WorldWideScience

Sample records for anti-cancer target gene

  1. Mitochondrial chaperones may be targets for anti-cancer drugs

    Science.gov (United States)

    Scientists at NCI have found that a mitochondrial chaperone protein, TRAP1, may act indirectly as a tumor suppressor as well as a novel target for developing anti-cancer drugs. Chaperone proteins, such as TRAP1, help other proteins adapt to stress, but sc

  2. Anti-cancer chalcones: Structural and molecular target perspectives.

    Science.gov (United States)

    Mahapatra, Debarshi Kar; Bharti, Sanjay Kumar; Asati, Vivek

    2015-06-15

    Chalcone or (E)-1,3-diphenyl-2-propene-1-one scaffold remained a fascination among researchers in the 21st century due to its simple chemistry, ease of synthesis and a wide variety of promising biological activities. Several natural and (semi) synthetic chalcones have shown anti-cancer activity due to their inhibitory potential against various targets namely ABCG2/P-gp/BCRP, 5α-reductase, aromatase, 17-β-hydroxysteroid dehydrogenase, HDAC/Situin-1, proteasome, VEGF, VEGFR-2 kinase, MMP-2/9, JAK/STAT signaling pathways, CDC25B, tubulin, cathepsin-K, topoisomerase-II, Wnt, NF-κB, B-Raf and mTOR etc. In this review, a comprehensive study on molecular targets/pathways involved in carcinogenesis, mechanism of actions (MOAs), structure activity relationships (SARs) and patents granted have been highlighted. With the knowledge of molecular targets, structural insights and SARs, this review may be helpful for (medicinal) chemists to design more potent, safe, selective and cost effective anti-cancer chalcones.

  3. uPAR as anti-cancer target

    DEFF Research Database (Denmark)

    Lund, Ida K; Illemann, Martin; Thurison, Tine

    2011-01-01

    , and a potential diagnostic and predictive impact of the different uPAR forms has been reported. Hence, pericellular proteolysis seems to be a suitable target for anti-cancer therapy and numerous approaches have been pursued. Targeting of this process may be achieved by preventing the binding of uPA to u......Degradation of proteins in the extracellular matrix is crucial for the multistep process of cancer invasion and metastasis. Compelling evidence has demonstrated the urokinase receptor (uPAR) and its cognate ligand, the urokinase plasminogen activator (uPA), to play critical roles in the concerted...... up-regulated during cancer progression and is primarily confined to the tumor-associated stromal compartment. Furthermore, both uPAR and uPA have proven to be prognostic markers in several types of cancer; high levels indicating poor survival. The cleaved forms of uPAR are also prognostic markers...

  4. Telomere and telomerase as targets for anti-cancer and regeneration therapies

    Institute of Scientific and Technical Information of China (English)

    Yi-hsin HSU; Jing-jer LIN

    2005-01-01

    Telomerase is a ribonucleoprotein that directs the synthesis of telomeric sequence.It is detected in majority of malignant tumors, but not in most normal somatic cells.Because telomerase plays a critical role in cell immortality and tumor formation, it has been one of the targets for anti-cancer and regeneration drug development. In this review, we will discuss therapeutic approaches based mainly on small molecules that have been developed to inhibit telomerase activity, modulate telomerase expression, and telomerase directed gene therapy.

  5. Folate receptor targeted liposomes encapsulating anti-cancer drugs.

    Science.gov (United States)

    Chaudhury, Anumita; Das, Surajit

    2015-01-01

    Among all available lipid based nanoparticulate systems, the success of liposomal drug delivery system is evident by the number of liposomal products available in the market or under advanced stages of preclinical and clinical trials. Liposome has the ability to deliver chemotherapeutic agents to the targeted tissues or even inside the cancerous cells by enhanced intracellular penetration or improved tumour targeting. In the last decade, folate receptor mediated tumour targeting has emerged as an attractive alternative method of active targeting of cancer cells through liposomes due to its numerous advantages over other targeting methods. Folate receptors, also known as folate binding proteins, allow the binding and internalization of folate or folic acid into the cells by a method called folate receptor mediated endocytosis. They have restricted presence in normal cells and are mostly expressed during malignant transformation. In this review article, folate receptor targeting capability of liposomes has been described. This review article has focussed on the different cancer drugs which have been encapsulated in folate receptor targeted liposomes and their in vitro as well as in vivo efficacies in several tumour models.

  6. Targeting Anti-Cancer Active Compounds: Affinity-Based Chromatographic Assays

    Science.gov (United States)

    de Moraes, Marcela Cristina; Cardoso, Carmen Lucia; Seidl, Claudia; Moaddel, Ruin; Cass, Quezia Bezerra

    2016-01-01

    Affinity-based chromatography assays encompass the use of solid supports containing immobilized biological targets to monitor binding events in the isolation , identification and/or characterization of bioactive compounds. This powerful bioanalytical technique allows the screening of potential binders through fast analyses that can be directly performed using isolated substances or complex matrices. An overview of the recent researches in frontal and zonal affinity-based chromatography screening assays, which has been used as a tool in the identification and characterization of new anti-cancer agents, is discussed. In addition, a critical evaluation of the recently emerged ligands fishing assays in complex mixtures is also discussed. PMID:27306095

  7. Logical design of an anti-cancer agent targeting the plant homeodomain in Pygopus2.

    Science.gov (United States)

    Ali, Ferdausi; Yamaguchi, Keiichi; Fukuoka, Mayuko; Elhelaly, Abdelazim Elsayed; Kuwata, Kazuo

    2016-09-01

    Pygopus2 (Pygo2) is a component of the Wnt signaling pathway, which is required for β-catenin mediated transcription. Plant homeodomain (PHD) finger in Pygo2 intercalates the methylated histone 3 (H3K4me) tail and HD1 domain of BCL9 that binds to β-catenin. Thus, PHD finger may be a potential target for the logical design of an anti-cancer drug. Here, we found that Spiro[2H-naphthol[1,2-b]pyran-2,4'-piperidine]-1'ethanol,3,4-dihydro-4-hydroxy-α-(6-methyl-1H-indol-3-yl)) termed JBC117 interacts with D339, A348, R356, V376 and A378 in PHD corresponding to the binding sites with H3K4me and/or HD1, and has strong anti-cancer effects. For colon (HCT116) and lung (A549) cancer cell lines, IC50 values were 2.6 ± 0.16 and 3.3 ± 0.14 μM, respectively, while 33.80 ± 0.15 μM for the normal human fibroblast cells. JBC117 potently antagonized the cellular effects of β-catenin-dependent activity and also inhibited the migration and invasion of cancer cells. In vivo studies showed that the survival time of mice was significantly prolonged by the subcutaneous injection of JBC117 (10 mg/kg/day). In conclusion, JBC117 is a novel anti-cancer lead compound targeting the PHD finger of Pygo2 and has a therapeutic effect against colon and lung cancer.

  8. Reverse screening approach to identify potential anti-cancer targets of dipyridamole

    Science.gov (United States)

    Ge, Shu-Min; Zhan, Dong-Ling; Zhang, Shu-Hua; Song, Li-Qiang; Han, Wei-Wei

    2016-01-01

    Dipyridamole (DIP) inhibits thrombus formation when given chronically, and causes vasodilation over a short time. To date, DIP can increase the anticancer drugs (5-fluorouracil, methotrexate, piperidine, vincristine) concentration in cancer cells and hence enhance the efficacy of treatment cancer. The inhibition of DIP may result in increased 5-fluorouracil efficacy and diminish the drug side effects. But the actual molecular targets remain unknown. In this study, reverse protein-ligands docking, and quantum mechanics were used to search for the potential molecular targets of DIP. The quantum mechanics calculation was performed by using Gaussian 03 program package. Reverse pharmacophore mapping was used to search for potential molecular target candidates for a given small molecule. The docking study was used for exploring the potential anti-cancer targets of dipyridamole. The two predicted binders with the statistically significant prediction are dihydropyrimidine dehydrogenase (DPD) (PDB Id: 1GTE) and human spindle checkpoint kinase Bub1 (PDB Id: 3E7E). Structure analysis suggests that electrostatic interaction and hydrogen bonding play an important role in their binding process. The strong functional linkage of DIP and 5FU supports our prediction. In conclusion, these results generate a tractable set of anticancer proteins. The exploration of polypharmacology will provide us new opportunities in treating systematic diseases, such as the cancers. The results would generate a tractable set of anticancer target proteins for future experimental validations. PMID:28077994

  9. Optimization of anti-cancer drugs and a targeting molecule on multifunctional gold nanoparticles

    Science.gov (United States)

    Rizk, Nahla; Christoforou, Nicolas; Lee, Sungmun

    2016-05-01

    Breast cancer is the most common and deadly cancer among women worldwide. Currently, nanotechnology-based drug delivery systems are useful for cancer treatment; however, strategic planning is critical in order to enhance the anti-cancer properties and reduce the side effects of cancer therapy. Here, we designed multifunctional gold nanoparticles (AuNPs) conjugated with two anti-cancer drugs, TGF-β1 antibody and methotrexate, and a cancer-targeting molecule, folic acid. First, optimum size and shape of AuNPs was selected by the highest uptake of AuNPs by MDA-MB-231, a metastatic human breast cancer cell line. It was 100 nm spherical AuNPs (S-AuNPs) that were used for further studies. A fixed amount (900 μl) of S-AuNP (3.8 × 108 particles/ml) was conjugated with folic acid-BSA or methotrexate-BSA. Methotrexate on S-AuNP induced cellular toxicity and the optimum amount of methotrexate-BSA (2.83 mM) was 500 μl. Uptake of S-AuNPs was enhanced by folate conjugation that binds to folate receptors overexpressed by MDA-MB-231 and the optimum uptake was at 500 μl of folic acid-BSA (2.83 mM). TGF-β1 antibody on S-AuNP reduced extracellular TGF-β1 of cancer cells by 30%. Due to their efficacy and tunable properties, we anticipate numerous clinical applications of multifunctional gold nanospheres in treating breast cancer.

  10. Targeting NK cells for anti-cancer immunotherapy: clinical and pre-clinical approaches

    Directory of Open Access Journals (Sweden)

    Sebastian eCarotta

    2016-04-01

    Full Text Available The recent success of checkpoint blockade has highlighted the potential of immunotherapy approaches for cancer treatment. While the majority of approved immunotherapy drugs target T cell subsets, it is appreciated that other components of the immune system have important roles in tumor immune-surveillance as well and thus represent promising additional targets for immunotherapy. Natural killer cells are the body’s first line of defense against infected or transformed cells as they kill target cells in an antigen-independent manner. Although several studies have clearly demonstrated the active role of NK cells in cancer-immune surveillance, only few clinically approved therapies currently exist that harness their potential. Our increased understanding of NK cell biology over the past few years has renewed the interest in NK cell based anti-cancer therapies, which has lead to a steady increase of NK cell based clinical and pre-clinical trials. Here, the role of NK cells in cancer immunesurveillance is summarized and several novel approaches to enhance NK cell cytotoxicity against cancer are discussed.

  11. Bridging academic science and clinical research in the search for novel targeted anti-cancer agents

    Institute of Scientific and Technical Information of China (English)

    Alex Matter

    2015-01-01

    This review starts with a brief history of drug discovery&development, and the place of Asia in this worldwide effort discussed. hTe conditions and constraints of a successful translational R&D involving academic basic research and clinical research are discussed and the Singapore model for pursuit of open R&D described. hTe importance of well-characterized, validated drug targets for the search for novel targeted anti-cancer agents is emphasized, as well as a structured, high quality translational R&D. Furthermore, the characteristics of an attractive preclinical development drug candidate are discussed laying the foundation of a successful preclinical development. hTe most frequent sources of failures are described and risk management at every stage is highly recommended. Organizational factors are also considered to play an important role. hTe factors to consider before starting a new drug discovery&development project are described, and an example is given of a successful clinical project that has had its roots in local universities and was carried through preclinical development into phase I clinical trials.

