562 and Jak2(V617F)+ HEL Leukemia Cells

    International Nuclear Information System (INIS)

    Signal transducers and activators of transcription (Stats) play central roles in the conversion of extracellular signals, e.g., cytokines, hormones and growth factors, into tissue and cell type specific gene expression patterns. In normal cells, their signaling potential is strictly limited in extent and duration. The persistent activation of Stat3 or Stat5 is found in many human tumor cells and contributes to their growth and survival. Stat5 activation plays a pivotal role in nearly all hematological malignancies and occurs downstream of oncogenic kinases, e.g., Bcr-Abl in chronic myeloid leukemias (CML) and Jak2(V617F) in other myeloproliferative diseases (MPD). We defined the mechanisms through which Stat5 affects growth and survival of K562 cells, representative of Bcr-Abl positive CML, and HEL cells, representative for Jak2(V617F) positive acute erythroid leukemia. In our experiments we suppressed the protein expression levels of Stat5a and Stat5b through shRNA mediated downregulation and demonstrated the dependence of cell survival on the presence of Stat5. Alternatively, we interfered with the functional capacities of the Stat5 protein through the interaction with a Stat5 specific peptide ligand. This ligand is a Stat5 specific peptide aptamer construct which comprises a 12mer peptide integrated into a modified thioredoxin scaffold, S5-DBD-PA. The peptide sequence specifically recognizes the DNA binding domain (DBD) of Stat5. Complex formation of S5-DBD-PA with Stat5 causes a strong reduction of P-Stat5 in the nuclear fraction of Bcr-Abl-transformed K562 cells and a suppression of Stat5 target genes. Distinct Stat5 mediated survival mechanisms were detected in K562 and Jak2(V617F)-transformed HEL cells. Stat5 is activated in the nuclear and cytosolic compartments of K562 cells and the S5-DBD-PA inhibitor most likely affects the viability of Bcr-Abl+ K562 cells through the inhibition of canonical Stat5 induced target gene transcription. In HEL cells, Stat5 is

  4. Stat5 Exerts Distinct, Vital Functions in the Cytoplasm and Nucleus of Bcr-Abl+ K562 and Jak2(V617F)+ HEL Leukemia Cells.

    Science.gov (United States)

    Weber, Axel; Borghouts, Corina; Brendel, Christian; Moriggl, Richard; Delis, Natalia; Brill, Boris; Vafaizadeh, Vida; Groner, Bernd

    2015-01-01

    Signal transducers and activators of transcription (Stats) play central roles in the conversion of extracellular signals, e.g., cytokines, hormones and growth factors, into tissue and cell type specific gene expression patterns. In normal cells, their signaling potential is strictly limited in extent and duration. The persistent activation of Stat3 or Stat5 is found in many human tumor cells and contributes to their growth and survival. Stat5 activation plays a pivotal role in nearly all hematological malignancies and occurs downstream of oncogenic kinases, e.g., Bcr-Abl in chronic myeloid leukemias (CML) and Jak2(V617F) in other myeloproliferative diseases (MPD). We defined the mechanisms through which Stat5 affects growth and survival of K562 cells, representative of Bcr-Abl positive CML, and HEL cells, representative for Jak2(V617F) positive acute erythroid leukemia. In our experiments we suppressed the protein expression levels of Stat5a and Stat5b through shRNA mediated downregulation and demonstrated the dependence of cell survival on the presence of Stat5. Alternatively, we interfered with the functional capacities of the Stat5 protein through the interaction with a Stat5 specific peptide ligand. This ligand is a Stat5 specific peptide aptamer construct which comprises a 12mer peptide integrated into a modified thioredoxin scaffold, S5-DBD-PA. The peptide sequence specifically recognizes the DNA binding domain (DBD) of Stat5. Complex formation of S5-DBD-PA with Stat5 causes a strong reduction of P-Stat5 in the nuclear fraction of Bcr-Abl-transformed K562 cells and a suppression of Stat5 target genes. Distinct Stat5 mediated survival mechanisms were detected in K562 and Jak2(V617F)-transformed HEL cells. Stat5 is activated in the nuclear and cytosolic compartments of K562 cells and the S5-DBD-PA inhibitor most likely affects the viability of Bcr-Abl+ K562 cells through the inhibition of canonical Stat5 induced target gene transcription. In HEL cells, Stat5 is

  5. Stat5 Exerts Distinct, Vital Functions in the Cytoplasm and Nucleus of Bcr-Abl+ K562 and Jak2(V617F+ HEL Leukemia Cells

    Directory of Open Access Journals (Sweden)

    Axel Weber

    2015-03-01

    Full Text Available Signal transducers and activators of transcription (Stats play central roles in the conversion of extracellular signals, e.g., cytokines, hormones and growth factors, into tissue and cell type specific gene expression patterns. In normal cells, their signaling potential is strictly limited in extent and duration. The persistent activation of Stat3 or Stat5 is found in many human tumor cells and contributes to their growth and survival. Stat5 activation plays a pivotal role in nearly all hematological malignancies and occurs downstream of oncogenic kinases, e.g., Bcr-Abl in chronic myeloid leukemias (CML and Jak2(V617F in other myeloproliferative diseases (MPD. We defined the mechanisms through which Stat5 affects growth and survival of K562 cells, representative of Bcr-Abl positive CML, and HEL cells, representative for Jak2(V617F positive acute erythroid leukemia. In our experiments we suppressed the protein expression levels of Stat5a and Stat5b through shRNA mediated downregulation and demonstrated the dependence of cell survival on the presence of Stat5. Alternatively, we interfered with the functional capacities of the Stat5 protein through the interaction with a Stat5 specific peptide ligand. This ligand is a Stat5 specific peptide aptamer construct which comprises a 12mer peptide integrated into a modified thioredoxin scaffold, S5-DBD-PA. The peptide sequence specifically recognizes the DNA binding domain (DBD of Stat5. Complex formation of S5-DBD-PA with Stat5 causes a strong reduction of P-Stat5 in the nuclear fraction of Bcr-Abl-transformed K562 cells and a suppression of Stat5 target genes. Distinct Stat5 mediated survival mechanisms were detected in K562 and Jak2(V617F-transformed HEL cells. Stat5 is activated in the nuclear and cytosolic compartments of K562 cells and the S5-DBD-PA inhibitor most likely affects the viability of Bcr-Abl+ K562 cells through the inhibition of canonical Stat5 induced target gene transcription. In HEL

  6. A report case of mixed phenotype acute leukemia with BCR - ABL positive and literature review%BCR - ABL 融合基因阳性的急性混合细胞白血病1例并文献复习

    Institute of Scientific and Technical Information of China (English)

    符爽; 张男; 陈芳; 刘璇; 胡延平; 张旻昱; 张继红

    2014-01-01

    Objective:To investigate the clinical characteristics,biological features,clinical curative effect and out-come of mixed PhenotyPe acute leukemia(MPAL). Methods:Results of clinical data,cell morPhology,immunoPheno-tyPe,cytogenetics and molecular genetics of the MPAL Patient with BCR - ABL Positive were analyzed with literature reviews. Results:The clinical and hematological findings were comPatible with the diagnosis of MPAL. LymPhoid and myeloid markers were Positive on the leukemia cells. KaryotyPe analysis showed that the Patient had t(9;22)(q34;q11)translocation. Quantitative reverse transcriPtion PCR results showed that BCR - ABL(P210)exPressed 100% . And no deletion,Point or insert mutation was found among C - Kit(8,17 exon),FLT3 - ITD/ TKD,NPM1 and CEBPα. Conclusion:Primary MPAL with BCR - ABL Positive is rarely rePorted which have a Poor Prognosis. Some of them are cases of chronic myeloid leukemia blast crisis. BCR - ABL gene exPression is closely related to Patients' Prognosis.%目的:探讨 BCR - ABL 融合基因阳性表达的急性混合细胞白血病(MPAL)的临床特点、生物学特性、临床疗效及预后。方法:对1例 BCR - ABL 融合基因阳性的急性混合细胞白血病患者的临床资料、细胞形态学、免疫分型、细胞遗传学及分子生物学的检测结果进行综合分析。结果:患者的临床表现和实验室检测结果均符合急性混合细胞白血病诊断标准,免疫表型分析白血病细胞同时表达髓系和淋系抗原,染色体核型为46,XX,t(9;22)(q34;q11),荧光定量 PCR 检测结果示 BCR - ABL(P210)融合基因表达100%,白血病预后基因检测结果示患者 C - Kit(8、17 exon)、FLT3- ITD/ TKD、NPM1和 CEBPα基因均为野生型。结论:初发的 BCR - ABL 融合基因阳性的急性混合细胞白血病病例罕见,部分病例由慢性粒细胞白血病急变而来,预后差。BCR - ABL 基因阳性与患者预后密切相关。

  7. Allogeneic stem cell transplantation for patients harboring T315I BCR-ABL mutated leukemias

    DEFF Research Database (Denmark)

    Nicolini, Franck Emmanuel; Basak, Grzegorz W; Soverini, Simona;

    2011-01-01

    , still alive]) for those with chronic myeloid leukemia in the blast phase and 7.4 months (range 1.4 months to not reached [ie, still alive]) for those with Philadelphia chromosome-positive acute lymphoblastic leukemia but has not yet been reached for those in the chronic and accelerated phases of chronic...... myeloid leukemia. The occurrence of chronic GVHD had a positive impact on overall survival (P = .047). Transplant-related mortality rates were low. Multivariate analysis identified only blast phase at transplantation (hazard ratio 3.68, P = .0011) and unrelated stem cell donor (hazard ratio 2.98, P = .011...

  8. Allogeneic stem cell transplantation for patients harboring T315I BCR-ABL mutated leukemias

    DEFF Research Database (Denmark)

    Nicolini, Franck Emmanuel; Basak, Grzegorz W; Soverini, Simona;

    2011-01-01

    , still alive]) for those with chronic myeloid leukemia in the blast phase and 7.4 months (range 1.4 months to not reached [ie, still alive]) for those with Philadelphia chromosome-positive acute lymphoblastic leukemia but has not yet been reached for those in the chronic and accelerated phases of chronic...

  9. Involvement of primary mesenchymal precursors and hematopoietic bone marrow cells from chronic myeloid leukemia patients by BCR-ABL1 fusion gene.

    Science.gov (United States)

    Chandia, Mauricio; Sayagués, José-María; Gutiérrez, María-Laura; Chillón, María-Laura; Aristizábal, José-Alejandro; Corrales, Alejandro; Castellanos, Marta; Melón, Alberto; Sánchez, María-Luz; Bárcena, Paloma; Matarraz, Sergio; González-González, María; Barrena, Susana; López, Antonio; Cañizo, María-Consuelo; Sánchez-Guijo, Fermín; Orfao, Alberto

    2014-03-01

    For decades now, it is well established that chronic myeloid leukemia (CML) is a hematopoietic stem cell(HPC) disorder. However, it remains to be determined whether BCR-ABL1 gene rearrangement occurs in a HPC or at an earlier stem cell and whether the degree of involvement of hematopoiesis by the BCR-ABL1 fusion gene relates to the response to therapy. Here, we have investigated by interphase fluorescence in situ hybridization (iFISH) the distribution of BCR-ABL1 fusion gene in FACS-sorted bone marrow (BM) populations of mesenchymal precursor cells (MPC) and other hematopoietic cell populations from 18 newly diagnosed CML patients. Overall, our results showed systematic involvement at relatively high percentages of BM maturing neutrophils (97%615%), basophils (95%612%), eosinophils (90%68%), CD341 precursors cells (90%67%),monocytes (84%630%), nucleated red blood cells (87%624%), and mast cells (77%633%). By contrast, MPC(30%634%), B-cells (15%627%), T-lymphocytes (50%626%), and NK-cells (35%634%) were involved at lower percentages. In 8/18 CML patients, 2 tumor BCR-ABL11 subclones were detected by iFISH. Of note, all tumor cell subclones were systematically detected in CD341 cells, whereas MPC were only involved by the ancestral tumor cell subclone. In summary, here we confirm the presence at diagnosis of the BCR-ABL1 fusion gene inMPC, CD341 precursors, and other different BM hematopoietic myeloid cell lineages from CML patients,including also in a significant fraction of cases, a smaller percentage of T, B, and NK lymphocytes.Interestingly, involvement of MPC was restricted to the ancestral BCR-ABL11 subclone. PMID:24779036

  10. Gads (Grb2-related adaptor downstream of Shc) is required for BCR-ABL-mediated lymphoid leukemia

    Science.gov (United States)

    Gillis, LC; Berry, DM; Minden, MD; McGlade, CJ; Barber, DL

    2016-01-01

    Philadelphia chromosome-positive leukemias, including chronic myeloid leukemia and B-cell acute lymphoblastic leukemia (B-ALL), are driven by the oncogenic BCR-ABL fusion protein. Animal modeling experiments utilizing retroviral transduction and subsequent bone marrow transplantation have demonstrated that BCR-ABL generates both myeloid and lymphoid disease in mice receiving whole bone marrow transduced with BCR-ABL. Y177 of BCR-ABL is critical to the development of myeloid disease, and phosphorylation of Y177 has been shown to induce GRB2 binding to BCR-ABL, followed by activation of the Ras and phosphoinositide 3 kinase signaling pathways. We show that the GRB2-related adapter protein, GADS, also associates with BCR-ABL, specifically through Y177 and demonstrate that BCR-ABL-driven lymphoid disease requires Gads. BCR-ABL transduction of Gads(−/−) bone marrow results in short latency myeloid disease within 3–4 weeks of transplant, while wild-type mice succumb to both a longer latency lymphoid and myeloid diseases. We report that GADS mediates a unique BCR-ABL complex with SLP-76 in BCR-ABL-positive cell lines and B-ALL patient samples. These data suggest that GADS mediates lymphoid disease downstream of BCR-ABL through the recruitment of specific signaling intermediates. PMID:23399893

  11. miR-29b suppresses CML cell proliferation and induces apoptosis via regulation of BCR/ABL1 protein

    International Nuclear Information System (INIS)

    MicroRNAs (miRNAs) are small RNAs that regulate gene expression posttranscriptionally and are critical for many cellular pathways. Recent evidence has shown that aberrant miRNA expression profiles and unique miRNA signaling pathways are present in many cancers. Here, we demonstrate that miR-29b is markedly lower expressed in CML patient samples. Bioinformatics analysis reveals a conserved target site for miR-29b in the 3′-untranslated region (UTR) of ABL1. miR-29b significantly suppresses the activity of a luciferase reporter containing ABL1-3′UTR and this activity is not observed in cells transfected with mutated ABL1-3′UTR. Enforced expression of miR-29b in K562 cells inhibits cell growth and colony formation ability thereby inducing apoptosis through cleavage of procaspase 3 and PARP. Furthermore, K562 cells transfected with a siRNA targeting ABL1 show similar growth and apoptosis phenotypes as cells overexpression of miR-29b. Collectively, our results suggest that miR-29b may function as a tumor suppressor by targeting ABL1 and BCR/ABL1. - Highlights: ► miR-29b expression was downregulated in CML patients. ► ABL1 was identified as a direct target gene of miR-29b. ► Enforced expression of miR-29b inhibits cell proliferation and induces apoptosis. ► miR-29b might be a therapeutic target to CML

  12. Allogeneic Stem Cell Transplantation for Patients with T315I BCR-ABL Mutated Chronic Myeloid Leukemia.

    Science.gov (United States)

    Xu, Lan-Ping; Xu, Zheng-Li; Zhang, Xiao-Hui; Chen, Huan; Chen, Yu-Hong; Han, Wei; Chen, Yao; Wang, Feng-Rong; Wang, Jing-Zhi; Wang, Yu; Yan, Chen-Hua; Mo, Xiao-Dong; Liu, Kai-Yan; Huang, Xiao-Jun

    2016-06-01

    Allogeneic stem cell transplantation (SCT) is currently the only curative treatment option for chronic myeloid leukemia (CML) patients with BCR-ABL T315I mutations. We report the outcome of SCT in 22 patients with T315I(+) CML, most (n = 16) from haploidentical family donors (HID-SCT). At the time the mutation was detected, 8 patients were in the chronic phase (CP), 7 in the accelerated phase (AP), and 7 in the blast phase (BP). At the time of SCT 7 were in the CP, 8 in the AP or returning to the CP post-AP (AP/AP-CPn), and 7 in the BP or returning to CP post-BP (BP/BP-CPn). The cumulative incidence of grades III to IV acute graft-versus-host disease was 9.1%. Chronic graft-versus-host disease was observed in 60.0% of patients, including 25.0% who suffered from severe disease. Four patients died of transplant-related complications at a median interval from SCT of 16.3 months. The estimated 2-year leukemia-free survival rate was 80.0%, 72.9%, and 0% in CP, AP/AP-CPn and BP/BP-CPn groups at the time of SCT, respectively. After a median follow-up of 17.3 months from SCT, 14 patients are alive, including 13 in complete molecular response and 1 with an extramedullary relapse. In conclusion, HID-SCT is a potentially curative treatment for T315I + CML patients. For patients in CP/AP, immediate SCT might result in promising survival. The outcome of patients in BP with T315I(+) mutation remains very poor. PMID:26995693

  13. MicroRNA-1301-Mediated RanGAP1 Downregulation Induces BCR-ABL Nuclear Entrapment to Enhance Imatinib Efficacy in Chronic Myeloid Leukemia Cells.

    Directory of Open Access Journals (Sweden)

    Tsung-Yao Lin

    Full Text Available Chronic myeloid leukemia (CML is a myeloproliferative disease. Imatinib (IM, the first line treatment for CML, is excessively expensive and induces various side effects in CML patients. Therefore, it is essential to investigate a new strategy for improving CML therapy. Our immunoblot data revealed that RanGTPase activating protein 1 (RanGAP1 protein levels increased by approximately 30-fold in K562 cells compared with those in normal cells. RanGAP1 is one of the important components of RanGTPase system, which regulates the export of nuclear protein. However, whether RanGAP1 level variation influences BCR-ABL nuclear export is still unknown. In this report, using shRNA to downregulate RanGAP1 expression level augmented K562 cell apoptosis by approximately 40% after treatment with 250 nM IM. Immunofluorescence assay also indicated that three-fold of nuclear BCR-ABL was detected. These data suggest that BCR-ABL nuclear entrapment induced by RanGAP1 downregulation can be used to improve IM efficacy. Moreover, our qRT-PCR data indicated a trend of inverse correlation between the RanGAP1 and microRNA (miR-1301 levels in CML patients. MiR-1301, targeting the RanGAP1 3' untranslated region, decreased by approximately 100-fold in K562 cells compared with that in normal cells. RanGAP1 downregulation by miR-1301 transfection impairs BCR-ABL nuclear export to increase approximately 60% of cell death after treatment of 250 nM IM. This result was almost the same as treatment with 1000 nM IM alone. Furthermore, immunofluorescence assay demonstrated that Tyr-99 of nuclear P73 was phosphorylated accompanied with nuclear entrapment of BCR-ABL after transfection with RanGAP1 shRNA or miR-1301 in IM-treated K562 cells. Altogether, we demonstrated that RanGAP1 downregulation can mediate BCR-ABL nuclear entrapment to activate P73-dependent apoptosis pathway which is a novel strategy for improving current IM treatment for CML.

  14. Cell sorting enables interphase fluorescence in situ hybridization detection of low BCR-ABL1 producing stem cells in chronic myeloid leukaemia patients beyond deep molecular remission

    DEFF Research Database (Denmark)

    van Kooten Niekerk, Peter Buur; Petersen, Charlotte Christie; Nyvold, Charlotte Guldborg;

    2014-01-01

    The exact disease state of chronic myeloid leukaemia (CML) patients in deep molecular remission is unknown, because even the most sensitive quantitative reverse transcription polymerase chain reaction (qPCR) methods cannot identify patients prone to relapse after treatment withdrawal. To elucidate......) ), n = 11) using both sensitive qPCR and interphase fluorescence in situ hybridization (iFISH). Despite evaluating fewer cells, iFISH proved superior to mRNA-based qPCR in detecting residual Ph(+) stem cells (P = 0·005), and detected Ph(+) stem- and progenitor cells in 9/10 patients at frequencies of 2......-14%. Moreover, while all qPCR(+) samples also were iFISH(+) , 9/33 samples were qPCR-/iFISH(+) , including all positive samples from MR(4) patients. Our findings show that residual Ph(+) cells are low BCR-ABL1 producers, and that DNA-based methods are required to assess the content of persisting Ph(+) stem...

  15. Allosteric inhibition enhances the efficacy of ABL kinase inhibitors to target unmutated BCR-ABL and BCR-ABL-T315I

    Directory of Open Access Journals (Sweden)

    Mian Afsar

    2012-09-01

    Full Text Available Abstract Background Chronic myelogenous leukemia (CML and Philadelphia chromosome-positive (Ph+ acute lymphatic leukemia (Ph + ALL are caused by the t(9;22, which fuses BCR to ABL resulting in deregulated ABL-tyrosine kinase activity. The constitutively activated BCR/ABL-kinase “escapes” the auto-inhibition mechanisms of c-ABL, such as allosteric inhibition. The ABL-kinase inhibitors (AKIs Imatinib, Nilotinib or Dasatinib, which target the ATP-binding site, are effective in Ph + leukemia. Another molecular therapy approach targeting BCR/ABL restores allosteric inhibition. Given the fact that all AKIs fail to inhibit BCR/ABL harboring the ‘gatekeeper’ mutation T315I, we investigated the effects of AKIs in combination with the allosteric inhibitor GNF2 in Ph + leukemia. Methods The efficacy of this approach on the leukemogenic potential of BCR/ABL was studied in Ba/F3 cells, primary murine bone marrow cells, and untransformed Rat-1 fibroblasts expressing BCR/ABL or BCR/ABL-T315I as well as in patient-derived long-term cultures (PDLTC from Ph + ALL-patients. Results Here, we show that GNF-2 increased the effects of AKIs on unmutated BCR/ABL. Interestingly, the combination of Dasatinib and GNF-2 overcame resistance of BCR/ABL-T315I in all models used in a synergistic manner. Conclusions Our observations establish a new approach for the molecular targeting of BCR/ABL and its resistant mutants using a combination of AKIs and allosteric inhibitors.

  16. Biodegradable charged polyester-based vectors (BCPVs) as an efficient non-viral transfection nanoagent for gene knockdown of the BCR-ABL hybrid oncogene in a human chronic myeloid leukemia cell line.

    Science.gov (United States)

    Yang, Chengbin; Panwar, Nishtha; Wang, Yucheng; Zhang, Butian; Liu, Maixian; Toh, Huiting; Yoon, Ho Sup; Tjin, Swee Chuan; Chong, Peter Han Joo; Law, Wing-Cheung; Chen, Chih-Kuang; Yong, Ken-Tye

    2016-04-28

    First-line therapy of chronic myelogenous leukemia (CML) has always involved the use of BCR-ABL tyrosine-kinase inhibitors which is associated with an abnormal chromosome called Philadelphia chromosome. Although the overall survival rate has been improved by the current therapeutic regime, the presence of resistance has resulted in limited efficacy. In this study, an RNA interference (RNAi)-based therapeutic regime is proposed with the aim to knockdown the BCR-ABL hybrid oncogene using small interfering RNA (siRNA). The siRNA transfection rates have usually been limited due to the declining contact probability among polyplexes and the non-adherent nature of leukemic cells. Our work aims at addressing this limitation by using a biodegradable charged polyester-based vector (BCPV) as a nanocarrier for the delivery of BCR-ABL-specific siRNA to the suspension culture of a K562 CML cell line. BCR-ABL siRNAs were encapsulated in the BCPVs by electrostatic force. Cell internalization was facilitated by the BCPV and assessed by confocal microscopy and flow cytometry. The regulation of the BCR-ABL level in K562 cells as a result of RNAi was analyzed by real-time polymerase chain reaction (RT-PCR). We observed that BCPV was able to form stable nanoplexes with siRNA molecules, even in the presence of fetal bovine serum (FBS), and successfully assisted in vitro siRNA transfection in the non-adherent K562 cells. As a consequence of downregulation of BCR-ABL, BCPV-siRNA nanoplexes inhibited cell proliferation and promoted cell apoptosis. All results were compared with a commercial transfection reagent, Lipofectamine2000™, which served as a positive control. More importantly, this class of non-viral vector exhibits biodegradable features and negligible cytotoxicity, thus providing a versatile platform to deliver siRNA to non-adherent leukemia cells with high transfection efficiency by effectively overcoming extra- and intra-cellular barriers. Due to the excellent in vitro

  17. Biodegradable charged polyester-based vectors (BCPVs) as an efficient non-viral transfection nanoagent for gene knockdown of the BCR-ABL hybrid oncogene in a human chronic myeloid leukemia cell line

    Science.gov (United States)

    Yang, Chengbin; Panwar, Nishtha; Wang, Yucheng; Zhang, Butian; Liu, Maixian; Toh, Huiting; Yoon, Ho Sup; Tjin, Swee Chuan; Chong, Peter Han Joo; Law, Wing-Cheung; Chen, Chih-Kuang; Yong, Ken-Tye

    2016-04-01

    First-line therapy of chronic myelogenous leukemia (CML) has always involved the use of BCR-ABL tyrosine-kinase inhibitors which is associated with an abnormal chromosome called Philadelphia chromosome. Although the overall survival rate has been improved by the current therapeutic regime, the presence of resistance has resulted in limited efficacy. In this study, an RNA interference (RNAi)-based therapeutic regime is proposed with the aim to knockdown the BCR-ABL hybrid oncogene using small interfering RNA (siRNA). The siRNA transfection rates have usually been limited due to the declining contact probability among polyplexes and the non-adherent nature of leukemic cells. Our work aims at addressing this limitation by using a biodegradable charged polyester-based vector (BCPV) as a nanocarrier for the delivery of BCR-ABL-specific siRNA to the suspension culture of a K562 CML cell line. BCR-ABL siRNAs were encapsulated in the BCPVs by electrostatic force. Cell internalization was facilitated by the BCPV and assessed by confocal microscopy and flow cytometry. The regulation of the BCR-ABL level in K562 cells as a result of RNAi was analyzed by real-time polymerase chain reaction (RT-PCR). We observed that BCPV was able to form stable nanoplexes with siRNA molecules, even in the presence of fetal bovine serum (FBS), and successfully assisted in vitro siRNA transfection in the non-adherent K562 cells. As a consequence of downregulation of BCR-ABL, BCPV-siRNA nanoplexes inhibited cell proliferation and promoted cell apoptosis. All results were compared with a commercial transfection reagent, Lipofectamine2000™, which served as a positive control. More importantly, this class of non-viral vector exhibits biodegradable features and negligible cytotoxicity, thus providing a versatile platform to deliver siRNA to non-adherent leukemia cells with high transfection efficiency by effectively overcoming extra- and intra-cellular barriers. Due to the excellent in vitro

  18. Outcome of treatment in adults with Philadelphia chromosome-positive and/or BCR-ABL--positive acute lymphoblastic leukemia-retrospective analysis of Polish Adult Leukemia Group (PALG).

    Science.gov (United States)

    Wrzesień-Kuś, A; Robak, T; Pluta, A; Zwolińska, M; Wawrzyniak, E; Wierzbowska, A; Skotnicki, A; Jakubas, B; Hołowiecki, J; Nowak, K; Kuliczkowski, K; Mazur, G; Haus, O; Dmoszyńska, A; Adamczyk-Cioch, M; Jedrzejczak, W W; Paluszewska, M; Konopka, L; Pałynyczko, G

    2006-06-01

    Patients with Philadelphia chromosome-positive (Ph+) and/or BCR-ABL+ acute lymphoblastic leukemia (ALL) have extremely poor prognoses. Most of these patients have additional, heterogenous karyotype abnormalities, the majority of which have uncertain clinical significance. In this study we analyzed the clinical characteristics, karyotype abnormalities, and outcome of 77 patients with Ph+ and/or BCR-ABL+ ALL registered in Poland in 1997-2004. In 31/55 patients with known karyotype, the sole t(9;22)(q34;q11) abnormality had been diagnosed; in one patient, variant translocation t(4;9;22)(q21q31.1;q34;q11), and additional abnormalities in 23 (42%) patients, had been diagnosed. The characteristics of the patients with Ph chromosome and additional abnormalities were not significantly different when compared with the entire analyzed group. Out of 77 patients, 54 (70%) achieved first complete remission (CR1) after one or more induction cycles. The overall survival (OS) probability of 2 years was 63, 43, and 17% for patients treated with allogeneic stem cell transplantation (alloSCT), autologous SCT, and chemotherapy, respectively (log rank p=0.002). Median OS from the time of alloSCT was significantly longer for patients transplanted in CR1 compared with alloSCT in CR >1 (p=0.032). There were no significant differences in CR rate, disease-free survival (DFS), and OS for patients with t(9;22) and additional abnormalities compared with the whole group. Only WBC >20 G/l at diagnosis adversely influenced OS probability (log rank p=0.0017). In conclusion, our data confirm poor outcome of Ph+ and/or BCR-ABL+ ALL. Only patients who received alloSCT in CR1 had longer DFS and OS. We have shown that additional karyotype abnormalities did not influence the clinical characteristics of the patients; however, their influence on treatment results needs to be further assessed. PMID:16523310

  19. Down-regulation of Thanatos-associated protein 11 by BCR-ABL promotes CML cell proliferation through c-Myc expression.

    Science.gov (United States)

    Nakamura, Satoki; Yokota, Daisuke; Tan, Lin; Nagata, Yasuyuki; Takemura, Tomonari; Hirano, Isao; Shigeno, Kazuyuki; Shibata, Kiyoshi; Fujisawa, Shinya; Ohnishi, Kazunori

    2012-03-01

    Bcr-Abl activates various signaling pathways in chronic myelogenous leukemia (CML) cells. The proliferation of Bcr-Abl transformed cells is promoted by c-Myc through the activation of Akt, JAK2 and NF-κB. However, the mechanism by which c-Myc regulates CML cell proliferation is unclear. In our study, we investigated the role of Thanatos-associated protein 11 (THAP11), which inhibits c-Myc transcription, in CML cell lines and in hematopoietic progenitor cells derived from CML patients. The induction of THAP11 expression by Abl kinase inhibitors in CML cell lines and in CML-derived hematopoietic progenitor cells resulted in the suppression of c-Myc. In addition, over-expression of THAP11 inhibited CML cell proliferation. In colony forming cells derived from CML-aldehyde dehydrogenase (ALDH)(hi) /CD34(+) cells, treatment with Abl kinase inhibitors and siRNA depletion of Bcr-Abl induced THAP11 expression and reduced c-Myc expression, resulting in inhibited colony formation. Moreover, overexpression of THAP11 significantly decreased the colony numbers, and also inhibited the expression of c-myc target genes such as Cyclin D1, ODC and induced the expression of p21(Cip1) . The depletion of THAP11 inhibited JAK2 or STAT5 inactivation-mediated c-Myc reduction in ALDH(hi) /CD34(+) CML cells. Thus, the induced THAP11 might be one of transcriptional regulators of c-Myc expression in CML cell. Therefore, the induction of THAP11 has a potential possibility as a target for the inhibition of CML cell proliferation. PMID:21400515

  20. Sensitive detection of pre-existing BCR-ABL kinase domain mutations in CD34+ cells of newly diagnosed chronic-phase chronic myeloid leukemia patients is associated with imatinib resistance: implications in the post-imatinib era.

    Directory of Open Access Journals (Sweden)

    Zafar Iqbal

    Full Text Available BACKGROUND: BCR-ABL kinase domain mutations are infrequently detected in newly diagnosed chronic-phase chronic myeloid leukemia (CML patients. Recent studies indicate the presence of pre-existing BCR-ABL mutations in a higher percentage of CML patients when CD34+ stem/progenitor cells are investigated using sensitive techniques, and these mutations are associated with imatinib resistance and disease progression. However, such studies were limited to smaller number of patients. METHODS: We investigated BCR-ABL kinase domain mutations in CD34+ cells from 100 chronic-phase CML patients by multiplex allele-specific PCR and sequencing at diagnosis. Mutations were re-investigated upon manifestation of imatinib resistance using allele-specific PCR and direct sequencing of BCR-ABL kinase domain. RESULTS: Pre-existing BCR-ABL mutations were detected in 32/100 patients and included F311L, M351T, and T315I. After a median follow-up of 30 months (range 8-48, all patients with pre-existing BCR-ABL mutations exhibited imatinib resistance. Of the 68 patients without pre-existing BCR-ABL mutations, 24 developed imatinib resistance; allele-specific PCR and BCR-ABL kinase domain sequencing detected mutations in 22 of these patients. All 32 patients with pre-existing BCR-ABL mutations had the same mutations after manifestation of imatinib-resistance. In imatinib-resistant patients without pre-existing BCR-ABL mutations, we detected F311L, M351T, Y253F, and T315I mutations. All imatinib-resistant patients except T315I and Y253F mutations responded to imatinib dose escalation. CONCLUSION: Pre-existing BCR-ABL mutations can be detected in a substantial number of chronic-phase CML patients by sensitive allele-specific PCR technique using CD34+ cells. These mutations are associated with imatinib resistance if affecting drug binding directly or indirectly. After the recent approval of nilotinib, dasatinib, bosutinib and ponatinib for treatment of chronic myeloid

  1. GENE THERAPY USING RETROVIRAL VECTOR OF bcr-abl SPECIFIC MULTI-UNIT RIBOZYMES COULD INHIBIT CML CELL GROWTH AND INDUCE APOPTOSIS

    Institute of Scientific and Technical Information of China (English)

    冯琦; 孙凯; 赵永同; 张涛; 尚振川; 王莎; 王玮; 赵宁; 颜真; 韩苇; 张英起; 孙秉中

    2003-01-01

    Objective: To investigate the in vitro cleavage ability and effects on apoptosis and cell growth of the bcr-abl fusion gene specific multi-unit ribozymes. Methods: Three fusion point specific ribozymes were designed and the multi-unit ribozymes' in vitro transcription vector and retroviral vector were constructed. The in vitro cleavage ability was tested. The retroviral vector was transfected into K562 cell and the effects on proliferation, apoptosis, cell cycle and cell structure were observed. Results: Multi-unit ribozymes had in vitro cleavage efficiency of 70.8%, which was more efficient than single-unit and double-unit ribozymes. Transfection of the retroviral vector of the ribozyme into K562 cells, induced inhibition of cell growth and apoptosis. The incorporation rate of DNA in ribozymes transfected K562 cells was greatly decreased along with time passed, with an inhibition rate of more than 50% after 96 h of transfection. Under FCM, 18.4% of the cells underwent apoptosis 72 h after transfection and more cells were blocked in G phase, with the ratio in S phase greatly decreased (41.9%). Under electron microscope, compaction of nuclear chromatin and apoptosis bodies were observed.Conclusion: Multi-unit ribozymes specific to bcr-abl fusion gene can be used to treat CML and to purge bone marrow for self-grafting.

  2. Induction of autophagy by Imatinib sequesters Bcr-Abl in autophagosomes and down-regulates Bcr-Abl protein.

    LENUS (Irish Health Repository)

    Elzinga, Baukje M

    2013-06-01

    Chronic Myeloid Leukemia (CML) is a disease of hematopoietic stem cells which harbor the chimeric gene Bcr-Abl. Expression levels of this constitutively active tyrosine kinase are critical for response to tyrosine kinase inhibitor treatment and also disease progression, yet the regulation of protein stability is poorly understood. We have previously demonstrated that imatinib can induce autophagy in Bcr-Abl expressing cells. Autophagy has been associated with the clearance of large macromolecular signaling complexes and abnormal proteins, however, the contribution of autophagy to the turnover of Bcr-Abl protein in imatinib treated cells is unknown. In this study, we show that following imatinib treatment, Bcr-Abl is sequestered into vesicular structures that co-localize with the autophagy marker LC3 or GABARAP. This association is inhibited by siRNA mediated knockdown of autophagy regulators (Beclin 1\\/ATG7). Pharmacological inhibition of autophagy also reduced Bcr-Abl\\/LC3 co-localization in both K562 and CML patient cells. Bcr-Abl protein expression was reduced with imatinib treatment. Inhibition of both autophagy and proteasome activity in imatinib treated cells was required to restore Bcr-Abl protein levels to those of untreated cells. This ability to down-regulate Bcr-Abl protein levels through the induction of autophagy may be an additional and important feature of the activity of imatinib.

  3. Stat5 Exerts Distinct, Vital Functions in the Cytoplasm and Nucleus of Bcr-Abl{sup +} K562 and Jak2(V617F){sup +} HEL Leukemia Cells

    Energy Technology Data Exchange (ETDEWEB)

    Weber, Axel [Georg-Speyer-Haus, Institute for Tumor Biology and Experimental Therapy, Frankfurt am Main 60596 (Germany); Borghouts, Corina [Ganymed Pharmaceuticals AG, Mainz 55131 (Germany); Brendel, Christian [Boston Children’s Hospital, Division of Hematology/Oncology, Boston, MA 02115 (United States); Moriggl, Richard [Ludwig Boltzmann Institute for Cancer Research (LBI-CR), Vienna 1090 (Austria); Delis, Natalia; Brill, Boris; Vafaizadeh, Vida; Groner, Bernd, E-mail: Groner@em.uni-frankfurt.de [Georg-Speyer-Haus, Institute for Tumor Biology and Experimental Therapy, Frankfurt am Main 60596 (Germany)

    2015-03-19

    Signal transducers and activators of transcription (Stats) play central roles in the conversion of extracellular signals, e.g., cytokines, hormones and growth factors, into tissue and cell type specific gene expression patterns. In normal cells, their signaling potential is strictly limited in extent and duration. The persistent activation of Stat3 or Stat5 is found in many human tumor cells and contributes to their growth and survival. Stat5 activation plays a pivotal role in nearly all hematological malignancies and occurs downstream of oncogenic kinases, e.g., Bcr-Abl in chronic myeloid leukemias (CML) and Jak2(V617F) in other myeloproliferative diseases (MPD). We defined the mechanisms through which Stat5 affects growth and survival of K562 cells, representative of Bcr-Abl positive CML, and HEL cells, representative for Jak2(V617F) positive acute erythroid leukemia. In our experiments we suppressed the protein expression levels of Stat5a and Stat5b through shRNA mediated downregulation and demonstrated the dependence of cell survival on the presence of Stat5. Alternatively, we interfered with the functional capacities of the Stat5 protein through the interaction with a Stat5 specific peptide ligand. This ligand is a Stat5 specific peptide aptamer construct which comprises a 12mer peptide integrated into a modified thioredoxin scaffold, S5-DBD-PA. The peptide sequence specifically recognizes the DNA binding domain (DBD) of Stat5. Complex formation of S5-DBD-PA with Stat5 causes a strong reduction of P-Stat5 in the nuclear fraction of Bcr-Abl-transformed K562 cells and a suppression of Stat5 target genes. Distinct Stat5 mediated survival mechanisms were detected in K562 and Jak2(V617F)-transformed HEL cells. Stat5 is activated in the nuclear and cytosolic compartments of K562 cells and the S5-DBD-PA inhibitor most likely affects the viability of Bcr-Abl{sup +} K562 cells through the inhibition of canonical Stat5 induced target gene transcription. In HEL cells

  4. High risk acute lymphoblastic leukemia cells with bcr-abl and INK4A/ARF mutations retain susceptibility to alloreactive T cells

    OpenAIRE

    Young, Faith M.; Campbell, Andrew; Emo, Kris Lambert; Jansson, Johan; Wang, Pin-Yi; Jordan, Craig T.; Mullen, Craig A.

    2008-01-01

    INK4A/ARF mutations are acquired in bcr/abl+ lymphoid blast phase chronic myelogenous leukemia (CML) and bcr/abl+ acute lymphoblastic leukemia (ALL). Donor lymphocyte infusion and graft versus leukemia are generally ineffective in such ALL’s, while GVL is highly active against bcr/abl+ CML that does not have a lesion in the INK4A/ARF locus. The mechanisms for the ineffectiveness of GVL are not fully known and it is possible that intrinsic resistance of acute lymphoid leukemias to immune effec...

  5. BCR-ABL fusion genes are inducible by X-irradiation in vitro

    International Nuclear Information System (INIS)

    The Philadelphia chromosome consists of a reciprocal translocation between the ABL oncogene at chromosome 9q34 and the BCR gene at chromosome 22q resulting in the expression of chimeric BCR-ABL mRNAs specific to chronic myelogenous leukemia (CML). The presence of the fusion genes can be detected with high specificity and sensitivity by means of reverse transcription and polymerase chain reaction. Using this assay, it was possible to detect BCR-ABL fusion genes induced among HL60 cells after 100 Gy of X-irradiation in vitro. A total of five fusion gene transcripts were obtained. These fusion genes contained not only CML-specific BCR-ABL rearrangements, but also other forms of BCR-ABL fusions. These latter genes had junctions of BCR exon 4/ABL exon 2 intervened by a segment of DNA of unknown origin, BCR exon 5/ABL exon 2, and BCR exon 4/ABL exon 2. The results appear to be the first evidence for the induction of the BCR-ABL fusion gene by X-irradiation. In terms of leukemogenesis, it is suggested that only those cells bearing certain CML-related BCR-ABL fusion genes are positively selected by virtue of a growth advantage in vivo. (author)

  6. Nilotinib (formerly AMN107), a highly selective BCR-ABL tyrosine kinase inhibitor, is effective in patients with Philadelphia chromosome-positive chronic myelogenous leukemia in chronic phase following imatinib resistance and intolerance.

    Science.gov (United States)

    Kantarjian, Hagop M; Giles, Francis; Gattermann, Norbert; Bhalla, Kapil; Alimena, Giuliana; Palandri, Francesca; Ossenkoppele, Gert J; Nicolini, Franck-Emmanuel; O'Brien, Stephen G; Litzow, Mark; Bhatia, Ravi; Cervantes, Francisco; Haque, Ariful; Shou, Yaping; Resta, Debra J; Weitzman, Aaron; Hochhaus, Andreas; le Coutre, Philipp

    2007-11-15

    Nilotinib, an orally bioavailable, selective Bcr-Abl tyrosine kinase inhibitor, is 30-fold more potent than imatinib in pre-clinical models, and overcomes most imatinib resistant BCR-ABL mutations. In this phase 2 open-label study, 400 mg nilotinib was administered orally twice daily to 280 patients with Philadelphia chromosome-positive (Ph(+)) chronic myeloid leukemia in chronic phase (CML-CP) after imatinib failure or intolerance. Patients had at least 6 months of follow-up and were evaluated for hematologic and cytogenetic responses, as well as for safety and overall survival. At 6 months, the rate of major cytogenetic response (Ph < or = 35%) was 48%: complete (Ph = 0%) in 31%, and partial (Ph = 1%-35%) in 16%. The estimated survival at 12 months was 95%. Nilotinib was effective in patients harboring BCR-ABL mutations associated with imatinib resistance (except T315I), and also in patients with a resistance mechanism independent of BCR-ABL mutations. Adverse events were mostly mild to moderate, and there was minimal cross-intolerance with imatinib. Grades 3 to 4 neutropenia and thrombocytopenia were observed in 29% of patients; pleural or pericardial effusions were observed in 1% (none were severe). In summary, nilotinib is highly active and safe in patients with CML-CP after imatinib failure or intolerance. This clinical trial is registered at http://clinicaltrials.gov as ID no. NCT00109707. PMID:17715389

  7. SGX393 inhibits the CML mutant Bcr-Abl[superscript T315I] and preempts in vitro resistance when combined with nilotinib or dasatinib

    Energy Technology Data Exchange (ETDEWEB)

    O' Hare, Thomas; Eide, Christopher A.; Tyner, Jeffrey W.; Corbin, Amie S.; Wong, Matthew J.; Buchanan, Sean; Holme, Kevin; Jessen, Katayoun A.; Tang, Crystal; Lewis, Hal A.; Romero, Richard D.; Burley, Stephen K.; Deininger, Michael W. (OHSU- Cancer Instit.); (SGX)

    2010-01-12

    Imatinib inhibits Bcr-Abl, the oncogenic tyrosine kinase that causes chronic myeloid leukemia. The second-line inhibitors nilotinib and dasatinib are effective in patients with imatinib resistance resulting from Bcr-Abl kinase domain mutations. Bcr-Abl{sup T315I}, however, is resistant to all Abl kinase inhibitors in clinical use and is emerging as the most frequent cause of salvage therapy failure. SGX393 is a potent inhibitor of native and T315I-mutant Bcr-Abl kinase that blocks the growth of leukemia cell lines and primary hematopoietic cells expressing Bcr-Abl{sup T315I}, with minimal toxicity against Bcr-Abl-negative cell lines or normal bone marrow. A screen for Bcr-Abl mutants emerging in the presence of SGX393 revealed concentration-dependent reduction in the number and range of mutations. Combining SGX393 with nilotinib or dasatinib preempted emergence of resistant subclones, including Bcr-Abl{sup T315I}. These findings suggest that combination of a T315I inhibitor with the current clinically used inhibitors may be useful for reduction of Bcr-Abl mutants in Philadelphia chromosome-positive leukemia.

  8. Association of HLA Class I and Class II genes with bcr-abl transcripts in leukemia patients with t(9;22 (q34;q11

    Directory of Open Access Journals (Sweden)

    Cano Pedro

    2004-06-01

    Full Text Available Abstract Background Based on the site of breakpoint in t(9;22 (q34;q11, bcr-abl fusion in leukemia patients is associated with different types of transcript proteins. In this study we have seen the association of HLA genes with different types of bcr-abl transcripts. The association could predict the bcr-abl peptide presentation by particular HLA molecules. Methods The study included a total of 189 patients of mixed ethnicity with chronic myelogenous leukemia and acute lymphocytic leukemia who were being considered for bone marrow transplantation. Typing of bcr-abl transcripts was done by reverse transcriptase PCR method. HLA typing was performed by molecular methods. The bcr-abl and HLA association was studied by calculating the relative risks and chi-square test. Results Significant negative associations (p Conclusions The negative associations of a particular bcr-abl transcript with specific HLA alleles suggests that these alleles play a critical role in presenting peptides derived from the chimeric proteins and eliciting a successful T-cell cytotoxic response. Knowledge of differential associations between HLA phenotypes and bcr-abl fusion transcript types would help in developing better strategies for immunization with the bcr-abl peptides against t(9;22 (q34;q11-positive leukemia.

  9. Nilotinib treatment in mouse models of P190 Bcr/Abl lymphoblastic leukemia

    Directory of Open Access Journals (Sweden)

    Groffen John

    2007-10-01

    Full Text Available Abstract Background Ph-positive leukemias are caused by the aberrant fusion of the BCR and ABL genes. Nilotinib is a selective Bcr/Abl tyrosine kinase inhibitor related to imatinib, which is widely used to treat chronic myelogenous leukemia. Because Ph-positive acute lymphoblastic leukemia only responds transiently to imatinib therapy, we have used mouse models to test the efficacy of nilotinib against lymphoblastic leukemia caused by the P190 form of Bcr/Abl. Results After transplant of 10,000 highly malignant leukemic cells into compatible recipients, untreated mice succumbed to leukemia within 21 days, whereas mice treated with 75 mg/kg nilotinib survived significantly longer. We examined cells from mice that developed leukemia while under treatment for Bcr/Abl kinase domain point mutations but these were not detected. In addition, culture of such cells ex vivo showed that they were as sensitive as the parental cell line to nilotinib but that the presence of stromal support allowed resistant cells to grow out. Nilotinib also exhibited impressive anti-leukemia activity in P190 Bcr/Abl transgenic mice that had developed overt leukemia/lymphoma masses and that otherwise would have been expected to die within 7 days. Visible lymphoma masses disappeared within six days of treatment and leukemic cell numbers in peripheral blood were significantly reduced. Treated mice survived more than 30 days. Conclusion These results show that nilotinib has very impressive anti-leukemia activity but that lymphoblastic leukemia cells can become unresponsive to it both in vitro and in vivo through mechanisms that appear to be Bcr/Abl independent.

  10. MPT0B169, a New Antitubulin Agent, Inhibits Bcr-Abl Expression and Induces Mitochondrion-Mediated Apoptosis in Nonresistant and Imatinib-Resistant Chronic Myeloid Leukemia Cells.

    Directory of Open Access Journals (Sweden)

    Shuit-Mun Wong

    Full Text Available Chronic myeloid leukemia (CML is a clonal disorder of hematopoietic stem/progenitor cells that is caused by the Bcr-Abl oncoprotein. Clinical resistance to the Bcr-Abl inhibitor imatinib is a critical problem in treating CML. This study investigated the antitumor effect and mechanism of MPT0B169, a new antitubulin agent, in K562 CML cells and their derived imatinib-resistant cells, IMR2 and IMR3. IMR2 and IMR3 cells showed complete resistance to imatinib-induced growth inhibition and apoptosis. Resistance involved ERK1/2 overactivation and MDR1 overexpression. MPT0B169 inhibited the growth of K562, IMR2, and IMR3 cells in a dose- and time-dependent manner. MPT0B169 substantially inhibited the mRNA and protein levels of Bcr-Abl, followed by its downstream pathways including Akt, ERK1/2, and STAT3 in these cells. MPT0B169 treatment resulted in a decrease in the polymer form of tubulin according to Western blot analysis. It triggered cell cycle arrest at the G2/M phase before apoptosis, which was related to the upregulation of the mitotic marker MPM2 and the cyclin B1 level, and a change in the phosphorylation of Cdk1. MPT0B169 induced apoptosis in nonresistant and imatinib-resistant cells via a mitochondrion-mediated caspase pathway. Further study showed that the agent led to a decrease in the antiapoptotic proteins Bcl-2, Bcl-xL, and Mcl-1 and an increase in the apoptotic protein Bax. Taken together, our results suggest that MPT0B169 might be a promising agent for overcoming imatinib resistance in CML cells.

  11. Association of HLA antigens and BCR-ABL transcripts in leukemia patients with the Philadelphia chromosome

    Directory of Open Access Journals (Sweden)

    Daiana Landenberger de Carvalho

    2012-01-01

    Full Text Available OBJECTIVE: This study aimed to verify the association between human leukocyte antigens and the bcr-abl fusion protein resulting from t(9;22(q34;q11 in chronic leukemia myeloid and acute lymphoblastic leukemia patients. METHODS: Forty-seven bcr-abl positive individuals were evaluated. Typing was performed bymicrolymphocytotoxicity and molecular biological methods (human leukocyte antigens Class I and Class II. A control group was obtained from the data of potential bone marrow donors registered in the Brazilian Bone Marrow Donor Registry (REDOME. RESULTS: Positive associations with HLA-A25 and HLA-B18 were found for the b2a2 transcript, as well as a tendency towards a positive association with HLA-B40 and a negative association with HLA-A68. The b3a2 transcript showed positive associations with HLA-B40 and HLA-DRB1*3. CONCLUSION: The negative association between human leukocyte antigens and the BCR-ABL transcript suggests that binding and presentation of peptides derived from the chimeric protein are effective to increase a cytotoxic T lymphocyte response appropriate for the destruction of leukemic cells.

  12. Association of HLA Class I and Class II genes with bcr-abl transcripts in leukemia patients with t(9;22) (q34;q11)

    International Nuclear Information System (INIS)

    Based on the site of breakpoint in t(9;22) (q34;q11), bcr-abl fusion in leukemia patients is associated with different types of transcript proteins. In this study we have seen the association of HLA genes with different types of bcr-abl transcripts. The association could predict the bcr-abl peptide presentation by particular HLA molecules. The study included a total of 189 patients of mixed ethnicity with chronic myelogenous leukemia and acute lymphocytic leukemia who were being considered for bone marrow transplantation. Typing of bcr-abl transcripts was done by reverse transcriptase PCR method. HLA typing was performed by molecular methods. The bcr-abl and HLA association was studied by calculating the relative risks and chi-square test. Significant negative associations (p < 0.05) were observed with HLA-A*02 (b2a2, e1a2), -A*68 (b2a2, b3a2, e1a2), -B*14 (b2a2, b3a2, e1a2), -B*15 (b2a2, b3a2), -B*40 (b2a2), -DQB1*0303 (b2a2, b3a2), -DQB1*0603 (b2a2), -DRB1*0401 (e1a2), -DRB1*0701 (b3a2), and -DRB1*1101 (b2a2). The negative associations of a particular bcr-abl transcript with specific HLA alleles suggests that these alleles play a critical role in presenting peptides derived from the chimeric proteins and eliciting a successful T-cell cytotoxic response. Knowledge of differential associations between HLA phenotypes and bcr-abl fusion transcript types would help in developing better strategies for immunization with the bcr-abl peptides against t(9;22) (q34;q11)-positive leukemia

  13. WT1-mediated repression of the proapoptotic transcription factor ZNF224 is triggered by the BCR-ABL oncogene

    Science.gov (United States)

    Montano, Giorgia; Vidovic, Karina; Palladino, Chiara; Cesaro, Elena; Sodaro, Gaetano; Quintarelli, Concetta; De Angelis, Biagio; Errichiello, Santa; Pane, Fabrizio; Izzo, Paola; Grosso, Michela; Gullberg, Urban; Costanzo, Paola

    2015-01-01

    The Kruppel-like protein ZNF224 is a co-factor of the Wilms’ tumor 1 protein, WT1. We have previously shown that ZNF224 exerts a specific proapoptotic role in chronic myelogenous leukemia (CML) K562 cells and contributes to cytosine arabinoside-induced apoptosis, by modulating WT1-dependent transcription of apoptotic genes. Here we demonstrate that ZNF224 gene expression is down-regulated both in BCR-ABL positive cell lines and in primary CML samples and is restored after imatinib and second generation tyrosine kinase inhibitors treatment. We also show that WT1, whose expression is positively regulated by BCR-ABL, represses transcription of the ZNF224 gene. Finally, we report that ZNF224 is significantly down-regulated in patients with BCR-ABL positive chronic phase-CML showing poor response or resistance to imatinib treatment as compared to high-responder patients. Taken as a whole, our data disclose a novel pathway activated by BCR-ABL that leads to inhibition of apoptosis through the ZNF224 repression. ZNF224 could thus represent a novel promising therapeutic target in CML. PMID:26320177

  14. Essential role for telomerase in chronic myeloid leukemia induced by BCR-ABL in mice

    OpenAIRE

    Vicente-Dueñas, Carolina; Barajas-Diego, Marcos; Romero-Camarero, Isabel; González-Herrero, Inés; Flores, Teresa; Sánchez García, Isidro

    2012-01-01

    The telomerase protein is constitutively activated in malignant cells from many patients with cancer, including the chronic myeloid leukemia (CML), but whether telomerase is essential for the pathogenesis of this disease is not known. Here, we used telomerase deficient mice to determine the requirement for telomerase in CML induced by BCR-ABL in mouse models of CML. Loss of one telomerase allele or complete deletion of telomerase prevented the development of leukemia induced by BCR-ABL. Howev...

  15. Accumulation of FGF-2 in CD34+ Bcr-Abl+ hematopoietic progenitor cells from chronic myeloid leukemia patients: relationship with malignant phenotype

    Czech Academy of Sciences Publication Activity Database

    Dvořák, Petr; Faitová, Jitka; Dvořáková, D.; Hampl, Aleš; Mayer, J.

    Steamboat Springs, 2003. s. -. [From Stem Cells to Therapy (Keystone Symposia). 29.03.2003-03.04.2003, Steamboat Springs] R&D Projects: GA MŠk LN00A065 Institutional research plan: CEZ:AV0Z5039906; CEZ:MSM 432100001 Keywords : leukemia Subject RIV: FH - Neurology

  16. Increased expression of fibroblast growth factor receptor 3 in CD34+ BCR-ABL+ cells from patients with chronic myeloid leukemia

    Czech Academy of Sciences Publication Activity Database

    Dvořák, Petr; Dvořáková, D.; Doubek, M.; Faitová, Jitka; Pacholíková, J.; Hampl, Aleš; Mayer, J.

    2003-01-01

    Roč. 17, - (2003), s. 1-8. ISSN 0887-6924 R&D Projects: GA ČR GA301/03/1122; GA MŠk LN00A065 Institutional research plan: CEZ:AV0Z5039906; CEZ:MSM 00065269705; CEZ:VZ432100001 Keywords : CD34+cells, imatinib Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.116, year: 2003

  17. A Requirement for SOCS-1 and SOCS-3 Phosphorylation in Bcr-Abl-Induced Tumorigenesis

    Directory of Open Access Journals (Sweden)

    Xiaoxue Qiu

    2012-06-01

    Full Text Available Suppressors of cytokine signaling 1 and 3 (SOCS-1 and SOCS-3 are inhibitors of the Janus tyrosine kinase (JAK/signal transducers and activators of transcription (STAT pathway and function in a negative feedback loop during cytokine signaling. Abl transformation is associated with constitutive activation of JAK/STAT-dependent signaling. However, the mechanism by which Abl oncoproteins bypass SOCS inhibitory regulation remains poorly defined. Here, we demonstrate that coexpression of Bcr-Abl with SOCS-1 or SOCS-3 results in tyrosine phosphorylation of these SOCS proteins. Interestingly, SOCS-1 is highly tyrosine phosphorylated in one of five primary chronic myelogenous leukemia samples. Bcr-Abl-dependent tyrosine phosphorylation of SOCS-1 and SOCS-3 occurs mainly on Tyr 155 and Tyr 204 residues of SOCS-1 and on Tyr 221 residue of SOCS-3. We observed that phosphorylation of these SOCS proteins was associated with their binding to Bcr-Abl. Bcr-Abl-dependent phosphorylation of SOCS-1 and SOCS-3 diminished their inhibitory effects on the activation of JAK and STAT5 and thereby enhanced JAK/STAT5 signaling. Strikingly, disrupting the tyrosine phosphorylation of SOCS-1 or SOCS-3 impaired the expression of Bcl-XL protein and sensitized K562 leukemic cells to undergo apoptosis. Moreover, selective mutation of tyrosine phosphorylation sites of SOCS-1 or SOCS-3 significantly blocked Bcr-Abl-mediated tumorigenesis in nude mice and inhibited Bcr-Abl-mediated murine bone marrow transformation. Together, these results reveal a mechanism of how Bcr-Abl may overcome SOCS-1 and SOCS-3 inhibition to constitutively activate the JAK/STAT-dependent signaling, and suggest that Bcr-Abl may critically requires tyrosine phosphorylation of SOCS-1 and SOCS-3 to mediate tumorigenesis when these SOCS proteins are present in cells.

  18. Molecular Detection of BCR-ABL in Chronic Myeloid Leukemia.

    Science.gov (United States)

    Qin, Ya-Zhen; Huang, Xiao-Jun

    2016-01-01

    All chronic myeloid leukemia (CML) patients have the BCR-ABL fusion gene. The constitutively activated BCR-ABL tyrosine kinase is a critical pathogenetic event in CML. Tyrosine kinase inhibitors (TKIs), such as imatinib, are synthesized small molecules that primarily target BCR-ABL tyrosine kinases and have become a first-line treatment for CML. Detection of BCR-ABL transcript level by real-time quantitative polymerase chain reaction (RQ-PCR) is a clinical routine for evaluating TKI treatment efficacy and predicting long-term response. Furthermore, because they are a main TKI resistance mechanism, the BCR-ABL tyrosine kinase domain (TKD) point mutations that are detected by Sanger sequencing can help clinicians make decisions on subsequent treatment selections. Here, we present protocols for the two abovementioned molecular methods for CML analysis. PMID:27581134

  19. Expression of p89c-Mybex9b, an alternatively spliced form of c-Myb, is required for proliferation and survival of p210BCR/ABL-expressing cells

    International Nuclear Information System (INIS)

    The c-Myb gene encodes the p75c-Myb isoform and less-abundant proteins generated by alternatively spliced transcripts. Among these, the best known is pc-Mybex9b, which contains 121 additional amino acids between exon 9 and 10, in a domain involved in protein–protein interactions and negative regulation. In hematopoietic cells, expression of pc-Mybex9b accounts for 10–15% of total c-Myb; these levels may be biologically relevant because modest changes in c-Myb expression affects proliferation and survival of leukemic cells and lineage choice and frequency of normal hematopoietic progenitors. In this study, we assessed biochemical activities of pc-Mybex9b and the consequences of perturbing its expression in K562 and primary chronic myeloid leukemia (CML) progenitor cells. Compared with p75c-Myb, pc-Mybex9b is more stable and more effective in transactivating Myb-regulated promoters. Ectopic expression of pc-Mybex9b enhanced proliferation and colony formation and reduced imatinib (IM) sensitivity of K562 cells; conversely, specific downregulation of pc-Mybex9b reduced proliferation and colony formation, enhanced IM sensitivity of K562 cells and markedly suppressed colony formation of CML CD34+ cells, without affecting the levels of p75c-Myb. Together, these studies indicate that expression of the low-abundance pc-Mybex9b isoform has an important role for the overall biological effects of c-Myb in BCR/ABL-transformed cells

  20. BCR-ABL Promotes PTEN Downregulation in Chronic Myeloid Leukemia

    OpenAIRE

    Cristina Panuzzo; Sabrina Crivellaro; Giovanna Carrà; Angelo Guerrasio; Giuseppe Saglio; Alessandro Morotti

    2014-01-01

    Chronic myeloid leukemia (CML) is a myeloproliferative disorder characterized by the t(9;22) translocation coding for the chimeric protein p210 BCR-ABL. The tumor suppressor PTEN plays a critical role in the pathogenesis of CML chronic phase, through non genomic loss of function mechanisms, such as protein down-regulation and impaired nuclear/cytoplasmic shuttling. Here we demonstrate that BCR-ABL promotes PTEN downregulation through a MEK dependent pathway. Furthermore, we describe a novel n...

  1. Targeting the SH2-Kinase Interface in Bcr-Abl Inhibits Leukemogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Grebien, Florian; Hantschel, Oliver; Wojcik, John; Kaupe, Ines; Kovacic, Boris; Wyrzucki, Arkadiusz M.; Gish, Gerald D.; Cerny-Reiterer, Sabine; Koide, Akiko; Beug, Hartmut; Pawson, Tony; Valent, Peter; Koide, Shohei; Superti-Furga, Giulio (AAS); (Mount Sinai Hospital); (Med U. Vienna); (UC); (IMP-CNRS)

    2012-10-25

    Chronic myelogenous leukemia (CML) is caused by the constitutively active tyrosine kinase Bcr-Abl and treated with the tyrosine kinase inhibitor (TKI) imatinib. However, emerging TKI resistance prevents complete cure. Therefore, alternative strategies targeting regulatory modules of Bcr-Abl in addition to the kinase active site are strongly desirable. Here, we show that an intramolecular interaction between the SH2 and kinase domains in Bcr-Abl is both necessary and sufficient for high catalytic activity of the enzyme. Disruption of this interface led to inhibition of downstream events critical for CML signaling and, importantly, completely abolished leukemia formation in mice. Furthermore, disruption of the SH2-kinase interface increased sensitivity of imatinib-resistant Bcr-Abl mutants to TKI inhibition. An engineered Abl SH2-binding fibronectin type III monobody inhibited Bcr-Abl kinase activity both in vitro and in primary CML cells, where it induced apoptosis. This work validates the SH2-kinase interface as an allosteric target for therapeutic intervention.

  2. p210 Bcr-Abl confers overexpression of inosine monophosphate dehydrogenase : an intrinsic pathway to drug resistance mediated by oncogene.

    Energy Technology Data Exchange (ETDEWEB)

    Gharehbaghi, K.; Burgess, G. S.; Collart, F. R.; Litz-Jackson, S.; Huberman, E.; Jayaram, H. N.; Boswell, H. S.; Center for Mechanistic Biology and Biotechnology; Lab. for Experimental Oncology; Indiana Univ. School of Medicine

    1994-01-01

    The p210 bcr-abl fusion protein tyrosine kinase oncogene has been implicated in the pathogenesis of chronic granulocytic leukemia (CGL). Specific intracellular functions performed by p210 bcr-abl have recently been delineated. We considered the possibility that p210 bcr-abl may also regulate the abundance of inosine 5'-monophosphate dehydrogenase (IMPDH) which is a rate-limiting enzyme for de novo guanylate synthesis. We performed studies of the inhibition of IMPDH by tiazofurin, which acts as a competitive inhibitor through its active species that mimics nicotinamide adenine dinucleotide (NAD), i.e. thiazole-4-carboxamide adenine dinucleotide (TAD). The mean inhibitory concentration (IC50) of tiazofurin for cellular proliferation inhibition was 2.3-2.8-fold greater in cells expressing p210 bcr-abl than in their corresponding parent cells proliferating under the influence of growth factors or in growth factor-independent derivative cells not expressing detectable p210 bcr-abl. IMPDH activity was 1.5-2.3-fold greater within cells expressing p210 bcr-abl than in their parent cells. This increase in enzyme activity was a result of 2-fold increased IMPDH protein as determined by immunoblotting. In addition, an increase in the Km value for NAD utilization by IMPDH was observed in p210 bcr-abl transformed cells, but this increase was within the range of resident NAD concentrations observed in the cells. Increased IMPDH protein in p210 bcr-abl transformed cells was traced to an increased level of IMP dehydrogenase II messenger RNA. Thus, regulation of IMPDH gene expression is mediated at least in part by the bcr-abl gene product and may therefore be indicative of a specific mechanism of intrinsic resistance to tiazofurin.

  3. p210 bcr-abl confers overexpression of inosine monophosphate dehydrogenase: an intrinsic pathway to drug resistance mediated by oncogene.

    Science.gov (United States)

    Gharehbaghi, K; Burgess, G S; Collart, F R; Litz-Jackson, S; Huberman, E; Jayaram, H N; Boswell, H S

    1994-08-01

    The p210 bcr-abl fusion protein tyrosine kinase oncogene has been implicated in the pathogenesis of chronic granulocytic leukemia (CGL). Specific intracellular functions performed by p210 bcr-abl have recently been delineated. We considered the possibility that p210 bcr-abl may also regulate the abundance of inosine 5'-monophosphate dehydrogenase (IMPDH) which is a rate-limiting enzyme for de novo guanylate synthesis. We performed studies of the inhibition of IMPDH by tiazofurin, which acts as a competitive inhibitor through its active species that mimics nicotinamide adenine dinucleotide (NAD), i.e. thiazole-4-carboxamide adenine dinucleotide (TAD). The mean inhibitory concentration (IC50) of tiazofurin for cellular proliferation inhibition was 2.3-2.8-fold greater in cells expressing p210 bcr-abl than in their corresponding parent cells proliferating under the influence of growth factors or in growth factor-independent derivative cells not expressing detectable p210 bcr-abl. IMPDH activity was 1.5-2.3-fold greater within cells expressing p210 bcr-abl than in their parent cells. This increase in enzyme activity was a result of 2-fold increased IMPDH protein as determined by immunoblotting. In addition, an increase in the Km value for NAD utilization by IMPDH was observed in p210 bcr-abl transformed cells, but this increase was within the range of resident NAD concentrations observed in the cells. Increased IMPDH protein in p210 bcr-abl transformed cells was traced to an increased level of IMP dehydrogenase II messenger RNA. Thus, regulation of IMPDH gene expression is mediated at least in part by the bcr-abl gene product and may therefore be indicative of a specific mechanism of intrinsic resistance to tiazofurin. PMID:7520100

  4. A BCR/ABL-hIL-2 DNA Vaccine Enhances the Immune Responses in BALB/c Mice

    Directory of Open Access Journals (Sweden)

    Yanan Qin

    2013-01-01

    Full Text Available The use of a DNA vaccine encoding the BCR/ABL fusion gene is thought to be a promising approach for patients with chronic myeloid leukemia (CML to eradicate minimal residual disease after treatment with chemotherapy or targeted therapy. In this study, our strategy employs genetic technology to create a DNA vaccine encoding the BCR/ABL fusion and human interleukin-2 (hIL-2 genes. The successfully constructed plasmids BCR/ABL-pIRES-hIL-2, BCR/ABL-pIRES, and pIRES-hIL-2 were delivered intramuscularly to BALB/c mice at 14-day intervals for three cycles. The transcription and expression of the BCR/ABL and hIL-2 genes were found in the injected muscle tissues. The interferon-γ (IFN-γ serum levels were increased, and the splenic CD4+/CD8+ T cell ratio was significantly decreased in the BCR/ABL-pIRES-hIL-2-injected mice. Furthermore, specific antibodies against K562 cells could be detected by indirect immunofluorescence. These results indicate that a DNA vaccine containing BCR/ABL and hIL-2 together may elicit increased in vivo humoral and cellular immune responses in BALB/c mice.

  5. Harmonization of BCR-ABL mRNA quantification using a uniform multifunctional control plasmid in 37 international laboratories

    DEFF Research Database (Denmark)

    Müller, M C; Erben, P; Saglio, G; Gottardi, E; Nyvold, Charlotte Guldborg; Schenk, T; Ernst, T; Lauber, S; Kruth, J; Hehlmann, R; Hochhaus, A

    2008-01-01

    Individualized PCR strategies hamper comparability of molecular results between different laboratories in several fields of medicine. To harmonize BCR-ABL mRNA quantification an international multicenter trial involving 37 laboratories in 14 countries was initiated using 10 samples, each containing...... various dilutions (10, 2, 1 and 0.1%) of b3a2 or b2a2 BCR-ABL positive in normal leukocytes and negative controls. A novel control plasmid (pME-2) was designed for external calibration containing BCR-ABL and glucuronidase-beta (GUS) sequences. Median BCR-ABL/ABL ratios were 9.1, 1.8, 0.85 and 0.11% in b3a...... participating laboratories (13%) detected low BCR-ABL copy numbers in negative control samples; one laboratory failed to detect BCR-ABL in a low-level sample. We conclude that the use of a common control plasmid does indeed improve comparability of BCR-ABL mRNA quantification results. However, further...

  6. BCR-ABL DERIVED PEPTIDE VACCINES FOR CHRONIC MYELOID LEUKAEMIA

    Directory of Open Access Journals (Sweden)

    M. Bocchia

    2012-01-01

    Full Text Available Chronic Myeloid Leukemia (CML is a myeloproliferative pluripotent stem cell disorder characterized by the presence of a cytogenetic hallmark, the Philadelphia (Ph chromosome, and accounts for 15% of adult leukemias. The disease progresses from a chronic phase through an accelerated phase to a blast phase and its natural course accounts for a median 4 years survival1. The Ph chromosome is derived by a reciprocal translocation termed t(9;22 in which the c-abl oncogene has moved from chromosome 9 into the breakpoint cluster region (bcr, within the bcr gene on chromosome 22, resulting in a chimeric bcr-abl fusion gene that encodes a 210 KD protein (p210 with constitutive tyrosine kinase activity. Two major alternative chimeric p210 can result from this fusion gene: p210-b2a2 where the junction occurs between bcr exon 2 (b2 and abl exon 2 (a2 and p210-b3a2 where the the junction occurs between bcr exon 3 (b3 and abl exon 2 (a2. About 40% of CML patients harbor the p210-b2a2 and about 60% of them show the p210-b3a2.

  7. The human leukemia oncogene bcr-abl abrogates the anchorage requirement but not the growth factor requirement for proliferation.

    OpenAIRE

    Renshaw, M W; McWhirter, J R; Wang, J Y

    1995-01-01

    Proliferation of normal cells in a multicellular organism requires not only growth factors but also the proper attachment to the extracellular matrix. A hallmark of neoplastic transformation is the loss of anchorage dependence which usually accompanies the loss of growth factor requirement. The Bcr-Abl tyrosine kinase of human leukemias is shown here to abrogate only the anchorage, not the growth factor, requirement. Bcr-Abl-transformed cells grow in soft agar but do not proliferate in serum-...

  8. AP24534, a Pan-BCR-ABL Inhibitor for Chronic Myeloid Leukemia, Potently Inhibits the T315I Mutant and Overcomes Mutation-Based Resistance

    Energy Technology Data Exchange (ETDEWEB)

    O’Hare, Thomas; Shakespeare, William C.; Zhu, Xiaotian; Eide, Christopher A.; Rivera, Victor M.; Wang, Frank; Adrian, Lauren T.; Zhou, Tianjun; Huang, Wei-Sheng; Xu, Qihong; Metcalf, III, Chester A.; Tyner, Jeffrey W.; Loriaux, Marc M.; Corbin, Amie S.; Wardwell, Scott; Ning, Yaoyu; Keats, Jeffrey A.; Wang, Yihan; Sundaramoorthi, Raji; Thomas, Mathew; Zhou, Dong; Snodgrass, Joseph; Commodore, Lois; Sawyer, Tomi K.; Dalgarno, David C.; Deininger, Michael W.N.; Druker, Brian J.; Clackson, Tim; (OHSU- Cancer Instit.); (ARIAD)

    2010-09-07

    Inhibition of BCR-ABL by imatinib induces durable responses in many patients with chronic myeloid leukemia (CML), but resistance attributable to kinase domain mutations can lead to relapse and a switch to second-line therapy with nilotinib or dasatinib. Despite three approved therapeutic options, the cross-resistant BCR-ABL{sup T315I} mutation and compound mutants selected on sequential inhibitor therapy remain major clinical challenges. We report design and preclinical evaluation of AP24534, a potent, orally available multitargeted kinase inhibitor active against T315I and other BCR-ABL mutants. AP24534 inhibited all tested BCR-ABL mutants in cellular and biochemical assays, suppressed BCR-ABL{sup T315I}-driven tumor growth in mice, and completely abrogated resistance in cell-based mutagenesis screens. Our work supports clinical evaluation of AP24534 as a pan-BCR-ABL inhibitor for treatment of CML.