  12. Landscape of Targeted Anti-Cancer Drug Synergies in Melanoma Identifies a Novel BRAF-VEGFR/PDGFR Combination Treatment.

    Directory of Open Access Journals (Sweden)

    Adam A Friedman

    Full Text Available A newer generation of anti-cancer drugs targeting underlying somatic genetic driver events have resulted in high single-agent or single-pathway response rates in selected patients, but few patients achieve complete responses and a sizeable fraction of patients relapse within a year. Thus, there is a pressing need for identification of combinations of targeted agents which induce more complete responses and prevent disease progression. We describe the results of a combination screen of an unprecedented scale in mammalian cells performed using a collection of targeted, clinically tractable agents across a large panel of melanoma cell lines. We find that even the most synergistic drug pairs are effective only in a discrete number of cell lines, underlying a strong context dependency for synergy, with strong, widespread synergies often corresponding to non-specific or off-target drug effects such as multidrug resistance protein 1 (MDR1 transporter inhibition. We identified drugs sensitizing cell lines that are BRAFV600E mutant but intrinsically resistant to BRAF inhibitor PLX4720, including the vascular endothelial growth factor receptor/kinase insert domain receptor (VEGFR/KDR and platelet derived growth factor receptor (PDGFR family inhibitor cediranib. The combination of cediranib and PLX4720 induced apoptosis in vitro and tumor regression in animal models. This synergistic interaction is likely due to engagement of multiple receptor tyrosine kinases (RTKs, demonstrating the potential of drug- rather than gene-specific combination discovery approaches. Patients with elevated biopsy KDR expression showed decreased progression free survival in trials of mitogen-activated protein kinase (MAPK kinase pathway inhibitors. Thus, high-throughput unbiased screening of targeted drug combinations, with appropriate library selection and mechanistic follow-up, can yield clinically-actionable drug combinations.

  13. RasGRPs are targets of the anti-cancer agent ingenol-3-angelate.

    Directory of Open Access Journals (Sweden)

    Xiaohua Song

    Full Text Available Ingenol-3-angelate (I3A is a non-tumor promoting phorbol ester-like compound identified in the sap of Euphoria peplus. Similar to tumor promoting phorbol esters, I3A is a diacylglycerol (DAG analogue that binds with high affinity to the C1 domains of PKCs, recruits PKCs to cellular membranes and promotes enzyme activation. Numerous anti-cancer activities have been attributed to I3A and ascribed to I3A's effects on PKCs. We show here that I3A also binds to and activates members of the RasGRP family of Ras activators leading to robust elevation of Ras-GTP and engagement of the Raf-Mek-Erk kinase cascade. In response to I3A, recombinant proteins consisting of GFP fused separately to full-length RasGRP1 and RasGRP3 were rapidly recruited to cell membranes, consistent with direct binding of the compound to RasGRP's C1 domain. In the case of RasGRP3, IA3 treatment led to positive regulatory phosphorylation on T133 and activation of the candidate regulatory kinase PKCδ. I3A treatment of select B non-Hodgkin's lymphoma cell lines resulted in quantitative and qualitative changes in Bcl-2 family member proteins and induction of apoptosis, as previously demonstrated with the DAG analogue bryostatin 1 and its synthetic analogue pico. Our results offer further insights into the anticancer properties of I3A, support the idea that RasGRPs represent potential cancer therapeutic targets along with PKC, and expand the known range of ligands for RasGRP regulation.

  14. Design, Synthesis and Biological Evaluation of Novel Rapamycin Benzothiazole Hybrids as mTOR Targeted Anti-cancer Agents.

    Science.gov (United States)

    Xie, Lijun; Huang, Jie; Chen, Xiaoming; Yu, Hui; Li, Kualiang; Yang, Dan; Chen, Xiaqin; Ying, Jiayin; Pan, Fusheng; Lv, Youbing; Cheng, Yuanrong

    2016-01-01

    The immunosuppressant drug rapamycin, was firstly identified as a mammalian target of rapamycin (mTOR) allosteric inhibitor, and its derivatives have been successfully developed as anti-cancer drugs. Therefore, finding rapamycin derivatives with better anti-cancer activity has been proved to be an effective way to discover new targeted anti-cancer drugs. In this paper, structure modification was performed at the C-43 position of rapamycin using bioisosterism and a hybrid approach: a series of novel rapamycin-benzothiazole hybrids 4a-e, 5a-c, and 9a, b have been designed, synthesized and evaluated for their anti-cancer activity against Caski, CNE-2, SGC-7901, PC-3, SK-NEP-1 and A-375 human cancer cell lines. Some of these compounds (4a-e, 9a, b) displayed good to excellent potency against the Caski and SK-NEP-1 cell line as compared with rapamycin. Compound 9b as the most active compound showed IC50 values of 8.3 (Caski) and 9.6 μM (SK-NEP-1), respectively. In addition, research on the mechanism showed that 9b was able to cause G1 phase arrest and induce apoptosis in the Caski cell line. Most importantly, it significantly decreased the phosphorylation of S6 ribosomal protein, p70S6K1 and 4EBP1, which indicated that 9b inhibited the cancer cell growth by blocking the mTOR pathway and may have the potential to become a new mTOR inhibitor.

  15. Natural product Celastrol destabilizes tubulin heterodimer and facilitates mitotic cell death triggered by microtubule-targeting anti-cancer drugs.

    Directory of Open Access Journals (Sweden)

    Hakryul Jo

    Full Text Available BACKGROUND: Microtubule drugs are effective anti-cancer agents, primarily due to their ability to induce mitotic arrest and subsequent cell death. However, some cancer cells are intrinsically resistant or acquire a resistance. Lack of apoptosis following mitotic arrest is thought to contribute to drug resistance that limits the efficacy of the microtubule-targeting anti-cancer drugs. Genetic or pharmacological agents that selectively facilitate the apoptosis of mitotic arrested cells present opportunities to strengthen the therapeutic efficacy. METHODOLOGY AND PRINCIPAL FINDINGS: We report a natural product Celastrol targets tubulin and facilitates mitotic cell death caused by microtubule drugs. First, in a small molecule screening effort, we identify Celastrol as an inhibitor of neutrophil chemotaxis. Subsequent time-lapse imaging analyses reveal that inhibition of microtubule-mediated cellular processes, including cell migration and mitotic chromosome alignment, is the earliest events affected by Celastrol. Disorganization, not depolymerization, of mitotic spindles appears responsible for mitotic defects. Celastrol directly affects the biochemical properties of tubulin heterodimer in vitro and reduces its protein level in vivo. At the cellular level, Celastrol induces a synergistic apoptosis when combined with conventional microtubule-targeting drugs and manifests an efficacy toward Taxol-resistant cancer cells. Finally, by time-lapse imaging and tracking of microtubule drug-treated cells, we show that Celastrol preferentially induces apoptosis of mitotic arrested cells in a caspase-dependent manner. This selective effect is not due to inhibition of general cell survival pathways or mitotic kinases that have been shown to enhance microtubule drug-induced cell death. CONCLUSIONS AND SIGNIFICANCE: We provide evidence for new cellular pathways that, when perturbed, selectively induce the apoptosis of mitotic arrested cancer cells, identifying a

  16. Translational approaches targeting the p53 pathway for anti-cancer therapy

    OpenAIRE

    2012-01-01

    The p53 tumour suppressor blocks cancer development by triggering apoptosis or cellular senescence in response to oncogenic stress or DNA damage. Consequently, the p53 signalling pathway is virtually always inactivated in human cancer cells. This unifying feature has commenced tremendous efforts to develop p53-based anti-cancer therapies. Different strategies exist that are adapted to the mechanisms of p53 inactivation. In p53-mutated tumours, delivery of wild-type p53 by adenovirus-based gen...

  17. Enzyme inhibition as a key target for the development of novel metal-based anti-cancer therapeutics.

    Science.gov (United States)

    Griffith, Darren; Parker, James P; Marmion, Celine J

    2010-06-01

    Historically, DNA has been the target for many metal-based anti-cancer drugs, but drawbacks of prevailing therapies have stimulated the search for new molecular targets which may present unique opportunities for therapeutic exploitation. Enzyme inhibition has recently been identified as an alternative and significant target. The pursuit of novel metallodrug candidates that selectively target enzymes is now the subject of intense investigation in medicinal bioinorganic chemistry and chemical biology. In the field of drug design, it is recognised by many that exploiting the structural and chemical diversity of metal ions for the identification of potential hit and lead candidates can dramatically increase the number of possible drug candidates that may be added to the already abundant armoury of chemotherapeutic agents. This review will focus on recent key advancements in enzyme inhibition as a key target for the development of novel metal-based anti-cancer therapeutics. The enormous clinical success of classical platinum drugs, amongst others, coupled with the wealth of knowledge accumulated in recent years on enzyme structure and function, has undoubtedly been the impetus behind the development of new metallodrug candidates with enzyme inhibitory properties. Recent trends in this field will be reviewed with a particular emphasis on metal complexes that inhibit protein and lipid kinases, matrix metalloproteases, telomerases, topoisomerases, glutathione-S-transferases, and histone deacetylases.

  18. Targeted anti-cancer prodrug based on carbon nanotube with photodynamic therapeutic effect and pH-triggered drug release

    Energy Technology Data Exchange (ETDEWEB)

    Fan, Jianquan; Zeng, Fang, E-mail: mcfzeng@scut.edu.cn; Xu, Jiangsheng; Wu, Shuizhu, E-mail: shzhwu@scut.edu.cn [South China University of Technology, College of Materials Science and Engineering, State Key Laboratory of Luminescent Materials and Devices (China)

    2013-09-15

    Herein, we describe a multifunctional anti-cancer prodrug system based on water-dispersible carbon nanotube (CNT); this prodrug system features active targeting, pH-triggered drug release, and photodynamic therapeutic properties. For this prodrug system (with the size of {approx}100-300 nm), an anti-cancer drug, doxorubicin (DOX), was incorporated onto CNT via a cleavable hydrazone bond; and a targeting ligand (folic acid) was also coupled onto CNT. This prodrug can preferably enter folate receptor (FR)-positive cancer cells and undergo intracellular release of the drug triggered by the reduced pH. The targeted CNT-based prodrug system can cause lower cell viability toward FR-positive cells compared to the non-targeted ones. Moreover, the CNT carrier exhibits photodynamic therapeutic (PDT) action; and the cell viability of FR-positive cancer cells can be further reduced upon light irradiation. The dual effects of pH-triggered drug release and PDT increase the therapeutic efficacy of the DOX-CNT prodrug. This study may offer some useful insights on designing and improving the applicability of CNT for other drug delivery systems.

  19. LGR5 expressing cells of hair follicle as potential targets for antibody mediated anti-cancer laser therapy

    Science.gov (United States)

    Popov, Boris V.

    2013-02-01

    Near infrared laser immunotherapy becomes now a new promising research field to cure the patients with cancers. One of the critical limitation in medical application of this treatment is availability of the specific markers for delivery of laser-sensitive nanoparticles. When coupled to antibodies to the cancer stem cells markers these nanoparticles may be delivered to the cancer tissue and mediate the laser induced thermolysis of the cancer stem cells that initiate and drive growth of cancer. This paper addresses the Lgr5 cell surface marker mediating the Wnt/β-catenin signal transduction as a potential target for anti-cancer laser immunotherapy of skin cancers.

  20. Structure and Potential Cellular Targets of HAMLET-like Anti-Cancer Compounds made from Milk Components.

    Science.gov (United States)

    Rath, Emma M; Duff, Anthony P; Håkansson, Anders P; Vacher, Catherine S; Liu, Guo Jun; Knott, Robert B; Church, William Bret

    2015-01-01

    The HAMLET family of compounds (Human Alpha-lactalbumin Made Lethal to Tumours) was discovered during studies on the properties of human milk, and is a class of protein-lipid complexes having broad spectrum anti-cancer, and some specific anti-bacterial properties. The structure of HAMLET-like compounds consists of an aggregation of partially unfolded protein making up the majority of the compound's mass, with fatty acid molecules bound in the hydrophobic core. This is a novel protein-lipid structure and has only recently been derived by small-angle X-ray scattering analysis. The structure is the basis of a novel cytotoxicity mechanism responsible for anti-cancer activity to all of the around 50 different cancer cell types for which the HAMLET family has been trialled. Multiple cytotoxic mechanisms have been hypothesised for the HAMLET-like compounds, but it is not yet clear which of those are the initiating cytotoxic mechanism(s) and which are subsequent activities triggered by the initiating mechanism(s). In addition to the studies into the structure of these compounds, this review presents the state of knowledge of the anti-cancer aspects of HAMLET-like compounds, the HAMLET-induced cytotoxic activities to cancer and non-cancer cells, and the several prospective cell membrane and intracellular targets of the HAMLET family. The emerging picture is that HAMLET-like compounds initiate their cytotoxic effects on what may be a cancer-specific target in the cell membrane that has yet to be identified. This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

  1. Anti-cancer drug loaded iron-gold core-shell nanoparticles (Fe@Au) for magnetic drug targeting.

    Science.gov (United States)

    Kayal, Sibnath; Ramanujan, Raju Vijayaraghavan

    2010-09-01

    Magnetic drug targeting, using core-shell magnetic carrier particles loaded with anti-cancer drugs, is an emerging and significant method of cancer treatment. Gold shell-iron core nanoparticles (Fe@Au) were synthesized by the reverse micelle method with aqueous reactants, surfactant, co-surfactant and oil phase. XRD, XPS, TEM and magnetic property measurements were utilized to characterize these core-shell nanoparticles. Magnetic measurements showed that the particles were superparamagnetic at room temperature and that the saturation magnetization decreased with increasing gold concentration. The anti-cancer drug doxorubicin (DOX) was loaded onto these Fe@Au nanoparticle carriers and the drug release profiles showed that upto 25% of adsorbed drug was released in 80 h. It was found that the amine (-NH2) group of DOX binds to the gold shell. An in vitro apparatus simulating the human circulatory system was used to determine the retention of these nanoparticle carriers when exposed to an external magnetic field. A high percentage of magnetic carriers could be retained for physiologically relevant flow speeds of fluid. The present findings show that DOX loaded gold coated iron nanoparticles are promising for magnetically targeted drug delivery.

  2. uPAR as anti-cancer target: evaluation of biomarker potential, histological localization, and antibody-based therapy

    DEFF Research Database (Denmark)

    Lund, Ida K; Illemann, Martin; Sørensen, Tine Thurison

    2011-01-01

    , and a potential diagnostic and predictive impact of the different uPAR forms has been reported. Hence, pericellular proteolysis seems to be a suitable target for anti-cancer therapy and numerous approaches have been pursued. Targeting of this process may be achieved by preventing the binding of uPA to u......Degradation of proteins in the extracellular matrix is crucial for the multistep process of cancer invasion and metastasis. Compelling evidence has demonstrated the urokinase receptor (uPAR) and its cognate ligand, the urokinase plasminogen activator (uPA), to play critical roles in the concerted...... up-regulated during cancer progression and is primarily confined to the tumor-associated stromal compartment. Furthermore, both uPAR and uPA have proven to be prognostic markers in several types of cancer; high levels indicating poor survival. The cleaved forms of uPAR are also prognostic markers...

  3. Molecular biology of cancer-associated fibroblasts: can these cells be targeted in anti-cancer therapy?

    Science.gov (United States)

    Gonda, Tamas A; Varro, Andrea; Wang, Timothy C; Tycko, Benjamin

    2010-02-01

    It is increasingly recognized that the non-neoplastic stromal compartment in most solid cancers plays an active role in tumor proliferation, invasion and metastasis. Cancer-associated fibroblasts (CAFs) are one of the most abundant cell types in the tumor stroma, and these cells are pro-tumorigenic. Evidence that CAFs are epigenetically and possibly also genetically distinct from normal fibroblasts is beginning to define these cells as potential targets of anti-cancer therapy. Here, we review the cell-of-origin and molecular biology of CAFs, arguing that such knowledge provides a rational basis for designing therapeutic strategies to coordinately and synergistically target both the stromal and malignant epithelial component of human cancers.

  4. Use of the Rad51 promoter for targeted anti-cancer therapy

    Science.gov (United States)

    Hine, Christopher M.; Seluanov, Andrei; Gorbunova, Vera

    2008-01-01

    Rad51 protein, involved in homologous recombination, is overexpressed in a variety of tumors, and its expression is correlated with a poor prognosis. Here we propose to exploit the overexpression of Rad51 in cancer cells to design a Rad51 promoter-based anticancer therapy. On average, Rad51 mRNA and protein levels are increased in cancer cells four- and sixfold, respectively. Serendipitously, we discovered that when the Rad51 ORF is replaced with another ORF, the difference in promoter activity between normal and cancer cells increases to an average of 840-fold with a maximum difference of 12,500-fold. This dramatic difference in activity has high therapeutic potential. We demonstrate that the fusion of Rad51 promoter to diphtheria toxin A (DTA) gene kills a variety of cancer cell types, including breast cancer, fibrosarcoma, and cervical cancer cells, with minimal effect on normal breast epithelial cells and normal fibroblasts. Our results suggest that therapies based on the Rad51 promoter will be highly tumor specific and open new avenues for targeting a broad range of cancers. PMID:19106292

  5. Network modelling reveals the mechanism underlying colitis-associated colon cancer and identifies novel combinatorial anti-cancer targets.

    Science.gov (United States)

    Lu, Junyan; Zeng, Hanlin; Liang, Zhongjie; Chen, Limin; Zhang, Liyi; Zhang, Hao; Liu, Hong; Jiang, Hualiang; Shen, Bairong; Huang, Ming; Geng, Meiyu; Spiegel, Sarah; Luo, Cheng

    2015-10-08

    The connection between inflammation and tumourigenesis has been well established. However, the detailed molecular mechanism underlying inflammation-associated tumourigenesis remains unknown because this process involves a complex interplay between immune microenvironments and epithelial cells. To obtain a more systematic understanding of inflammation-associated tumourigenesis as well as to identify novel therapeutic approaches, we constructed a knowledge-based network describing the development of colitis-associated colon cancer (CAC) by integrating the extracellular microenvironment and intracellular signalling pathways. Dynamic simulations of the CAC network revealed a core network module, including P53, MDM2, and AKT, that may govern the malignant transformation of colon epithelial cells in a pro-tumor inflammatory microenvironment. Furthermore, in silico mutation studies and experimental validations led to a novel finding that concurrently targeting ceramide and PI3K/AKT pathway by chemical probes or marketed drugs achieves synergistic anti-cancer effects. Overall, our network model can guide further mechanistic studies on CAC and provide new insights into the design of combinatorial cancer therapies in a rational manner.

  6. Recursive Random Lasso (RRLasso) for Identifying Anti-Cancer Drug Targets.

    Science.gov (United States)

    Park, Heewon; Imoto, Seiya; Miyano, Satoru

    2015-01-01

    Uncovering driver genes is crucial for understanding heterogeneity in cancer. L1-type regularization approaches have been widely used for uncovering cancer driver genes based on genome-scale data. Although the existing methods have been widely applied in the field of bioinformatics, they possess several drawbacks: subset size limitations, erroneous estimation results, multicollinearity, and heavy time consumption. We introduce a novel statistical strategy, called a Recursive Random Lasso (RRLasso), for high dimensional genomic data analysis and investigation of driver genes. For time-effective analysis, we consider a recursive bootstrap procedure in line with the random lasso. Furthermore, we introduce a parametric statistical test for driver gene selection based on bootstrap regression modeling results. The proposed RRLasso is not only rapid but performs well for high dimensional genomic data analysis. Monte Carlo simulations and analysis of the "Sanger Genomics of Drug Sensitivity in Cancer dataset from the Cancer Genome Project" show that the proposed RRLasso is an effective tool for high dimensional genomic data analysis. The proposed methods provide reliable and biologically relevant results for cancer driver gene selection.

  7. Recursive Random Lasso (RRLasso for Identifying Anti-Cancer Drug Targets.

    Directory of Open Access Journals (Sweden)

    Heewon Park

    Full Text Available Uncovering driver genes is crucial for understanding heterogeneity in cancer. L1-type regularization approaches have been widely used for uncovering cancer driver genes based on genome-scale data. Although the existing methods have been widely applied in the field of bioinformatics, they possess several drawbacks: subset size limitations, erroneous estimation results, multicollinearity, and heavy time consumption. We introduce a novel statistical strategy, called a Recursive Random Lasso (RRLasso, for high dimensional genomic data analysis and investigation of driver genes. For time-effective analysis, we consider a recursive bootstrap procedure in line with the random lasso. Furthermore, we introduce a parametric statistical test for driver gene selection based on bootstrap regression modeling results. The proposed RRLasso is not only rapid but performs well for high dimensional genomic data analysis. Monte Carlo simulations and analysis of the "Sanger Genomics of Drug Sensitivity in Cancer dataset from the Cancer Genome Project" show that the proposed RRLasso is an effective tool for high dimensional genomic data analysis. The proposed methods provide reliable and biologically relevant results for cancer driver gene selection.

  8. IGF-1R as an anti-cancer target-trials and tribulations

    Institute of Scientific and Technical Information of China (English)

    Helen X.Chen; Elad Sharon

    2013-01-01

    Type Ⅰ insulin-like growth factor receptor (IGF-1R) has long been recognized for its role in tumorigenesis and growth,but only recently have the tools for targeting the IGF pathway become available.More than 10 IGF/IGF-1R inhibitors have entered clinical trials,and these belong to three main classes:(1)monoclonal antibodies against IGF-1R,(2) monoclonal antibodies against IGF-1R ligands (IGF-1 and IGF-2),and (3) IGF-1R tyrosine kinase inhibitors.These IGF-1R-targeting agents share common effects on IGF-1R signaling but differ in mechanisms of action,spectrum of target inhibition,and pharmacological features.Clinical activity of IGF-1R inhibitors has been demonstrated with sustained responses in a small number of patients with select tumor types,such as Ewing sarcoma and thymoma.However,many large clinical trials involving patients with adult tumors,including non-small cell lung cancer,breast cancer,and pancreatic cancer,failed to show clinical benefit in the overall patient population.Possible reasons for failure include the complexity of the IGF-1R/insulin receptor system and parallel growth and survival pathways,as well as a lack of patient selection markers.While IGF-1R remains a valid target for selected tumor types,identification of predictive markers and rational combinations will be critical to success in future development.

  9. Molecular docking based screening of novel designed chalcone series of compounds for their anti-cancer activity targeting EGFR kinase domain

    Science.gov (United States)

    Rao, Chennu Maruthi Malya Prasada; Yejella, Rajendra Prasad; Rehman, Rehman Shaik Abdul; Basha, Syed Hussain

    2015-01-01

    Epidermal growth factor receptors (EGFR) are critical for the growth of many tumors and expressed at high levels in about one third of epithelial cancers. Hence, blockade of the binding sites for EGFR has been hypothesized as an effective anti-cancer therapy. Chalcone derivative compounds have been shown to be highly effective anti-cancer agents, however there are still so many novel derivatives possible, one of which might get us the best targeted EGFR inhibitor. In this effort directed towards the discovery of novel, potent anti-tumor agents for the treatment of cancer, in the present study a library of novel chalcone series of compounds has been designed and evaluated for their anti-cancer activity targeting EGFR kinase domain using various computational approaches. Among the twenty five novel designed chalcone series of compounds, all of them have found to be successfully docking inside the active binding domain of EGFR receptor target with a binding energy in a range of -6.10 to -9.25 Kcal/mol with predicted IC50 value range of 33.50 micor molar to 164.66 nano molar respectively. On the other hand, calculated 2DQSAR molecular descriptor properties of the compounds showed promising ADME parameters and found to be well in compliance with Lipinski׳s rule of five. Among all the twenty five compounds tested, compound 21 ((2E)-3-(anthracen-9-yl)-1-phenylprop-2-2n-1- one) was found to be the best lead like molecule with a binding energy of -9.25 kcal/mol with predicted IC50 value of 164.66 nano molar. Conclusively, novel designed compound 21 of the present study have shown promising anti-cancer potential worth considering for further evaluations. PMID:26339147

  10. Exploiting developments in nanotechnology for the preferential delivery of platinum-based anti-cancer agents to tumours: targeting some of the hallmarks of cancer.