  9. Frequency of BCR-ABL Transcript Types in Syrian CML Patients

    Science.gov (United States)

    Farhat-Maghribi, Sulaf; Habbal, Wafa; Monem, Fawza

    2016-01-01

    Background. In Syria, CML patients are started on tyrosine kinase inhibitors (TKIs) and monitored until complete molecular response is achieved. BCR-ABL mRNA transcript type is not routinely identified, contrary to the recommendations. In this study we aimed to identify the frequency of different BCR-ABL transcripts in Syrian CML patients and highlight their significance on monitoring and treatment protocols. Methods. CML patients positive for BCR-ABL transcripts by quantitative RT-PCR were enrolled. BCR-ABL transcript types were investigated using a home-made PCR method that was adapted from published protocols and optimized. The transcript types were then confirmed using a commercially available research kit. Results. Twenty-four transcripts were found in 21 patients. The most common was b2a2, followed by b3a2, b3a3, and e1a3 present solely in 12 (57.1%), 3 (14.3%), 2 (9.5%), and 1 (4.8%), respectively. Three samples (14.3%) contained dual transcripts. While b3a2 transcript was apparently associated with warning molecular response to imatinib treatment, b2a2, b3a3, and e1a3 transcripts collectively proved otherwise (P = 0.047). Conclusion. It might be advisable to identify the BCR-ABL transcript type in CML patients at diagnosis, using an empirically verified method, in order to link the detected transcript with the clinical findings, possible resistance to treatment, and appropriate monitoring methods. PMID:27313614

  10. A critical role of CDKN3 in Bcr-Abl-mediated tumorigenesis.

    Directory of Open Access Journals (Sweden)

    Qinghuang Chen

    Full Text Available CDKN3 (cyclin-dependent kinase inhibitor 3, a dual specificity protein phosphatase, dephosphorylates cyclin-dependent kinases (CDKs and thus functions as a key negative regulator of cell cycle progression. Deregulation or mutations of CDNK3 have been implicated in various cancers. However, the role of CDKN3 in Bcr-Abl-mediated chronic myelogenous leukemia (CML remains unknown. Here we found that CDKN3 acts as a tumor suppressor in Bcr-Abl-mediated leukemogenesis. Overexpression of CDKN3 sensitized the K562 leukemic cells to imanitib-induced apoptosis and dramatically inhibited K562 xenografted tumor growth in nude mouse model. Ectopic expression of CDKN3 significantly reduced the efficiency of Bcr-Abl-mediated transformation of FDCP1 cells to growth factor independence. In contrast, depletion of CDKN3 expression conferred resistance to imatinib-induced apoptosis in the leukemic cells and accelerated the growth of xenograph leukemia in mice. In addition, we found that CDKN3 mutant (CDKN3-C140S devoid of the phosphatase activity failed to affect the K562 leukemic cell survival and xenografted tumor growth, suggesting that the phosphatase of CDKN3 was required for its tumor suppressor function. Furthermore, we observed that overexpression of CDKN3 reduced the leukemic cell survival by dephosphorylating CDK2, thereby inhibiting CDK2-dependent XIAP expression. Moreover, overexpression of CDKN3 delayed G1/S transition in K562 leukemic cells. Our results highlight the importance of CDKN3 in Bcr-Abl-mediated leukemogenesis, and provide new insights into diagnostics and therapeutics of the leukemia.

  11. Identification of drug combinations containing imatinib for treatment of BCR-ABL+ leukemias.

    Directory of Open Access Journals (Sweden)

    Yunyi Kang

    Full Text Available The BCR-ABL translocation is found in chronic myeloid leukemia (CML and in Ph+ acute lymphoblastic leukemia (ALL patients. Although imatinib and its analogues have been used as front-line therapy to target this mutation and control the disease for over a decade, resistance to the therapy is still observed and most patients are not cured but need to continue the therapy indefinitely. It is therefore of great importance to find new therapies, possibly as drug combinations, which can overcome drug resistance. In this study, we identified eleven candidate anti-leukemic drugs that might be combined with imatinib, using three approaches: a kinase inhibitor library screen, a gene expression correlation analysis, and literature analysis. We then used an experimental search algorithm to efficiently explore the large space of possible drug and dose combinations and identified drug combinations that selectively kill a BCR-ABL+ leukemic cell line (K562 over a normal fibroblast cell line (IMR-90. Only six iterations of the algorithm were needed to identify very selective drug combinations. The efficacy of the top forty-nine combinations was further confirmed using Ph+ and Ph- ALL patient cells, including imatinib-resistant cells. Collectively, the drug combinations and methods we describe might be a first step towards more effective interventions for leukemia patients, especially those with the BCR-ABL translocation.

  12. Molecular Detection of BCR/ ABL Fusion Gene in Saudi Acute Lymphoblastic Leukemia Patients

    International Nuclear Information System (INIS)

    Background: Molecular cytogenetics is becoming one of the most useful tools targeting some genes which are generally considered to lead to leukemic transformation (as well as for numerical abnormalities). A fraction of acute lymphoblastic leukemia (ALL) cases carry the translocation t(9;22) (q34;ql1.2) which juxtaposes the ABL proto-oncogene to the BCR gene generating a chimeric gene, BCR/ABL. This aberration is more frequent in adult ALL (20%-40%) than in pediatric ALL >)5%), and predicts poor clinical outcome. Aim of our Work: Is to study BCR/ A BL fusion gene in ALL cases using fluorescent in situ hybridization. Patients and Methods: Twenty newly diagnosed ALL patients, 16 adult and 4 paediatric cases, were included in the study, 11 cases (55%) were of precursor B phenotype, S cases (40%) belonged to T lineage, while one case was bi phenotypic expressing mainly precursor B cell markers tether with CD13, CD33, CD117, Detection of BCR/ABL fusion gene was done using interphase FISH technique and was confirmed molecularly using the RT-PCR technique. Results: BCR/ ABL fusion gene was negative in all the examined cases, yet abnormality involving 9q34, ABL gene, either by addition or deletion was detected in three cases (15%). Two of these cases were associated with BCR gene extra copies (three and four copies, respectively). Conclusion: This may reflect the frequency of association of ABL gene and BCR gene abnormality in our cases, and that absence of fusion gene BCR/ABL does not exclude their role in the leukomogenic process, yet a larger study is required to confirm and detect the prevalence of these gene disturbances in ALL and their association

  13. Loss of the xeroderma pigmentosum group B protein binding site impairs p210 BCR/ABL1 leukemogenic activity

    International Nuclear Information System (INIS)

    Previous studies have demonstrated that p210 BCR/ABL1 interacts directly with the xeroderma pigmentosum group B (XPB) protein, and that XPB is phosphorylated on tyrosine in cells that express p210 BCR/ABL1. In the current study, we have constructed a p210 BCR/ABL1 mutant that can no longer bind to XPB. The mutant has normal kinase activity and interacts with GRB2, but can no longer phosphorylate XPB. Loss of XPB binding is associated with reduced expression of c-MYC and reduced transforming potential in ex-vivo clonogenicity assays, but does not affect nucleotide excision repair in lymphoid or myeloid cells. When examined in a bone marrow transplantation (BMT) model for chronic myelogenous leukemia, mice that express the mutant exhibit attenuated myeloproliferation and lymphoproliferation when compared with mice that express unmodified p210 BCR/ABL1. Thus, the mutant-transplanted mice show predominantly neutrophilic expansion and altered progenitor expansion, and have significantly extended lifespans. This was confirmed in a BMT model for B-cell acute lymphoblastic leukemia, wherein the majority of the mutant-transplanted mice remain disease free. These results suggest that the interaction between p210 BCR/ABL1 and XPB can contribute to disease progression by influencing the lineage commitment of lymphoid and myeloid progenitors

  14. Beneficial effects of combining nilotinib and imatinib in preclinical models of BCR-ABL+ leukemias

    OpenAIRE

    Weisberg, Ellen; Catley, Laurie; Wright, Renee D.; Moreno, Daisy; Banerji, Lolita; Ray, Arghya; Manley, Paul W; Mestan, Juergen; Fabbro, Doriano; Jiang, Jingrui; Hall-Meyers, Elizabeth; Callahan, Linda; DellaGatta, Jamie L.; Kung, Andrew L.; Griffin, James D.

    2007-01-01

    Drug resistance resulting from emergence of imatinib-resistant BCR-ABL point mutations is a significant problem in advanced-stage chronic myelogenous leukemia (CML). The BCR-ABL inhibitor, nilotinib (AMN107), is significantly more potent against BCR-ABL than imatinib, and is active against many imatinib-resistant BCR-ABL mutants. Phase 1/2 clinical trials show that nilotinib can induce remissions in patients who have previously failed imatinib, indicating that sequential therapy with these 2 ...

  15. Distinct GAB2 signaling pathways are essential for myeloid and lymphoid transformation and leukemogenesis by BCR-ABL1.

    Science.gov (United States)

    Gu, Shengqing; Chan, Wayne W; Mohi, Golam; Rosenbaum, Joel; Sayad, Azin; Lu, Zhibin; Virtanen, Carl; Li, Shaoguang; Neel, Benjamin G; Van Etten, Richard A

    2016-04-01

    Tyrosine kinase inhibitors (TKIs) directed against BCR-ABL1, the product of the Philadelphia (Ph) chromosome, have revolutionized treatment of patients with chronic myeloid leukemia (CML). However, acquired resistance to TKIs is a significant clinical problem in CML, and TKI therapy is much less effective against Ph(+)B-cell acute lymphoblastic leukemia (B-ALL). BCR-ABL1, via phosphorylated Tyr177, recruits the adapter GRB2-associated binding protein 2 (GAB2) as part of a GRB2/GAB2 complex. We showed previously that GAB2 is essential for BCR-ABL1-evoked myeloid transformation in vitro. Using a genetic strategy and mouse models of CML and B-ALL, we show here that GAB2 is essential for myeloid and lymphoid leukemogenesis by BCR-ABL1. In the mouse model, recipients of BCR-ABL1-transducedGab2(-/-)bone marrow failed to develop CML-like myeloproliferative neoplasia. Leukemogenesis was restored by expression of GAB2 but not by GAB2 mutants lacking binding sites for its effectors phosphatidylinositol 3-kinase (PI3K) or SRC homology 2-containing phosphotyrosine phosphatase 2 (SHP2). GAB2 deficiency also attenuated BCR-ABL1-induced B-ALL, but only the SHP2 binding site was required. The SHP2 and PI3K binding sites were differentially required for signaling downstream of GAB2. Hence, GAB2 transmits critical transforming signals from Tyr177 to PI3K and SHP2 for CML pathogenesis, whereas only the GAB2-SHP2 pathway is essential for lymphoid leukemogenesis. Given that GAB2 is dispensable for normal hematopoiesis, GAB2 and its effectors PI3K and SHP2 represent promising targets for therapy in Ph(+)hematologic neoplasms. PMID:26773044

  16. A multiple reaction monitoring (MRM method to detect Bcr-Abl kinase activity in CML using a peptide biosensor.

    Directory of Open Access Journals (Sweden)

    Tzu-Yi Yang

    Full Text Available The protein kinase Bcr-Abl plays a major role in the pathogenesis of chronic myelogenous leukemia (CML, and is the target of the breakthrough drug imatinib (Gleevec™. While most patients respond well to imatinib, approximately 30% never achieve remission or develop resistance within 1-5 years of starting imatinib treatment. Evidence from clinical studies suggests that achieving at least 50% inhibition of a patient's Bcr-Abl kinase activity (relative to their level at diagnosis is associated with improved patient outcomes, including reduced occurrence of resistance and longer maintenance of remission. Accordingly, sensitive assays for detecting Bcr-Abl kinase activity compatible with small amounts of patient material are desirable as potential companion diagnostics for imatinib. Here we report the detection of Bcr-Abl activity and inhibition by imatinib in the human CML cell line K562 using a cell-penetrating peptide biosensor and multiple reaction monitoring (MRM on a triple quadrupole mass spectrometer. MRM enabled reproducible, selective detection of the peptide biosensor at fmol levels from aliquots of cell lysate equivalent to ~15,000 cells. This degree of sensitivity will facilitate the miniaturization of the entire assay procedure down to cell numbers approaching 15,000, making it practical for translational applications in patient cells in which the limited amount of available patient material often presents a major challenge.

  17. A clinical and laboratory study of chronic myeloid leukemia with atypical BCR-ABL fusion gene subtypes

    Institute of Scientific and Technical Information of China (English)

    桂晓敏

    2014-01-01

    Objective To explore the clinical and laboratory features of chronic myeloid leukemia(CML)with atypical e14a3 and e19a2 BCR-ABL fusion gene subtypes.Methods We retrospectively analyzed a cohort of CML patients with Ph chromosome positive confirmed by cytogenetic and FISH but classical e13a3(b2a2),e14a2(b3a2)and e1a2 fusion transcripts negative identified by

  18. Frequency of p190 and p210 BCR-ABL rearrangements and survival in Brazilian adult patients with acute lymphoblastic leukemia

    Directory of Open Access Journals (Sweden)

    Ilana de França Azevedo

    2014-10-01

    Full Text Available Objective: This study investigated the occurrence of the p190 and p210 break point clusterregion-Abelson (BCR-ABL rearrangements in adults with acute lymphoblastic leukemia and possible associations with clinical and laboratory characteristics and survival. Methods: Forty-one over 18-year-old patients with acute lymphoblastic leukemia of both genders followed-up between January 2008 and May 2012 were included in this study. Clinical and laboratory data were obtained from the medical charts of the patients. Reverse transcription polymerase chain reaction (RT-PCR using specific primers was employed to identify molecular rearrangements. Results: At diagnosis, the median age was 33 years, and there was a predominance of males (61%. The most common immunophenotype was B lineage (76%. BCR-ABL rearrangements was detected in 14 (34% patients with the following distribution: p190 (28%, p210 (50% and double positive (22%. Overall survival of patients with a mean/median of 331/246 days of follow up was 39%, respectively, negative BCR-ABL (44% and positive BCR-ABL (28%. Conclusion: These results confirm the high frequency of BCR-ABL rearrangements and the low survival rate of adult Brazilian patients with acute lymphoblastic leukemia.

  19. The diagnosis of BCR/ABL-negative chronic myeloproliferative diseases (CMPD): a comprehensive approach based on morphology, cytogenetics, and molecular markers.

    Science.gov (United States)

    Haferlach, Torsten; Bacher, Ulrike; Kern, Wolfgang; Schnittger, Susanne; Haferlach, Claudia

    2008-01-01

    Recent years showed significant progress in the molecular characterization of the chronic myeloproliferative disorders (CMPD) which are classified according to the WHO classification of 2001 as polycythemia vera (PV), chronic idiopathic myelofibrosis (CIMF), essential thrombocythemia (ET), CMPD/unclassifiable (CMPD-U), chronic neutrophilic leukemia, and chronic eosinophilic leukemia (CEL)/hypereosinophilic syndrome, all to be delineated from BCR/ABL-positive chronic myeloid leukemia (CML). After 2001, the detection of the high frequency of the JAK2V617F mutation in PV, CIMF, and ET, and of the FIP1L1-PDGFRA fusion gene in CEL further added important information in the diagnosis of CMPD. These findings also enhanced the importance of tyrosine kinase mutations in CMPD and paved the way to a more detailed classification and to an improved definition of prognosis using also novel minimal residual disease (MRD) markers. Simultaneously, the broadening of therapeutic strategies in the CMPD, e.g., due to reduced intensity conditioning in allogeneic hematopoietic stem cell transplantation and the introduction of tyrosine kinase inhibitors in CML, in CEL, and in other ABL and PDGRFB rearrangements, increased the demands to diagnostics. Therefore, today, a multimodal diagnostic approach combining cytomorphology, cytogenetics, and individual molecular methods is needed in BCR/ABL-negative CMPD. A stringent diagnostic algorithm for characterization, choice of treatment, and monitoring of MRD will be proposed in this review. PMID:17938925

  20. The chimeric ubiquitin ligase SH2-U-box inhibits the growth of imatinib-sensitive and resistant CML by targeting the native and T315I-mutant BCR-ABL.

    Science.gov (United States)

    Ru, Yi; Wang, Qinhao; Liu, Xiping; Zhang, Mei; Zhong, Daixing; Ye, Mingxiang; Li, Yuanchun; Han, Hua; Yao, Libo; Li, Xia

    2016-01-01

    Chronic myeloid leukemia (CML) is characterized by constitutively active fusion protein tyrosine kinase BCR-ABL. Although the tyrosine kinase inhibitor (TKI) against BCR-ABL, imatinib, is the first-line therapy for CML, acquired resistance almost inevitably emerges. The underlying mechanism are point mutations within the BCR-ABL gene, among which T315I is notorious because it resists to almost all currently available inhibitors. Here we took use of a previously generated chimeric ubiquitin ligase, SH2-U-box, in which SH2 from the adaptor protein Grb2 acts as a binding domain for activated BCR-ABL, while U-box from CHIP functions as an E3 ubiquitin ligase domain, so as to target the ubiquitination and degradation of both native and T315I-mutant BCR-ABL. As such, SH2-U-box significantly inhibited proliferation and induced apoptosis in CML cells harboring either the wild-type or T315I-mutant BCR-ABL (K562 or K562R), with BCR-ABL-dependent signaling pathways being repressed. Moreover, SH2-U-box worked in concert with imatinib in K562 cells. Importantly, SH2-U-box-carrying lentivirus could markedly suppress the growth of K562-xenografts in nude mice or K562R-xenografts in SCID mice, as well as that of primary CML cells. Collectively, by degrading the native and T315I-mutant BCR-ABL, the chimeric ubiquitin ligase SH2-U-box may serve as a potential therapy for both imatinib-sensitive and resistant CML. PMID:27329306

  1. Mutations in the BCR-ABL1 Kinase Domain and Elsewhere in Chronic Myeloid Leukemia.

    Science.gov (United States)

    Soverini, Simona; de Benedittis, Caterina; Mancini, Manuela; Martinelli, Giovanni

    2015-06-01

    Chronic myeloid leukemia (CML) has been the first human malignancy to be associated, more than 50 years ago, with a consistent chromosomal abnormality--the t(9;22)(q34;q11) chromosomal translocation. The resulting BCR-ABL1 fusion gene, encoding a tyrosine kinase with deregulated activity, has a central role in the pathogenesis of CML. Ancestral or additional genetic events necessary for CML to develop have long been hypothesized but never really demonstrated. CML can successfully be treated with tyrosine kinase inhibitors (TKIs). Mutations in the BCR-ABL1 kinase domain might arise, however, that confer resistance to 1 or more of the currently available TKIs. Hence, the critical role of BCR-ABL1 mutation screening for optimal therapeutic management, with the current gold standard technique, conventional sequencing, likely to be replaced soon by ultra-deep sequencing. Mutations in genes other than BCR-ABL1 include ASXL1, TET2, RUNX1, DNMT3A, EZH2, and TP53 in chronic phase patients and RUNX1, ASXL1, IKZF1, WT1, TET2, NPM1, IDH1, IDH2, NRAS, KRAS, CBL, TP53, CDKN2A, RB1, and GATA-2 mutations in advanced phase patients. The latter also display additional cytogenetic abnormalities, including submicroscopic regions of gain or loss that only single nucleotide polymorphism arrays or array comparative genomic hybridization can detect. Whether whole genome/exome sequencing studies will uncover novel mutations relevant for pathogenesis, progression, and risk-adapted therapy is still unclear. PMID:26297264

  2. Low expression of miR-196b enhances the expression of BCR-ABL1 and HOXA9 oncogenes in chronic myeloid leukemogenesis.

    Directory of Open Access Journals (Sweden)

    Yue Liu

    Full Text Available MicroRNAs (miRNAs can function as tumor suppressors or oncogene promoters during tumor development. In this study, low levels of expression of miR-196b were detected in patients with chronic myeloid leukemia. Bisulfite genomic sequencing PCR and methylation-specific PCR were used to examine the methylation status of the CpG islands in the miR-196b promoter in K562 cells, patients with leukemia and healthy individuals. The CpG islands showed more methylation in patients with chronic myeloid leukemia compared with healthy individuals (P<0.05, which indicated that low expression of miR-196b may be associated with an increase in the methylation of CpG islands. The dual-luciferase reporter assay system demonstrated that BCR-ABL1 and HOXA9 are the target genes of miR-196b, which was consistent with predictions from bioinformatics software analyses. Further examination of cell function indicated that miR-196b acts to reduce BCR-ABL1 and HOXA9 protein levels, decrease cell proliferation rate and retard the cell cycle. A low level of expression of miR-196b can cause up-regulation of BCR-ABL1 and HOXA9 expression, which leads to the development of chronic myeloid leukemia. MiR-196b may represent an effective target for chronic myeloid leukemia therapy.

  3. Extramedullary blast crisis as initial presentation in chronic myeloid leukemia with the e1a2 BCR-ABL1 transcript: A case report

    Science.gov (United States)

    AI, DI; LIU, WEI; LU, GARY; PATEL, KEYUR PRAVINCHANDRA; CHEN, ZI

    2015-01-01

    A 23-year-old woman presented with enlarged right inguinal lymph nodes. The pathological examination of the nodes revealed infiltration by myeloid sarcoma. A bone marrow smear and biopsy revealed cytogenetic abnormalities, with 46,XX,t(9;22) and chronic myeloid leukemia (CML) was diagnosed. The e1a2 BCR-ABL1 fusion transcript was detected. The patient received imatinib-based combined chemotherapy, allogeneic hematopoietic stem cell transplantation, donor lymphocyte infusions and dasatinib treatment. The patient achieved complete response and has remained leukemia-free for >48 months. To the best of our knowledge, this is the first case report of CML with the e1a2 BCR-ABL1 transcript, with extramedullary blast crisis as the initial presentation. The aim of the present study was to discuss this special case with reference to the literature. PMID:26807241

  4. Molecular Imaging of Bcr-Abl Phosphokinase in a Xenograft Model*

    OpenAIRE

    Wu, Ji Yuan; Yang, David J.; Angelo, Laura S.; Kohanim, Saady; Kurzrock, Razelle

    2009-01-01

    The purpose of this study was to determine whether the Bcr-Abl tyrosine kinase can be assessed by gamma imaging using an 111Indium-labeled anti-phosphotyrosine antibody, and if the response to treatment with imatinib could be detected using this imaging technique. Anti-phosphotyrosine antibody (APT) was labeled with indium (111In) using ethylenedicysteine (EC) as a chelator. To determine if 111In-EC-APT could assess a non-receptor tyrosine kinase, xenografts of the human chronic myelogenous l...

  5. Expression of p190 BCR-ABL fusion gene in a patient with chronic myeloid leukemia Expressão do rearranjo gênico BCR-ABL com ponto de quebra na região menor do gene BCR em um paciente com leucemia mielóide crônica

    Directory of Open Access Journals (Sweden)

    P. V. B. Carvalho

    2003-01-01

    Full Text Available A minority of chronic myeloid leukemia cases have breakpoints in the minor cluster region (m-bcr of the BCR-ABL gene. We report on a patient with Ph-positive and m-bcr breakpoint at diagnosis. She was treated with hydroxyurea and interferon-alpha. Two years later, she developed a lymphoid blast crisis and died shortly after. We discuss herein the different forms of the BCR-ABL oncogene, its products, and the possible influence of them on the clinical outcome of patients with the disease.A leucemia mielóide crônica (LMC é uma doença mieloproliferativa clonal e caracteriza-se pela presença da translocação cromossômica entre os braços longos dos cromossomos 9 e 22, o denominado cromossomo Ph. Esta translocação determina a fusão dos genes BCR e ABL. Os diferentes pontos de quebra no gene BCR determinam a síntese de proteínas com diferentes pesos moleculares pelo gene BCR-ABL. Nós relatamos o caso de uma paciente portadora de LMC com ponto de quebra cromossômico na região menor do gene BCR. Foi tratada com hidroxiuréia e interferon alfa. Dois anos após o diagnóstico desenvolveu crise blástica linfóide e evoluiu rapidamente para o óbito. Nós discutimos nesta apresentação as diferentes formas do gene BCR-ABL e seus produtos e a possível influência dos mesmos na evolução clínica dos pacientes com a doença.

  6. In-silico identification of inhibitors against mutated BCR-ABL protein of chronic myeloid leukemia: a virtual screening and molecular dynamics simulation study.

    Science.gov (United States)

    Kumar, Himansu; Raj, Utkarsh; Gupta, Saurabh; Varadwaj, Pritish Kumar

    2016-10-01

    Aberrant and proliferative expression of the oncogene BCR-ABL in the bone marrow cells had been proven as the prime cause of chronic myeloid leukemia (CML). It has been established that tyrosine kinase domain of BCR-ABL protein is a potential therapeutic target for the treatment of CML. Imatinib is considered as a first-generation drug that can inhibit the enzymatic action by inhibiting the ATP binding with BCR-ABL protein. Later on, insensitivity of CML cells towards Imatinib has been observed may be due to mutation in tyrosine kinase domain of the ABL receptor. Subsequently, some other second-generation drugs have also been reported viz. Baustinib, Nilotinib, Dasatinib, Ponatinib, Bafetinib, etc., which can able to combat against mutated domain of ABL tyrosine kinase protein. By taking into account of bioavailability and resistance developed, there is an utmost need to find some more inhibitors for the mutated ABL tyrosine kinase protein. For virtual screening, a data-set has been generated by collecting the all available drug like natural compounds from ZINC and Drug Bank databases. Comparative docking analysis was also carried out on the active site of ABL tyrosine kinase receptor with reported reference inhibitors. Molecular dynamics simulation of the best screened interacting complex was done for 50 ns to validate the stability of the system. These selected inhibitors were further validated and analyzed through pharmacokinetics properties and series of ADMET parameters by in silico methods. Considering the above said parameters proposed molecules are concluded as potential leads for drug designing pipeline against CML. PMID:26479578

  7. Increased acetylation of lysine 317/320 of p53 caused by BCR-ABL protects from cytoplasmic translocation of p53 and mitochondria-dependent apoptosis in response to DNA damage

    NARCIS (Netherlands)

    Kusio-Kobialka, Monika; Wolanin, Kamila; Podszywalow-Bartnicka, Paulina; Sikora, Ewa; Skowronek, Krzysztof; McKenna, Sharon L.; Ghizzoni, Massimo; Dekker, Frank J.; Piwocka, Katarzyna

    2012-01-01

    Chronic myeloid leukemia (CML) is a disorder of hematopoietic stem cells caused by the expression of BCR-ABL. Loss of p53 has not been implicated as important for the development of CML. Mutations in p53 protein are infrequent, however they correlate with the disease progression. The absence of p53

  8. Functionally deregulated AML1/RUNX1 cooperates with BCR-ABL to induce a blastic phase-like phenotype of chronic myelogenous leukemia in mice.

    Directory of Open Access Journals (Sweden)

    Kiyoko Yamamoto

    Full Text Available Patients in the chronic phase (CP of chronic myelogenous leukemia (CML have been treated successfully following the advent of ABL kinase inhibitors, but once they progress to the blast crisis (BC phase the prognosis becomes dismal. Although mechanisms underlying the progression are largely unknown, recent studies revealed the presence of alterations of key molecules for hematopoiesis, such as AML1/RUNX1. Our analysis of 13 BC cases revealed that three cases had AML1 mutations and the transcript levels of wild-type (wt. AML1 were elevated in BC compared with CP. Functional analysis of representative AML1 mutants using mouse hematopoietic cells revealed the possible contribution of some, but not all, mutants for the BC-phenotype. Specifically, K83Q and R139G, but neither R80C nor D171N mutants, conferred upon BCR-ABL-expressing cells a growth advantage over BCR-ABL-alone control cells in cytokine-free culture, and the cells thus grown killed mice upon intravenous transfer. Unexpectedly, wt.AML1 behaved similarly to K83Q and R139G mutants. In a bone marrow transplantation assay, K83Q and wt.AML1s induced the emergence of blast-like cells. The overall findings suggest the roles of altered functions of AML1 imposed by some, but not all, mutants, and the elevated expression of wt.AML1 for the disease progression of CML.

  9. Detection of BCR-ABL Fusion mRNA Using Reverse Transcriptase Loop-mediated Isothermal Amplification

    Energy Technology Data Exchange (ETDEWEB)

    Dugan, L C; Hall, S; Kohlgruber, A; Urbin, S; Torres, C; Wilson, P

    2011-12-08

    RT-PCR is commonly used for the detection of Bcr-Abl fusion transcripts in patients diagnosed with chronic myelogenous leukemia, CML. Two fusion transcripts predominate in CML, Br-Abl e13a2 and e14a2. They have developed reverse transcriptase isothermal loop-mediated amplification (RT-LAMP) assays to detect these two fusion transcripts along with the normal Bcr transcript.

  10. Structural Mechanism of the Pan-BCR-ABL Inhibitor Ponatinib (AP24534): Lessons for Overcoming Kinase Inhibitor Resistance

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Tianjun; Commodore, Lois; Huang, Wei-Sheng; Wang, Yihan; Thomas, Mathew; Keats, Jeff; Xu, Qihong; Rivera, Victor M.; Shakespeare, William C.; Clackson, Tim; Dalgarno, David C.; Zhu, Xiaotian (ARIAD)

    2012-01-20

    The BCR-ABL inhibitor imatinib has revolutionized the treatment of chronic myeloid leukemia. However, drug resistance caused by kinase domain mutations has necessitated the development of new mutation-resistant inhibitors, most recently against the T315I gatekeeper residue mutation. Ponatinib (AP24534) inhibits both native and mutant BCR-ABL, including T315I, acting as a pan-BCR-ABL inhibitor. Here, we undertook a combined crystallographic and structure-activity relationship analysis on ponatinib to understand this unique profile. While the ethynyl linker is a key inhibitor functionality that interacts with the gatekeeper, virtually all other components of ponatinib play an essential role in its T315I inhibitory activity. The extensive network of optimized molecular contacts found in the DFG-out binding mode leads to high potency and renders binding less susceptible to disruption by single point mutations. The inhibitory mechanism exemplified by ponatinib may have broad relevance to designing inhibitors against other kinases with mutated gatekeeper residues.

  11. Determination of lactate dehydrogenase (LDH and Bcr-Abl transcript in the follow-up of patients with chronic myeloid leukemia = Determinação da lactate desidrogenase (LDH e do transcrito Bcr-Abl em pacientes com leucemia mielóide crônica

    Directory of Open Access Journals (Sweden)

    Roberto Iemitsu Tatakihara

    2010-07-01

    Full Text Available Chronic myeloid leukemia (CML is a malignant myeloproliferative disorder that originates from a pluripotent stem cell characterized by abnormal release of the expanded, malignant stem cell clone from the bone marrow into the bloodstream. The vast majority of patients with CML present Bcr-Abl transcripts. Lactate dehydrogenase (LDH is considered a biochemical marker common for tumor growth, anaerobic glycolysis and has been considered a poor prognostic factor for acute myeloid leukemia. Therefore, this study aimed to evaluate the concentration of LDH in plasma and the detection of the Bcr-Abl transcripts in patients with CML and healthy donors. We analyzed 22 patients demonstrably diagnosed with CML and 56 healthy donors. LDH concentration in plasma was higher in patients with CML. All patients with CML in this study were under treatment, but even so four patients had the Bcr-Abl (b3a2 transcript in peripheral blood. Two out of the four patients with b3a2 showed higher LDH (486 U L-1 and 589 U L-1. Thus, although the study was conducted with small numbers of samples, it is possible to suggest therapy alteration for two patients who presented transcript b3a2 in the peripheral blood samples and whose LDH concentration was high, in order to improve the disease. Leucemia mieloide crônica (LMC é uma desordem mieloproliferativa maligna que é originada de célula-tronco pluripotente caracterizada por expansão anormal, maligna de clones de células tronco da medula óssea na circulação. A grande maioria dos pacientes com LMC apresentam transcritos Bcr-Abl. Lactato desidrogenase (LDH,considerado um marcador bioquímico para crescimento tumoral, glicólise anaeróbica, e tem sido considerado um fator de pior prognóstico da LMC. Portanto, este estudo visa avaliar a concentraçãode LDH no plasma e a detecção do transcrito Bcr-Abl em 22 pacientes com LMC e 56 indivíduos saudáveis. Foram avaliados 22 pacientes com LMC e 56 doadores saudáveis. A

  12. Conventional and fluorescence in situ hybridization analysis of three-way complex BCR-ABL rearrangement in a chronic myeloid leukemia patient

    Directory of Open Access Journals (Sweden)

    Ganguly Bani

    2007-01-01

    Full Text Available Chromosomal analysis was carried out in bone marrow sample of an 11-year-old girl suspected of myeloproliferative disorder. Conventional G-banding study detected a complex three-way translocation involving 7, 9 and 22, which has resulted in the formation of a variant Philadelphia chromosome causing rearrangement of abl and bcr genes in 87% cells. Fluorescence in situ hybridization (FISH confirmed the fusion of bcr-abl oncogene. Thus the bone marrow karyotype was observed as 46,XX (13% / 46,XX,t(7;9;22(q11;q34;q11 (87%. Hyperdiploidy was present in two cells. In this study, both conventional cytogenetic and FISH diagnosis proved to be significant to identify the variant nature of the Philadelphia chromosome and hyperdiploid condition for introduction of a suitable treatment regimen and estimation of life expectancy of the young girl.

  13. Synthesis and characterization of a BODIPY conjugate of the BCR-ABL kinase inhibitor Tasigna® (Nilotinib): Evidence for transport of Tasigna® and its fluorescent derivative by ABC drug transporters

    OpenAIRE

    Shukla, Suneet; Skoumbourdis, Amanda P.; Walsh, Martin J.; Hartz, Anika M. S.; Fung, King Leung; Wu, Chung-pu; Gottesman, Michael M.; Bauer, Björn; Thomas, Craig J.; Suresh V Ambudkar

    2011-01-01

    Tasigna® (Nilotinib) is a recently approved BCR-ABL kinase inhibitor by the Food and Drug Administration, which is indicated for the treatment of drug-resistant chronic myelogenous leukemia (CML). The efflux of tyrosine kinase inhibitors by ATP-binding cassette (ABC) drug transporters, which actively pump these drugs out of cells utilizing ATP as an energy source, has been linked to the development of drug resistance in CML patients. We report here synthesis and characterization of a fluoresc...

  14. Next-generation sequencing for sensitive detection of BCR-ABL1 mutations relevant to tyrosine kinase inhibitor choice in imatinib-resistant patients

    Science.gov (United States)

    Soverini, Simona; De Benedittis, Caterina; Polakova, Katerina Machova; Linhartova, Jana; Castagnetti, Fausto; Gugliotta, Gabriele; Papayannidis, Cristina; Mancini, Manuela; Klamova, Hana; Salvucci, Marzia; Crugnola, Monica; Iurlo, Alessandra; Albano, Francesco; Russo, Domenico; Rosti, Gianantonio; Cavo, Michele; Baccarani, Michele; Martinelli, Giovanni

    2016-01-01

    In chronic myeloid leukemia (CML) and Philadelphia-positive (Ph+) acute lymphoblastic leukemia (ALL) patients who fail imatinib treatment, BCR-ABL1 mutation profiling by Sanger sequencing (SS) is recommended before changing therapy since detection of specific mutations influences second-generation tyrosine kinase inhibitor (2GTKI) choice. We aimed to assess i) in how many patients who relapse on second-line 2GTKI therapy next generation sequencing (NGS) may track resistant mutations back to the sample collected at the time of imatinib resistance, before 2GTKI start (switchover sample) and ii) whether low level mutations identified by NGS always undergo clonal expansion. To this purpose, we used NGS to retrospectively analyze 60 imatinib-resistant patients (CML, n = 45; Ph+ ALL, n = 15) who had failed second-line 2GTKI therapy and had acquired BCR-ABL1 mutations (Group 1) and 25 imatinib-resistant patients (CML, n = 21; Ph+ ALL, n = 4) who had responded to second-line 2GTKI therapy, for comparison (Group 2). NGS uncovered that in 26 (43%) patients in Group 1, the 2GTKI-resistant mutations that triggered relapse were already detectable at low levels in the switchover sample (median mutation burden, 5%; range 1.1%–18.4%). Importantly, none of the low level mutations detected by NGS in switchover samples failed to expand whenever the patient received the 2GTKI to whom they were insensitive. In contrast, no low level mutation that was resistant to the 2GTKI the patients subsequently received was detected in the switchover samples from Group 2. NGS at the time of imatinib failure reliably identifies clinically relevant mutations, thus enabling a more effective therapeutic tailoring. PMID:26980736

  15. 用COLD-PCR方法提高BCR-ABL1激酶区耐药突变检测灵敏度%Application of COLD-PCR for improved detection of BCR-ABL1 kinase domain drug-resistant mutation

    Institute of Scientific and Technical Information of China (English)

    李庆; 刘红星; 孙慧; 王倩; 张之芬; 武焕玲; 李元堂; 陈永金; 田文君

    2013-01-01

    Objective To evaluate the feasibility of the co-amplification using a low denaturation temperature PCR (COLD-PCR) method in improving the detection of BCR-ABL1 fusion gene kinase domain drug-resistant mutation.Methods Conventional nested PCR (CN-PCR) and COLD-PCR protocol were designed to amplify full-length of BCR-ABL1 kinase domain.PCR products were purified and then Sanger sequenced to detect BCR-ABL1 kinase domain mutation (KDM).20 newly diagnosed CML patients and 32 Imatinib (IM) resistant patients were chosen and peripheral blood or bone marrow samples were collected.PCR products of the clinical specimens were purified and then cloned to evaluate the proportion of the KDM allele.Standard samples with different proportion of KDMs were prepared using T-A vectors with KDM and wild type BCR-ABL1 kinase domain sequence,and then CN-PCR,COLD-PCR and Sanger sequencing were performed to compare the detection sensitivity.Results In the 32 IM-resistant patients,23 were identified KDM-positive using CN-PCR and 28 KDM-positive using COLD-PCR.In the 20 newly diagnosed CML patients,none was identified KDM-positive using CN-PCR and 1 was identified carrying a lower proportion of KDM using COLD-PCR.For 16 different types of KDMs,COLD-PCR method could improve the detection sensitivity 4-12 times.Conclusion Detection sensitivity of BCR-ABL1 drug-resistant kinase domain mutation can be effectively improved by using COLD-PCR method and may contribute to early detection of resistance KDMs.%目的 探讨用低变性温度共扩增PCR(COLD-PCR)方法提高BCR-ABL1融合基因激酶区耐药突变检测灵敏度的可行性.方法 本研究为实验诊断研究.设计常规巢式PCR(CN-PCR)和COLD-PCR方案扩增BCR-ABL1激酶区全长,扩增产物纯化后用Sanger测序法检测激酶区突变(KDM).采集32例经伊马替尼(IM)治疗并出现耐药的慢性粒细胞性白血病(CML)患者和20例初诊CML患者外周血或骨髓标本,用上述2种方案检测标本中的KDM.

  16. Selective Depletion of CD45RA+ T Cells From Allogeneic Peripheral Blood Stem Cell Grafts From HLA-Matched Related and Unrelated Donors in Preventing GVHD

    Science.gov (United States)

    2016-07-08

    Accelerated Phase Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Acute Biphenotypic Leukemia; Acute Leukemia of Ambiguous Lineage; Acute Undifferentiated Leukemia; Adult Acute Lymphoblastic Leukemia in Remission; Adult Acute Myeloid Leukemia in Remission; Blast Phase Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Blastic Plasmacytoid Dendritic Cell Neoplasm; Childhood Acute Lymphoblastic Leukemia in Remission; Childhood Acute Myeloid Leukemia in Remission; Graft Versus Host Disease; Lymphoblastic Lymphoma; Recurrent Adult Acute Lymphoblastic Leukemia; Recurrent Adult Acute Myeloid Leukemia; Recurrent Childhood Acute Lymphoblastic Leukemia; Recurrent Childhood Acute Myeloid Leukemia; Recurrent Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Refractory Anemia With Excess Blasts

  17. Allelic expression imbalance of JAK2 V617F mutation in BCR-ABL negative myeloproliferative neoplasms.

    Directory of Open Access Journals (Sweden)

    Hye-Ran Kim

    Full Text Available The discovery of a single point mutation in the JAK2 gene in patients with BCR/ABL-negative myeloproliferative neoplasms (MPNs has not only brought new insights and pathogenesis, but also has made the diagnosis of MPNs much easier. Although, to date, several mechanisms for the contribution of single JAK2V617F point mutation to phenotypic diversity of MPNs have been suggested in multiple studies, but it is not clear how a unique mutation can cause the phenotypic diversity of MPNs. In this study, our results show that allelic expression imbalance of JAK2 V617F mutant frequently occurs and contributes to phenotypic diversity of BCR-ABL-negative MPNs. The proportion of JAK2 V617F mutant allele was significantly augmented in RNA levels as compared with genomic DNA differently by distinct MPNs subtypes. In detail, preferential expression of JAK2 mutant allele showed threefold increase from the cDNA compared with the genomic DNA from patients with essential thrombocythemia and twofold increase in polycythemia vera. In conclusion, allelic expression imbalance of JAK2 V617F mutant proposes another plausible mechanism for the contribution of single JAK2 point mutation to phenotypic diversity of MPNs.

  18. Sphingomyelin synthase 1 activity is regulated by the BCR-ABL oncogene[S

    OpenAIRE

    Burns, Tara Ann; Subathra, Marimuthu; signorelli, Paola; Choi, Young; Yang, Xiaofeng; Wang, Yong; Villani, Maristella; Bhalla, Kapil; Zhou, Daohong; Luberto, Chiara

    2013-01-01

    Sphingomyelin synthase (SMS) produces sphingomyelin while consuming ceramide (a negative regulator of cell proliferation) and forming diacylglycerol (DAG) (a mitogenic factor). Therefore, enhanced SMS activity could favor cell proliferation. To examine if dysregulated SMS contributes to leukemogenesis, we measured SMS activity in several leukemic cell lines and found that it is highly elevated in K562 chronic myelogenous leukemia (CML) cells. The increased SMS in K562 cells was caused by the ...

  19. Donor Lymphocyte Infusion in Treating Patients With Persistent, Relapsed, or Progressing Cancer After Donor Hematopoietic Cell Transplant

    Science.gov (United States)

    2016-07-25

    Blast Phase Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Recurrent Adult Acute Lymphoblastic Leukemia; Recurrent Adult Acute Myeloid Leukemia; Recurrent Hodgkin Lymphoma; Recurrent Non-Hodgkin Lymphoma; Recurrent Plasma Cell Myeloma

  20. Pentostatin and Lymphocyte Infusion in Preventing Graft Rejection in Patients Who Have Undergone Donor Stem Cell Transplant

    Science.gov (United States)

    2016-02-29

    Acute Lymphoblastic Leukemia; Acute Myeloid Leukemia; Chronic Lymphocytic Leukemia; Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Graft Versus Host Disease; Hodgkin Lymphoma; Myelodysplastic/Myeloproliferative Neoplasm; Non-Hodgkin Lymphoma; Plasma Cell Myeloma; Waldenstrom Macroglobulinemia

  1. Molecular approach to diagnose BCR/ABL negative chronic myeloproliferative neoplasms

    Directory of Open Access Journals (Sweden)

    Michelle Maccarini Barcelos

    2011-01-01

    Full Text Available Chronic myeloproliferative neoplasms arise from clonal proliferation of hematopoietic stem cells. According to the World Health Organization myeloproliferative neoplasms are classified as: chronic myelogenous leukemia, polycythemia vera, essential thrombocythemia, primary myelofibrosis, chronic neutrophilic leukemia, chronic eosinophilic leukemia, hypereosinophilic syndrome, mast cell disease, and unclassifiable myeloproliferative neoplasms. In the revised 2008 WHO diagnostic criteria for myeloproliferative neoplasms, mutation screening for JAK2V617F is considered a major criterion for polycythemia vera diagnosis and also for essential thrombocythemia and primary myelofibrosis, the presence of this mutation represents a clonal marker. There are currently two hypotheses explaining the role of the JAK2V617F mutation in chronic myeloproliferative neoplasms. According to these theories, the mutation plays either a primary or secondary role in disease development. The discovery of the JAK2V617F mutation has been essential in understanding the genetic basis of chronic myeloproliferative neoplasms, providing some idea on how a single mutation can result in three different chronic myeloproliferative neoplasm phenotypes. But there are still some issues to be clarified. Thus, studies are still needed to determine specific molecular markers for each subtype of chronic myeloproliferative neoplasm.

  2. BCR-ABL negative myeloproliferative neoplasia: a review of involved molecular mechanisms.

    Science.gov (United States)

    Koopmans, Suzanne M; Schouten, Harry C; van Marion, Ariënne M W

    2015-02-01

    The clonal bone marrow stem cell disorders essential thrombocythemia (ET), polycythemia vera (PV) and primary myelofibrosis (PMF) belong to the group of Philadelphia chromosome negative myeloproliferative neoplasia (Ph- MPN). In 2005 the JAK2(V617F) mutation was discovered which has generated more insight in the pathogenetic mechanism of the MPNs. More mutations have been detected in MPN patients since. However, the underlying cause of MPN has not been discovered so far. The mechanism of increased angiogenesis in MPNs and the development of fibrosis in the bone marrow in PMF patients and in some ET and PV patients is still not known. This review will focus on the most important molecular pathogenetic mechanisms in MPN patients. PMID:25196073

  3. Recent advances in the bcr-abl negative chronic myeloproliferative diseases

    Directory of Open Access Journals (Sweden)

    Stroncek David F

    2006-10-01

    Full Text Available Abstract The chronic myeloproliferative disorders are clonal hematopoietic stem cell disorders of unknown etiology. In one of these (chronic myeloid leukemia, there is an associated pathognomonic chromosomal abnormality known as the Philadelphia chromosome. This leads to constitutive tyrosine kinase activity which is responsible for the disease and is used as a target for effective therapy. This review concentrates on the search in the other conditions (polycythemia vera, essential thrombocythemia and idiopathic mylofibrosis for a similar biological marker with therapeutic potential. There is no obvious chromosomal marker in these conditions and yet evidence of clonality can be obtained in females by the use of X-inactivation patterns. PRV-1mRNA over expression, raised vitamin B12 levels and raised neutrophil alkaline phosphatase scores are evidence that cells in these conditions have received excessive signals for proliferation, maturation and reduced apoptosis. The ability of erythroid colonies to grow spontaneously without added external erythropoietin in some cases, provided a useful marker and a clue to this abnormal signaling. In the past year several important discoveries have been made which go a long way in elucidating the involved pathways. The recently discovered JAK2 V617F mutation which occurs in the majority of cases of polycythemia vera and in about half of the cases with the two other conditions, enables constitutive tyrosine kinase activity without the need for ligand binding to hematopoietic receptors. This mutation has become the biological marker for these conditions and has spurred the development of a specific therapy to neutralize its effects. The realization that inherited mutations in the thrombopoietin receptor (c-Mpl can cause a phenotype of thrombocytosis such as in Mpl Baltimore (K39N and in a Japanese family with S505A, has prompted the search for acquired mutations in this receptor in chronic myeloproliferative

  4. BCR/ABL-negative primitive progenitors suitable for transplantation can be selected from the marrow of most early-chronic phase but not accelerated-phase chronic myelogenous leukemia patients

    OpenAIRE

    Verfaillie, Catherine; R Bhatia; Miller, W.; F. Mortari; Van Roy, V.; Burger, S.; Mccullough, J; Stieglbauer, K; Dewald, G; Heimfeld, S; Miller, J. S.; McGlave, P B

    1996-01-01

    We have previously reported that selection of marrow cells on the basis of the CD34+HLA-DR- phenotype (34+DR-) may result in the recovery of Philadelphia chromosome (Ph)- and BCR/ABL-negative long-term culture-initiating cells (LTC-IC) in selected patients with chronic myelogenous leukemia (CML). We now present data on 27 early chronic-phase ([ECP] studied within 1 year after diagnosis) and 23 advanced-phase ([AP] late chronic phase, ie, studied >1 year from diagnosis, or accelerated phase) C...

  5. A certified plasmid reference material for the standardisation of BCR-ABL1 mRNA quantification by real-time quantitative PCR

    OpenAIRE

    White, Helen; DEPREZ LIESBET; Corbisier, Philippe; HALL Victoria; Lin, Feng; Mazoua, Stephane; TRAPMANN Stefanie; AGGERHOLM Annie; Andrikovics, Hajnalka; AKIKI Susanna; Barbany, Gisela; Boeckx, Nancy; Bench, Anthony; CATHERWOOD Mark; Cayuela, Jean-Michel

    2014-01-01

    Serial quantification of BCR-ABL1 mRNA is an important therapeutic indicator in chronic myeloid leukaemia, but there is a substantial variation in results reported by different laboratories. To improve comparability, an internationally accepted plasmid certified reference material (CRM) was developed according to ISO Guide 34:2009. Fragments of BCR-ABL1 (e14a2 mRNA fusion), BCR and GUSB transcripts were amplified and cloned into pUC18 to yield plasmid pIRMM0099. Six different linearised plasm...

  6. Myeloproliferative neoplasms (BCR-ABL1 negative) and myelodysplastic/myeloproliferative neoplasms: current diagnostic principles and upcoming updates.

    Science.gov (United States)

    Geyer, J T; Orazi, A

    2016-05-01

    Since the publication of the latest World Health Organization (WHO) classification in 2008, there has been a significant effort for clarification of unresolved questions, especially with the help of the rapidly developing field of molecular genetic studies, next-generation sequencing in particular. Numerous entities within the WHO categories of myeloproliferative neoplasms (MPNs) and myelodysplastic (MDS)/MPNs have been extensively studied, with large published series attempting to characterize and better define their morphologic and molecular genetic features. This emerging genetic landscape maintains a robust correlation with the various disease entities recognized by the WHO classification scheme based on a careful integration of detailed clinical information, bone marrow and peripheral blood morphology, immunohistology, and genomics. This brief review summarizes the current guidelines as they apply to diagnosing both the classical BCR-ABL1 negative MPN (polycythemia vera, essential thrombocythemia, and primary myelofibrosis) and the more common subtypes of MDS/MPN overlap syndromes. The more important recent molecular updates as well as the upcoming changes to the current WHO classification, expected to be published in late 2016, will also be briefly reviewed. PMID:27161873

  7. Epidemiologic study on survival of chronic myeloid leukemia and Ph(+) acute lymphoblastic leukemia patients with BCR-ABL T315I mutation

    DEFF Research Database (Denmark)

    Nicolini, Franck E; Mauro, Michael J; Martinelli, Giovanni;

    2009-01-01

    The BCR-ABL T315I mutation represents a major mechanism of resistance to tyrosine kinase inhibitors (TKIs). The objectives of this retrospective observational study were to estimate overall and progression-free survival for chronic myeloid leukemia in chronic-phase (CP), accelerated-phase (AP), o...

  8. Rapid Evolution to Blast Crisis Associated with a Q252H ABL1 Kinase Domain Mutation in e19a2 BCR-ABL1 Chronic Myeloid Leukaemia

    Directory of Open Access Journals (Sweden)

    Sarah L. McCarron

    2013-01-01

    Full Text Available A minority of chronic myeloid leukaemia (CML patients express variant transcripts of which the e19a2 BCR-ABL1 fusion is the most common. Instances of tyrosine kinase inhibitor (TKI resistance in e19a2 BCR-ABL1 CML patients have rarely been reported. A case of e19a2 BCR-ABL1 CML is described in whom imatinib resistance, associated with a Q252H ABL1 kinase domain mutation, became apparent soon after initiation of TKI therapy. The patient rapidly transformed to myeloid blast crisis (BC with considerable bone marrow fibrosis and no significant molecular response to a second generation TKI. The clinical course was complicated by comorbidities with the patient rapidly succumbing to advanced disease. This scenario of Q252H-associated TKI resistance with rapid BC transformation has not been previously documented in e19a2 BCR-ABL1 CML. This case highlights the considerable challenges remaining in the management of TKI-resistant BC CML, particularly in the elderly patient.

  9. SPR Detection and Discrimination of the Oligonucleotides Related to the Normal and the Hybrid bcr-abl Genes by Two Stringency Control Strategies.

    Science.gov (United States)

    Matsishin, M J; Ushenin, Iu V; Rachkov, A E; Solatkin, A P

    2016-12-01

    In this study, we applied two stringency control strategies for surface plasmon resonance (SPR) detection of DNA hybridization and discrimination of completely and partially complementary 24-mer sequences. These sequences are specific to the human normal bcr and the hybrid bcr-abl genes, protein products of which are responsible for some leukemia. SPR sensors based on resonance phenomena in nanoscale gold films are well suited for label-free, real-time investigations of the macromolecule interactions. Thermodynamic parameters obtained using the web server DINAMelt allowed supposing the possibility for realization (a) stringency control based on the ionic strength of the hybridization buffer and (b) stringency control based on the temperature elevation. The first one resulted in that the discrimination index of completely complementary and partially complementary oligonucleotides depending on the target concentration varied from 1.3 to 1.8 in 2 × SSC and from 2.0 to 2.9 in 0.5 × SSC. For implementation of the second stringency control strategy, SPR spectrometer measuring flow cell with built-in high-precision temperature control and regulation as well as corresponding software was created. It is shown that the duplexes formed by the immobilized probes mod-Ph and completely complementary oligonucleotides P1 remained without significant changes until ~50 °C, while the duplexes formed with partially complementary oligonucleotide Bcrex14 almost entirely disrupted at 40 °C. Thus, the absolutely effective thermodiscrimination of this pair of oligonucleotides was achieved in this temperature range (40-50 °C). PMID:26759355

  10. SPR Detection and Discrimination of the Oligonucleotides Related to the Normal and the Hybrid bcr-abl Genes by Two Stringency Control Strategies

    Science.gov (United States)

    Matsishin, M. J.; Ushenin, Iu. V.; Rachkov, A. E.; Solatkin, A. P.

    2016-01-01

    In this study, we applied two stringency control strategies for surface plasmon resonance (SPR) detection of DNA hybridization and discrimination of completely and partially complementary 24-mer sequences. These sequences are specific to the human normal bcr and the hybrid bcr-abl genes, protein products of which are responsible for some leukemia. SPR sensors based on resonance phenomena in nanoscale gold films are well suited for label-free, real-time investigations of the macromolecule interactions. Thermodynamic parameters obtained using the web server DINAMelt allowed supposing the possibility for realization (a) stringency control based on the ionic strength of the hybridization buffer and (b) stringency control based on the temperature elevation. The first one resulted in that the discrimination index of completely complementary and partially complementary oligonucleotides depending on the target concentration varied from 1.3 to 1.8 in 2 × SSC and from 2.0 to 2.9 in 0.5 × SSC. For implementation of the second stringency control strategy, SPR spectrometer measuring flow cell with built-in high-precision temperature control and regulation as well as corresponding software was created. It is shown that the duplexes formed by the immobilized probes mod-Ph and completely complementary oligonucleotides P1 remained without significant changes until ~50 °C, while the duplexes formed with partially complementary oligonucleotide Bcrex14 almost entirely disrupted at 40 °C. Thus, the absolutely effective thermodiscrimination of this pair of oligonucleotides was achieved in this temperature range (40-50 °C).

  11. BCR-ABL transcript variations in chronic phase chronic myelogenous leukemia patients on imatinib first-line: Possible role of the autologous immune system.

    Science.gov (United States)

    Clapp, Geoffrey D; Lepoutre, Thomas; Nicolini, Franck E; Levy, Doron

    2016-05-01

    Many chronic myelogenous leukemia (CML) patients in chronic phase who respond well to imatinib therapy show fluctuations in their leukemic loads in the long-term. We developed a mathematical model of CML that incorporates the intervention of an autologous immune response. Our results suggest that the patient's immune system plays a crucial role in imatinib therapy in maintaining disease control over time. The observed BCR-ABL/ABL oscillations in such patients provide a signature of the autologous immune response. PMID:27467931

  12. Coupling a universal DNA circuit with graphene sheets/polyaniline/AuNPs nanocomposites for the detection of BCR/ABL fusion gene

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Xueping [Key Laboratory of Laboratory Medical Diagnostics of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016 (China); Wang, Li [Key Laboratory of Laboratory Medical Diagnostics of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016 (China); Department of Medical Laboratory, Chongqing Emergency Medical Center (Chongqing The Fourth Hospital), Chongqing, 400016 (China); Sheng, Shangchun [The No.2 Peoples' Hospital of Yibin, Sichuan, 644000 (China); Wang, Teng; Yang, Juan [Key Laboratory of Laboratory Medical Diagnostics of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016 (China); Xie, Guoming, E-mail: guomingxie@cqmu.edu.cn [Key Laboratory of Laboratory Medical Diagnostics of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016 (China); Feng, Wenli, E-mail: fengwlcqmu@sina.com [Key Laboratory of Laboratory Medical Diagnostics of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016 (China)

    2015-08-19

    This article described a novel method by coupling a universal DNA circuit with graphene sheets/polyaniline/AuNPs nanocomposites (GS/PANI/AuNPs) for highly sensitive and specific detection of BCR/ABL fusion gene (bcr/abl) in chronic myeloid leukemia (CML). DNA circuit known as catalyzed hairpin assembly (CHA) is enzyme-free and can be simply operated to achieve exponential amplification, which has been widely employed in biosensing. However, application of CHA has been hindered by the need of specially redesigned sequences for each single-stranded DNA input. Herein, a transducer hairpin (HP) was designed to obtain a universal DNA circuit with favorable signal-to-background ratio. To further improve signal amplification, GS/PANI/AuNPs with excellent conductivity and enlarged effective area were introduced into this DNA circuit. Consequently, by combining the advantages of CHA and GS/PANI/AuNPs, bcr/abl could be detected in a linear range from 10 pM to 20 nM with a detection limit of 1.05 pM. Moreover, this protocol showed excellent specificity, good stability and was successfully applied for the detection of real sample, which demonstrated its great potential in clinical application. - Highlights: • A transducer hairpin was designed to improve the versatility of DNA circuit. • GS/PANI/AuNPs were introduced to the DNA circuit for further signal amplification. • The established biosensor displayed high sensitivity and good specificity.

  13. Coupling a universal DNA circuit with graphene sheets/polyaniline/AuNPs nanocomposites for the detection of BCR/ABL fusion gene

    International Nuclear Information System (INIS)

    This article described a novel method by coupling a universal DNA circuit with graphene sheets/polyaniline/AuNPs nanocomposites (GS/PANI/AuNPs) for highly sensitive and specific detection of BCR/ABL fusion gene (bcr/abl) in chronic myeloid leukemia (CML). DNA circuit known as catalyzed hairpin assembly (CHA) is enzyme-free and can be simply operated to achieve exponential amplification, which has been widely employed in biosensing. However, application of CHA has been hindered by the need of specially redesigned sequences for each single-stranded DNA input. Herein, a transducer hairpin (HP) was designed to obtain a universal DNA circuit with favorable signal-to-background ratio. To further improve signal amplification, GS/PANI/AuNPs with excellent conductivity and enlarged effective area were introduced into this DNA circuit. Consequently, by combining the advantages of CHA and GS/PANI/AuNPs, bcr/abl could be detected in a linear range from 10 pM to 20 nM with a detection limit of 1.05 pM. Moreover, this protocol showed excellent specificity, good stability and was successfully applied for the detection of real sample, which demonstrated its great potential in clinical application. - Highlights: • A transducer hairpin was designed to improve the versatility of DNA circuit. • GS/PANI/AuNPs were introduced to the DNA circuit for further signal amplification. • The established biosensor displayed high sensitivity and good specificity

  14. RT-PCR ANALYSIS OF E2A-PBX1, TEL-AML1, BCR-ABL AND MLL-AF4 FUSION GENE TRANSCRIPTS IN B-LINEAGE ACUTE LYMPHOBLASTIC LEUKEMIA

    Directory of Open Access Journals (Sweden)

    Iuliu-Cristian Ivanov

    2013-11-01

    Full Text Available Acute lymphoblastic leukemia represents a heterogeneous group of hematological malignancies, defined by clonal proliferation of lymphoid cells. Immunophenotyping by flow cytometry and molecular analysis for the detection of genetic anomalies are clinical standard procedures for diagnosis, sub-classification and post-therapeutic evaluation. Samples from 105 patients diagnosed with acute lymphoblastic leukemia were immunophenotyped at diagnosis and were investigated by molecular analysis in order to identify the occurrence of four fusion genes: MLL-AF4, TEL-AML-1, BCR-ABL-p190, E2A-PBX-1. There were no associations found between the immunophenotype and the presence of any fusion genes evaluated. Both methods in combination remain a prerequisite for an improved subclassification of hematological malignancies, therapeutic decision, and evaluation of treatment response.

  15. Leucine replaced by methionine at 273 position in chronic myeloid leukemia: Knowns and unknowns…

    Directory of Open Access Journals (Sweden)

    Aditi Harsh Thanky

    2016-01-01

    Full Text Available Chronic myeloid leukemia is a clonal bone marrow stem cell disorder characterized by the presence of Philadelphia chromosome t(9;22(q34;q11 leading to fusion oncogene BCR-ABL. Tyrosine kinase inhibitors (TKIs act by competitively inhibiting BCR-ABL oncoprotein with significant response rates. However, up to 30% of patients fail to achieve complete cytogenetic remission on 1st line TKI imatinib, one of the reasons being mutations in BCR-ABL kinase domain leading to imatinib resistance. Over 80 such mutations have been documented in the literature; however, some of the rare mutations still remain to be studied for their impact in development of resistance and their responsiveness to currently available therapeutic options. Here, we report one such case of a rare mutation leucine replaced by methionine at 273 position and its clinical implications.

  16. Optimization of novel combi-molecules: identification of balanced and mixed bcr-abl/DNA targeting properties.

    Science.gov (United States)

    Rachid, Zakaria; Katsoulas, Athanasia; Williams, Christopher; Larroque, Anne-Laure; McNamee, James; Jean-Claude, Bertrand J

    2007-08-01

    Steps toward the identification of combi-molecules with strong abl tyrosine kinase (TK) inhibitory property and significant DNA damaging potential are described. The optimized combi-molecule 13a was shown to induce approximately twofold stronger abl TK inhibitory activity than Gleevec and high levels of DNA damage in chronic myelogenous leukemic cells. PMID:17572088

  17. A combination of STI571 and BCR-ABL1 siRNA with overexpressed p15INK4B induced enhanced proliferation inhibition and apoptosis in chronic myeloid leukemia

    Energy Technology Data Exchange (ETDEWEB)

    Xia, D.Y.; Liu, L.; Hao, M.W.; Liu, Q.; Chen, R.A.; Liang, Y.M. [Department of Hematology, Tangdu Hospital, Fourth Military Medical University, Xi' an (China)

    2014-10-14

    p15INK4B, a cyclin-dependent kinase inhibitor, has been recognized as a tumor suppressor. Loss of or methylation of the p15INK4B gene in chronic myeloid leukemia (CML) cells enhances myeloid progenitor formation from common myeloid progenitors. Therefore, we examined the effects of overexpressed p15INK4B on proliferation and apoptosis of CML cells. Overexpression of p15INK4B inhibited the growth of K562 cells by downregulation of cyclin-dependent kinase 4 (CDK4) and cyclin D1 expression. Overexpression of p15INK4B also induced apoptosis of K562 cells by upregulating Bax expression and downregulating Bcl-2 expression. Overexpression of p15INK4B together with STI571 (imatinib) or BCR-ABL1 small interfering RNA (siRNA) also enhanced growth inhibition and apoptosis induction of K562 cells. The enhanced effect was also mediated by reduction of cyclin D1 and CDK4 and regulation of Bax and Bcl-2. In conclusion, our study may provide new insights into the role of p15INK4B in CML and a potential therapeutic target for overcoming tyrosine kinase inhibitor resistance in CML.

  18. BCR-ABL1: Test

    Science.gov (United States)

    ... Complete Blood Count ; Bone Marrow Aspiration and Biopsy ; Blood Smear ; WBC Differential At a Glance Test Sample The ... are often evaluated as part of the initial diagnosis, but the majority of follow-up monitoring is performed on blood samples. There is significant test variability among laboratories ...

  19. Standardization of real-time quantitative polymerase chain reaction for detection of the JAK2V617F mutation in BCR-ABL1 negative myeloproliferative neoplasms: A tertiary care centre experience

    Directory of Open Access Journals (Sweden)

    Tathagat Chatterjee

    2015-01-01

    Full Text Available Background: Identification of JAK2V617F mutations has led to a significant development in our understanding of the pathogenesis and therapy of BCR-ABL1 negative myeloproliferative neoplasms (MPNs. However, not all cases of BCR-ABL1 negative MPNs carry JAK2V617F mutations. The present study was undertaken with an aim to standardize the real-time quantitative polymerase chain reaction (PCR for the detection of JAK2V617F mutations and to find the prevalence of Janus kinase 2 (JAK2 mutations in MPNs in the Indian scenario. Materials and Methods: Real-time quantitative PCR was used to detect the JAK2V617F mutation. Standardization of the detection procedure was carried out using recommended guidelines to ensure the accuracy and reproducibility of results. Forty-nine patients of BCR-ABL1 negative MPNs were included in the study. Results: The JAK2V617F mutation was detected in 63.3% patients of BCR-ABL1 negative MPNs. On classification of these BCR-ABL1 negative MPNs, JAK2V617F mutation was detected in 78.3% patients with polycythemia vera (PV, 62.5% patients with essential thrombocythemia (ET, and 44.4% patients with Primary myelofibrosis (PMF. Conclusion: Role of detection of JAK2 mutations in BCR-ABL1 negative MPN has begun to be described and can be used in the diagnosis of PV, ET, and PMF along with other criterias. In future, it may be suitable for treatment monitoring and prognostication.

  20. Total-Body Irradiation With or Without Fludarabine Phosphate Followed By Donor Stem Cell Transplant in Treating Patients With Hematologic Cancer

    Science.gov (United States)

    2016-02-02

    Acute Lymphoblastic Leukemia in Remission; Acute Myeloid Leukemia in Remission; Aggressive Non-Hodgkin Lymphoma; Chronic Phase Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Diffuse Large B-Cell Lymphoma; Hematopoietic and Lymphoid Cell Neoplasm; Indolent Non-Hodgkin Lymphoma; Mantle Cell Lymphoma; Myelodysplastic/Myeloproliferative Neoplasm; Plasma Cell Myeloma; Refractory Chronic Lymphocytic Leukemia; Refractory Hodgkin Lymphoma; Waldenstrom Macroglobulinemia

  1. Relationship between JAK2 gene V617F mutation and clinical characteristics in patients with BCR-ABL-negative myeloproliferative neoplasms%BCR-ABL阴性的骨髓增殖性肿瘤JAK2基因V617F突变分析

    Institute of Scientific and Technical Information of China (English)

    张燕香; 魏蓉

    2013-01-01

    ,42 subjects harbored V617F mutation (64.62%).The positive rates of V617F mutation in PV and ET patients were 75.00% (21 subjects) and 56.67% (17 subjects),respectively.Four of the 7 patients with IMF were V617F mutation positive.For PV patients,the levels of white blood cell count and platelet count in V617F mutation carriers were significantly higher than that in non-carriers (P<0.05).For ET patients,the levels of white blood cell count and hemoglobin in V617F mutation carriers were significantly higher than that in non-carriers (P<0.05).For IMF patients,however,no significant differences were found between mutation carriers and non-carriers.The incidence of thrombosis in V617F mutation carriers was higher than that in non-carriers (P<0.001).Conclusions:V617F mutation in JAK2 is a major molecular genetic marker for Chinese patients with BCR-ABL-negative myeloproliferative neoplasms.This mutation has impacts on clinical characteristics and can be used as a criterion for the diagnosis of BCR-ABL-negative myeloproliferative neoplasm.

  2. Failure of CDKN2A/B (INK4A/B–ARF)-mediated tumor suppression and resistance to targeted therapy in acute lymphoblastic leukemia induced by BCR-ABL

    OpenAIRE

    Mullighan, Charles G.; Williams, Richard T.; Downing, James R.; Sherr, Charles J.

    2008-01-01

    Deletions of the CDKN2A/B tumor suppressor locus and of the IKAROS and PAX5 genes that promote B-lineage development occur frequently in lymphoid, but not myeloid leukemias initiated by the BCR-ABL tyrosine kinase. Why is this the case, and how do these genetic lesions contribute to an aggressive disease that fails to durably respond to targeted kinase inhibitors?

  3. A phase 2 study of MK-0457 in patients with BCR-ABL T315I mutant chronic myelogenous leukemia and philadelphia chromosome-positive acute lymphoblastic leukemia

    DEFF Research Database (Denmark)

    Seymour, J F; Kim, D W; Rubin, E;

    2014-01-01

    achieved major hematologic response. The most common adverse event (AE) was neutropenia (50%). The most common grade 3/4 AEs were neutropenia (46%) and febrile neutropenia (35%). MK-0457 demonstrated minimal efficacy and only at higher, intolerable doses; lower doses were tolerated and no unexpected...

  4. Pharmacophore Modeling of Nilotinib as an Inhibitor of ATP-Binding Cassette Drug Transporters and BCR-ABL Kinase Using a Three-Dimensional Quantitative Structure–Activity Relationship Approach

    OpenAIRE

    Shukla, Suneet; Kouanda, Abdul; Silverton, Latoya; Talele, Tanaji T.; Suresh V Ambudkar

    2014-01-01

    Nilotinib (Tasigna) is a tyrosine kinase inhibitor approved by the FDA to treat chronic phase chronic myeloid leukemia patients. It is also a transport substrate of the ATP-binding cassette (ABC) drug efflux transporters ABCB1 (P-glycoprotein, P-gp) and ABCG2 (BCRP), which may have an effect on the pharmacokinetics and toxicity of this drug. The goal of this study was to identify pharmacophoric features of nilotinib in order to potentially develop specific inhibitors of BCR-ABL kinase with mi...

  5. The mTOR inhibitor, everolimus (RAD001), overcomes resistance to imatinib in quiescent Ph-positive acute lymphoblastic leukemia cells

    International Nuclear Information System (INIS)

    In Ph-positive (Ph+) leukemia, the quiescent cell state is one of the reasons for resistance to the BCR-ABL-kinase inhibitor, imatinib. In order to examine the mechanisms of resistance due to quiescence and the effect of the mammalian target of rapamycin inhibitor, everolimus, for such a resistant population, we used Ph+ acute lymphoblastic leukemia patient cells serially xenotransplanted into NOD/SCID/IL2rγnull (NOG) mice. Spleen cells from leukemic mice showed a higher percentage of slow-cycling G0 cells in the CD34+CD38− population compared with the CD34+CD38+ and CD34− populations. After ex vivo imatinib treatment, more residual cells were observed in the CD34+CD38− population than in the other populations. Although slow-cycling G0 cells were insensitive to imatinib in spite of BCR-ABL and CrkL dephosphorylation, combination treatment with everolimus induced substantial cell death, including that of the CD34+CD38− population, with p70-S6 K dephosphorylation and decrease of MCL-1 expression. The leukemic non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mouse system with the in vivo combination treatment with imatinib and everolimus showed a decrease of tumor burden including CD34+ cells. These results imply that treatment with everolimus can overcome resistance to imatinib in Ph+ leukemia due to quiescence

  6. Over-expression of fibroblast growth factor receptor 3 in CD34+BCR-ABL+ cells from patients with chronic myeloid leukemia

    Czech Academy of Sciences Publication Activity Database

    Dvořáková, D.; Mayer, J.; Doubek, M.; Faitová, Jitka; Pacholíková, J.; Hampl, Aleš; Dvořák, Petr

    Istanbul : Nature Publishing Group, 2003. s. 246. [Annual Meeting of the European Group for Blood and Marrow Transplantation /29./. 23.03.2003-26.03.2003, Istanbul] R&D Projects: GA MŠk LN00A065; GA MŠk ME 198 Institutional research plan: CEZ:MSM 432100001; CEZ:MZd 00065269705; CEZ:AV0Z5039906 Keywords : leukemia * fibroblast Subject RIV: FH - Neurology

  7. Graft-Versus-Host Disease Prophylaxis in Treating Patients With Hematologic Malignancies Undergoing Unrelated Donor Peripheral Blood Stem Cell Transplant

    Science.gov (United States)

    2016-08-18

    Acute Lymphoblastic Leukemia; Acute Myeloid Leukemia; Aggressive Non-Hodgkin Lymphoma; Chronic Lymphocytic Leukemia; Diffuse Large B-Cell Lymphoma; Hematopoietic and Lymphoid Cell Neoplasm; Indolent Non-Hodgkin Lymphoma; Mantle Cell Lymphoma; Myelodysplastic Syndrome; Myeloproliferative Neoplasm; Prolymphocytic Leukemia; Recurrent Chronic Lymphocytic Leukemia; Recurrent Plasma Cell Myeloma; Refractory Chronic Lymphocytic Leukemia; Refractory Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Refractory Hodgkin Lymphoma; Small Lymphocytic Lymphoma; T-Cell Chronic Lymphocytic Leukemia; Waldenstrom Macroglobulinemia

  8. Uptake of synthetic Low Density Lipoprotein by leukemic stem cells--a potential stem cell targeted drug delivery strategy.