    Science.gov (United States)

    Parker, James P; Ude, Ziga; Marmion, Celine J

    2016-01-01

    Platinum drugs as anti-cancer therapeutics are held in extremely high regard. Despite their success, there are drawbacks associated with their use; their dose-limiting toxicity, their limited activity against an array of common cancers and patient resistance to Pt-based therapeutic regimes. Current investigations in medicinal inorganic chemistry strive to offset these shortcomings through selective targeting of Pt drugs and/or the development of Pt drugs with new or multiple modes of action. A comprehensive overview showcasing how liposomes, nanocapsules, polymers, dendrimers, nanoparticles and nanotubes may be employed as vehicles to selectively deliver cytotoxic Pt payloads to tumour cells is provided.

  11. Detecting the effect of targeted anti-cancer medicines on single cancer cells using a poly-silicon wire ion sensor integrated with a confined sensitive window.

    Science.gov (United States)

    Wu, You-Lin; Hsu, Po-Yen; Hsu, Chung-Ping; Lin, Jing-Jenn

    2012-10-01

    A mold-cast polydimethylsiloxane (PDMS) confined window was integrated with a poly-silicon wire (PSW) ion sensor. The PSW sensor surface inside the confined window was coated with a 3-aminopropyltriethoxysilane (γ-APTES) sensitive layer which allowed a single living cell to be cultivated. The change in the microenvironment due to the extracellular acidification of the single cell could then be determined by measuring the current flowing through the PSW channel. Based on this, the PSW sensor integrated with a confined sensitive window was used to detect the apoptosis as well as the effect of anti-cancer medicines on the single living non-small-lung-cancer (NSLC) cells including lung adenocarcinoma cancer cells A549 and H1299, and lung squamous-cell carcinoma CH27 cultivated inside the confined window. Single human normal cells including lung fibroblast cells WI38, lung fibroblast cells MRC5, and bronchial epithelium cell Beas-2B were tested for comparison. Two targeted anti-NSCLC cancer medicines, Iressa and Staurosporine, were used in the present study. It was found that the PSW sensor can be used to accurately detect the apoptosis of single cancer cells after the anti-cancer medicines were added. It was also found that Staurosporine is more effective than Iressa in activating the apoptosis of cancer cells.

  12. Development of PIK-75 nanosuspension formulation with enhanced delivery efficiency and cytotoxicity for targeted anti-cancer therapy.

    Science.gov (United States)

    Talekar, Meghna; Ganta, Srinivas; Amiji, Mansoor; Jamieson, Stephen; Kendall, Jackie; Denny, William A; Garg, Sanjay

    2013-06-25

    PIK-75 is a phosphatidylinositol 3-kinase (PI3K) inhibitor that shows selectivity toward p110-α over the other PI3K class Ia isoforms p110-β and p110-δ, but it lacks solubility, stability and other kinase selectivity. The purpose of this study was to develop folate-targeted PIK-75 nanosuspension for tumor targeted delivery and to improve therapeutic efficacy in human ovarian cancer model. High pressure homogenization was used to prepare the non-targeted and targeted PIK-75 nanosuspensions which were characterized for size, zeta potential, entrapment efficiency, morphology, saturation solubility and dissolution velocity. In vitro analysis of drug uptake, cell viability and cell survival was conducted in SKOV-3 cells. Drug pharmacokinetics and pAkt expression were determined in SKOV-3 tumor bearing mice. PIK-75 nanosuspensions showed an improvement in dissolution velocity and an 11-fold increase in saturation solubility over pre-milled PIK-75. In vitro studies in SKOV-3 cells indicated a 2-fold improvement in drug uptake and 0.4-fold decrease in IC50 value of PIK-75 following treatment with targeted nanosuspension compared to non-targeted nanosuspension. The improvement in cytotoxicity was attributed to an increase in caspase 3/7 and hROS activity. In vivo studies indicated a 5-10-fold increased PIK-75 accumulation in the tumor with both the nanosuspension formulations compared to PIK-75 suspension. The targeted nanosuspension showed an enhanced downregulation of pAkt compared to non-targeted formulation system. These results illustrate the opportunity to formulate PIK-75 as a targeted nanosuspension to enhance uptake and cytotoxicity of the drug in tumor.

  13. Mapping Novel Metabolic Nodes Targeted by Anti-Cancer Drugs that Impair Triple-Negative Breast Cancer Pathogenicity.

    Science.gov (United States)

    Roberts, Lindsay S; Yan, Peter; Bateman, Leslie A; Nomura, Daniel K

    2017-03-08

    Triple-negative breast cancers (TNBCs) are estrogen receptor, progesterone receptor, and HER2 receptor-negative subtypes of breast cancers that show the worst prognoses and lack targeted therapies. Here, we have coupled the screening of ∼400 anticancer agents that are under development or in the clinic with chemoproteomic and metabolomic profiling to identify novel metabolic mechanisms for agents that impair TNBC pathogenicity. We identify 20 anticancer compounds that significantly impaired cell survival across multiple types of TNBC cells. Among these 20 leads, the phytoestrogenic natural product licochalcone A was of interest, since TNBCs are unresponsive to estrogenic therapies, indicating that licochalcone A was likely acting through another target. Using chemoproteomic profiling approaches, we reveal that licochalcone A impairs TNBC pathogenicity, not through modulating estrogen receptor activity but rather through inhibiting prostaglandin reductase 1, a metabolic enzyme involved in leukotriene B4 inactivation. We also more broadly performed metabolomic profiling to map additional metabolic mechanisms of compounds that impair TNBC pathogenicity. Overlaying lipidomic profiling with drug responses, we find that deubiquitinase inhibitors cause dramatic elevations in acyl carnitine levels, which impair mitochondrial respiration and contribute to TNBC pathogenic impairments. We thus put forth two unique metabolic nodes that are targeted by drugs or drug candidates that impair TNBC pathogenicity. Our results also showcase the utility of coupling drug screens with chemoproteomic and metabolomic profiling to uncover unique metabolic drivers of TNBC pathogenicity.

  14. Cancer metabolic reprogramming:impor tance, main features, and potentials for precise targeted anti-cancer therapies

    Institute of Scientific and Technical Information of China (English)

    Liem Minh Phan; Sai-Ching Jim Yeung; Mong-Hong Lee

    2014-01-01

    Cancer cells are well documented to rewire their metabolism and energy production networks to support and enable rapid proliferation, continuous growth, survival in harsh conditions, invasion, metastasis, and resistance to cancer treatments. Since Dr. Otto Warburg’s discovery about altered cancer cell metabolism in 1930, thousands of studies have shed light on various aspects of cancer metabolism with a common goal to find new ways for effectively eliminating tumor cells by targeting their energy metabolism. hTis review highlights the importance of the main features of cancer metabolism, summarizes recent remarkable advances in this ifeld, and points out the potentials to translate these scientiifc ifndings into life-saving diagnosis and therapies to help cancer patients.

  15. Chemical Genetics Identify eIF2α Kinase Heme Regulated Inhibitor as Anti-Cancer Target

    Science.gov (United States)

    Chen, Ting; Ozel, Duygu; Qiao, Yuan; Harbinski, Fred; Chen, Limo; Denoyelle, Séverine; He, Xiaoying; Zvereva, Nela; Supko, Jeffrey G.; Chorev, Michael; Halperin, Jose A.; Aktas, Bertal H.

    2013-01-01

    Translation initiation plays a critical role in cellular homeostasis, proliferation, differentiation and malignant transformation. Consistently, increasing the abundance of the eIF2·GTP·Met-tRNAi translation initiation complex transforms normal cells and contributes to cancer initiation and the severity of some anemia. The chemical modifiers of the eIF2·GTP·Met-tRNAi ternary complex are therefore invaluable tools for studying its role in the pathobiology of human disorders and for determining if this complex can be pharmacologically targeted for therapeutic purposes. Using a cell based assay, we identified N,N’-diarylureas as novel inhibitors of the ternary complex abundance. Direct functional-genetics and biochemical evidence demonstrated that the N,N’-diarylureas activate heme regulated inhibitor kinase, thereby phosphorylate eIF2α and reduce abundance of the ternary complex. Using tumor cell proliferation in vitro and tumor growth in vivo as paradigms, we demonstrate that N,N’-diarylureas are potent and specific tools for studying the role eIF2·GTP·Met-tRNAi ternary complex in the pathobiology of human disorders. PMID:21765405

  16. Glutathione-responsive nano-vehicles as a promising platform for targeted intracellular drug and gene delivery

    NARCIS (Netherlands)

    Cheng, Ru; Feng, Fang; Meng, Fenghua; Deng, Chao; Feijen, Jan; Zhong, Zhiyuan

    2011-01-01

    The past couple of years have witnessed a tremendous progress in the development of glutathione-responsive nano-vehicles for targeted intracellular drug and gene delivery, as driven by the facts that (i) many therapeutics (e.g. anti-cancer drugs, photosensitizers, and anti-oxidants) and biotherapeut

  17. Nanopharmaceutical Approach for Enhanced Anti-cancer Activity of Betulinic Acid in Lung-cancer Treatment via Activation of PARP: Interaction with DNA as a Target -Anti-cancer Potential of Nano-betulinic Acid in Lung Cancer-

    Directory of Open Access Journals (Sweden)

    Jayeeta Das

    2016-03-01

    Full Text Available Objectives: This study examined the relative efficacies of a derivative of betulinic acid (dBA and its poly (lactide- co-glycolide (PLGA nano-encapsulated form in A549 lung cancer cells in vivo and in co-mutagen [sodium arsenite (SA + benzo]undefined[a]pyrene (BaP]-induced lung cancer in mice in vivo. Methods: dBA was loaded with PLGA nanoparticles by using the standard solvent displacement method. The sizes and morphologies of nano-dBA (NdBA were determined by using transmission electron microscopy (TEM, and their intracellular localization was verified by using confocal microscopy. The binding and interaction of NdBA with calf thymus deoxyribonucleic acid (CT-DNA as a target were analyzed by using conventional circular dichroism (CD and melting temperature (Tm profile data. Apoptotic signalling cascades in vitro and in vivo were studied by using an enzyme-linked immunosorbent assay (ELISA; the ability of NdBA to cross the blood-brain barrier (BBB was also examined. The stage of cell cycle arrest was confirmed by using a fluorescence-activated cell-sorting (FACS data analysis. Results: The average size of the nanoparticles was ~ 110 nm. Confocal microscopy images confirmed the presence of NdBA in the cellular cytoplasm. The bio-physical properties of dBA and NdBA ascertained from the CD and the Tm profiles revealed that NdBA had greater interaction with the target DNA than dBA did. Both dBA and NdBA arrested cell proliferation at G0/G1, NdBA showing the greater effect. NdBA also induced a greater degree of cytotoxicity in A549 cells, but it had an insignificant cytotoxic effect in normal L6 cells. The results of flow cytometric, cytogenetial and histopathological studies in mice revealed that NdBA caused less nuclear condensation and DNA damage than dBA did. TEM images showed the presence of NdBA in brain samples of NdBA fed mice, indicating its ability to cross the BBB. Conclusion: Thus, compared to dBA, NdBA appears to have greater