    Science.gov (United States)

    Zhou, Peixun; Hatziieremia, Sophia; Elliott, Moira A; Scobie, Linda; Crossan, Claire; Michie, Alison M; Holyoake, Tessa L; Halbert, Gavin W; Jørgensen, Heather G

    2010-12-20

    Chronic Myeloid Leukemia (CML) stem/progenitor cells, which over-express Bcr-Abl, respond to imatinib by a reversible block in proliferation without significant apoptosis. As a result, patients are unlikely to be cured owing to the persistence of leukemic quiescent stem cells (QSC) capable of initiating relapse. Previously, we have reported that intracellular levels of imatinib in primary primitive CML cells (CD34+38(lo/⁻)), are significantly lower than in CML progenitor cells (total CD34+) and leukemic cell lines. The aim of this study was to determine if potentially sub-therapeutic intracellular drug concentrations in persistent leukemic QSC may be overcome by targeted drug delivery using synthetic Low Density Lipoprotein (sLDL) particles. As a first step towards this goal, however, the extent of uptake of sLDL by leukemic cell lines and CML patient stem/progenitor cells was investigated. Results with non-drug loaded particles have shown an increased and preferential uptake of sLDL by Bcr-Abl positive cell lines in comparison to Bcr-Abl negative. Furthermore, CML CD34+ and primitive CD34+38(lo/⁻) cells accumulated significantly higher levels of sLDL when compared with non-CML CD34+ cells. Thus, drug-loading the sLDL nanoparticles could potentially enhance intracellular drug concentrations in primitive CML cells and thus aid their eradication. PMID:20869412

  9. Fludarabine Phosphate, Cyclophosphamide, Total-Body Irradiation, and Donor Bone Marrow Transplant Followed by Donor Natural Killer Cell Therapy, Mycophenolate Mofetil, and Tacrolimus in Treating Patients With Hematologic Cancer

    Science.gov (United States)

    2016-06-07

    Acute Lymphoblastic Leukemia; Acute Myeloid Leukemia; Aggressive Non-Hodgkin Lymphoma; Diffuse Large B-Cell Lymphoma; Previously Treated Myelodysplastic Syndrome; Recurrent Chronic Lymphocytic Leukemia; Recurrent Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Recurrent Indolent Adult Non-Hodgkin Lymphoma; Recurrent Mantle Cell Lymphoma; Recurrent Plasma Cell Myeloma; Recurrent Small Lymphocytic Lymphoma; Refractory Chronic Lymphocytic Leukemia; Refractory Hodgkin Lymphoma; Refractory Plasma Cell Myeloma; Refractory Small Lymphocytic Lymphoma; Waldenstrom Macroglobulinemia

  10. Interferon-α Revisited: Individualized Treatment Management Eased the Selective Pressure of Tyrosine Kinase Inhibitors on BCR-ABL1 Mutations Resulting in a Molecular Response in High-Risk CML Patients.

    Science.gov (United States)

    Polivkova, Vaclava; Rohon, Peter; Klamova, Hana; Cerna, Olga; Divoka, Martina; Curik, Nikola; Zach, Jan; Novak, Martin; Marinov, Iuri; Soverini, Simona; Faber, Edgar; Machova Polakova, Katerina

    2016-01-01

    Bone marrow transplantation or ponatinib treatment are currently recommended strategies for management of patients with chronic myeloid leukemia (CML) harboring the T315I mutation and compound or polyclonal mutations. However, in some individual cases, these treatment scenarios cannot be applied. We used an alternative treatment strategy with interferon-α (IFN-α) given solo, sequentially or together with TKI in a group of 6 cases of high risk CML patients, assuming that the TKI-independent mechanism of action may lead to mutant clone repression. IFN-α based individualized therapy decreases of T315I or compound mutations to undetectable levels as assessed by next-generation deep sequencing, which was associated with a molecular response in 4/6 patients. Based on the observed results from immune profiling, we assumed that the principal mechanism leading to the success of the treatment was the immune activation induced with dasatinib pre-treatment followed by restoration of immunological surveillance after application of IFN-α therapy. Moreover, we showed that sensitive measurement of mutated BCR-ABL1 transcript levels augments the safety of this individualized treatment strategy. PMID:27214026

  11. Interferon-α Revisited: Individualized Treatment Management Eased the Selective Pressure of Tyrosine Kinase Inhibitors on BCR-ABL1 Mutations Resulting in a Molecular Response in High-Risk CML Patients.

    Directory of Open Access Journals (Sweden)

    Vaclava Polivkova

    Full Text Available Bone marrow transplantation or ponatinib treatment are currently recommended strategies for management of patients with chronic myeloid leukemia (CML harboring the T315I mutation and compound or polyclonal mutations. However, in some individual cases, these treatment scenarios cannot be applied. We used an alternative treatment strategy with interferon-α (IFN-α given solo, sequentially or together with TKI in a group of 6 cases of high risk CML patients, assuming that the TKI-independent mechanism of action may lead to mutant clone repression. IFN-α based individualized therapy decreases of T315I or compound mutations to undetectable levels as assessed by next-generation deep sequencing, which was associated with a molecular response in 4/6 patients. Based on the observed results from immune profiling, we assumed that the principal mechanism leading to the success of the treatment was the immune activation induced with dasatinib pre-treatment followed by restoration of immunological surveillance after application of IFN-α therapy. Moreover, we showed that sensitive measurement of mutated BCR-ABL1 transcript levels augments the safety of this individualized treatment strategy.

  12. Interferon-α Revisited: Individualized Treatment Management Eased the Selective Pressure of Tyrosine Kinase Inhibitors on BCR-ABL1 Mutations Resulting in a Molecular Response in High-Risk CML Patients

    Science.gov (United States)

    Polivkova, Vaclava; Rohon, Peter; Klamova, Hana; Cerna, Olga; Divoka, Martina; Curik, Nikola; Zach, Jan; Novak, Martin; Marinov, Iuri; Soverini, Simona; Faber, Edgar; Machova Polakova, Katerina

    2016-01-01

    Bone marrow transplantation or ponatinib treatment are currently recommended strategies for management of patients with chronic myeloid leukemia (CML) harboring the T315I mutation and compound or polyclonal mutations. However, in some individual cases, these treatment scenarios cannot be applied. We used an alternative treatment strategy with interferon-α (IFN-α) given solo, sequentially or together with TKI in a group of 6 cases of high risk CML patients, assuming that the TKI-independent mechanism of action may lead to mutant clone repression. IFN-α based individualized therapy decreases of T315I or compound mutations to undetectable levels as assessed by next-generation deep sequencing, which was associated with a molecular response in 4/6 patients. Based on the observed results from immune profiling, we assumed that the principal mechanism leading to the success of the treatment was the immune activation induced with dasatinib pre-treatment followed by restoration of immunological surveillance after application of IFN-α therapy. Moreover, we showed that sensitive measurement of mutated BCR-ABL1 transcript levels augments the safety of this individualized treatment strategy. PMID:27214026

  13. Nilotinib (formerly AMN107), a highly selective BCR-ABL tyrosine kinase inhibitor, is active in patients with imatinib-resistant or -intolerant accelerated-phase chronic myelogenous leukemia.

    Science.gov (United States)

    le Coutre, Philipp; Ottmann, Oliver G; Giles, Francis; Kim, Dong-Wook; Cortes, Jorge; Gattermann, Norbert; Apperley, Jane F; Larson, Richard A; Abruzzese, Elisabetta; O'Brien, Stephen G; Kuliczkowski, Kazimierz; Hochhaus, Andreas; Mahon, Francois-Xavier; Saglio, Giuseppe; Gobbi, Marco; Kwong, Yok-Lam; Baccarani, Michele; Hughes, Timothy; Martinelli, Giovanni; Radich, Jerald P; Zheng, Ming; Shou, Yaping; Kantarjian, Hagop

    2008-02-15

    Patients with imatinib-resistant or -intolerant accelerated-phase chronic myelogenous leukemia (CML-AP) have very limited therapeutic options. Nilotinib is a highly selective BCR-ABL tyrosine kinase inhibitor. This phase 2 trial was designed to characterize the efficacy and safety of nilotinib (400 mg twice daily) in this patient population with hematologic response (HR) as primary efficacy endpoint. A total of 119 patients were enrolled and had a median duration of treatment of 202 days (range, 2-611 days). An HR was observed in 56 patients (47%; 95% confidence interval [CI], 38%-56%). Major cytogenetic response (MCyR) was observed in 35 patients (29%; 95% CI, 21%-39%). The median duration of HR has not been reached. Overall survival rate among the 119 patients after 12 months of follow-up was 79% (95% CI, 70%-87%). Nonhematologic adverse events were mostly mild to moderate. Severe peripheral edema and pleural effusions were not observed. The most common grade 3 or higher hematologic adverse events were thrombocytopenia (35%) and neutropenia (21%). Grade 3 or higher bilirubin and lipase elevations occurred in 9% and 18% of patients, respectively, resulting in treatment discontinuation in one patient. In conclusion, nilotinib is an effective and well-tolerated treatment in imatinib-resistant and -intolerant CML-AP. This trial is registered at www.clinicaltrials.gov as NCT00384228. PMID:18048643

  14. Estandarización de la TR-PCR para la detección de las fusiones génicas BCR-ABL en pacientes con leucemia Mieloide Crónica (LMC y Linfoide Aguda (LLA provenientes de HUSVP y Clíncia León XIII

    Directory of Open Access Journals (Sweden)

    Gonzálo Vásquez Palacio

    2006-04-01

    Full Text Available La translocación recíproca t(9:22(q34;q11 involucra el proto-oncogen ABL y el gen BCR, originando un gen de fusión BCR-ABL, que codifica una proteína con elevada actividad tirosina quinasa, implicada en la leucemogénesis.

  15. Estandarización de la TR-PCR para la detección de las fusiones génicas BCR-ABL en pacientes con leucemia Mieloide Crónica (LMC) y Linfoide Aguda (LLA) provenientes de HUSVP y Clíncia León XIII

    OpenAIRE

    Gonzálo Vásquez Palacio; José Domingo Torres Hernández; Francisco Cuéllar Ambrossi; Carlos Enrique Muskus; Carlos Alberto Paya

    2006-01-01

    La translocación recíproca t(9:22)(q34;q11) involucra el proto-oncogen ABL y el gen BCR, originando un gen de fusión BCR-ABL, que codifica una proteína con elevada actividad tirosina quinasa, implicada en la leucemogénesis.

  16. Rapid identification of compound mutations in patients with Ph-positive leukemias by long-range next generation sequencing

    Science.gov (United States)

    Kastner, R.; Zopf, A.; Preuner, S.; Pröll, J.; Niklas, N.; Foskett, P.; Valent, P.; Lion, T.; Gabriel, C.

    2016-01-01

    An emerging problem in patients with Ph-positive leukemias is the occurrence of cells with multiple mutations in the BCR-ABL1 tyrosine kinase domain (TKD) associated with high resistance to different tyrosine kinase inhibitors. Rapid and sensitive detection of leukemic subclones carrying such changes, referred to as compound mutations, is therefore of increasing clinical relevance. However, current diagnostic methods including next generation sequencing (NGS) of short fragments do not optimally meet these requirements. We have therefore established a long-range (LR) NGS approach permitting massively parallel sequencing of the entire TKD length of 933bp in a single read using 454 sequencing with the GS FLX+ instrument (454 Life Sciences). By testing a series of individual and consecutive specimens derived from six patients with chronic myeloid leukemia, we demonstrate that long-range NGS analysis permits sensitive identification of mutations and their assignment to the same or to separate subclones. This approach also facilitates readily interpretable documentation of insertions and deletions in the entire BCR-ABL1 TKD. The long-range NGS findings were reevaluated by an independent technical approach in select cases. PCR amplicons of the BCR-ABL1 TKD derived from individual specimens were subcloned into pGEM®-T plasmids, and >100 individual clones were subjected to analysis by Sanger sequencing. The NGS results were confirmed, thus documenting the reliability of the new technology. Long-range NGS analysis therefore provides an economic approach to the identification of compound mutations and other genetic alterations in the entire BCR-ABL1 TKD, and represents an important advancement of the diagnostic armamentarium for rapid assessment of impending resistant disease. PMID:24365090

  17. ATG7 regulates energy metabolism, differentiation and survival of Philadelphia-chromosome-positive cells.

    Science.gov (United States)

    Karvela, Maria; Baquero, Pablo; Kuntz, Elodie M; Mukhopadhyay, Arunima; Mitchell, Rebecca; Allan, Elaine K; Chan, Edmond; Kranc, Kamil R; Calabretta, Bruno; Salomoni, Paolo; Gottlieb, Eyal; Holyoake, Tessa L; Helgason, G Vignir

    2016-06-01

    A major drawback of tyrosine kinase inhibitor (TKI) treatment in chronic myeloid leukemia (CML) is that primitive CML cells are able to survive TKI-mediated BCR-ABL inhibition, leading to disease persistence in patients. Investigation of strategies aiming to inhibit alternative survival pathways in CML is therefore critical. We have previously shown that a nonspecific pharmacological inhibition of autophagy potentiates TKI-induced death in Philadelphia chromosome-positive cells. Here we provide further understanding of how specific and pharmacological autophagy inhibition affects nonmitochondrial and mitochondrial energy metabolism and reactive oxygen species (ROS)-mediated differentiation of CML cells and highlight ATG7 (a critical component of the LC3 conjugation system) as a potential specific therapeutic target. By combining extra- and intracellular steady state metabolite measurements by liquid chromatography-mass spectrometry with metabolic flux assays using labeled glucose and functional assays, we demonstrate that knockdown of ATG7 results in decreased glycolysis and increased flux of labeled carbons through the mitochondrial tricarboxylic acid cycle. This leads to increased oxidative phosphorylation and mitochondrial ROS accumulation. Furthermore, following ROS accumulation, CML cells, including primary CML CD34(+) progenitor cells, differentiate toward the erythroid lineage. Finally, ATG7 knockdown sensitizes CML progenitor cells to TKI-induced death, without affecting survival of normal cells, suggesting that specific inhibitors of ATG7 in combination with TKI would provide a novel therapeutic approach for CML patients exhibiting persistent disease. PMID:27168493

  18. Effective Concentration of a Multikinase Inhibitor within Bone Marrow Correlates with In Vitro Cell Killing in Therapy-Resistant Chronic Myeloid Leukemia.

    Science.gov (United States)

    Mu, Chaofeng; Wu, Xiaoyan; Ma, Helen; Tao, Wenjing; Zhang, Guodong; Xia, Xiaojun; Shen, Jianliang; Mai, Junhua; Sun, Tong; Sun, Xiaoping; Arlinghaus, Ralph B; Shen, Haifa

    2016-05-01

    Leukemia cells escape BCR-ABL-targeted therapy by developing mutations, such as T315I, in the p210(BCR-ABL) fusion protein in Philadelphia chromosome-positive chronic myeloid leukemia (CML). Although most effort has been focused on development of new tyrosine kinase inhibitors, enrichment of these small-molecule inhibitors in the tumor tissue can also have a profound impact on treatment outcomes. Here, we report that a 2-hour exposure of the T315I-mutant CML cells to 10 μmol/L of the multikinase inhibitor TG101209 suppressed BCR-ABL-independent signaling and caused cell-cycle arrest at G2-M. Further increase in drug concentration to 17.5 μmol/L blocked phosphorylation of the mutant BCR-ABL kinase and its downstream JAK2 and STAT5. The effective dosage to overcome therapy resistance identified in an in vitro setting serves as a guidance to develop the proper drug formulation for in vivo efficacy. A targeted formulation was developed to achieve sustained bone marrow TG101209 concentration at or above 17.5 μmol/L for effective killing of CML cells in vivo Potent inhibition of leukemia cell growth and extended survival were observed in two murine models of CML treated with 40 mg/kg intravenously administered targeted TG101209, but not with the untargeted drug at the same dosage. Our finding provides a unique approach to develop treatments for therapy-resistant CML. Mol Cancer Ther; 15(5); 899-910. ©2016 AACR. PMID:26846820

  19. Selective Depletion of CD45RA+ T Cells From Allogeneic Peripheral Blood Stem Cell Grafts in Preventing GVHD in Children

    Science.gov (United States)

    2016-06-21

    Accelerated Phase Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Acute Biphenotypic Leukemia; Acute Leukemia of Ambiguous Lineage; Acute Myeloid Leukemia in Remission; Acute Undifferentiated Leukemia; Adult Acute Lymphoblastic Leukemia in Remission; Blast Phase Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Childhood Acute Lymphoblastic Leukemia in Remission; Chronic Myelogenous Leukemia, BCR-ABL1 Positive; RAEB-1; RAEB-2; Recurrent Adult Acute Myeloid Leukemia; Recurrent Childhood Acute Lymphoblastic Leukemia; Recurrent Childhood Acute Myeloid Leukemia; Refractory Adult Acute Lymphoblastic Leukemia; Refractory Childhood Acute Lymphoblastic Leukemia

  20. Open Label, Phase II Study to Evaluate Efficacy and Safety of Oral Nilotinib in Philadelphia Positive (Ph+) Chronic Myelogenous Leukemia (CML) Pediatric Patients.

    Science.gov (United States)

    2016-08-05

    Leukemia; Leukemia,Pediatric; Leukemia, Myleiod; Leukemia, Mylegenous, Chronic; Leukemia, Mylegenous, Accelerated; BCR-ABL Positive; Myeloproliferative Disorder; Bone Marrow Disease; Hematologic Diseases; Neoplastic Processes; Imatinib; Dasatinib; Enzyme Inhibitor; Protein Kinase Inhibitor

  1. Sirolimus, Cyclosporine, and Mycophenolate Mofetil in Preventing Graft-versus-Host Disease in Treating Patients With Hematologic Malignancies Undergoing Donor Peripheral Blood Stem Cell Transplant

    Science.gov (United States)

    2016-03-09

    Adult Acute Lymphoblastic Leukemia; Adult Acute Myeloid Leukemia; Adult Diffuse Large B-Cell Lymphoma; Adult Myelodysplastic Syndrome; Adult Non-Hodgkin Lymphoma; Aggressive Non-Hodgkin Lymphoma; Childhood Acute Lymphoblastic Leukemia; Childhood Acute Myeloid Leukemia; Childhood Diffuse Large B -Cell Lymphoma; Childhood Myelodysplastic Syndrome; Childhood Non-Hodgkin Lymphoma; Chronic Lymphocytic Leukemia; Chronic Lymphocytic Leukemia in Remission; Chronic Phase Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Hematopoietic and Lymphoid Cell Neoplasm; Mantle Cell Lymphoma; Plasma Cell Myeloma; Prolymphocytic Leukemia; Recurrent Chronic Lymphocytic Leukemia; Recurrent Diffuse Large B-Cell Lymphoma; Recurrent Hodgkin Lymphoma; Refractory Chronic Lymphocytic Leukemia; T-Cell Chronic Lymphocytic Leukemia; T-Cell Prolymphocytic Leukemia; Waldenstrom Macroglobulinemia

  2. A combination of STI571 and BCR-ABL1 siRNA with overexpressed p15INK4B induced enhanced proliferation inhibition and apoptosis in chronic myeloid leukemia

    OpenAIRE

    Xia, D.Y.; Liu, L; Hao, M.W.; Liu, Q; Chen, R A; Y.M. Liang

    2014-01-01

    p15INK4B, a cyclin-dependent kinase inhibitor, has been recognized as a tumor suppressor. Loss of or methylation of the p15INK4B gene in chronic myeloid leukemia (CML) cells enhances myeloid progenitor formation from common myeloid progenitors. Therefore, we examined the effects of overexpressed p15INK4B on proliferation and apoptosis of CML cells. Overexpression of p15INK4B inhibited the growth of K562 cells by downregulation of cyclin-dependent kinase 4 (CDK4) and cyclin D1 expression. Over...

  3. Signal transduction factors on the modulation of radiosusceptibility in K562 cells

    International Nuclear Information System (INIS)

    The human chronic myelogenous leukemia cell line, K562, expresses the chimeric bcr-abl oncoprotein, whose deregulated protein tyrosine kinase activity antagonizes the induction of apoptosis via DNA damaging agents. Previous experiments have shown that nanomolar concentrations of herbimycin A [HMA] coupled with X-irradiation have a synergistic effect in inducing apoptosis in the Ph-positive K562 leukemia cell line, but genistein, a PTK inhibitor, is non selective for the radiation-induced apoptosis of p210bcr/abl protected K562 cells. In these experiments, the cytoplasmic signal transduction pathways, the induction of a number of transcription factors and the differential gene expression in this model were investigated. K562 cells in the exponential growth phase were used in this study. The cells were irradiated with 0.5-12 Gy, using a 6 MeV Linac (Clinac 1800, Varian, USA). Immediately after irradiation, the cells were treated with 0.25μM of HMA and 25μM of genistein, and the expressions and the activities of ablkinase, MAPK family, NF-KB, c-fos, c-myc, and thymidine kinase1 (TK1) were examined. The differential gene expressions induced by PTK inhibitors were also investigated. The modulating effects of herbimycin A and genistein on the radiosensitivity of K562 cells were not related to the bcr-abl kinase activity. The signaling responses through the MAPK family of proteins, were not involved either. In association with the radiation-induced apoptosis, which is accelerated by HMA, the expression of c-myc was increased. The combined treatment of genistein, with irradiation, enhanced NF-KB activity and the TK 1 expression and activity. The effects of HMA and genistein on the radiosensitivity of the K562 cells were not related to the bcr-abl kinase activity. In this study, another signaling pathway, besides the MAPK family responses to radiation to K562 cells, was found. Further evaluation using this model will provide valuable information for the optional

  4. Dasatinib inhibits proliferation and activation of CD8+ T-lymphocytes by down-regulation of the T-cell receptor signaling

    OpenAIRE

    Fei, Fei

    2007-01-01

    The novel Src/Abl inhibitor dasatinib (BMS-354825) is a promising therapeutic agent with imatinib-resistant BCR-ABL mutations in patients with chronic myelogenous leukemia (CML) or Ph-positive acute lymphoblastic leukemia (ALL). However, little is known about its effects on T-cell function, especially for patients undergoing allogeneic transplantation for leukemia. Here, we demonstrate that dasatinib at a concentration of 5 nM to 250 nM inhibits the proliferation and activation of CD8+ T-lymp...

  5. Tacrolimus and Mycophenolate Mofetil With or Without Sirolimus in Preventing Acute Graft-Versus-Host Disease in Patients Who Are Undergoing Donor Stem Cell Transplant for Hematologic Cancer

    Science.gov (United States)

    2015-10-14

    Myelodysplastic/Myeloproliferative Neoplasm, Unclassifiable; Previously Treated Myelodysplastic Syndrome; Refractory Chronic Lymphocytic Leukemia; Refractory Plasma Cell Myeloma; Waldenstrom Macroglobulinemia; Accelerated Phase Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Adult Acute Lymphoblastic Leukemia in Remission; Adult Acute Myeloid Leukemia in Remission; Adult Acute Myeloid Leukemia With t(9;11)(p22;q23); MLLT3-; Adult Acute Myeloid Leukemia With Inv(16)(p13.1q22); CBFB-MYH11; Adult Acute Promyelocytic Leukemia With t(15;17)(q22;q12); PML-RARA; Adult Acute Myeloid Leukemia With t(8;21)(q22;q22); RUNX1-RUNX1T1; Atypical Chronic Myeloid Leukemia, BCR-ABL1 Negative; Blast Phase Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Childhood Acute Lymphoblastic Leukemia in Remission; Childhood Acute Myeloid Leukemia in Remission; Childhood Burkitt Lymphoma; Childhood Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Childhood Diffuse Large Cell Lymphoma; Childhood Immunoblastic Lymphoma; Childhood Myelodysplastic Syndrome; Stage II Contiguous Adult Burkitt Lymphoma; Stage II Contiguous Adult Diffuse Large Cell Lymphoma; Stage II Contiguous Adult Diffuse Mixed Cell Lymphoma; Stage II Contiguous Adult Diffuse Small Cleaved Cell Lymphoma; Stage II Adult Contiguous Immunoblastic Lymphoma; Stage II Contiguous Adult Lymphoblastic Lymphoma; Stage II Grade 1 Contiguous Follicular Lymphoma; Stage II Grade 2 Contiguous Follicular Lymphoma; Stage II Grade 3 Contiguous Follicular Lymphoma; Stage II Contiguous Mantle Cell Lymphoma; Stage II Non-Contiguous Adult Burkitt Lymphoma; Stage II Non-Contiguous Adult Diffuse Large Cell Lymphoma; Stage II Non-Contiguous Adult Diffuse Mixed Cell Lymphoma; Stage II Non-Contiguous Adult Diffuse Small Cleaved Cell Lymphoma; Stage II Adult Non-Contiguous Immunoblastic Lymphoma; Stage II Non-Contiguous Adult Lymphoblastic Lymphoma; Stage II Grade 1 Non-Contiguous Follicular Lymphoma; Stage II Grade 2 Non-Contiguous Follicular Lymphoma; Stage

  6. Sustaining integrating imatinib and interferon-α into maintenance therapy improves survival of patients with Philadelphia positive acute lymphoblastic leukemia ineligible for allogeneic stem cell transplantation.

    Science.gov (United States)

    Kuang, Pu; Liu, Ting; Pan, Ling; Zhu, Huanling; Wu, Yu; Ye, Yuanxin; Xiang, Bing; Ma, Hongbing; Chang, Hong; Niu, Ting; Cui, Xu; He, Chuan; Li, Jianjun; Ji, Jie; Huang, Jie; Dong, Tian; Dai, Yang; Lu, Xiaojun; Qing, Shenglan; Wu, Huaxin; Liang, Xiaogong; Wang, Xiaoyu; Wu, Chunnong

    2016-10-01

    We report the clinical results of sustainedly integrating imatinib and interferon-α into maintenance therapy in the patients ineligible for allogeneic hematopoietic stem cell transplantation (allo-HSCT). Maintenance therapy lasted for 5 years with imatinib 400 mg daily, interferon-α 3 million units, 2∼3 doses per week, and chemotherapy including vindesine and dexamethasone scheduled monthly in first year, once every 2 months in second year, and once every 3 months in third year. The chemotherapy was discontinued after 3 years and the imatinib and interferon-α continued for another 2 years. For 41 patients without allo-HSCT with a median follow-up of 32 months, the 3-year DFS and OS were 42.7  ± 8.6% and 57.9  ± 8.4%, respectively. Our study suggests that sustaining maintenance with low-dose chemotherapy, imatinib and interferon-α improved survival of adult Philadelphia-positive acute lymphoblastic leukemia (Ph + ALL) patients ineligible for allo-HSCT, and even provided an opportunity for cure. BCR/ABL persistent negativity at 6 and 9 months may have benefit to choose suitable patients for the imatinib/interferon-α maintenance strategy. PMID:26879808

  7. Efficacy of ponatinib against ABL tyrosine kinase inhibitor-resistant leukemia cells

    Energy Technology Data Exchange (ETDEWEB)

    Okabe, Seiichi, E-mail: okabe@tokyo-med.ac.jp; Tauchi, Tetsuzo; Tanaka, Yuko; Ohyashiki, Kazuma

    2013-06-07

    Highlights: •Efficacy of ponatinib against ABL tyrosine kinase inhibitor-resistant leukemia cells okabe et al. •Imatinib or nilotinib resistance was involved Src family kinase. •The BCR-ABL point mutation (E334V) was highly resistant to imatinib or nilotinib. •Ponatinib was a powerful strategy against imatinib or nilotinib resistant Ph-positive cells. -- Abstract: Because a substantial number of patients with chronic myeloid leukemia acquire resistance to ABL tyrosine kinase inhibitors (TKIs), their management remains a challenge. Ponatinib, also known as AP24534, is an oral multi-targeted TKI. Ponatinib is currently being investigated in a pivotal phase 2 clinical trial. In the present study, we analyzed the molecular and functional consequences of ponatinib against imatinib- or nilotinib-resistant (R) K562 and Ba/F3 cells. The proliferation of imatinib- or nilotinib-resistant K562 cells did not decrease after treatment with imatinib or nilotinib. Src family kinase Lyn was activated. Point mutation Ba/F3 cells (E334 V) were also highly resistant to imatinib and nilotinib. Treatment with ponatinib for 72 h inhibited the growth of imatinib- and nilotinib-resistant cells. The phosphorylation of BCR-ABL, Lyn, and Crk-L was reduced. This study demonstrates that ponatinib has an anti-leukemia effect by reducing ABL and Lyn kinase activity and this information may be of therapeutic relevance.

  8. Quantitative Proteomics Analysis of Leukemia Cells.

    Science.gov (United States)

    Halbach, Sebastian; Dengjel, Jörn; Brummer, Tilman

    2016-01-01

    Chronic myeloid leukemia (CML) is driven by the oncogenic fusion kinase Bcr-Abl, which organizes its own signaling network with various proteins. These proteins, their interactions, and their role in relevant signaling pathways can be analyzed by quantitative mass spectrometry (MS) approaches in various models systems, e.g., in cell culture models. In this chapter, we describe in detail immunoprecipitations and quantitative proteomics analysis using stable isotope labeling by amino acids in cell culture (SILAC) of components of the Bcr-Abl signaling pathway in the human CML cell line K562. PMID:27581145

  9. An Adult Male Presenting with Concurrent Plasma Cell Myeloma Involving a CCND1-IGH Translocation and Chronic Myelogenous Leukemia with a Variant (9;22) Translocation.

    Science.gov (United States)

    Lee, Peter M; Siangchin, Ken; Song, Sophie; Shabsovich, David; Naeini, Yalda; Tirado, Carlos A

    2016-01-01

    The t(11;14)(q13;q32) involving IGH and CCND1 a nd t(9;22) (q34;q11.2) involving BCR and ABL1 are common abnormalities in plasma cell myeloma (PCM) and chronic myelogenous leukemia (CML), respectively. However, the concurrence of the two malignancies is extremely rare. Herein, we present a case of an 87-year-old male who presented with anemia and monocytosis. FISH studies on a bone marrow sample enriched for plasma cells detected a t(11;14) positive for IGH and CCND1 fusion in 92% of nuclei. However, cytogenetic analysis of the bone marrow revealed a t(9;22)(q34;q11.2) in 40% of the metaphases. Interphase and metaphase FISH studies on the sample confirmed the presence of the BCR-ABL1 fusion in 88% of nuclei but did not show any signals corresponding to the derivative 9, suggesting a variant t(9;22) with a deletion or additional material of unknown origin at the 9q34 band of the derivative 9 and a derivative 22 bearing the BCR-ABL1 fusion gene. The concurrence of plasma cell myeloma and chronic myelogenous leukemia is extremely rare with less than 20 cases reported. The molecular pathway in which the multiple malignancies arise is still poorly understood, and this case provides insight into the concurrence of PCM and CML. PMID:27584682

  10. Modeling chronic myeloid leukemia in immunodeficient mice reveals expansion of aberrant mast cells and accumulation of pre-B cells

    International Nuclear Information System (INIS)

    Chronic myeloid leukemia (CML) is a myeloproliferative neoplasm that, if not treated, will progress into blast crisis (BC) of either myeloid or B lymphoid phenotype. The BCR-ABL1 fusion gene, encoding a constitutively active tyrosine kinase, is thought to be sufficient to cause chronic phase (CP) CML, whereas additional genetic lesions are needed for progression into CML BC. To generate a humanized CML model, we retrovirally expressed BCR-ABL1 in the cord blood CD34+ cells and transplanted these into NOD-SCID (non-obese diabetic/severe-combined immunodeficient) interleukin-2-receptor γ-deficient mice. In primary mice, BCR-ABL1 expression induced an inflammatory-like state in the bone marrow and spleen, and mast cells were the only myeloid lineage specifically expanded by BCR-ABL1. Upon secondary transplantation, the pronounced inflammatory phenotype was lost and mainly human mast cells and macrophages were found in the bone marrow. Moreover, a striking block at the pre-B-cell stage was observed in primary mice, resulting in an accumulation of pre-B cells. A similar block in B-cell differentiation could be confirmed in primary cells from CML patients. Hence, this humanized mouse model of CML reveals previously unexplored features of CP CML and should be useful for further studies to understand the disease pathogenesis of CML

  11. 16-Hydroxycleroda-3,13-dien-15,16-olide deregulates PI3K and Aurora B activities that involve in cancer cell apoptosis

    International Nuclear Information System (INIS)

    Highlights: → Natural product PL3 induced cancer cell apoptosis and cell cycle arrest. → PL3 targeted and de-regulated PI3K pathway activities. → De-activated PI3K pathway caused Aurora B degradation and spindle dysfunction. → PL3 could reverse the drug-resistance of Bcr-ABL T315I-positive cells to Imatinib. -- Abstract: The PI3K-AKT pathway and Aurora kinase play essential roles in such cellular processes as cell survival, angiogenesis, and differentiation, and are usually expressed at maximum levels during cancer cell proliferation. The present study investigated the effect of the natural compound, 16-hydroxycleroda-3,13-dien-15,16-olide (PL3), on regulating the PI3K-AKT pathway and Aurora B, which led to cancer cell apoptosis. PL3 acts as a PI3K inhibitor by influencing cell survival, signaling transduction, and cell cycle progression. It was observed that PL3 targeted and induced dephosphorylation of the PI3K pathway, degradation of Aurora B and mitotic-related gene expressions, and sequentially shut down the cell cycle. This eventually resulted in cell death. As Aurora B was downregulated, spindle dysfunction and destruction of the G2/M phase checkpoint resulted in DNA-damaged cells undergoing apoptosis. Moreover, PL3 also resensitized T315I-mutated Bcr-ABL+ BA/F3 cells to improve the cytotoxicity of Imatinib in Imatinib-resistant cell line. Taken together, PL3 can perturb the PI3K-AKT pathway and Aurora B resulting in gene silencing and cell cycle disturbance. It was demonstrated that PL3 acted like a novel small-molecule PI3K modulator, thereby potentially contributing to cancer chemotherapy and combination medication.

  12. Ipilimumab After Allogeneic Stem Cell Transplant in Treating Patients With Persistent or Progressive Cancer

    Science.gov (United States)

    2013-03-26

    Adult Acute Myeloid Leukemia With 11q23 (MLL) Abnormalities; Adult Acute Myeloid Leukemia With Inv(16)(p13;q22); Adult Acute Myeloid Leukemia With t(15;17)(q22;q12); Adult Acute Myeloid Leukemia With t(16;16)(p13;q22); Adult Acute Myeloid Leukemia With t(8;21)(q22;q22); Atypical Chronic Myeloid Leukemia, BCR-ABL1 Negative; Childhood Myelodysplastic Syndromes; Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Disseminated Neuroblastoma; Malignant Neoplasm; Ovarian Choriocarcinoma; Ovarian Embryonal Carcinoma; Ovarian Immature Teratoma; Ovarian Mature Teratoma; Ovarian Mixed Germ Cell Tumor; Ovarian Monodermal and Highly Specialized Teratoma; Ovarian Polyembryoma; Ovarian Yolk Sac Tumor; Previously Treated Myelodysplastic Syndromes; Recurrent Adult Acute Lymphoblastic Leukemia; Recurrent Adult Acute Myeloid Leukemia; Recurrent Adult Burkitt Lymphoma; Recurrent Adult Diffuse Large Cell Lymphoma; Recurrent Adult Diffuse Mixed Cell Lymphoma; Recurrent Adult Hodgkin Lymphoma; Recurrent Adult Immunoblastic Large Cell Lymphoma; Recurrent Adult Lymphoblastic Lymphoma; Recurrent Grade 3 Follicular Lymphoma; Recurrent Malignant Testicular Germ Cell Tumor; Recurrent Mantle Cell Lymphoma; Recurrent Neuroblastoma; Recurrent Ovarian Epithelial Cancer; Recurrent Ovarian Germ Cell Tumor; Refractory Chronic Lymphocytic Leukemia; Refractory Multiple Myeloma; Relapsing Chronic Myelogenous Leukemia; Stage I Multiple Myeloma; Stage II Multiple Myeloma; Stage II Ovarian Epithelial Cancer; Stage III Malignant Testicular Germ Cell Tumor; Stage III Multiple Myeloma; Stage III Ovarian Epithelial Cancer; Stage IIIA Breast Cancer; Stage IIIB Breast Cancer; Stage IIIC Breast Cancer; Stage IV Breast Cancer; Stage IV Ovarian Epithelial Cancer; Testicular Choriocarcinoma; Testicular Choriocarcinoma and Embryonal Carcinoma; Testicular Choriocarcinoma and Seminoma; Testicular Choriocarcinoma and Teratoma; Testicular Choriocarcinoma and Yolk Sac Tumor; Testicular Embryonal Carcinoma; Testicular

  13. Fludarabine Phosphate and Total-Body Irradiation Followed by Donor Peripheral Blood Stem Cell Transplant in Treating Patients With Acute Lymphoblastic Leukemia or Chronic Myelogenous Leukemia That Has Responded to Treatment With Imatinib Mesylate, Dasatinib, or Nilotinib

    Science.gov (United States)

    2015-07-20

    Adult Acute Lymphoblastic Leukemia in Remission; Blastic Phase Chronic Myelogenous Leukemia; Childhood Acute Lymphoblastic Leukemia in Remission; Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Chronic Phase Chronic Myelogenous Leukemia; Philadelphia Chromosome Positive Adult Precursor Acute Lymphoblastic Leukemia; Philadelphia Chromosome Positive Childhood Precursor Acute Lymphoblastic Leukemia; Recurrent Adult Acute Lymphoblastic Leukemia; Recurrent Childhood Acute Lymphoblastic Leukemia; Relapsing Chronic Myelogenous Leukemia

  14. Novel Acylguanidine Derivatives Targeting Smoothened Induce Antiproliferative and Pro-Apoptotic Effects in Chronic Myeloid Leukemia Cells

    Science.gov (United States)

    Chiarenza, Alessandra; Manetti, Fabrizio; Petricci, Elena; Ruat, Martial; Naldini, Antonella; Taddei, Maurizio; Carraro, Fabio

    2016-01-01

    The most relevant therapeutic approaches to treat CML rely on the administration of tyrosine kinase inhibitors (TKIs) like Imatinib, which are able to counteract the activity of Bcr-Abl protein increasing patient’s life expectancy and survival. Unfortunately, there are some issues TKIs are not able to address; first of all TKIs are not so effective in increasing survival of patients in blast crisis, second they are not able to eradicate leukemic stem cells (LSC) which represent the major cause of disease relapse, and third patients often develop resistance to TKIs due to mutations in the drug binding site. For all these reasons it’s of primary interest to find alternative strategies to treat CML. Literature shows that Hedgehog signaling pathway is involved in LSC maintenance, and pharmacological inhibition of Smoothened (SMO), one of the key molecules of the pathway, has been demonstrated to reduce Bcr-Abl positive bone marrow cells and LSC. Consequently, targeting SMO could be a promising way to develop a new treatment strategy for CML overcoming the limitations of current therapies. In our work we have tested some compounds able to inhibit SMO, and among them MRT92 appears to be a very potent SMO antagonist. We found that almost all our compounds were able to reduce Gli1 protein levels in K-562 and in KU-812 CML cell lines. Furthermore, they were also able to increase Gli1 and SMO RNA levels, and to reduce cell proliferation and induce apoptosis/autophagy in both the tested cell lines. Finally, we demonstrated that our compounds were able to modulate the expression of some miRNAs related to Hedgehog pathway such as miR-324-5p and miR-326. Being Hedgehog pathway deeply implicated in the mechanisms of CML we may conclude that it could be a good therapeutic target for CML and our compounds seem to be promising antagonists of such pathway. PMID:26934052

  15. Tyrosine kinase inhibitors influence ABCG2 expression in EGFR-positive MDCK BCRP cells via the PI3K/Akt signaling pathway.

    Science.gov (United States)

    Pick, Anne; Wiese, Michael

    2012-04-01

    Multidrug resistance observed in cancer chemotherapy is commonly attributed to overexpression of efflux transporter proteins. These proteins act as ATP-dependent drug efflux pumps, actively extruding chemotherapeutic agents from cells and causing a decrease in intracellular drug accumulation. Besides the well-recognized role of P-glycoprotein (P-gp, ABCB1), the breast cancer resistance protein (BCRP, ABCG2) is becoming increasingly accepted as playing an important role in multidrug resistance. In contrast to P-glycoprotein, only a few inhibitors of ABCG2 are known. According to the literature, tyrosine kinase inhibitors (TKIs) can be considered to be broad-spectrum inhibitors, interacting with ABCB1, ABCC1 and ABCG2. Here, we investigated seven different TKIs, gefitinib, erlotinib, AG1478, PD158780, PD153035, nilotinib and imatinib, for their potential to restore ABCG2 sensitivity to cells. Furthermore, we analyzed the alteration of ABCG2 expression caused by TKIs and demonstrated that EGFR inhibitors such as gefitinib and PD158780 reduced both total and surface expression of ABCG2 in EGRF-positive MDCK BCRP cells by interaction with the PI3K/Akt signaling pathway. The reduced ABCG2 content led to an increased effect of XR9577, a well-known ABCG2 modulator, lowering the concentration required for half maximal inhibition. On the other hand, BCR-ABL inhibitors had no influence on ABCG2 expression and modulator activity. Interestingly, a combination of an EGFR inhibitor with the PI3K/Akt inhibitor LY294002 led to a significant reduction of ABCG2 expression at low concentrations of the drugs. Based on our results, we assume that EGFR exerts a post-transcriptional enhancing effect on ABCG2 expression via the PI3K/Akt signaling pathway, which can be attenuated by EGFR inhibitors. Blocking the key signaling pathway regulating ABCG2 expression with EGFR inhibitors, combined with the inhibition of ABCG2 with potent modulators might be a promising approach to circumvent MDR

  16. Donor Umbilical Cord Blood Transplant With or Without Ex-vivo Expanded Cord Blood Progenitor Cells in Treating Patients With Acute Myeloid Leukemia, Acute Lymphoblastic Leukemia, Chronic Myelogenous Leukemia, or Myelodysplastic Syndromes

    Science.gov (United States)

    2016-08-10

    Acute Biphenotypic Leukemia; Acute Lymphoblastic Leukemia in Remission; Acute Myeloid Leukemia Arising From Previous Myelodysplastic Syndrome; Acute Myeloid Leukemia in Remission; Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Mixed Phenotype Acute Leukemia; Myelodysplastic Syndrome; Pancytopenia; Refractory Anemia; Refractory Anemia With Excess Blasts; Refractory Anemia With Excess Blasts in Transformation; Secondary Acute Myeloid Leukemia

  17. Automated detection of residual cells after sex-mismatched stem-cell transplantation – evidence for presence of disease-marker negative residual cells

    Directory of Open Access Journals (Sweden)

    Johannes Tilman

    2009-05-01

    Full Text Available Abstract Background A new chimerism analysis based on automated interphase fluorescence in situ hybridization (FISH evaluation was established to detect residual cells after allogene sex-mismatched bone marrow or blood stem-cell transplantation. Cells of 58 patients were characterized as disease-associated due to presence of a bcr/abl-gene-fusion or a trisomy 8 and/or a simultaneous hybridization of gonosome-specific centromeric probes. The automatic slide scanning platform Metafer with its module MetaCyte was used to analyse 3,000 cells per sample. Results Overall 454 assays of 58 patients were analyzed. 13 of 58 patients showed residual recipient cells at one stage of more than 4% and 12 of 58 showed residual recipient cells less than 4%, respectively. As to be expected, patients of the latter group were associated with a higher survival rate (48 vs. 34 month. In only two of seven patients with disease-marker positive residual cells between 0.1–1.3% a relapse was observed. Besides, disease-marker negative residual cells were found in two patients without relapse at a rate of 2.8% and 3.3%, respectively. Conclusion The definite origin and meaning of disease-marker negative residual cells is still unclear. Overall, with the presented automatic chimerism analysis of interphase FISH slides, a sensitive method for detection of disease-marker positive residual cells is on hand.

  18. AUTOLOGOUS HEMATOPOIETIC STEM CELL TRANSPLANTATION FOR HIGH-RISK ACUTE LYMPHOBLASTIC LEUKEMIA: NON-RANDOMIZED STUDY WITH A MAXIMUM FOLLOW-UP OF MORE THAN 22 YEARS

    Directory of Open Access Journals (Sweden)

    Grzegorz Helbig

    2014-06-01

    Full Text Available Objective. To evaluate the efficacy and toxicity of autologous hematopoietic stem cell transplantation (AHSCT for high-risk acute lymphoblastic leukemia (ALL. Material and methods. Overall, 128 high-risk ALL patients at a median age of 26 years (range 18-56 years at diagnosis received AHSCT between 1991-2008. Induction treatment was anthracycline-based in all patients. Conditioning regimen consisted of CAV (cyclophosphamide, cytarabine, etoposide in 125 patients whereas 3 subjects received cyclophosphamide and TBI (total body irridation. Bone marrow was stored for 72 hours in 4oC and re-infused 24 hours after conditioning completion. Bone marrow was a source of stem cells in 119 patients, peripheral blood in 2 and 7 subjects received both bone marrow and peripheral blood. Results. With a median follow-up after AHSCT of 1.6 years (range 0.1-22.3 years, the probability of leukemia-free survival (LFS for the whole group at 10 years was 27% and 23% at 20 years. Transplant-related mortality at 100 days after AHSCT was 3.2%.. There was a strong tendency for better LFS for MRD-negative patients if compared with patients who had positive or unknown MRD status at AHSCT (32% vs 23% and 25%, respectively; p=0.06. There was no difference in LFS between B- and T-lineage ALL as well as between patients transplanted in first complete remission (CR1 and CR2. LFS at 10 years for patients with detectable BCR-ABL at transplant was 20% and this was comparable with subjects with negative and missing BCR-ABL status (26% and 28%; p=0.97. Conclusions. The results of AHSCT for high-risk ALL remains unsatisfactory with low probability of long-term LFS.

  19. CML Mouse Model Generated from Leukemia Stem Cells.

    Science.gov (United States)

    Hu, Yiguo

    2016-01-01

    Chronic myeloid leukemia (CML) is a myeloproliferative disorder with a high number of well-differentiated neutrophils in peripheral blood and myeloid cells in bone marrow (BM). CML is derived from the hematopoietic stem cells (HSCs) with the Philadelphia chromosome (Ph(+), t(9;22)-(q34;q11)), resulting in generating a fusion oncogene, BCR/ABL1. HSCs with Ph(+) are defined as leukemia stem cells (LSCs), a subpopulation cell at the apex of hierarchies in leukemia cells and responsible for the disease continuous propagation. Several kinds of CML models have been developed to reveal the mechanism of CML pathogenesis and evaluate therapeutic drugs in the past three decades. Here, we describe the procedures to generate a CML mouse model by introducing BCR/ABL1 into Lin(-)Sca1(+) cKit(+) population cells purified from mouse bone marrow. In CML retroviral transduction/transplantation mouse models, this modified model can mimic CML pathogenesis on high fidelity. PMID:27581136

  20. Coexistance of JAK2V617F mutation and BCR/ABL translocation in one patient

    Directory of Open Access Journals (Sweden)

    Murat Albayrak

    2010-09-01

    Full Text Available Dear Editor,The myeloproliferative disorders (MPDs constitute a subcategory of chronic myeloid disorders and include chronic myeloid leukemia (CML, essential thrombocytemia (ET, polycythemia vera (PV and myelofibrosis (MF. In 1960, the discovery of the Philadelphia chromosome (Ph became a cornerstone in CML treatment and led to the development of moleculary targeted therapy. Recently, an acquired mutation in the Janus kinase 2 (JAK2 gene has been discovered in nearly all patents with PV and approximately half of the patients with primary MF and ET. Subsequently, the mutation has been demonstrated in atypical MPDs (chronic neutrophilic leukemia, unclassified, de novo myelodysplastic syndrome or acute myeloid leukemia.1 It has been hoped that targeted inhibition of JAK2V617F should achieve similar disease control as thyrosine kinases has produced in CML.

  1. Targeted destruction of HIV-positive cells

    Directory of Open Access Journals (Sweden)

    Jyoti R Sharma

    2014-11-01

    Full Text Available Introduction: HIV/AIDS is now a global epidemic that has become the leading infectious killer of adults worldwide. Although antiretroviral (ARV therapy has dramatically improved the quality of life and increased the life expectancy of those infected with HIV but frequency of dosing and drug toxicity as well as the development of viral resistance pose additional limitations. The rapidly expanding field of nanotechnology has vast potential to radically advance the treatment and prevention of HIV/AIDS. Nanoparticles can provide improved drug delivery, by virtue of their small size, robustness, safety, multimodality or multifunctionality. Aims and objectives: Since HIV primarily infects CD4+ cells; we aim to use CD4 as a selectable target to deliver a pro-apoptotic protein to HIV-infected cells using nanoparticles as carriers. The aim of study was to develop a nanotechnology-based death inducing delivery system for the destruction of CD4+HIV infected cells through the activation of caspase-3. Methodology: A modified caspase-3 protein (Mut-3 was engineered, which is cleavable only by HIV-1 protease. Mut-3 can activate apoptosis in the presence of HIV-1 protease, consequently killing HIV-positive cells. Mut-3 protein was conjugated to gold nanoparticles together with a CD4-targeting peptide. The efficacy of the gold nanoparticles was tested on CHO cells that were genetically engineered to express GFP labelled CD4 and HIV-1 protease. Results: Mut-3 was expressed in bacterial cells and purified. CHO cells that stably over express CD4-GFP and HIV-1 protease were selected using Fluorescence Activated Cell Sorting. Dose response cell culture experiments showed that gold nanoparticles without Mut-3 and CD4-targeting peptide did not induce cell death in CHO cells, while gold nanoparticles that was conjugated with Mut-3 and the CD4-targeting peptide rapidly induced cell death in CHO cells. Conclusions: Our results suggest that gold nanoparticles conjugated

  2. Pharmacokinetics of dasatinib for Philadelphia-positive acute lymphocytic leukemia with acquired T315I mutation

    Directory of Open Access Journals (Sweden)

    Takahashi Naoto

    2012-05-01

    Full Text Available Abstract Background The BCR-ABL T315I kinase domain mutation is insensitive to dasatinib therapy for Philadelphia-positive acute lymphoid leukemia (Ph + ALL patients. Resistant T315I clone may be present prior to initiating dasatinib, which could expand under selective pressures during treatment. However, it is also possible that Ph + ALL patients newly acquire the T315I mutation during dasatinib therapy. Despite the potent inhibition of BCR-ABL kinase by dasatinib, little is known about the relationship between dasatinib pharmacokinetics and the emergence of kinase domain mutations in vivo. Methods To determine whether plasma dasatinib pharmacokinetics influences the emergence of BCR-ABL mutations, we measured plasma dasatinib levels in 11 Ph + ALL patients undergoing dasatinib monotherapy. Results Bone marrow relapse occurred in 5 of the 11 Ph + ALL patients (45%. Importantly, a T315I mutation was detected in 4 of the 5 relapsed patients, despite the absence of BCR-ABL mutations in any patient at baseline. The median plasma concentration at 2 hours (C2h, the median plasma maximum concentration (Cmax, and the median area under the observed plasma concentration-time curve from 0 to 4 hours (AUC0-4 were all significantly lower in patients with T315I than those without the mutation (C2h, 22.3 ng/mL vs. 111.6 ng/mL, P = 0.0242; Cmax, 43.8 ng/mL vs. 112.4 ng/mL, P = 0.0242; AUC0-4, 108.3 ng·h/mL vs. 268.3 ng·h/mL, P = 0.0061, respectively. Conclusions These data indicate that the emergence of the T315I mutation among Ph + ALL patients treated with dasatinib is, in part, dependent on plasma dasatinib pharmacokinetics. Notably, these data also suggest that newly acquired BCR-ABL mutations may be inhibited by an increased exposure of dasatinib.

  3. Treatment of human pre-B acute lymphoblastic leukemia with the Aurora kinase inhibitor PHA-739358 (Danusertib

    Directory of Open Access Journals (Sweden)

    Fei Fei

    2012-06-01

    Full Text Available Abstract Background Treatment of Philadelphia chromosome-positive acute lymphoblastic leukemias (Ph-positive ALL with clinically approved inhibitors of the Bcr/Abl tyrosine kinase frequently results in the emergence of a leukemic clone carrying the T315I mutation in Bcr/Abl, which confers resistance to these drugs. PHA-739358, an Aurora kinase inhibitor, was reported to inhibit the Bcr/Abl T315I mutant in CML cells but no preclinical studies have examined this in detail in human ALL. Results We compared the sensitivity of human Bcr/Abl T315I, Bcr/Abl wild type and non-Bcr/Abl ALL cells to this drug. PHA-739358 inhibited proliferation and induced apoptosis independently of Bcr/Abl, the T315I mutation, or presence of the tumor suppressor p53, but the degree of effectiveness varied between different ALL samples. Since short-term treatment with a single dose of drug only transiently inhibited proliferation, we tested combination treatments of PHA-739358 with the farnesyltransferase inhibitor Lonafarnib, with vincristine and with dasatinib. All combinations reduced viability and cell numbers compared to treatment with a single drug. Clonogenic assays showed that 25 nM PHA-739358 significantly reduced the colony growth potential of Ph-positive ALL cells, and combined treatment with a second drug abrogated colony growth in this assay. PHA-739358 further effectively blocked Bcr/Abl tyrosine kinase activity and Aurora kinase B in vivo, and mice transplanted with human Bcr/Abl T315I ALL cells treated with a 3x 7-day cycle of PHA-739358 as mono-treatment had significantly longer survival. Conclusions PHA-739358 represents an alternative drug for the treatment of both Ph-positive and negative ALL, although combined treatment with a second drug may be needed to eradicate the leukemic cells.

  4. Aging-associated inflammation promotes selection for adaptive oncogenic events in B cell progenitors

    OpenAIRE

    Henry, C J; Casas-Selves, M.; Kim, J; Zaberezhnyy, V.; Aghili, L.; Daniel, A.E.; Jimenez, L; Azam, T.; McNamee, E.N.; Clambey, E.T.; Klawitter, J; Serkova, N.J.; Tan, A.C.; Dinarello, C A; DeGregori, J.

    2015-01-01

    The incidence of cancer is higher in the elderly; however, many of the underlying mechanisms for this association remain unexplored. Here, we have shown that B cell progenitors in old mice exhibit marked signaling, gene expression, and metabolic defects. Moreover, B cell progenitors that developed from hematopoietic stem cells (HSCs) transferred from young mice into aged animals exhibited similar fitness defects. We further demonstrated that ectopic expression of the oncogenes BCR-ABL, NRASV1...

  5. Hypoxia selects bortezomib-resistant stem cells of chronic myeloid leukemia.

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    Michele Tanturli

    Full Text Available We previously demonstrated that severe hypoxia inhibits growth of Chronic Myeloid Leukemia (CML cells and selects stem cells where BCR/Abl(protein is suppressed, although mRNA is not, so that hypoxia-selected stem cells, while remaining leukemic, are independent of BCR/Abl signaling and thereby refractory to Imatinib-mesylate. The main target of this study was to address the effects of the proteasome inhibitor Bortezomib (BZ on the maintenance of stem or progenitor cells in hypoxic primary cultures (LC1, by determining the capacity of LC1 cells to repopulate normoxic secondary cultures (LC2 and the kinetics of this repopulation. Unselected K562 cells from day-2 hypoxic LC1 repopulated LC2 with rapid, progenitor-type kinetics; this repopulation was suppressed by BZ addition to LC1 at time 0, but completely resistant to day-1 BZ, indicating that progenitors require some time to adapt to stand hypoxia. K562 cells selected in hypoxic day-7 LC1 repopulated LC2 with stem-type kinetics, which was largely resistant to BZ added at either time 0 or day 1, indicating that hypoxia-selectable stem cells are BZ-resistant per se, i.e. before their selection. Furthermore, these cells were completely resistant to day-6 BZ, i.e. after selection. On the other hand, hypoxia-selected stem cells from CD34-positive cells of blast-crisis CML patients appeared completely resistant to either time-0 or day-1 BZ. To exploit in vitro the capacity of CML cells to adapt to hypoxia enabled to detect a subset of BZ-resistant leukemia stem cells, a finding of particular relevance in light of the fact that our experimental system mimics the physiologically hypoxic environment of bone marrow niches where leukemia stem cells most likely home and sustain minimal residual disease in vivo. This suggests the use of BZ as an enhanced strategy to control CML. in particular to prevent relapse of disease, to be considered with caution and to need further deepening.

  6. Gravity and positional homeostasis of the cell

    Science.gov (United States)

    Nace, G. W.

    1983-01-01

    The effect of gravity upon cytoplasmic aggregates of the size present in eggs and upon cells is investigated. An expression is developed to describe the tendency of torque to rotate the egg and reorganize its constituents. This expression provides the net torque resulting from buoyancy and gravity acting upon a dumbbell-shaped cell, with heavy and light masses at either end and floating in a medium. Torques of approximately 2.5 x 10 to the -13th to 0.85 dyne-cm are found to act upon cells ranging from 6.4 microns to 31 mm (chicken egg). It is noted that cells must expend energy to maintain positional homeostasis against gravity, as demonstrated by results from Skylab 3, where tissue cultures used 58 percent more glucose on earth than in space. The implications for developmental biology, physiology, genetics, and evolution are discussed. It is argued that at the cellular and tissue levels the concept of gravity receptors may be unnecessary.

  7. Gut microbiota-derived propionate reduces cancer cell proliferation in the liver

    OpenAIRE

    Bindels, Laure B.; Porporato, Paolo; Dewulf, Evelyne; Verrax, Julien; Audrey M Neyrinck; Martin, J C; Scott, K P; Buc Calderon, Pedro; Feron, Olivier; Muccioli, Giulio; Sonveaux, Pierre; Cani, Patrice D.; Delzenne, Nathalie M

    2012-01-01

    BACKGROUND: Metabolites released by the gut microbiota may influence host metabolism and immunity. We have tested the hypothesis that inulin-type fructans (ITF), by promoting microbial production of short-chain fatty acids (SCFA), influence cancer cell proliferation outside the gut. METHODS: Mice transplanted with Bcr-Abl-transfected BaF3 cells, received ITF in their drinking water. Gut microbiota was analysed by 16S rDNA polymerase chain reaction (PCR)-denaturing gradient gel electrophoresis...

  8. CDKIs p18INK4c and p57Kip2 are involved in quiescence of CML leukemic stem cells after treatment with TKI

    Science.gov (United States)

    Moreno-Lorenzana, Dafne; Avilés-Vazquez, Sócrates; Sandoval Esquivel, Miguel Angel; Alvarado-Moreno, Antonio; Ortiz-Navarrete, Vianney; Torres-Martínez, Héctor; Ayala-Sánchez, Manuel; Mayani, Héctor; Chavez-Gonzalez, Antonieta

    2016-01-01

    ABSTRACT Chronic Myeloid Leukemia (CML) is sustained by a small population of cells with stem cell characteristics known as Leukemic Stem Cells that are positive to BCR-ABL fusion protein, involved with several abnormalities in cell proliferation, expansion, apoptosis and cell cycle regulation. Current treatment options for CML involve the use of Tirosine Kinase Inhibitor (Imatinib, Nilotinib and Dasatinib), that efficiently reduce proliferation proliferative cells but do not kill non proliferating CML primitive cells that remain and contributes to the persistence of the disease. In order to understand the role of Cyclin Dependent Kinase Inhibitors in CML LSC permanence after TKI treatment, in this study we analyzed cell cycle status, the levels of several CDKIs and the subcellular localization of such molecules in different CML cell lines, as well as primary CD34+CD38−lin− LSC and HSC. Our results demonstrate that cellular location of p18INK4c and p57Kip2 seems to be implicated in the antiproliferative activity of Imatinib and Dasatinib in CML cells and also suggest that the permanence of quiescent stem cells after TKI treatment could be associated with a decrease in p18INK4c and p57Kip2 nuclear location. The differences in p18INK4cand p57Kip2activities in CML and normal stem cells suggest a different cell cycle regulation and provide a platform that could be considered in the development of new therapeutic options to eliminate LSC. PMID:26985855

  9. CD34-positive interstitial cells of the human detrusor

    DEFF Research Database (Denmark)

    Rasmussen, Helle; Hansen, Alastair; Smedts, Frank; Rumessen, Jüri J; Horn, Thomas

    2007-01-01

    Interstitial cells of Cajal (ICC) are well described in the bowel wall. They are c-kit positive and play a role as pacemaker cells. Similar c-kit-positive cells have recently been described in the human bladder. The aim of this study was to characterize interstitial cells of the bladder detrusor ...

  10. Chronic Myelogenous Leukemia Cells Contribute to the Stromal Myofibroblasts in Leukemic NOD/SCID Mouse In Vivo

    Directory of Open Access Journals (Sweden)

    Ryosuke Shirasaki

    2012-01-01

    Full Text Available We recently reported that chronic myelogenous leukemia (CML cells converted into myofibroblasts to create a microenvironment for proliferation of CML cells in vitro. To analyze a biological contribution of CML-derived myofibroblasts in vivo, we observed the characters of leukemic nonobese diabetes/severe combined immunodeficiency (NOD/SCID mouse. Bone marrow nonadherent mononuclear cells as well as human CD45-positive cells obtained from CML patients were injected to the irradiated NOD/SCID mice. When the chimeric BCR-ABL transcript was demonstrated in blood, human CML cells were detected in NOD/SCID murine bone marrow. And CML-derived myofibroblasts composed with the bone marrow-stroma, which produced significant amounts of human vascular endothelial growth factor A. When the parental CML cells were cultured with myofibroblasts separated from CML cell-engrafted NOD/SCID murine bone marrow, CML cells proliferated significantly. These observations indicate that CML cells make an adequate microenvironment for their own proliferation in vivo.

  11. Production of Replication-Defective Retrovirus by Transient Transfection of 293T cells

    OpenAIRE

    Gavrilescu, L Cristina; Richard A Van Etten

    2007-01-01

    Our lab studies human myeloproliferative diseases induced by such oncogenes as Bcr-Abl or growth factor receptor-derived oncogenes (ZNF198-FGFR1, Bcr-PDGFRα, etc.). We are able to model and study a human-like disease in our mouse model, by transplanting bone marrow cells previously infected with a retrovirus expressing the oncogene of interest. Replication-defective retrovirus encoding a human oncogene and a marker (GFP, RFP, antibiotic resistance gene, etc.) is produced by a transient transf...

  12. Heterogeneity of leukemia-initiating capacity of chronic myelogenous leukemia stem cells.

    Science.gov (United States)

    Zhang, Bin; Li, Ling; Ho, Yinwei; Li, Min; Marcucci, Guido; Tong, Wei; Bhatia, Ravi

    2016-03-01

    Chronic myelogenous leukemia (CML) results from transformation of a long-term hematopoietic stem cell (LTHSC) by expression of the BCR-ABL fusion gene. However, BCR-ABL-expressing LTHSCs are heterogeneous in their capacity as leukemic stem cells (LSCs). Although discrepancies in proliferative, self-renewal, and differentiation properties of normal LTHSCs are being increasingly recognized, the mechanisms underlying heterogeneity of leukemic LTHSCs are poorly understood. Using a CML mouse model, we identified gene expression differences between leukemic and nonleukemic LTHSCs. Expression of the thrombopoietin (THPO) receptor MPL was elevated in leukemic LTHSC populations. Compared with LTHSCs with low MPL expression, LTHSCs with high MPL expression showed enhanced JAK/STAT signaling and proliferation in response to THPO in vitro and increased leukemogenic capacity in vivo. Although both G0 and S phase subpopulations were increased in LTHSCs with high MPL expression, LSC capacity was restricted to quiescent cells. Inhibition of MPL expression in CML LTHSCs reduced THPO-induced JAK/STAT signaling and leukemogenic potential. These same phenotypes were also present in LTHSCs from patients with CML, and patient LTHSCs with high MPL expression had reduced sensitivity to BCR-ABL tyrosine kinase inhibitor treatment but increased sensitivity to JAK inhibitors. Together, our studies identify MPL expression levels as a key determinant of heterogeneous leukemia-initiating capacity and drug sensitivity of CML LTHSCs and suggest that high MPL-expressing CML stem cells are potential targets for therapy. PMID:26878174

  13. Influence of different chromosomal abnormalities in Ph-positive bone marrow cells on the chronic myeloid leukemia course during tyrosine kinase inhibitors therapy

    Directory of Open Access Journals (Sweden)

    O. Yu. Vinogradova

    2012-01-01

    Full Text Available The additional molecular and chromosomal abnormalities (ACA in Phositive cells usually considered as a genetic marker of chronic myeloid leukemia (CML progression. 457 patients in different CML phases received tyrosine kinase inhibitors (1st and 2nd generation were studied. During therapy 50 cases with additional chromosomal abnormalities in Ph+ clone (22 of them in chronic CML phase were revealed (median follow-up from CML diagnosis – 117 months, median imatinib therapy – 62 months. 86 % of patients in chronic phase with Ph+- cell abnormalities were cytogenetic resistance, and their 5-years overall survival was 80 % which was significantly lower than in patients without ACA (p < 0.005. The treatment results depend on chromosomal abnormalities detected. In patients with additional chromosome 8 imatinib therapy is effective, although complete cytogenetic response (CCR is achieved only in the later therapy stages. In patients with additional translocations CCR also achieved with imatinib or 2nd generation TKI. Only a third of patients with additional Ph-chromosome or BCR/ABL amplification achieved complete suppression of Ph+ clone using 2nd generation TKI. The presence of additional chromosome 7 abnormalities and complex karyotype disorders involving isochromosome i(17(q10 are poor prognostic factors of TKI treatment failures.

  14. Influence of different chromosomal abnormalities in Ph-positive bone marrow cells on the chronic myeloid leukemia course during tyrosine kinase inhibitors therapy

    Directory of Open Access Journals (Sweden)

    O. Yu. Vinogradova

    2014-07-01

    Full Text Available The additional molecular and chromosomal abnormalities (ACA in Phositive cells usually considered as a genetic marker of chronic myeloid leukemia (CML progression. 457 patients in different CML phases received tyrosine kinase inhibitors (1st and 2nd generation were studied. During therapy 50 cases with additional chromosomal abnormalities in Ph+ clone (22 of them in chronic CML phase were revealed (median follow-up from CML diagnosis – 117 months, median imatinib therapy – 62 months. 86 % of patients in chronic phase with Ph+- cell abnormalities were cytogenetic resistance, and their 5-years overall survival was 80 % which was significantly lower than in patients without ACA (p < 0.005. The treatment results depend on chromosomal abnormalities detected. In patients with additional chromosome 8 imatinib therapy is effective, although complete cytogenetic response (CCR is achieved only in the later therapy stages. In patients with additional translocations CCR also achieved with imatinib or 2nd generation TKI. Only a third of patients with additional Ph-chromosome or BCR/ABL amplification achieved complete suppression of Ph+ clone using 2nd generation TKI. The presence of additional chromosome 7 abnormalities and complex karyotype disorders involving isochromosome i(17(q10 are poor prognostic factors of TKI treatment failures.

  15. BM microenvironmental protection of CML cells from imatinib through Stat5/NF-κB signaling and reversal by Wogonin.

    Science.gov (United States)

    Xu, Xuefen; Zhang, Xiaobo; Liu, Yicheng; Yang, Lin; Huang, Shaoliang; Lu, Lu; Wang, Shuhao; Guo, Qinglong; Zhao, Li

    2016-04-26

    Constitutive Stat5 activation enhanced cell survival and resistance to imatinib (IM) in chronic myelogenous leukemia (CML) cells. However, the mechanism of Stat5 activation in mediating resistance to IM in bone marrow (BM) microenvironment has not been evaluated precisely. In this study, we reported HS-5-derived conditioned medium (CM) significantly enhanced IM resistance in K562 and KU812. Interestingly, upregulation of the proportion of CD34+ subpopulation was found in CML cells. Subsequently, the BCR/ABL-independent activation of Stat5 increased P-glycoprotein (P-gp) activity in CM-mediated protection of CML stem cells (LSCs) from IM. Further research revealed Stat5 activation increased the DNA binding activity of NF-κB though binding of p-Stat5 and p-RelA in nucleus. Moreover, highly acetylated RelA was required for Stat5-mediated RelA nuclear binding. The study further confirmed that Wogonin potentiated the inhibitory effects of IM on leukemia development by suppressing Stat5 pathway both in CM model and the K562 xenograft model. In summary, results clearly demonstrated BCR/ABL-independent Stat5 survival pathway could contribute to resistance of CML LSCs to IM in BM microenvironment and suggested that natural durgs effectively inhibiting Stat5 may be an attractive approach to overcome resistance to BCR/ABL kinase inhibitors. PMID:27027438

  16. Specific Antileukemic Activity of PD0332991, a CDK4/6 Inhibitor, against Philadelphia Chromosome-Positive Lymphoid Leukemia.

    Science.gov (United States)

    Nemoto, Atsushi; Saida, Satoshi; Kato, Itaru; Kikuchi, Jiro; Furukawa, Yusuke; Maeda, Yasuhiro; Akahane, Koshi; Honna-Oshiro, Hiroko; Goi, Kumiko; Kagami, Keiko; Kimura, Shinya; Sato, Yuko; Okabe, Seiichi; Niwa, Akira; Watanabe, Kenichiro; Nakahata, Tatsutoshi; Heike, Toshio; Sugita, Kanji; Inukai, Takeshi

    2016-01-01

    S-phase progression of the cell cycle is accelerated in tumors through various genetic abnormalities, and, thus, pharmacologic inhibition of altered cell-cycle progression would be an effective strategy to control tumors. In the current study, we analyzed the antileukemic activity of three available small molecules targeting CDK4/CDK6 against lymphoid crisis of chronic myeloid leukemia (CML-LC) and Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph(+) ALL), and found that all three molecules showed specific activities against leukemic cell lines derived from CML-LC and Ph(+) ALL. In particular, PD0332991 exhibited extremely high antileukemic activity against CML-LC and Ph(+) ALL cell lines in the nanomolar range by the induction of G0-G1 arrest and partially cell death through dephosphorylation of pRb and downregulation of the genes that are involved in S-phase transition. As an underlying mechanism for favorable sensitivity to the small molecules targeting CDK4/CDK6, cell-cycle progression of Ph(+) lymphoid leukemia cells was regulated by transcriptional and posttranscriptional modulation of CDK4 as well as Cyclin D2 gene expression under the control of BCR-ABL probably through the PI3K pathway. Consistently, the gene expression level of Cyclin D2 in Ph(+) lymphoid leukemia cells was significantly higher than that in Ph(-) lymphoid leukemia cells. Of note, three Ph(+) ALL cell lines having the T315I mutation also showed sensitivity to PD0332991. In a xenograft model, PD0332991, but not imatinib, suppressed dissemination of Ph(+) ALL having the T315I mutation and prolonged survival, demonstrating that this reagent would be a new therapeutic modality for relapsed CML-LC and Ph(+) ALL patients after treatment with tyrosine kinase inhibitors. PMID:26637365

  17. Reciprocal t(9;22 ABL/BCR fusion proteins: leukemogenic potential and effects on B cell commitment.

    Directory of Open Access Journals (Sweden)

    Xiaomin Zheng

    Full Text Available BACKGROUND: t(9;22 is a balanced translocation, and the chromosome 22 breakpoints (Philadelphia chromosome--Ph+ determine formation of different fusion genes that are associated with either Ph+ acute lymphatic leukemia (Ph+ ALL or chronic myeloid leukemia (CML. The "minor" breakpoint in Ph+ ALL encodes p185(BCR/ABL from der22 and p96(ABL/BCR from der9. The "major" breakpoint in CML encodes p210(BCR/ABL and p40(ABL/BCR. Herein, we investigated the leukemogenic potential of the der9-associated p96(ABL/BCR and p40(ABL/BCR fusion proteins and their roles in the lineage commitment of hematopoietic stem cells in comparison to BCR/ABL. METHODOLOGY: All t(9;22 derived proteins were retrovirally expressed in murine hematopoietic stem cells (SL cells and human umbilical cord blood cells (UCBC. Stem cell potential was determined by replating efficiency, colony forming--spleen and competitive repopulating assays. The leukemic potential of the ABL/BCR fusion proteins was assessed by in a transduction/transplantation model. Effects on the lineage commitment and differentiation were investigated by culturing the cells under conditions driving either myeloid or lymphoid commitment. Expression of key factors of the B-cell differentiation and components of the preB-cell receptor were determined by qRT-PCR. PRINCIPAL FINDINGS: Both p96(ABL/BCR and p40(ABL/BCR increased proliferation of early progenitors and the short term stem cell capacity of SL-cells and exhibited own leukemogenic potential. Interestingly, BCR/ABL gave origin exclusively to a myeloid phenotype independently from the culture conditions whereas p96(ABL/BCR and to a minor extent p40(ABL/BCR forced the B-cell commitment of SL-cells and UCBC. CONCLUSIONS/SIGNIFICANCE: Our here presented data establish the reciprocal ABL/BCR fusion proteins as second oncogenes encoded by the t(9;22 in addition to BCR/ABL and suggest that ABL/BCR contribute to the determination of the leukemic phenotype through their

  18. Current Concepts in Pediatric Philadelphia Chromosome Positive Acute Lymphoblastic Leukemia (Ph+ ALL

    Directory of Open Access Journals (Sweden)

    KathrinM.Bernt

    2014-03-01

    Full Text Available The t(9;22(q34;q11 or Philadelphia chromosome that creates a BCR-ABL1 fusion gene encoding for a chimeric BCR-ABL1 protein is present in 3-4% of pediatric acute lymphoblastic leukemia (Ph+ ALL, and about 25% of adult ALL cases. Prior to the advent of tyrosine kinase inhibitors (TKI, Ph+ ALL was associated with a very poor prognosis despite use of intensive chemotherapy and frequently hematopoietic stem cell transplantation (HSCT in first remission. The development of TKIs revolutionized the therapy of Ph+ ALL. Addition of the first generation ABL1 class TKI imatinib to intensive chemotherapy dramatically increased survival for children with Ph+ ALL and established that many patients can be cured without HSCT. In parallel, the mechanistic understanding of Ph+ ALL expanded exponentially through careful mapping of pathways downstream of BCR-ABL1, the discovery of mutations in master regulators of B-cell development such as IKZF1 (Ikaros, PAX5 and EBF, the recognition of the complex clonal architecture of Ph+ ALL, and the delineation of genomic, epigenetic and signaling abnormalities contributing to relapse and resistance. Still, many important basic and clinical questions remain unanswered. Current clinical trials are testing second generation TKIs in patients with newly diagnosed Ph+ ALL. Neither the optimal duration of therapy nor the optimal chemotherapy backbone are currently defined. The role of HSCT in first remission and post-transplant TKI therapy also require further study. In addition, it will be crucial to continue to dig deeper into understanding Ph+ ALL at a mechanistic level, and translate findings into complementary targeted approaches. Expanding targeted therapies holds great promise to decrease toxicity and improve survival in this high risk disease, which provides a paradigm for how targeted therapies can be incorporated into treatment of other high risk leukemias.

  19. Distinct graft-versus-leukemic stem cell effects of early or delayed donor leukocyte infusions in a mouse chronic myeloid leukemia model

    OpenAIRE

    Lu, Yi-Fen; Gavrilescu, L Cristina; Betancur, Monica; Lazarides, Katherine; Klingemann, Hans; Van Etten, Richard A.