  18. Personalizing Anti-Cancer Treatment from Genetic and Pharmacokinetic Perspective

    NARCIS (Netherlands)

    S. Bins (Sander)

    2017-01-01

    markdownabstractOnly recently, systemic anti-cancer treatment consisted of little more than chemotherapy, targeting mitosis in rapidly dividing cells such as cancer cells. Increasing biological insight has led to the development of more biology driven treatments, e.g. tyrosine kinase inhibitors and

  19. Genome-wide transcriptional effects of the anti-cancer agent camptothecin.

    Directory of Open Access Journals (Sweden)

    Artur Veloso

    Full Text Available The anti-cancer drug camptothecin inhibits replication and transcription by trapping DNA topoisomerase I (Top1 covalently to DNA in a "cleavable complex". To examine the effects of camptothecin on RNA synthesis genome-wide we used Bru-Seq and show that camptothecin treatment primarily affected transcription elongation. We also observed that camptothecin increased RNA reads past transcription termination sites as well as at enhancer elements. Following removal of camptothecin, transcription spread as a wave from the 5'-end of genes with no recovery of transcription apparent from RNA polymerases stalled in the body of genes. As a result, camptothecin preferentially inhibited the expression of large genes such as proto-oncogenes, and anti-apoptotic genes while smaller ribosomal protein genes, pro-apoptotic genes and p53 target genes showed relative higher expression. Cockayne syndrome group B fibroblasts (CS-B, which are defective in transcription-coupled repair (TCR, showed an RNA synthesis recovery profile similar to normal fibroblasts suggesting that TCR is not involved in the repair of or RNA synthesis recovery from transcription-blocking Top1 lesions. These findings of the effects of camptothecin on transcription have important implications for its anti-cancer activities and may aid in the design of improved combinatorial treatments involving Top1 poisons.

  20. Targeted deletion of the ara operon of Salmonella typhimurium enhances L-arabinose accumulation and drives PBAD-promoted expression of anti-cancer toxins and imaging agents.

    Science.gov (United States)

    Hong, Hyun; Lim, Daejin; Kim, Geun-Joong; Park, Seung-Hwan; Sik Kim, Hyeon; Hong, Yeongjin; Choy, Hyon E; Min, Jung-Joon

    2014-01-01

    Tumor-specific expression of antitumor drugs can be achieved using attenuated Salmonella typhimurium harboring the PBAD promoter, which is induced by L-arabinose. However, L-arabinose does not accumulate because it is metabolized to D-xylulose-5-P by enzymes encoded by the ara operon in Salmonellae. To address this problem, we developed an engineered strain of S. typhimurium in which the ara operon is deleted. Linear DNA transformation was performed using λ red recombinase to exchange the ara operon with linear DNA carrying an antibiotic-resistance gene with homology to regions adjacent to the ara operon. The ara operon-deleted strain and its parental strain were transformed with a plasmid encoding Renilla luciferase variant 8 (RLuc8) or cytolysin A (clyA) under the control of the PBAD promoter. Luciferase assays demonstrated that RLuc8 expression was 49-fold higher in the ara operon-deleted S. typhimurium than in the parental strain after the addition of L-arabinose. In vivo bioluminescence imaging showed that the tumor tissue targeted by the ara operon-deleted Salmonella had a stronger imaging signal (~30-fold) than that targeted by the parental strain. Mice with murine colon cancer (CT26) that had been injected with the ara operon-deleted S. typhimurium expressing clyA showed significant tumor suppression. The present report demonstrates that deletion of the ara operon of S. typhimurium enhances L-arabinose accumulation and thereby drives PBAD-promoted expression of cytotoxic agents and imaging agents. This is a promising approach for tumor therapy and imaging.

  1. Geldanamycin and its anti-cancer activities.

    Science.gov (United States)

    Fukuyo, Yayoi; Hunt, Clayton R; Horikoshi, Nobuo

    2010-04-01

    Geldanamycin is a benzoquinone ansamycin antibiotic that manifests anti-cancer activity through the inhibition of HSP90-chaperone function. The HSP90 molecular chaperone is expressed at high levels in a wide variety of human cancers including melanoma, leukemia, and cancers in colon, prostate, lung, and breast. In cancer cells dependent upon mutated and/or over-expressed oncogene proteins, HSP90 is thought to have a critical role in regulating the stability, folding, and activity of HSP90-associated proteins, so-called "client proteins". These client proteins include the growth-stimulating proteins and kinases that support malignant transformation. Recently, oncogenic activating BRAF mutants have been identified in variety of cancers where constitutive activation of the MEK/ERK MAPK signaling pathway is the key for tumorigenesis, and they have been shown to be client proteins for HSP90. Accordingly, HSP90 inhibition can suppress certain cancer-causing client proteins and therefore represents an important therapeutic target. The molecular mechanism underlying the anti-cancer effect of HSP90 inhibition is complicated. Geldanamycin and its derivatives have been shown to induce the depletion of mutationally-activated BRAF through several mechanisms. In this review, we will describe the HSP90-inhibitory mechanism, focusing on recent progress in understanding HSP90 chaperone structure-function relationships, the identification of new HSP90 client proteins and the development of HSP90 inhibitors for clinical applications.

  2. Toward Repurposing Metformin as a Precision Anti-Cancer Therapy Using Structural Systems Pharmacology

    Science.gov (United States)

    Hart, Thomas; Dider, Shihab; Han, Weiwei; Xu, Hua; Zhao, Zhongming; Xie, Lei

    2016-01-01

    Metformin, a drug prescribed to treat type-2 diabetes, exhibits anti-cancer effects in a portion of patients, but the direct molecular and genetic interactions leading to this pleiotropic effect have not yet been fully explored. To repurpose metformin as a precision anti-cancer therapy, we have developed a novel structural systems pharmacology approach to elucidate metformin’s molecular basis and genetic biomarkers of action. We integrated structural proteome-scale drug target identification with network biology analysis by combining structural genomic, functional genomic, and interactomic data. Through searching the human structural proteome, we identified twenty putative metformin binding targets and their interaction models. We experimentally verified the interactions between metformin and our top-ranked kinase targets. Notably, kinases, particularly SGK1 and EGFR were identified as key molecular targets of metformin. Subsequently, we linked these putative binding targets to genes that do not directly bind to metformin but whose expressions are altered by metformin through protein-protein interactions, and identified network biomarkers of phenotypic response of metformin. The molecular targets and the key nodes in genetic networks are largely consistent with the existing experimental evidence. Their interactions can be affected by the observed cancer mutations. This study will shed new light into repurposing metformin for safe, effective, personalized therapies. PMID:26841718

  3. Gene targeting with retroviral vectors

    Energy Technology Data Exchange (ETDEWEB)

    Ellis, J.; Bernstein, A. (Toronto Univ., ON (Canada))

    1989-04-01

    The authors have designed and constructed integration-defective retroviral vectors to explore their potential for gene targeting in mammalian cells. Two nonoverlapping deletion mutants of the bacterial neomycin resistance (neo) gene were used to detect homologous recombination events between viral and chromosomal sequences. Stable neo gene correction events were selected at a frequency of approximately 1 G418/sup r/ cell per 3 x 10/sup 6/ infected cells. Analysis of the functional neo gene in independent targeted cell clones indicated that unintegrated retroviral linear DNA recombined with the target by gene conversion for variable distances into regions of nonhomology. In addition, transient neo gene correction events which were associated with the complete loss of the chromosomal target sequences were observed. These results demonstrated that retroviral vectors can recombine with homologous chromosomal sequences in rodent and human cells.

  4. Gene targeting in malaria parasites.

    Science.gov (United States)

    Ménard, R; Janse, C

    1997-10-01

    Gene targeting, which permits alteration of a chosen gene in a predetermined way by homologous recombination, is an emerging technology in malaria research. Soon after the development of techniques for stable transformation of red blood cell stages of Plasmodium falciparum and Plasmodium berghei, genes of interest were disrupted in the two species. The main limitations of gene targeting in malaria parasites result from the intracellular growth and slow replication of these parasites. On the other hand, the technology is facilitated by the very high rate of homologous recombination following transformation with targeting constructs (approximately 100%). Here, we describe (i) the vector design and the type of mutation that may be generated in a target locus, (ii) the selection and screening strategies that can be used to identify clones with the desired modification, and (iii) the protocol that was used for disrupting the circumsporozoite protein (CS) and thrombospondin-related anonymous protein (TRAP) genes of P. berghei.

  5. Tanshinones: Sources, Pharmacokinetics and Anti-Cancer Activities

    Directory of Open Access Journals (Sweden)

    Sung-Hoon Kim

    2012-10-01

    Full Text Available Tanshinones are a class of abietane diterpene compound isolated from Salvia miltiorrhiza (Danshen or Tanshen in Chinese, a well-known herb in Traditional Chinese Medicine (TCM. Since they were first identified in the 1930s, more than 40 lipophilic tanshinones and structurally related compounds have been isolated from Danshen. In recent decades, numerous studies have been conducted to investigate the isolation, identification, synthesis and pharmacology of tanshinones. In addition to the well-studied cardiovascular activities, tanshinones have been investigated more recently for their anti-cancer activities in vitro and in vivo. In this review, we update the herbal and alternative sources of tanshinones, and the pharmacokinetics of selected tanshinones. We discuss anti-cancer properties and identify critical issues for future research. Whereas previous studies have suggested anti-cancer potential of tanshinones affecting multiple cellular processes and molecular targets in cell culture models, data from in vivo potency assessment experiments in preclinical models vary greatly due to lack of uniformity of solvent vehicles and routes of administration. Chemical modifications and novel formulations had been made to address the poor oral bioavailability of tanshinones. So far, human clinical trials have been far from ideal in their design and execution for the purpose of supporting an anti-cancer indication of tanshinones.

  6. Effects of JS-K, a novel anti-cancer nitric oxide prodrug, on gene expression in human hepatoma Hep3B cells.

    Science.gov (United States)

    Dong, Ray; Wang, Xueqian; Wang, Huan; Liu, Zhengyun; Liu, Jie; Saavedra, Joseph E

    2017-04-01

    JS-K is a novel anticancer nitric oxide (NO) prodrug effective against a variety of cancer cells, including the inhibition of AM-1 hepatoma cell growth in rats. To further evaluate anticancer effects of JS-K, human hepatoma Hep3B cells were treated with JS-K and the compound control JS-43-126 at various concentrations (0-100μM) for 24h, and cytotoxicity was determined by the MTS assay. The compound control JS-43-126 was not cytotoxic to Hep3B cells at concentrations up to 100μM, while the LC50 for JS-K was about 10μM. To examine the molecular mechanisms of antitumor effects of JS-K, Hep3B cells were treated with 1-10μM of JS-K for 24h, and then subjected to gene expression analysis via real time RT-PCR and protein immunostain via confocal images. JS-K is a GST-α targeting NO prodrug, and decreased immunostaining for GST-α was associated with JS-K treatment. JS-K activated apoptosis pathways in Hep3B cells, including induction of caspase-3, caspase-9, Bax, TNF-α, and IL-1β, and immunostaining for caspase-3 was intensified. The expressions of thrombospondin-1 (TSP-1) and the tissue inhibitors of metalloproteinase-1 (TIMP-1) were increased by JS-K at both transcript and protein levels. JS-K treatment also increased the expression of differentiation-related genes CD14 and CD11b, and depressed the expression of c-myc in Hep3B cells. Thus, multiple molecular events appear to be associated with anticancer effects of JS-K in human hepatoma Hep3B cells, including activation of genes related to apoptosis and induction of genes involved in antiangiogenesis and tumor cell migration.