    2012-01-01

    Among hematologic neoplasms, chronic myeloid leukemia (CML) is exquisitely sensitive to graft-versus-leukemia (GVL) because patients relapsing after allogeneic hematopoietic stem-cell transplantation (alloHSCT) can be cured by donor leukocyte infusion (DLI); however, the cellular mechanisms and strategies to separate GVL from GVHD are unclear. We used a BCR-ABL1 transduction/transplantation mouse model to study the mechanisms of DLI in MHC-matched, minor histocompatibility antigen–mismatched ...

  20. T Cell Adolescence: Maturation Events Beyond Positive Selection.

    Science.gov (United States)

    Hogquist, Kristin A; Xing, Yan; Hsu, Fan-Chi; Shapiro, Virginia Smith

    2015-08-15

    Single-positive thymocytes that successfully complete positive and negative selection must still undergo one final step, generally termed T cell maturation, before they gain functional competency and enter the long-lived T cell pool. Maturation initiates after positive selection in single-positive thymocytes and continues in the periphery in recent thymic emigrants, before these newly produced T cells gain functional competency and are ready to participate in the immune response as peripheral naive T cells. Recent work using genetically altered mice demonstrates that T cell maturation is not a single process, but a series of steps that occur independently and sequentially after positive selection. This review focuses on the changes that occur during T cell maturation, as well as the molecules and pathways that are critical at each step. PMID:26254267

  1. Marker-free cell discrimination by holographic optical tweezers

    Science.gov (United States)

    Schaal, F.; Warber, M.; Zwick, S.; van der Kuip, H.; Haist, T.; Osten, W.

    2009-06-01

    We introduce a method for marker-free cell discrimination based on optical tweezers. Cancerous, non-cancerous, and drug-treated cells could be distinguished by measuring the trapping forces using holographic optical tweezers. We present trapping force measurements on different cell lines: normal pre-B lymphocyte cells (BaF3; "normal cells"), their Bcr-Abl transformed counterparts (BaF3-p185; "cancer cells") as a model for chronic myeloid leukaemia (CML) and Imatinib treated BaF3-p185 cells. The results are compared with reference measurements obtained by a commercial flow cytometry system.

  2. Clinical impact of ABL1 kinase domain mutations and IKZF1 deletion in adults under age 60 with Philadelphia chromosome-positive (Ph+) acute lymphoblastic leukemia (ALL): molecular analysis of CALGB (Alliance) 10001 and 9665.

    Science.gov (United States)

    DeBoer, Rebecca; Koval, Gregory; Mulkey, Flora; Wetzler, Meir; Devine, Steven; Marcucci, Guido; Stone, Richard M; Larson, Richard A; Bloomfield, Clara D; Geyer, Susan; Mullighan, Charles G; Stock, Wendy

    2016-10-01

    Recent studies have identified oncogenic lesions in Philadelphia chromosome-positive (Ph+)  acute lymphoblastic leukemia (ALL) and ABL1 kinase mutations that confer resistance to tyrosine kinase inhibitors. We sought to determine the prevalence and clinical impact of these lesions in patients on CALGB 10001, a previously reported Phase II study of imatinib, chemotherapy, and hematopoietic cell transplant in adult Ph + ALL. Of the 58 enrolled, 22 relapsed. By direct sequencing, an ABL1 kinase mutation known to induce imatinib resistance was present at relapse in 13 of 20. Using quantitative PCR assays, the mutations were detectable at diagnosis or early during treatment in most (62%) relapsed patients. Aberrations in IKZF1, CDKN2A/B, and PAX5 were assessed in 28 samples using SNP arrays and genomic DNA sequencing. Of these, 22 (79%) had IKZF1 deletion. The combination of IKZF1 deletion and p210 BCR-ABL1 (p < 0.0001), high white blood cell count (p = 0.021), and minimal residual disease (p = 0.013) were associated with worse disease-free survival. PMID:26892479

  3. T cell depleted haploidentical transplantation: positive selection

    Directory of Open Access Journals (Sweden)

    Franco Aversa

    2011-06-01

    Full Text Available Interest in mismatched transplantation arises from the fact that a suitable one-haplotype mismatched donor is immediately available for virtually all patients, particularly for those who urgently need an allogenic transplant. Work on one haplotype-mismatched transplants has been proceeding for over 20 years all over the world and novel transplant techniques have been developed. Some centres have focused on the conditioning regimens and post transplant immune suppression; others have concentrated on manipulating the graft which may be a megadose of extensively T celldepleted or unmanipulated progenitor cells. Excellent engraftment rates are associated with a very low incidence of acute and chronic GVHD and regimen-related mortality even in patients who are over 50 years old. Overall, event-free survival and transplant-related mortality compare favourably with reports on transplants from sources of stem cells other than the matched sibling.

  4. Novel Serial Positive Enrichment Technology Enables Clinical Multiparameter Cell Sorting

    OpenAIRE

    Christian Stemberger; Stefan Dreher; Claudia Tschulik; Christine Piossek; Jeannette Bet; Yamamoto, Tori N.; Matthias Schiemann; Michael Neuenhahn; Klaus Martin; Martin Schlapschy; Arne Skerra; Thomas Schmidt; Matthias Edinger; Riddell, Stanley R.; Lothar Germeroth

    2012-01-01

    A general obstacle for clinical cell preparations is limited purity, which causes variability in the quality and potency of cell products and might be responsible for negative side effects due to unwanted contaminants. Highly pure populations can be obtained best using positive selection techniques. However, in many cases target cell populations need to be segregated from other cells by combinations of multiple markers, which is still difficult to achieve--especially for clinical cell product...

  5. Activity of ladanein on leukemia cell lines and its occurrence in Marrubium vulgare.

    Science.gov (United States)

    Alkhatib, Racha; Joha, Sami; Cheok, Meyling; Roumy, Vincent; Idziorek, Thierry; Preudhomme, Claude; Quesnel, Bruno; Sahpaz, Sevser; Bailleul, François; Hennebelle, Thierry

    2010-01-01

    Three methoxylated flavones isolated from Marrubium peregrinum - ladanein, scutellarein-5,7,4'-trimethyl ether, and scutellarein-5,6,7,4'-tetramethyl ether - were assayed for their cytotoxicity towards a recently developed dasatinib-resistant murine leukemia cell line (DA1-3b/M2 (BCR-ABL)), together with the structurally related non-methylated flavone scutellarein. The most active compound, ladanein, was looked for in 20 common Lamiaceae species by a quick HPLC screening. Among the possible positive results, the most interesting source was found to be Marrubium vulgare, which led to the isolation and identification of ladanein for the first time in this species. Ladanein also displayed moderate (20-40 microM) activities against K562, K562R (imatinib-resistant), and 697 human leukemia cell lines but was toxic neither to MOLM13 nor to human peripheral blood mononuclear cells. This work provides a common natural source for the hemi-synthesis of future ladanein-derived flavones and the study of their antileukemic activity. PMID:19644796

  6. Identification of galectin-1 as a novel mediator for chemoresistance in chronic myeloid leukemia cells.

    Science.gov (United States)

    Luo, Wu; Song, Li; Chen, Xi-Lei; Zeng, Xiang-Feng; Wu, Jian-Zhang; Zhu, Cai-Rong; Huang, Tao; Tan, Xiang-Peng; Lin, Xiao-Mian; Yang, Qi; Wang, Ji-Zhong; Li, Xiao-Kun; Wu, Xiao-Ping

    2016-05-01

    Multidrug resistance protein-1 (MDR1) has been proven to be associated with the development of chemoresistance to imatinib (Glivec, STI571) which displays high efficacy in treatment of BCR-ABL-positive chronic myelogenous leukemia (CML). However, the possible mechanisms of MDR1 modulation in the process of the resistance development remain to be defined. Herein, galectin-1 was identified as a candidate modulator of MDR1 by proteomic analysis of a model system of leukemia cell lines with a gradual increase of MDR1 expression and drug resistance. Coincidently, alteration of galectin-1 expression triggers the change of MDR1 expression as well as the resistance to the cytotoxic drugs, suggesting that augment of MDR1 expression engages in galectin-1-mediated chemoresistance. Moreover, we provided the first data showing that NF-κB translocation induced by P38 MAPK activation was responsible for the modulation effect of galectin-1 on MDR1 in the chronic myelogenous leukemia cells. Galectin-1 might be considered as a novel target for combined modality therapy for enhancing the efficacy of CML treatment with imatinib. PMID:27050374

  7. HER2-positive circulating tumor cells in breast cancer.

    Directory of Open Access Journals (Sweden)

    Michail Ignatiadis

    Full Text Available PURPOSE: Circulating Tumor Cells (CTCs detection and phenotyping are currently evaluated in Breast Cancer (BC. Tumor cell dissemination has been suggested to occur early in BC progression. To interrogate dissemination in BC, we studied CTCs and HER2 expression on CTCs across the spectrum of BC staging. METHODS: Spiking experiments with 6 BC cell lines were performed and blood samples from healthy women and women with BC were analyzed for HER2-positive CTCs using the CellSearch®. RESULTS: Based on BC cell lines experiments, HER2-positive CTCs were defined as CTCs with HER2 immunofluorescence intensity that was at least 2.5 times higher than the background. No HER2-positive CTC was detected in 42 women without BC (95% confidence interval (CI 0-8.4% whereas 4.1% (95%CI 1.4-11.4% of 73 patients with ductal/lobular carcinoma in situ (DCIS/LCIS had 1 HER2-positive CTC/22.5 mL, 7.9%, (95%CI 4.1-14.9% of 101 women with non metastatic (M0 BC had ≥1 HER2-positive CTC/22.5 mL (median 1 cell, range 1-3 cells and 35.9% (95%CI 22.7-51.9% of 39 patients with metastatic BC had ≥1 HER2-positive CTC/7.5 mL (median 1.5 cells, range 1-42 cells. In CTC-positive women with DCIS/LCIS or M0 BC, HER2-positive CTCs were more commonly detected in HER2-positive (5 of 5 women than HER2-negative BC (5 of 12 women (p = 0.03. CONCLUSION: HER2-positive CTCs were detected in DCIS/LCIS or M0 BC irrespective of the primary tumor HER2 status. Nevertheless, their presence was more common in women with HER2-positive disease. Monitoring of HER2 expression on CTCs might be useful in trials with anti-HER2 therapies.

  8. Hurdles overcome in technology transfer for AIET and Positive outcome in Indian patients

    Directory of Open Access Journals (Sweden)

    Dedeepiya V

    2012-01-01

    perigastric peritoneal deposits with a marked decrease in the size of the inguinal lymph nodes from and no recurrence in the pelvic region. There was also improvement in appetite and quality of life of the patient. There were no adverse reactions following the AIET infusions. [8] The latest follow-up in May 2012 revealed static non progressive disease with all the parameters including tumor markers within normal range. Case 2: A 15 year old girl presented with complaints of diffuse bone pain for 3 months, intermittent fever for 1 month and increasing pallor in October 2004. On examination, she was found to have pallor with hepatosplenomegaly. A low haemoglobin level (9.3 mg/dl, low platelets (21000 and a WBC count of 13600 with 83% lymphoblasts was revealed in her blood picture. Bone marrow examination and flow cytometry were suggestive of Acute Lymphoblastic Leukemia – CD 10+ CD 19+. Cytogenetics revealed Philadelphia chromosome and BCR-ABL positivity. Qualitative BCR-ABL was done serially to assess disease status. She was started on chemotherapy as per UK MRC protocol – Regimen B along with Imatinib Mesylate (in view of her BCR-ABL positivity and prophylactic cranial radiotherapy of 12 Gy at the end of delayed intensification. She had persistent disease with low positive BCR-ABL at 0.04% at the end of delayed intensification. As there were no matched HLA donors as she had high risk disease she was suggested for AIET using autologous natural killer (NK cells (CD3-CD56+ in January 2007. 562 x 106 in vitro expanded NK cells were infused to the patient intravenously and four weeks later, the BCR-ABL in the PB turned negative. She completed her treatment in October 2007. She has been on regular follow up for the last five years and she continues to be in remission. Case 3: A female patient aged 64 years with advanced serous papillary Adenocarcinoma of both ovaries with liver metastasis underwent omentectomy and resection of the liver nodule in the month of January 2011 and

  9. Novel serial positive enrichment technology enables clinical multiparameter cell sorting.

    Directory of Open Access Journals (Sweden)

    Christian Stemberger

    Full Text Available A general obstacle for clinical cell preparations is limited purity, which causes variability in the quality and potency of cell products and might be responsible for negative side effects due to unwanted contaminants. Highly pure populations can be obtained best using positive selection techniques. However, in many cases target cell populations need to be segregated from other cells by combinations of multiple markers, which is still difficult to achieve--especially for clinical cell products. Therefore, we have generated low-affinity antibody-derived Fab-fragments, which stain like parental antibodies when multimerized via Strep-tag and Strep-Tactin, but can subsequently be removed entirely from the target cell population. Such reagents can be generated for virtually any antigen and can be used for sequential positive enrichment steps via paramagnetic beads. First protocols for multiparameter enrichment of two clinically relevant cell populations, CD4(high/CD25(high/CD45RA(high 'regulatory T cells' and CD8(high/CD62L(high/CD45RA(neg 'central memory T cells', have been established to determine quality and efficacy parameters of this novel technology, which should have broad applicability for clinical cell sorting as well as basic research.

  10. A positive feedback cell signaling nucleation model of astrocyte dynamics

    OpenAIRE

    MacDonald, Christopher L.; Silva, Gabriel A.

    2013-01-01

    We constructed a model of calcium signaling in astrocyte neural glial cells that incorporates a positive feedback nucleation mechanism, whereby small microdomain increases in local calcium can stochastically produce global cellular and intercellular network scale dynamics. The model is able to simultaneously capture dynamic spatial and temporal heterogeneities associated with intracellular calcium transients in individual cells and intercellular calcium waves (ICW) in spatially realistic netw...

  11. Novel Serial Positive Enrichment Technology Enables Clinical Multiparameter Cell Sorting

    Science.gov (United States)

    Tschulik, Claudia; Piossek, Christine; Bet, Jeannette; Yamamoto, Tori N.; Schiemann, Matthias; Neuenhahn, Michael; Martin, Klaus; Schlapschy, Martin; Skerra, Arne; Schmidt, Thomas; Edinger, Matthias; Riddell, Stanley R.; Germeroth, Lothar; Busch, Dirk H.

    2012-01-01

    A general obstacle for clinical cell preparations is limited purity, which causes variability in the quality and potency of cell products and might be responsible for negative side effects due to unwanted contaminants. Highly pure populations can be obtained best using positive selection techniques. However, in many cases target cell populations need to be segregated from other cells by combinations of multiple markers, which is still difficult to achieve – especially for clinical cell products. Therefore, we have generated low-affinity antibody-derived Fab-fragments, which stain like parental antibodies when multimerized via Strep-tag and Strep-Tactin, but can subsequently be removed entirely from the target cell population. Such reagents can be generated for virtually any antigen and can be used for sequential positive enrichment steps via paramagnetic beads. First protocols for multiparameter enrichment of two clinically relevant cell populations, CD4high/CD25high/CD45RAhigh ‘regulatory T cells’ and CD8high/CD62Lhigh/CD45RAneg ‘central memory T cells’, have been established to determine quality and efficacy parameters of this novel technology, which should have broad applicability for clinical cell sorting as well as basic research. PMID:22545138

  12. Vimentin positive acantholytic penile squamous cell carcinoma with rhabdoid features

    Directory of Open Access Journals (Sweden)

    Ramesh Y Chavan

    2014-01-01

    Full Text Available Acantholytic variant of penile squamous cell carcinoma (SCC is an exceedingly rare and associated with bad prognosis. Histologically it mimics angiosarcoma due to pseudovascular spaces. Vimentin immunopositivity in such cases represent epithelial to mesenchymal transition manifested by cellular discohesion. We describe a case of vimentin positive acantholytic penile SCC in a 55-year-old patient.

  13. Donor Umbilical Cord Blood Transplant in Treating Patients With Hematologic Cancer

    Science.gov (United States)

    2016-08-05

    Acute Biphenotypic Leukemia; Acute Lymphoblastic Leukemia; Acute Myeloid Leukemia; Aggressive Non-Hodgkin Lymphoma; Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Chronic Phase Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Indolent Non-Hodgkin Lymphoma; Malignant Lymphoma, Large Cell Type; Mixed Phenotype Acute Leukemia; Myelodysplastic Syndrome; Myeloproliferative Neoplasm; Recurrent Chronic Lymphocytic Leukemia; Recurrent Follicular Lymphoma; Recurrent Lymphoplasmacytic Lymphoma; Recurrent Mantle Cell Lymphoma; Recurrent Marginal Zone Lymphoma; Recurrent Plasma Cell Myeloma; Recurrent Small Lymphocytic Lymphoma; Recurrent T-Cell Non-Hodgkin Lymphoma; Refractory Chronic Lymphocytic Leukemia; Refractory Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Refractory Follicular Lymphoma; Refractory Hodgkin Lymphoma; Refractory Lymphoplasmacytic Lymphoma; Refractory Mantle Cell Lymphoma; Refractory Small Lymphocytic Lymphoma; T-Cell Non-Hodgkin Lymphoma

  14. The conformational control inhibitor of tyrosine kinases DCC-2036 is effective for imatinib-resistant cells expressing T674I FIP1L1-PDGFRα.

    Directory of Open Access Journals (Sweden)

    Yingying Shen

    Full Text Available The cells expressing the T674I point mutant of FIP1-like-1-platelet-derived growth factor receptor alpha (FIP1L1-PDGFRα in hypereosinophilics syndrome (HES are resistant to imatinib and some second-generation tyrosine kinase inhibitors (TKIs. There is a desperate need to develop therapy to combat this acquired drug resistance. DCC-2036 has been synthesized as a third-generation TKI to combat especially the Bcr-Abl T315I mutant in chronic myeloid leukemia. This study evaluated the effect of DCC-2036 on FIP1L1-PDGFRα-positive cells, including the wild type (WT and the T674I mutant. The in vitro effects of DCC-2036 on the PDGFRα signal pathways, proliferation, cell cycling and apoptosis of FIP1L1-PDGFRα-positive cells were investigated, and a nude mouse xenograft model was employed to assess the in vivo antitumor activity. We found that DCC-2036 decreased the phosphorylated levels of PDGFRα and its downstream targets without apparent effects on total protein levels. DCC-2036 inhibited proliferation, and induced apoptosis with MEK-dependent up-regulation of the pro-apoptotic protein Bim in FIP1L1-PDGFRα-positive cells. DCC-2036 also exhibited in vivo antineoplastic activity against cells with T674I FIP1L1-PDGFRα. In summary, FIP1L1-PDGFRα-positive cells are sensitive to DCC-2036 regardless of their sensitivity to imatinib. DCC-2036 may be a potential compound to treat imatinib-resistant HES.

  15. Positioning.

    Science.gov (United States)

    Conone, Ruth M.

    The key to positioning is the creation of a clear benefit image in the consumer's mind. One positioning strategy is creating in the prospect's mind a position that takes into consideration the company's or agency's strengths and weaknesses as well as those of its competitors. Another strategy is to gain entry into a position ladder owned by…

  16. Highly parallel identification of essential genes in cancer cells.

    Science.gov (United States)

    Luo, Biao; Cheung, Hiu Wing; Subramanian, Aravind; Sharifnia, Tanaz; Okamoto, Michael; Yang, Xiaoping; Hinkle, Greg; Boehm, Jesse S; Beroukhim, Rameen; Weir, Barbara A; Mermel, Craig; Barbie, David A; Awad, Tarif; Zhou, Xiaochuan; Nguyen, Tuyen; Piqani, Bruno; Li, Cheng; Golub, Todd R; Meyerson, Matthew; Hacohen, Nir; Hahn, William C; Lander, Eric S; Sabatini, David M; Root, David E

    2008-12-23

    More complete knowledge of the molecular mechanisms underlying cancer will improve prevention, diagnosis and treatment. Efforts such as The Cancer Genome Atlas are systematically characterizing the structural basis of cancer, by identifying the genomic mutations associated with each cancer type. A powerful complementary approach is to systematically characterize the functional basis of cancer, by identifying the genes essential for growth and related phenotypes in different cancer cells. Such information would be particularly valuable for identifying potential drug targets. Here, we report the development of an efficient, robust approach to perform genome-scale pooled shRNA screens for both positive and negative selection and its application to systematically identify cell essential genes in 12 cancer cell lines. By integrating these functional data with comprehensive genetic analyses of primary human tumors, we identified known and putative oncogenes such as EGFR, KRAS, MYC, BCR-ABL, MYB, CRKL, and CDK4 that are essential for cancer cell proliferation and also altered in human cancers. We further used this approach to identify genes involved in the response of cancer cells to tumoricidal agents and found 4 genes required for the response of CML cells to imatinib treatment: PTPN1, NF1, SMARCB1, and SMARCE1, and 5 regulators of the response to FAS activation, FAS, FADD, CASP8, ARID1A and CBX1. Broad application of this highly parallel genetic screening strategy will not only facilitate the rapid identification of genes that drive the malignant state and its response to therapeutics but will also enable the discovery of genes that participate in any biological process. PMID:19091943

  17. Fulvestrant radiosensitizes human estrogen receptor-positive breast cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Jing, E-mail: wangstella5@163.com [Department of Breast Surgery, Qilu Hospital, Shandong Univeristy, Wenhua Xi Road 107, Shandong Province (China); Department of Oncology, Affiliated Hospital of Qingdao University Medical College, Shandong Province (China); Yang, Qifeng, E-mail: qifengy@gmail.com [Department of Breast Surgery, Qilu Hospital, Shandong Univeristy, Wenhua Xi Road 107, Shandong Province (China); Haffty, Bruce G., E-mail: hafftybg@umdnj.edu [Department of Radiation Oncology, UMDNJ-Robert Wood Johnson School of Medicine, Cancer Institute of New Jersey, NB (United States); Li, Xiaoyan, E-mail: xiaoyanli1219@gmail.com [Department of Oncology, Affiliated Hospital of Qingdao University Medical College, Shandong Province (China); Moran, Meena S., E-mail: meena.moran@yale.edu [Department of Therapeutic Radiology, Yale University School of Medicine, New Haven, CT (United States)

    2013-02-08

    Highlights: ► Fulvestrant radiosensitizes MCF-7 cells. ► Fulvestrant increases G1 arrest and decreases S phase in MCF-7 cells. ► Fulvestrant down-regulates DNA-PKcs and RAD51 in MCF-7 cells. -- Abstract: The optimal sequencing for hormonal therapy and radiation are yet to be determined. We utilized fulvestrant, which is showing promise as an alternative to other agents in the clinical setting of hormonal therapy, to assess the cellular effects of concomitant anti-estrogen therapy (fulvestrant) with radiation (F + RT). This study was conducted to assess the effects of fulvestrant alone vs. F + RT on hormone-receptor positive breast cancer to determine if any positive or negative combined effects exist. The effects of F + RT on human breast cancer cells were assessed using MCF-7 clonogenic and tetrazolium salt colorimetric (MTT) assays. The assays were irradiated with a dose of 0, 2, 4, 6 Gy ± fulvestrant. The effects of F + RT vs. single adjuvant treatment alone on cell-cycle distribution were assessed using flow cytometry; relative expression of repair proteins (Ku70, Ku80, DNA-PKcs, Rad51) was assessed using Western Blot analysis. Cell growth for radiation alone vs. F + RT was 0.885 ± 0.013 vs. 0.622 ± 0.029 @2 Gy, 0.599 ± 0.045 vs. 0.475 ± 0.054 @4 Gy, and 0.472 ± 0.021 vs. 0.380 ± 0.018 @6 Gy RT (p = 0.003). While irradiation alone induced G2/M cell cycle arrest, the combination of F + RT induced cell redistribution in the G1 phase and produced a significant decrease in the proportion of cells in G2 phase arrest and in the S phase in breast cancer cells (p < 0.01). Furthermore, levels of repair proteins DNA-PKcs and Rad51 were significantly decreased in the cells treated with F + RT compared with irradiation alone. F + RT leads to a decrease in the surviving fraction, increased cell cycle arrest, down regulating of nonhomologous repair protein DNA-PKcs and homologous recombination repair protein RAD51. Thus, our findings suggest that F + RT

  18. Fulvestrant radiosensitizes human estrogen receptor-positive breast cancer cells

    International Nuclear Information System (INIS)

    Highlights: ► Fulvestrant radiosensitizes MCF-7 cells. ► Fulvestrant increases G1 arrest and decreases S phase in MCF-7 cells. ► Fulvestrant down-regulates DNA-PKcs and RAD51 in MCF-7 cells. -- Abstract: The optimal sequencing for hormonal therapy and radiation are yet to be determined. We utilized fulvestrant, which is showing promise as an alternative to other agents in the clinical setting of hormonal therapy, to assess the cellular effects of concomitant anti-estrogen therapy (fulvestrant) with radiation (F + RT). This study was conducted to assess the effects of fulvestrant alone vs. F + RT on hormone-receptor positive breast cancer to determine if any positive or negative combined effects exist. The effects of F + RT on human breast cancer cells were assessed using MCF-7 clonogenic and tetrazolium salt colorimetric (MTT) assays. The assays were irradiated with a dose of 0, 2, 4, 6 Gy ± fulvestrant. The effects of F + RT vs. single adjuvant treatment alone on cell-cycle distribution were assessed using flow cytometry; relative expression of repair proteins (Ku70, Ku80, DNA-PKcs, Rad51) was assessed using Western Blot analysis. Cell growth for radiation alone vs. F + RT was 0.885 ± 0.013 vs. 0.622 ± 0.029 @2 Gy, 0.599 ± 0.045 vs. 0.475 ± 0.054 @4 Gy, and 0.472 ± 0.021 vs. 0.380 ± 0.018 @6 Gy RT (p = 0.003). While irradiation alone induced G2/M cell cycle arrest, the combination of F + RT induced cell redistribution in the G1 phase and produced a significant decrease in the proportion of cells in G2 phase arrest and in the S phase in breast cancer cells (p < 0.01). Furthermore, levels of repair proteins DNA-PKcs and Rad51 were significantly decreased in the cells treated with F + RT compared with irradiation alone. F + RT leads to a decrease in the surviving fraction, increased cell cycle arrest, down regulating of nonhomologous repair protein DNA-PKcs and homologous recombination repair protein RAD51. Thus, our findings suggest that F + RT

  19. Imatinib triggers mesenchymal-like conversion of CML cells associated with increased aggressiveness

    Institute of Scientific and Technical Information of China (English)

    Alexandre Puissant; Yann Chéli; Jill-Patrice Cassuto; Sophie Raynaud; Laurence Legros; Jean-Max Pasquet; Francois-Xavier Mahon; Frédéric Luciano; Patrick Auberger; Maeva Dufies; Nina Fenouille; Issam Ben Sahra; Arnaud Jacquel; Guillaume Robert; Thomas Cluzeau; Marcel Deckert; Mélanie Tichet

    2012-01-01

    Chronic myelogenous leukemia (CML) is a cytogenetic disorder resulting from the expression of p210BCR-ABL Imatinib,an inhibitor of BCR-ABL,has emerged as the leading compound to treat CML patients.Despite encouraging clinical results,resistance to imatinib represents a major drawback for therapy,as a substantial proportion of patients are refractory to this treatment.Recent publications have described the existence of a small cancer cell population with the potential to exhibit the phenotypic switch responsible for chemoresistance.To investigate the existence of such a chemoresistant cellular subpopulation in CML,we used a two-step approach of pulse and continuous selection by imatinib in different CML cell lines that allowed the emergence of a subpopulation of adherent cells (IM-R Adh) displaying an epithelial-mesenchymal transition (EMT)-like phenotype.Overexpression of several EMT markers was observed in this CML subpopulation,as well as in CD34+ CML primary cells from patients who responded poorly to imatinib treatment.In response to imatinib,this CD44high/CD24low IM-R Adh subpopulation exhibited increased adhesion,transmigration and invasion in vitro and in vivo through specific overexpression of the αVβ3 receptor.FAK/Akt pathway activation following integrin β3 (ITGβ3) engagement mediated the migration and invasion of IM-R Adh cells,whereas persistent activation of ERK counteracted BCR-ABL inhibition by imatinib,promoting cell adhesion-mediated resistance.

  20. FOXP3 positive regulatory T-cells in cutaneous and systemic CD30 positive T-cell lymphoproliferations

    DEFF Research Database (Denmark)

    Gjerdrum, Lise Mette; Woetmann, Anders; Ødum, Niels;

    2008-01-01

    -cells (Tregs) may be implicated. In this study, skin biopsies from lymphomatoid papulosis (LyP) (n = 14), primary cutaneous anaplastic large cells lymphoma (C-ALCL) (n = 13) and systemic anaplastic large cells lymphoma (S-ALCL) with (n = 9) or without (n = 6) ALK expression were examined by immunohistology for...... FOXP3 expression in tumour cells and tumour infiltrating Tregs. Labelling of a majority of the neoplastic cells was seen in one case of C-ALCL. Another three cases (one LyP and two C-ALCL) displayed weak labelling of very occasional atypical T-cells. In the remaining 38 cases the atypical lymphoid...... infiltrate was FOXP3 negative. By contrast, all biopsies contained tumour infiltrating FOXP3-positive Tregs. Significant higher numbers were recorded in ALK negative S-ALCL and LyP than in C-ALCL and S-ALCL positive for ALK. In conclusion, it is shown that FOXP3 expression in cutaneous and systemic CD30...

  1. Octreotide scintigraphy localizes somatostatin receptor-positive islet cell carcinomas

    International Nuclear Information System (INIS)

    Tyr-3-octreotide is a synthetic derivative of somatostatin and a somatostatin-receptor analogue. The iodine-123-labelled compound localizes somatostatin-receptor-positive tumours. In this paper two patients are reported in whom somatostatin receptors were demonstrated in vitro. In a 60-year-old female with an islet cell carcinoma of the pancreas, multiple liver metastases and previously uncrecognized bone metastases in the right acetabulum could be diagnosed as the reason for a persistent hypoglycaemia. In a 60-year-old male an islet cell carcinoma of the pancreas was localized with 123I-Tyr-3-octreotide. The somatostatin receptors were demonstrated in vitro and the tumour was successfully treated with somatostatin. These studies demonstrate that 123I-Tyr-3-octreotide offers the possibility of localizing somatostatin-receptor-positive tumours and their metastases. Moreover the method makes it possible to determine the receptor status of a tumour in vivo. (orig.)

  2. Segmentation of breast cancer cells positive 1+ and 3+ immunohistochemistry

    Science.gov (United States)

    Labellapansa, Ause; Muhimmah, Izzati; Indrayanti

    2016-03-01

    Breast cancer is a disease occurs as a result of uncontrolled cells growth. One examination method of breast cancer cells is using Immunohistochemistry (IHC) to determine status of Human Epidermal Growth Factor Receptor2 (HER2) protein. This study helps anatomic pathologist to determine HER2 scores using image processing techniques to obtain HER2 overexpression positive area percentages of 1+ and 3+ scores. This is done because the score of 0 is HER2 negative cells and 2+ scores have equivocal results, which means it could not be determined whether it is necessary to give targeted therapy or not. HER2 overexpression positive area percentage is done by dividing the area with a HER2 positive tumor area. To obtain better tumor area, repair is done by eliminating lymphocytes area which is not tumor area using morphological opening. Results of 10 images IHC scores of 1+ and 3+ and 10 IHC images testing without losing lymphocytes area in tumor area, has proven that the system has been able to provide an overall correct classification in accordance with the experts analysis. However by doing operation to remove non-tumor areas, classification can be done correctly 100% for scores of 3+ and 65% for scores of 1+.

  3. A positive feedback cell signaling nucleation model of astrocyte dynamics

    Directory of Open Access Journals (Sweden)

    Gabriel A Silva

    2013-07-01

    Full Text Available We constructed a model of calcium signaling in astrocyte neural glial cells that incorporates a positive feedback nucleation mechanism, whereby small microdomain increases in local calcium can stochastically produce global cellular and intercellular network scale dynamics. The model is able to simultaneously capture dynamic spatial and temporal heterogeneities associated with intracellular calcium transients in individual cells and intercellular calcium waves (ICW in spatially realistic networks of astrocytes, i.e. networks where the positions of cells were taken from real in vitro experimental data of spontaneously forming sparse networks, as opposed to artificially constructed grid networks or other non-realistic geometries. This is the first work we are aware of where an intracellular model of calcium signaling that reproduces intracellular dynamics inherently accounts for intercellular network dynamics. These results suggest that a nucleation type mechanism should be further investigated experimentally in order to test its contribution to calcium signaling in astrocytes and in other cells more broadly. It may also be of interest in engineered neuromimetic network systems that attempt to emulate biological signaling and information processing properties in synthetic hardwired neuromorphometric circuits or coded algorithms.

  4. Micro-magnet arrays for specific single bacterial cell positioning

    Energy Technology Data Exchange (ETDEWEB)

    Pivetal, Jérémy, E-mail: jeremy.piv@netcmail.com [Ecole Centrale de Lyon, CNRS UMR 5005, Laboratoire Ampère, F-69134 Écully (France); Royet, David [Ecole Centrale de Lyon, CNRS UMR 5005, Laboratoire Ampère, F-69134 Écully (France); Ciuta, Georgeta [Univ. Grenoble Alpes, Inst NEEL, F-38042 Grenoble (France); CNRS, Inst NEEL, F-38042 Grenoble (France); Frenea-Robin, Marie [Université de Lyon, Université Lyon 1, CNRS UMR 5005, Laboratoire Ampère, F-69622 Villeurbanne (France); Haddour, Naoufel [Ecole Centrale de Lyon, CNRS UMR 5005, Laboratoire Ampère, F-69134 Écully (France); Dempsey, Nora M. [Univ. Grenoble Alpes, Inst NEEL, F-38042 Grenoble (France); CNRS, Inst NEEL, F-38042 Grenoble (France); Dumas-Bouchiat, Frédéric [Univ Limoges, CNRS, SPCTS UMR 7513, 12 Rue Atlantis, F-87068 Limoges (France); Simonet, Pascal [Ecole Centrale de Lyon, CNRS UMR 5005, Laboratoire Ampère, F-69134 Écully (France)

    2015-04-15

    In various contexts such as pathogen detection or analysis of microbial diversity where cellular heterogeneity must be taken into account, there is a growing need for tools and methods that enable microbiologists to analyze bacterial cells individually. One of the main challenges in the development of new platforms for single cell studies is to perform precise cell positioning, but the ability to specifically target cells is also important in many applications. In this work, we report the development of new strategies to selectively trap single bacterial cells upon large arrays, based on the use of micro-magnets. Escherichia coli bacteria were used to demonstrate magnetically driven bacterial cell organization. In order to provide a flexible approach adaptable to several applications in the field of microbiology, cells were magnetically and specifically labeled using two different strategies, namely immunomagnetic labeling and magnetic in situ hybridization. Results show that centimeter-sized arrays of targeted, isolated bacteria can be successfully created upon the surface of a flat magnetically patterned hard magnetic film. Efforts are now being directed towards the integration of a detection tool to provide a complete micro-system device for a variety of microbiological applications. - Highlights: 1.We report a new approach to selectively micropattern bacterial cells individually upon micro-magnet arrays. 2.Permanent micro-magnets of a size approaching that of bacteria could be fabricated using a Thermo-Magnetic Patterning process. 3.Bacterial cells were labeled using two different magnetic labeling strategies providing flexible approach adaptable to several applications in the field of microbiology.

  5. Micro-magnet arrays for specific single bacterial cell positioning

    International Nuclear Information System (INIS)

    In various contexts such as pathogen detection or analysis of microbial diversity where cellular heterogeneity must be taken into account, there is a growing need for tools and methods that enable microbiologists to analyze bacterial cells individually. One of the main challenges in the development of new platforms for single cell studies is to perform precise cell positioning, but the ability to specifically target cells is also important in many applications. In this work, we report the development of new strategies to selectively trap single bacterial cells upon large arrays, based on the use of micro-magnets. Escherichia coli bacteria were used to demonstrate magnetically driven bacterial cell organization. In order to provide a flexible approach adaptable to several applications in the field of microbiology, cells were magnetically and specifically labeled using two different strategies, namely immunomagnetic labeling and magnetic in situ hybridization. Results show that centimeter-sized arrays of targeted, isolated bacteria can be successfully created upon the surface of a flat magnetically patterned hard magnetic film. Efforts are now being directed towards the integration of a detection tool to provide a complete micro-system device for a variety of microbiological applications. - Highlights: 1.We report a new approach to selectively micropattern bacterial cells individually upon micro-magnet arrays. 2.Permanent micro-magnets of a size approaching that of bacteria could be fabricated using a Thermo-Magnetic Patterning process. 3.Bacterial cells were labeled using two different magnetic labeling strategies providing flexible approach adaptable to several applications in the field of microbiology

  6. Embryonic and foetal Islet-1 positive cells in human hearts are also positive to c-Kit

    Directory of Open Access Journals (Sweden)

    C. Serradifalco

    2011-12-01

    Full Text Available During embryogenesis, the mammalian heart develops from a primitive heart tube originating from two bilateral primary heart fields located in the lateral plate mesoderm. Cells belongings to the pre-cardiac mesoderm will differentiate into early cardiac progenitors, which express early transcription factors which are also common to the Isl-1 positive cardiac progenitor cells isolated from the developing pharyngeal mesoderm and the foetal and post-natal mice hearts. A second population of cardiac progenitor cells positive to c-Kit has been abundantly isolated from adult hearts. Until now, these two populations have been considered two different sets of progenitor cells present in the heart in different stages of an individual life. In the present study we collected embryonic, foetal and infant hearts, and we tested the hypotheses that c-Kit positive cells, usually isolated from the adult heart, are also present in the intra-uterine life and persist in the adult heart after birth, and that foetal Isl-1 positive cells are also positive to c-Kit. Using immunohistochemistry we studied the temporal distribution of Isl-1 positive and c-Kit/CD105 double positive cells, and by immunofluorescence and confocal analysis we studied the co-localization of c-Kit and Isl-1 positive cells. The results indicated that cardiomyocytes and interstitial cells were positive for c-Kit from the 9th to the 19th gestational week, that cells positive for both c-Kit and CD105 appeared in the interstitium at the 17th gestational week and persisted in the postnatal age, and that the Isl-1 positive cells were a subset of the c-Kit positive population.

  7. On the height of cell aggregates formed with positive dielectrophoresis

    International Nuclear Information System (INIS)

    The influence of a number of parameters on the height of cell aggregates formed by positive dielectrophoresis was systematically investigated. It was found that the aggregate height could be increased by following a number of simple rules. Interdigitated electrodes with oppositely placed castellations gave higher aggregate heights than interdigitated parallel electrodes. The optimal frequency was identified to be 1 MHz. To obtain the highest aggregates, the conductivity of the suspending medium should be kept to the lowest value obtainable, as should the fluid flow rate through the chamber. Aggregate height increased with increasing voltage, but the effect of increasing the voltage diminished as higher aggregate heights were reached. Optima were observed in the aggregate height as a function of the electrode characteristic size, which depended on the cell type and cell size. It was shown to be possible to create aggregate heights of over 150 μm for all the three cell types (bacteria, yeasts and mammalian cells) employed, using voltages of only 20 Vpk-pk

  8. A cell cycle timer for asymmetric spindle positioning.

    Directory of Open Access Journals (Sweden)

    Erin K McCarthy Campbell

    2009-04-01

    Full Text Available The displacement of the mitotic spindle to one side of a cell is important for many cells to divide unequally. While recent progress has begun to unveil some of the molecular mechanisms of mitotic spindle displacement, far less is known about how spindle displacement is precisely timed. A conserved mitotic progression mechanism is known to time events in dividing cells, although this has never been linked to spindle displacement. This mechanism involves the anaphase-promoting complex (APC, its activator Cdc20/Fizzy, its degradation target cyclin, and cyclin-dependent kinase (CDK. Here we show that these components comprise a previously unrecognized timer for spindle displacement. In the Caenorhabditis elegans zygote, mitotic spindle displacement begins at a precise time, soon after chromosomes congress to the metaphase plate. We found that reducing the function of the proteasome, the APC, or Cdc20/Fizzy delayed spindle displacement. Conversely, inactivating CDK in prometaphase caused the spindle to displace early. The consequence of experimentally unlinking spindle displacement from this timing mechanism was the premature displacement of incompletely assembled components of the mitotic spindle. We conclude that in this system, asymmetric positioning of the mitotic spindle is normally delayed for a short time until the APC inactivates CDK, and that this delay ensures that the spindle does not begin to move until it is fully assembled. To our knowledge, this is the first demonstration that mitotic progression times spindle displacement in the asymmetric division of an animal cell. We speculate that this link between the cell cycle and asymmetric cell division might be evolutionarily conserved, because the mitotic spindle is displaced at a similar stage of mitosis during asymmetric cell divisions in diverse systems.

  9. Impaired DNA replication within progenitor cell pools promotes leukemogenesis.

    Directory of Open Access Journals (Sweden)

    Ganna Bilousova

    2005-12-01

    Full Text Available Impaired cell cycle progression can be paradoxically associated with increased rates of malignancies. Using retroviral transduction of bone marrow progenitors followed by transplantation into mice, we demonstrate that inhibition of hematopoietic progenitor cell proliferation impairs competition, promoting the expansion of progenitors that acquire oncogenic mutations which restore cell cycle progression. Conditions that impair DNA replication dramatically enhance the proliferative advantage provided by the expression of Bcr-Abl or mutant p53, which provide no apparent competitive advantage under conditions of healthy replication. Furthermore, for the Bcr-Abl oncogene the competitive advantage in contexts of impaired DNA replication dramatically increases leukemogenesis. Impaired replication within hematopoietic progenitor cell pools can select for oncogenic events and thereby promote leukemia, demonstrating the importance of replicative competence in the prevention of tumorigenesis. The demonstration that replication-impaired, poorly competitive progenitor cell pools can promote tumorigenesis provides a new rationale for links between tumorigenesis and common human conditions of impaired DNA replication such as dietary folate deficiency, chemotherapeutics targeting dNTP synthesis, and polymorphisms in genes important for DNA metabolism.

  10. Umbilical Cord Blood Transplant, Cyclophosphamide, Fludarabine Phosphate, and Total-Body Irradiation in Treating Patients With Hematologic Disease

    Science.gov (United States)

    2016-04-18

    Acute Biphenotypic Leukemia; Acute Myeloid Leukemia Arising From Previous Myelodysplastic Syndrome; Acute Myeloid Leukemia in Remission; Adult Acute Lymphoblastic Leukemia in Complete Remission; Aggressive Non-Hodgkin Lymphoma; B Acute Lymphoblastic Leukemia With t(1;19)(q23;p13.3); E2A-PBX1 (TCF3-PBX1); B Acute Lymphoblastic Leukemia With t(9;22)(q34;q11.2); BCR-ABL1; Burkitt Lymphoma; Childhood Acute Lymphoblastic Leukemia in Complete Remission; Chronic Phase Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Lymphoblastic Lymphoma; Mantle Cell Lymphoma; Myelofibrosis; Pancytopenia; Plasma Cell Myeloma; Prolymphocytic Leukemia; Recurrent Childhood Acute Myeloid Leukemia; Recurrent Chronic Lymphocytic Leukemia; Recurrent Chronic Myelogenous Leukemia, BCR-ABL1 Positive; Recurrent Follicular Lymphoma; Recurrent Lymphoplasmacytic Lymphoma; Recurrent Marginal Zone Lymphoma; Recurrent Small Lymphocytic Lymphoma; Refractory Anemia With Excess Blasts

  11. Small molecule ErbB inhibitors decrease proliferative signaling and promote apoptosis in philadelphia chromosome-positive acute lymphoblastic leukemia.

    Directory of Open Access Journals (Sweden)

    Mary E Irwin

    Full Text Available The presence of the Philadelphia chromosome in patients with acute lymphoblastic leukemia (Ph(+ALL is a negative prognostic indicator. Tyrosine kinase inhibitors (TKI that target BCR/ABL, such as imatinib, have improved treatment of Ph(+ALL and are generally incorporated into induction regimens. This approach has improved clinical responses, but molecular remissions are seen in less than 50% of patients leaving few treatment options in the event of relapse. Thus, identification of additional targets for therapeutic intervention has potential to improve outcomes for Ph+ALL. The human epidermal growth factor receptor 2 (ErbB2 is expressed in ~30% of B-ALLs, and numerous small molecule inhibitors are available to prevent its activation. We analyzed a cohort of 129 ALL patient samples using reverse phase protein array (RPPA with ErbB2 and phospho-ErbB2 antibodies and found that activity of ErbB2 was elevated in 56% of Ph(+ALL as compared to just 4.8% of Ph(-ALL. In two human Ph+ALL cell lines, inhibition of ErbB kinase activity with canertinib resulted in a dose-dependent decrease in the phosphorylation of an ErbB kinase signaling target p70S6-kinase T389 (by 60% in Z119 and 39% in Z181 cells at 3 µM. Downstream, phosphorylation of S6-kinase was also diminished in both cell lines in a dose-dependent manner (by 91% in both cell lines at 3 µM. Canertinib treatment increased expression of the pro-apoptotic protein Bim by as much as 144% in Z119 cells and 49% in Z181 cells, and further produced caspase-3 activation and consequent apoptotic cell death. Both canertinib and the FDA-approved ErbB1/2-directed TKI lapatinib abrogated proliferation and increased sensitivity to BCR/ABL-directed TKIs at clinically relevant doses. Our results suggest that ErbB signaling is an additional molecular target in Ph(+ALL and encourage the development of clinical strategies combining ErbB and BCR/ABL kinase inhibitors for this subset of ALL patients.

  12. Ocular adnexal marginal zone B cell lymphoma infiltrated by IgG4-positive plasma cells

    OpenAIRE

    KUBOTA, TOSHINOBU; Moritani, Suzuko; Yoshino, Tadashi; Nagai, Hirokazu; Terasaki, Hiroko

    2010-01-01

    Aims To report the clinicopathological characteristics of patients with ocular adnexal marginal zone B cell lymphoma (MZBL) with IgG4-positive plasma cells. Methods 114 biopsy samples of ocular adnexal MZBLs were analysed. MZBLs with IgG4-positive plasma cells were included when the IgG4:IgG ratio was >40% (IgG4-related group). The serum levels of each subclass of immunoglobulins and soluble interleukin-2 receptor in the IgG4-related group were compared with those in 61 consecutive patients h...

  13. False positive paediatric labelled white blood cell study

    International Nuclear Information System (INIS)

    Full text: An eight-month-old female presented for a technetium labelled white blood cell study (LWBC) to exclude an intra-abdominal abscess. Born premature, the child had surgery to repair a perforated bowel and had repeated presentations with diarrhoea, fevers, a tender right upper quadrant and a raised leucocyte count. Multiple imaging modalities failed to demonstrate recurrent bowel perforation, ischaemia or an intra-abdominal mass. A LWBC study was performed with whole body imaging at 1 and 5 hours post re-injection of the radiolabelled blood. No abnormal uptake was visualised in the abdomen but abnormal white cell accumulation was noted in the right hind foot and the length of the right lower leg. This activity appeared to lie along the course of the right tibia. Plain X-ray demonstrated no evidence of tibial osteomyelitis. Concern that the LWBC may be falsely negative in a patient on antibiotics, a gallium scan was immediately performed to re-examine the abdomen. The whole body gallium images demonstrated normal physiological uptake in the abdomen and no evidence of infection in the right leg. The patient had no clinical features to support right leg pathology. The abnormal LWBC localisation in the right lower leg/foot was therefore falsely positive. The most likely explanation is increased activation of the autologous LWBC by 'rough' handling during difficult venesection and re-injection through small veins and needles/cannulas. The slow flow through the veins draining the foot injection site would contribute to margination in these vessel walls. This is a potential cause for false positive LWBC studies- with significant implications for patient care. Copyright (2002) The Australian and New Zealand Society of Nuclear Medicine Inc

  14. Identification of uPAR-positive chemoresistant cells in small cell lung cancer.

    Directory of Open Access Journals (Sweden)

    Margarita Gutova

    Full Text Available BACKGROUND: The urokinase plasminogen activator (uPA and its receptor (uPAR/CD87 are major regulators of extracellular matrix degradation and are involved in cell migration and invasion under physiological and pathological conditions. The uPA/uPAR system has been of great interest in cancer research because it is involved in the development of most invasive cancer phenotypes and is a strong predictor of poor patient survival. However, little is known about the role of uPA/uPAR in small cell lung cancer (SCLC, the most aggressive type of lung cancer. We therefore determined whether uPA and uPAR are involved in generation of drug resistant SCLC cell phenotype. METHODS AND FINDINGS: We screened six human SCLC cell lines for surface markers for putative stem and cancer cells. We used fluorescence-activated cell sorting (FACS, fluorescence microscopy and clonogenic assays to demonstrate uPAR expression in a subpopulation of cells derived from primary and metastatic SCLC cell lines. Cytotoxic assays were used to determine the sensitivity of uPAR-positive and uPAR-negative cells to chemotherapeutic agents. The uPAR-positive cells in all SCLC lines demonstrated multi-drug resistance, high clonogenic activity and co-expression of CD44 and MDR1, putative cancer stem cell markers. CONCLUSIONS: These data suggest that uPAR-positive cells may define a functionally important population of cancer cells in SCLC, which are resistant to traditional chemotherapies, and could serve as critical targets for more effective therapeutic interventions in SCLC.

  15. Plastid position in Arabidopsis columella cells is similar in microgravity and on a random-positioning machine

    Science.gov (United States)

    Kraft, T. F.; van Loon, J. J.; Kiss, J. Z.

    2000-01-01

    In order to study gravity effects on plant structure and function, it may become necessary to remove the g-stimulus. On Earth, various instruments such as clinostats have been used by biologists in an attempt to neutralize the effects of gravity. In this study, the position of amyloplasts was assayed in columella cells in the roots of Arabidopsis thaliana (L.) Heynh. seedlings grown in the following conditions: on Earth, on a two-dimensional clinostat at 1 rpm, on a three-dimensional clinostat (also called a random-positioning machine, or an RPM), and in space (true microgravity). In addition, the effects of these gravity treatments on columella cell area and plastid area also were measured. In terms of the parameters measured, only amyloplast position was affected by the gravity treatments. Plastid position was not significantly different between spaceflight and RPM conditions but was significantly different between spaceflight and the classical two-dimensional clinostat treatments. Flanking columella cells showed a greater susceptibility to changes in gravity compared to the central columella cells. In addition, columella cells of seedlings that were grown on the RPM did not exhibit deleterious effects in terms of their ultrastructure as has been reported previously for seedlings grown on a two-dimensional clinostat. This study supports the hypothesis that the RPM provides a useful simulation of weightlessness.

  16. GVL against mouse BC-CML and CP-CML: Shared mechanisms of T cell killing, but PD-ligands render CP-CML and not BC-CML GVL-resistant*

    OpenAIRE

    Matte-Martone, Catherine; Venkatesan, Srividhya; Tan, Hung Sheng; Athanasiadis, Ioanna; Chang, Julia; Pavisic, Jovana; Shlomchik, Warren

    2011-01-01

    GVL against chronic phase CML (CP-CML) is potent, but it is less efficacious against acute leukemias and blast crisis CML (BC-CML). The mechanisms underlying GVL-resistance are unknown. Previously, we found that alloreactive T cell targeting of GVL-sensitive bcr-abl-induced mouse CP-CML (mCP-CML) required TCR:MHC interactions and that multiple and redundant killing mechanisms were in play. To better understand why BC-CML is resistant to GVL, we performed a comprehensive analysis of GVL agains...

  17. Molecular pathways of early CD105-positive erythroid cells as compared with CD34-positive common precursor cells by flow cytometric cell-sorting and gene expression profiling

    International Nuclear Information System (INIS)

    Special attention has recently been drawn to the molecular network of different genes that are responsible for the development of erythroid cells. The aim of the present study was to establish in detail the immunophenotype of early erythroid cells and to compare the gene expression profile of freshly isolated early erythroid precursors with that of the CD34-positive (CD34+) compartment. Multiparameter flow cytometric analyses of human bone marrow mononuclear cell fractions (n=20) defined three distinct early erythroid stages. The gene expression profile of sorted early erythroid cells was analyzed by Affymetrix array technology. For 4524 genes, a differential regulation was found in CD105-positive erythroid cells as compared with the CD34+ progenitor compartment (2362 upregulated genes). A highly significant difference was observed in the expression level of genes involved in transcription, heme synthesis, iron and mitochondrial metabolism and transforming growth factor-β signaling. A comparison with recently published data showed over 1000 genes that as yet have not been reported to be upregulated in the early erythroid lineage. The gene expression level within distinct pathways could be illustrated directly by applying the Ingenuity software program. The results of gene expression analyses can be seen at the Gene Expression Omnibus repository

  18. Synthetic Quorum Sensing and Cell-Cell Communication in Gram-Positive Bacillus megaterium.

    Science.gov (United States)

    Marchand, Nicholas; Collins, Cynthia H

    2016-07-15

    The components of natural quorum-sensing (QS) systems can be used to engineer synthetic communication systems that regulate gene expression in response to chemical signals. We have used the machinery from the peptide-based agr QS system from Staphylococcus aureus to engineer a synthetic QS system in Bacillus megaterium to enable autoinduction of a target gene at high cell densities. Growth and gene expression from these synthetic QS cells were characterized in both complex and minimal media. We also split the signal production and sensing components between two strains of B. megaterium to produce sender and receiver cells and characterized the resulting communication in liquid media and on semisolid agar. The system described in this work represents the first synthetic QS and cell-cell communication system that has been engineered to function in a Gram-positive host, and it has the potential to enable the generation of dynamic gene regulatory networks in B. megaterium and other Gram-positive organisms. PMID:26203497

  19. ADAR1 Activation Drives Leukemia Stem Cell Self-Renewal by Impairing Let-7 Biogenesis.

    Science.gov (United States)

    Zipeto, Maria Anna; Court, Angela C; Sadarangani, Anil; Delos Santos, Nathaniel P; Balaian, Larisa; Chun, Hye-Jung; Pineda, Gabriel; Morris, Sheldon R; Mason, Cayla N; Geron, Ifat; Barrett, Christian; Goff, Daniel J; Wall, Russell; Pellecchia, Maurizio; Minden, Mark; Frazer, Kelly A; Marra, Marco A; Crews, Leslie A; Jiang, Qingfei; Jamieson, Catriona H M

    2016-08-01

    Post-transcriptional adenosine-to-inosine RNA editing mediated by adenosine deaminase acting on RNA1 (ADAR1) promotes cancer progression and therapeutic resistance. However, ADAR1 editase-dependent mechanisms governing leukemia stem cell (LSC) generation have not been elucidated. In blast crisis chronic myeloid leukemia (BC CML), we show that increased JAK2 signaling and BCR-ABL1 amplification activate ADAR1. In a humanized BC CML mouse model, combined JAK2 and BCR-ABL1 inhibition prevents LSC self-renewal commensurate with ADAR1 downregulation. Lentiviral ADAR1 wild-type, but not an editing-defective ADAR1(E912A) mutant, induces self-renewal gene expression and impairs biogenesis of stem cell regulatory let-7 microRNAs. Combined RNA sequencing, qRT-PCR, CLIP-ADAR1, and pri-let-7 mutagenesis data suggest that ADAR1 promotes LSC generation via let-7 pri-microRNA editing and LIN28B upregulation. A small-molecule tool compound antagonizes ADAR1's effect on LSC self-renewal in stromal co-cultures and restores let-7 biogenesis. Thus, ADAR1 activation represents a unique therapeutic vulnerability in LSCs with active JAK2 signaling. PMID:27292188

  20. CD163 positive subsets of blood dendritic cells

    DEFF Research Database (Denmark)

    Maniecki, Maciej Bogdan; Møller, Holger Jon; Moestrup, Søren Kragh; Møller, Bjarne Kuno

    CD163 and CD91 are scavenging receptors with highly increased expression during the differentiation of monocytes into the anti-inflammatory macrophage phenotype. In addition, CD91 is expressed in monocyte-derived dendritic cells (MoDCs), where the receptor is suggested to be important for...... internalization of CD91-targeted antigens to be presented on the dendritic cell surface for T-cell stimulation. Despite their overlap in functionality, the expression of CD91 and CD163 has never been compared and the expression of CD163 in the monocyte-dendritic cell lineage is not yet characterized. CD163...... expression in dendritic cells (DCs) was investigated using multicolor flow cytometry in peripheral blood from 31 healthy donors and 15 HIV-1 patients in addition to umbilical cord blood from 5 newborn infants. Total RNA was isolated from MACS purified DCs and CD163 mRNA was determined with real-time reverse...

  1. Conductivity and Dielectric Dispersion of Gram-Positive Bacterial Cells

    Science.gov (United States)

    van der Wal A; Minor; Norde; Zehnder; Lyklema

    1997-02-01

    The conductivity of bacterial cell suspensions has been studied over a wide range of ionic strengths and is interpreted in terms of their cell wall properties. The experimental data have been analyzed after improving the high kappaa double-layer theory of Fixman, by accounting for ionic mobility in the hydrodynamically stagnant layer, i.e., in the bacterial wall. Static conductivity and dielectric dispersion measurements both show that the counterions in the porous gel-like cell wall give rise to a considerable surface conductance. From a comparison of the mobile charge with the total cell wall charge it is inferred that the mobilities of the ions in the bacterial wall are of the same order but somewhat lower than those in the bulk electrolyte solution. The occurrence of surface conductance reduces the electrophoretic mobility in electrophoresis studies. If this effect is not taken into account, the zeta-potential will be underestimated, especially at low electrolyte concentrations. PMID:9056304

  2. Predomination of IL-17-producing tryptase-positive/chymase-positive mast cells in azoospermic chronic testicular inflammation.

    Science.gov (United States)

    Chen, S-J; Duan, Y-G; Haidl, G; Allam, J-P

    2016-08-01

    Chronic testicular inflammation and infection have been regarded as important factors in the pathogenesis of azoospermia. As key effector cells in innate and adaptive immune system, mast cells (MCs) were observed in inflammation and autoimmune disease. Furthermore, increased expression of tryptase-positive MCs has been reported in testicular disorders associated with male infertility/subfertility. However, little is known about the potential relationship between MCs and chronic testicular inflammation in azoospermic patients. Moreover, the preferential expression of MCs' subtypes in testis of these patients is still far from being understood. Thus, this study aimed to investigate characteristics of testicular MCs as well as their subtypes in azoospermic men with chronic testicular inflammation (AZI, n = 5) by immunohistochemical techniques. Our results showed significant increase of MCs in AZI, and more importantly, considerable numbers of tryptase-positive/chymase-positive MCs could also be demonstrated in AZI, when compared to control groups representing azoospermia without chronic testicular inflammation (AZW, n = 5) and normal spermatogenesis (NT, n = 5) respectively. Most interestingly, immunofluorescence staining revealed autoimmune-associated interleukin (IL)-17-producing MCs in AZI, whereas co-expression of MC markers with tumour necrosis factor (TNF)-α, IL-10 and IL-1β could not be detected. In conclusion, AZI is associated with significant increase of tryptase-positive/chymase-positive MCs expressing IL-17, and these MCs might contribute to the pathogenesis of AZI. PMID:26420243

  3. Radiation-induced apoptosis in differentially modulated by PTK inhibitora in K562 cells

    International Nuclear Information System (INIS)

    The effect of PTK inhibitors (herbimycin A and genistein) on the induction of radiation-induced apoptosis in Ph-positive K562 leukemia cell line was investigated. K562 cells in exponential growth phase were irradiated with a linear accelerator at room temperature. For 6 MV X-ray irradiation and drug treatment, cultures were initiated at 2x106 cells/ml. The cells were irradiated with 10Gy. Stock solutions of herbimycin A and genistein were prepared in dimethyl sulphoxide (DMSO). After incubation at 37 .deg. for 0-48 h, the extent of apoptosis was determined using agarose gel electrophoresis and TUNEL assay. The progression of cells through the cell cycle after irradiation and drug treatment was also determined with flow cytometry. Western blot analysis was used to monitor bcl-2, bcl-X-L and bax protein levels. Treatment with 10 Gy X-irradiation did not result in the induction of apoptosis. The HMA alone (500 nM) also failed to induce apoptosis. By contrast, incubation of K562 cells with HMA after irradiation resulted in a substantial induction of nuclear condensation and fragmentation by agarose gel electrophoresis and TUNEL assay. Genistein failed to enhance the ability of X-irradiation to induce DNA fragmentation. Enhancement of apoptosis by HMA was not attributable to downregulation of the bcl-2 or bcl-X-L anti-apoptotic proteins. When the cells were irradiated and maintained with HMA, the percentage of cells in G2/M phase decreased to 30-40% at 48 h. On the other hand, cells exposed to 10 Gy X-irradiation alone or maintained with genistein did not show marked cell cycle redistribution. We have shown that nanomolar concentrations of the PTK inhibitor HMA synergize with X-irradiation in inducing the apoptosis in Ph (+) K562 leukemia cell line. While, genistein, a PTK inhibitor which is not selective for p210bcr/abl failed to enhance the radiation induced apoptosis in K562 cells. It is unlikely that the ability of HMA to enhance apoptosis in K562 cells is

  4. The Culture Repopulation Ability (CRA) Assay and Incubation in Low Oxygen to Test Antileukemic Drugs on Imatinib-Resistant CML Stem-Like Cells.

    Science.gov (United States)

    Cheloni, Giulia; Tanturli, Michele

    2016-01-01

    Chronic myeloid leukemia (CML) is a stem cell-driven disorder caused by the BCR/Abl oncoprotein, a constitutively active tyrosine kinase (TK). Chronic-phase CML patients are treated with impressive efficacy with TK inhibitors (TKi) such as imatinib mesylate (IM). However, rather than definitively curing CML, TKi induces a state of minimal residual disease, due to the persistence of leukemia stem cells (LSC) which are insensitive to this class of drugs. LSC persistence may be due to different reasons, including the suppression of BCR/Abl oncoprotein. It has been shown that this suppression follows incubation in low oxygen under appropriate culture conditions and incubation times.Here we describe the culture repopulation ability (CRA) assay, a non-clonogenic assay capable - together with incubation in low oxygen - to reveal in vitro stem cells endowed with marrow repopulation ability (MRA) in vivo. The CRA assay can be used, before moving to animal tests, as a simple and reliable method for the prescreening of drugs potentially active on CML and other leukemias with respect to their activity on the more immature leukemia cell subsets. PMID:27581140

  5. CD4+/CD8+ double-positive T cells

    DEFF Research Database (Denmark)

    Overgaard, Nana H; Jung, Ji-Won; Steptoe, Raymond J;

    2015-01-01

    lymphoid tissues of numerous species, as well as in numerous disease settings, including cancer. The expression of CD4 and CD8 is regulated by a very strict transcriptional program involving the transcription factors Runx3 and ThPOK. Initially thought to be mutually exclusive within CD4(+) and CD8(+) T...... reports describing cytotoxic or suppressive roles for these cells. In this review, we describe how transcriptional regulation, lineage of origin, heterogeneity of CD4 and CD8 expression, age, species, and specific disease settings influence the functionality of this rarely studied T cell population....

  6. Clinical Utility of Circulating Tumor Cells in ALK-Positive Non-Small-Cell Lung Cancer.

    Science.gov (United States)

    Faugeroux, Vincent; Pailler, Emma; Auger, Nathalie; Taylor, Melissa; Farace, Françoise

    2014-01-01

    The advent of rationally targeted therapies such as small-molecule tyrosine kinase inhibitors (TKIs) has considerably transformed the therapeutic management of a subset of patients with non-small-cell lung cancer (NSCLC) harboring defined molecular abnormalities. When such genetic molecular alterations are detected the use of specific TKI has demonstrated better results (overall response rate, progression free survival) compared to systemic therapy. However, the detection of such molecular abnormalities is complicated by the difficulty in obtaining sufficient tumor material, in terms of quantity and quality, from a biopsy. Here, we described how circulating tumor cells (CTCs) can have a clinical utility in anaplastic lymphoma kinase (ALK) positive NSCLC patients to diagnose ALK-EML4 gene rearrangement and to guide therapeutic management of these patients. The ability to detect genetic abnormalities such ALK rearrangement in CTCs shows that these cells could offer new perspectives both for the diagnosis and the monitoring of ALK-positive patients eligible for treatment with ALK inhibitors. PMID:25414829

  7. Clinical Utility of Circulating Tumor Cells in ALK-Positive Non-Small-Cell Lung Cancer

    Science.gov (United States)

    Faugeroux, Vincent; Pailler, Emma; Auger, Nathalie; Taylor, Melissa; Farace, Françoise

    2014-01-01

    The advent of rationally targeted therapies such as small-molecule tyrosine kinase inhibitors (TKIs) has considerably transformed the therapeutic management of a subset of patients with non-small-cell lung cancer (NSCLC) harboring defined molecular abnormalities. When such genetic molecular alterations are detected the use of specific TKI has demonstrated better results (overall response rate, progression free survival) compared to systemic therapy. However, the detection of such molecular abnormalities is complicated by the difficulty in obtaining sufficient tumor material, in terms of quantity and quality, from a biopsy. Here, we described how circulating tumor cells (CTCs) can have a clinical utility in anaplastic lymphoma kinase (ALK) positive NSCLC patients to diagnose ALK-EML4 gene rearrangement and to guide therapeutic management of these patients. The ability to detect genetic abnormalities such ALK rearrangement in CTCs shows that these cells could offer new perspectives both for the diagnosis and the monitoring of ALK-positive patients eligible for treatment with ALK inhibitors. PMID:25414829

  8. The equatorial position of the metaphase plate ensures symmetric cell divisions.

    Science.gov (United States)

    Tan, Chia Huei; Gasic, Ivana; Huber-Reggi, Sabina P; Dudka, Damian; Barisic, Marin; Maiato, Helder; Meraldi, Patrick

    2015-01-01

    Chromosome alignment in the middle of the bipolar spindle is a hallmark of metazoan cell divisions. When we offset the metaphase plate position by creating an asymmetric centriole distribution on each pole, we find that metaphase plates relocate to the middle of the spindle before anaphase. The spindle assembly checkpoint enables this centering mechanism by providing cells enough time to correct metaphase plate position. The checkpoint responds to unstable kinetochore-microtubule attachments resulting from an imbalance in microtubule stability between the two half-spindles in cells with an asymmetric centriole distribution. Inactivation of the checkpoint prior to metaphase plate centering leads to asymmetric cell divisions and daughter cells of unequal size; in contrast, if the checkpoint is inactivated after the metaphase plate has centered its position, symmetric cell divisions ensue. This indicates that the equatorial position of the metaphase plate is essential for symmetric cell divisions. PMID:26188083

  9. Increased numbers of P63-positive/CD117-positive cells in advanced adenoid cystic carcinoma give a poorer prognosis

    Directory of Open Access Journals (Sweden)

    Zhou Quan

    2012-09-01

    Full Text Available Abstract Objectives This study consisted of two parts. One part was to analyze the survival rates of adenoid cystic carcinoma (ACC in Chinese and explain the difference between our data and the literature. The other was to analyze the relationship between the expression of CD117 and the histological grade and the prognosis. Methods A retrospective study of 80 ACC patients was performed. Clinical data were collected, and p63, CD117 were detected by immunohistochemical staining. Results Eighty patients received follow-ups 3 to 216 months after initial diagnosis. ACC occurred in the lacrimal gland (26.3%, n = 21, nasal cavity and parasinus (33.8%, n = 27 and other sites (40.0%, n = 33. The 5-year and 10-year survival rates were 66.41% and 10.16%, respectively. Over expression of CD117 was detected in p63-negative cells in 94.3% of cases and in p63-positive cells in 45.8%. The expression of CD117 in p63-positive cells was significantly associated with the histological grade (P Conclusions ACC had a good 5-year survival but poor 10-year survival in Chinese, which differed from the occidental data. More p63+/CD117+ cells were associated with a higher histological grade and poorer outcome. Virtual slides The virtual slide(s for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1701457278762097

  10. Human papillomavirus 52 positive squamous cell carcinoma of the conjunctiva

    Directory of Open Access Journals (Sweden)

    Silvia Olivia Salceanu

    2015-01-01

    Full Text Available Human papillomavirus (HPV infection is strongly associated with several human cancers; the most known genotypes involved being HPV 16 and HPV 18. We report the detection of HPV 52 in a sample taken from a 47-year-old patient with squamous cell carcinoma of the conjunctiva of the left eye. The method used for the detection of HPV was real time polymerase chain reaction. The evolution was favorable after surgical removal of the tumor and the patient was explained that long-term follow-up is essential to avoid recurrence.

  11. T cells expressing CD19-specific Engager Molecules for the Immunotherapy of CD19-positive Malignancies

    OpenAIRE

    Mireya Paulina Velasquez; David Torres; Kota Iwahori; Sunitha Kakarla; Caroline Arber; Tania Rodriguez-Cruz; Arpad Szoor; Bonifant, Challice L.; Claudia Gerken; Cooper, Laurence J.N.; Xiao-Tong Song; Stephen Gottschalk

    2016-01-01

    T cells expressing chimeric antigen receptors (CARs) or the infusion of bispecific T-cell engagers (BITEs) have shown antitumor activity in humans for CD19-positive malignancies. While BITEs redirect the large reservoir of resident T cells to tumors, CAR T cells rely on significant in vivo expansion to exert antitumor activity. We have shown that it is feasible to modify T cells to secrete solid tumor antigen-specific BITEs, enabling T cells to redirect resident T cells to tumor cells. To ada...

  12. Positional information is reprogrammed in blastema cells of the regenerating limb of the axolotl (Ambystoma mexicanum.

    Directory of Open Access Journals (Sweden)

    Catherine D McCusker

    Full Text Available The regenerating region of an amputated salamander limb, known as the blastema, has the amazing capacity to replace exactly the missing structures. By grafting cells from different stages and regions of blastemas induced to form on donor animals expressing Green Fluorescent Protein (GFP, to non-GFP host animals, we have determined that the cells from early stage blastemas, as well as cells at the tip of late stage blastemas are developmentally labile such that their positional identity is reprogrammed by interactions with more proximal cells with stable positional information. In contrast, cells from the adjacent, more proximal stump tissues as well as the basal region of late bud blastemas are positionally stable, and thus form ectopic limb structures when grafted. Finally, we have found that a nerve is required to maintain the blastema cells in a positionally labile state, thus indicating a role for reprogramming cues in the blastema microenvironment.

  13. Increased intratumoral FOXP3-positive regulatory immune cells during interleukin-2 treatment in metastatic renal cell carcinoma

    DEFF Research Database (Denmark)

    Jensen, Hanne Krogh; Donskov, Frede; Nordsmark, Marianne; Marcussen, Niels; von der Maase, Hans

    2009-01-01

    PURPOSE: The administration of interleukin-2 (IL-2) may increase the frequency of peripherally circulating FOXP3-positive regulatory immune cells, thus potentially compromising this treatment option for patients with metastatic renal cell carcinoma. The impact of IL-2-based therapy on the...... accumulation of FOXP3-positive immune cells in the tumor microenvironment in metastatic renal cell carcinoma is unknown. EXPERIMENTAL DESIGN: Baseline (n = 58) and on-treatment (n = 42) tumor core biopsies were prospectively obtained from patients with clear cell metastatic renal cell carcinoma before and...... during IL-2-based immunotherapy. Immunohistochemical expression of FOXP3 was estimated by stereological counting technique and correlated with other immune cell subsets and overall survival. RESULTS: A significant increase in absolute intratumoral FOXP3-positive immune cells was observed comparing...

  14. Computational model of cell positioning: directed and collective migration in the intestinal crypt epithelium

    OpenAIRE

    Wong, Shek Yoon; Chiam, K.-H.; Lim, Chwee Teck; Matsudaira, Paul

    2010-01-01

    The epithelium of the intestinal crypt is a dynamic tissue undergoing constant regeneration through cell growth, cell division, cell differentiation and apoptosis. How the epithelial cells maintain correct positioning and how they migrate in a directed and collective fashion are still not well understood. In this paper, we developed a computational model to elucidate these processes. We show that differential adhesion between epithelial cells, caused by the differential activation of EphB rec...

  15. Bacterial glycobiology: rhamnose-containing cell wall polysaccharides in Gram-positive bacteria.

    OpenAIRE

    Mistou, Michel-Yves; Sutcliffe, Iain; van Sorge, Nina

    2016-01-01

    The composition of the Gram-positive cell wall is typically described as containing peptidoglycan, proteins and essential secondary cell wall structures called teichoic acids, which comprise approximately half of the cell wall mass. The cell walls of many species within the genera Streptococcus, Enterococcus and Lactococcus contain large amounts of the sugar rhamnose, which is incorporated in cell wall-anchored polysaccharides (CWP) that possibly function as homologues of well-studied wall te...

  16. Cell wall sorting signals in surface proteins of gram-positive bacteria.

    OpenAIRE

    Schneewind, O; Mihaylova-Petkov, D; Model, P

    1993-01-01

    Staphylococcal protein A is anchored to the cell wall, a unique cellular compartment of Gram-positive bacteria. The sorting signal sufficient for cell wall anchoring consists of an LPXTG motif, a C-terminal hydrophobic domain and a charged tail. Homologous sequences are found in many surface proteins of Gram-positive bacteria and we explored the universality of these sequences to serve as cell wall sorting signals. We show that several signals are able to anchor fusion proteins to the staphyl...

  17. Nestin expression in osteosarcomas and derivation of nestin/CD133 positive osteosarcoma cell lines

    International Nuclear Information System (INIS)

    Nestin was originally identified as a class VI intermediate filament protein that is expressed in stem cells and progenitor cells in the mammalian CNS during development. This protein is replaced in the adult organism by other intermediate filament proteins; however, nestin may be re-expressed under certain pathological conditions such as ischemia, inflammation, brain injury, and neoplastic transformation. Nestin has been detected in many kinds of tumors, especially in tumors derived from the CNS. Co-expression of nestin and the CD133 surface molecule is considered to be a marker for cancer stem cells in neurogenic tumors. Our work was aimed at a detailed study of nestin expression in osteosarcomas and osteosarcoma-derived cell lines. Using immunodetection methods, we examined nestin in tumor tissue samples from 18 patients with osteosarcomas. We also successfully established permanent cell lines from the tumor tissue of 4 patients and immunodetection of nestin and CD133 was performed on these cell lines. Nestin-positive tumor cells were immunohistochemically detected in all of the examined osteosarcomas, but the proportion of these cells that were positively stained as well as the intensity of staining varied. Nestin-positive cells were rarely observed in 2 tumor samples, and the remaining 16 tumor samples showed various nestin expression patterns ranging from very sporadic occurrence to an overwhelming proportion of cells with strong positive staining. Three of the established osteosarcoma cell lines were demonstrated to be nestin-positive, and only one cell line showed no expression of nestin; this finding corresponds with the rare occurrence of nestin-positive cells in the respective tumor sample. Moreover, three of these osteosarcoma cell lines were undoubtedly proven to be Nes+/CD133+. Our results represent the first evidence of nestin expression in osteosarcomas and suggest the possible occurrence of cells with a stem-like phenotype in these tumors

  18. Deletions of Immunoglobulin heavy chain and T cell receptor gene regions are uniquely associated with lymphoid blast transformation of chronic myeloid leukemia

    Directory of Open Access Journals (Sweden)

    Valgañon Mikel

    2010-01-01

    Full Text Available Abstract Background Chronic myelogenous leukemia (CML results from the neoplastic transformation of a haematopoietic stem cell. The hallmark genetic abnormality of CML is a chimeric BCR/ABL1 fusion gene resulting from the Philadelphia chromosome rearrangement t(9;22(q34;q11. Clinical and laboratory studies indicate that the BCR/ABL1 fusion protein is essential for initiation, maintenance and progression of CML, yet the event(s driving the transformation from chronic phase to blast phase are poorly understood. Results Here we report multiple genome aberrations in a collection of 78 CML and 14 control samples by oligonucleotide array comparative genomic hybridization. We found a unique signature of genome deletions within the immunoglobulin heavy chain (IGH and T cell receptor regions (TCR, frequently accompanied by concomitant loss of sequences within the short arm regions of chromosomes 7 and 9, including IKZF1, HOXA7, CDKN2A/2B, MLLT3, IFNA/B, RNF38, PAX5, JMJD2C and PDCD1LG2 genes. Conclusions None of these genome losses were detected in any of the CML samples with myeloid transformation, chronic phase or controls, indicating that their presence is obligatory for the development of a malignant clone with a lymphoid phenotype. Notably, the coincidental deletions at IGH and TCR regions appear to precede the loss of IKZF1 and/or p16 genes in CML indicating a possible involvement of RAG in these deletions.

  19. Cell shape impacts on the positioning of the mitotic spindle with respect to the substratum.

    Science.gov (United States)

    Lázaro-Diéguez, Francisco; Ispolatov, Iaroslav; Müsch, Anne

    2015-04-01

    All known mechanisms of mitotic spindle orientation rely on astral microtubules. We report that even in the absence of astral microtubules, metaphase spindles in MDCK and HeLa cells are not randomly positioned along their x-z dimension, but preferentially adopt shallow β angles between spindle pole axis and substratum. The nonrandom spindle positioning is due to constraints imposed by the cell cortex in flat cells that drive spindles that are longer and/or wider than the cell's height into a tilted, quasidiagonal x-z position. In rounder cells, which are taller, fewer cortical constraints make the x-z spindle position more random. Reestablishment of astral microtubule-mediated forces align the spindle poles with cortical cues parallel to the substratum in all cells. However, in flat cells, they frequently cause spindle deformations. Similar deformations are apparent when confined spindles rotate from tilted to parallel positions while MDCK cells progress from prometaphase to metaphase. The spindle disruptions cause the engagement of the spindle assembly checkpoint. We propose that cell rounding serves to maintain spindle integrity during its positioning. PMID:25657320

  20. ATP-ase positive cells in human oral mucosa transplanted to nude mice

    DEFF Research Database (Denmark)

    Dabelsteen, E; Kirkeby, S

    1981-01-01

    , identified as ATP-ase positive dendritic cells, have almost disappeared from the transplanted epithelium whereas at day 21 after transplantation such cells were abundant. It is suggested that the ATP-ase positive cells which reappear in the transplanted epithelium are of mouse origin.......A model to study the differentiation of human oral epithelium in vivo utilizing transplantation of human tissue to nude mice has been described. Previous studies have described the epithelial cells in this model. In this study we demonstrate that 8 d after transplantation, Langerhans cells...

  1. Cellular uptake and cytotoxicity of positively charged chitosan gold nanoparticles in human lung adenocarcinoma cells

    International Nuclear Information System (INIS)

    Cellular uptake, cytotoxicity, and mechanisms of cytotoxicity of the positively charged Au nanoparticles (NPs) were examined in A549 cells, which are one of the most characterized pulmonary cellular systems. Positively charged Au NPs were prepared by chemical reduction using chitosan. The dimension and surface charge of Au NPs were examined by transmission electron microscopy (TEM), dynamic light scattering, and zeta potential measurements. The uptake of Au NPs into A549 cells was also monitored using TEM and dark-field microscopy (DFM) and z-stack confocal microRaman spectroscopy. DFM live cell imaging was also performed to monitor the entry of chitosan Au NPs in real time. The cytotoxic assay, using both methylthiazol tetrazolium and lactate dehydrogenase assays revealed that positively charged Au NPs decreased cell viability. Flow cytometry, DNA fragmentation, real-time PCR, and western blot analysis suggest that positively charged chitosan Au NPs provoke cell damage through both apoptotic and necrotic pathways.

  2. CD34-positive cell yield in peripheral blood of cancer patients during radiation therapy

    International Nuclear Information System (INIS)

    For the purpose of evaluating the feasibility of inducing CD34-positive cells in peripheral blood by radiation therapy, we examined the changes in CD34-positive cells in 21 cancer patients (16 with lung cancer and five with esophageal cancer) during thoracic radiotherapy using flow cytometry and CD34 monoclonal antibody. Although assays of granulocyte-colony-forming units (G-CFU) and granulocyte-monocyte-colony-forming units (GM-CFU) were also performed for 16 of the patients during radiation therapy, in most of these cases there was approximately one G-CFU or GM-CFU for every 100 CD34-positive cells. Seven of the 21 cancer patients showed percentages of CD34-positive cells of more than 3% (more than mean+2 S.D.; standard deviation) of mononuclear cells in peripheral blood in at least one of the examinations. However, six of these seven patients were in stage IV with distant metastases (four with lung cancer and two with esophageal cancer), and another lung cancer patient was in stage III. Therefore, CD-34 positive cells were shown to appear in peripheral blood especially in the patients with advanced stages of malignancy, and further examinations are needed to induce CD34-positive cells by radiation therapy for possible application of ultrahigh dose chemotherapy supported by peripheral blood stem cell transplantation. (author)

  3. Computational model of cell positioning: directed and collective migration in the intestinal crypt epithelium

    Science.gov (United States)

    Wong, Shek Yoon; Chiam, K.-H.; Lim, Chwee Teck; Matsudaira, Paul

    2010-01-01

    The epithelium of the intestinal crypt is a dynamic tissue undergoing constant regeneration through cell growth, cell division, cell differentiation and apoptosis. How the epithelial cells maintain correct positioning and how they migrate in a directed and collective fashion are still not well understood. In this paper, we developed a computational model to elucidate these processes. We show that differential adhesion between epithelial cells, caused by the differential activation of EphB receptors and ephrinB ligands along the crypt axis, is necessary to regulate cell positioning. Differential cell adhesion has been proposed previously to guide cell movement and cause cell sorting in biological tissues. The proliferative cells and the differentiated post-mitotic cells do not intermingle as long as differential adhesion is maintained. We also show that, without differential adhesion, Paneth cells are randomly distributed throughout the intestinal crypt. In addition, our model suggests that, with differential adhesion, cells migrate more rapidly as they approach the top of the intestinal crypt. Finally, by calculating the spatial correlation function of the cell velocities, we observe that differential adhesion results in the differentiated epithelial cells moving in a coordinated manner, where correlated velocities are maintained at large distances, suggesting that differential adhesion regulates coordinated migration of cells in tissues. PMID:20356873

  4. Development of an Integrated Chip for Automatic Tracking and Positioning Manipulation for Single Cell Lysis

    Directory of Open Access Journals (Sweden)

    Chao-Wang Young

    2012-02-01

    Full Text Available This study adopted a microelectromechanical fabrication process to design a chip integrated with electroosmotic flow and dielectrophoresis force for single cell lysis. Human histiocytic lymphoma U937 cells were driven rapidly by electroosmotic flow and precisely moved to a specific area for cell lysis. By varying the frequency of AC power, 15 V AC at 1 MHz of frequency configuration achieved 100% cell lysing at the specific area. The integrated chip could successfully manipulate single cells to a specific position and lysis. The overall successful rate of cell tracking, positioning, and cell lysis is 80%. The average speed of cell driving was 17.74 μm/s. This technique will be developed for DNA extraction in biomolecular detection. It can simplify pre-treatment procedures for biotechnological analysis of samples.

  5. Age-related changes in proliferation, the numbers of mast cells, eosinophils, and cd45-positive cells in human dermis.

    Science.gov (United States)

    Gunin, Andrei G; Kornilova, Natalia K; Vasilieva, Olga V; Petrov, Vadim V

    2011-04-01

    Skin aging is an extremely important medical and social problem in the modern world. Therefore, a goal of the present work was to estimate changes in the numbers of fibroblast-like cells, proliferating cells nuclear antigen-positive cells, CD45-positive cells, mast cells, and eosinophils in human dermis at different ages. Skin specimens from human fetuses that died antenatally from 20 to 40 weeks of pregnancy and humans who died from different causes from 1 day to 85 years of life were used for the study. Results showed a decrease in a total number and the number of proliferating cells nuclear antigen-positive fibroblast-like cells in dermis with progression of age. The numbers of CD45-positive cells and mast cells are gradually increased with aging. Eosinophils are almost absent in dermis independently on age. Mast cells are probably a main factor that potentially can be involved in tissue damage and aging changes in skin. Mast cells should be regarded as an important target for anti-aging therapy. PMID:21106704

  6. Protein transport across the cell wall of monoderm Gram-positive bacteria

    OpenAIRE

    Forster, Brian M.; Marquis, Hélène

    2012-01-01

    In monoderm (single membrane) Gram-positive bacteria, the majority of secreted proteins are first translocated across the cytoplasmic membrane into the inner wall zone. For a subset of these proteins, final destination is within the cell envelope either as membrane-anchored or cell wall-anchored proteins, whereas another subset of proteins is destined to be transported across the cell wall into the extracellular milieu. Although the cell wall is a porous structure, there is evidence that, for...

  7. Multifunctionalized iron oxide nanoparticles for selective drug delivery to CD44-positive cancer cells.

    Science.gov (United States)

    Aires, Antonio; Ocampo, Sandra M; Simões, Bruno M; Josefa Rodríguez, María; Cadenas, Jael F; Couleaud, Pierre; Spence, Katherine; Latorre, Alfonso; Miranda, Rodolfo; Somoza, Álvaro; Clarke, Robert B; Carrascosa, José L; Cortajarena, Aitziber L

    2016-02-12

    Nanomedicine nowadays offers novel solutions in cancer therapy and diagnosis by introducing multimodal treatments and imaging tools in one single formulation. Nanoparticles acting as nanocarriers change the solubility, biodistribution and efficiency of therapeutic molecules, reducing their side effects. In order to successfully  apply these novel therapeutic approaches, efforts are focused on the biological functionalization of the nanoparticles to improve the selectivity towards cancer cells. In this work, we present the synthesis and characterization of novel multifunctionalized iron oxide magnetic nanoparticles (MNPs) with antiCD44 antibody and gemcitabine derivatives, and their application for the selective treatment of CD44-positive cancer cells. The lymphocyte homing receptor CD44 is overexpressed in a large variety of cancer cells, but also in cancer stem cells (CSCs) and circulating tumor cells (CTCs). Therefore, targeting CD44-overexpressing cells is a challenging and promising anticancer strategy. Firstly, we demonstrate the targeting of antiCD44 functionalized MNPs to different CD44-positive cancer cell lines using a CD44-negative non-tumorigenic cell line as a control, and verify the specificity by ultrastructural characterization and downregulation of CD44 expression. Finally, we show the selective drug delivery potential of the MNPs by the killing of CD44-positive cancer cells using a CD44-negative non-tumorigenic cell line as a control. In conclusion, the proposed multifunctionalized MNPs represent an excellent biocompatible nanoplatform for selective CD44-positive cancer therapy in vitro. PMID:26754042

  8. Multifunctionalized iron oxide nanoparticles for selective drug delivery to CD44-positive cancer cells

    Science.gov (United States)

    Aires, Antonio; Ocampo, Sandra M.; Simões, Bruno M.; Josefa Rodríguez, María; Cadenas, Jael F.; Couleaud, Pierre; Spence, Katherine; Latorre, Alfonso; Miranda, Rodolfo; Somoza, Álvaro; Clarke, Robert B.; Carrascosa, José L.; Cortajarena, Aitziber L.

    2016-02-01

    Nanomedicine nowadays offers novel solutions in cancer therapy and diagnosis by introducing multimodal treatments and imaging tools in one single formulation. Nanoparticles acting as nanocarriers change the solubility, biodistribution and efficiency of therapeutic molecules, reducing their side effects. In order to successfully apply these novel therapeutic approaches, efforts are focused on the biological functionalization of the nanoparticles to improve the selectivity towards cancer cells. In this work, we present the synthesis and characterization of novel multifunctionalized iron oxide magnetic nanoparticles (MNPs) with antiCD44 antibody and gemcitabine derivatives, and their application for the selective treatment of CD44-positive cancer cells. The lymphocyte homing receptor CD44 is overexpressed in a large variety of cancer cells, but also in cancer stem cells (CSCs) and circulating tumor cells (CTCs). Therefore, targeting CD44-overexpressing cells is a challenging and promising anticancer strategy. Firstly, we demonstrate the targeting of antiCD44 functionalized MNPs to different CD44-positive cancer cell lines using a CD44-negative non-tumorigenic cell line as a control, and verify the specificity by ultrastructural characterization and downregulation of CD44 expression. Finally, we show the selective drug delivery potential of the MNPs by the killing of CD44-positive cancer cells using a CD44-negative non-tumorigenic cell line as a control. In conclusion, the proposed multifunctionalized MNPs represent an excellent biocompatible nanoplatform for selective CD44-positive cancer therapy in vitro.

  9. Radiosensitivity and effect of hypoxia in HPV positive head and neck cancer cells

    International Nuclear Information System (INIS)

    Background and purpose: HPV associated Head and Neck Squamous Cell Carcinoma (HNSCC) represents a distinct subgroup of HNSCC characterized by a favorable prognosis and a distinct molecular biology. Previous data from the randomized DAHANCA 5 trial indicated that HPV positive tumors did not benefit from hypoxic modifications by Nimorazole during radiotherapy, whereas a significant benefit was observed in the HPV negative tumors. However, more studies have demonstrated equal frequencies of hypoxic tumors among HPV-positive and HPV-negative tumors. The aim of the present study was to determine radiosensitivity, the impact of hypoxia and the effect of Nimorazole in HPV positive and HPV negative cell lines. Materials and method: The used cell lines were: UDSCC2, UMSCC47 and UPCISCC90 (HPV positive) and FaDuDD, UTSCC33 and UTSCC5 (HPV negative). Cells were cultured under normoxic or hypoxic conditions, and gene expression levels of previously established hypoxia induced genes were assessed by qPCR. Cells were irradiated with various doses under normoxia, hypoxia or hypoxia +1 mM Nimorazole, and the clonogenic survival was determined. Results: The HPV positive and HPV negative cell lines exhibited similar patterns of upregulation of hypoxia induced genes in response to hypoxia. The HPV positive cell lines were up to 2.4 times more radiation sensitive than HPV negative cell lines. However, all HPV positive cells displayed the same response to hypoxia in radiosensitivity, with an OER in the range 2.3–2.9, and a sensitizer effect of Nimorazole of 1.13–1.29, similar to HPV negative cells. Conclusions: Although HPV positive cells had a markedly higher radiosensitivity compared to HPV negative cells, they displayed the same relative radioresistance under hypoxia and the same relative sensitizer effect of Nimorazole. The clinical observation that HPV positive patients do not seem to benefit from Nimorazole treatment is not due to inherent differences in hypoxia sensitivity

  10. A comparative proteomic study identified LRPPRC and MCM7 as putative actors in imatinib mesylate cross-resistance in Lucena cell line

    Directory of Open Access Journals (Sweden)

    Corrêa Stephany

    2012-03-01

    Full Text Available Abstract Background Although chronic myeloid leukemia (CML treatment has improved since the introduction of imatinib mesylate (IM, cases of resistance have been reported. This resistance has been associated with the emergence of multidrug resistance (MDR phenotype, as a BCR-ABL independent mechanism. The classic pathway studied in MDR promotion is ATP-binding cassette (ABC family transporters expression, but other mechanisms that drive drug resistance are largely unknown. To better understand IM therapy relapse due to the rise of MDR, we compared the proteomic profiles of K562 and Lucena (K562/VCR cells. Results The use of 2-DE coupled with a MS approach resulted in the identification of 36 differentially expressed proteins. Differential mRNA levels of leucine-rich PPR motif-containing (LRPPRC protein, minichromosome maintenance complex component 7 (MCM7 and ATP-binding cassette sub-family B (MDR/TAP member 1 (ABCB1 were capable of defining samples from CML patients as responsive or resistant to therapy. Conclusions Through the data presented in this work, we show the relevance of MDR to IM therapy. In addition, our proteomic approach identified candidate actors involved in resistance, which could lead to additional information on BCR-ABL-independent molecular mechanisms.

  11. DNA methylation of the CpG island at the Pa promoter within the bcr-abl locus followed in CML patients

    Czech Academy of Sciences Publication Activity Database

    Fajkusová, L.; Fajkus, Jiří; Dvořáková, D.; Krahulcová, E.; Fulneček, Jaroslav; Berková, K.; Mayer, J.

    1998, s. 90. [Meeting on dynamic organization of nuclear function. Cold Spring Harbor (US), 07.10.1998-11.10.1998] R&D Projects: GA MZd IZ3722; GA MŠk VS97032 Institutional research plan: CEZ:A17/98:Z5-004-9-ii Subject RIV: EB - Genetics ; Molecular Biology

  12. BCR-ABL isoforms associated with intrinsic or acquired resistance to imatinib : more heterogeneous than just ABL kinase domain point mutations?

    NARCIS (Netherlands)

    Gruber, Franz X.; Lundan, Tuija; Goll, Rasmus; Silye, Aleksandra; Mikkola, Ingvild; Rekvig, Ole Petter; Knuutila, Sakari; Remes, Kari; Gedde-Dahl, Tobias; Porkka, Kimmo; Hjorth-Hansen, Henrik

    2012-01-01

    Imatinib, a small molecule inhibitor of ABL, PDGFR and C-KIT, has revolutionized treatment of chronic myeloid leukaemia (CML). However, resistance to treatment is of increasing importance and often is due to point mutations in the Abl kinase domain (Abl KD). Here, we analysed clinical outcome and mu

  13. Kinasen der Src-Familie vermitteln die zytoplasmatische Retention von aktiviertem STAT5A in BCR-ABL positiven Leukämiezellen

    OpenAIRE

    Chatain, Nicolas

    2012-01-01

    The transcription factor STAT5 belongs to the family of Signal Transducer and Activator of Transcription and subdivides into STAT5A and STAT5B which share 96% sequence homology on the amino acid level. STAT5A is involved in hematopoiesis, immunity and mammary differentiation. The phosphorylation of a conserved tyrosine is essential for the activation of STAT transcription factors leading to the dimerization of two STAT proteins through reciprocal SH2 domain-phosphotyrosine interactions. The p...

  14. Kidney specific protein-positive cells derived from embryonic stem cells reproduce tubular structures in vitro and differentiate into renal tubular cells.

    Directory of Open Access Journals (Sweden)

    Ryuji Morizane

    Full Text Available Embryonic stem cells and induced pluripotent stem cells have the ability to differentiate into various organs and tissues, and are regarded as new tools for the elucidation of disease mechanisms as well as sources for regenerative therapies. However, a method of inducing organ-specific cells from pluripotent stem cells is urgently needed. Although many scientists have been developing methods to induce various organ-specific cells from pluripotent stem cells, renal lineage cells have yet to be induced in vitro because of the complexity of kidney structures and the diversity of kidney-component cells. Here, we describe a method of inducing renal tubular cells from mouse embryonic stem cells via the cell purification of kidney specific protein (KSP-positive cells using an anti-KSP antibody. The global gene expression profiles of KSP-positive cells derived from ES cells exhibited characteristics similar to those of cells in the developing kidney, and KSP-positive cells had the capacity to form tubular structures resembling renal tubular cells when grown in a 3D culture in Matrigel. Moreover, our results indicated that KSP-positive cells acquired the characteristics of each segment of renal tubular cells through tubular formation when stimulated with Wnt4. This method is an important step toward kidney disease research using pluripotent stem cells, and the development of kidney regeneration therapies.

  15. Autocrine regulation of cell proliferation by estrogen receptor-alpha in estrogen receptor-alpha-positive breast cancer cell lines

    Directory of Open Access Journals (Sweden)

    Pan Zhongzong

    2009-01-01

    Full Text Available Abstract Background Estrogen receptor-α (ERα is essential for mammary gland development and is a major oncogene in breast cancer. Since ERα is not colocalized with the cell proliferation marker Ki-67 in the normal mammary glands and the majority of primary breast tumors, it is generally believed that paracrine regulation is involved in ERα mediated cell proliferation. In the paracrine model, ERα-positive cells don't proliferate but will release some paracrine growth factors to stimulate the neighboring cells to proliferate. In a subpopulation of cancer cells in some primary breast tumors, however, ERα does colocalize with the cell proliferation marker Ki-67, suggesting an autocrine regulation by ERα in some primary breast tumors. Methods Colocalization of ERα with Ki-67 in ERα-positive breast cancer cell lines (MCF-7, T47D, and ZR75-1 was evaluated by immunofluorescent staining. Cell cycle phase dependent expression of ERα was determined by co-immunofluorescent staining of ERα and the major cyclins (D, E, A, B, and by flow cytometry analysis of ERαhigh cells. To further confirm the autocrine action of ERα, MCF-7 cells were growth arrested by ICI182780 treatment, followed by treatment with EGFR inhibitor, before estrogen stimulation and analyses for colocalization of Ki-67 and ERα and cell cycle progression. Results Colocalization of ERα with Ki-67 was present in all three ERα-positive breast cancer cell lines. Unlike that in the normal mammary glands and the majority of primary breast tumors, ERα is highly expressed throughout the cell cycle in MCF-7 cells. Without E2 stimulation, MCF-7 cells released from ICI182780 treatment remain at G1 phase. E2 stimulation of ICI182780 treated cells, however, promotes the expression and colocalization of ERα and Ki-67 as well as the cell cycle progressing through the S and G2/M phases. Inhibition of EGFR signaling does not inhibit the autocrine action of ERα. Conclusion Our data indicate

  16. Autocrine regulation of cell proliferation by estrogen receptor-alpha in estrogen receptor-alpha-positive breast cancer cell lines

    International Nuclear Information System (INIS)

    Estrogen receptor-α (ERα) is essential for mammary gland development and is a major oncogene in breast cancer. Since ERα is not colocalized with the cell proliferation marker Ki-67 in the normal mammary glands and the majority of primary breast tumors, it is generally believed that paracrine regulation is involved in ERα mediated cell proliferation. In the paracrine model, ERα-positive cells don't proliferate but will release some paracrine growth factors to stimulate the neighboring cells to proliferate. In a subpopulation of cancer cells in some primary breast tumors, however, ERα does colocalize with the cell proliferation marker Ki-67, suggesting an autocrine regulation by ERα in some primary breast tumors. Colocalization of ERα with Ki-67 in ERα-positive breast cancer cell lines (MCF-7, T47D, and ZR75-1) was evaluated by immunofluorescent staining. Cell cycle phase dependent expression of ERα was determined by co-immunofluorescent staining of ERα and the major cyclins (D, E, A, B), and by flow cytometry analysis of ERαhigh cells. To further confirm the autocrine action of ERα, MCF-7 cells were growth arrested by ICI182780 treatment, followed by treatment with EGFR inhibitor, before estrogen stimulation and analyses for colocalization of Ki-67 and ERα and cell cycle progression. Colocalization of ERα with Ki-67 was present in all three ERα-positive breast cancer cell lines. Unlike that in the normal mammary glands and the majority of primary breast tumors, ERα is highly expressed throughout the cell cycle in MCF-7 cells. Without E2 stimulation, MCF-7 cells released from ICI182780 treatment remain at G1 phase. E2 stimulation of ICI182780 treated cells, however, promotes the expression and colocalization of ERα and Ki-67 as well as the cell cycle progressing through the S and G2/M phases. Inhibition of EGFR signaling does not inhibit the autocrine action of ERα. Our data indicate that ERα can mediate estrogen-induced cell proliferation in

  17. Rare Association of Anti-Hu Antibody Positive Paraneoplastic Neurological Syndrome and Transitional Cell Bladder Carcinoma

    OpenAIRE

    S. Lukacs; Szabo, N; Woodhams, S

    2012-01-01

    Introduction. Paraneoplastic encephalomyelitis (PEM) and subacute sensory neuronopathy (SSN) are remote effects of cancer, usually associated with small-cell lung carcinoma and positive anti-Hu antibody. We describe the rare association of bladder transitional cell carcinoma (TCC) with anti-Hu antibody positivity resulting in this paraneoplastic neurological syndrome. Patient. A 76-year-old female presented with bilateral muscle weakness and paraesthesia of the upper and lower limbs in a leng...

  18. Design of a Single-Cell Positioning Controller Using Electroosmotic Flow and Image Processing

    OpenAIRE

    Jhong-Yin Chen; Chao-Wang Young; Chyung Ay

    2013-01-01

    The objective of the current research was not only to provide a fast and automatic positioning platform for single cells, but also improved biomolecular manipulation techniques. In this study, an automatic platform for cell positioning using electroosmotic flow and image processing technology was designed. The platform was developed using a PCI image acquisition interface card for capturing images from a microscope and then transferring them to a computer using human-machine interface softwar...

  19. 酪氨酸激酶抑制剂联合异基因造血干细胞移植治疗费城染色体阳性急性淋巴细胞白血病的研究进展%Research progress of tyrosine kinase inhibitor combining with allogeneic hematopoietic stem cell transplantation in treatment of Philadelphia chromosome positive acute lymphoblastic leukemia

    Institute of Scientific and Technical Information of China (English)

    王静静; 江明

    2016-01-01

    费城染色体阳性(Ph+)急性淋巴细胞白血病(ALL)是成年人ALL的常见类型.在酪氨酸激酶抑制剂(TKI)问世前,ph+是成年人ALL的不良预后因素之一.在TKI应用于Ph+ ALL治疗后显著改善其疗效,完全缓解(CR)率可达90%以上,缓解持续时间较前明显延长,从而使更多Ph+ ALL患者能够行异基因造血干细胞移植(allo-HSCT).采用以TKI为基础化疗获得CR后行allo-HSCT是Ph+ ALL患者的标准治疗方案.目前,移植前微小残留疾病(MRD)检测是否对该病的治疗具有临床指导意义,尚存争议.替代供者移植治疗Ph+ ALL在临床取得理想疗效,但需要前瞻性随机对照试验来证实其疗效.移植后联合TKI治疗可明显改善Ph+ ALL患者的长期生存,但其具体使用方案仍需多中心、大样本量的临床随机对照实验证实,而移植后复发仍是Ph+ ALL患者面临的难题之一.%Philadelphia chromosome positive (Ph+) acute lymphoblastic leukemia (ALL) is the most common type of adult acute lymphoblastic leukemia(ALL).Before the advent of tyrosine kinase inhibitors (TKI),Ph+ ALL carried a dismal prognosis.Outcomes for patients with Ph+ ALL improved substantially with the administration of TKI,and the TKI induced complete remissions(CR) in more than 90% patients,with remission duration prolonging,making more patients able to undergo allogeneic hematopoietic stem cell transplantation(allo-HSCT).Currently,TKI-based chemotherapy following allo-HSCT is established as the first-line strategy.Breakpoint cluster region-abelson leukemia virus(BCR-ABL) monitoring appears to relate to prognostic relevance and may be as guiding treatment before allo-HSCT,but clinical benefit is controversial in the TKI era.Alternative donor transplantation obtains promising effect in clinical practice,but needs more research.The administration of TKI after allo-HSCT may improve quality of life and prolong life span,but relapse remains a major problem of treatment failure

  20. Aurora A kinase regulates proper spindle positioning in C. elegans and in human cells.

    Science.gov (United States)

    Kotak, Sachin; Afshar, Katayon; Busso, Coralie; Gönczy, Pierre

    2016-08-01

    Accurate spindle positioning is essential for error-free cell division. The one-cell Caenorhabditis elegans embryo has proven instrumental for dissecting mechanisms governing spindle positioning. Despite important progress, how the cortical forces that act on astral microtubules to properly position the spindle are modulated is incompletely understood. Here, we report that the PP6 phosphatase PPH-6 and its associated subunit SAPS-1, which positively regulate pulling forces acting on spindle poles, associate with the Aurora A kinase AIR-1 in C. elegans embryos. We show that acute inactivation of AIR-1 during mitosis results in excess pulling forces on astral microtubules. Furthermore, we uncover that AIR-1 acts downstream of PPH-6-SAPS-1 in modulating spindle positioning, and that PPH-6-SAPS-1 negatively regulates AIR-1 localization at the cell cortex. Moreover, we show that Aurora A and the PP6 phosphatase subunit PPP6C are also necessary for spindle positioning in human cells. There, Aurora A is needed for the cortical localization of NuMA and dynein during mitosis. Overall, our work demonstrates that Aurora A kinases and PP6 phosphatases have an ancient function in modulating spindle positioning, thus contributing to faithful cell division. PMID:27335426

  1. Surface cell differentiation controls tissue surface tension and tissue positioning during zebrafish gastrulation

    Science.gov (United States)

    Krens, S. F. G.

    2011-03-01

    Differences in tissue surface tension (TST) between different tissue types are thought to guide tissue organization and cell sorting in development. Measurements of TST have been useful to predict the outcome of in vitro cell sorting and envelopment experiments. However, the outcome of cell sorting experiments in vitro often substantially differs from tissue positioning in vivo, raising questions as to the actual contribution of TST to tissue positioning within the developing embryo. Here, we show that surface tension of germ layer tissues during zebrafish gastrulation critically relies on the differentiation of their surface cells. We also show that surface differentiation of the different germ layer tissues varies and is considerably different between the situation in vitro and in vivo, explaining the apparent dissimilar outcome of cell segregation between these two situations. To analyze germ layer TST as a function of surface cell differentiation, we interfere with surface cell properties of germ layer aggregates by misexpressing genes involved in surface cell differentiation specifically within surface cells using the GAL4-UAS system, and measure tissue surface tension using both parallel plate compression and micropipette aspiration techniques. Our data provides evidence in favor of a critical function of surface cell differentiation in modulating TST and subsequently tissue positioning within the developing embryo.

  2. Calpain-Mediated Positional Information Directs Cell Wall Orientation to Sustain Plant Stem Cell Activity, Growth and Development.

    Science.gov (United States)

    Liang, Zhe; Brown, Roy C; Fletcher, Jennifer C; Opsahl-Sorteberg, Hilde-Gunn

    2015-09-01

    Eukaryotic development and stem cell control depend on the integration of cell positional sensing with cell cycle control and cell wall positioning, yet few factors that directly link these events are known. The DEFECTIVE KERNEL1 (DEK1) gene encoding the unique plant calpain protein is fundamental for development and growth, being essential to confer and maintain epidermal cell identity that allows development beyond the globular embryo stage. We show that DEK1 expression is highest in the actively dividing cells of seeds, meristems and vasculature. We further show that eliminating Arabidopsis DEK1 function leads to changes in developmental cues from the first zygotic division onward, altered microtubule patterns and misshapen cells, resulting in early embryo abortion. Expression of the embryonic marker genes WOX2, ATML1, PIN4, WUS and STM, related to axis organization, cell identity and meristem functions, is also altered in dek1 embryos. By monitoring cell layer-specific DEK1 down-regulation, we show that L1- and 35S-induced down-regulation mainly affects stem cell functions, causing severe shoot apical meristem phenotypes. These results are consistent with a requirement for DEK1 to direct layer-specific cellular activities and set downstream developmental cues. Our data suggest that DEK1 may anchor cell wall positions and control cell division and differentiation, thereby balancing the plant's requirement to maintain totipotent stem cell reservoirs while simultaneously directing growth and organ formation. A role for DEK1 in regulating microtubule-orchestrated cell wall orientation during cell division can explain its effects on embryonic development, and suggests a more general function for calpains in microtubule organization in eukaryotic cells. PMID:26220906

  3. High expression of hTERT and stemness genes in BORIS/CTCFL positive cells isolated from embryonic cancer cells.

    Directory of Open Access Journals (Sweden)

    Loredana Alberti

    Full Text Available BORIS/CTCFL is a member of cancer testis antigen family normally expressed in germ cells. In tumors, it is aberrantly expressed although its functions are not completely well-defined. To better understand the functions of BORIS in cancer, we selected the embryonic cancer cells as a model. Using a molecular beacon, which specifically targets BORIS mRNA, we demonstrated that BORIS positive cells are a small subpopulation of tumor cells (3-5% of total. The BORIS-positive cells isolated using BORIS-molecular beacon, expressed higher telomerase hTERT, stem cell (NANOG, OCT4, SOX2 and cancer stem cell marker genes (CD44 and ALDH1 compared to the BORIS-negative tumor cells. In order to define the functional role of BORIS, stable BORIS-depleted embryonic cancer cells were generated. BORIS silencing strongly down-regulated the expression of hTERT, stem cell and cancer stem cell marker genes. Moreover, the BORIS knockdown increased cellular senescence in embryonic cancer cells, revealing a putative role of BORIS in the senescence biological program. Our data indicate an association of BORIS expressing cells subpopulation with the expression of stemness genes, highlighting the critical role played by BORIS in embryonic neoplastic disease.

  4. 3D Image-Guided Automatic Pipette Positioning for Single Cell Experiments in vivo

    OpenAIRE

    Brian Long; Lu Li; Ulf Knoblich; Hongkui Zeng; Hanchuan Peng

    2015-01-01

    We report a method to facilitate single cell, image-guided experiments including in vivo electrophysiology and electroporation. Our method combines 3D image data acquisition, visualization and on-line image analysis with precise control of physical probes such as electrophysiology microelectrodes in brain tissue in vivo. Adaptive pipette positioning provides a platform for future advances in automated, single cell in vivo experiments.

  5. IgG4-positive plasma cell infiltration in explanted livers for primary sclerosing cholangitis

    OpenAIRE

    Zen, Yoh; Quaglia, Alberto; Portmann, Bernard

    2011-01-01

    Abstract Aims: To explore whether IgG4-related sclerosing cholangitis (IgG4-SC) contributes to end-stage primary sclerosing cholangitis (PSC) in UK. Methods and Results: This study consisted of 41 patients who underwent liver transplantation for advanced PSC. Explanted livers were histologically examined with an emphasis on IgG4-positive plasma cell infiltration. Thirty-nine cases (95%) had minimal or mild infiltration of IgG4-positive plasma cells (? 30 cells/high power field)...

  6. Holoprosencephaly and Pure Red Cell Aplasia in a Feline Leukaemia Virus-Positive Kitten.

    Science.gov (United States)

    Southard, T L; Rodriguez-Ramos Fernandez, J; Priest, H; Stokol, T

    2016-01-01

    A 9-month-old, female, domestic longhair cat with severe anaemia tested positive for feline leukaemia virus (FeLV) and was humanely destroyed and submitted for necropsy examination. Gross findings included a non-divided rostral telencephalon, consistent with semilobar holoprosencephaly. Histological examination of the bone marrow revealed an almost complete absence of erythroid precursor cells, consistent with pure red cell aplasia, and mild to moderate myelofibrosis. This case demonstrates a very unusual central nervous system defect, as well as an atypical presentation of pure red cell aplasia, in a FeLV-positive kitten. PMID:26897097

  7. Galiellalactone Inhibits Stem Cell-Like ALDH-Positive Prostate Cancer Cells

    OpenAIRE

    Hellsten, Rebecka; Johansson, Martin; Dahlman, Anna; Sterner, Olov; Bjartell, Anders

    2011-01-01

    Galiellalactone is a potent and specific inhibitor of STAT3 signaling which has been shown to possess growth inhibitory effects on prostate cancer cells expressing active STAT3. In this study we aimed to investigate the effect of galiellalactone on prostate cancer stem cell-like cells. We explored the expression of aldehyde dehydrogenase (ALDH) as a marker for cancer stem cell-like cells in different human prostate cancer cell lines and the effects of galiellalactone on ALDH expressing (ALDH+...

  8. Allo-hematopoietic stem cell transplantation is a potential treatment for a patient with a combined disorder of hereditary spherocytosis

    Institute of Scientific and Technical Information of China (English)

    ZHANG Xiao-hui; WANG Yu; ZHAO Ting; LIU Kai-yan; HUANG Xiao-jun; FU Hai-xia; XU Lan-ping; LIU Dai-hong; CHEN Huan; HAN Wei; CHEN Yu-hong; WANG Feng-rong; WANG Jing-zhi

    2012-01-01

    Both human hereditary spherocytosis (HS) and chronic myelogenous leukemia (CML) are life threatening.Herein we have reported the case of a woman with a combined disorder of HS and CML who underwent the matched sibling allogeneic stem cell transplantation.The complete donor erythroid cells were obtained.The red blood cell counts significantly improved throughout life comparing with pre-hematopoietic stem cell transplantation (HSCT).Reticulocyte counts normalized,and BCR-ABL was cleared away.The total bilirubin level was also corrected in this recipient.Our case is a rare example with a combined disorder of HS and CML following allogeneic stem cell transplantation.HS was not a contraindication for patient in the matched sibling transplant setting.

  9. Peroxidase-positive Auer bodies in plasma cells in multiple myeloma: a case report

    Science.gov (United States)

    Zhu, Lin; An, Li; Zhang, Xiao-Yan; Ren, Xue-Rui; Song, Jing-Wen

    2015-01-01

    Reports of clinical cases with Auer bodies in the plasma cells in multiple myeloma (MM) are rare; however, most of those reported contain peroxidase (POX)-negative Auer bodies rather than the POX-positive Auer bodies observed in myeloid progenitors, indicating differences in their chemical properties. Furthermore, the cases with POX-positive Auer bodies similar to those observed in myeloid cells are extremely rare in non-myeloid cells. Here, we report the clinical features, laboratory investigations, diagnosis and treatment of a case of MM with POX-positive Auer bodies in plasma cells and review related the literature to advance the prognostic evaluation, diagnosis and treatment of similar cases. PMID:26823884

  10. Phototheranostics of CD44-positive cell populations in triple negative breast cancer

    Science.gov (United States)

    Jin, Jiefu; Krishnamachary, Balaji; Mironchik, Yelena; Kobayashi, Hisataka; Bhujwalla, Zaver M.

    2016-01-01

    Triple-negative breast cancer (TNBC) is one of the most lethal subtypes of breast cancer that has limited treatment options. Its high rates of recurrence and metastasis have been associated, in part, with a subpopulation of breast cancer stem-like cells that are resistant to conventional therapies. A compendium of markers such as CD44high/CD24low, and increased expression of the ABCG2 transporter and increased aldehyde dehydrogenase (ALDH1), have been associated with these cells. We developed a CD44-targeted monoclonal antibody photosensitizer conjugate for combined fluorescent detection and photoimmunotherapy (PIT) of CD44 expressing cells in TNBC. The CD44-targeted conjugate demonstrated acute cell killing of breast cancer cells with high CD44 expression. This cell death process was dependent upon CD44-specific cell membrane binding combined with near-infrared irradiation. The conjugate selectively accumulated in CD44-positive tumors and caused dramatic tumor shrinkage and efficient elimination of CD44-positive cell populations following irradiation. This novel phototheranostic strategy provides a promising opportunity for the destruction of CD44-positive populations that include cancer stem-like cells, in locally advanced primary and metastatic TNBC. PMID:27302409

  11. Phototheranostics of CD44-positive cell populations in triple negative breast cancer.

    Science.gov (United States)

    Jin, Jiefu; Krishnamachary, Balaji; Mironchik, Yelena; Kobayashi, Hisataka; Bhujwalla, Zaver M

    2016-01-01

    Triple-negative breast cancer (TNBC) is one of the most lethal subtypes of breast cancer that has limited treatment options. Its high rates of recurrence and metastasis have been associated, in part, with a subpopulation of breast cancer stem-like cells that are resistant to conventional therapies. A compendium of markers such as CD44(high)/CD24(low), and increased expression of the ABCG2 transporter and increased aldehyde dehydrogenase (ALDH1), have been associated with these cells. We developed a CD44-targeted monoclonal antibody photosensitizer conjugate for combined fluorescent detection and photoimmunotherapy (PIT) of CD44 expressing cells in TNBC. The CD44-targeted conjugate demonstrated acute cell killing of breast cancer cells with high CD44 expression. This cell death process was dependent upon CD44-specific cell membrane binding combined with near-infrared irradiation. The conjugate selectively accumulated in CD44-positive tumors and caused dramatic tumor shrinkage and efficient elimination of CD44-positive cell populations following irradiation. This novel phototheranostic strategy provides a promising opportunity for the destruction of CD44-positive populations that include cancer stem-like cells, in locally advanced primary and metastatic TNBC. PMID:27302409

  12. Clinicopathological Analysis of Ocular Adnexal Extranodal Marginal Zone B-Cell Lymphoma with IgG4-Positive Cells

    OpenAIRE

    Lee, Min Joung; Kim, Namju; Choe, Ji-Young; Khwarg, Sang In; Jeon, Yoon Kyung; Choung, Ho-Kyung; Kim, Ji Eun

    2015-01-01

    This study aims to analyze clinical and pathological characteristics of ocular adnexal extranodal marginal zone B-cell lymphoma (EMZL) accompanying IgG4-positive cells. Fifty patients with a diagnosis of primary non-conjunctival ocular adnexal EMZL were enrolled in this study. The number of IgG4-positive cells and the ratio of IgG/IgG4 were evaluated by immunohistochemistry in the biopsy specimens. The patients were divided into two groups based on the absolute number and the ratio of IgG4-po...

  13. Label-free quantitative proteomics of CD133-positive liver cancer stem cells

    Directory of Open Access Journals (Sweden)

    Tsai Sheng-Ta

    2012-11-01

    Full Text Available Abstract Background CD133-positive liver cancer stem cells, which are characterized by their resistance to conventional chemotherapy and their tumor initiation ability at limited dilutions, have been recognized as a critical target in liver cancer therapeutics. In the current work, we developed a label-free quantitative method to investigate the proteome of CD133-positive liver cancer stem cells for the purpose of identifying unique biomarkers that can be utilized for targeting liver cancer stem cells. Label-free quantitation was performed in combination with ID-based Elution time Alignment by Linear regression Quantitation (IDEAL-Q and MaxQuant. Results Initially, IDEAL-Q analysis revealed that 151 proteins were differentially expressed in the CD133-positive hepatoma cells when compared with CD133-negative cells. We then analyzed these 151 differentially expressed proteins by MaxQuant software and identified 10 significantly up-regulated proteins. The results were further validated by RT-PCR, western blot, flow cytometry or immunofluorescent staining which revealed that prominin-1, annexin A1, annexin A3, transgelin, creatine kinase B, vimentin, and EpCAM were indeed highly expressed in the CD133-positive hepatoma cells. Conclusions These findings confirmed that mass spectrometry-based label-free quantitative proteomics can be used to gain insights into liver cancer stem cells.

  14. Cytokines inducing bone marrow SCA+ cells migration into pancreatic islet and conversion into insulin-positive cells in vivo.

    Directory of Open Access Journals (Sweden)

    LuGuang Luo

    Full Text Available We hypothesize that specific bone marrow lineages and cytokine treatment may facilitate bone marrow migration into islets, leading to a conversion into insulin producing cells in vivo. In this study we focused on identifying which bone marrow subpopulations and cytokine treatments play a role in bone marrow supporting islet function in vivo by evaluating whether bone marrow is capable of migrating into islets as well as converting into insulin positive cells. We approached this aim by utilizing several bone marrow lineages and cytokine-treated bone marrow from green fluorescent protein (GFP positive bone marrow donors. Sorted lineages of Mac-1(+, Mac-1(-, Sca(+, Sca(-, Sca(-/Mac-1(+ and Sca(+/Mac-1(- from GFP positive mice were transplanted to irradiated C57BL6 GFP negative mice. Bone marrow from transgenic human ubiquitin C promoter GFP (uGFP, with strong signal C57BL6 mice was transplanted into GFP negative C57BL6 recipients. After eight weeks, migration of GFP positive donor' bone marrow to the recipient's pancreatic islets was evaluated as the percentage of positive GFP islets/total islets. The results show that the most effective migration comes from the Sca(+/Mac(- lineage and these cells, treated with cytokines for 48 hours, were found to have converted into insulin positive cells in pancreatic islets in vivo. This study suggests that bone marrow lineage positive cells and cytokine treatments are critical factors in determining whether bone marrow is able to migrate and form insulin producing cells in vivo. The mechanisms causing this facilitation as well as bone marrow converting to pancreatic beta cells still need to be investigated.

  15. Proteomic analysis of imatinib-resistant CML-T1 cells reveals calcium homeostasis as a potential therapeutic target

    Science.gov (United States)

    Toman, O.; Kabickova, T.; Vit, O.; Fiser, R.; Polakova, K. Machova; Zach, J.; Linhartova, J.; Vyoral, D.; Petrak, J.

    2016-01-01

    Chronic myeloid leukemia (CML) therapy has markedly improved patient prognosis after introduction of imatinib mesylate for clinical use. However, a subset of patients develops resistance to imatinib and other tyrosine kinase inhibitors (TKIs), mainly due to point mutations in the region encoding the kinase domain of the fused BCR-ABL oncogene. To identify potential therapeutic targets in imatinib-resistant CML cells, we derived imatinib-resistant CML-T1 human cell line clone (CML-T1/IR) by prolonged exposure to imatinib in growth media. Mutational analysis revealed that the Y235H mutation in BCR-ABL is probably the main cause of CML-T1/IR resistance to imatinib. To identify alternative therapeutic targets for selective elimination of imatinib-resistant cells, we compared the proteome profiles of CML-T1 and CML-T1/IR cells using 2-DE-MS. We identified eight differentially expressed proteins, with strongly upregulated Na+/H+ exchanger regulatory factor 1 (NHERF1) in the resistant cells, suggesting that this protein may influence cytosolic pH, Ca2+ concentration or signaling pathways such as Wnt in CML-T1/IR cells. We tested several compounds including drugs in clinical use that interfere with the aforementioned processes and tested their relative toxicity to CML-T1 and CML-T1/IR cells. Calcium channel blockers, calcium signaling antagonists and modulators of calcium homeostasis, namely thapsigargin, ionomycin, verapamil, carboxyamidotriazole and immunosuppressive drugs cyclosporine A and tacrolimus (FK-506) were selectively toxic to CML-T1/IR cells. The putative cellular targets of these compounds in CML-T1/IR cells are postulated in this study. We propose that Ca2+ homeostasis can be a potential therapeutic target in CML cells resistant to TKIs. We demonstrate that a proteomic approach may be used to characterize a TKI-resistant population of CML cells enabling future individualized treatment options for patients. PMID:27430982

  16. A positive feedback regulation of ISL-1 in DLBCL but not in pancreatic β-cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Qiao, E-mail: zhangqiao200824@126.com [Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Key Laboratory of Molecular Cardiovascular Sciences (Ministry of Education), Peking University, 38 Xueyuan Road, 100191 Beijing (China); Yang, Zhe, E-mail: zheyang@bjmu.edu.cn [Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Key Laboratory of Molecular Cardiovascular Sciences (Ministry of Education), Peking University, 38 Xueyuan Road, 100191 Beijing (China); Wang, Weiping, E-mail: wwp@bjmu.edu.cn [Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Key Laboratory of Molecular Cardiovascular Sciences (Ministry of Education), Peking University, 38 Xueyuan Road, 100191 Beijing (China); Guo, Ting, E-mail: luckyguoting@bjmu.edu.cn [Department of Gastrointestinal Translation Research, Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education), Peking University Cancer Hospital, 52 Fucheng Road, 100142 Beijing (China); Jia, Zhuqing, E-mail: zhuqingjia@126.com [Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Key Laboratory of Molecular Cardiovascular Sciences (Ministry of Education), Peking University, 38 Xueyuan Road, 100191 Beijing (China); Ma, Kangtao, E-mail: makangtao11@126.com [Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Key Laboratory of Molecular Cardiovascular Sciences (Ministry of Education), Peking University, 38 Xueyuan Road, 100191 Beijing (China); Zhou, Chunyan, E-mail: chunyanzhou@bjmu.edu.cn [Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Key Laboratory of Molecular Cardiovascular Sciences (Ministry of Education), Peking University, 38 Xueyuan Road, 100191 Beijing (China)

    2014-07-04

    Highlights: • ISL-1 is highly expressed in human pancreatic β-cells and DLBCL. • ISL-1 accelerates the tumorigenesis of DLBCL in vivo. • c-Myc positively regulates ISL-1 expression in DLBCL but not in pancreatic β-cells. • ISL-1 and c-Myc forms an ISL-1/c-Myc transcriptional complex only in DLBCL. • Positive feedback regulation of ISL-1 does not exist in normal pancreatic β-cell. - Abstract: Insulin enhancer binding protein-1 (ISL-1), a LIM-homeodomain transcription factor, has been reported to play essential roles in promoting adult pancreatic β-cells proliferation. Recent studies indicate that ISL-1 may also involve in the occurrence of a variety of tumors. However, whether ISL-1 has any functional effect on tumorigenesis, and what are the differences on ISL-1 function in distinct conditions, are completely unknown. In this study, we found that ISL-1 was highly expressed in human pancreatic β-cells, as well as in diffuse large B cell lymphoma (DLBCL), but to a much less extent in other normal tissues or tumor specimens. Further study revealed that ISL-1 promoted the proliferation of pancreatic β-cells and DLBCL cells, and also accelerated the tumorigenesis of DLBCL in vivo. We also found that ISL-1 could activate c-Myc transcription not only in pancreatic β-cells but also in DLBCL cells. However, a cell-specific feedback regulation was detectable only in DLBCL cells. This auto-regulatory loop was established by the interaction of ISL-1 and c-Myc to form an ISL-1/c-Myc transcriptional complex, and synergistically to promote ISL-1 transcription through binding on the ISL-1 promoter. Taken together, our results demonstrate a positive feedback regulation of ISL-1 in DLBCL but not in pancreatic β-cells, which might result in the functional diversities of ISL-1 in different physiological and pathological processes.

  17. A positive feedback regulation of ISL-1 in DLBCL but not in pancreatic β-cells

    International Nuclear Information System (INIS)

    Highlights: • ISL-1 is highly expressed in human pancreatic β-cells and DLBCL. • ISL-1 accelerates the tumorigenesis of DLBCL in vivo. • c-Myc positively regulates ISL-1 expression in DLBCL but not in pancreatic β-cells. • ISL-1 and c-Myc forms an ISL-1/c-Myc transcriptional complex only in DLBCL. • Positive feedback regulation of ISL-1 does not exist in normal pancreatic β-cell. - Abstract: Insulin enhancer binding protein-1 (ISL-1), a LIM-homeodomain transcription factor, has been reported to play essential roles in promoting adult pancreatic β-cells proliferation. Recent studies indicate that ISL-1 may also involve in the occurrence of a variety of tumors. However, whether ISL-1 has any functional effect on tumorigenesis, and what are the differences on ISL-1 function in distinct conditions, are completely unknown. In this study, we found that ISL-1 was highly expressed in human pancreatic β-cells, as well as in diffuse large B cell lymphoma (DLBCL), but to a much less extent in other normal tissues or tumor specimens. Further study revealed that ISL-1 promoted the proliferation of pancreatic β-cells and DLBCL cells, and also accelerated the tumorigenesis of DLBCL in vivo. We also found that ISL-1 could activate c-Myc transcription not only in pancreatic β-cells but also in DLBCL cells. However, a cell-specific feedback regulation was detectable only in DLBCL cells. This auto-regulatory loop was established by the interaction of ISL-1 and c-Myc to form an ISL-1/c-Myc transcriptional complex, and synergistically to promote ISL-1 transcription through binding on the ISL-1 promoter. Taken together, our results demonstrate a positive feedback regulation of ISL-1 in DLBCL but not in pancreatic β-cells, which might result in the functional diversities of ISL-1 in different physiological and pathological processes

  18. Islet Cells Serve as Cells of Origin of Pancreatic Gastrin-Positive Endocrine Tumors

    DEFF Research Database (Denmark)

    Bonnavion, Rémy; Teinturier, Romain; Jaafar, Rami;

    2015-01-01

    The cells of origin of pancreatic gastrinomas remain an enigma, since no gastrin-expressing cells are found in the normal adult pancreas. It was proposed that the cellular origin of pancreatic gastrinomas may come from either the pancreatic cells themselves or gastrin-expressing cells which have...

  19. Reversible immortalization of Nestin-positive precursor cells from pancreas and differentiation into insulin-secreting cells

    Energy Technology Data Exchange (ETDEWEB)

    Wei, Pei; Li, Li; Qi, Hui [The Clinical Medical Research Center, The Second Clinical Medical College (Shenzhen People' s Hospital), Jinan University, 518020 Shenzhen (China); Zhou, Han-xin [Department of General Surgery, First Hospital (Shenzhen Second People' s Hospital) of Shenzhen University, 518020 Shenzhen (China); Deng, Chun-yan [The Clinical Medical Research Center, The Second Clinical Medical College (Shenzhen People' s Hospital), Jinan University, 518020 Shenzhen (China); Li, Fu-rong, E-mail: frli62@yahoo.com [The Clinical Medical Research Center, The Second Clinical Medical College (Shenzhen People' s Hospital), Jinan University, 518020 Shenzhen (China); Shenzhen Institution of Gerontology, 518020 Shenzhen (China)

    2012-02-10

    Highlights: Black-Right-Pointing-Pointer The NPPCs from mouse pancreas were isolated. Black-Right-Pointing-Pointer Tet-on system for SV40 large in NPPCs was used to get RINPPCs. Black-Right-Pointing-Pointer The RINPPCs can undergo at least 80 population doublings without senescence. Black-Right-Pointing-Pointer The RINPPCs can be induced to differentiate into insulin-producing cells. Black-Right-Pointing-Pointer The combination of GLP-1 and sodium butyrate promoted the differentiation process. -- Abstract: Pancreatic stem cells or progenitor cells posses the ability of directed differentiation into pancreatic {beta} cells. However, these cells usually have limited proliferative capacity and finite lifespan in vitro. In the present study, Nestin-positive progenitor cells (NPPCs) from mouse pancreas that expressed the pancreatic stem cells or progenitor cell marker Nestin were isolated to obtain a sufficient number of differentiated pancreatic {beta} cells. Tet-on system for SV40 large T-antigen expression in NPPCs was used to achieve reversible immortalization. The reversible immortal Nestin-positive progenitor cells (RINPPCs) can undergo at least 80 population doublings without senescence in vitro while maintaining their biological and genetic characteristics. RINPPCs can be efficiently induced to differentiate into insulin-producing cells that contain a combination of glucagon-like peptide-1 (GLP-1) and sodium butyrate. The results of the present study can be used to explore transplantation therapy of type I diabetes mellitus.

  20. Reversible immortalization of Nestin-positive precursor cells from pancreas and differentiation into insulin-secreting cells

    International Nuclear Information System (INIS)

    Highlights: ► The NPPCs from mouse pancreas were isolated. ► Tet-on system for SV40 large in NPPCs was used to get RINPPCs. ► The RINPPCs can undergo at least 80 population doublings without senescence. ► The RINPPCs can be induced to differentiate into insulin-producing cells. ► The combination of GLP-1 and sodium butyrate promoted the differentiation process. -- Abstract: Pancreatic stem cells or progenitor cells posses the ability of directed differentiation into pancreatic β cells. However, these cells usually have limited proliferative capacity and finite lifespan in vitro. In the present study, Nestin-positive progenitor cells (NPPCs) from mouse pancreas that expressed the pancreatic stem cells or progenitor cell marker Nestin were isolated to obtain a sufficient number of differentiated pancreatic β cells. Tet-on system for SV40 large T-antigen expression in NPPCs was used to achieve reversible immortalization. The reversible immortal Nestin-positive progenitor cells (RINPPCs) can undergo at least 80 population doublings without senescence in vitro while maintaining their biological and genetic characteristics. RINPPCs can be efficiently induced to differentiate into insulin-producing cells that contain a combination of glucagon-like peptide-1 (GLP-1) and sodium butyrate. The results of the present study can be used to explore transplantation therapy of type I diabetes mellitus.

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  9. Canine CD4(+)CD8(+) double-positive T cells can develop from CD4(+) and CD8(+) T cells.

    Science.gov (United States)

    Bismarck, Doris; Moore, Peter F; Alber, Gottfried; von Buttlar, Heiner

    2014-12-15

    For a long time the expression of the CD4 and CD8 receptor on peripheral blood T cells was thought to be mutually exclusive. However, in canine peripheral blood, similar to other species as swine or human for example, mature CD4(+)CD8(+) double-positive (dp) T cells exist which simultaneously express both surface receptors and have features of activated T cells. Canine CD4(+)CD8(+)dp T cells are heterogeneous and can be divided into three subpopulations by their intensity of CD4 and CD8α expression: CD4(bright)CD8α(bright), CD4(dim)CD8α(bright) and CD4(dim)CD8α(dim). The number of CD4(+)CD8α(+)dp T cells increases after in vitro stimulation of canine peripheral blood mononuclear cells (PBMC) raising the question of their progenitor(s). Thus, the aim of our study was to characterize the progenitor(s) of canine CD4(+)CD8α(+)dp T cells. By cell tracing experiments we identified both CD4(+) single-positive (sp) and also CD8α(+)sp T cells as progenitors of canine CD4(+)CD8α(+)dp T cells after in vitro stimulation. CD4(+)sp T cells almost exclusively upregulate a CD8αα homodimer, whereas CD8α(+)sp T cells can become CD4(+)CD8αβ(+) or CD4(+)CD8αα(+). Even in the absence of other cells, highly purified CD4(+)sp T cells can become double-positive upon in vitro stimulation, whereas highly purified CD8α(+)sp T cells fail to do so. However, CD8α(+)sp T cells can additionally express CD4 when stimulated in the presence of CD4(-)CD8α(-) double-negative (dn) cells or more efficiently when stimulated in the presence of CD4(+)sp T cells. Soluble factors secreted by CD4(+)sp T cells are sufficient for the upregulation of CD4 on CD8α(+)sp T cells, but direct cell-cell contact between CD4(+)sp and CD8α(+)sp T cells is more efficient. mRNA analysis shows that additional CD4 expression on CD8α(+)sp T cells results from de novo synthesis. Thus, uptake of soluble CD4 or trogocytosis is less likely as mechanism for generation of canine double-positive T cells. CD4(+)CD

  10. Liver tryptase-positive mast cells and fibrosis in children with hepatic echinococcosis

    Directory of Open Access Journals (Sweden)

    Gulubova Maya

    2005-01-01

    Full Text Available The hepatic echinococcosis in children is a serious surgical problem. The aim of this study is to investigate the participation of mast cells in liver inflammatory reactions triggered by echinococcal cysts. Liver biopsy samples were collected from the tissue surrounding the cysts from 16 sick children (11 boys and 5 girls in the course of abdominal surgery and from 5 controls. Light and ultrastructural immunocytochemistry was performed using monoclonal antibody against tryptase. Light microscopical immunocytochemistry revealed abundance of tryptase-positive (MCT mast cells in the capsules of the cysts (43.58 cells/mm2. There were also observed greatly increased numbers of mast cells in portal tracts surrounding the cyst, compared to those of control biopsies (26.49 vs. 1.78 cells/mm2, p=0.0009, Mann-Whitney U test. Based on the ultrastructural appearance of tryptase-positive mast cell granules, morphological sings of activation of most of the mast cells were distinguished. In conclusion, we suggest that the accumulated and activated tryptase-positive mast cells in liver tissues surrounding the echinococcal cysts play a crucial role in modulation of the inflammatory liver response and could induce chronic inflammation and fibrogenesis, resulting in serious liver injury such as nonspecific reactive hepatitis.

  11. Phosphatidylserine-positive particles in the apical domain of sensory hair cells

    Institute of Scientific and Technical Information of China (English)

    SHI Xiao-rui; Alfred Nuttall

    2006-01-01

    Apical membrane recycling has been proposed to be important for normal hair cell function. The current study reports an in vitro work that demonstrates the presence of phosphatidylserine (PS) and PS-positive vesicles labeled by Annexin V in the apical portion of hair cells. The following characteristics of the PS-positive vesicles were noticed using scanning confocal fluorescence microscopy: (1) variable sizes around 200 nm; (2)variable distribution patterns (either uniformly along individual stereocilia in the hair bundle or irregular) in the stereocilia from cell to cell; (3) variable sizes and numbers at locations along the border of the cuticular plate (CP),with a large number of them located at the vestigal kinocilial location; (4) motility with some of the vesicles during the observation period; (5) increase in PS labeling and the number of PS-positive vesicles after loud sound stimulation; and (6) decreased PS labeling and PS-positive vesicle numbers following treatment with LY-294002, a PI3 -kinase inhibitor. These results suggest that the presence of PS-positive vesicles at the apical area of hair cells may be indicative of vesicle shedding or transportation of a protein or rafts.

  12. Isolation and analysis of SSEA-4 positive cells derived from fetal marrow mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    LIU Daqing; PEI Xuetao; YANG Yinxiang; GAO Yanhong; YUAN Hongfeng; QIN Lipeng; WANG Yunfang; NAN Xue; SHI Shuangshuang; YUE Wen

    2006-01-01

    A big issue in stem cell research is to derive prospective totipotential stem cells. In this study, fMSC-SSEA-4 cells expressing SSEA-4 antigen were isolated from fetal marrow masenchymal stem cells (fMSCs) using immunomagnetic bead sorting technique. The totipotent cells were identified and their biological characteristics were further studied. The expression of Oct-4 and SSEA-4, carcino- genicity, and the ability to differentiation of fMSC- SSEA-4 cells were evaluated to verify the totipotent potential. fMSC-SSEA-4 cells were isolated successfully from fMSCs (2.5% among fMSCs), while no obvious differences were seen in morphology, growth curve, cell cycle and immunophenotype, Oct-4 and SSEA-4 expression between fMSC-SSEA-4 cells and fMSCs. fMSC-SSEA-4 cells showed normal diploid chromosome karyotype and no carcinoma was induced after inoculation into nude mice. fMSC- SSEA-4 cells could be induced to fat cells, osteogenic cells and neuron-like cells in vitro with different induced factors. The results indicated that there may be a few totipotent cells among the fMSCs and it may offer the experimental basis for the further study and application of fMSCs.

  13. From elasticity to inelasticity in cancer cell mechanics: A loss of scale-invariance

    Science.gov (United States)

    Laperrousaz, B.; Drillon, G.; Berguiga, L.; Nicolini, F.; Audit, B.; Satta, V. Maguer; Arneodo, A.; Argoul, F.

    2016-08-01

    Soft materials such as polymer gels, synthetic biomaterials and living biological tissues are generally classified as viscoelastic or viscoplastic materials, because they behave neither as pure elastic solids, nor as pure viscous fluids. When stressed beyond their linear viscoelastic regime, cross-linked biopolymer gels can behave nonlinearly (inelastically) up to failure. In living cells, this type of behavior is more frequent because their cytoskeleton is basically made of cross-linked biopolymer chains with very different structural and flexibility properties. These networks have high sensitivity to stress and great propensity to local failure. But in contrast to synthetic passive gels, they can "afford" these failures because they have ATP driven reparation mechanisms which often allow the recovery of the original texture. A cell pressed in between two plates for a long period of time may recover its original shape if the culture medium brings all the nutrients for keeping it alive. When the failure events are too frequent or too strong, the reparation mechanisms may abort, leading to an irreversible loss of mechanical homeostasis and paving the way for chronic diseases such as cancer. To illustrate this discussion, we consider a model of immature cell transformation during cancer progression, the chronic myelogenous leukemia (CML), where the formation of the BCR-ABL oncogene results from a single chromosomal translocation t(9; 22). Within the assumption that the cell response to stress is scale invariant, we show that the power-law exponent that characterizes their mechanosensitivity can be retrieved from AFM force indentation curves. Comparing control and BCR-ABL transduced cells, we observe that in the later case, one month after transduction, a small percentage the cancer cells no longer follows the control cell power law, as an indication of disruption of the initial cytoskeleton network structure.

  14. Biochanin A Modulates Cell Viability, Invasion, and Growth Promoting Signaling Pathways in HER-2-Positive Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Vikas Sehdev

    2009-01-01

    Full Text Available Overexpression of HER-2 receptor is associated with poor prognosis and aggressive forms of breast cancer. Scientific literature indicates a preventive role of isoflavones in cancer. Since activation of HER-2 receptor initiates growth-promoting events in cancer cells, we studied the effect of biochanin A (an isoflavone on associated signaling events like receptor activation, downstream signaling, and invasive pathways. HER-2-positive SK-BR-3 breast cancer cells, MCF-10A normal breast epithelial cells, and NIH-3T3 normal fibroblast cells were treated with biochanin A (2–100 μM for 72 hours. Subsequently cell viability assay, western blotting and zymography were carried out. The data indicate that biochanin A inhibits cell viability, signaling pathways, and invasive enzyme expression and activity in SK-BR-3 cancer cells. Biochanin A did not inhibit MCF-10A and NIH-3T3 cell viability. Therefore, biochanin A could be a unique natural anticancer agent which can selectively target cancer cells and inhibit multiple signaling pathways in HER-2-positive breast cancer cells.

  15. Synergistic apoptosis induction in leukemic cells by the phosphatase inhibitor salubrinal and proteasome inhibitors.

    Directory of Open Access Journals (Sweden)

    Hannes C A Drexler

    Full Text Available BACKGROUND: Cells adapt to endoplasmic reticulum (ER-stress by arresting global protein synthesis while simultaneously activating specific transcription factors and their downstream targets. These processes are mediated in part by the phosphorylation-dependent inactivation of the translation initiation factor eIF2alpha. Following restoration of homeostasis protein synthesis is resumed when the serine/threonine-protein phosphatase PP1 dephosphorylates and reactivates eIF2alpha. Proteasome inhibitors, used to treat multiple myeloma patients evoke ER-stress and apoptosis by blocking the ER-associated degradation of misfolded proteins (ERAD, however, the role of eIF2alpha phosphorylation in leukemic cells under conditions of proteasome inhibitor-mediated ER stress is currently unclear. METHODOLOGY AND PRINCIPAL FINDINGS: Bcr-Abl-positive and negative leukemic cell lines were used to investigate the functional implications of PP1-related phosphatase activities on eIF2alpha phosphorylation in proteasome inhibitor-mediated ER stress and apoptosis. Rather unexpectedly, salubrinal, a recently identified PP1 inhibitor capable to protect against ER stress in various model systems, strongly synergized with proteasome inhibitors to augment apoptotic death of different leukemic cell lines. Salubrinal treatment did not affect the phosphorlyation status of eIF2alpha. Furthermore, the proapoptotic effect of salubrinal occurred independently from the chemical nature of the proteasome inhibitor, was recapitulated by a second unrelated phosphatase inhibitor and was unaffected by overexpression of a dominant negative eIF2alpha S51A variant that can not be phosphorylated. Salubrinal further aggravated ER-stress and proteotoxicity inflicted by the proteasome inhibitors on the leukemic cells since characteristic ER stress responses, such as ATF4 and CHOP synthesis, XBP1 splicing, activation of MAP kinases and eventually apoptosis were efficiently abrogated by the

  16. PDGFBB promotes PDGFRα-positive cell migration into artificial bone in vivo

    International Nuclear Information System (INIS)

    Highlights: ► We examined effects of PDGFBB in PDGFRα positive cell migration in artificial bones. ► PDGFBB was not expressed in osteoblastic cells but was expressed in peripheral blood cells. ► PDGFBB promoted PDGFRα positive cell migration into artificial bones but not osteoblast proliferation. ► PDGFBB did not inhibit osteoblastogenesis. -- Abstract: Bone defects caused by traumatic bone loss or tumor dissection are now treated with auto- or allo-bone graft, and also occasionally by artificial bone transplantation, particularly in the case of large bone defects. However, artificial bones often exhibit poor affinity to host bones followed by bony union failure. Thus therapies combining artificial bones with growth factors have been sought. Here we report that platelet derived growth factor bb (PDGFBB) promotes a significant increase in migration of PDGF receptor α (PDGFRα)-positive mesenchymal stem cells/pre-osteoblastic cells into artificial bone in vivo. Growth factors such as transforming growth factor beta (TGFβ) and hepatocyte growth factor (HGF) reportedly inhibit osteoblast differentiation; however, PDGFBB did not exhibit such inhibitory effects and in fact stimulated osteoblast differentiation in vitro, suggesting that combining artificial bones with PDGFBB treatment could promote host cell migration into artificial bones without inhibiting osteoblastogenesis.

  17. Valproic acid triggers differentiation and apoptosis in AML1/ETO-positive leukemic cells specifically.

    Science.gov (United States)

    Zapotocky, Michal; Mejstrikova, Ester; Smetana, Karel; Stary, Jan; Trka, Jan; Starkova, Julia

    2012-06-28

    Valproic acid (VPA) has extensive effects on leukemic blasts through its inhibition of histone deacetylases. The main goal of this study was to identify the subgroup of patients who may benefit most from VPA treatment. We examined the significance of t(8;21) chromosomal aberration for VPA treatment response among acute myeloid leukemia (AML) patients by direct comparison of AML1/ETO-negative vs. positive leukemic cell-lines as well as bone marrow blasts from AML patients. In t(8;21) AML, leukemogenesis is supposed to be induced via aberrant recruitment of histone deacetylases. AML cell lines of different genotypes (Kasumi-1, Kasumi-6, MV4;11, K562) and diagnostic bone marrow samples from patients were treated with VPA. VPA induced apoptosis in AML1/ETO-positive and MLL-AF4-positive cells in a dose-dependent manner. Differentiation, as indicated by changes in immunophenotype, was observed only in AML1/ETO-positive cells. VPA increased the expression of AML1 target genes - PU.1, C/EBPa, BPI and IGFBP7 only in AML1/ETO-positive cells. This AML1/ETO-specific effect was confirmed also using patient blasts isolated at the time of diagnosis. AML1/ETO-positive leukemia shows specific mechanism of VPA residing from differentiation followed by apoptosis that is accompanied by an increase in the expression of repressed AML1 target genes. Our data suggest that AML1/ETO-positive patients might derive the greatest benefit from VPA treatment. PMID:22261333

  18. Identification and analysis of CXCR4-positive synovial sarcoma-initiating cells.

    Science.gov (United States)

    Kimura, T; Wang, L; Tabu, K; Tsuda, M; Tanino, M; Maekawa, A; Nishihara, H; Hiraga, H; Taga, T; Oda, Y; Tanaka, S

    2016-07-28

    Synovial sarcoma accounts for almost 10% of all soft tissue sarcomas, and its prognosis is poor with 5-year survival rates at 36%. Thus, new treatments and therapeutic targets for synovial sarcoma are required. Tumor-initiating cells have been defined by the ability for self-renewal and multipotent differentiation, and they exhibit higher tumorigenic capacity, chemoresistance and radiation resistance, expecting to be a new therapeutic target. In synovial sarcoma, the presence of such stemness remains largely unclear; thus, we analyzed whether synovial sarcoma possessed tumor-initiating cells and explored specific markers, and we discovered that synovial sarcoma cell lines possessed heterogeneity by way of containing a sphere-forming subpopulation highly expressing NANOG, OCT4 and SOX2. By expression microarray analysis, CXCR4 was identified to be highly expressed in the sphere subpopulation and correlated with stem-cell-associated markers. Inhibition of CXCR4 suppressed the cell proliferation of synovial sarcoma cell lines in vitro. The tumor-initiating ability of CXCR4-positive cells was demonstrated by xenograft propagation assay. CXCR4-positive cells showed higher tumorigenicity than negative ones and possessed both self-renewal and multipotent differentiation ability. Immunohistochemical analysis of 39 specimens of synovial sarcoma patients revealed that CXCR4 strongly correlated with poor prognosis of synovial sarcoma. Thus, we conclude that CXCR4 is the marker of synovial sarcoma-initiating cells, a new biomarker for prognosis and a new potential therapeutic target. PMID:26640147

  19. Positional variation of antipodal cells in polyembryonic rice ApⅢ before and after fertilization

    Institute of Scientific and Technical Information of China (English)

    YUAN Liangchen; CHEN Yongzhe; MU Xijin; LIN Jinxing

    2003-01-01

    The variation in the position of antipodal cells before and after fertilization in polyembryonic rice strain ApⅢ was studied by using conventional sectioning technique. It was shown that initially the three antipodal cells lie at the chalazal end of the young embryo sacs. Along with the development of embryo sac, the antipodal cells proliferate into a multicellular structure containing up to about 20 cells. Most of the cells migrate to its dorsal side when the embryo sacs turn into mature. In a few embryo sacs the antipodal cells are situated in the positions adjacent to the egg apparatus at the micropylar end, while in other sacs, they spread from the midst of the embryo sac to the micropylar end clinging to the nucellar tissue. Furthermore, it is reported for the first time that cells of some egg apparatus or proembryos disorganize when antipodal cells lie at the micropylar end, indicating that the function of the antipodal cells may vary during the embryogenesis in rice ApⅢ.