  7. Anti-cancer Lead Molecule

    KAUST Repository

    Sagar, Sunil

    2014-04-17

    Derivatives of plumbagin can be selectively cytotoxic to breast cancer cells. Derivative `A` (Acetyl Plumbagin) has emerged as a lead molecule for testing against estrogen positive breast cancer and has shown low hepatotoxicity as well as overall lower toxicity in nude mice model. The toxicity of derivative `A` was determined to be even lower than vehicle control (ALT and AST markers). The possible mechanism of action identified based on the microarray experiments and pathway mapping shows that derivative `A` could be acting by altering the cholesterol-related mechanisms. The low toxicity profile of derivative `A` highlights its possible role\\'as future anti-cancer drug and/or as an adjuvant drug to reduce the toxicity of highly toxic chemotherapeutic\\'drugs

  8. Targeted delivery and pH-responsive release of stereoisomeric anti-cancer drugs using β-cyclodextrin assemblied Fe3O4 nanoparticles

    Science.gov (United States)

    Wang, Congli; Huang, Lizhen; Song, Shengmei; Saif, Bassam; Zhou, Yehong; Dong, Chuan; Shuang, Shaomin

    2015-12-01

    The β-cyclodextrin assemblied magnetic Fe3O4 nanoparticles (β-CD-MNPs) were successfully fabricated via a layer-by-layer method. Possessing an average size 14 nm, good stability and super-paramagnetic response (Ms 64 emu/g), the resultant nanocomposites could be served as a versatile biocompatible platform for selective loading, targeted delivery and pH-responsive release of stereoisomeric doxorubicin (DOX) and epirubicin (EPI). 1H-nuclear magnetic resonance (1H NMR) and the computer simulation further give the evidence that partial anthracene ring of drug molecule is included by β-CD. In addition, non-toxic β-CD-MNPs have excellent biocompatibility on MCF-7 cells, and cellular uptake indicate that different amounts of DOX or EPI can be transported to targeting site and released from the internalized carriers. The results demonstrate that as-prepared β-CD-MNPs could be a very promising vehicle for DOX and EPI.

  9. Gene Targeting in Neuroendocrinology.

    Science.gov (United States)

    Candlish, Michael; De Angelis, Roberto; Götz, Viktoria; Boehm, Ulrich

    2015-09-20

    Research in neuroendocrinology faces particular challenges due to the complex interactions between cells in the hypothalamus, in the pituitary gland and in peripheral tissues. Within the hypothalamus alone, attempting to target a specific neuronal cell type can be problematic due to the heterogeneous nature and level of cellular diversity of hypothalamic nuclei. Because of the inherent complexity of the reproductive axis, the use of animal models and in vivo experiments are often a prerequisite in reproductive neuroendocrinology. The advent of targeted genetic modifications, particularly in mice, has opened new avenues of neuroendocrine research. Within this review, we evaluate various mouse models used in reproductive neuroendocrinology and discuss the different approaches to generate genetically modified mice, along with their inherent advantages and disadvantages. We also discuss a variety of versatile genetic tools with a focus on their potential use in reproductive neuroendocrinology.

  10. Strategic development on generic anti-cancer drugs Bevacizumab and Erlotinib Hydrochloride for Harbin Pharmaceutical Group

    Institute of Scientific and Technical Information of China (English)

    Cheung Fat Ping

    2011-01-01

    @@ With improved economy, changing life styles, aging population and health care reform, China had a very potential anti-cancer drug market.The patents of popular anti-cancer drugs Avastin and Tarceva would expire in few years.Generic versions of Avastin and Tarceva were Bevacizumab and Erlotinib Hydrochloride respectively.Harbin Pharmaceutical Group was proposed to develop strategically both generic medicines to enter the high-end anti-cancer drug market for targeted cancer therapies.The vital to success of developing the generic drugs were discussed.

  11. Progress of gene targeting in mouse

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Gene targeting is a powerful approach of study- ing the genefunction in vivo. Specific genetic modifications, including simple gene disruption, point mutations, large chromosomal deletions and rearrangements, targeted incor- poration of foreign genes, could be introduced into the mouse genome by gene targeting. Recent studies make it possible to do the gene targeting with temporal and spatial control.

  12. Anti-cancer activity of an osthole derivative, NBM-T-BMX-OS01: targeting vascular endothelial growth factor receptor signaling and angiogenesis.

    Directory of Open Access Journals (Sweden)

    Hung-Yu Yang

    Full Text Available Angiogenesis occurs during tissue growth, development and wound healing. It is also required for tumor progression and represents a rational target for therapeutic intervention. NBM-T-BMX-OS01 (BMX, derived from the semisynthesis of osthole, an active ingredient isolated from Chinese herb Cnidium monnieri (L. Cuss., was recently shown to enhance learning and memory in rats. In this study, we characterized the anti-angiogenic activities of NBM-T-BMX-OS01 (BMX in an effort to develop novel inhibitors to suppress angiogenesis and tumor growth. BMX inhibited vascular endothelial growth factor (VEGF-induced proliferation, migration and endothelial tube formation in human umbilical endothelial cells (HUVECs. BMX also attenuated VEGF-induced microvessel sprouting from aortic rings ex vivo and reduced HCT116 colorectal cancer cells-induced angiogenesis in vivo. Moreover, BMX inhibited the phosphorylation of VEGFR2, FAK, Akt and ERK in HUVECs exposed to VEGF. BMX was also shown to inhibit HCT116 cell proliferation and to suppress the growth of subcutaneous xenografts of HCT116 cells in vivo. Taken together, this study provides evidence that BMX modulates vascular endothelial cell remodeling and leads to the inhibition of tumor angiogenesis. These results also support the role of BMX as a potential drug candidate and warrant the clinical development in the treatment of cancer.

  13. Anti-cancer activity of an osthole derivative, NBM-T-BMX-OS01: targeting vascular endothelial growth factor receptor signaling and angiogenesis.

    Science.gov (United States)

    Yang, Hung-Yu; Hsu, Ya-Fen; Chiu, Pei-Ting; Ho, Shiau-Jing; Wang, Chi-Han; Chi, Chih-Chin; Huang, Yu-Han; Lee, Cheng-Feng; Li, Ying-Shiuan; Ou, George; Hsu, Ming-Jen

    2013-01-01

    Angiogenesis occurs during tissue growth, development and wound healing. It is also required for tumor progression and represents a rational target for therapeutic intervention. NBM-T-BMX-OS01 (BMX), derived from the semisynthesis of osthole, an active ingredient isolated from Chinese herb Cnidium monnieri (L.) Cuss., was recently shown to enhance learning and memory in rats. In this study, we characterized the anti-angiogenic activities of NBM-T-BMX-OS01 (BMX) in an effort to develop novel inhibitors to suppress angiogenesis and tumor growth. BMX inhibited vascular endothelial growth factor (VEGF)-induced proliferation, migration and endothelial tube formation in human umbilical endothelial cells (HUVECs). BMX also attenuated VEGF-induced microvessel sprouting from aortic rings ex vivo and reduced HCT116 colorectal cancer cells-induced angiogenesis in vivo. Moreover, BMX inhibited the phosphorylation of VEGFR2, FAK, Akt and ERK in HUVECs exposed to VEGF. BMX was also shown to inhibit HCT116 cell proliferation and to suppress the growth of subcutaneous xenografts of HCT116 cells in vivo. Taken together, this study provides evidence that BMX modulates vascular endothelial cell remodeling and leads to the inhibition of tumor angiogenesis. These results also support the role of BMX as a potential drug candidate and warrant the clinical development in the treatment of cancer.

  14. Profound activity of the anti-cancer drug bortezomib against Echinococcus multilocularis metacestodes identifies the proteasome as a novel drug target for cestodes.

    Directory of Open Access Journals (Sweden)

    Britta Stadelmann

    2014-12-01

    Full Text Available A library of 426 FDA-approved drugs was screened for in vitro activity against E. multilocularis metacestodes employing the phosphoglucose isomerase (PGI assay. Initial screening at 20 µM revealed that 7 drugs induced considerable metacestode damage, and further dose-response studies revealed that bortezomib (BTZ, a proteasome inhibitor developed for the chemotherapy of myeloma, displayed high anti-metacestodal activity with an EC50 of 0.6 µM. BTZ treatment of E. multilocularis metacestodes led to an accumulation of ubiquinated proteins and unequivocally parasite death. In-gel zymography assays using E. multilocularis extracts demonstrated BTZ-mediated inhibition of protease activity in a band of approximately 23 kDa, the same size at which the proteasome subunit beta 5 of E. multilocularis could be detected by Western blot. Balb/c mice experimentally infected with E. multilocularis metacestodes were used to assess BTZ treatment, starting at 6 weeks post-infection by intraperitoneal injection of BTZ. This treatment led to reduced parasite weight, but to a degree that was not statistically significant, and it induced adverse effects such as diarrhea and neurological symptoms. In conclusion, the proteasome was identified as a drug target in E. multilocularis metacestodes that can be efficiently inhibited by BTZ in vitro. However, translation of these findings into in vivo efficacy requires further adjustments of treatment regimens using BTZ, or possibly other proteasome inhibitors.

  15. Profound activity of the anti-cancer drug bortezomib against Echinococcus multilocularis metacestodes identifies the proteasome as a novel drug target for cestodes.

    Science.gov (United States)

    Stadelmann, Britta; Aeschbacher, Denise; Huber, Cristina; Spiliotis, Markus; Müller, Joachim; Hemphill, Andrew

    2014-12-01

    A library of 426 FDA-approved drugs was screened for in vitro activity against E. multilocularis metacestodes employing the phosphoglucose isomerase (PGI) assay. Initial screening at 20 µM revealed that 7 drugs induced considerable metacestode damage, and further dose-response studies revealed that bortezomib (BTZ), a proteasome inhibitor developed for the chemotherapy of myeloma, displayed high anti-metacestodal activity with an EC50 of 0.6 µM. BTZ treatment of E. multilocularis metacestodes led to an accumulation of ubiquinated proteins and unequivocally parasite death. In-gel zymography assays using E. multilocularis extracts demonstrated BTZ-mediated inhibition of protease activity in a band of approximately 23 kDa, the same size at which the proteasome subunit beta 5 of E. multilocularis could be detected by Western blot. Balb/c mice experimentally infected with E. multilocularis metacestodes were used to assess BTZ treatment, starting at 6 weeks post-infection by intraperitoneal injection of BTZ. This treatment led to reduced parasite weight, but to a degree that was not statistically significant, and it induced adverse effects such as diarrhea and neurological symptoms. In conclusion, the proteasome was identified as a drug target in E. multilocularis metacestodes that can be efficiently inhibited by BTZ in vitro. However, translation of these findings into in vivo efficacy requires further adjustments of treatment regimens using BTZ, or possibly other proteasome inhibitors.

  16. 靶向血管内皮生长因子及其受体的抗肿瘤药物研究进展%Anti-cancer drugs targeting vascular endothelial growth factors and receptors: research advances

    Institute of Scientific and Technical Information of China (English)

    张娜; 姚文兵; 徐晨

    2012-01-01

    Angiogenesis plays a critical rede in the process of tumor growth and metastasis, and vascular endothelial growth factor and its' receptor (VEGF/VEGFR) signaling pathway is an important mechanism of neovascularization, At present, drug inhibition of angiogenesis has become a significant research topic and a variety of anti-angiogenesis agents aimed at blocking VECF or its receptor-signaling system have been marketed or issued to the clinical trials. The main purpose of this review is to summarize the available information regarding the importance of VEGF/VEGFR in cancer therapy, with a focus on the latest development, clinical use and challenges of the anti-cancer drugs targeting VEGF/VEGFR.%血管生成对肿瘤的生长和转移起着关键作用,血管内皮生长因子(VEGF)及其受体信号通路是调节肿瘤新生血管生成的重要途径,因此,近年来以VEGF及其受体为作用靶标的抗肿瘤血管生成治疗已经成为研究热点,目前已有多种药物上市或处于临床试验阶段.本文主要综述了VEGF及其受体在肿瘤血管生成调节机制中的作用,同时着重介绍靶向VEGF及其受体的抗肿瘤药物的新近研究进展、临床应用及存在的问题.