  20. Antibody to a molecular marker of cell position inhibits synapse formation in retina.

    OpenAIRE

    Trisler, D.; Bekenstein, J; Daniels, M P

    1986-01-01

    A topographic gradient of TOP molecules in retina can be used to identify neuron position. Antibody to TOP from hybridoma cells that were injected into in vivo embryo eyes diffused into the retina and bound in a topographic gradient of [antibody.TOP] ([Ab.TOP]) complexes. Synapse formation in retina was inhibited in the presence of anti-TOP antibody. This suggests that TOP is involved in synapse formation and that recognition of position by neurons is necessary for normal synapse formation.

  1. Activating β-catenin signaling in CD133-positive dermal papilla cells increases hair inductivity.

    Science.gov (United States)

    Zhou, Linli; Yang, Kun; Xu, Mingang; Andl, Thomas; Millar, Sarah E; Boyce, Steven; Zhang, Yuhang

    2016-08-01

    Bioengineering hair follicles using cells isolated from human tissue remains a difficult task. Dermal papilla (DP) cells are known to guide the growth and cycling activities of hair follicles by interacting with keratinocytes. However, DP cells quickly lose their inductivity during in vitro passaging. Rodent DP cell cultures need external addition of growth factors, including WNT and BMP molecules, to maintain the hair inductive property. CD133 is expressed by a subpopulation of DP cells that are capable of inducing hair follicle formation in vivo. We report here that expression of a stabilized form of β-catenin promoted clonal growth of CD133-positive (CD133+) DP cells in in vitro three-dimensional hydrogel culture while maintaining expression of DP markers, including alkaline phosphatase (AP), CD133, and integrin α8. After a 2-week in vitro culture, cultured CD133+ DP cells with up-regulated β-catenin activity led to an accelerated in vivo hair growth in reconstituted skin compared to control cells. Further analysis showed that matrix cell proliferation and differentiation were significantly promoted in hair follicles when β-catenin signaling was up-regulated in CD133+ DP cells. Our data highlight an important role for β-catenin signaling in promoting the inductive capability of CD133+ DP cells for in vitro expansion and in vivo hair follicle regeneration, which could potentially be applied to cultured human DP cells. PMID:27312243

  2. Design of a single-cell positioning controller using electroosmotic flow and image processing.

    Science.gov (United States)

    Ay, Chyung; Young, Chao-Wang; Chen, Jhong-Yin

    2013-01-01

    The objective of the current research was not only to provide a fast and automatic positioning platform for single cells, but also improved biomolecular manipulation techniques. In this study, an automatic platform for cell positioning using electroosmotic flow and image processing technology was designed. The platform was developed using a PCI image acquisition interface card for capturing images from a microscope and then transferring them to a computer using human-machine interface software. This software was designed by the Laboratory Virtual Instrument Engineering Workbench, a graphical language for finding cell positions and viewing the driving trace, and the fuzzy logic method for controlling the voltage or time of an electric field. After experiments on real human leukemic cells (U-937), the success of the cell positioning rate achieved by controlling the voltage factor reaches 100% within 5 s. A greater precision is obtained when controlling the time factor, whereby the success rate reaches 100% within 28 s. Advantages in both high speed and high precision are attained if these two voltage and time control methods are combined. The control speed with the combined method is about 5.18 times greater than that achieved by the time method, and the control precision with the combined method is more than five times greater than that achieved by the voltage method. PMID:23698272

  3. Design of a Single-Cell Positioning Controller Using Electroosmotic Flow and Image Processing

    Directory of Open Access Journals (Sweden)

    Jhong-Yin Chen

    2013-05-01

    Full Text Available The objective of the current research was not only to provide a fast and automatic positioning platform for single cells, but also improved biomolecular manipulation techniques. In this study, an automatic platform for cell positioning using electroosmotic flow and image processing technology was designed. The platform was developed using a PCI image acquisition interface card for capturing images from a microscope and then transferring them to a computer using human-machine interface software. This software was designed by the Laboratory Virtual Instrument Engineering Workbench, a graphical language for finding cell positions and viewing the driving trace, and the fuzzy logic method for controlling the voltage or time of an electric field. After experiments on real human leukemic cells (U-937, the success of the cell positioning rate achieved by controlling the voltage factor reaches 100% within 5 s. A greater precision is obtained when controlling the time factor, whereby the success rate reaches 100% within 28 s. Advantages in both high speed and high precision are attained if these two voltage and time control methods are combined. The control speed with the combined method is about 5.18 times greater than that achieved by the time method, and the control precision with the combined method is more than five times greater than that achieved by the voltage method.

  4. Tuberculosis-specific CD8 cells in HLA A*02-positive TB- and LTBI patients

    DEFF Research Database (Denmark)

    Fløe, Andreas; Brix, Liselotte; Wejse, Christian;

    Background: Understanding the CD8+ response against Mycobacterium tuberculosis (MTB) may be a key to improved TB diagnostics and vaccine development. Aims and Objectives: To detect a CD8+ T-cell response against Mycobacterium tuberculosis (MTB) in active tuberculosis (TB) and latent TB (LTBI), in...... candidates, from which we constructed MHC multimers (Dextramers). Peripheral blood mononuclear cells (PBMC) from 7 TB-patients, 16 LTBI patients and 8 MTB-exposed, IGRA-negative, healthy subjects (HE), all HLA A*02 positive, were stained with the Dextramers and with anti-CD8 and anti-CD3, and analyzed on a...... flow cytometer. The MTB epitopes were analyzed in 5 pools (3-7 epitopes each). Positive responses included >0.001 % of CD8+, CD3+ cells, supported by inspection of flow cytometry plots. Results: MTB-specific CD8+ T-cells were detected more often in TB patients (57%) than in LTBI patients (41%) and in...

  5. Canine CD4+CD8+ double positive T cells in peripheral blood have features of activated T cells.

    Science.gov (United States)

    Bismarck, Doris; Schütze, Nicole; Moore, Peter; Büttner, Mathias; Alber, Gottfried; Buttlar, Heiner v

    2012-10-15

    In dogs a CD4(+)CD8(+) double positive T cell subpopulation exists that has not been phenotypically defined yet. We demonstrate that canine CD4(+)CD8(+) T cells are mature CD1a(-) and TCRαβ(+) T cells. To analyse the activation potential of CD4(+)CD8(+) T cells, PBMC from dogs vaccinated against canine distemper virus (CDV) were re-stimulated with CDV. Upon antigen-specific stimulation, the CD4(+)CD8(+) T cell fraction increases and consists nearly exclusively of proliferated cells. Similarly, other features of activated effector/memory T cells such as up-regulation of CD25 and MHC-II as well as down-regulation of CD62L (L-selectin) were observed in CD4(+)CD8(+) T cells after stimulation. Canine CD4(+)CD8(+) T cells are less abundant, but more heterogeneous than porcine ones, comprising a small proportion expressing the β chain of CD8 in addition to the CD8α chain, like human CD4(+)CD8(+) T cells. In summary, this analysis provides the basis for functional characterisation of the in vivo relevance of CD4(+)CD8(+) T cells in T-cell mediated immunity. PMID:22789871

  6. Subculture of Germ Cell-Derived Colonies with GATA4-Positive Feeder Cells from Neonatal Pig Testes

    Directory of Open Access Journals (Sweden)

    Kyung Hoon Lee

    2016-01-01

    Full Text Available Enrichment of spermatogonial stem cells is important for studying their self-renewal and differentiation. Although germ cell-derived colonies (GDCs have been successfully cultured from neonatal pig testicular cells under 31°C conditions, the short period of in vitro maintenance (<2 months limited their application to further investigations. To develop a culture method that allows for in vitro maintenance of GDCs for long periods, we subcultured the GDCs with freshly prepared somatic cells from neonatal pig testes as feeder cells. The subcultured GDCs were maintained up to passage 13 with the fresh feeder cells (FFCs and then frozen. Eight months later, the frozen GDCs could again form the colonies on FFCs as shown in passages 1 to 13. Immunocytochemistry data revealed that the FFCs expressed GATA-binding protein 4 (GATA4, which is also detected in the cells of neonatal testes and total testicular cells, and that the expression of GATA4 was decreased in used old feeder cells. The subcultured GDCs in each passage had germ and stem cell characteristics, and flow cytometric analyses revealed that ~60% of these cells were GFRα-1 positive. In conclusion, neonatal pig testes-derived GDCs can be maintained for long periods with GATA4-expressing testicular somatic cells.

  7. Combinatorial treatment of mammospheres with trastuzumab and salinomycin efficiently targets HER2-positive cancer cells and cancer stem cells.

    Science.gov (United States)

    Oak, Prajakta S; Kopp, Florian; Thakur, Chitra; Ellwart, Joachim W; Rapp, Ulf R; Ullrich, Axel; Wagner, Ernst; Knyazev, Pjotr; Roidl, Andreas

    2012-12-15

    A major obstacle in the successful treatment of cancer is the occurrence of chemoresistance. Cancer cells surviving chemotherapy and giving rise to a recurrence of the tumor are termed cancer stem cells and can be identified by elevated levels of certain stem cell markers. Eradication of this cell population is a priority objective in cancer therapy. Here, we report elevated levels of stem cell markers in MCF-7 mammospheres. Likewise, an upregulation of HER2 and its differential expression within individual cells of mammospheres was observed. Sorting for HER2(high) and HER2(low) cells revealed an upregulation of stem cell markers NANOG, OCT4 and SOX2 in the HER2(low) cell fraction. Accordingly, HER2(low) cells also showed reduced proliferation, ductal-like outgrowths and an increased number of colonies in matrigel. Xenografts from subcutaneously injected HER2(low) sorted cells exihibited earlier onset but slower growth of tumors and an increase in stem cell markers compared to tumors developed from the HER2(high) fraction. Treatment of mammospheres with salinomycin reduced the expression of SOX2 indicating a selective targeting of cancer stem cells. Trastuzumab however, did not reduce the expression of SOX2 in mammospheres. Furthermore, a combinatorial treatment of mammospheres with trastuzumab and salinomycin was superior to single treatment with each drug. Thus, targeting HER2 expressing tumors with anti-HER2 therapies will not necessarily eliminate cancer stem cells and may lead to a more aggressive cancer cell phenotype. Our study demonstrates efficient killing of both HER2 positive cells and cancer stem cells, hence opening a possibility for a new combinatorial treatment strategy. PMID:22511343

  8. Chemokine-guided cell positioning in the lymph node orchestrates the generation of adaptive immune responses.

    Science.gov (United States)

    Lian, Jeffrey; Luster, Andrew D

    2015-10-01

    The generation of adaptive immune responses occurs in the lymph node (LN) and requires that lymphocytes locate and interact with cognate antigen-bearing dendritic cells. This process requires the coordinated movement of both innate and adaptive immune cells, and is orchestrated by the chemokine family of chemotactic cytokines. Upon initiation of inflammation, the LN undergoes dramatic changes that include the marked induction of specific chemokines in distinct regions of the reactive LN. These chemokine rich domains establish LN niches that facilitate the differentiation of CD4+ T cells into effector cell subsets and the rapid activation of memory CD8+ T cells. This review will focus on recent advances highlighting the importance of LN chemokines for shaping adaptive immune responses by controlling immune cell migration, positioning, and interactions in the reactive LN. PMID:26067148

  9. Cytokeratin-positive interstitial reticulum cell sarcoma: extranodal presentations mimicking carcinoma.

    Science.gov (United States)

    Sundersingh, Shirley; Majhi, Urmila; Krishnamurthy, Arvind; Velusami, Sri Devi

    2013-01-01

    Cytokeratin-positive interstitial reticulum cell (CIRC) sarcoma is a rare type of dendritic cell tumor derived from a subset of fibroblastic reticular cells. Expression of cytokeratins and extranodal location of these tumors can lead to a misdiagnosis of carcinoma. We report two cases of CIRC sarcomas primarily involving the scalp and breast. Patients were referred with an initial diagnosis of carcinoma. Case 1 underwent wide local excision of the scalp tumor with left posterolateral neck dissection. Case 2 had modified radical mastectomy for the tumor in left breast. Histopathological examination of both specimens showed an epithelioid to spindle cell malignant tumor that co-expressed CK 8, CK 18, vimentin, and smooth muscle actin. A diagnosis of CIRC sarcoma was made. Pathologists should be aware of this subset of dendritic cell sarcoma. Carcinomas, other sarcomas of the accessory dendritic cell family, and poorly differentiated malignant tumors have to be ruled out by combination of morphology, immunohistochemistry, and electron microscopic studies. PMID:24056661

  10. Retinal Targets ALDH Positive Cancer Stem Cell and Alters the Phenotype of Highly Metastatic Osteosarcoma Cells

    Directory of Open Access Journals (Sweden)

    Xiaodong Mu

    2015-01-01

    Full Text Available Aldehyde dehydrogenase (ALDH is a cancer stem cell marker. Retinoic acid has antitumor properties, including the induction of apoptosis and inhibition of proliferation. Retinal, the precursor of retinoic acid, can be oxidized to retinoic acid by dehydrogenases, including ALDH. We hypothesized that retinal could potentially be transformed to retinoic acid with higher efficiency by cancer stem cells, due to the higher ALDH activity. We previously observed that ALDH activity is greater in highly metastatic K7M2 osteosarcoma (OS cells than in nonmetastatic K12 OS cells. We also demonstrated that ALDH activity correlates with clinical metastases in bone sarcoma patients, suggesting that ALDH may be a therapeutic target specific to cells with high metastatic potential. Our current results demonstrated that retinal preferentially affected the phenotypes of ALDH-high K7M2 cells in contrast to ALDH-low K12 cells, which could be mediated by the more efficient transformation of retinal to retinoic acid by ALDH in K7M2 cells. Retinal treatment of highly metastatic K7M2 cells decreased their proliferation, invasion capacity, and resistance to oxidative stress. Retinal altered the expression of metastasis-related genes. These observations indicate that retinal may be used to specifically target metastatic cancer stem cells in OS.

  11. Argyrophilic nucleolar organizer region in MIB-1 positive cells in non-small cell lung cancer: clinicopathological significance and survival

    International Nuclear Information System (INIS)

    To evaluate the relation between argyrophilic nucleolar organizer region (AgNOR)-associated proteins and clinicopathological parameters and survival in non-small-cell lung cancer (NSCLC). A total of 207 surgical specimens diagnosed as NSCLC were included in this study. Double-staining procedures were performed using antigen Ki-67 (clone MIB-1) and silver nitrate by immunohistochemical and AgNOR-staining methods. The AgNOR area in MIB-1-positive cells of NSCLC is related to clinicopathological parameters under the TNM (tumor, node, and metastasis) system. The survival of patients with small AgNOR area in MIB-1-positive cells is better than that of patients with large AgNOR area. Molecular, biological (AgNOR area in MIB-1-positive cells), and clinicopathological (greatest tumor dimension, metastases to regional lymph nodes, histology, and differentiation) parameters are independent prognostic factors of NSCLC. The AgNOR area in MIB-1-positive cells is related to clinicopathological parameters and survival in NSCLC

  12. Proteomic Profiling of Ex Vivo Expanded CD34-Positive Haematopoetic Cells Derived from Umbilical Cord Blood

    Directory of Open Access Journals (Sweden)

    Heiner Falkenberg

    2013-01-01

    Full Text Available Ex vivo expansion of haematopoetic cells by application of specific cytokines is one approach to overcome boundaries in cord blood transplantation due to limited numbers of haematopoetic stem cells. While many protocols describe an effective increase of total cell numbers and the amount of CD34-positive cells, it still remains unclear if and how the procedure actually affects the cells’ properties. In the presented publications, CD34-positive cells were isolated from cord blood and expanded for up to 7 days in media supplemented with stem cell factor (SCF, thrombopoietin (THPO, interleukin 6 (IL-6, and fms-related tyrosine kinase 3 ligand (FLT3lg. At days 3 and 7, expanded cells were harvested and analyzed by flow cytometry and quantitative proteomics. 2970 proteins were identified, whereof proteomic analysis showed 440 proteins significantly changed in abundance during ex vivo expansion. Despite the fact that haematopoetic cells still expressed CD34 on the surface after 3 days, major changes in regard to the protein profile were observed, while further expansion showed less effect on the proteome level. Enrichment analysis of biological processes clearly showed a proteomic change toward a protein biosynthesis phenotype already within the first three days of expression.

  13. Generation model of positional values as cell operation during the development of multicellular organisms.

    Science.gov (United States)

    Ogawa, Ken-ichiro; Miyake, Yoshihiro

    2011-03-01

    Many conventional models have used the positional information hypothesis to explain each elementary process of morphogenesis during the development of multicellular organisms. Their models assume that the steady concentration patterns of morphogens formed in an extracellular environment have an important property of positional information, so-called "robustness". However, recent experiments reported that a steady morphogen pattern, the concentration gradient of the Bicoid protein, during early Drosophila embryonic development is not robust for embryo-to-embryo variability. These reports encourage a reconsideration of a long-standing problem in systematic cell differentiation: what is the entity of positional information for cells? And, what is the origin of the robust boundary of gene expression? To address these problems at a cellular level, in this article we pay attention to the re-generative phenomena that show another important property of positional information, "size invariance". In view of regenerative phenomena, we propose a new mathematical model to describe the generation mechanism of a spatial pattern of positional values. In this model, the positional values are defined as the values into which differentiable cells transform a spatial pattern providing positional information. The model is mathematically described as an associative algebra composed of various terms, each of which is the multiplication of some fundamental operators under the assumption that the operators are derived from the remarkable properties of cell differentiation on an amputation surface in regenerative phenomena. We apply this model to the concentration pattern of the Bicoid protein during the anterior-posterior axis formation in Drosophila, and consider the conditions needed to establish the robust boundary of the expression of the hunchback gene. PMID:21167904

  14. Cytotoxic effect of lapatinib is restricted to human papillomavirus-positive head and neck squamous cell carcinoma cell lines

    Directory of Open Access Journals (Sweden)

    Fumagalli I

    2015-02-01

    Full Text Available Ingrid Fumagalli,1,2 Delphine Dugue,1 Jean Emmanuel Bibault,1,2 Céline Clémenson,1 Marie Catherine Vozenin,1 Michele Mondini,1,* Eric Deutsch1,3,*1Inserm U1030, Molecular Radiotherapy, LABEX LERMIT, Gustave Roussy, University Paris XI, Villejuif, France; 2Radiation Therapy Department, Oscar Lambret Comprehensive Cancer Center, Lille, France; 3Department of Radiation Oncology, Gustave Roussy, University Paris XI, Villejuif, France*These authors share senior authorshipBackground: Lapatinib is a dual epidermal growth factor receptor (EGFR and HER2 inhibitor. Overexpression of these receptors is frequently observed in head and neck squamous cell carcinoma (HNSCC. As growing proportion of HNSCC is characterized by human papillomavirus (HPV infection, we aimed at evaluating the efficacy of lapatinib as function of HPV status in HNSCC cell lines.Methods: Two HPV-positive and two HPV-negative HNSCC cell lines were used. Proliferation, cell cycle, and Annexin V assays were performed to test their sensitivity to lapatinib. Combination of lapatinib and ionizing radiation was evaluated with clonogenic survival assays. Akt, EGFR and HER2, and E6/E7 expression and activation were analyzed by immunoblotting and quantitative reverse transcription polymerase chain reaction.Results: Lapatinib reduced E6 and E7 expression and Akt phosphorylation, inhibited cell proliferation and induced cell death in HPV-positive cell lines. An additive effect of lapatinib with radiation was observed in these cells. Lapatinib had no effect on HPV-negative cells.Conclusion: Lapatinib efficacy restricted to the HPV-positive cells suggests that HPV status could be a potential marker for enhanced response to lapatinib in HNSCC.Keywords: HPV, lapatinib, ionizing radiation, EGFR, HER2, tyrosine kinase inhibitor

  15. Positive and negative selection of T cells in T-cell receptor transgenic mice expressing a bcl-2 transgene.

    OpenAIRE

    Strasser, A.; Harris, A W; von Boehmer, H; Cory, S

    1994-01-01

    To explore the role of bcl-2 in T-cell development, a bcl-2 transgene was introduced into mice expressing a T-cell receptor (TCR) transgene encoding reactivity for the mouse male antigen HY presented by the H-2Db class I antigen of the major histocompatibility complex (MHC). Normal thymic development is contingent on the ability of immature thymocytes to interact with self-MHC molecules presented by thymic stroma (positive selection). Thus, thymocyte numbers are low in femal...

  16. Common Effects on Cancer Cells Exerted by a Random Positioning Machine and a 2D Clinostat

    OpenAIRE

    Svejgaard, Benjamin; Wehland, Markus; Ma, Xiao; Kopp, Sascha; Sahana, Jayashree; Warnke, Elisabeth; Aleshcheva, Ganna; Hemmersbach, Ruth; Hauslage, Jens; Grosse, Jirka; Bauer, Johann; Corydon, Thomas Juhl; Islam, Tawhidul; Infanger, Manfred; Grimm, Daniela

    2015-01-01

    In this study we focused on gravity-sensitive proteins of two human thyroid cancer cell lines (ML-1; RO82-W-1), which were exposed to a 2D clinostat (CLINO), a random positioning machine (RPM) and to normal 1g-conditions. After a three (3d)- or seven-day-culture (7d) on the two devices, we found both cell types growing three-dimensionally within multicellular spheroids (MCS) and also cells remaining adherent (AD) to the culture flask, while 1g-control cultures only formed adherent monolayers,...

  17. Common Effects on Cancer Cells Exerted by a Random Positioning Machine and a 2D Clinostat

    OpenAIRE

    Svejgaard, Benjamin; Wehland, Markus; Ma, Xiao; Kopp, Sascha; Sahana, Jayashree; Warnke, Elisabeth; Aleshcheva, Ganna; Hemmersbach, Ruth; Hauslage, Jens; Grosse, Jirka; Bauer, Johann; Corydon, Thomas Juhl; Islam, Tawhidul; Infanger, Manfred; Grimm, Daniela

    2015-01-01

    In this study we focused on gravity-sensitive proteins of two human thyroid cancer cell lines (ML-1; RO82-W-1), which were exposed to a 2D clinostat (CLINO), a random positioning machine (RPM) and to normal 1g-conditions. After a three (3d)- or seven-day-culture (7d) on the two devices, we found both cell types growing three-dimensionally within multicellular spheroids (MCS) and also cells remaining adherent (AD) to the culture flask, while 1gcontrol cultures only formed adherent monolayers, ...

  18. Prevalence of t(12;21)[ETV6-RUNX1]-positive cells in healthy neonates

    DEFF Research Database (Denmark)

    Lausten-Thomsen, Ulrik; Madsen, Hans O.; Vestergaard, Therese Risom;

    2011-01-01

    t(12;21)(p13;q22)[ETV6-RUNX1] is the most common chromosomal translocation in childhood acute lymphoblastic leukemia, and it can often be backtracked to Guthrie cards supporting prenatal initiation and high levels of circulating t(12;21)-positive cells at birth. To explore the prevalence of ETV6...

  19. Positioning of polarity formation by extracellular signaling during asymmetric cell division.

    Science.gov (United States)

    Seirin Lee, Sungrim

    2016-07-01

    Anterior-posterior (AP) polarity formation of cell membrane proteins plays a crucial role in determining cell asymmetry, which ultimately generates cell diversity. In Caenorhabditis elegans, a single fertilized egg cell (P0), its daughter cell (P1), and the germline precursors (P2 and P3 cells) form two exclusive domains of different PAR proteins on the membrane along the anterior-posterior axis. However, the phenomenon of polarity reversal has been observed in which the axis of asymmetric cell division of the P2 and P3 cells is formed in an opposite manner to that of the P0 and P1 cells. The extracellular signal MES-1/SRC-1 has been shown to induce polarity reversal, but the detailed mechanism remains elusive. Here, using a mathematical model, I explore the mechanism by which MES-1/SRC-1 signaling can induce polarity reversal and ultimately affect the process of polarity formation. I show that a positive correlation between SRC-1 and the on-rate of PAR-2 is the essential mechanism underlying polarity reversal, providing a mathematical basis for the orientation of cell polarity patterns. PMID:27086039

  20. CD13-positive anaplastic large cell lymphoma of T-cell origin--a diagnostic and histogenetic problem.

    Science.gov (United States)

    Popnikolov, N K; Payne, D A; Hudnall, S D; Hawkins, H K; Kumar, M; Norris, B A; Elghetany, M T

    2000-12-01

    The expression of myelomonocytic-associated antigens in anaplastic large cell lymphomas (ALCLs), particularly those presenting in extranodal sites, can make their distinction from extramedullary myeloid cell tumors (EMCTs) or histiocytic tumors problematic. Yet, this distinction is clinically significant because of its therapeutic and prognostic implications. Herein, we describe a case of extranodal anaplastic lymphoma kinase-positive CD30-positive ALCL of T-cell origin in a 12-year-old boy, which was initially called an EMCT because of the expression of CD13 and HLA-DR detected by flow cytometry and the absence of other T-cell-related surface markers. However, the detection of cytoplasmic CD3 by flow cytometry prompted further studies. The tumor was composed of large cells with abundant slightly eosinophilic vacuolated cytoplasm and ovoid or reniform nuclei with a few small nucleoli. Using immunohistochemistry, the tumor was positive for CD45, CD30, CD45RO, and CD43 with a strong cytoplasmic and nuclear anaplastic lymphoma kinase stain. The tumor cells showed a T-cell clonal genotype. Electron microscopy revealed no ultrastructural features of myelomonocytic or histiocytic origin. The patient responded well to the chemotherapy and was in complete remission for 10 months at the time of submission of this manuscript. Review of the literature showed inconsistencies regarding the diagnosis, nomenclature, and, therefore, treatment and prognosis of these tumors. In addition, the CD13 expression in ALCL raises some histogenetic questions and may indicate origin from a pluripotent stem cell, misprogramming during malignant transformation, or a microenvironmental effect on lymphoid cell expression of surface antigens. Therefore, ALCL should be considered in the differential diagnosis of EMCTs or histiocytic tumors, particularly when surface marker lineage assignment is ambiguous. PMID:11100061

  1. False Positive B-Cells Crossmatch after Prior Rituximab Exposure of the Kidney Donor

    Directory of Open Access Journals (Sweden)

    Judith Desoutter

    2016-01-01

    Full Text Available Crossmatching is essential prior to kidney transplantation to confirm compatibility between the donor and the recipient, particularly to prevent acute antibody-mediated rejection. An unexpected positive crossmatch may be obtained in recipients with an autoimmune disease or preexisting antibodies not detected by single-antigen bead array due to complement interference or who have been previously treated by desensitization protocols such as rituximab, antithymocyte globulin, or intravenous immunoglobulins. We report donor and recipient investigations that revealed unexpected positive B-cells crossmatch, probably due to donor cells, as the donor had received rituximab therapy shortly before organ harvesting, in a context of severe idiopathic thrombocytopenic purpura. We consequently detected unexpected Class II IgG complement-dependent cytotoxicity for all sera tested. Other laboratory investigations failed to elucidate the reasons for this recipient-related positivity.

  2. Imaging- and Flow Cytometry-based Analysis of Cell Position and the Cell Cycle in 3D Melanoma Spheroids.

    Science.gov (United States)

    Beaumont, Kimberley A; Anfosso, Andrea; Ahmed, Farzana; Weninger, Wolfgang; Haass, Nikolas K

    2015-01-01

    Three-dimensional (3D) tumor spheroids are utilized in cancer research as a more accurate model of the in vivo tumor microenvironment, compared to traditional two-dimensional (2D) cell culture. The spheroid model is able to mimic the effects of cell-cell interaction, hypoxia and nutrient deprivation, and drug penetration. One characteristic of this model is the development of a necrotic core, surrounded by a ring of G1 arrested cells, with proliferating cells on the outer layers of the spheroid. Of interest in the cancer field is how different regions of the spheroid respond to drug therapies as well as genetic or environmental manipulation. We describe here the use of the fluorescence ubiquitination cell cycle indicator (FUCCI) system along with cytometry and image analysis using commercial software to characterize the cell cycle status of cells with respect to their position inside melanoma spheroids. These methods may be used to track changes in cell cycle status, gene/protein expression or cell viability in different sub-regions of tumor spheroids over time and under different conditions. PMID:26779761

  3. CCL21/IL21-armed oncolytic adenovirus enhances antitumor activity against TERT-positive tumor cells.

    Science.gov (United States)

    Li, Yang; Li, Yi-Fei; Si, Chong-Zhan; Zhu, Yu-Hui; Jin, Yan; Zhu, Tong-Tong; Liu, Ming-Yuan; Liu, Guang-Yao

    2016-07-15

    Multigene-armed oncolytic adenoviruses are capable of efficiently generating a productive antitumor immune response. The chemokine (C-C motif) ligand 21 (CCL21) binds to CCR7 on naïve T cells and dendritic cells (DCs) to promote their chemoattraction to the tumor and resultant antitumor activity. Interleukin 21 (IL21) promotes survival of naïve T cells while maintaining their CCR7 surface expression, which increases their capacity to transmigrate in response to CCL21 chemoattraction. IL21 is also involved in NK cell differentiation and B cell activation and proliferation. The generation of effective antitumor immune responses is a complex process dependent upon coordinated interactions of various subsets of effector cells. Using the AdEasy system, we aimed to construct an oncolytic adenovirus co-expressing CCL21 and IL21 that could selectively replicate in TERTp-positive tumor cells (Ad-CCL21-IL21 virus). The E1A promoter of these oncolytic adenoviruses was replaced by telomerase reverse transcriptase promoter (TERTp). Ad-CCL21-IL21 was constructed from three plasmids, pGTE-IL21, pShuttle-CMV-CCL21 and AdEasy-1 and was homologously recombined and propagated in the Escherichia coli strain BJ5183 and the packaging cell line HEK-293, respectively. Our results showed that our targeted and armed oncolytic adenoviruses Ad-CCL21-IL21 can induce apoptosis in TERTp-positive tumor cells to give rise to viral propagation, in a dose-dependent manner. Importantly, we confirm that these modified oncolytic adenoviruses do not replicate efficiently in normal cells even under high viral loads. Additionally, we investigate the role of Ad-CCL21-IL21 in inducing antitumor activity and tumor specific cytotoxicity of CTLs in vitro. This study suggests that Ad-CCL21-IL21 is a promising targeted tumor-specific oncolytic adenovirus. PMID:27157859

  4. Programming of donor T cells using allogeneic δ-like ligand 4-positive dendritic cells to reduce GVHD in mice.

    Science.gov (United States)

    Mochizuki, Kazuhiro; Meng, Lijun; Mochizuki, Izumi; Tong, Qing; He, Shan; Liu, Yongnian; Purushe, Janaki; Fung, Henry; Zaidi, M Raza; Zhang, Yanyun; Reshef, Ran; Blazar, Bruce R; Yagita, Hideo; Mineishi, Shin; Zhang, Yi

    2016-06-23

    Alloreactive T cells play a critical role in eliminating hematopoietic malignant cells but are also the mediators of graft-versus-host disease (GVHD), a major complication that subverts the success of allogeneic hematopoietic stem cell transplantation (HSCT). However, induction of alloreactive T cells does not necessarily lead to GVHD. Here we report the development of a cellular programming approach to render alloreactive T cells incapable of causing severe GVHD in both major histocompatibility complex (MHC)-mismatched and MHC-identical but minor histocompatibility antigen-mismatched mouse models. We established a novel platform that produced δ-like ligand 4-positive dendritic cells (Dll4(hi)DCs) from murine bone marrow using Flt3 ligand and Toll-like receptor agonists. Upon allogeneic Dll4(hi)DC stimulation, CD4(+) naïve T cells underwent effector differentiation and produced high levels of interferon γ (IFN-γ) and interleukin-17 in vitro, depending on Dll4 activation of Notch signaling. Following transfer, allogeneic Dll4(hi)DC-induced T cells were unable to mediate severe GVHD but preserved antileukemic activity, significantly improving the survival of leukemic mice undergoing allogeneic HSCT. This effect of Dll4(hi)DC-induced T cells was associated with their impaired expansion in GVHD target tissues. IFN-γ was important for Dll4(hi)DC programming to reduce GVHD toxicities of alloreactive T cells. Absence of T-cell IFN-γ led to improved survival and expansion of Dll4(hi)DC-induced CD4(+) T cells in transplant recipients and caused lethal GVHD. Our findings demonstrate that Dll4(hi)DC programming can overcome GVHD toxicity of donor T cells and produce leukemia-reactive T cells for effective immunotherapy. PMID:27143255

  5. Isolation and characterization of portal branch ligation-stimulated Hmga2-positive bipotent hepatic progenitor cells

    Energy Technology Data Exchange (ETDEWEB)

    Sakai, Hiroshi [Department of Surgery, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621 (Japan); Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 B51, Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8503 (Japan); Tagawa, Yoh-ichi, E-mail: ytagawa@bio.titech.ac.jp [Frontier Research Center, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8503 (Japan); Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 B51, Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8503 (Japan); PRESTO, Japan Science and Technology Agency, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012 (Japan); Tamai, Miho [Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 B51, Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8503 (Japan); Motoyama, Hiroaki [Department of Surgery, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621 (Japan); Ogawa, Shinichiro [Department of Surgery, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621 (Japan); McEwen Center for Regenerative Medicine, University Health Network, 190 Elizabeth Street, Toronto, Ont., Canada M5G 2C4 (Canada); Soeda, Junpei; Nakata, Takenari; Miyagawa, Shinichi [Department of Surgery, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621 (Japan)

    2010-12-17

    Research highlights: {yields} Hepatic progenitor cells were isolated from the portal branch-ligated liver of mice. {yields} Portal branch ligation-stimulated hepatic progenitor cells (PBLHCs) express Hmga2. {yields} PBLHCs have bidirectional differentiation capability in vitro. -- Abstract: Hepatic stem/progenitor cells are one of several cell sources that show promise for restoration of liver mass and function. Although hepatic progenitor cells (HPCs), including oval cells, are induced by administration of certain hepatotoxins in experimental animals, such a strategy would be inappropriate in a clinical setting. Here, we investigated the possibility of isolating HPCs in a portal branch-ligated liver model without administration of any chemical agents. A non-parenchymal cell fraction was prepared from the portal branch-ligated or non-ligated lobe, and seeded onto plates coated with laminin. Most of the cells died, but a small number were able to proliferate. These proliferating cells were cloned as portal branch ligation-stimulated hepatic cells (PBLHCs) by the limiting dilution method. The PBLHCs expressed cytokeratin19, albumin, and Hmga2. The PBLHCs exhibited metabolic functions such as detoxification of ammonium ions and synthesis of urea on Matrigel-coated plates in the presence of oncostatin M. In Matrigel mixed with type I collagen, the PBLHCs became rearranged into cystic and tubular structures. Immunohistochemical staining demonstrated the presence of Hmga2-positive cells around the interlobular bile ducts in the portal branch-ligated liver lobes. In conclusion, successful isolation of bipotent hepatic progenitor cell clones, PBLHCs, from the portal branch-ligated liver lobes of mice provides the possibility of future clinical application of portal vein ligation to induce hepatic progenitor cells.

  6. Depletion of γ-catenin by Histone Deacetylase Inhibition Confers Elimination of CML Stem Cells in Combination with Imatinib.

    Science.gov (United States)

    Jin, Yanli; Yao, Yiwu; Chen, Li; Zhu, Xiaohui; Jin, Bei; Shen, Yingying; Li, Juan; Du, Xin; Lu, Yuhong; Jiang, Sheng; Pan, Jingxuan

    2016-01-01

    Quiescent leukemia stem cells (LSCs) that are insensitive to BCR-ABL tyrosine kinase inhibitors confer resistance to imatinib in chronic myelogenous leukemia (CML). Identifying proteins to regulate survival and stemness of LSCs is urgently needed. Although histone deacetylase inhibitors (HDACis) can eliminate quiescent LSCs in CML, little is known about the underlying mechanism that HDACis kill LSCs. By fishing with a biotin-labeled probe, we identified that HDACi JSL-1 bound to the protein γ-catenin. γ-Catenin expression was higher in LSCs from CML patients than normal hematopoietic stem cells. Silencing γ-catenin in human CML CD34(+) bone-marrow (BM) cells sufficiently eliminated LSCs, which suggests that γ-catenin is required for survival of CML LSCs. Pharmacological inhibition of γ-catenin thwarted survival and self-renewal of human CML CD34(+) cells in vitro, and of murine LSCs in BCR-ABL-driven CML mice. γ-Catenin inhibition reduced long-term engraftment of human CML CD34(+) cells in NOD.Cg-Prkdc (scid) II2rg (tm1Sug)/JicCrl (NOG) mice. Silencing γ-catenin by shRNA in human primary CD34(+) cells did not alter β-catenin, implying a β-catenin-independent role of γ-catenin in survival and self-renewal of CML LSCs. Taken together, our findings validate that γ-catenin may be a novel therapeutic target of LSCs, and suppression of γ-catenin by HDACi may explain elimination of CML LSCs. PMID:27570562

  7. Reciprocal positive regulation between TRPV6 and NUMB in PTEN-deficient prostate cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Sung-Young; Hong, Chansik; Wie, Jinhong [Department of Physiology, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of); Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of); Kim, Euiyong [Department of Physiology, College of Medicine, Inje University, Busan 614-735 (Korea, Republic of); Kim, Byung Joo [Division of Longevity and Biofunctional Medicine, Pusan National University School of Korean Medicine, Yangsan 626-870 (Korea, Republic of); Ha, Kotdaji [Department of Physiology, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of); Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of); Cho, Nam-Hyuk; Kim, In-Gyu [Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of); Jeon, Ju-Hong [Department of Physiology, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of); Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of); So, Insuk, E-mail: insuk@snu.ac.kr [Department of Physiology, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of); Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul 110-799 (Korea, Republic of)

    2014-04-25

    Highlights: • TRPV6 interacts with tumor suppressor proteins. • Numb has a selective effect on TRPV6, depending on the prostate cancer cell line. • PTEN is a novel regulator of TRPV6–Numb complex. - Abstract: Calcium acts as a second messenger and plays a crucial role in signaling pathways involved in cell proliferation. Recently, calcium channels related to calcium influx into the cytosol of epithelial cells have attracted attention as a cancer therapy target. Of these calcium channels, TRPV6 is overexpressed in prostate cancer and is considered an important molecule in the process of metastasis. However, its exact role and mechanism is unclear. NUMB, well-known tumor suppressor gene, is a novel interacting partner of TRPV6. We show that NUMB and TRPV6 have a reciprocal positive regulatory relationship in PC-3 cells. We repeated this experiment in two other prostate cancer cell lines, DU145 and LNCaP. Interestingly, there were no significant changes in TRPV6 expression following NUMB knockdown in DU145. We revealed that the presence or absence of PTEN was the cause of NUMB–TRPV6 function. Loss of PTEN caused a positive correlation of TRPV6–NUMB expression. Collectively, we determined that PTEN is a novel interacting partner of TRPV6 and NUMB. These results demonstrated a novel relationship of NUMB–TRPV6 in prostate cancer cells, and show that PTEN is a novel regulator of this complex.

  8. Reciprocal positive regulation between TRPV6 and NUMB in PTEN-deficient prostate cancer cells

    International Nuclear Information System (INIS)

    Highlights: • TRPV6 interacts with tumor suppressor proteins. • Numb has a selective effect on TRPV6, depending on the prostate cancer cell line. • PTEN is a novel regulator of TRPV6–Numb complex. - Abstract: Calcium acts as a second messenger and plays a crucial role in signaling pathways involved in cell proliferation. Recently, calcium channels related to calcium influx into the cytosol of epithelial cells have attracted attention as a cancer therapy target. Of these calcium channels, TRPV6 is overexpressed in prostate cancer and is considered an important molecule in the process of metastasis. However, its exact role and mechanism is unclear. NUMB, well-known tumor suppressor gene, is a novel interacting partner of TRPV6. We show that NUMB and TRPV6 have a reciprocal positive regulatory relationship in PC-3 cells. We repeated this experiment in two other prostate cancer cell lines, DU145 and LNCaP. Interestingly, there were no significant changes in TRPV6 expression following NUMB knockdown in DU145. We revealed that the presence or absence of PTEN was the cause of NUMB–TRPV6 function. Loss of PTEN caused a positive correlation of TRPV6–NUMB expression. Collectively, we determined that PTEN is a novel interacting partner of TRPV6 and NUMB. These results demonstrated a novel relationship of NUMB–TRPV6 in prostate cancer cells, and show that PTEN is a novel regulator of this complex

  9. Distinct and conserved prominin-1/CD133-positive retinal cell populations identified across species.

    Directory of Open Access Journals (Sweden)

    József Jászai

    Full Text Available Besides being a marker of various somatic stem cells in mammals, prominin-1 (CD133 plays a role in maintaining the photoreceptor integrity since mutations in the PROM1 gene are linked with retinal degeneration. In spite of that, little information is available regarding its distribution in eyes of non-mammalian vertebrates endowed with high regenerative abilities. To address this subject, prominin-1 cognates were isolated from axolotl, zebrafish and chicken, and their retinal compartmentalization was investigated and compared to that of their mammalian orthologue. Interestingly, prominin-1 transcripts--except for the axolotl--were not strictly restricted to the outer nuclear layer (i.e., photoreceptor cells, but they also marked distinct subdivisions of the inner nuclear layer (INL. In zebrafish, where the prominin-1 gene is duplicated (i.e., prominin-1a and prominin-1b, a differential expression was noted for both paralogues within the INL being localized either to its vitreal or scleral subdivision, respectively. Interestingly, expression of prominin-1a within the former domain coincided with Pax-6-positive cells that are known to act as progenitors upon injury-induced retino-neurogenesis. A similar, but minute population of prominin-1-positive cells located at the vitreal side of the INL was also detected in developing and adult mice. In chicken, however, prominin-1-positive cells appeared to be aligned along the scleral side of the INL reminiscent of zebrafish prominin-1b. Taken together our data indicate that in addition to conserved expression of prominin-1 in photoreceptors, significant prominin-1-expressing non-photoreceptor retinal cell populations are present in the vertebrate eye that might represent potential sources of stem/progenitor cells for regenerative therapies.

  10. Placental Kisspeptins Differentially Modulate Vital Parameters of Estrogen Receptor-Positive and -Negative Breast Cancer Cells.

    Science.gov (United States)

    Rasoulzadeh, Zahra; Ghods, Roya; Kazemi, Tohid; Mirzadegan, Ebrahim; Ghaffari-Tabrizi-Wizsy, Nassim; Rezania, Simin; Kazemnejad, Somaieh; Arefi, Soheila; Ghasemi, Jamileh; Vafaei, Sedigheh; Mahmoudi, Ahmad-Reza; Zarnani, Amir-Hassan

    2016-01-01

    Kisspeptins (KPs) are major regulators of trophoblast and cancer invasion. Thus far, limited and conflicting data are available on KP-mediated modulation of breast cancer (BC) metastasis; mostly based on synthetic KP-10, the most active fragment of KP. Here, we report for the first time comprehensive functional effects of term placental KPs on proliferation, adhesion, Matrigel invasion, motility, MMP activity and pro-inflammatory cytokine production in MDA-MB-231 (estrogen receptor-negative) and MCF-7 (estrogen receptor-positive). KPs were expressed at high level by term placental syncytiotrophoblasts and released in soluble form. Placental explant conditioned medium containing KPs (CM) significantly reduced proliferation of both cell types compared to CM without (w/o) KP (CM-w/o KP) in a dose- and time-dependent manner. In MDA-MB-231 cells, placental KPs significantly reduced adhesive properties, while increased MMP9 and MMP2 activity and stimulated invasion. Increased invasiveness of MDA-MB-231 cells after CM treatment was inhibited by KP receptor antagonist, P-234. CM significantly reduced motility of MCF-7 cells at all time points (2-30 hr), while it stimulated motility of MDA-MB-231 cells. These effects were reversed by P-234. Co-treatment with selective ER modulators, Tamoxifen and Raloxifene, inhibited the effect of CM on motility of MCF-7 cells. The level of IL-6 in supernatant of MCF-7 cells treated with CM was higher compared to those treated with CM-w/o KP. Both cell types produced more IL-8 after treatment with CM compared to those treated with CM-w/o KP. Taken together, our observations suggest that placental KPs differentially modulate vital parameters of estrogen receptor-positive and -negative BC cells possibly through modulation of pro-inflammatory cytokine production. PMID:27101408

  11. Quantum dot labeling using positive charged peptides in human hematopoetic and mesenchymal stem cells.

    Science.gov (United States)

    Ranjbarvaziri, Sarah; Kiani, Sahar; Akhlaghi, Aliasghar; Vosough, Ahmad; Baharvand, Hossein; Aghdami, Nasser

    2011-08-01

    Quantum dots (QDs), as new and promising fluorescent probes, hold great potential in long term non-invasive bio-imaging, however there are many uncovered issues regarding their competency. In the present study, different QDs (525, 585 and 800 nm) were used to label CD133, CD34, CD14 and mesenchymal stem cells (MSCs) using positively charged peptides. Results demonstrated highly efficient internalization with the possible involvement of macropinocytosis. As indicated by LDH release and the TUNEL assay, no measurable effects on cell viability were detected at a concentration of 10 nM. QDs did not have any deleterious effects on normal cell functionality where both labeled CD133(+) cells and MSCs remarkably differentiated along multiple lineages with the use of the colony forming assay and adipo/osteo induction, respectively. Our results regarding QD maintenance revealed that these nano-particles are not properly stable and various excretion times have been observed depending on particle size and cell type. In vitro co-culture system and transplantation of labeled cells to an animal model showed that QDs leaked out from labeled cells and the released nano-particles were able to re-enter adjacent cells over time. These data suggest that before any utilization of QDs in bio-imaging and related applications, an efficient intra-cellular delivery technique should be considered to preserve QDs for a prolonged time as well as eliminating their leakage. PMID:21549422

  12. Controlled Positioning of Cells in Biomaterials—Approaches Towards 3D Tissue Printing

    Directory of Open Access Journals (Sweden)

    Sandra Hofmann

    2011-08-01

    Full Text Available Current tissue engineering techniques have various drawbacks: they often incorporate uncontrolled and imprecise scaffold geometries, whereas the current conventional cell seeding techniques result mostly in random cell placement rather than uniform cell distribution. For the successful reconstruction of deficient tissue, new material engineering approaches have to be considered to overcome current limitations. An emerging method to produce complex biological products including cells or extracellular matrices in a controlled manner is a process called bioprinting or biofabrication, which effectively uses principles of rapid prototyping combined with cell-loaded biomaterials, typically hydrogels. 3D tissue printing is an approach to manufacture functional tissue layer-by-layer that could be transplanted in vivo after production. This method is especially advantageous for stem cells since a controlled environment can be created to influence cell growth and differentiation. Using printed tissue for biotechnological and pharmacological needs like in vitro drug-testing may lead to a revolution in the pharmaceutical industry since animal models could be partially replaced by biofabricated tissues mimicking human physiology and pathology. This would not only be a major advancement concerning rising ethical issues but would also have a measureable impact on economical aspects in this industry of today, where animal studies are very labor-intensive and therefore costly. In this review, current controlled material and cell positioning techniques are introduced highlighting approaches towards 3D tissue printing.

  13. Positive association of long telomeres with the invasive capacity of hepatocellular carcinoma cells.

    Science.gov (United States)

    Ko, Eunkyong; Jung, Guhung

    2014-05-01

    Invasion, the representative feature of malignant tumors, leads to an increase in mortality. The malignant liver tumor - hepatocellular carcinoma (HCC) - has an enhanced invasive capacity that results in increased patient mortality. Moreover, this enhanced invasive capacity is due to the up-regulation of invasion promoters such as zinc finger protein SNAI1 (Snail) and matrix metalloproteinases (MMPs), and the down-regulation of invasion suppressor molecules such as E-cadherin. Telomerase reverse transcriptase (TERT), which encodes the catalytic subunit of telomerase, is highly expressed in a variety of invasive cancers, including HCC. Telomerase activation induces telomere elongation, thereby leading to cell immortalization during malignant tumor progression. However, the relationship between telomere length and invasion is yet to be experimentally corroborated. In this paper, we revealed that invasive HCC cells passing through the Matrigel display significantly longer telomeres than non-invasive HCC cells. Moreover, we established a method that can distinguish and sort cells containing long telomeres and short telomeres. Using this system, we observed that the HCC cells containing long telomeres had a high-level expression of invasion-promoting genes and a low-level expression of invasion-suppressing E-cadherin. Furthermore, HCC cells containing long telomeres exhibited a higher invasive capacity than HCC cells containing short telomeres. Taken together, our findings suggest that long telomeres are positively associated with the invasive capacity of HCC cells and may be a potent target for malignant liver cancer treatment. PMID:24732358

  14. Indium-111-oxine labeled leukocyte uptake in Ki-1-positive anaplastic large cell lymphoma

    Energy Technology Data Exchange (ETDEWEB)

    Chiu, W.; Amodio, J.B.; Scharf, S.C. (New York Univ., NY (United States). Dept. of Radiology); Rivlin, K.A. (Department of Hematology and Oncology, New York Univ. Medical Center, NY (United States)); Desai, P. (Department of Pathology, Hospital for Joint Diseases-Orthopaedic Inst., New York, NY (United States)); Breuer, F. (Department of Pathology, Lenox Hill Hospital, New York, NY (United States))

    1999-08-01

    Indium-111-oxine labeled leukocyte ([sup 111]In-WBC) scintigraphy is well known for its ability to localize in areas of active infection, but not in areas of lymphomatous involvement. We present a case of Ki-1-positive anaplastic large-cell lymphoma that was initially thought to be a case of multifocal osteomyelitis because of positive uptake on a [sup 111]In-WBC scan. The areas of abnormal uptake on the indium scan were demonstrated histopathologically to be sites of lymphomatous involvement in bone. (orig.) With 4 figs., 3 refs.

  15. Positively charged supported lipid bilayer formation on gold surfaces for neuronal cell culture.

    Science.gov (United States)

    Choi, Sung-Eun; Greben, Kyrylo; Wördenweber, Roger; Offenhäusser, Andreas

    2016-06-01

    Supported lipid bilayers are widely used as cell membrane models and sensor platforms, but the usage on gold surface needs additional surface modification or optimized experimental conditions. In this work, the authors show lipid bilayer formation on plasma activated gold surfaces in physiological conditions without any other modification if at least 30% positively charged lipids are present. Details of bilayer formation from small unilamellar vesicles were monitored using quartz crystal microbalance with dissipation in both basic and acidic environment. The authors also confirmed that this positively charged bilayer system can sustain primary cortical neuron growth and lipid transfer. This method will provide simple means to construct biomimetic interface on gold electrodes. PMID:27052005

  16. Indium-111-oxine labeled leukocyte uptake in Ki-1-positive anaplastic large cell lymphoma

    International Nuclear Information System (INIS)

    Indium-111-oxine labeled leukocyte (111In-WBC) scintigraphy is well known for its ability to localize in areas of active infection, but not in areas of lymphomatous involvement. We present a case of Ki-1-positive anaplastic large-cell lymphoma that was initially thought to be a case of multifocal osteomyelitis because of positive uptake on a 111In-WBC scan. The areas of abnormal uptake on the indium scan were demonstrated histopathologically to be sites of lymphomatous involvement in bone. (orig.)

  17. An epidermis-driven mechanism positions and scales stem cell niches in plants

    Science.gov (United States)

    Gruel, Jérémy; Landrein, Benoit; Tarr, Paul; Schuster, Christoph; Refahi, Yassin; Sampathkumar, Arun; Hamant, Olivier; Meyerowitz, Elliot M.; Jönsson, Henrik

    2016-01-01

    How molecular patterning scales to organ size is highly debated in developmental biology. We explore this question for the characteristic gene expression domains of the plant stem cell niche residing in the shoot apical meristem. We show that a combination of signals originating from the epidermal cell layer can correctly pattern the key gene expression domains and notably leads to adaptive scaling of these domains to the size of the tissue. Using live imaging, we experimentally confirm this prediction. The identified mechanism is also sufficient to explain de novo stem cell niches in emerging flowers. Our findings suggest that the deformation of the tissue transposes meristem geometry into an instructive scaling and positional input for the apical plant stem cell niche.

  18. An epidermis-driven mechanism positions and scales stem cell niches in plants.

    Science.gov (United States)

    Gruel, Jérémy; Landrein, Benoit; Tarr, Paul; Schuster, Christoph; Refahi, Yassin; Sampathkumar, Arun; Hamant, Olivier; Meyerowitz, Elliot M; Jönsson, Henrik

    2016-01-01

    How molecular patterning scales to organ size is highly debated in developmental biology. We explore this question for the characteristic gene expression domains of the plant stem cell niche residing in the shoot apical meristem. We show that a combination of signals originating from the epidermal cell layer can correctly pattern the key gene expression domains and notably leads to adaptive scaling of these domains to the size of the tissue. Using live imaging, we experimentally confirm this prediction. The identified mechanism is also sufficient to explain de novo stem cell niches in emerging flowers. Our findings suggest that the deformation of the tissue transposes meristem geometry into an instructive scaling and positional input for the apical plant stem cell niche. PMID:27152324

  19. The distribution and number of Leu-7 (CD57) positive cells in lung tissue from patients with pulmonary fibrosis.

    OpenAIRE

    Yamanouchi H; Ohtsuki Y; Fujita J; Bandoh S; Yoshinouchi T; Ishida T

    2002-01-01

    Leu-7 positive lymphocytes, including natural killer cells, play an important role in the immune system's surveillance function to prevent the development of cancer. The incidence of lung cancer is significantly high in patients with end-stage pulmonary fibrosis. We hypothesized that the number of Leu-7 positive cells may be decreased in areas of severe pulmonary fibrosis. To demonstrate this, Leu-7 positive cells were immunohistochemically stained in 41 lung specimens obtained from patients ...

  20. Impact of CD133 positive stem cell proportion on survival in patients with glioblastoma multiforme

    International Nuclear Information System (INIS)

    The aim of the study was to assess the impact of CD133-positive (CD133+) cancer stem cell proportions on treatment results of glioblastoma multiforme (GBM) patients. Patients with GBM (n = 42) received postoperative radiotherapy (± chemotherapy). Surgically excised GBM tissue sections were immunohistochemically examined for CD133 expression. The proportions of CD133+ GBM cells were determined (%). The proportion of CD133+ GBM stem cells was established by 2 independent researchers whose results were in good accordance (R = 0.8, p < 0.01). Additionally, CD133 expression levels were correlated with patients overall survival. The proportion of CD133+ cells varied between patients, being from 0.5% to 82%. Mean and median proportions of CD133+ cells of the entire study group were 33% ± 24% (mean ± SD) and 28%, respectively. Clinical data do not support the association between higher proportion of stem cells and the aggressiveness of GBM. Median survival time of the study group was 10.0 months (95% CI 9.0–11.0). The survival time clearly depended on the proportion of CD133+ cells (log rank test, p = 0.02). Median survival times for patients with low (< median) and high (≥ median) proportion of CD133+ cells were 9.0 months (95% CI 7.6–10.5) and 12.0 months (95% CI 9.3–14.7), respectively. In multivariate analysis, the proportion of CD133+ cells emerged as a significant independent predictor for longer overall survival (HR 2.0, 95% CI 1.0–3.8, p = 0.04). In patients with higher stem cell proportion, significantly longer survival times after postoperative radiotherapy were achieved. Underlying reasons and possible higher sensitivity of GBM stem cells to fractionated radio-therapy should be clarified in further studies

  1. High myeloperoxidase positive cell infiltration in colorectal cancer is an independent favorable prognostic factor.

    Directory of Open Access Journals (Sweden)

    Raoul A Droeser

    Full Text Available BACKGROUND: Colorectal cancer (CRC infiltration by adaptive immune system cells correlates with favorable prognosis. The role of the innate immune system is still debated. Here we addressed the prognostic impact of CRC infiltration by neutrophil granulocytes (NG. METHODS: A TMA including healthy mucosa and clinically annotated CRC specimens (n = 1491 was stained with MPO and CD15 specific antibodies. MPO+ and CD15+ positive immune cells were counted by three independent observers. Phenotypic profiles of CRC infiltrating MPO+ and CD15+ cells were validated by flow cytometry on cell suspensions derived from enzymatically digested surgical specimens. Survival analysis was performed by splitting randomized data in training and validation subsets. RESULTS: MPO+ and CD15+ cell infiltration were significantly correlated (p<0.0001; r = 0.76. However, only high density of MPO+ cell infiltration was associated with significantly improved survival in training (P = 0.038 and validation (P = 0.002 sets. In multivariate analysis including T and N stage, vascular invasion, tumor border configuration and microsatellite instability status, MPO+ cell infiltration proved an independent prognostic marker overall (P = 0.004; HR = 0.65; CI:±0.15 and in both training (P = 0.048 and validation (P = 0.036 sets. Flow-cytometry analysis of CRC cell suspensions derived from clinical specimens showed that while MPO+ cells were largely CD15+/CD66b+, sizeable percentages of CD15+ and CD66b+ cells were MPO-. CONCLUSIONS: High density MPO+ cell infiltration is a novel independent favorable prognostic factor in CRC.

  2. Cytotoxicity and apoptosis induced by a plumbagin derivative in estrogen positive MCF-7 breast cancer cells

    KAUST Repository

    Sagar, Sunil

    2014-01-31

    Plumbagin [5-hydroxy- 2-methyl-1, 4-naphthaquinone] is a well-known plant derived anticancer lead compound. Several efforts have been made to synthesize its analogs and derivatives in order to increase its anticancer potential. In the present study, plumbagin and its five derivatives have been evaluated for their antiproliferative potential in one normal and four human cancer cell lines. Treatment with derivatives resulted in dose- and time-dependent inhibition of growth of various cancer cell lines. Prescreening of compounds led us to focus our further investigations on acetyl plumbagin, which showed remarkably low toxicity towards normal BJ cells and HepG2 cells. The mechanisms of apoptosis induction were determined by APOPercentage staining, caspase-3/7 activation, reactive oxygen species production and cell cycle analysis. The modulation of apoptotic genes (p53, Mdm2, NF-kB, Bad, Bax, Bcl-2 and Casp-7) was also measured using real time PCR. The positive staining using APOPercentage dye, increased caspase-3/7 activity, increased ROS production and enhanced mRNA expression of proapoptotic genes suggested that acetyl plumbagin exhibits anticancer effects on MCF-7 cells through its apoptosis-inducing property. A key highlighting point of the study is low toxicity of acetyl plumbagin towards normal BJ cells and negligible hepatotoxicity (data based on HepG2 cell line). Overall results showed that acetyl plumbagin with reduced toxicity might have the potential to be a new lead molecule for testing against estrogen positive breast cancer. 2014 Bentham Science Publishers.

  3. Spatial distribution of prominin-1 (CD133-positive cells within germinative zones of the vertebrate brain.

    Directory of Open Access Journals (Sweden)

    József Jászai

    Full Text Available BACKGROUND: In mammals, embryonic neural progenitors as well as adult neural stem cells can be prospectively isolated based on the cell surface expression of prominin-1 (CD133, a plasma membrane glycoprotein. In contrast, characterization of neural progenitors in non-mammalian vertebrates endowed with significant constitutive neurogenesis and inherent self-repair ability is hampered by the lack of suitable cell surface markers. Here, we have investigated whether prominin-1-orthologues of the major non-mammalian vertebrate model organisms show any degree of conservation as for their association with neurogenic geminative zones within the central nervous system (CNS as they do in mammals or associated with activated neural progenitors during provoked neurogenesis in the regenerating CNS. METHODS: We have recently identified prominin-1 orthologues from zebrafish, axolotl and chicken. The spatial distribution of prominin-1-positive cells--in comparison to those of mice--was mapped in the intact brain in these organisms by non-radioactive in situ hybridization combined with detection of proliferating neural progenitors, marked either by proliferating cell nuclear antigen or 5-bromo-deoxyuridine. Furthermore, distribution of prominin-1 transcripts was investigated in the regenerating spinal cord of injured axolotl. RESULTS: Remarkably, a conserved association of prominin-1 with germinative zones of the CNS was uncovered as manifested in a significant co-localization with cell proliferation markers during normal constitutive neurogenesis in all species investigated. Moreover, an enhanced expression of prominin-1 became evident associated with provoked, compensatory neurogenesis during the epimorphic regeneration of the axolotl spinal cord. Interestingly, significant prominin-1-expressing cell populations were also detected at distinct extraventricular (parenchymal locations in the CNS of all vertebrate species being suggestive of further, non

  4. Leukemia Mediated Endothelial Cell Activation Modulates Leukemia Cell Susceptibility to Chemotherapy through a Positive Feedback Loop Mechanism.

    Directory of Open Access Journals (Sweden)

    Bahareh Pezeshkian

    Full Text Available In acute myeloid leukemia (AML, the chances of achieving disease-free survival are low. Studies have demonstrated a supportive role of endothelial cells (ECs in normal hematopoiesis. Here we show that similar intercellular relationships exist in leukemia. We demonstrate that leukemia cells themselves initiate these interactions by directly modulating the behavior of resting ECs through the induction of EC activation. In this inflammatory state, activated ECs induce the adhesion of a sub-set of leukemia cells through the cell adhesion molecule E-selectin. These adherent leukemia cells are sequestered in a quiescent state and are unaffected by chemotherapy. The ability of adherent cells to later detach and again become proliferative following exposure to chemotherapy suggests a role of this process in relapse. Interestingly, differing leukemia subtypes modulate this process to varying degrees, which may explain the varied response of AML patients to chemotherapy and relapse rates. Finally, because leukemia cells themselves induce EC activation, we postulate a positive-feedback loop in leukemia that exists to support the growth and relapse of the disease. Together, the data defines a new mechanism describing how ECs and leukemia cells interact during leukemogenesis, which could be used to develop novel treatments for those with AML.

  5. New Insights on Human Nestin-positive Bone Marrow-derived Multipotent Cells

    Directory of Open Access Journals (Sweden)

    SimonePacini

    2014-05-01

    Full Text Available Recent investigations have made considerable progress in the understanding of tissue regeneration driven by mesenchymal stromal cells (MSCs. Data indicate the anatomical location of MSC as residing in the “perivascular” space of blood vessels dispersed across the whole body. This histological localization suggests that MSCs contribute to the formation of new blood vessels in vivo. Indeed, MSCs can release angiogenic factors and protease to facilitate blood vessel formation and in vitro are able to promote/support angiogenesis. However, the direct differentiation of MCSs into endothelial cells is still matter of debate. Most of the conflicting data might arise from the presence of multiple subtypes of cells with heterogeneous morph-functional features within the MSC cultures. According to this scenario, we hypothesize that the presence of the recently described Mesodermal Progenitor Cells (MPCs within the MSCs cultures is responsible for their variable angiogenic potential. Indeed, MPCs are Nestin-positive CD31-positive cells exhibiting angiogenic potential that differentiate in MSC upon proper stimuli. The ISCT criteria do not account for the presence of MPC within MSC culture generating confusion in the interpretation of MSC angiogenic potential. In conclusion, the discovery of MPC gives new insight in defining MSC ancestors in human bone marrow, and indicates the tunica intima as a further, and previously overlooked, possible additional source of MSC.

  6. Activated WNT signaling in postnatal SOX2-positive dental stem cells can drive odontoma formation.

    Science.gov (United States)

    Xavier, Guilherme M; Patist, Amanda L; Healy, Chris; Pagrut, Ankita; Carreno, Gabriela; Sharpe, Paul T; Martinez-Barbera, Juan Pedro; Thavaraj, Selvam; Cobourne, Martyn T; Andoniadou, Cynthia L

    2015-01-01

    In common with most mammals, humans form only two dentitions during their lifetime. Occasionally, supernumerary teeth develop in addition to the normal complement. Odontoma represent a small group of malformations containing calcified dental tissues of both epithelial and mesenchymal origin, with varying levels of organization, including tooth-like structures. The specific cell type responsible for the induction of odontoma, which retains the capacity to re-initiate de novo tooth development in postnatal tissues, is not known. Here we demonstrate that aberrant activation of WNT signaling by expression of a non-degradable form of β-catenin specifically in SOX2-positive postnatal dental epithelial stem cells is sufficient to generate odontoma containing multiple tooth-like structures complete with all dental tissue layers. Genetic lineage-tracing confirms that odontoma form in a similar manner to normal teeth, derived from both the mutation-sustaining epithelial stem cells and adjacent mesenchymal tissues. Activation of the WNT pathway in embryonic SOX2-positive progenitors results in ectopic expression of secreted signals that promote odontogenesis throughout the oral cavity. Significantly, the inductive potential of epithelial dental stem cells is retained in postnatal tissues, and up-regulation of WNT signaling specifically in these cells is sufficient to promote generation and growth of ectopic malformations faithfully resembling human odontoma. PMID:26411543

  7. Cytokeratin-positive cells in preoperative peripheral blood and bone marrow aspirates of patients with colorectal cancer

    DEFF Research Database (Denmark)

    Werther, K; Normark, M; Brünner, N;

    2002-01-01

    Detection of cytokeratin-positive cells in bone marrow and peripheral blood may have prognostic significance in cancer patients. Furthermore, a correlation between uPAR expression on micrometastases and patient prognosis has been suggested. However, in patients with colorectal cancer, preoperative...... cancer, were immunocytochemically screened for cytokeratin-positive cells. Where cytokeratin-positive cells were observed, an additional microslide was double immunostained for simultaneous detection of cytokeratin and uPAR/CD87. RESULTS: Cytokeratin-positive cells were observed in 4 out of 41 bone...... preoperatively obtained bone marrow aspirates or peripheral blood from patients with colorectal cancer....

  8. Autoimmune pancreatitis metachronously associated with retroperitoneal fibrosis with IgG4-positive plasma cell infiltration

    Institute of Scientific and Technical Information of China (English)

    Terumi Kamisawa; Pong Yui Chen; Yuyang Tu; Hitoshi Nakajima; Naoto Egawa

    2006-01-01

    Retroperitoneal fibrosis is an uncommon disorder characterized by the formation of a dense plaque of fibrous tissue in the retroperitoneum, and its etiology remains unknown. Autoimmune pancreatitis is a rare type of chronic pancreatitis characterized by fibrosis with abundant infiltration of IgG4-positive plasma cells and lymphocytes and obliterative phlebitis in the pancreas. We present a case of autoimmune pancreatitis that developed 10 mo after the occurrence of retroperitoneal fibrosis. Histological findings of the resected retroperitoneal mass were marked periureteral fibrosis with abundant infiltration of IgG4-positive plasma cells and lymphocytes and obliterative phlebitis.These findings suggest a common pathophysiological mechanism for retroperitoneal fibrosis and autoimmune pancreatitis in this case. Some cases of retroperitoneal fibrosis might be a retroperitoneal lesion of IgG4-related sclerosing disease.

  9. MMP7 expression regulated by endocrine therapy in ERβ-positive colon cancer cells

    Directory of Open Access Journals (Sweden)

    Zhang Li-Yi

    2009-09-01

    Full Text Available Abstract Background Many studies have shown that colon cancer is an estrogen-dependent carcinoma. This study explored the efficacy of endocrine therapy in colon cancer cells with high metastatic potential (HT29. We investigated the proliferation of HT29 cells after exposure to endocrine therapy (tamoxifen and 5-FU. Methods Apoptosis was evaluated using flow cytometry. The expression of matrix metalloproteinases 7 (MMP-7 and estrogen receptor beta (ERβ was measured by reverse transcription-polymerase chain reaction (RT-PCR and western blot. The migration capability of treated cells was determined with wound scratch assay. Results Tamoxifen alone, 5-FU alone, and the combination of the two drugs can significantly inhibit HT29 cell proliferation and migration, block the cells in G2/M phase and induce cell apoptosis. These drugs also can down-regulate MMP7 and ERβ expression. Conclusion Our findings suggest that endocrine therapy is an efficient therapy for inhibiting ERβ-positive colon cancer cell proliferation and migration via down-regulation of MMP7.

  10. Are MSCs angiogenic cells? New insights on human nestin-positive bone marrow-derived multipotent cells.

    Science.gov (United States)

    Pacini, Simone; Petrini, Iacopo

    2014-01-01

    Recent investigations have made considerable progress in the understanding of tissue regeneration driven by mesenchymal stromal cells (MSCs). Data indicate the anatomical location of MSC as residing in the "perivascular" space of blood vessels dispersed across the whole body. This histological localization suggests that MSCs contribute to the formation of new blood vessels in vivo. Indeed, MSCs can release angiogenic factors and protease to facilitate blood vessel formation and in vitro are able to promote/support angiogenesis. However, the direct differentiation of MCSs into endothelial cells is still matter of debate. Most of the conflicting data might arise from the presence of multiple subtypes of cells with heterogeneous morpho functional features within the MSC cultures. According to this scenario, we hypothesize that the presence of the recently described Mesodermal Progenitor Cells (MPCs) within the MSCs cultures is responsible for their variable angiogenic potential. Indeed, MPCs are Nestin-positive CD31-positive cells exhibiting angiogenic potential that differentiate in MSC upon proper stimuli. The ISCT criteria do not account for the presence of MPC within MSC culture generating confusion in the interpretation of MSC angiogenic potential. In conclusion, the discovery of MPC gives new insight in defining MSC ancestors in human bone marrow, and indicates the tunica intima as a further, and previously overlooked, possible additional source of MSC. PMID:25364727

  11. Relationship between squamous cell carcinoma of the tongue and the position of dental prosthesis

    OpenAIRE

    Fan, Huan; Yoon, Ki-Yong; Kim, Soung-Min; Myoung, Hoon; Lee, Jong-Ho; Kim, Myung-Jin

    2015-01-01

    PURPOSE Squamous cell carcinoma (SCC) of the tongue has a relatively high incidence of all oral cancers. Some studies have reported a relationship between intraoral dental prosthesis and SCC of the tongue; however, this relationship remains controversial. The purpose of this study was to investigate the relationship between SCC of the tongue and the positional aspects of dental prosthesis using a retrospective analysis. MATERIALS AND METHODS A total of 439 patients with SCC of the tongue were...

  12. Positive-unlabeled learning for the prediction of conformational B-cell epitopes

    OpenAIRE

    Ren, Jing; Liu, Qian; Ellis, John; Li, Jinyan

    2015-01-01

    Background The incomplete ground truth of training data of B-cell epitopes is a demanding issue in computational epitope prediction. The challenge is that only a small fraction of the surface residues of an antigen are confirmed as antigenic residues (positive training data); the remaining residues are unlabeled. As some of these uncertain residues can possibly be grouped to form novel but currently unknown epitopes, it is misguided to unanimously classify all the unlabeled residues as negati...

  13. Clinical utility of Circulating Tumor Cells in ALK-Positive Non-Small-Cell Lung Cancer

    OpenAIRE

    Vincent eFaugeroux; Emma ePailler; Nathalie eAuger; Meilissa eTaylor; Françoise eFarace

    2014-01-01

    The advent of rationally targeted therapies such as small-molecule tyrosine kinase inhibitors (TKIs) has considerably transformed the therapeutic management of a subset of patients with non-small cell lung cancer (NSCLC) harboring defined molecular abnormalities. When such genetic molecular alterations are detected the use of specific TKI has demonstrated better results (overall response rate (ORR), progression free survival (PFS)) compared to systemic therapy. However the detection of such m...

  14. Clinical Utility of Circulating Tumor Cells in ALK-Positive Non-Small-Cell Lung Cancer

    OpenAIRE

    Faugeroux, Vincent; Pailler, Emma; Auger, Nathalie; Taylor, Melissa; Farace, Françoise

    2014-01-01

    The advent of rationally targeted therapies such as small-molecule tyrosine kinase inhibitors (TKIs) has considerably transformed the therapeutic management of a subset of patients with non-small-cell lung cancer (NSCLC) harboring defined molecular abnormalities. When such genetic molecular alterations are detected the use of specific TKI has demonstrated better results (overall response rate, progression free survival) compared to systemic therapy. However, the detection of such molecular ab...

  15. Rare Association of Anti-Hu Antibody Positive Paraneoplastic Neurological Syndrome and Transitional Cell Bladder Carcinoma

    Directory of Open Access Journals (Sweden)

    S. Lukacs

    2012-01-01

    Full Text Available Introduction. Paraneoplastic encephalomyelitis (PEM and subacute sensory neuronopathy (SSN are remote effects of cancer, usually associated with small-cell lung carcinoma and positive anti-Hu antibody. We describe the rare association of bladder transitional cell carcinoma (TCC with anti-Hu antibody positivity resulting in this paraneoplastic neurological syndrome. Patient. A 76-year-old female presented with bilateral muscle weakness and paraesthesia of the upper and lower limbs in a length-dependent “glove and stocking” distribution. Central nervous system symptoms included cognitive problems, personality change, and truncal ataxia. Case notes and the literature were reviewed. Result. Autoantibody screening was positive for anti-Hu antibody (recently renamed antineuronal nuclear antibody 1, ANNA-1. The diagnosis of PEM and SSN was supported by MRI and lumbar puncture results. A superficial bladder TCC was demonstrated on CT and subsequently confirmed on histology. No other primary neoplasm was found on full-body imaging. The neurological symptoms were considered to be an antibody-mediated paraneoplastic neurological syndrome and improved after resection of the tumour. Discussion. The association of anti-Hu positive paraneoplastic neurological syndrome and TCC has not been described in the literature previously. We emphasize the need for detailed clinical examination and the importance of a multidisciplinary thought process and encourage further awareness of this rare association.

  16. High False-Positive Rate of Cytokeratin-19 in Detecting Circulating Tumor Cells for Nasopharyngeal Carcinoma

    Directory of Open Access Journals (Sweden)

    Ruey-Ho Kao

    2002-04-01

    Full Text Available Background: Nasopharyngeal carcinoma (NPC harbors a higher metastatic potential thanother head and neck cancers. In order to seek a possible surrogate marker forearly detection of recurrent or metastatic disease, we tested the feasibility ofcytokeratin-19 (CK-19-nested reverse-transcription polymerase chain reaction(RT-PCR for detecting circulating tumor cells in NPC patients.Methods: Two tubes of blood were sequentially collected in individual draws from 7NPC patients and 15 healthy persons. Total ribonucleic acid (RNA wasextracted from blood cells and treated with deoxyribonuclease. The RNAwas then subjected to RT and nested PCR with specific CK-19 primers. Thereaction products were run on an agarose gel and visualized under UV light.The sequences of the products were determined using an ABI377 automaticsequencer.Results: Among the 7 NPC cases, 4 cases presented CK-19 expression with 2 in bothtubes, 1 in the first tube, and 1 in the second tube. In the control group, 8 of15 cases also presented CK-19 expression with 6 in both tubes and 2 in thesecond tube resulting in a 53.3% false-positive rate. Incidentally, an aberrantsplicing product lacking exon 4 of CK-19 messenger RNA was discovered.Conclusion: Results of the present study indicate that the CK-19-nested RT-PCR is notsuitable for detecting circulating tumor cells in NPC patients because of ahigh false-positive rate in the control group. The reason for the high rate offalse-positives may be attributed to pseudogenes, different blood cell separationmethods, or illegitimate expression of CK-19 in blood cells.

  17. Galpha/LGN-mediated asymmetric spindle positioning does not lead to unequal cleavage of the mother cell in 3-D cultured MDCK cells

    OpenAIRE

    Xiao, Zhuoni; Wan, Qingwen; Du, Quansheng; Zheng, Zhen

    2012-01-01

    The position of the mitotic spindle plays a key role in spatial control of cell division. It is generally believed that when a spindle is positioned asymmetrically in a dividing cell, the resulting daughter cells are usually unequal in size due to eccentric cleavage of the mother cell. Molecular mechanisms underlying the generation of unequal sized daughter cells have been extensively studied in Drosophila neuroblast and C elegans zygote where the Gα subunit of the heterotrimeric G proteins a...

  18. Effects of soft X-ray irradiation on NK cell activity and the percentage of asialo GM1-positive cells in spleen cells of mice

    International Nuclear Information System (INIS)

    Effects of soft X-ray irradiation on the natural killer (NK) cell activity and the percentage of asialo GM1-positive cells of spleen cells in mice were investigated by using a soft X-ray generator intended for non-destructive radiological examination. Soft X-ray irradiation in graded doses of more than 25.8 mC/kg indicated dose dependent reductions in the NK cell activity in the spleen of irradiated mice. Significant reductions in the population of asialo GM1-positive cells in spleen cells were also observed. These results suggest that a soft X-ray generator could also be useful in immuno-irradiation studies. (author)

  19. Functional screening identifies CRLF2 in precursor B-cell acute lymphoblastic leukemia.

    Science.gov (United States)

    Yoda, Akinori; Yoda, Yuka; Chiaretti, Sabina; Bar-Natan, Michal; Mani, Kartik; Rodig, Scott J; West, Nathan; Xiao, Yun; Brown, Jennifer R; Mitsiades, Constantine; Sattler, Martin; Kutok, Jeffrey L; DeAngelo, Daniel J; Wadleigh, Martha; Piciocchi, Alfonso; Dal Cin, Paola; Bradner, James E; Griffin, James D; Anderson, Kenneth C; Stone, Richard M; Ritz, Jerome; Foà, Robin; Aster, Jon C; Frank, David A; Weinstock, David M

    2010-01-01

    The prognosis for adults with precursor B-cell acute lymphoblastic leukemia (B-ALL) remains poor, in part from a lack of therapeutic targets. We identified the type I cytokine receptor subunit CRLF2 in a functional screen for B-ALL-derived mRNA transcripts that can substitute for IL3 signaling. We demonstrate that CRLF2 is overexpressed in approximately 15% of adult and high-risk pediatric B-ALL that lack MLL, TCF3, TEL, and BCR/ABL rearrangements, but not in B-ALL with these rearrangements or other lymphoid malignancies. CRLF2 overexpression can result from translocation with the IGH locus or intrachromosomal deletion and is associated with poor outcome. CRLF2 overexpressing B-ALLs share a transcriptional signature that significantly overlaps with a BCR/ABL signature, and is enriched for genes involved in cytokine receptor and JAK-STAT signaling. In a subset of cases, CRLF2 harbors a Phe232Cys gain-of-function mutation that promotes constitutive dimerization and cytokine independent growth. A mutually exclusive subset harbors activating mutations in JAK2. In fact, all 22 B-ALLs with mutant JAK2 that we analyzed overexpress CRLF2, distinguishing CRLF2 as the key scaffold for mutant JAK2 signaling in B-ALL. Expression of WT CRLF2 with mutant JAK2 also promotes cytokine independent growth that, unlike CRLF2 Phe232Cys or ligand-induced signaling by WT CRLF2, is accompanied by JAK2 phosphorylation. Finally, cells dependent on CRLF2 signaling are sensitive to small molecule inhibitors of either JAKs or protein kinase C family kinases. Together, these findings implicate CRLF2 as an important factor in B-ALL with diagnostic, prognostic, and therapeutic implications. PMID:20018760

  20. STEREOLOGIC ESTIMATION OF KI-67, CASPASE 3 AND GSTP1 POSITIVE CELLS IN PROSTATE LESIONS

    Directory of Open Access Journals (Sweden)

    Luis Santamaría

    2011-05-01

    Full Text Available Cell proliferation, caspase 3 and pi-form of glutathione S transferase (GSTP1 were evaluated in prostate carcinoma (PCA, proliferative inflammatory atrophy (PIA and prostate intraepithelial neoplasia (PIN. Forty biopsies were classified as: without morphological lesions (controls: CTR, PIA, PIN and PCA. Ki67, caspase3 and GSTP1 were immunostained. The following estimates were performed: Numerical densities of Ki67+ cells (NVEPKi67, of all epithelial cells (NVEPtotal and of GSTP1+ cells (NVEPGSTP1; labelling index for Ki67 (LIKi67; volume fraction to caspase 3 positive tissue (VVcaspase 3 and of GSTP1 positive tissue (VVGSTP1. ANOVA was performed to compare the groups. NVEPtotal and NVEPKi67 were increased in PIA. LIKi67 was only increased in PCA. VVcaspase 3 was decreased in PIN and PCA. VVEGSTP1 was decreased in PCA. In our results PIA lacks the characteristics of a premalignant lesion. The result may be explained by the use of unbiased quantitative methods, the inadequate definition of PIA and the scarce inflammation observed in the samples with PIA included in this study.

  1. Quality Indicator Development for Positive Screen Follow-up for Sickle Cell Disease and Trait.

    Science.gov (United States)

    Faro, Elissa Z; Wang, C Jason; Oyeku, Suzette O

    2016-07-01

    Extensive variation exists in the follow-up of positive screens for sickle cell disease. Limited quality indicators exist to measure if the public health goals of screening-early initiation of treatment and enrollment to care-are being achieved. This manuscript focuses on the development of quality indicators related to the follow-up care for individuals identified with sickle cell disease and trait through screening processes. The authors used a modified Delphi method to develop the indicators. The process included a comprehensive literature review with rating of the evidence followed by ratings of draft indicators by an expert panel held in September 2012. The expert panel was nominated by leaders of various professional societies, the Health Resources and Services Administration, and the National Heart, Lung, and Blood Institute and met face to face to discuss and rate each indicator. The panel recommended nine quality indicators focused on key aspects of follow-up care for individuals with positive screens for sickle cell disease and trait. Public health programs and healthcare institutions can use these indicators to assess the quality of follow-up care and provide a basis for improvement efforts to ensure appropriate family education, early initiation of treatment, and appropriate referral to care for individuals identified with sickle cell disease and trait. PMID:27320465

  2. Active diffusion and microtubule-based transport oppose myosin forces to position organelles in cells

    Science.gov (United States)

    Lin, Congping; Schuster, Martin; Guimaraes, Sofia Cunha; Ashwin, Peter; Schrader, Michael; Metz, Jeremy; Hacker, Christian; Gurr, Sarah Jane; Steinberg, Gero

    2016-06-01

    Even distribution of peroxisomes (POs) and lipid droplets (LDs) is critical to their role in lipid and reactive oxygen species homeostasis. How even distribution is achieved remains elusive, but diffusive motion and directed motility may play a role. Here we show that in the fungus Ustilago maydis ~95% of POs and LDs undergo diffusive motions. These movements require ATP and involve bidirectional early endosome motility, indicating that microtubule-associated membrane trafficking enhances diffusion of organelles. When early endosome transport is abolished, POs and LDs drift slowly towards the growing cell end. This pole-ward drift is facilitated by anterograde delivery of secretory cargo to the cell tip by myosin-5. Modelling reveals that microtubule-based directed transport and active diffusion support distribution, mobility and mixing of POs. In mammalian COS-7 cells, microtubules and F-actin also counteract each other to distribute POs. This highlights the importance of opposing cytoskeletal forces in organelle positioning in eukaryotes.

  3. Genistein inhibits the proliferation of human HER2-positive cancer cells by downregulating HER2 receptor

    Directory of Open Access Journals (Sweden)

    Guodong Shen

    2013-07-01

    Full Text Available Functional Foods in Health and Disease 2013; 3(7:291-299Research Article Open AccessGenistein inhibits the proliferation of human HER2-positive cancer cells by downregulating HER2 receptorGuodong Shen, Haiying Yu, Geng Bian, Min Gao, Lingqing Liu, Min Cheng, Gan Shen, Shilian HuGeriatrics Department, Gerontology Institute, Anhui Provincial Hospital; Anhui Provincial Key Laboratory of Tumor Immunotherapy and Nutrition Therapy, Hefei 230001, ChinaCorresponding Author: Shilian Hu, Department of Geriatrics, Anhui Provincial Hospital, No. 17 Lujiang Road, Hefei 230001, China Submission date: June 9, 2013; Acceptance date: July 19, 2013; Publication date: July 20, 2013ABSTRACTBackground: It was well studied that HER2/ErbB2/p185 overexpression in human malignant cancers correlates with poor prognosis and chemo-resistance. Meanwhile, Genistein (4,5,7-trihydroxyisoflavone, a major isoflavone component of soybeans and other leguminous plants, has been shown to exhibit a potent anti-proliferative effect on some sex hormone dependent cancers. Objective: The effects of genistein on the proliferation of human HER2-overexpressing breast and ovarian cancer cell lines were investigated, and the action mechanism was explored.Methods: Western blotting, fluorescence-activated cell sorting (FACS and immunofluorescence methods, cell proliferation assay kit from Promega and cell apoptosis assay kit from Biolegend were used. The dose- or time-response relationship of genistein were observed on the HER2-negative breast cancer cell line MCF-7 or HER2-positive breast cancer cell lines BT-474 and MCF-7/Her2 derived from MCF-7, and ovarian cancer cell line SKOV-3.Results: The addition of genistein ranged from 1-10g/ml in the medium for 48 hours had a marked inhibition on the proliferation of HER2-positive cancer cell lines MCF-7/Her2, BT-474 and SKOV-3, compared with tamoxifen and DMSO control (P<0.01, and a dose-dependent response was presented. However, genistein

  4. Active diffusion and microtubule-based transport oppose myosin forces to position organelles in cells.

    Science.gov (United States)

    Lin, Congping; Schuster, Martin; Guimaraes, Sofia Cunha; Ashwin, Peter; Schrader, Michael; Metz, Jeremy; Hacker, Christian; Gurr, Sarah Jane; Steinberg, Gero

    2016-01-01

    Even distribution of peroxisomes (POs) and lipid droplets (LDs) is critical to their role in lipid and reactive oxygen species homeostasis. How even distribution is achieved remains elusive, but diffusive motion and directed motility may play a role. Here we show that in the fungus Ustilago maydis ∼95% of POs and LDs undergo diffusive motions. These movements require ATP and involve bidirectional early endosome motility, indicating that microtubule-associated membrane trafficking enhances diffusion of organelles. When early endosome transport is abolished, POs and LDs drift slowly towards the growing cell end. This pole-ward drift is facilitated by anterograde delivery of secretory cargo to the cell tip by myosin-5. Modelling reveals that microtubule-based directed transport and active diffusion support distribution, mobility and mixing of POs. In mammalian COS-7 cells, microtubules and F-actin also counteract each other to distribute POs. This highlights the importance of opposing cytoskeletal forces in organelle positioning in eukaryotes. PMID:27251117

  5. Notch and TGFβ form a positive regulatory loop and regulate EMT in epithelial ovarian cancer cells.

    Science.gov (United States)

    Zhou, Jiesi; Jain, Saket; Azad, Abul K; Xu, Xia; Yu, Hai Chuan; Xu, Zhihua; Godbout, Roseline; Fu, YangXin

    2016-08-01

    Epithelial-mesenchymal transition (EMT) plays a critical role in the progression of epithelial ovarian cancer (EOC). However, the mechanisms that regulate EMT in EOC are not fully understood. Here, we report that activation of Notch1 induces EMT in EOC cells as evidenced by downregulation of E-cadherin and cytokeratins, upregulation of Slug and Snail, as well as morphological changes. Interestingly, activation of Notch1 increases TGFβ/Smad signaling by upregulating the expression of TGFβ and TGFβ type 1 receptor. Time course experiments demonstrate that inhibition of Notch by DAPT (a γ-secretase inhibitor) decreases TGFβ-induced phosphorylation of receptor Smads at late, but not at early, timepoints. These results suggest that Notch activation plays a role in sustaining TGFβ/Smad signaling in EOC cells. Furthermore, inhibition of Notch by DAPT decreases TGFβ induction of Slug and repression of E-cadherin and knockdown of Notch1 decreases TGFβ-induced repression of E-cadherin, indicating that Notch is required, at least in part, for TGFβ-induced EMT in EOC cells. On the other hand, TGFβ treatment increases the expression of Notch ligand Jagged1 and Notch target gene HES1 in EOC cells. Functionally, the combination of Notch1 activation and TGFβ treatment is more potent in promoting motility and migration of EOC cells than either stimulation alone. Taken together, our results indicate that Notch and TGFβ form a reciprocal positive regulatory loop and cooperatively regulate EMT and promote EOC cell motility and migration. PMID:27075926

  6. Deficiency of platelet-derived growth factor receptor-α-positive cells in Hirschsprung's disease colon

    Science.gov (United States)

    O’Donnell, Anne-Marie; Coyle, David; Puri, Prem

    2016-01-01

    AIM: To investigate whether the expression of platelet-derived growth factor receptor-α-positive (PDGFRα+)-cells is altered in Hirschsprung’s disease (HD). METHODS: HD tissue specimens (n = 10) were collected at the time of pull-through surgery, while colonic control samples were obtained at the time of colostomy closure in patients with imperforate anus (n = 10). Immunolabelling of PDGFRα+-cells was visualized using confocal microscopy to assess the distribution of these cells, while Western blot analysis was undertaken to quantify PDGFRα protein expression. RESULTS: Confocal microscopy revealed PDGFRα+-cells within the mucosa, myenteric plexus and smooth muscle in normal controls, with a marked reduction in PDGFRα+-cells in the HD specimens. Western blotting revealed high levels of PDGFRα protein expression in normal controls, while there was a striking decrease in PDGFRα protein expression in the HD colon. CONCLUSION: These findings suggest that the altered distribution of PDGFRα+-cells in both the aganglionic and ganglionic HD bowel may contribute to the motility dysfunction in HD. PMID:27022215

  7. Cytokeratin-positive interstitial reticulum cell sarcoma: Extranodal presentations mimicking carcinoma

    Directory of Open Access Journals (Sweden)

    Shirley Sundersingh

    2013-01-01

    Full Text Available Cytokeratin-positive interstitial reticulum cell (CIRC sarcoma is a rare type of dendritic cell tumor derived from a subset of fibroblastic reticular cells. Expression of cytokeratins and extranodal location of these tumors can lead to a misdiagnosis of carcinoma. We report two cases of CIRC sarcomas primarily involving the scalp and breast. Patients were referred with an initial diagnosis of carcinoma. Case 1 underwent wide local excision of the scalp tumor with left posterolateral neck dissection. Case 2 had modified radical mastectomy for the tumor in left breast. Histopathological examination of both specimens showed an epithelioid to spindle cell malignant tumor that co-expressed CK 8, CK 18, vimentin, and smooth muscle actin. A diagnosis of CIRC sarcoma was made. Pathologists should be aware of this subset of dendritic cell sarcoma. Carcinomas, other sarcomas of the accessory dendritic cell family, and poorly differentiated malignant tumors have to be ruled out by combination of morphology, immunohistochemistry, and electron microscopic studies.

  8. Effect of Weight Reduction on Cardiovascular Risk Factors and CD34-positive Cells in Circulation

    Directory of Open Access Journals (Sweden)

    Nina A Mikirova, Joseph J Casciari, Ronald E Hunninghake, Margaret M Beezley

    2011-01-01

    Full Text Available Being overweight or obese is associated with an increased risk for the development of non-insulin-dependent diabetes mellitus, hypertension, and cardiovascular disease. Dyslipidemia of obesity is characterized by elevated fasting triglycerides and decreased high-density lipoprotein-cholesterol concentrations. Endothelial damage and dysfunction is considered to be a major underlying mechanism for the elevated cardiovascular risk associated with increased adiposity. Alterations in endothelial cells and stem/endothelial progenitor cell function associated with overweight and obesity predispose to atherosclerosis and thrombosis.In our study, we analyzed the effect of a low calorie diet in combination with oral supplementation by vitamins, minerals, probiotics and human chorionic gonadotropin (hCG, 125-180 IUs on the body composition, lipid profile and CD34-positive cells in circulation.During this dieting program, the following parameters were assessed weekly for all participants: fat free mass, body fat, BMI, extracellular/intracellular water, total body water and basal metabolic rate. For part of participants blood chemistry parameters and circulating CD34-positive cells were determined before and after dieting.The data indicated that the treatments not only reduced body fat mass and total mass but also improved the lipid profile. The changes in body composition correlated with the level of lipoproteins responsible for the increased cardiovascular risk factors. These changes in body composition and lipid profile parameters coincided with the improvement of circulatory progenitor cell numbers.As the result of our study, we concluded that the improvement of body composition affects the number of stem/progenitor cells in circulation.

  9. In Vitro Differentiation of Insulin Secreting Cells from Mouse Bone Marrow Derived Stage-Specific Embryonic Antigen 1 Positive Stem Cells

    Directory of Open Access Journals (Sweden)

    Morteza Abouzaripour

    2016-02-01

    Full Text Available Objective: Bone marrow has recently been recognized as a novel source of stem cells for the treatment of wide range of diseases. A number of studies on murine bone marrow have shown a homogenous population of rare stage-specific embryonic antigen 1 (SSEA-1 positive cells that express markers of pluripotent stem cells. This study focuses on SSEA-1 positive cells isolated from murine bone marrow in an attempt to differentiate them into insulin-secreting cells (ISCs in order to investigate their differentiation potential for future use in cell therapy. Materials and Methods: This study is an experimental research. Mouse SSEA-1 positive cells were isolated by Magnetic-activated cell sorting (MACS followed by characterization with flow cytometry. Induced SSEA-1 positive cells were differentiated into ISCs with specific differentiation media. In order to evaluate differentiation quality and analysis, dithizone (DTZ staining was use, followed by reverse transcription polymerase chain reaction (RT-PCR, immunocytochemistry and insulin secretion assay. Statistical results were analyzed by one-way ANOVA. Results: The results achieved in this study reveal that mouse bone marrow contains a population of SSEA-1 positive cells that expresses pluripotent stem cells markers such as SSEA-1, octamer-binding transcription factor 4 (OCT-4 detected by immunocytochemistry and C-X-C chemokine receptor type 4 (CXCR4 and stem cell antigen-1 (SCA-1 detected by flow cytometric analysis. SSEA-1 positive cells can differentiate into ISCs cell clusters as evidenced by their DTZ positive staining and expression of genes such as Pdx1 (pancreatic transcription factors, Ngn3 (endocrine progenitor marker, Insulin1 and Insulin2 (pancreaticβ-cell markers. Additionally, our results demonstrate expression of PDX1 and GLUT2 protein and insulin secretion in response to a glucose challenge in the differentiated cells. Conclusion: Our study clearly demonstrates the potential of SSEA-1

  10. Common Effects on Cancer Cells Exerted by a Random Positioning Machine and a 2D Clinostat.

    Science.gov (United States)

    Svejgaard, Benjamin; Wehland, Markus; Ma, Xiao; Kopp, Sascha; Sahana, Jayashree; Warnke, Elisabeth; Aleshcheva, Ganna; Hemmersbach, Ruth; Hauslage, Jens; Grosse, Jirka; Bauer, Johann; Corydon, Thomas Juhl; Islam, Tawhidul; Infanger, Manfred; Grimm, Daniela

    2015-01-01

    In this study we focused on gravity-sensitive proteins of two human thyroid cancer cell lines (ML-1; RO82-W-1), which were exposed to a 2D clinostat (CLINO), a random positioning machine (RPM) and to normal 1g-conditions. After a three (3d)- or seven-day-culture (7d) on the two devices, we found both cell types growing three-dimensionally within multicellular spheroids (MCS) and also cells remaining adherent (AD) to the culture flask, while 1g-control cultures only formed adherent monolayers, unless the bottom of the culture dish was covered by agarose. In this case, the cytokines IL-6 and IL-8 facilitated the formation of MCS in both cell lines using the liquid-overlay technique at 1g. ML-1 cells grown on the RPM or the CLINO released amounts of IL-6 and MCP-1 into the supernatant, which were significantly elevated as compared to 1g-controls. Release of IL-4, IL-7, IL-8, IL-17, eotaxin-1 and VEGF increased time-dependently, but was not significantly influenced by the gravity conditions. After 3d on the RPM or the CLINO, an accumulation of F-actin around the cellular membrane was detectable in AD cells of both cell lines. IL-6 and IL-8 stimulation of ML-1 cells for 3d and 7d influenced the protein contents of ß1-integrin, talin-1, Ki-67, and beta-actin dose-dependently in adherent cells. The ß1-integrin content was significantly decreased in AD and MCS samples compared with 1g, while talin-1 was higher expressed in MCS than AD populations. The proliferation marker Ki-67 was elevated in AD samples compared with 1g and MCS samples. The ß-actin content of R082-W-1 cells remained unchanged. ML-1 cells exhibited no change in ß-actin in RPM cultures, but a reduction in CLINO samples. Thus, we concluded that simulated microgravity influences the release of cytokines in follicular thyroid cancer cells, and the production of ß1-integrin and talin-1 and predicts an identical effect under real microgravity conditions. PMID:26274317

  11. Fluorescence intensity positivity classification of Hep-2 cells images using fuzzy logic

    Science.gov (United States)

    Sazali, Dayang Farzana Abang; Janier, Josefina Barnachea; May, Zazilah Bt.

    2014-10-01

    Indirect Immunofluorescence (IIF) is a good standard used for antinuclear autoantibody (ANA) test using Hep-2 cells to determine specific diseases. Different classifier algorithm methods have been proposed in previous works however, there still no valid set as a standard to classify the fluorescence intensity. This paper presents the use of fuzzy logic to classify the fluorescence intensity and to determine the positivity of the Hep-2 cell serum samples. The fuzzy algorithm involves the image pre-processing by filtering the noises and smoothen the image, converting the red, green and blue (RGB) color space of images to luminosity layer, chromaticity layer "a" and "b" (LAB) color space where the mean value of the lightness and chromaticity layer "a" was extracted and classified by using fuzzy logic algorithm based on the standard score ranges of antinuclear autoantibody (ANA) fluorescence intensity. Using 100 data sets of positive and intermediate fluorescence intensity for testing the performance measurements, the fuzzy logic obtained an accuracy of intermediate and positive class as 85% and 87% respectively.

  12. Significance of a positive Clostridium difficile toxin test after hematopoietic stem cell transplantation.

    Science.gov (United States)

    Akahoshi, Yu; Kimura, Shun-Ichi; Nakano, Hirofumi; Harada, Naonori; Kameda, Kazuaki; Ugai, Tomotaka; Wada, Hidenori; Yamasaki, Ryoko; Ishihara, Yuko; Kawamura, Koji; Sakamoto, Kana; Ashizawa, Masahiro; Sato, Miki; Terasako-Saito, Kiriko; Nakasone, Hideki; Kikuchi, Misato; Yamazaki, Rie; Kanda, Junya; Kako, Shinichi; Nishida, Junji; Kanda, Yoshinobu

    2016-06-01

    Patients with hematological malignancies show a high prevalence of asymptomatic colonization with Clostridium difficile (CD colonization). Therefore, it is difficult to distinguish CD colonization with diarrhea induced by a conditioning regimen from true Clostridium difficile infection (CDI) in hematopoietic stem cell transplantation (HSCT) recipients. We retrospectively analyzed 308 consecutive patients who underwent a CD toxin A/B enzyme immunoassay test for diarrhea within 100 d after HSCT from November 2007 to May 2014. Thirty patients (9.7%) had positive CD toxin results, and 11 of these had positive results in subsequent tests after an initial negative result. Allogeneic HSCT, total body irradiation, stem cell source, acute leukemia, and the duration of neutropenia were significantly correlated with positive CD toxin results. In a logistic regression model, allogeneic HSCT was identified as a significant risk factor (odds ratio 18.6, p < 0.01). In an analysis limited to within 30 d after the conditioning regimen, the duration of neutropenia was the sole risk factor (odds ratio 10.4, p < 0.01). There were no distinctive clinical features for CDI, including the onset or duration of diarrhea. In conclusion, although CDI may be overdiagnosed in HSCT recipients, it is difficult to clinically distinguish between CDI and CD colonization. PMID:27019071

  13. Simultaneous positioning of cells into two-dimensional arrays using ultrasound.

    Science.gov (United States)

    Neild, Adrian; Oberti, Stefano; Radziwill, Gerald; Dual, Jürg

    2007-08-01

    Contactless simultaneous positioning of micrometer-sized particles in suspension (e.g., copolymer beads, living cells, silicon microparts) can be performed using ultrasound. Current devices are capable of collecting particles into planes or lines by exciting a resonance in the fluid by means of a piezoelectric transducer located beneath the fluidic cavity and are designed such that a one-dimensional pressure field is created. The focus of this work is to collect cells in distinct point locations for potential drug screening array applications. A device to create two-dimensional arrays of cells within a micromachined chamber is described. The chamber is etched into a silicon wafer and sealed with glass; on the underside of the silicon layer a piezoelectric actuator is attached. A signal is applied to each of two orthogonally aligned strips electrodes defined on the surface of the piezoelectric plate. These two strip electrodes create independently addressable approximately one-dimensional pressure fields. It is shown that by applying the same signal to each electrode a diagonally aligned grid of cells can be produced. However, the independence of the two electrodes allows the application of two signals with slightly different frequencies to be applied which creates a grid of circular cell clumps highly suitable for the identified application. (c) PMID:17187440

  14. Mesotheliomas show higher hyaluronan positivity around tumor cells than metastatic pulmonary adenocarcinomas.

    Science.gov (United States)

    Törrönen, Kari; Soini, Ylermi; Pääkkö, Paavo; Parkkinen, Jyrki; Sironen, Reijo; Rilla, Kirsi

    2016-10-01

    Hyaluronan is a unique glycosaminoglycan of the extracellular matrix, abundant in normal connective tissues but highly increased in many pathological conditions like cancer. Mesothelioma, one of the most malignant cancer types, is associated with high content of hyaluronan, with elevated levels of hyaluronan in pleural effusions and serum of the patients. Metastatic lung adenocarcinomas are typically less aggressive and have a better prognosis as compared to mesotheliomas, a reason why it is highly important to find reliable tools to differentiate these cancer types. The main purpose of this study was to evaluate the amount of hyaluronan, hyaluronan producing synthases (HAS's) and hyaluronan receptor CD44, in mesothelioma and metastatic lung adenocarcinomas. Furthermore, we wanted to clarify the role of hyaluronan, CD44 and HAS's as putative markers for differentiating malignant mesothelioma from metastatic lung adenocarcinomas. The main finding of this study was that mesotheliomas are significantly more positive for hyaluronan staining than metastatic adenocarcinomas. Unexceptionally, a trend of CD44 positivity of stromal cells was higher in adenocarcinomas as compared to mesotheliomas. However, no statistically significant differences were found between the staining of any of the HAS isoenzymes either in tumor cells or stromal cells of different groups of cases. The results show that there are significant differences in hyaluronan content between metastatic lung adenocarcinomas and mesotheliomas. However, as previous studies have suggested, hyaluronan alone is not a sufficient independent marker for diagnostic differentiation of these cancer types, but could be utilized as a combination together with other specific markers. PMID:26912058

  15. HPV vaccine stimulates cytotoxic activity of killer dendritic cells and natural killer cells against HPV-positive tumour cells

    OpenAIRE

    Van den Bergh, Johan M J; Guerti, Khadija; Willemen, Yannick; Lion, Eva; Cools, Nathalie; Goossens, Herman; Vorsters, Alex; Van Tendeloo, Viggo F.I.; Anguille, Sébastien; Van Damme, Pierre; Smits, Evelien L. J. M.

    2014-01-01

    Cervarix™ is approved as a preventive vaccine against infection with the human papillomavirus (HPV) strains 16 and 18, which are causally related to the development of cervical cancer. We are the first to investigate in vitro the effects of this HPV vaccine on interleukin (IL)-15 dendritic cells (DC) as proxy of a naturally occurring subset of blood DC, and natural killer (NK) cells, two innate immune cell types that play an important role in antitumour immunity. Our results show that exposur...

  16. [Novel calretinin-positive cells with polymorphous spines in the mouse forebrain during early postnatal ontogenesis].

    Science.gov (United States)

    Revishchin, A V; Okhotin, V E; Pavlova, G V

    2009-01-01

    Using an immunocytochemical method for calretinin (CR) detection, we have earlier described (Morfologiya, 2009 v. 135. No. 3, p. 7-19) the population of previously unknown mono- and bipolar cells with polymorphous spines (PS) covering their cell bodies and processes, in adult mice forebrain structures adjacent to anterior horn of lateral ventricle. CR-positive spiny (CR+PS) cells were negative to GAD67 and were detected in the white matter and in layers V and VI of frontal area of dorsomedial cortex close to the cingulum, in in rostro-dorsal part of the caudate nucleus-putamen complex, anterior olfactory nucleus and in subependymal layer of the dorso-lateral angle of the lateral ventricle. In this work, the distribution of these cells in 7-day-old mice was studied. Comparative topographical analysis of definitive and early CR+PS cells demonstrated that in 7-day-old mice CR+PS cells were absent from the areas of their localization in adult animals - anterior olfactory nucleus, cortical plate and inner portion of neostriatum. Meanwhile, some CR+PS-like cells were detected in 7-day-old mice inside the rostral migratory route, close to neostriatum anterior boundary, along the dorsal border between neostriatum and corpus callosum, subependymal layer of lateral wall of the lateral ventricle, and in the cingulum area. These findings are indicative of the possible postnatal appearance of CR+PS cells. To test this hypothesis, the experiments were conducted in which bromodeoxyuridine (BrdU) was administered to the mice on their postnatal days 2-4 with the subsequent study of the brain sections of these animals sacrificed on their postnatal day 20. Double immunolabeling of these sections for CR and BrdU has detected the presence of CR+PS cells that contained postnatally administered BrdU. These results strongly suggest that, at least, some portion of CR+PS cells have their mitosis postnatally. It may be assumed, that CR+PS cells migrate to the sites of their distribution in

  17. Graft-versus-leukemia effects of Wilms' tumor 1 protein-specific cytotoxic T lymphocytes in patients with chronic myeloid leukemia after allogeneic hematopoietic stem cell transplantation

    Institute of Scientific and Technical Information of China (English)

    WANG Zhi-dong; LI Dan; HUANG Xiao-jun

    2010-01-01

    Background The role of Wilms' tumor 1 protein (WT1)-specific cytotoxic T cells (CTL) in eradicating chronic myeloid leukemia (CML) cells is to be established. The aim of this study was to determine whether WT1 contributed to the graft-versus-leukemia effects (GVLE) for CML following allogeneic hematopoietic stem cell transplantation (HSCT). Methods High-resolution human leukocyte antigen (HLA) class I genotyping was performed by sequence-specific polymerase chain reaction (PCR). Fifteen HLA-A~*2402 patients with CML who underwent allogeneic HSCT were enrolled in this study. We monitored the frequency of WT1-specific CTL by pentamer assay and the molecular minimal residual disease by real-time quantitative PCR.Results A CD8~+ T-cell response to WT1 was observed in 14 of 15 patients after HSCT. The median frequencies of WT1-CTL were 0.54%, 0.62%, 0.81% and 1.28% (%CD8) on days 30, 60, 90 and 180, respectively. The median frequency of WT1-CTL (1.38%) in patients with molecular remission (MoR) was significantly higher than that in those without MoR (0.38%) on day 30, while no significant differences between them were detected on days 60, 90 and 180. The increase of WT1-CTL was associated with a decrease in bcr-abl expression and MoR; and the decrease of WT1-CTL was associated with an increase in bcr-abl expression, suggesting a WT1 -driven GVL effect. WT1-CTL had a predominant effector-memory phenotype (CD45RO~+CD27~-CD57~+).Conclusions The emergence of WT1-CTL with an effector-memory phenotype is associated with GVLE in CML patients after HSCT. This will pave the way for the WT1 vaccines to enhance GVLE after HSCT in CML.

  18. Dermatopathic Lymphadenopathy With Increased IgG4-Positive Plasma Cells

    OpenAIRE

    Nakayama, Shoko; Yokote, Taiji; Akioka, Toshikazu; Miyoshi, Takuji; Hirata, Yuji; Hiraoka, Nobuya; Iwaki, Kazuki; Takayama, Ayami; Nishiwaki, Uta; Masuda, Yuki; Hanafusa, Toshiaki; Nishimura, Yasuichiro; Tsuji, Motomu

    2015-01-01

    Abstract Both dermatopathic lymphadenopathy (DL) and immunoglobulin G4-related disease (IgG4-RD) are frequently complicated with allergic diseases. However, the relationship between DL and IgG4-RD is not well known. To clarify this relationship on the basis of clinical and pathological findings, including IgG4-positive (IgG4+) plasma cell infiltration in lymph nodes (LNs) of DL patients, we analyzed LNs of 11 DL patients using immunostaining of IgG, IgG4, forkhead box P3 (FOXP3), transforming...

  19. Positioning of the NOR-bearing chromosomes in relation to nucleoli in daughter cells after mitosis

    Czech Academy of Sciences Publication Activity Database

    Kalmárová, Markéta; Smirnov, Evgeny; Kováčik, Lubomír; Popov, Alexey; Raška, Ivan

    2008-01-01

    Roč. 57, č. 3 (2008), s. 421-425. ISSN 0862-8408 R&D Projects: GA ČR(CZ) GA304/06/1691 Grant ostatní: GA ČR(CZ) GA304/06/1662; GA MŠk(CZ) LC535; Wellcome Trust(XE) 075834/04/Z Institutional research plan: CEZ:AV0Z50110509 Keywords : chromosome positioning * NORs * daughter cells Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.653, year: 2008

  20. False positive indium-111 white blood cell scan in a closed clavicle fracture

    International Nuclear Information System (INIS)

    Aggressive treatment of the multiply injured patient often requires early fixation of many fractures, some of which may be open. Often, patients develop postoperative fevers requiring a thorough workup to rule out infection. Recently, indium-111 white blood cell (WBC) imaging has become a valuable adjunct in the diagnosis of acute infection. The patient described had a simple, closed clavicle fracture with markedly increased activity on an indium-111 WBC scan obtained for fever workup. This subsequently proved to be a normal, healing, noninfected fracture by other diagnostic techniques. Noninfected, simple closed fractures should be added to the list of causes for a false-positive indium-111 WBC scan

  1. BRAF V600E-Positive Multisite Langerhans Cell Histiocytosis in a Preterm Neonate

    Directory of Open Access Journals (Sweden)

    Sara V. Bates

    2013-10-01

    Full Text Available Hemorrhagic pustules with a “blueberry muffin” appearance accompanied by respiratory failure in a neonate present a challenging differential diagnosis that includes infections and neoplasms. We present a case of multiorgan, multisite Langerhans cell histiocytosis (LCH, positive for the oncogenic BRAF V600E mutation, in a preterm neonate. Infants with LCH pose a diagnostic challenge due to their heterogeneous presentations. This case is unusual in that the newborn presented with severe multiorgan involvement. Due to the rare incidence, wide spectrum of clinical manifestations, and high mortality rate, clinicians must maintain a high index of suspicion for LCH.

  2. New calretinin-positive cells with polymorphous spines in the mouse forebrain during early postnatal ontogeny.

    Science.gov (United States)

    Revishchin, A V; Okhotin, V E; Pavlova, G V

    2010-10-01

    Immunohistochemical studies of calretinin (CR) in forebrain structures adjacent to the anterior horn of the lateral ventricle in adult mice allowed us to detect a population of previously unknown mono- and bipolar cells whose bodies and processes were coated with polymorphous spines (PS) (Morfologiya, 135, No. 3, 7-19 (2009)). CR-positive spiny (CR(+)PS) cells did not contain GAD67 and were located in the white matter and layers V-VI of the frontal area of the dorsomedial cortex close to the cingulum, the rostrodorsal part of the caudate-putamen, the anterior olfactory nucleus, and the subependyma of the dorsolateral angle of the lateral ventricle. We report here studies of the distribution of these cells in seven-day-old mice. Comparative topographic analysis of definitive and early CR(+)PS cells showed that in seven-day-old mice, CR(+)PS cells were absent from the sites at which they were seen in adults, i.e., the anterior olfactory cortex, the cortical plate, and the inner part of the neostriatum. In addition, small numbers of CR(+)PS-like cells were seen at this age within the dorsal migration pathway, at the anterior margin of the neostriatum, along the dorsal border of the neostriatum with the corpus callosum, in the subependymal layer of the lateral wall of the lateral ventricle, and in the cingulum area. These data demonstrate that CR(+)PS cells may have a postnatal origin. Experiments to verify this hypothesis were performed using postnatal administration of bromodeoxyuridine (BrdU) to mice aged 2-4 days, followed by assessment of brain sections fixed at age 20 days. Double immunolabeling of sections for CR and BrdU demonstrated the presence of CR(+)PS cells containing postnatally supplied BrdU. These data provide evidence that at least some CR(+)PS cells undergo mitosis at postnatal age. In all probability, during the period from 7 to 20 days of postnatal development, CR(+)PS cells migrate to the sites that they occupy in adult animals. PMID:20721693

  3. Position-effect variegation revisited: HUSHing up heterochromatin in human cells.

    Science.gov (United States)

    Timms, Richard T; Tchasovnikarova, Iva A; Lehner, Paul J

    2016-04-01

    Much of what we understand about heterochromatin formation in mammals has been extrapolated from forward genetic screens for modifiers of position-effect variegation (PEV) in the fruit fly Drosophila melanogaster. The recent identification of the HUSH (Human Silencing Hub) complex suggests that more recent evolutionary developments contribute to the mechanisms underlying PEV in human cells. Although HUSH-mediated repression also involves heterochromatin spreading through the reading and writing of the repressive H3K9me3 histone modification, clear orthologues of HUSH subunits are not found in Drosophila but are conserved in vertebrates. Here we compare the insights into the mechanisms of PEV derived from genetic screens in the fly, the mouse and in human cells, review what is currently known about the HUSH complex and discuss the implications of HUSH-mediated silencing for viral latency. Future studies will provide mechanistic insight into HUSH complex function and reveal the relationship between HUSH and other epigenetic silencing complexes. PMID:26853531

  4. Anti-oxidative therapy with oral dapsone improved HCV antibody positive annular elastolytic giant cell granuloma.

    Science.gov (United States)

    Igawa, K; Maruyama, R; Katayama, I; Nishioka, K

    1997-05-01

    A 72-year-old fisherman who was positive for the HCV antibody developed an annular, erythematous, infiltrated lesions on sun-exposed areas. The lesions were diagnosed as annular elastolytic giant cell granuloma both clinically and histologically. Topical corticosteroid and cryotherapy with liquid nitrogen for several months failed to improve the lesions. We then started dapsone, a known anti-oxidant, at 50 mg/day. A month later, the margins of the erythematous lesions faded, and the infiltration gradually decreased. No recurrence has been observed for one year after the start of the therapy. Anti-oxidative therapy appears to be effective for annular elastolytic giant cell granuloma and could be an alternate therapy for refractory granulomatous disease. PMID:9198323

  5. ALK-positive anaplastic large cell lymphoma with soft tissue involvement in a young woman

    Directory of Open Access Journals (Sweden)

    Gao KH

    2016-07-01

    Full Text Available Kehai Gao, Hongtao Li, Caihong Huang, Huazhuang Li, Jun Fang, Chen Tian Department of Orthopaedics, Yidu Central Hospital, Shandong, People’s Republic of China Introduction: Anaplastic large cell lymphoma (ALCL is a type of non-Hodgkin lymphoma that has strong expression of CD30. ALCL can sometimes involve the bone marrow, and in advanced stages, it can produce destructive extranodal lesions. But anaplastic large cell lymphoma kinase (ALK+ ALCL with soft tissue involvement is very rare.Case report: A 35-year-old woman presented with waist pain for over 1 month. The biopsy of soft tissue lesions showed that these cells were positive for ALK-1, CD30, TIA-1, GranzymeB, CD4, CD8, and Ki67 (90%+ and negative for CD3, CD5, CD20, CD10, cytokeratin (CK, TdT, HMB-45, epithelial membrane antigen (EMA, and pan-CK, which identified ALCL. After six cycles of Hyper-CVAD/MA regimen, she achieved partial remission. Three months later, she died due to disease progression.Conclusion: This case illustrates the unusual presentation of ALCL in soft tissue with a bad response to chemotherapy. Because of the tendency for rapid progression, ALCL in young adults with extranodal lesions are often treated with high-grade chemotherapy, such as Hyper-CVAD/MA. Keywords: anaplastic large cell lymphoma, ALK+, soft tissue involvement, Hyper-CVAD/MA

  6. A positive feedback pathway of estrogen biosynthesis in breast cancer cells is contained by resveratrol

    International Nuclear Information System (INIS)

    Cytochrome P450 (CYP) 19 enzyme or aromatase catalyses the rate-determining step of estrogen synthesis. The transcriptional control of CYP19 gene is highly specific in different cell types, for instance, Promoter I.3/II is commonly used for regulation in breast cancer cells. Recently, a positive feedback pathway for estrogen synthesis has been identified in ERα expressing SK-BR-3 cells. CYP19 mRNA abundance and activity are increased in this pathway and the promoter usage is switched from Promoter I.3/II to I.1 through a non-genomic process. In the present study, effect of the phytocompound resveratrol on this Promoter I.1-controlled expression of aromatase was investigated. Results indicated that resveratrol reduced the estradiol-induced mRNA abundance in SK-BR-3 cells expressing ERα. Luciferase reporter gene assays revealed that resveratrol could also repress the transcriptional control dictated by Promoter I.1. Since the ERE-driven luciferase activity was not repressed by resveratrol, the nuclear events of estrogen were unlikely to be suppressed by resveratrol. Instead the phytochemical reduced the amount of ERK activated by estradiol, which could be the pathway responsible for Promoter I.1 transactivation and the induced CYP19 expression. The present study illustrated that resveratrol impeded the non-genomic induction of estrogen on CYP19

  7. Live Cell Reporter Systems for Positive-Sense Single Strand RNA Viruses.

    Science.gov (United States)

    Ren, Linzhu; Peng, Zhiyuan; Chen, Xinrong; Ouyang, Hongsheng

    2016-04-01

    Cell-based reporter systems have facilitated studies of viral replication and pathogenesis, virus detection, and drug susceptibility testing. There are three types of cell-based reporter systems that express certain reporter protein for positive-sense single strand RNA virus infections. The first type is classical reporter system, which relies on recombinant virus, reporter virus particle, or subgenomic replicon. During infection with the recombinant virus or reporter virus particle, the reporter protein is expressed and can be detected in real time in a dose-dependent manner. Using subgenomic replicon, which are genetically engineered viral RNA molecules that are capable of replication but incapable of producing virions, the translation and replication of the replicon could be tracked by the accumulation of reporter protein. The second type of reporter system involves genetically engineered cells bearing virus-specific protease cleavage sequences, which can sense the incoming viral protease. The third type is based on viral replicase, which can report the specific virus infection via detection of the incoming viral replicase. This review specifically focuses on the major technical breakthroughs in the design of cell-based reporter systems and the application of these systems to the further understanding and control of viruses over the past few decades. PMID:26728654

  8. Real-time single-ion hit position detecting system for cell irradiation

    International Nuclear Information System (INIS)

    We have developed a real-time single-ion hit position detecting system to replace a CR-39 solid-state nuclear-track detector for cell irradiation experiments because the CR-39 takes several minutes for off-line etching. The new real-time system consists of a 500-μm-thick CaF2(Eu) scintillator, an optical microscope with a 10× objective lens, and a high-gain charge-coupled device camera. Each of the 260-MeV neon ions passing through a 100-μm-thick CR-39 sheet was detected using the real-time system in a performance test for the spatial resolution. The full width at half maxima (FWHMs) of the distances between positions detected by the real-time system and the centers of the etch pits on CR-39 were 6.5 and 6.9 μm in the x and y directions, respectively. The result shows that the system is useful for typical cultured cells of a few tens of micrometers in size

  9. SPOT14-Positive Neural Stem/Progenitor Cells in the Hippocampus Respond Dynamically to Neurogenic Regulators

    Directory of Open Access Journals (Sweden)

    Marlen Knobloch

    2014-11-01

    Full Text Available Proliferation of neural stem/progenitor cells (NSPCs in the adult brain is tightly controlled to prevent exhaustion and to ensure proper neurogenesis. Several extrinsic stimuli affect NSPC regulation. However, the lack of unique markers led to controversial results regarding the in vivo behavior of NSPCs to different stimuli. We recently identified SPOT14, which controls NSPC proliferation through regulation of de novo lipogenesis, selectively in low-proliferating NSPCs. Whether SPOT14-expressing (SPOT14+ NSPCs react in vivo to neurogenic regulators is not known. We show that aging is accompanied by a marked disappearance of SPOT14+ NSPCs, whereas running, a positive neurogenic stimulus, increases proliferation of SPOT14+ NSPCs. Furthermore, transient depletion of highly proliferative cells recruits SPOT14+ NSPCs into the proliferative pool. Additionally, we have established endogenous SPOT14 protein staining, reflecting transgenic SPOT14-GFP expression. Thus, our data identify SPOT14 as a potent marker for adult NSPCs that react dynamically to positive and negative neurogenic regulators.

  10. Real-time single-ion hit position detecting system for cell irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Satoh, Takahiro, E-mail: satoh.takahiro37@jaea.go.jp [Takasaki Advanced Radiation Research Institute, Japan Atomic Energy Agency (JAEA), 1233 Watanuki-machi, Takasaki, Gunma 370-1292 (Japan); Koka, Masahi [Takasaki Advanced Radiation Research Institute, Japan Atomic Energy Agency (JAEA), 1233 Watanuki-machi, Takasaki, Gunma 370-1292 (Japan); Kada, Wataru [Faculty of Science and Technology, Gunma University, 1-5-1 Tenjin-machi, Kiryu, Gunma 376-8515 (Japan); Yokoyama, Akihito; Kamiya, Tomihiro [Takasaki Advanced Radiation Research Institute, Japan Atomic Energy Agency (JAEA), 1233 Watanuki-machi, Takasaki, Gunma 370-1292 (Japan)

    2014-08-01

    We have developed a real-time single-ion hit position detecting system to replace a CR-39 solid-state nuclear-track detector for cell irradiation experiments because the CR-39 takes several minutes for off-line etching. The new real-time system consists of a 500-μm-thick CaF{sub 2}(Eu) scintillator, an optical microscope with a 10× objective lens, and a high-gain charge-coupled device camera. Each of the 260-MeV neon ions passing through a 100-μm-thick CR-39 sheet was detected using the real-time system in a performance test for the spatial resolution. The full width at half maxima (FWHMs) of the distances between positions detected by the real-time system and the centers of the etch pits on CR-39 were 6.5 and 6.9 μm in the x and y directions, respectively. The result shows that the system is useful for typical cultured cells of a few tens of micrometers in size.

  11. Interferon-α/β enhances temozolomide activity against MGMT-positive glioma stem-like cells.

    Science.gov (United States)

    Shen, Dong; Guo, Cheng-Cheng; Wang, Jing; Qiu, Zhi-Kun; Sai, Ke; Yang, Qun-Ying; Chen, Yin-Sheng; Chen, Fu-Rong; Wang, Jie; Panasci, Lawrence; Chen, Zhong-Ping

    2015-11-01

    Glioma is one of the most common primary tumors of the central nervous system in adults. Glioblastoma (GBM) is the most lethal type of glioma, whose 5-year survival is 9.8% at best. Glioma stem-like cells (GSCs) play an important role in recurrence and treatment resistance. MGMT is a DNA repair protein that removes DNA adducts and therefore attenuates treatment efficiency. It has been reported that interferon-α/β (IFN-α/β) downregulates the level of MGMT and sensitizes glioma cells to temozolomide. In the present study, we assessed whether IFN-α/β is able to sensitize GSCs to temozolomide by modulating MGMT expression. Upon the treatment of IFN-α/β, the efficacy of temozolomide against MGMT‑positive GSCs was markedly enhanced by combination treatment with IFN-α/β when compared with the temozolomide single agent group, and MGMT expression was markedly decreased at the same time. Further mechanistic study showed that IFN-α/β suppressed the NF-κB activity, which further mediated the sensitization of MGMT‑positive GSCs to temozolomide. Our data therefore demonstrated that the application of IFN-α/β is a promising agent with which to enhance temozolomide efficiency and reduce drug resistance, and our findings shed light on improving clinical outcomes and prolonging the survival of patients with malignant gliomas. PMID:26329778

  12. A highly restricted T-cell receptor dominates the CD8+ T-cell response to parvovirus B19 infection in HLA-A*2402-positive individuals

    DEFF Research Database (Denmark)

    Kasprowicz, V; Jeffery, K; Broliden, K;

    2006-01-01

    Six of seven HLA-A*2402-positive individuals with acute parvovirus B19 infections made vigorous CD8-positive cytotoxic T-cell (CTL) responses to the viral epitope FYTPLADQF. All responders showed highly focused T-cell receptor (TCR) usage, using almost exclusively BV5.1. The BV5.1 TCR dominated the...

  13. PIK3CA, HRAS and PTEN in human papillomavirus positive oropharyngeal squamous cell carcinoma

    International Nuclear Information System (INIS)

    Recent genomic evidence suggests frequent phosphatidylinositide 3-kinase (PI3K) pathway activation in human papillomavirus (HPV) positive oropharyngeal squamous cell carcinoma. Mutations/amplification of the gene encoding p110α catalytic subunit of phosphoinositide 3-kinase (PIK3CA), loss of phosphatase and tensin homolog (PTEN) and HRAS mutations are known to activate PI3K pathway. PIK3CA mutations were identified by Sanger sequencing in 23 of 75 (31%) HPV-positive oropharyngeal carcinomas, including exon 9 (p.E545K [n = 10] and p.E542K [n = 5]) or exon 20 (p.H1047Y, n = 2) mutations. Five rare and one novel (p.R537Q) PIK3CA mutations were identified. HRAS mutation (p.Q61L) was detected in 1 of 62 tested cases. PIK3CA amplification by fluorescence in situ hybridization (FISH) was identified in 4 cases (4/21, 20%), while PTEN loss was seen in 7 (7/21, 33%) cases (chromosome 10 monosomy [n = 4], homozygous deletion [n = 3]). Overall, genetic alterations that likely lead to PI3K pathway activation were identified in 34 of 75 cases (45%) and did not correlate with disease specific survival. These findings offer a molecular rationale for therapeutic targeting of PI3K pathway in patients with HPV-positive oropharyngeal carcinoma

  14. Detection of (Leu-7)-positive cells with NK activity in human gingival tissues from patients with periodontitis

    Energy Technology Data Exchange (ETDEWEB)

    Komiyama, K.; Hirsch, H.Z.; Mestecky, J.; Moro, I.

    1986-03-05

    Natural killer (NK) cells have been identified in peripheral blood, lymphoid tissue and more recently in gut mucosa and may be involved in the regulation of immunoglobulin synthesis. They have assayed gingival tissues obtained from 25 periodontitis patients, for the presence and activity of NK cells. Routine histological techniques demonstrated an inflammatory infiltrate dominated by plasma cells and B lymphocytes. Indirect staining procedures with a biotin-labeled mouse anti-human, Leu-7 antibody revealed the presence of numerous positive cells accompanying the inflammatory cellular infiltrate in perivascular areas. Several specimens demonstrated positive-staining cells in the epithelium as well. Few cells were observed in histologically uninflammed areas. Single cell suspension obtained by collagenase digestion of 5 gingival samples were used in /sup 51/Cr release cytotoxicity assay against K562 cells. Three of the five samples were positive in this assay. The finding of Leu-7-positive cells in areas of intense plasma cell foci but not in uninflammed areas, may support a role for these cells in the regulation of immunoglobulin synthesis in oral mucosal tissues.

  15. Detection of (Leu-7)-positive cells with NK activity in human gingival tissues from patients with periodontitis

    International Nuclear Information System (INIS)

    Natural killer (NK) cells have been identified in peripheral blood, lymphoid tissue and more recently in gut mucosa and may be involved in the regulation of immunoglobulin synthesis. They have assayed gingival tissues obtained from 25 periodontitis patients, for the presence and activity of NK cells. Routine histological techniques demonstrated an inflammatory infiltrate dominated by plasma cells and B lymphocytes. Indirect staining procedures with a biotin-labeled mouse anti-human, Leu-7 antibody revealed the presence of numerous positive cells accompanying the inflammatory cellular infiltrate in perivascular areas. Several specimens demonstrated positive-staining cells in the epithelium as well. Few cells were observed in histologically uninflammed areas. Single cell suspension obtained by collagenase digestion of 5 gingival samples were used in 51Cr release cytotoxicity assay against K562 cells. Three of the five samples were positive in this assay. The finding of Leu-7-positive cells in areas of intense plasma cell foci but not in uninflammed areas, may support a role for these cells in the regulation of immunoglobulin synthesis in oral mucosal tissues

  16. Vitamin D status is positively correlated with regulatory T cell function in patients with multiple sclerosis.

    Directory of Open Access Journals (Sweden)

    Joost Smolders

    Full Text Available BACKGROUND: In several autoimmune diseases, including multiple sclerosis (MS, a compromised regulatory T cell (Treg function is believed to be critically involved in the disease process. In vitro, the biologically active metabolite of vitamin D has been shown to promote Treg development. A poor vitamin D status has been linked with MS incidence and MS disease activity. In the present study, we assess a potential in vivo correlation between vitamin D status and Treg function in relapsing remitting MS (RRMS patients. METHODOLOGY/PRINCIPAL FINDINGS: Serum levels of 25-hydroxyvitamin D (25(OHD were measured in 29 RRMS patients. The number of circulating Tregs was assessed by flow-cytometry, and their functionality was tested in vitro in a CFSE-based proliferation suppression assay. Additionally, the intracellular cytokine profile of T helper cells was determined directly ex-vivo by flow-cytometry. Serum levels of 25(OHD correlated positively with the ability of Tregs to suppress T cell proliferation (R = 0.590, P = 0.002. No correlation between 25(OHD levels and the number of Tregs was found. The IFN-gamma/IL-4 ratio (Th1/Th2-balance was more directed towards IL-4 in patients with favourable 25(OHD levels (R = -0.435, P = 0.023. CONCLUSIONS/SIGNIFICANCE: These results show an association of high 25(OHD levels with an improved Treg function, and with skewing of the Th1/Th2 balance towards Th2. These findings suggest that vitamin D is an important promoter of T cell regulation in vivo in MS patients. It is tempting to speculate that our results may not only hold for MS, but also for other autoimmune diseases. Future intervention studies will show whether modulation of vitamin D status results in modulation of the T cell response and subsequent amelioration of disease activity.

  17. Low dose irradiation of thyroid cells reveals a unique transcriptomic and epigenetic signature in RET/PTC-positive cells

    Energy Technology Data Exchange (ETDEWEB)

    Abou-El-Ardat, Khalil, E-mail: kabouela@sckcen.be [Radiobiology Unit, Molecular and Cellular Biology, GKD Building, Studiecentrum voor Kernenergie - Centre d' Etude de l' Energie Nucleaire (SCK-CEN), Boeretang 200, 2400 Mol (Belgium); Department of Molecular Biotechnology, Faculty of Bioscience Engineering, Universiteit Gent, 9000 Ghent (Belgium); Monsieurs, Pieter [Radiobiology Unit, Molecular and Cellular Biology, GKD Building, Studiecentrum voor Kernenergie - Centre d' Etude de l' Energie Nucleaire (SCK-CEN), Boeretang 200, 2400 Mol (Belgium); Anastasov, Natasa; Atkinson, Mike [Department of Radiation Sciences, Helmholtz Zentrum Muenchen, Munich (Germany); Derradji, Hanane [Radiobiology Unit, Molecular and Cellular Biology, GKD Building, Studiecentrum voor Kernenergie - Centre d' Etude de l' Energie Nucleaire (SCK-CEN), Boeretang 200, 2400 Mol (Belgium); De Meyer, Tim [Department of Molecular Biotechnology, Faculty of Bioscience Engineering, Universiteit Gent, 9000 Ghent (Belgium); Department of Applied Mathematics, Biometrics and Process Control, Faculty of Bioscience Engineering, Universiteit Gent, 9000 Ghent (Belgium); Bekaert, Sofie [Clinical Research Center, Faculty for Medicine and Health Sciences, Universiteit Gent, 185 De Pintelaan, 9000 Ghent (Belgium); Van Criekinge, Wim [Department of Molecular Biotechnology, Faculty of Bioscience Engineering, Universiteit Gent, 9000 Ghent (Belgium); and others

    2012-03-01

    The high doses of radiation received in the wake of the Chernobyl incident and the atomic bombing of Hiroshima and Nagasaki have been linked to the increased appearance of thyroid cancer in the children living in the vicinity of the site. However, the data gathered on the effect of low doses of radiation on the thyroid remain limited. We have examined the genome wide transcriptional response of a culture of TPC-1 human cell line of papillary thyroid carcinoma origin with a RET/PTC1 translocation to various doses (0.0625, 0.5, and 4 Gy) of X-rays and compared it to response of thyroids with a RET/PTC3 translocation and against wild-type mouse thyroids irradiated with the same doses using Affymetrix microarrays. We have found considerable overlap at a high dose of 4 Gy in both RET/PTC-positive systems but no common genes at 62.5 mGy. In addition, the response of RET/PTC-positive system at all doses was distinct from the response of wild-type thyroids with both systems signaling down different pathways. Analysis of the response of microRNAs in TPC-1 cells revealed a radiation-responsive signature of microRNAs in addition to dose-responsive microRNAs. Our results point to the fact that a low dose of X-rays seems to have a significant proliferative effect on normal thyroids. This observation should be studied further as opposed to its effect on RET/PTC-positive thyroids which was subtle, anti-proliferative and system-dependent.

  18. Low dose irradiation of thyroid cells reveals a unique transcriptomic and epigenetic signature in RET/PTC-positive cells.

    Science.gov (United States)

    Abou-El-Ardat, Khalil; Monsieurs, Pieter; Anastasov, Nataša; Atkinson, Mike; Derradji, Hanane; De Meyer, Tim; Bekaert, Sofie; Van Criekinge, Wim; Baatout, Sarah

    2012-03-01

    The high doses of radiation received in the wake of the Chernobyl incident and the atomic bombing of Hiroshima and Nagasaki have been linked to the increased appearance of thyroid cancer in the children living in the vicinity of the site. However, the data gathered on the effect of low doses of radiation on the thyroid remain limited. We have examined the genome wide transcriptional response of a culture of TPC-1 human cell line of papillary thyroid carcinoma origin with a RET/PTC1 translocation to various doses (0.0625, 0.5, and 4Gy) of X-rays and compared it to response of thyroids with a RET/PTC3 translocation and against wild-type mouse thyroids irradiated with the same doses using Affymetrix microarrays. We have found considerable overlap at a high dose of 4Gy in both RET/PTC-positive systems but no common genes at 62.5mGy. In addition, the response of RET/PTC-positive system at all doses was distinct from the response of wild-type thyroids with both systems signaling down different pathways. Analysis of the response of microRNAs in TPC-1 cells revealed a radiation-responsive signature of microRNAs in addition to dose-responsive microRNAs. Our results point to the fact that a low dose of X-rays seems to have a significant proliferative effect on normal thyroids. This observation should be studied further as opposed to its effect on RET/PTC-positive thyroids which was subtle, anti-proliferative and system-dependent. PMID:22027090

  19. Interleukin 13-positive mast cells are increased in immunoglobulin G4-related sialadenitis.

    Science.gov (United States)

    Takeuchi, Mai; Ohno, Kyotaro; Takata, Katsuyoshi; Gion, Yuka; Tachibana, Tomoyasu; Orita, Yorihisa; Yoshino, Tadashi; Sato, Yasuharu

    2015-01-01

    Interleukin (IL)-13 is a T helper 2 (Th2) cytokine that plays important roles in the pathogenesis of asthma. IL-13 induces hypersensitivity of the airways, increased mucous production, elevated serum immunoglobulin (Ig) E levels, and increased numbers of eosinophils. Many patients with IgG4-related disease have allergic backgrounds and show elevated serum IgE levels and an increase in the number of eosinophils. Upregulation of Th2/regulatory T (Treg) cytokines, including IL-13, has been detected in affected tissues of patients with IgG4-related disease. We previously reported that mast cells might be responsible for the production of the Th2/Treg cytokines IL-4, IL-10, and transforming growth factor (TGF)-β1 in IgG4-related disease. In this study, immunohistochemical analysis showed increased numbers of IL-13-positive mast cells in IgG4-related disease, which suggests that mast cells also produce IL-13 and contribute to elevation of serum IgE levels and eosinophil infiltration in IgG4-related disease. PMID:25571893

  20. GATA Factor-Dependent Positive-Feedback Circuit in Acute Myeloid Leukemia Cells

    Directory of Open Access Journals (Sweden)

    Koichi R. Katsumura

    2016-08-01

    Full Text Available The master regulatory transcription factor GATA-2 triggers hematopoietic stem and progenitor cell generation. GATA2 haploinsufficiency is implicated in myelodysplastic syndrome (MDS and acute myeloid leukemia (AML, and GATA2 overexpression portends a poor prognosis for AML. However, the constituents of the GATA-2-dependent genetic network mediating pathogenesis are unknown. We described a p38-dependent mechanism that phosphorylates GATA-2 and increases GATA-2 target gene activation. We demonstrate that this mechanism establishes a growth-promoting chemokine/cytokine circuit in AML cells. p38/ERK-dependent GATA-2 phosphorylation facilitated positive autoregulation of GATA2 transcription and expression of target genes, including IL1B and CXCL2. IL-1β and CXCL2 enhanced GATA-2 phosphorylation, which increased GATA-2-mediated transcriptional activation. p38/ERK-GATA-2 stimulated AML cell proliferation via CXCL2 induction. As GATA2 mRNA correlated with IL1B and CXCL2 mRNAs in AML-M5 and high expression of these genes predicted poor prognosis of cytogenetically normal AML, we propose that the circuit is functionally important in specific AML contexts.

  1. Nuclear Positioning

    OpenAIRE

    Gundersen, Gregg G.; Worman, Howard J.

    2013-01-01

    The nucleus is the largest organelle and is commonly depicted in the center of the cell. Yet during cell division, migration and differentiation, it frequently moves to an asymmetric position aligned with cell function. We consider the toolbox of proteins that move and anchor the nucleus within the cell and how forces generated by the cytoskeleton are coupled to the nucleus to move it. The significance of proper nuclear positioning is underscored by numerous diseases resulting from genetic al...

  2. Microchannel-connected SU-8 honeycombs by single-step projection photolithography for positioning cells on silicon oxide nanopillar arrays

    International Nuclear Information System (INIS)

    We report on the fabrication, functionalization and testing of SU-8 microstructures for cell culture and positioning over large areas. The microstructure consists of a honeycomb arrangement of cell containers interconnected by microchannels and centered on nanopillar arrays designed for promoting cell positioning. The containers have been dimensioned to trap single cells and, with a height of 50 µm, prevent cells from escaping. The structures are fabricated using a single ultraviolet photolithography exposure with focus depth in the lower part of the SU-8 resist. With optimized process parameters, microchannels of various aspect ratios are thus produced. The cell containers and microchannels serve for the organization of axonal growth between neurons. The roughly 2 µm-high and 500 nm-wide nanopillars are made of silicon oxide structured by deep reactive ion etching. In future work, beyond their cell positioning purpose, the nanopillars could be functionalized as sensors. The proof of concept of the novel microstructure for organized cell culture is given by the successful growth of interconnected PC12 cells. Promoted by the honeycomb geometry, a dense network of interconnections between the cells has formed and the intended intimate contact of cells with the nanopillar arrays was observed by scanning electron microscopy. This proves the potential of these new devices as tools for the controlled cell growth in an interconnected container system with well-defined 3D geometry. (paper)

  3. Microchannel-connected SU-8 honeycombs by single-step projection photolithography for positioning cells on silicon oxide nanopillar arrays

    Science.gov (United States)

    Larramendy, Florian; Charline Blatche, Marie; Mazenq, Laurent; Laborde, Adrian; Temple-Boyer, Pierre; Paul, Oliver

    2015-04-01

    We report on the fabrication, functionalization and testing of SU-8 microstructures for cell culture and positioning over large areas. The microstructure consists of a honeycomb arrangement of cell containers interconnected by microchannels and centered on nanopillar arrays designed for promoting cell positioning. The containers have been dimensioned to trap single cells and, with a height of 50 µm, prevent cells from escaping. The structures are fabricated using a single ultraviolet photolithography exposure with focus depth in the lower part of the SU-8 resist. With optimized process parameters, microchannels of various aspect ratios are thus produced. The cell containers and microchannels serve for the organization of axonal growth between neurons. The roughly 2 µm-high and 500 nm-wide nanopillars are made of silicon oxide structured by deep reactive ion etching. In future work, beyond their cell positioning purpose, the nanopillars could be functionalized as sensors. The proof of concept of the novel microstructure for organized cell culture is given by the successful growth of interconnected PC12 cells. Promoted by the honeycomb geometry, a dense network of interconnections between the cells has formed and the intended intimate contact of cells with the nanopillar arrays was observed by scanning electron microscopy. This proves the potential of these new devices as tools for the controlled cell growth in an interconnected container system with well-defined 3D geometry.

  4. CAR-pNK Cell Immunotherapy in CD7 Positive Leukemia and Lymphoma

    Science.gov (United States)

    2016-07-11

    Acute Myeloid Leukemia; Precursor T-Cell Lymphoblastic Leukemia-Lymphoma; T-cell Prolymphocytic Leukemia; T-cell Large Granular Lymphocytic Leukemia; Peripheral T-cell Lymphoma, NOS; Angioimmunoblastic T-cell Lymphoma; Extranodal NK/T-cell Lymphoma, Nasal Type; Enteropathy-type Intestinal T-cell Lymphoma; Hepatosplenic T-cell Lymphoma

  5. Without peripheral interference, thymic deletion is mediated in a cohort of double-positive cells without classical activation

    Science.gov (United States)

    Zhan, Yifan; Purton, Jared F.; Godfrey, Dale I.; Cole, Timothy J.; Heath, William R.; Lew, Andrew M.

    2003-01-01

    Peripheral activation can cause bystander thymocyte death by eliciting a “cytokine storm.” This event complicates in vivo studies using exogenous ligand-induced models of negative selection. A stable transgenic model that selectively eliminates peripheral CD4 cells has allowed us to analyze negative selection as direct cognate events in two T cell receptor transgenic mice, OT-II and DO11. Whereas cognate peptide induced a massive deletion in double-positive (DP) cells in mice with peripheral CD4 cells, this DP deletion was modest in mice lacking peripheral CD4 cells. Using BrdUrd and annexin V staining, we found that negative selection primarily occurs in a cohort of DP cells and the absence of single-positive (SP) cells is largely caused by reduction in the cohort of DP precursors. Moreover, the fates of DP cells and SP cells after antigen exposure were vastly different. Whereas SP cells up-regulated uniformly their CD69 and CD44 levels, increased their cell size, and survived after antigen exposure, DP cells had less CD69 and CD44 up-regulation, no size change, and promptly died. Thus, negative selection represents an “abortive” activation different from activation-induced cell death of mature T cells. PMID:12538873

  6. The distribution and number of Leu-7 (CD57 positive cells in lung tissue from patients with pulmonary fibrosis.

    Directory of Open Access Journals (Sweden)

    Yamanouchi H

    2002-04-01

    Full Text Available Leu-7 positive lymphocytes, including natural killer cells, play an important role in the immune system's surveillance function to prevent the development of cancer. The incidence of lung cancer is significantly high in patients with end-stage pulmonary fibrosis. We hypothesized that the number of Leu-7 positive cells may be decreased in areas of severe pulmonary fibrosis. To demonstrate this, Leu-7 positive cells were immunohistochemically stained in 41 lung specimens obtained from patients with idiopathic pulmonary fibrosis and pulmonary fibrosis associated with collagen vascular disorders. The number of Leu-7 positive cells was evaluated according to the pathological findings. In pathologically normal lung, Leu-7 positive cells were mostly found within the capillaries of the septa and rarely in the alveolar space or the stroma. The number of Leu-7 positive cells was 0.69 +/- 0.15 in areas of advanced fibrosis (n = 41, 2.39 +/- 0.60 in areas that had newly developeing fibrosis (n = 41, 1.14 +/- 0.57 in bronchiolitis obliterans organizing pneumonia (n = 9, and 1.35 +/- 0.87 in diffuse alveolar damage (DAD (n = 11. The number of Leu-7 positive cells in areas of newly developing fibrosis (2.39 +/- 0.60 was significantly higher than that in areas of established fibrosis (0.69 +/- 0.15, P < 0.05. Our present study demonstrates a significant decrease in the number of Leu-7 positive cells in areas of advanced fibrosis. This evidence may partly explain the high incidence of lung cancer associated with pulmonary fibrosis.

  7. Proliferation and survival signaling from both Jak2-V617F and Lyn involving GSK3 and mTOR/p70S6K/4EBP1 in PVTL-1 cell line newly established from acute myeloid leukemia transformed from polycythemia vera.

    Directory of Open Access Journals (Sweden)

    Toshikage Nagao

    Full Text Available The gain of function mutation JAK2-V617F is very frequently found in myeloproliferative neoplasms (MPNs and is strongly implicated in pathogenesis of these and other hematological malignancies. Here we report establishment of a new leukemia cell line, PVTL-1, homozygous for JAK2-V617F from a 73-year-old female patient with acute myeloid leukemia (AML transformed from MPN. PVTL-1 is positive for CD7, CD13, CD33, CD34, CD117, HLA-DR, and MPO, and has complex karyotypic abnormalities, 44,XX,-5q,-7,-8,add(11(p11.2,add(11(q23,-16,+21,-22,+mar1. Sequence analysis of JAK2 revealed only the mutated allele coding for Jak2-V617F. Proliferation of PVTL-1 was inhibited and apoptosis was induced by the pan-Jak inhibitor Jak inhibitor-1 (JakI-1 or dasatinib, which inhibits the Src family kinases as well as BCR/ABL. Consistently, the Src family kinase Lyn was constitutively activated with phosphorylation of Y396 in the activation loop, which was inhibited by dasatinib but not by JakI-1. Further analyses with JakI-1 and dasatinib indicated that Jak2-V617F phosphorylated STAT5 and SHP2 while Lyn phosphorylated SHP1, SHP2, Gab-2, c-Cbl, and CrkL to induce the SHP2/Gab2 and c-Cbl/CrkL complex formation. In addition, JakI-1 and dasatinib inactivated the mTOR/p70S6K/4EBP1 pathway and reduced the inhibitory phosphorylation of GSK3 in PVTL-1 cells, which correlated with their effects on proliferation and survival of these cells. Furthermore, inhibition of GSK3 by its inhibitor SB216763 mitigated apoptosis induced by dasatinib but not by JakI-1. Together, these data suggest that apoptosis may be suppressed in PVTL-1 cells through inactivation of GSK3 by Lyn as well as Jak2-V617F and additionally through activation of STAT5 by Jak2-V617F. It is also speculated that activation of the mTOR/p70S6K/4EBP1 pathway may mediate proliferation signaling from Jak2-V617F and Lyn. PVTL-1 cells may provide a valuable model system to elucidate the molecular mechanisms involved in

  8. Socioeconomic position and surgery for early-stage non-small-cell lung cancer

    DEFF Research Database (Denmark)

    Starr, Laila Kærgaard; Osler, Merete; Steding-Jessen, Marianne;

    2012-01-01

    explain the association with income or living alone for early-stage NSCLC patients. CONCLUSION: Early-stage NSCLC patients with low income or who live alone are less likely to undergo surgery than those with a high income or who live with a partner, even after control for possible explanatory factors......AIM: To examine possible associations between socioeconomic position and surgical treatment of patients with early-stage non-small-cell lung cancer (NSCLC). METHODS: In a register-based clinical cohort study, patients with early-stage (stages I-IIIa) NSCLC were identified in the Danish Lung Cancer...... and separately for stages I, II and IIIa. RESULTS: Of the 5538 eligible patients with stages I-IIIa NSCLC diagnosed 2001-2008, 53% underwent surgery. Higher stage, older age, being female and diagnosis early in the study period were associated with higher odds for not receiving surgery. Low disposable...