  17. PEGylation in anti-cancer therapy: An overview

    Directory of Open Access Journals (Sweden)

    Prajna Mishra

    2016-06-01

    Full Text Available Advanced drug delivery systems using poly(ethylene glycol (PEG is an important development in anti-cancer therapy. PEGylation has the ability to enhance the retention time of the therapeutics like proteins, enzymes small molecular drugs, liposomes and nanoparticles by protecting them against various degrading mechanisms active inside a tissue or cell, which consequently improves their therapeutic potential. PEGylation effectively alters the pharmacokinetics (PK of a variety of drugs and dramatically improves the pharmaceutical values; recent development of which includes fabrication of stimuli-sensitive polymers/smart polymers and polymeric micelles to cope of with the pathophysiological environment of targeted site with less toxic effects and more effectiveness. This overview discusses PEGylation involving proteins, enzymes, low molecular weight drugs, liposomes and nanoparticles that has been developed, clinically tried for anti-cancer therapy during the last decade.

  18. Copper and conquer: copper complexes of di-2-pyridylketone thiosemicarbazones as novel anti-cancer therapeutics.

    Science.gov (United States)

    Park, Kyung Chan; Fouani, Leyla; Jansson, Patric J; Wooi, Danson; Sahni, Sumit; Lane, Darius J R; Palanimuthu, Duraippandi; Lok, Hiu Chuen; Kovačević, Zaklina; Huang, Michael L H; Kalinowski, Danuta S; Richardson, Des R

    2016-09-01

    Copper is an essential trace metal required by organisms to perform a number of important biological processes. Copper readily cycles between its reduced Cu(i) and oxidised Cu(ii) states, which makes it redox active in biological systems. This redox-cycling propensity is vital for copper to act as a catalytic co-factor in enzymes. While copper is essential for normal physiology, enhanced copper levels in tumours leads to cancer progression. In particular, the stimulatory effect of copper on angiogenesis has been established in the last several decades. Additionally, it has been demonstrated that copper affects tumour growth and promotes metastasis. Based on the effects of copper on cancer progression, chelators that bind copper have been developed as anti-cancer agents. In fact, a novel class of thiosemicarbazone compounds, namely the di-2-pyridylketone thiosemicarbazones that bind copper, have shown great promise in terms of their anti-cancer activity. These agents have a unique mechanism of action, in which they form redox-active complexes with copper in the lysosomes of cancer cells. Furthermore, these agents are able to overcome P-glycoprotein (P-gp) mediated multi-drug resistance (MDR) and act as potent anti-oncogenic agents through their ability to up-regulate the metastasis suppressor protein, N-myc downstream regulated gene-1 (NDRG1). This review provides an overview of the metabolism and regulation of copper in normal physiology, followed by a discussion of the dysregulation of copper homeostasis in cancer and the effects of copper on cancer progression. Finally, recent advances in our understanding of the mechanisms of action of anti-cancer agents targeting copper are discussed.

  19. Phenethyl isothiocyanate: a comprehensive review of anti-cancer mechanisms.

    Science.gov (United States)

    Gupta, Parul; Wright, Stephen E; Kim, Sung-Hoon; Srivastava, Sanjay K

    2014-12-01

    The epidemiological evidence suggests a strong inverse relationship between dietary intake of cruciferous vegetables and the incidence of cancer. Among other constituents of cruciferous vegetables, isothiocyanates (ITC) are the main bioactive chemicals present. Phenethyl isothiocyanate (PEITC) is present as gluconasturtiin in many cruciferous vegetables with remarkable anti-cancer effects. PEITC is known to not only prevent the initiation phase of carcinogenesis process but also to inhibit the progression of tumorigenesis. PEITC targets multiple proteins to suppress various cancer-promoting mechanisms such as cell proliferation, progression and metastasis. Pre-clinical evidence suggests that combination of PEITC with conventional anti-cancer agents is also highly effective in improving overall efficacy. Based on accumulating evidence, PEITC appears to be a promising agent for cancer therapy and is already under clinical trials for leukemia and lung cancer. This is the first review which provides a comprehensive analysis of known targets and mechanisms along with a critical evaluation of PEITC as a future anti-cancer agent.

  20. Recent insights in nanotechnology-based drugs and formulations designed for effective anti-cancer therapy.

    Science.gov (United States)

    Piktel, Ewelina; Niemirowicz, Katarzyna; Wątek, Marzena; Wollny, Tomasz; Deptuła, Piotr; Bucki, Robert

    2016-05-26

    The rapid development of nanotechnology provides alternative approaches to overcome several limitations of conventional anti-cancer therapy. Drug targeting using functionalized nanoparticles to advance their transport to the dedicated site, became a new standard in novel anti-cancer methods. In effect, the employment of nanoparticles during design of antineoplastic drugs helps to improve pharmacokinetic properties, with subsequent development of high specific, non-toxic and biocompatible anti-cancer agents. However, the physicochemical and biological diversity of nanomaterials and a broad spectrum of unique features influencing their biological action requires continuous research to assess their activity. Among numerous nanosystems designed to eradicate cancer cells, only a limited number of them entered the clinical trials. It is anticipated that progress in development of nanotechnology-based anti-cancer materials will provide modern, individualized anti-cancer therapies assuring decrease in morbidity and mortality from cancer diseases. In this review we discussed the implication of nanomaterials in design of new drugs for effective antineoplastic therapy and describe a variety of mechanisms and challenges for selective tumor targeting. We emphasized the recent advantages in the field of nanotechnology-based strategies to fight cancer and discussed their part in effective anti-cancer therapy and successful drug delivery.

  1. Potential anti-cancer drugs commonly used for other indications.

    Science.gov (United States)

    Hanusova, Veronika; Skalova, Lenka; Kralova, Vera; Matouskova, Petra

    2015-01-01

    An increasing resistance of mammalian tumor cells to chemotherapy along with the severe side effects of commonly used cytostatics has raised the urgency in the search for new anti-cancer agents. Several drugs originally approved for indications other than cancer treatment have recently been found to have a cytostatic effect on cancer cells. These drugs could be expediently repurposed as anti-cancer agents, since they have already been tested for toxicity in humans and animals. The groups of newly recognized potential cytostatics discussed in this review include benzimidazole anthelmintics (albendazole, mebendazole, flubendazole), anti-hypertensive drugs (doxazosin, propranolol), psychopharmaceuticals (chlorpromazine, clomipramine) and antidiabetic drugs (metformin, pioglitazone). All these drugs have a definite potential to be used especially in combinations with other cytostatics; the chemotherapy targeting of multiple sites now represents a promising approach in cancer treatment. The present review summarizes recent information about the anti-cancer effects of selected drugs commonly used for other medical indications. Our aim is not to collect all the reported results, but to present an overview of various possibilities. Advantages, disadvantages and further perspectives regarding individual drugs are discussed and evaluated.

  2. Targeted gene flow for conservation.

    Science.gov (United States)

    Kelly, Ella; Phillips, Ben L

    2016-04-01

    Anthropogenic threats often impose strong selection on affected populations, causing rapid evolutionary responses. Unfortunately, these adaptive responses are rarely harnessed for conservation. We suggest that conservation managers pay close attention to adaptive processes and geographic variation, with an eye to using them for conservation goals. Translocating pre-adapted individuals into recipient populations is currently considered a potentially important management tool in the face of climate change. Targeted gene flow, which involves moving individuals with favorable traits to areas where these traits would have a conservation benefit, could have a much broader application in conservation. Across a species' range there may be long-standing geographic variation in traits or variation may have rapidly developed in response to a threatening process. Targeted gene flow could be used to promote natural resistance to threats to increase species resilience. We suggest that targeted gene flow is a currently underappreciated strategy in conservation that has applications ranging from the management of invasive species and their impacts to controlling the impact and virulence of pathogens.

  3. Structure Identification and Anti-Cancer Pharmacological Prediction of Triterpenes from Ganoderma lucidum

    Directory of Open Access Journals (Sweden)

    Yanyan Shao

    2016-05-01

    Full Text Available Ganoderma triterpenes (GTs are the major secondary metabolites of Ganoderma lucidum, which is a popularly used traditional Chinese medicine for complementary cancer therapy. In the present study, systematic isolation, and in silico pharmacological prediction are implemented to discover potential anti-cancer active GTs from G. lucidum. Nineteen GTs, three steroids, one cerebroside, and one thymidine were isolated from G. lucidum. Six GTs were first isolated from the fruiting bodies of G. lucidum, including 3β,7β,15β-trihydroxy-11,23-dioxo-lanost-8,16-dien-26-oic acid methyl ester (1, 3β,7β,15β-trihydroxy-11,23-dioxo-lanost-8,16-dien-26-oic acid (2, 3β,7β,15α,28-tetrahydroxy-11,23-dioxo-lanost-8,16-dien-26-oic acid (3, ganotropic acid (4, 26-nor-11,23-dioxo-5α-lanost-8-en-3β,7β,15α,25-tetrol (5 and (3β,7α-dihydroxy-lanosta-8,24-dien- 11-one (6. (4E,8E-N-d-2′-hydroxypalmitoyl-l-O-β-d-glucopyranosyl-9-methyl-4,8-spingodienine (7, and stigmasta-7,22-dien-3β,5α,6α-triol (8 were first reported from the genus Ganodema. By using reverse pharmacophoric profiling of the six GTs, thirty potential anti-cancer therapeutic targets were identified and utilized to construct their ingredient-target interaction network. Then nineteen high frequency targets of GTs were selected from thirty potential targets to construct a protein interaction network (PIN. In order to cluster the pharmacological activity of GTs, twelve function modules were identified by molecular complex detection (MCODE and gene ontology (GO enrichment analysis. The results indicated that anti-cancer effect of GTs might be related to histone acetylation and interphase of mitotic cell cycle by regulating general control non-derepressible 5 (GCN5 and cyclin-dependent kinase-2 (CDK2, respectively. This research mode of extraction, isolation, pharmacological prediction, and PIN analysis might be beneficial to rapidly predict and discover pharmacological activities of novel

  4. Structure Identification and Anti-Cancer Pharmacological Prediction of Triterpenes from Ganoderma lucidum.

    Science.gov (United States)

    Shao, Yanyan; Qiao, Liansheng; Wu, Lingfang; Sun, Xuefei; Zhu, Dan; Yang, Guanghui; Zhang, Xiaoxue; Mao, Xin; Chen, Wenjing; Liang, Wenyi; Zhang, Yanling; Zhang, Lanzhen

    2016-05-21

    Ganoderma triterpenes (GTs) are the major secondary metabolites of Ganoderma lucidum, which is a popularly used traditional Chinese medicine for complementary cancer therapy. In the present study, systematic isolation, and in silico pharmacological prediction are implemented to discover potential anti-cancer active GTs from G. lucidum. Nineteen GTs, three steroids, one cerebroside, and one thymidine were isolated from G. lucidum. Six GTs were first isolated from the fruiting bodies of G. lucidum, including 3β,7β,15β-trihydroxy-11,23-dioxo-lanost-8,16-dien-26-oic acid methyl ester (1), 3β,7β,15β-trihydroxy-11,23-dioxo-lanost-8,16-dien-26-oic acid (2), 3β,7β,15α,28-tetrahydroxy-11,23-dioxo-lanost-8,16-dien-26-oic acid (3), ganotropic acid (4), 26-nor-11,23-dioxo-5α-lanost-8-en-3β,7β,15α,25-tetrol (5) and (3β,7α)-dihydroxy-lanosta-8,24-dien- 11-one (6). (4E,8E)-N-d-2'-hydroxypalmitoyl-l-O-β-d-glucopyranosyl-9-methyl-4,8-spingodienine (7), and stigmasta-7,22-dien-3β,5α,6α-triol (8) were first reported from the genus Ganodema. By using reverse pharmacophoric profiling of the six GTs, thirty potential anti-cancer therapeutic targets were identified and utilized to construct their ingredient-target interaction network. Then nineteen high frequency targets of GTs were selected from thirty potential targets to construct a protein interaction network (PIN). In order to cluster the pharmacological activity of GTs, twelve function modules were identified by molecular complex detection (MCODE) and gene ontology (GO) enrichment analysis. The results indicated that anti-cancer effect of GTs might be related to histone acetylation and interphase of mitotic cell cycle by regulating general control non-derepressible 5 (GCN5) and cyclin-dependent kinase-2 (CDK2), respectively. This research mode of extraction, isolation, pharmacological prediction, and PIN analysis might be beneficial to rapidly predict and discover pharmacological activities of novel compounds.