  9. Immunohistochemical positive stained p53 protein in bladder transitional cell carcinoma

    Directory of Open Access Journals (Sweden)

    Halimi Monireh

    2009-04-01

    Full Text Available Background: Molecular genetics and immunopathologic analysis of bladder cancer have shown some abnormalities in a number of genes and proteins that have been implicated in the development and progression of such tumors, mainly in the p53 pathway. Aims: To investigate the rate of positively stained p53 protein in patients with urothelial papillary carcinoma of the bladder (UCB by immunohistochemistry and its relationship with tumor grade, gender and age of the patients. Settings and Design: During the present cross-sectional study, 100 paraffin-embedded specimens of UCB, which were provided from biopsies of the bladder by transurethral access, were immunohistochemically stained and studied for p53 protein from May 2006 to May 2007 in our referral center pathology laboratory. Materials and Methods: First, 4 µm slices of paraffin sections were provided and then stained by the avidin-biotin peroxidase method. The rate of positively stained p53 protein (defined as positive nuclear staining in over 10% of the cells was assessed. This rate was also estimated and compared between grades, genders and age-related groups (< 70 years, ≥70 years. Statistical Analysis: The χ2 , Fisher′s exact test and Mann-Whitney U test were used for comparing. Results: The overall rate of positively stained specimens was 11% for nuclear p53 protein. This rate was significantly higher in females (10/29 vs. 1/71; P < 0.001; odds ratio [OR]: 0.23; 95% confidence interval [CI]: 4.43-306.08, patients with 70 or older than 70 years (8/42 vs. 3/58; P = 0.04; OR: 0.55; 95% CI: 1.07-17.39 and in high-grade tumors (10/58 vs. 1/42; P = 0.02; OR: 0.59; 95% CI: 0.01-0.95. Conclusions: The rate of positively stained p53 protein for UCB was lower in our population. This rate was also higher in females, patients with 70 or older than 70 years and high grade of UCB.

  10. Quantitative trait gene Slit2 positively regulates murine hematopoietic stem cell numbers

    Science.gov (United States)

    Waterstrat, Amanda; Rector, Kyle; Geiger, Hartmut; Liang, Ying

    2016-01-01

    Hematopoietic stem cells (HSC) demonstrate natural variation in number and function. The genetic factors responsible for the variations (or quantitative traits) are largely unknown. We previously identified a gene whose differential expression underlies the natural variation of HSC numbers in C57BL/6 (B6) and DBA/2 (D2) mice. We now report the finding of another gene, Slit2, on chromosome 5 that also accounts for variation in HSC number. In reciprocal chromosome 5 congenic mice, introgressed D2 alleles increased HSC numbers, whereas B6 alleles had the opposite effect. Using gene array and quantitative polymerase chain reaction, we identified Slit2 as a quantitative trait gene whose expression was positively correlated with the number of HSCs. Ectopic expression of Slit2 not only increased the number of the long-term colony forming HSCs, but also enhanced their repopulation capacity upon transplantation. Therefore, Slit2 is a novel quantitative trait gene and a positive regulator of the number and function of murine HSCs. This finding suggests that Slit2 may be a potential therapeutic target for the effective in vitro and in vivo expansion of HSCs without compromising normal hematopoiesis. PMID:27503415

  11. Quantitative trait gene Slit2 positively regulates murine hematopoietic stem cell numbers.

    Science.gov (United States)

    Waterstrat, Amanda; Rector, Kyle; Geiger, Hartmut; Liang, Ying

    2016-01-01

    Hematopoietic stem cells (HSC) demonstrate natural variation in number and function. The genetic factors responsible for the variations (or quantitative traits) are largely unknown. We previously identified a gene whose differential expression underlies the natural variation of HSC numbers in C57BL/6 (B6) and DBA/2 (D2) mice. We now report the finding of another gene, Slit2, on chromosome 5 that also accounts for variation in HSC number. In reciprocal chromosome 5 congenic mice, introgressed D2 alleles increased HSC numbers, whereas B6 alleles had the opposite effect. Using gene array and quantitative polymerase chain reaction, we identified Slit2 as a quantitative trait gene whose expression was positively correlated with the number of HSCs. Ectopic expression of Slit2 not only increased the number of the long-term colony forming HSCs, but also enhanced their repopulation capacity upon transplantation. Therefore, Slit2 is a novel quantitative trait gene and a positive regulator of the number and function of murine HSCs. This finding suggests that Slit2 may be a potential therapeutic target for the effective in vitro and in vivo expansion of HSCs without compromising normal hematopoiesis. PMID:27503415

  12. Positive feedback can lead to dynamic nanometer-scale clustering on cell membranes

    Energy Technology Data Exchange (ETDEWEB)

    Wehrens, Martijn; Rein ten Wolde, Pieter [FOM Institute AMOLF, Science Park 104, 1098 XG Amsterdam (Netherlands); Mugler, Andrew, E-mail: amugler@purdue.edu [FOM Institute AMOLF, Science Park 104, 1098 XG Amsterdam (Netherlands); Department of Physics, Purdue University, West Lafayette, Indiana 47907 (United States)

    2014-11-28

    Clustering of molecules on biological membranes is a widely observed phenomenon. A key example is the clustering of the oncoprotein Ras, which is known to be important for signal transduction in mammalian cells. Yet, the mechanism by which Ras clusters form and are maintained remains unclear. Recently, it has been discovered that activated Ras promotes further Ras activation. Here we show using particle-based simulation that this positive feedback is sufficient to produce persistent clusters of active Ras molecules at the nanometer scale via a dynamic nucleation mechanism. Furthermore, we find that our cluster statistics are consistent with experimental observations of the Ras system. Interestingly, we show that our model does not support a Turing regime of macroscopic reaction-diffusion patterning, and therefore that the clustering we observe is a purely stochastic effect, arising from the coupling of positive feedback with the discrete nature of individual molecules. These results underscore the importance of stochastic and dynamic properties of reaction diffusion systems for biological behavior.

  13. Squamous cell carcinoma of the anus-an opportunistic cancer in HIV-positive male homosexuals

    Institute of Scientific and Technical Information of China (English)

    Pascal Gervaz; Alexandra Calmy; Ymer Durmishi; Abdelkarim S Allal; Philippe Morel

    2011-01-01

    Squamous cell carcinoma of the anus (SCCA) is a common cancer in the human immunodeficiency virus (HIV)- infected population, and its incidence continues to increase in male homosexuals. Combined chemoradiation with mitomycin C and 5-fluorouracil was poorly tolerated by severely immunocompromised patients in the early 1990s. In the era of highly active antiretroviral therapy (HAART), however, recent data indicate that: (1) most HIV patients with anal cancer can tolerate standard chemotherapy regimens; and (2) this approach is associated with survival rates similar to those of HIV-negative patients. However, HIV-positive patients with SCCA are much younger, more likely to develop local tumor recurrence, and ultimately die from anal cancer than immune competent patients. Taken together, these findings suggest that anal cancer is an often fatal neoplasia in middle- aged HIV-positive male homosexuals. In this population, SCCA is an opportunistic disease resulting in patients with suboptimal immune function from persistent papillomaviruses (HPVs). Large-scale cancer-prevention strategies (routine anuscopy and anal papanicolaou testing) should be implemented in this population. In addition, definitive eradication of oncogenic HPVs within the anogenital mucosa of high-risk individuals might require a proactive approach with repeated vaccination.

  14. Positioning of follicular dendritic cells within the spleen controls prion neuroinvasion.

    Science.gov (United States)

    Prinz, Marco; Heikenwalder, Mathias; Junt, Tobias; Schwarz, Petra; Glatzel, Markus; Heppner, Frank L; Fu, Yang-Xin; Lipp, Martin; Aguzzi, Adriano

    2003-10-30

    Peripheral infection is the natural route of transmission in most prion diseases. Peripheral prion infection is followed by rapid prion replication in lymphoid organs, neuroinvasion and progressive neurological disease. Both immune cells and nerves are involved in pathogenesis, but the mechanisms of prion transfer from the immune to the nervous system are unknown. Here we show that ablation of the chemokine receptor CXCR5 juxtaposes follicular dendritic cells (FDCs) to major splenic nerves, and accelerates the transfer of intraperitoneally administered prions into the spinal cord. Neuroinvasion velocity correlated exclusively with the relative locations of FDCs and nerves: transfer of CXCR5-/- bone marrow to wild-type mice induced perineural FDCs and enhanced neuroinvasion, whereas reciprocal transfer to CXCR5-/- mice abolished them and restored normal efficiency of neuroinvasion. Suppression of lymphotoxin signalling depleted FDCs, abolished splenic infectivity, and suppressed acceleration of pathogenesis in CXCR5-/- mice. This suggests that prion neuroimmune transition occurs between FDCs and sympathetic nerves, and relative positioning of FDCs and nerves controls the efficiency of peripheral prion infection. PMID:14562059

  15. CD8 positive T cells express IL-17 in patients with chronic obstructive pulmonary disease

    Directory of Open Access Journals (Sweden)

    Eidelman David H

    2011-04-01

    Full Text Available Abstract Background Chronic obstructive pulmonary disease (COPD is a progressive and irreversible chronic inflammatory disease of the lung. The nature of the immune reaction in COPD raises the possibility that IL-17 and related cytokines may contribute to this disorder. This study analyzed the expression of IL-17A and IL-17F as well as the phenotype of cells producing them in bronchial biopsies from COPD patients. Methods Bronchoscopic biopsies of the airway were obtained from 16 COPD subjects (GOLD stage 1-4 and 15 control subjects. Paraffin sections were used for the investigation of IL-17A and IL-17F expression in the airways by immunohistochemistry, and frozen sections were used for the immunofluorescence double staining of IL-17A or IL-17F paired with CD4 or CD8. In order to confirm the expression of IL-17A and IL-17F at the mRNA level, a quantitative RT-PCR was performed on the total mRNA extracted from entire section or CD8 positive cells selected by laser capture microdissection. Results IL-17F immunoreactivity was significantly higher in the bronchial biopsies of COPD patients compared to control subjects (P P P P Conclusion These findings support the notion that Th17 cytokines could play important roles in the pathogenesis of COPD, raising the possibility of using this mechanism as the basis for novel therapeutic approaches.

  16. Niemann-Pick disease, type B with TRAP-positive storage cells and secondary sea blue histiocytosis

    Directory of Open Access Journals (Sweden)

    R. Saxena

    2009-09-01

    Full Text Available We present 2 cases of Niemann Pick disease, type B with secondary sea-blue histiocytosis. Strikingly, in both cases the Pick cells were positive for tartrate resistant acid phosphatase, a finding hitherto described only in Gaucher cells. This report highlights the importance of this finding as a potential cytochemical diagnostic pitfall in the diagnosis of Niemann Pick disease.

  17. Niemann-Pick disease, type B with TRAP-positive storage cells and secondary sea blue histiocytosis

    OpenAIRE

    R. Saxena; Pati, H. P.; Dutta, S.; Kar, R.; Sharma, P.

    2009-01-01

    We present 2 cases of Niemann Pick disease, type B with secondary sea-blue histiocytosis. Strikingly, in both cases the Pick cells were positive for tartrate resistant acid phosphatase, a finding hitherto described only in Gaucher cells. This report highlights the importance of this finding as a potential cytochemical diagnostic pitfall in the diagnosis of Niemann Pick disease.

  18. Erythropoietin Receptor Positive Circulating Progenitor Cells and Endothelial Progenitor Cells in Patients with Different Stages of Diabetic Retinopathy

    Institute of Scientific and Technical Information of China (English)

    Liu-mei Hu; Guo-xu Xu; Guo-tong XU; Wei-ye Li; Xia Lei; Bo Ma; Yu Zhang; Yan Yan; Ya-lan Wu; Ge-zhi Xu; Wen Ye; Ling Wang

    2011-01-01

    Objective To investigate the possible involvement of erythropoietin (EPO)/erythropoietin receptor(EPOR) system in neovascularization and vascular regeneration in diabetic retinopathy (DR).Methods EPOR positive circulating progenitor cells (CPCs: CD34+) and endothelial progenitor cells (EPCs: CD34+KDR+) were assessed by flow cytometry in type 2 diabetic patients with different stages of DR. The cohort consisted of age- and sex-matched control patients without diabetes (n=7), non-prolif-erative DR (NPDR, n=7), proliferative DR (PDR, n=8), and PDR complicated with diabetic nephropathy (PDR-DN, n=7). Results The numbers of EPOR+ CPCs and EPOR+ EPCs were reduced remarkably in NPDR compared with the control group (both P<0.01), whereas rebounded in PDR and PDR-DN groups in varying degrees. Similar changes were observed in respect of the proportion of EPOR+ CPCs in CPCs (NPDR vs.control, P< 0.01) and that of EPOR+ EPCs in EPCs (NPDR vs. control, P< 0.05). Conclusion Exogenous EPO, mediated via the EPO/EPOR system of EPCs, may alleviate the im-paired vascular regeneration in NPDR, whereas it might aggravate retinal neovascularization in PDR due to a rebound of EPOR+ EPCs associated with ischemia.

  19. Without peripheral interference, thymic deletion is mediated in a cohort of double-positive cells without classical activation

    OpenAIRE

    Zhan, Yifan; Purton, Jared F.; Godfrey, Dale I.; Cole, Timothy J.; Heath, William R.; Lew, Andrew M.

    2003-01-01

    Peripheral activation can cause bystander thymocyte death by eliciting a “cytokine storm.” This event complicates in vivo studies using exogenous ligand-induced models of negative selection. A stable transgenic model that selectively eliminates peripheral CD4 cells has allowed us to analyze negative selection as direct cognate events in two T cell receptor transgenic mice, OT-II and DO11. Whereas cognate peptide induced a massive deletion in double-positive (DP) cells in mice with peripheral ...

  20. Positional Isomers of Aspirin Are Equally Potent in Inhibiting Colon Cancer Cell Growth: Differences in Mode of Cyclooxygenase Inhibition

    OpenAIRE

    Kodela, Ravinder; Chattopadhyay, Mitali; Goswami, Satindra; Gan, Zong Yuan; Rao, Praveen P.N.; Nia, Kamran V.; Velázquez-Martínez, Carlos A.; Kashfi, Khosrow

    2013-01-01

    We compared the differential effects of positional isomers of acetylsalicylic acid (o-ASA, m-ASA, and p-ASA) on cyclooxygenase (COX) inhibition, gastric prostaglandin E2 (PGE2), malondialdehyde, tumor necrosis factor-alpha (TNF-α) levels, superoxide dismutase (SOD) activity, human adenocarcinoma colon cancer cell growth inhibition, cell proliferation, apoptosis, and cell-cycle progression. We also evaluated the gastric toxicity exerted by ASA isomers. All ASA isomers inhibit COX enzymes, but ...

  1. Hyperfractionated high-dose total body irradiation in bone marrow transplantation for Ph{sup 1}-positive acute lymphoblastic leukemia

    Energy Technology Data Exchange (ETDEWEB)

    Kikuchi, Akira; Ebihara, Yasuhiro; Mitsui, Tetsuo [Tokyo Univ. (Japan). Hospital of the Institute of Medical Science] [and others

    1998-12-01

    In two cases of Philadelphia-positive childhood acute lymphoblastic leukemia (Ph{sup 1} ALL), we performed allogeneic bone marrow transplantation (AlloBMT) with preconditioning regimen, including hyperfractionated high-dose total body irradiation (TBI) (13.5 Gy, in 9 fractions). Their disease statuses at BMT were hematological relapse in case 1 and molecular relapse in case 2. Bone marrow donors were unrelated in case 1, and HLA was a partially mismatched mother in case 2. Regimen-related toxicity was tolerable in both cases. Hematological recovery was rapid, and engraftment was obtained on day 14 in case 1 and on day 12 in case 2. BCR/ABL message in bone marrow disappeared on day 89 in case 1 and on day 19 in case 2 and throughout their subsequent clinical courses. Although short-term MTX and Cy-A continuous infusion were used for GVHD prophylaxis, grade IV GVHD was observed in case 1 and grade III in case 2. Both cases experienced hemorrhagic cystitis because of adenovirus type 11 infection. Although case 1 died of interstitial pneumonitis on day 442, case 2 has been free of disease through day 231. AlloBMT for Ph{sup 1} ALL with preconditioning regimen including hyperfractionated high-dose TBI is considered to be worth further investigation. (author)

  2. [Research Progress on CALR Mutation in the Myeloproliterative Neoplasm -Review].

    Science.gov (United States)

    Yuan, Jun; Hao, Hong-Ling; Li, Yan; Wang, Rui-Cang; Zhang, Xiao-Xia

    2016-08-01

    There is no gold diagnostic standard for BCR-ABL fusion gene negative chronic myeloproliterative neoplasm(cMPN). The following detection methods such as comprehensive bone marrow cell morphology, bone marrow pathology, genetic mutation, flow cytometry and immunohistochemical are needed to diagnose the BCR-ABL fusion gene positive cMPN. The JAK2 mutation can be used as a specific diagnostic criteria for polycythemia vera (PV), but there is no specific and sensitive indication for the JAK2 mutation-negative MPN. CALR mutation would be an indication in a certain extent. In this review, the CALR mutation detection, detection mean and its correlation with disease diagnosis and prognosis etc were summarized. PMID:27531810

  3. Autoimmune pancreatitis results from loss of TGFβ signalling in S100A4-positive dendritic cells

    Science.gov (United States)

    Boomershine, C S; Chamberlain, A; Kendall, P; Afshar-Sharif, A-R; Huang, H; Washington, M K; Lawson, W E; Thomas, J W; Blackwell, T S; Bhowmick, N A

    2009-01-01

    Background and aims: Autoimmune pancreatitis (AIP) is a poorly understood human disease affecting the exocrine pancreas. The goal of the present study was to elucidate the pathogenic mechanisms underlying pancreatic autoimmunity in a murine disease model. Methods: A transgenic mouse with an S100A4/fibroblast-specific protein 1 (FSP1) Cre-mediated conditional knockout of the transforming growth factor β (TGFβ) type II receptor, termed Tgfbr2fspKO, was used to determine the direct role of TGFβ in S100A4+ cells. Immunohistochemical studies suggested that Tgfbr2fspKO mice develop mouse AIP (mAIP) characterised by interlobular ductal inflammatory infiltrates and pancreatic autoantibody production. Fluorescence-activated cell sorting (FACS)-isolated dendritic cells (DCs) from diseased pancreata were verified to have S100A4-Cre-mediated DNA recombination. Results: The Tgfbr2fspKO mice spontaneously developed mAIP by 6 weeks of age. DCs were confirmed to express S100A4, a previously reported protein expressed by fibroblasts. Adoptive transfer of bone marrow-derived DCs from Tgfbr2fspKO mice into 2-week-old syngenic wild-type C57BL/6 mice resulted in reproduction of pancreatitis within 6 weeks. Similar adoptive transfer of wild-type DCs had no effect on pancreas pathology of the host mice. The inability to induce pancreatitis by adoptive transfer of Tgfbr2fspKO DCs in adult mice suggested a developmental event in mAIP pathogenesis. Tgfbr2fspKO DCs undergo elevated maturation in response to antigen and increased activation of naïve CD4-positive T cells. Conclusion: The development of mAIP in the Tgfbr2fspKO mouse model illustrates the role of TGFβ in maintaining myeloid DC immune tolerance. The loss of immune tolerance in myeloid S100A4+ DCs can mediate mAIP and may explain some aspects of AIP disease pathogenesis. PMID:19625278

  4. A novel fixative for immunofluorescence staining of CD133-positive glioblastoma stem cells

    OpenAIRE

    Sherman, Jonathan H.; Redpath, Gerard T.; Redick, Jan A.; Purow, Benjamin W.; Laws, Edward R.; Jane, John A.; Shaffrey, Mark E.; Hussaini, Isa M.

    2011-01-01

    Isolation of glioblastoma stem cells requires incubation of tumor cells in a neural stem cell media. Neurospheres containing these glioblastoma stem cells are formed after approximately a five-day period. These cells can then be analyzed for the presence of stem cell markers. Immunofluorescence staining for these markers can serve as a valuable tool for analyzing the intact neurosphere directly in stem cell media. Here we present the use of a novel fixative (1,4-benzoquinone) for immunofloure...

  5. P-glycoprotein is expressed and causes resistance to chemotherapy in EBV-positive T-cell lymphoproliferative diseases

    International Nuclear Information System (INIS)

    Epstein–Barr virus-positive T-cell lymphoproliferative diseases (EBV-T-LPDs) are rare lymphomas with poor prognosis. Although chemotherapeutic strategies such as CHOP have been often selected, they have exhibited only limited efficacy. To clarify the mechanism of chemoresistance, we examined P-glycoprotein (P-gp) expression. P-gp acts as an energy-dependent efflux pump that excretes drugs from the cytoplasm, resulting in low-intracellular drug concentrations and poor sensitivity to chemotherapy. We examined P-gp expression in EBV-positive cells by immunohistochemistry staining in three patients of EBV-T-LPDs and the expression was detected in all patients. We also examined mdr1 mRNA expression by reverse-transcriptase polymerase-chain reaction (RT-PCR) in EBV-positive tumor cells from these patients and additional three patients. The expression was detected in all examined patients. In five EBV-T-LPDs patients, P-gp function was detected by Rhodamine-123 efflux assay in these cells. The efflux was inhibited by treatment with a P-gp inhibitor, cyclosporine A (CsA). We also examined and detected P-gp expression in EBV-positive T-cell lines SNT8 and SNT16 established from EBV-T-LPDs patients, by RT-PCR and western blotting. The function was also detected by Rhodamine-123 efflux in these cell lines. Inhibition and knock down of P-gp by CsA and siRNA, respectively, enhanced etoposide- and doxorubicin-induced cell death in the EBV-positive T-cell lines. Finally, we infected the T-cell line MOLT4 with EBV, and found that mdr1 mRNA expression and Rhodamine 123 efflux were upregulated after infection. These results indicated that enhanced P-gp expression contributed to the chemoresistance of EBV-T-LPDs

  6. Diethylstilbestrol alters positive and negative selection of T cells in the thymus and modulates T-cell repertoire in the periphery

    International Nuclear Information System (INIS)

    Prenatal exposure to diethylstilbestrol (DES) is known to cause altered immune functions and increased susceptibility to autoimmune disease in humans. In the current study, we investigated the effects of DES on T-cell differentiation in the thymus using the HY-TCR transgenic (Tg) mouse model in which the female mice exhibit positive selection of T cells bearing the Tg TCR, while the male mice show negative selection of such T cells. In female HY-TCR-Tg mice, exposure to DES showed more pronounced decrease in thymic cellularity when compared to male mice. Additionally, female mice also showed a significant decrease in the proportion of double-positive (DP) T cells in the thymus and HY-TCR-specific CD8+ T cells in the periphery. Male mice exhibiting negative selection also showed decreased thymic cellularity following DES exposure. Moreover, the male mice showed increased proportion of double-negative (DN) T cells in the thymus and decreased proportion of CD8+ T cells. The density of expression of HY-TCR on CD8+ cells was increased following DES exposure in both females and males. Finally, the proliferative response of thymocytes to mitogens and peripheral lymph node T cells to male H-Y antigen was significantly altered in female and male mice following DES treatment. Taken together, these data suggest that DES alters T-cell differentiation in the thymus by interfering with positive and negative selection processes, which in turn modulates the T-cell repertoire in the periphery

  7. File list: Unc.Bld.20.AllAg.CD4_CD8_double_positive_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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    Lifescience Database Archive (English)

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  1. Position of fuel cells in Italy; Situation des piles a combustible en Italie

    Energy Technology Data Exchange (ETDEWEB)

    Janot-Giorgetti, M.; Mottini, N.

    2000-02-01

    The main researches concerning the fuel cells in Italy are the PEFC (Polymer Electrolyte Fuel Cell) and the MCFC (Molten Carbonate Fuel Cell). This reports takes stock of these two techniques in Italy, explaining the running of these two types of cells and relating the Italian situation (development and research program, development programs of fuel cells vehicles). (O.M.)

  2. High level PHGDH expression in breast is predominantly associated with keratin 5-positive cell lineage independently of malignancy

    DEFF Research Database (Denmark)

    Gromova, Irina; Gromov, Pavel; Honma, Naoko; Kumar, Sudha; Rimm, David; Talman, Maj-Lis Møller; Wielenga, Vera Timmermans; Moreira, José

    2015-01-01

    showed high-level expression of Phgdh in normal CK5-positive mammary epithelial cells, indicating that expression of this protein was not associated with malignancy, but rather with cell lineage. However, proteomic profiling of Phgdh showed it to be expressed in two major protein forms, and that the...... in TNBC samples. One such protein was D-3-phosphoglycerate dehydrogenase (Phgdh), a candidate oncogene. We analysed expression of Phgdh in normal and TNBC mammary tissue samples by 2D gel-based proteomics and immunohistochemistry (IHC), and show here that high-level expression of Phgdh in mammary...... epithelial cells is primarily associated with cell lineage, as we found that Phgdh expression was predominant in CK5-positive cells, normal as well as malignant, thus identifying an association of this protein with the basal phenotype. Quantitative IHC analysis of Phgdh expression in normal breast tissue...

  3. A biocompatible microchip and methodology for efficiently trapping and positioning living cells into array based on negative dielectrophoresis

    Science.gov (United States)

    Guo, Xiaoliang; Zhu, Rong

    2015-06-01

    We present a microchip and trapping methodology based on negative dielectrophoresis (nDEP), whereby living cells were manipulated and positioned into an array with high trapping efficiency while maintaining good viability. The main factors that ensured good viability of cells were investigated including temperature of medium, extra transmembrane potential on cells, and electrolysis effect in DEP-based trapping. Optimum DEP conditions for the microchip were determined by considering both biocompatibility and trapping efficiency. Experiments demonstrated that under a voltage of 3.6-4 Vpp and at a frequency of 100 kHz, HeLa cells could be trapped and positioned into an array in less than 10 s while maintaining good viability. The normal adherence morphology and fluorescence of the cells, dyed with propidium iodide and Calcein-AM, were observed and verified the biocompatibility of the microchip and trapping methodology.

  4. 亲缘异基因造血干细胞移植联合甲磺酸伊马替尼治疗Ph阳性白血病%Hematopoietic stem cell transplantation in combination with imatinib mesylate for treatment of Philadelphia-positive leukemia

    Institute of Scientific and Technical Information of China (English)

    李梦醒; 王季石; 张燕; 孙志强; 赵鹏; 卢英豪

    2014-01-01

    BACKGROUND:Hematopoietic stem cel transplantation is recognized as the only method of curing Philadelphia-positive leukemia. Imatinib mesylate is a competitive inhibitor of the BCR/ABL tyrosine kinase, which has been used more and more in the treatment of Philadelphia-positive leukemia. OBJECTIVE:To observe the curative effect of imatinib mesylate combined with alogeneic hematopoietic stem cel transplantation for treatment of Philadelphia-positive leukemia. METHODS: We retrospectively analyzed the clinical effect of 12 patients with Philadelphia-positive leukemia who were treated with the combined therapy of imatinib mesylate and alogeneic hematopoietic stem cel transplantation from January 2011 to October 2012, and reviewed the relevant literatures. RESULTS AND CONCLUSION: Hematopoietic reconstitution was achieved in al 12 patients and the median times of neutrophil recovery and platelet recovery were 15 and 18 days, respectively. Of the 12 patients, 7 patients developed acute graft-versus-host disease II, 1 developed acute graft-versus-host disease III, 7 developed localized chronic graft-versus-host disease, and 3 developed extensive chronic graft-versus-host disease. Leukemia-free survival rate was 66.7%, and transplant-related mortality was 25.0%. The overal survival rate of HLA-matched sibling donors was 75.0%. The mean disease-free survival period was 8.5 months (7-17 months), and the time needed for BCR/ABL becoming negative was 2-5 months. As an effective and safe method for Philadelphia-positive leukemia, alogeneic hematopoietic stem cel transplantation combined with imatinib mesylate before and after transplantation reduces the leukemia cel load before transplantation, inhibits the proliferation of residual leukemia cels, and promotes ful chimerism change.%背景:造血干细胞移植是可以治愈Ph+白血病有效方法,甲磺酸伊马替尼是一种高度特异的酪氨酸激酶抑制剂,能抑制BCR/ABL酪氨酸激酶活性,在Ph+白血病

  5. HPV-positive HNSCC cell lines but not primary human fibroblasts are radiosensitized by the inhibition of Chk1

    International Nuclear Information System (INIS)

    Purpose: Despite the comparably high cure rates observed for HPV-positive HNSCC, there is still a great need for specific tumor radiosensitization due to the often severe side effects resulting from intense radiochemotherapy. We recently demonstrated that HPV-positive HNSCC cell lines are characterized by a defect in DNA double-strand break repair associated with a pronounced G2-arrest. Here we tested whether abrogation of this radiation-induced G2-arrest by the inhibition of Chk1 results in specific radiosensitization of HPV-positive HNSCC cells. Materials and methods: Experiments were performed with five HPV and p16-positive (93-VU-147T, UM-SCC-47, UT-SCC-45, UD-SCC-2, UPCI-SCC-154) and two HPV and p16-negative HNSCC cell lines, as well as two normal human fibroblast strains. Chk1 was inhibited by the selective inhibitor PF-00477736. Cell cycle distribution was determined by flow cytometry, Chk1-activity via Western blot and cell survival by colony formation assay. Results: With the exception of UPCI-SCC-154, the inhibition of Chk1 was found to abolish the pronounced radiation-induced G2-arrest in all HPV-positive cells utilized. All tumor cell lines that demonstrated the abrogation of G2-arrest also demonstrated radiosensitization. Notably, in G1-arrest-proficient normal human fibroblasts no radiosensitization was induced. Conclusion: Abrogation of the G2 checkpoint through the inhibition of Chk1 may be used to selectively increase the cellular radiosensitivity of HPV-positive HNSCC without affecting the surrounding normal tissue

  6. Antitumour effects of combined radio- and parvovirotherapy in N-Ras-positive tumour cells

    International Nuclear Information System (INIS)

    Many innovative approaches have attempted to challenge the dominant position of surgery, chemotherapy and radiotherapy as the major therapeutic modalities to treat cancer. However, only recently some new agents have appeared that achieve effects comparable to the ones of these established treatments. A novel class of such therapeutics consists of oncolytic viruses with several main representatives, namely adeno-, herpes, reo- and parvovirus. The latter class of anticancer agents has proven to be quite effective in many preclinical models and is now considered for a phase I clinical trial in patients with glioma and pancreatic cancer. An important question to be answered is whether virotherapy can be combined with standard treatments to achieve an enhanced antitumor effect still at a low level of toxicity. Here we present data concerning the application of irradiation together with H-1PV parvovirus infection comparing their toxic effects on normal and Ras -transformed fibroblasts. This proved to be a potent combination achieving high level of synergistic toxicity on transformed cells at doses, which did not affect the viability of normal human fibroblasts. This strategy holds promise to become a part of the clinical treatment of radioresistant tumours. (authors)

  7. Positive feedback loop between cancer stem cells and angiogenesis in hepatocellular carcinoma.

    Science.gov (United States)

    Yao, Hong; Liu, Nianli; Lin, Marie C; Zheng, Junnian

    2016-09-01

    Anti-angiogenesis-related therapies have become the standard care for patients with advanced hepatocellular carcinoma (HCC), as HCC is a highly vascularized solid tumor. Unfortunately, only modest and limited efficacies are observed. Emerging evidence have attributed to the limited efficacy to the presence of cancer stem cells (CSCs) in the tumor. CSCs predominantly drives angiogenesis via releasing proangiogenic factors and exosomes. They have the ability to resistant intratumoral hypoxia via autophagy or by directly forming the tubular structure to obtain blood. On the other hand, the vascular niche in tumor microenvironment also releases growth factors via juxtacrine and paracrine mechanisms to support the growth of CSCs and maintain its stemness features. This positive feedback loop between angiogenesis and CSCs exists in liver tumor microenvironment that is responsible for the development and poor prognosis of HCC. In this review, we summarize recent advances in our understanding of the crosstalks between angiogenesis and CSCs, and their interactions in liver tumor microenvironment and their purpose that an effective anti-angiogenic therapy should also target CSCs for HCC treatment. PMID:27108065

  8. Cronkhite-Canada syndrome polyps infiltrated with IgG4-positive plasma cells.

    Science.gov (United States)

    Fan, Ru-Ying; Wang, Xiao-Wei; Xue, Li-Jun; An, Ran; Sheng, Jian-Qiu

    2016-08-16

    Cronkhite-Canada syndrome (CCS) is a rare but serious protein-losing enteropathy, but little is known about the mechanism. Further more, misdiagnosis is common due to non-familiarity of its clinical manifestation. A 40-year-old male patient was admitted to our hospital because of diarrhea and hypogeusia associated with weight loss for 4 mo. On physical examination, skin pigmentation, dystrophic nail changes and alopecia were noted. He had no alike family history. Laboratory results revealed low levels of serum albumin (30.1 g/L, range: 35.0-55.0 g/L), serum potassium (2.61 mmol/L, range: 3.5-5.5 mmol/L) and blood glucose (2.6 mmol/L, range: 3.9-6.1 mmol/L). The erythrocyte sedimentation rate was elevated to 17 mm/h (range: 0-15 mm/h). X-ray of chest and mandible was normal. The endoscopic examination showed multiple sessile polyps in the stomach, small bowel and colorectum. Histopathologic examination of biopsies obtained from those polyps showed hyperplastic change, cystic dilatation and distortion of glands with inflammatory infiltration, eosinophilic predominance and stromal edema. Immune staining for IgG4 plasma cells was positive in polyps of stomach and colon. The patient was diagnosed of CCS and treated with steroid, he had a good response to steroid. Both histologic findings and treatment response to steroid suggested an autoimmune mechanism underling CCS. PMID:27574615

  9. Nestin-positive mesenchymal stem cells favour the astroglial lineage in neural progenitors and stem cells by releasing active BMP4

    Directory of Open Access Journals (Sweden)

    Leprince Pierre

    2004-09-01

    Full Text Available Abstract Background Spontaneous repair is limited after CNS injury or degeneration because neurogenesis and axonal regrowth rarely occur in the adult brain. As a result, cell transplantation has raised much interest as potential treatment for patients with CNS lesions. Several types of cells have been considered as candidates for such cell transplantation and replacement therapies. Foetal brain tissue has already been shown to have significant effects in patients with Parkinson's disease. Clinical use of the foetal brain tissue is, however, limited by ethical and technical problems as it requires high numbers of grafted foetal cells and immunosuppression. Alternatively, several reports suggested that mesenchymal stem cells, isolated from adult bone marrow, are multipotent cells and could be used in autograft approach for replacement therapies. Results In this study, we addressed the question of the possible influence of mesenchymal stem cells on neural stem cell fate. We have previously reported that adult rat mesenchymal stem cells are able to express nestin in defined culture conditions (in the absence of serum and after 25 cell population doublings and we report here that nestin-positive (but not nestin-negative mesenchymal stem cells are able to favour the astroglial lineage in neural progenitors and stem cells cultivated from embryonic striatum. The increase of the number of GFAP-positive cells is associated with a significant decrease of the number of Tuj1- and O4-positive cells. Using quantitative RT-PCR, we demonstrate that mesenchymal stem cells express LIF, CNTF, BMP2 and BMP4 mRNAs, four cytokines known to play a role in astroglial fate decision. In this model, BMP4 is responsible for the astroglial stimulation and oligodendroglial inhibition, as 1 this cytokine is present in a biologically-active form only in nestin-positive mesenchymal stem cells conditioned medium and 2 anti-BMP4 antibodies inhibit the nestin-positive mesenchymal

  10. Effects of cell positive cans and separators on the performance of high-voltage Li-ion batteries

    Science.gov (United States)

    Chen, Xilin; Xu, Wu; Xiao, Jie; Engelhard, Mark H.; Ding, Fei; Mei, Donghai; Hu, Dehong; Zhang, Jian; Zhang, Ji-Guang

    2012-09-01

    The effects of different cell positive cans and separators on first-cycle Coulombic efficiency and long-term cycling stability of a high-voltage spinel cathode are investigated systematically. Compared to stainless steel (SS) positive cans, aluminum (Al)-clad SS-316 positive cans are much more resistant to oxidation at high voltages; therefore, the initial Coulombic efficiency of the batteries with Al-clad can is improved by more than 13%. Among the five separators studied in this work, the polyethylene (PE) separator exhibits the best electrochemical stability. The cells using LiCr0.05Ni0.45Mn1.5O4 as the cathode, an Al-clad positive can, and a PE separator exhibits a first-cycle Coulombic efficiency of about 90% and a capacity fading of only 0.01% per cycle.

  11. Implications of a positive sentinel node in oral squamous cell carcinoma

    DEFF Research Database (Denmark)

    Gurney, Benjamin A S; Schilling, Clare; Putcha, Venkata;

    2012-01-01

    = 109) from 15 European centers, with cT1/2,N0 tumors, and a positive sentinel lymph node were identified. Kaplan-Meier and univariate and multivariate logistic regression analysis were used to identify variables that predicted for additional positive nodes and their position within the neck. RESULTS: A...

  12. Prognosis of patients with esophageal squamous cell carcinoma after esophagectomy using the log odds of positive lymph nodes

    OpenAIRE

    Wu, San-Gang; Sun, Jia-Yuan; Yang, Li-Chao; Zhou, Juan; Li, Feng-Yan; Li, Qun; Lin, Huan-xin; Lin, Qin; He, Zhen-Yu

    2015-01-01

    To compare the log odds of positive lymph nodes (LODDS) with the number of positive lymph nodes (pN), lymph node ratio (LNR), removed lymph node (RLN) count, and negative lymph node (NLN) count in determining the prognosis of patients with esophageal squamous cell carcinoma (ESCC) after esophagectomy. The records of patients with ESCC who received esophagectomy were retrospectively reviewed. The log-rank test was used to compare curves for overall survival (OS), and Cox regression analysis wa...

  13. Murine CD83-positive T cells mediate suppressor functions in vitro and in vivo.

    Science.gov (United States)

    Kreiser, Simon; Eckhardt, Jenny; Kuhnt, Christine; Stein, Marcello; Krzyzak, Lena; Seitz, Christine; Tucher, Christine; Knippertz, Ilka; Becker, Christoph; Günther, Claudia; Steinkasserer, Alexander; Lechmann, Matthias

    2015-02-01

    The CD83 molecule (CD83) is a well-known surface marker present on mature dendritic cells (mDC). In this study, we show that CD83 is also expressed on a subset of T cells which mediate regulatory T cell (Treg)-like suppressor functions in vitro and in vivo. Treg-associated molecules including CD25, cytotoxic T lymphocyte antigen-4 (CTLA-4), glucocorticoid-induced TNFR family-related gene (GITR), Helios and neuropilin-1 (NRP-1) as well as forkhead box protein 3 (FOXP3) were specifically expressed by these CD83(+) T cells. In contrast, CD83(-) T cells showed a naive T cell phenotype with effector T cell properties upon activation. Noteworthy, CD83(-) T cells were not able to upregulate CD83 despite activation. Furthermore, CD83(+) T cells suppressed the proliferation and inflammatory cytokine release of CD83(-) T cells in vitro. Strikingly, stimulated CD83(+) T cells released soluble CD83 (sCD83), which has been reported to possess immunosuppressive properties. In vivo, using the murine transfer colitis model we could show that CD83(+) T cells were able to suppress colitis symptoms while CD83(-) T cells possessed effector functions. In addition, this CD83 expression is also conserved on expanded human Treg. Thus, from these studies we conclude that CD83(+) T cells share important features with regulatory T cells, identifying CD83 as a novel lineage marker to discriminate between different T cell populations. PMID:25151500

  14. Controlled positioning of cells in biomaterials - approaches towards 3D tissue printing

    OpenAIRE

    Sandra Hofmann; Ralph Müller; Silke Wüst

    2011-01-01

    Current tissue engineering techniques have various drawbacks: they often incorporate uncontrolled and imprecise scaffold geometries, whereas the current conventional cell seeding techniques result mostly in random cell placement rather than uniform cell distribution. For the successful reconstruction of deficient tissue, new material engineering approaches have to be considered to overcome current limitations. An emerging method to produce complex biological products including cells or extrac...

  15. Enforced expression of GATA-3 during T cell development inhibits maturation of CD8 single-positive cells and induces thymic lymphoma in transgenic mice

    OpenAIRE

    Nawijn, Martijn,; Ferreira, Rita; Dingjan, Gemma; Kahre, O; Drabek, Dubravka; Karis, Alar; Hendriks, Rudi; Grosveld, Frank

    2001-01-01

    textabstractThe zinc finger transcription factor GATA-3 is of critical importance for early T cell development and commitment of Th2 cells. To study the role of GATA-3 in early T cell development, we analyzed and modified GATA-3 expression in vivo. In mice carrying a targeted insertion of a lacZ reporter on one allele, we found that GATA-3 transcription in CD4(+)CD8(+) double-positive thymocytes correlated with the onset of positive selection events, i.e., TCRalphabeta up-regulation and CD69 ...

  16. Preferential antitumor effect of the Src inhibitor dasatinib associated with a decreased proportion of aldehyde dehydrogenase 1-positive cells in breast cancer cells of the basal B subtype

    Directory of Open Access Journals (Sweden)

    Watanabe Mika

    2010-10-01

    Full Text Available Abstract Background Recent studies have suggested that the Src inhibitor dasatinib preferentially inhibits the growth of breast cancer cells of the basal-like subtype. To clarify this finding and further investigate combined antitumor effects of dasatinib with cytotoxic agents, a panel of breast cancer cell lines of various subtypes was treated with dasatinib and/or chemotherapeutic agents. Methods Seven human breast cancer cell lines were treated with dasatinib and/or seven chemotherapeutic agents. Effects of the treatments on c-Src activation, cell growth, cell cycle, apoptosis and the proportion of aldehyde dehydrogenase (ALDH 1-positive cells were examined. Results The 50%-growth inhibitory concentrations (IC50s of dasatinib were much lower in two basal B cell lines than those in the other cell lines. The IC50s of chemotherapeutic agents were not substantially different among the cell lines. Dasatinib enhanced antitumor activity of etoposide in the basal B cell lines. Dasatinib induced a G1-S blockade with a slight apoptosis, and a combined treatment of dasatinib with etoposide also induced a G1-S blockade in the basal B cell lines. Dasatinib decreased the expression levels of phosphorylated Src in all cell lines. Interestingly, dasatinib significantly decreased the proportion of ALDH1-positive cells in the basal B cell lines but not in the other cell lines. Conclusions The present study indicates that dasatinib preferentially inhibits the growth of breast cancer cells of the basal B subtype associated with a significant loss of putative cancer stem cell population. A combined use of dasatinib with etoposide additively inhibits their growth. Further studies targeting breast cancers of the basal B subtype using dasatinib with cytotoxic agents are warranted.

  17. Therapeutic Potential of Adipose-Derived SSEA-3-Positive Muse Cells for Treating Diabetic Skin Ulcers

    OpenAIRE

    Kinoshita, Kahori; Kuno, Shinichiro; Ishimine, Hisako; Aoi, Noriyuki; Mineda, Kazuhide; Kato, Harunosuke; Doi, Kentaro; Kanayama, Koji; Feng, Jingwei; Mashiko, Takanobu; Kurisaki, Akira; Yoshimura, Kotaro

    2015-01-01

    Refractory skin ulcers were generated in severe combined immunodeficiency (SCID) mice with type 1 diabetes and delayed wound healing compared with nondiabetic SCID mice. Treatment with a multilineage differentiating stress-enduring (Muse)-rich cell population significantly accelerated wound healing compared with the Muse-poor cell population, and these cells be achieved in large amounts with minimal morbidity. Adipose-derived Muse cells could be a practical tool for a variety of stem cell-dep...

  18. Insulin-positive, Glut2-low cells present within mouse pancreas exhibit lineage plasticity and are enriched within extra-islet endocrine cell clusters.

    Science.gov (United States)

    Beamish, Christine A; Strutt, Brenda J; Arany, Edith J; Hill, David J

    2016-04-18

    Regeneration of insulin-producing β-cells from resident pancreas progenitors requires an understanding of both progenitor identity and lineage plasticity. One model suggested that a rare β-cell sub-population within islets demonstrated multi-lineage plasticity. We hypothesized that β-cells from young mice (postnatal day 7, P7) exhibit such plasticity and used a model of islet dedifferentiation toward a ductal epithelial-cell phenotype to test this theory. RIPCre;Z/AP(+/+) mice were used to lineage trace the fate of β-cells during dedifferentiation culture by a human placental alkaline phosphatase (HPAP) reporter. There was a significant loss of HPAP-expressing β-cells in culture, but remaining HPAP(+) cells lost insulin expression while gaining expression of the epithelial duct cell marker cytokeratin-19 (Ck19). Flow cytometry and recovery of β-cell subpopulations from whole pancreas vs. islets suggest that the HPAP(+)Ck19(+) cells had derived from insulin-positive, glucose-transporter-2-low (Ins(+)Glut2(LO)) cells, representing 3.5% of all insulin-expressing cells. The majority of these cells were found outside of islets within clusters of <5 β-cells. These insulin(+)Glut2(LO) cells demonstrated a greater proliferation rate in vivo and in vitro as compared to insulin(+)Glut2(+) cells at P7, were retained into adulthood, and a subset differentiated into endocrine, ductal, and neural lineages, illustrating substantial plasticity. Results were confirmed using RIPCre;ROSA- eYFP mice. Quantitative PCR data indicated these cells possess an immature β-cell phenotype. These Ins(+)Glut2(LO) cells may represent a resident population of cells capable of forming new, functional β-cells, and which may be potentially exploited for regenerative therapies in the future. PMID:27010375

  19. Proliferation of Estrogen Receptor alpha Positive Mammary Epithelial Cells is Restrained by TGFbeta1 in Adult Mice

    Energy Technology Data Exchange (ETDEWEB)

    Ewan, Kenneth B.R.; Oketch-Rabah, Hellen A.; Ravani, Shraddha A.; Shyamala, G.; Moses, Harold L.; Barcellos-Hoff, Mary Helen

    2005-03-03

    Transforming growth factor {beta}1 (TGF{beta}1) is a potent inhibitor of mammary epithelial proliferation. In human breast, estrogen receptor {alpha} (ER{alpha}) cells rarely co-localize with markers of proliferation, but their increased frequency correlates with breast cancer risk. To determine whether TGF{beta}1 is necessary for the quiescence of ER{alpha}-positive population, we examined mouse mammary epithelial gland at estrus. Approximately 35% of cells showed TGF{beta}1 activation, which co-localized with nuclear receptor-phosphorylated Smad 2/3, indicating that TGF{beta} signaling is autocrine. Furthermore, nuclear Smad co-localized with nuclear ER{alpha}. To test whether TGF{beta} was functional, we examined genetically engineered mice with different levels of TGF{beta}1. ER{alpha} co-localization with markers of proliferation (i.e. Ki-67 or BrdU) at estrus was significantly increased in the mammary glands of Tgf{beta}1 C57/bl/129SV heterozygote mice. This relationship was maintained following pregnancy, but was absent at puberty. Conversely, mammary epithelial expression of constitutively active TGF{beta}1 via the MMTV promoter suppressed proliferation of ER{alpha} positive cells. Thus, TGF{beta}1 activation functionally restrains ER{alpha} positive cells from proliferating in adult mammary gland. Accordingly, we propose that TGF{beta}1 dysregulation may promote proliferation of ER{alpha} positive cells associated with breast cancer risk in humans.

  20. Short-term environmental enrichment exposure induces proliferation and maturation of doublecortin-positive cells in the prefrontal cortex

    Institute of Scientific and Technical Information of China (English)

    Chunling Fan; Mengqi Zhang; Lei Shang; Ngobe Akume Cynthia; Zhi Li; Zhenyu Yang; Dan Chen; Jufang Huang; Kun Xiong

    2014-01-01

    Previous studies have demonstrated that doublecortin-positive immature neurons exist pre-dominantly in the superficial layer of the cerebral cortex of adult mammals such as guinea pigs, and these neurons exhibit very weak properties of self-proliferation during adulthood under physiological conditions. To verify whether environmental enrichment has an impact on the proliferation and maturation of these immature neurons in the prefrontal cortex of adult guinea pigs, healthy adult guinea pigs were subjected to short-term environmental enrichment. Animals were allowed to play with various cognitive and physical stimulating objects over a period of 2 weeks, twice per day, for 60 minutes each. Immunolfuorescence staining results indicated that the number of doublecortin-positive cells in layer II of the prefrontal cortex was signiifcantly increased after short-term environmental enrichment exposure. In addition, these doublecortin-positive cells co-expressed 5-bromo-2-deoxyuridine (a marker of cell prolifera-tion), c-Fos (a marker of cell viability) and NeuN (a marker of mature neurons). Experimental ifndings showed that short-term environmental enrichment can induce proliferation, activation and maturation of doublecortin-positive cells in layer II of the prefrontal cortex of adult guinea pigs.

  1. Gastrin: a distinct fate of neurogenin3 positive progenitor cells in the embryonic pancreas.

    Directory of Open Access Journals (Sweden)

    Yaron Suissa

    Full Text Available Neurogenin3(+ (Ngn3(+ progenitor cells in the developing pancreas give rise to five endocrine cell types secreting insulin, glucagon, somatostatin, pancreatic polypeptide and ghrelin. Gastrin is a hormone produced primarily by G-cells in the stomach, where it functions to stimulate acid secretion by gastric parietal cells. Gastrin is expressed in the embryonic pancreas and is common in islet cell tumors, but the lineage and regulators of pancreatic gastrin(+ cells are not known. We report that gastrin is abundantly expressed in the embryonic pancreas and disappears soon after birth. Some gastrin(+ cells in the developing pancreas co-express glucagon, ghrelin or pancreatic polypeptide, but many gastrin(+ cells do not express any other islet hormone. Pancreatic gastrin(+ cells express the transcription factors Nkx6.1, Nkx2.2 and low levels of Pdx1, and derive from Ngn3(+ endocrine progenitor cells as shown by genetic lineage tracing. Using mice deficient for key transcription factors we show that gastrin expression depends on Ngn3, Nkx2.2, NeuroD1 and Arx, but not Pax4 or Pax6. Finally, gastrin expression is induced upon differentiation of human embryonic stem cells to pancreatic endocrine cells expressing insulin. Thus, gastrin(+ cells are a distinct endocrine cell type in the pancreas and an alternative fate of Ngn3+ cells.

  2. Fluorescence in situ hybridization and immunohistochemistry as diagnostic methods for ALK positive non-small cell lung cancer patients.

    Directory of Open Access Journals (Sweden)

    Pablo Martinez

    Full Text Available BACKGROUND: Anaplastic Lymphoma Kinase (ALK positivity represents a novel molecular target in a subset of Non-Small Cell Lung Cancers (NSCLC. We explore Fluorescence in situ Hybridization (FISH and Immunohistochemistry (IHC as diagnostic methods for ALK positive patients and to describe its prevalence and outcomes in a population of NSCLC patients. METHODS: NSCLC patients previously screened for Epidermal Growth Factor Receptor (EGFR at our institution were selected. ALK positive patients were identified by FISH and the value of IHC (D5F3 was explored. RESULTS: ninety-nine patients were identified. Median age was 61.5 years (range 35-83, all were caucasians, eighty percent were adenocarcinomas, fifty-one percent were male and thirty-eight percent were current smokers. Seven (7.1% patients were ALK positive by FISH, thirteen (13.1% were EGFR mutant, and 65 (65.6% were negative/Wild Type (WT for both ALK and EGFR. ALK positivity and EGFR mutations were mutually exclusive. ALK positive patients tend to be younger than EGFR mutated or wt patients. ALK positive patients were predominantly never smokers (71.4% and adenocarcinoma (71.4%. ALK positive and EGFR mutant patients have a better outcome than negative/WT. All patients with ALK FISH negative tumours were negative for ALK IHC. Out of 6 patients positive for ALK FISH with more tissue available, 5 were positive for ALK IHC and 1 negative. CONCLUSIONS: ALK positive patients represent 7.1% of a population of selected NSCLC. ALK positive patients have different clinical features and a better outcome than EGFR WT and ALK negative patients. IHC is a promising method for detecting ALK positive NSCLC patients.

  3. BIM mediates synergistic killing of B-cell acute lymphoblastic leukemia cells by BCL-2 and MEK inhibitors.

    Science.gov (United States)

    Korfi, K; Smith, M; Swan, J; Somervaille, T C P; Dhomen, N; Marais, R

    2016-01-01

    B-cell acute lymphoblastic leukemia (B-ALL) is an aggressive hematological disease that kills ~50% of adult patients. With the exception of some BCR-ABL1(+) patients who benefit from tyrosine kinase inhibitors, there are no effective targeted therapies for adult B-ALL patients and chemotherapy remains first-line therapy despite adverse side effects and poor efficacy. We show that, although the MEK/ERK pathway is activated in B-ALL cells driven by different oncogenes, MEK inhibition does not suppress B-ALL cell growth. However, MEK inhibition synergized with BCL-2/BCL-XL family inhibitors to suppress proliferation and induce apoptosis in B-ALL cells. We show that this synergism is mediated by the pro-apoptotic factor BIM, which is dephosphorylated as a result of MEK inhibition, allowing it to bind to and neutralize MCL-1, thereby enhancing BCL-2/BCL-XL inhibitor-induced cell death. This cooperative effect is observed in B-ALL cells driven by a range of genetic abnormalities and therefore has significant therapeutic potential. PMID:27054332

  4. Different distribution of c-kit positive interstitial cells of Cajal-like in children’s urinary bladders

    OpenAIRE

    Marian Danilewicz; Andrzej Kulig; Monika Dzieniecka; Anna Małgorzata Piaseczna-Piotrowska; Andrzej Chilarski

    2011-01-01

    We describe the presence of c-kit positive interstitial cells of Cajal-like (ICCs-like) in the walls of the urinary bladders of children. An immunohistochemical study of specimens, obtained at autopsy from either the trigonum (Group A) or the corpus (Group B), was performed using antibodies against c-kit (CD 117). Histological morphometry of the immunoexpression of c-kit positive ICCs-like was performed by means of image analysis system. The c-kit positive ICCs-like were identified by their m...

  5. Different distribution of c-kit positive interstitial cells of Cajal-like in children’s urinary bladders

    OpenAIRE

    Anna Małgorzata Piaseczna-Piotrowska; Monika Dzieniecka; Andrzej Kulig; Marian Danilewicz; Andrzej Chilarski

    2011-01-01

    We describe the presence of c-kit positive interstitial cells of Cajal-like (ICCs-like) in the walls of theurinary bladders of children. An immunohistochemical study of specimens, obtained at autopsy from either thetrigonum (Group A) or the corpus (Group B), was performed using antibodies against c-kit (CD 117). Histologicalmorphometry of the immunoexpression of c-kit positive ICCs-like was performed by means of image analysissystem. The c-kit positive ICCs-like were identified by their morph...

  6. Gene expression profiling of CD133-positive cells in coronary artery disease.

    Science.gov (United States)

    Li, Jiayu; Zhou, Changyu; Li, Jiarui; Wan, Yingchun; Li, Tao; Ma, Piyong; Wang, Yingjian; Sang, Haiyan

    2015-11-01

    Gene expression profiles of CD133-positive cells from patients with coronary artery disease (CAD) were analyzed to identify key genes associated with cardiac therapy. Furthermore, the effect of exercise on gene expression was also investigated. Gene expression data set (accession number: GSE18608) was downloaded from the Gene Expression Omnibus, including blood samples from four healthy subjects (H), and from 10 patients with coronary artery disease at baseline (B) and after 3 months (3M) of exercise. Differential analysis was performed for H vs. B and H vs. 3M using limma package of R. Two‑way cluster analysis was performed using the expression levels of the differentially expressed genes (DEGs) by package pheatmap of R. Functional enrichment analysis was applied on the DEGs using the Database for Annotation, Visualization and Integrated Discovery. Relevant small molecules were predicted using the Connectivity map database (cMap). A total of 131 and 71 DEGs were identified in patients with CAD prior to and following 3 months of exercise. The two groups of DEGs were compared and 44 genes overlapped. In cluster analysis with the expression levels of the common DEGs, patients with CAD could be well separated from the healthy controls. Functional enrichment analysis showed that response to peptide hormone stimulus and anti‑apoptosis pathways were significantly enriched in the common DEGs. A total of 12 relevant small molecules were revealed by cMap based upon the expression levels of common DEGs, such as 5252917 and MG‑262. Three months of exercise in part normalized the gene expression in CAD patients. The genes not altered by exercise may be the targets of small molecules, such as 5252917 and MG-262. PMID:26458356

  7. CD44 expression positively correlates with Foxp3 expression and suppressive function of CD4+ Treg cells

    Directory of Open Access Journals (Sweden)

    König Rolf

    2009-10-01

    Full Text Available Abstract Background CD4+CD25+ regulatory T (Treg cells develop in the thymus and can suppress T cell proliferation, modulated by Foxp3 and cytokines; however, the relevance of CD44 in Treg cell development is less clear. To address this issue, we analyzed Foxp3 expression in CD44+ Treg cells by using multiple parameters, measured the levels of the immunoregulatory cytokine interleukin (IL-10 in various thymocyte subsets, and determined the suppressor activity in different splenic Treg cell populations. Results Within mouse thymocytes, we detected Treg cells with two novel phenotypes, namely the CD4+CD8-CD25+CD44+ and CD4+CD8-CD25+CD44- staining features. Additional multi-parameter analyses at the single-cell and molecular levels suggested to us that CD44 expression positively correlated with Foxp3 expression in thymocytes, the production of IL-10, and Treg activity in splenic CD4+CD25+ T cells. This suppressive effect of Treg cells on T cell proliferation could be blocked by using anti-IL-10 neutralizing antibodies. In addition, CD4+CD25+CD44+ Treg cells expressed higher levels of IL-10 and were more potent in suppressing effector T cell proliferation than were CD4+CD25+CD44- cells. Conclusion This study indicates the presence of two novel phenotypes of Treg cells in the thymus, the functional relevance of CD44 in defining Treg cell subsets, and the role of both IL-10 and Foxp3 in modulating the function of Treg cells. Reviewers This article was reviewed by Dr. M. Lenardo, Dr. L. Klein & G. Wirnsberger (nominated by Dr. JC Zungia-Pfluker, and Dr. E.M. Shevach.

  8. Immunocapture of CD133-positive cells from human cancer cell lines by using monodisperse magnetic poly(glycidyl methacrylate) microspheres containing amino groups

    International Nuclear Information System (INIS)

    Magnetic poly(glycidyl methacrylate)-based macroporous microspheres with an average particle size of 4.2 μm were prepared using a modified multi-step swelling polymerization method and by introducing amino functionality on their surfaces. Antibody molecules were oxidized on their carbohydrate moieties and bound to the amino-containing magnetic microspheres via a site-directed procedure. CD133-positive cells could be effectively captured from human cancer cell lines (HepG2, HCT116, MCF7, and IMR-32) by using magnetic microspheres conjugated to an anti-human CD133 antibody. After further culture, the immunocaptured CD133-expressing cells from IMR-32 proliferated and gradually detached from the magnetic microspheres. Flow-cytometric analysis confirmed the enrichment of CD133-expressing cells by using the antibody-bound magnetic microspheres. Such microspheres suitable for immunocapture are very promising for cancer diagnosis because the CD133-expressing cells in cancer cell lines have been suggested to be cancer stem cells. - Highlights: • Multi-step swelling polymerization produced poly(glycidyl methacrylate) microspheres. • Anti-human CD133 antibodies were bound to the amino-containing magnetic microspheres. • CD133-positive cells were effectively captured from human cancer cell lines. • Immunocaptured CD133-expressing cells proliferated and were detached from microspheres. • Enrichment of CD133-expressing cells was confirmed by flow-cytometric analysis

  9. Different distribution of c-kit positive interstitial cells of Cajal-like in children’s urinary bladders

    Directory of Open Access Journals (Sweden)

    Marian Danilewicz

    2011-10-01

    Full Text Available We describe the presence of c-kit positive interstitial cells of Cajal-like (ICCs-like in the walls of the urinary bladders of children. An immunohistochemical study of specimens, obtained at autopsy from either the trigonum (Group A or the corpus (Group B, was performed using antibodies against c-kit (CD 117. Histological morphometry of the immunoexpression of c-kit positive ICCs-like was performed by means of image analysis system. The c-kit positive ICCs-like were identified by their morphology and counted in the vesical muscle layer in ten adjacent high power fields, each of 0.0479 mm2. The areas of the epithelial and subepithelial layers containing c-kit positive mast cells (rounded body with no dendritic processes were neglected. The results were expressed as the number of ICCs-like cells per mm2. Differences between groups were tested using unpaired Student’s t-test preceded by evaluation of normality and Levene’s test. Results were considered statistically significant if p < 0.05. In Group A, the mean number of ICCs-like cells was statistically significantly higher (41.5 cells/mm2 than in Group B (30.4 cells/mm2, p < 0.05. ICCs-like cells were found within the smooth muscle layer of the urinary bladder. There was a different distribution of these cells in particular parts of the bladder, which was probably due to the different roles of the trigonum and the corpus in the bladders of children. (Folia Histochemica et Cytobiologica 2011; Vol. 49, No. 3, pp. 431–435

  10. Cell-wall determinants of the bactericidal action of group IIA phospholipase A2 against Gram-positive bacteria

    OpenAIRE

    Foreman-Wykert, Amy K.; Weinrauch, Yvette; Elsbach, Peter; Weiss, Jerrold

    1999-01-01

    We have shown previously that a group IIA phospholipase A2 (PLA2) is responsible for the potent bactericidal activity of inflammatory fluids against many Gram-positive bacteria. To exert its antibacterial activity, this PLA2 must first bind and traverse the bacterial cell wall to produce the extensive degradation of membrane phospholipids (PL) required for bacterial killing. In this study, we have examined the properties of the cell-wall that may determine the potency of group IIA PLA2 action...

  11. Blinded search for varicella zoster virus in giant cell arteritis (GCA)-positive and GCA-negative temporal arteries.

    Science.gov (United States)

    Gilden, Don; White, Teresa; Khmeleva, Nelly; Katz, Bradley J; Nagel, Maria A

    2016-05-15

    Recent analysis of archived temporal arteries (TAs) acquired from 13 pathology laboratories in the US, Canada, Iceland, France, Germany and Israel from patients with pathologically-verified giant cell arteritis (GCA-positive) and TAs from patients with clinical features and laboratory abnormalities of GCA but whose TAs were pathologically negative (GCA-negative) revealed VZV antigen in most TAs from both groups. Despite formalin-fixation, VZV DNA was also found in many VZV-antigen positive sections that were scraped, subjected to DNA extraction, and examined by PCR with VZV-specific primers. Importantly, in past studies, the pathological diagnosis (GCA-positive or -negative) was known to the neurovirology laboratory. Herein, GCA-positive and GCA-negative TAs were provided by an outside institution and examined by 4 investigators blinded to the pathological diagnoses. VZV antigen was found in 3/3 GCA-positive TAs and in 4/6 GCA-negative TAs, and VZV DNA in 1/3 VZV antigen-positive, GCA-positive TAs and in 3/4 VZV antigen-positive, GCA-negative TAs. VZV DNA was also detected in one GCA-negative, VZV-antigen negative TA. Overall, the detection of VZV antigen in 78% of GCA-positive and GCA-negative TAs is consistent with previous reports on the prevalence of VZV antigen in patients with clinically suspect GCA. PMID:27084233

  12. Tracking of [18F]FDG-labeled natural killer cells to HER2/neu-positive tumors

    International Nuclear Information System (INIS)

    Introduction: The objective of this study was to label the human natural killer (NK) cell line NK-92 with [18F]fluoro-deoxy-glucose (FDG) for subsequent in vivo tracking to HER2/neu-positive tumors. Methods: NK-92 cells were genetically modified to NK-92-scFv(FRP5)-zeta cells, which express a chimeric antigen receptor that is specific to the tumor-associated ErbB2 (HER2/neu) antigen. NK-92 and NK-92-scFv(FRP5)-zeta cells were labeled with [18F]FDG by simple incubation at different settings. Labeling efficiency was evaluated by a gamma counter. Subsequently, [18F]FDG-labeled parental NK-92 or NK-92-scFv(FRP5)-zeta cells were intravenously injected into mice with implanted HER2/neu-positive NIH/3T3 tumors. Radioactivity in tumors was quantified by digital autoradiography and correlated with histopathology. Results: The NK-92 and NK-92-scFv(FRP5)-zeta cells could be efficiently labeled with [18F]FDG by simple incubation. Optimal labeling efficiencies (80%) were achieved using an incubation period of 60 min and additional insulin (10 IU/ml). After injection of 5x106 [18F]FDG-labeled NK-92-scFv(FRP5)-zeta cells into tumor-bearing mice, digital autoradiography showed an increased uptake of radioactivity in HER2/neu-positive tumors at 60 min postinjection. Conversely, injection of 5x106 NK-92 cells not directed against HER2/neu receptors did not result in increased uptake of radioactivity in the tumors. Histopathology confirmed an accumulation of the NK-92-scFv(FRP5)-zeta cells, but not the parental NK cells, in tumor tissues. Conclusion: The human NK cell line NK-92 can be directed against HER2/neu antigens by genetic modification. The genetically modified NK cells can be efficiently labeled with [18F]FDG, and the accumulation of these labeled NK cells in HER2/neu-positive tumors can be monitored with autoradiography

  13. CXCR4-positive subset of glioma is enriched for cancer stem cells.

    Science.gov (United States)

    Zheng, Xuesheng; Xie, QingSong; Li, Shiting; Zhang, Wenchuan

    2011-01-01

    CXC chemokine receptor 4 (CXCR4) is a cell surface molecule expressed in a distinct subset of glioma cells with enhanced tumorigenicity, and it is related to many important biological activities of the tumor. We supposed that this receptor might be a cell surface "marker" for glioma stem cells. This hypothesis was tested both in vitro and in vivo. The CXCR4+ and CXCR4- subsets were sorted from three human malignant glioma specimens. They were tested for the capability of colony formation in soft agar, generation of tumorosphere, self-renewal, and multipotent differentiation in vitro, and the capability of xenograft tumor in vivo. Drug and radiation resistance and coexpression with CD133 were studied for each subset. CXCR4+ glioma cells, but not CXCR4- cells, were capable of generating tumorospheres in serum-free medium. In addition, these spheres were able to self-renew after passage, and had multipotent differentiation after being induced in serum-containing medium. In soft agar assay, CXCR4+ cells generate much more colonies. The animal experiment revealed that CXCR4+ subpopulation had stronger tumorigenicity than the unsorted parental glioma cells, while the CXCR4- cells did not generate xenograft tumor. CXCR4-possitive cells were more resistant to temozolomide and radiation treatment. Both CXCR4+ and CXCR4- subsets contained very few CD133+ cells. The CXCR4+ subsets of glioma cells fulfill the standard of "cancer stem cell". PMID:22812188

  14. Nck adaptors are positive regulators of the size and sensitivity of the T-cell repertoire.

    Science.gov (United States)

    Roy, Edwige; Togbe, Dieudonnée; Holdorf, Amy D; Trubetskoy, Dmitry; Nabti, Sabrina; Küblbeck, Günter; Klevenz, Alexandra; Kopp-Schneider, Annette; Leithäuser, Frank; Möller, Peter; Bladt, Friedhelm; Hämmerling, Günter; Arnold, Bernd; Pawson, Tony; Tafuri, Anna

    2010-08-31

    The size and sensitivity of the T-cell repertoire governs the effectiveness of immune responses against invading pathogens. Both are modulated by T-cell receptor (TCR) activity through molecular mechanisms, which remain unclear. Here, we provide genetic evidence that the SH2/SH3 domain containing proteins Nck lower the threshold of T-cell responsiveness. The hallmarks of Nck deletion were T-cell lymphopenia and hyporeactivity to TCR-mediated stimulation. In the absence of the Nck adaptors, peripheral T cells expressing a TCR with low avidity for self-antigens were strongly reduced, whereas an overall impairment of T-cell activation by weak antigenic stimulation was observed. Mechanistically, Nck deletion resulted in a significant decrease in calcium mobilization and ERK phosphorylation upon TCR engagement. Taken together, our findings unveil a crucial role for the Nck adaptors in shaping the T-cell repertoire to ensure maximal antigenic coverage and optimal T cell excitability. PMID:20709959

  15. Probing interaction of Gram-positive and Gram-negative bacterial cells with ZnO nanorods

    Energy Technology Data Exchange (ETDEWEB)

    Jain, Aanchal; Bhargava, Richa; Poddar, Pankaj, E-mail: p.poddar@ncl.res.in

    2013-04-01

    In the present work, the physiological effects of the ZnO nanorods on the Gram positive (Staphylococcus aureus and Bacillus subtilis) and Gram-negative (Escherichia coli and Aerobacter aerogenes) bacterial cells have been studied. The analysis of bacterial growth curves for various concentrations of ZnO nanorods indicates that Gram positive and Gram negative bacterial cells show inhibition at concentrations of ∼ 64 and ∼ 256 μg/mL respectively. The marked difference in susceptibility towards nanorods was also validated by spread plate and disk diffusion methods. In addition, the scanning electron micrographs show a clear damage to the cells via changed morphology of the cells from rod to coccoid etc. The confocal optical microscopy images of these cells also demonstrate the reduction in live cell count in the presence of ZnO nanorods. These, results clearly indicate that the antibacterial activity of ZnO nanorods is higher towards Gram positive bacterium than Gram negative bacterium which indicates that the structure of the cell wall might play a major role in the interaction with nanostructured materials and shows high sensitivity to the particle concentration. Highlights: ► Effect of ZnO nanorods on the growth cycles of four bacterial strains. ► A relation has been established between growth rate of bacteria and concentration. ► Serious damage in the morphology of bacterial cells in the presence of ZnO nanorods. ► Microscopic studies to see the time dependent effect on bacterial cells.

  16. CellSense: A Probabilistic RSSI-based GSM Positioning System

    OpenAIRE

    Ibrahim, Mohamed; Youssef, Moustafa

    2010-01-01

    Context-aware applications have been gaining huge interest in the last few years. With cell phones becoming ubiquitous computing devices, cell phone localization has become an important research problem. In this paper, we present CellSense, a prob- abilistic RSSI-based fingerprinting location determina- tion system for GSM phones.We discuss the challenges of implementing a probabilistic fingerprinting local- ization technique in GSM networks and present the details of the CellSense system and...

  17. Nck adaptors are positive regulators of the size and sensitivity of the T-cell repertoire

    OpenAIRE

    Roy, Edwige; Togbe, Dieudonnée; Holdorf, Amy D.; Trubetskoy, Dmitry; Nabti, Sabrina; Küblbeck, Günter; Klevenz, Alexandra; Kopp-Schneider, Annette; Leithäuser, Frank; Möller, Peter; Bladt, Friedhelm; Hämmerling, Günter; Arnold, Bernd; Pawson, Tony; Tafuri, Anna

    2010-01-01

    The size and sensitivity of the T-cell repertoire governs the effectiveness of immune responses against invading pathogens. Both are modulated by T-cell receptor (TCR) activity through molecular mechanisms, which remain unclear. Here, we provide genetic evidence that the SH2/SH3 domain containing proteins Nck lower the threshold of T-cell responsiveness. The hallmarks of Nck deletion were T-cell lymphopenia and hyporeactivity to TCR-mediated stimulation. In the absence of the Nck adaptors, pe...

  18. CellSense: An Accurate Energy-Efficient GSM Positioning System

    OpenAIRE

    Ibrahim, Mohamed; Youssef, Moustafa

    2011-01-01

    Context-aware applications have been gaining huge interest in the last few years. With cell phones becoming ubiquitous computing devices, cell phone localization has become an important research problem. In this paper, we present CellSense, a prob- abilistic RSSI-based fingerprinting location determi- nation system for GSM phones. We discuss the chal- lenges of implementing a probabilistic fingerprinting localization technique in GSM networks and present the details of the CellSense systemand...

  19. Myeloid-derived suppressor cells in Chronic myeloid leukemia

    Directory of Open Access Journals (Sweden)

    Cesarina eGiallongo

    2015-05-01

    Full Text Available The suppression of the immune system create a permissive environment for development and progression of cancer. One population of immunosuppressive cells that have become the focus of intense study is myeloid derived suppressor cells (MDSCs, immature myeloid cells able to induce immune-escape, angiogenesis and tumor progression. Two different subpopulations have been identified and studied: granulocytic and monocytic MDSCs, with a different immunophenotype and immunosuppressive properties. Recently, an accumulation of both Gr-MDSCs and Mo-MDSCs cells has been found in the peripheral blood of chronic myeloid leukemia (CML patients. They are part of the tumor clone showing BCR/ABL expression. Imatinib therapy decreases both MDSCs and arginase 1 levels to normal ones. This review will focus on actual knowledge for human MDSCs and their immunosuppressive activity in CML patients with a critical attention to comparison of Gr-MDSCs and polymorphonuclear cells (PMNs. We will then suggest the monitoring of MDSCs in patients who have discontinued tyrosine kinase inhibitors (TKIs therapy to evaluate if their increase could correlate with disease relapse.

  20. CXCR3-dependent recruitment and CCR6-mediated positioning of Th-17 cells in the inflamed liver

    OpenAIRE

    Oo, Ye Htun; Banz, Vanessa; Kavanagh, Dean; Liaskou, Evaggelia; Withers, David R; Humphreys, Elizabeth; Reynolds, Gary M; Lee-Turner, Laura; Kalia, Neena; Hubscher, Stefan G.; Klenerman, Paul; Eksteen, Bertus; Adams, David H

    2012-01-01

    Background & Aims IL-17 secreting CD4 (Th17) and CD8 (Tc17) T cells have been implicated in immune-mediated liver diseases, but the molecular basis for their recruitment and positioning within the liver is unknown. Methods The phenotype and migratory behaviour of human liver-derived Th17 and Tc17 cells were investigated by flow cytometry and chemotaxis and flow-based adhesion assays. The recruitment of murine Th17 cells to the liver was studied in vivo using intra-vital microscopy. Results IL...