  5. Methods for predicting anti-cancer response

    DEFF Research Database (Denmark)

    2013-01-01

    The present invention relates to methods for predicting response of a cancer in a subject to anti-cancer therapies based upon a determination and analysis of a chromosomal aberration score, such as the number of allelic imbalance or the number of telomeric allelic imbalance in the chromosomes...

  6. Are isothiocyanates potential anti-cancer drugs?

    Institute of Scientific and Technical Information of China (English)

    Xiang WU; Qing-hua ZHOU; Ke XU

    2009-01-01

    Isothiocyanates are naturally occurring small molecules that are formed from glucosinolate precursors of cruciferous vegetables. Many isothiocyanates, both natural and synthetic, display anticarcinogenic activity because they reduce activation of carcinogens and increase their detoxification. Recent studies show that they exhibit anti-tumor activity by affecting multiple pathways including apoptosis, MAPK signaling, oxidative stress, and cell cycle progression. This review summarizes the current knowledge on isothiocyanates and focuses on their role as potential anti-cancer agents.

  7. The promising alliance of anti-cancer electrochemotherapy with immunotherapy.

    Science.gov (United States)

    Calvet, Christophe Y; Mir, Lluis M

    2016-06-01

    Anti-tumor electrochemotherapy, which consists in increasing anti-cancer drug uptake by means of electroporation, is now implanted in about 140 cancer treatment centers in Europe. Its use is supported by the English National Institute for Health and Care Excellence for the palliative treatment of skin metastases, and about 13,000 cancer patients were treated by this technology by the end of 2015. Efforts are now focused on turning this local anti-tumor treatment into a systemic one. Electrogenetherapy, that is the electroporation-mediated transfer of therapeutic genes, is currently under clinical evaluation and has brought excitement to enlarge the anti-cancer armamentarium. Among the promising electrogenetherapy strategies, DNA vaccination and cytokine-based immunotherapy aim at stimulating anti-tumor immunity. We review here the interests and state of development of both electrochemotherapy and electrogenetherapy. We then emphasize the potent beneficial outcome of the combination of electrochemotherapy with immunotherapy, such as immune checkpoint inhibitors or strategies based on electrogenetherapy, to simultaneously achieve excellent local debulking anti-tumor responses and systemic anti-metastatic effects.

  8. The drug target genes show higher evolutionary conservation than non-target genes.

    Science.gov (United States)

    Lv, Wenhua; Xu, Yongdeng; Guo, Yiying; Yu, Ziqi; Feng, Guanglong; Liu, Panpan; Luan, Meiwei; Zhu, Hongjie; Liu, Guiyou; Zhang, Mingming; Lv, Hongchao; Duan, Lian; Shang, Zhenwei; Li, Jin; Jiang, Yongshuai; Zhang, Ruijie

    2016-01-26

    Although evidence indicates that drug target genes share some common evolutionary features, there have been few studies analyzing evolutionary features of drug targets from an overall level. Therefore, we conducted an analysis which aimed to investigate the evolutionary characteristics of drug target genes. We compared the evolutionary conservation between human drug target genes and non-target genes by combining both the evolutionary features and network topological properties in human protein-protein interaction network. The evolution rate, conservation score and the percentage of orthologous genes of 21 species were included in our study. Meanwhile, four topological features including the average shortest path length, betweenness centrality, clustering coefficient and degree were considered for comparison analysis. Then we got four results as following: compared with non-drug target genes, 1) drug target genes had lower evolutionary rates; 2) drug target genes had higher conservation scores; 3) drug target genes had higher percentages of orthologous genes and 4) drug target genes had a tighter network structure including higher degrees, betweenness centrality, clustering coefficients and lower average shortest path lengths. These results demonstrate that drug target genes are more evolutionarily conserved than non-drug target genes. We hope that our study will provide valuable information for other researchers who are interested in evolutionary conservation of drug targets.

  9. Double layered hydroxides as potential anti-cancer drug delivery agents.

    Science.gov (United States)

    Riaz, Ufana; Ashraf, S M

    2013-04-01

    The emergence of nanotechnology has changed the scenario of the medical world by revolutionizing the diagnosis, monitoring and treatment of cancer. This nanotechnology has been proved miraculous in detecting cancer cells, delivering chemotherapeutic agents and monitoring treatment from non-specific to highly targeted killing of tumor cells. In the past few decades, a number of inorganic materials have been investigated such as calcium phosphate, gold, carbon materials, silicon oxide, iron oxide, and layered double hydroxide (LDH) for examining their efficacy in targeting drug delivery. The reason behind the selection of these inorganic materials was their versatile and unique features efficient in drug delivery, such as wide availability, rich surface functionality, good biocompatibility, potential for target delivery, and controlled release of the drug from these inorganic nanomaterials. Although, the drug-LDH hybrids are found to be quite instrumental because of their application as advanced anti-cancer drug delivery systems, there has not been much research on them. This mini review is set to highlight the advancement made in the use of layered double hydroxides (LDHs) as anti-cancer drug delivery agents. Along with the advantages of LDHs as anti-cancer drug delivery agents, the process of interaction of some of the common anti-cancer drugs with LDH has also been discussed.

  10. Anti-Cancer Properties of the Naturally Occurring Aphrodisiacs: Icariin and Its Derivatives

    Science.gov (United States)

    Tan, Hui-Li; Chan, Kok-Gan; Pusparajah, Priyia; Saokaew, Surasak; Duangjai, Acharaporn; Lee, Learn-Han; Goh, Bey-Hing

    2016-01-01

    Epimedium (family Berberidaceae), commonly known as Horny Goat Weed or Yin Yang Huo, is commonly used as a tonic, aphrodisiac, anti-rheumatic and anti-cancer agent in traditional herbal formulations in Asian countries such as China, Japan, and Korea. The major bioactive compounds present within this plant include icariin, icaritin and icariside II. Although it is best known for its aphrodisiac properties, scientific and pharmacological studies suggest it possesses broad therapeutic capabilities, especially for enhancing reproductive function and osteoprotective, neuroprotective, cardioprotective, anti-inflammatory and immunoprotective effects. In recent years, there has been great interest in scientific investigation of the purported anti-cancer properties of icariin and its derivatives. Data from in vitro and in vivo studies suggests these compounds demonstrate anti-cancer activity against a wide range of cancer cells which occurs through various mechanisms such as apoptosis, cell cycle modulation, anti-angiogenesis, anti-metastasis and immunomodulation. Of note, they are efficient at targeting cancer stem cells and drug-resistant cancer cells. These are highly desirable properties to be emulated in the development of novel anti-cancer drugs in combatting the emergence of drug resistance and overcoming the limited efficacy of current standard treatment. This review aims to summarize the anti-cancer mechanisms of icariin and its derivatives with reference to the published literature. The currently utilized applications of icariin and its derivatives in cancer treatment are explored with reference to existing patents. Based on the data compiled, icariin and its derivatives are shown to be compounds with tremendous potential for the development of new anti-cancer drugs. PMID:27445824

  11. Anti-Cancer Properties of the Naturally Occurring Aphrodisiacs: Icariin and Its Derivatives.

    Science.gov (United States)

    Tan, Hui-Li; Chan, Kok-Gan; Pusparajah, Priyia; Saokaew, Surasak; Duangjai, Acharaporn; Lee, Learn-Han; Goh, Bey-Hing

    2016-01-01

    Epimedium (family Berberidaceae), commonly known as Horny Goat Weed or Yin Yang Huo, is commonly used as a tonic, aphrodisiac, anti-rheumatic and anti-cancer agent in traditional herbal formulations in Asian countries such as China, Japan, and Korea. The major bioactive compounds present within this plant include icariin, icaritin and icariside II. Although it is best known for its aphrodisiac properties, scientific and pharmacological studies suggest it possesses broad therapeutic capabilities, especially for enhancing reproductive function and osteoprotective, neuroprotective, cardioprotective, anti-inflammatory and immunoprotective effects. In recent years, there has been great interest in scientific investigation of the purported anti-cancer properties of icariin and its derivatives. Data from in vitro and in vivo studies suggests these compounds demonstrate anti-cancer activity against a wide range of cancer cells which occurs through various mechanisms such as apoptosis, cell cycle modulation, anti-angiogenesis, anti-metastasis and immunomodulation. Of note, they are efficient at targeting cancer stem cells and drug-resistant cancer cells. These are highly desirable properties to be emulated in the development of novel anti-cancer drugs in combatting the emergence of drug resistance and overcoming the limited efficacy of current standard treatment. This review aims to summarize the anti-cancer mechanisms of icariin and its derivatives with reference to the published literature. The currently utilized applications of icariin and its derivatives in cancer treatment are explored with reference to existing patents. Based on the data compiled, icariin and its derivatives are shown to be compounds with tremendous potential for the development of new anti-cancer drugs.

  12. Targeting Gene-Virotherapy for Cancer

    Institute of Scientific and Technical Information of China (English)

    Xin-Yuan LIU; Jing-Fa GU; Wen-Fang SHI

    2005-01-01

    Gene therapy and viral therapy for cancer have therapeutic effects, but there has been no significant breakthrough in these two forms of therapy. Therefore, a new strategy called "targeting genevirotherapy", which combines the advantages of gene therapy and viral therapy, has been formulated. This new therapy has stronger antitumor effects than either gene therapy or viral therapy. A tumor-specific replicative adenovirus vector ZD55 (E1B55KD deleted Adv.) was constructed and various single therapeutic genes were inserted into ZD55 to form ZD55-gene. These are the targeting gene-virotherapy genes. But experiments showed that a single gene was not effective in eliminating the tumor mass, and therefore two genes were separately inserted into ZD55. This strategy is called "targeting dual gene-virotherapy" (with PCT patent). Better results were obtained with this strategy, and all the xenograft tumor masses were completely eliminated in all mice when two suitable genes producing a synergetic or compensative effect were chosen. Twenty-six papers on these strategies have been published by researchers in our laboratory.Furthermore, an adenoviral vector with two targeting promoters harboring two antitumor genes has been constructed for cancer therapy. Promising results have been obtained with this adenoviral vectorand another patent has been applied for. This antitumor strategy can be used to kill tumor cells completely with minimum damage to normal cells.

  13. Targeting tumor suppressor genes for cancer therapy.

    Science.gov (United States)

    Liu, Yunhua; Hu, Xiaoxiao; Han, Cecil; Wang, Liana; Zhang, Xinna; He, Xiaoming; Lu, Xiongbin

    2015-12-01

    Cancer drugs are broadly classified into two categories: cytotoxic chemotherapies and targeted therapies that specifically modulate the activity of one or more proteins involved in cancer. Major advances have been achieved in targeted cancer therapies in the past few decades, which is ascribed to the increasing understanding of molecular mechanisms for cancer initiation and progression. Consequently, monoclonal antibodies and small molecules have been developed to interfere with a specific molecular oncogenic target. Targeting gain-of-function mutations, in general, has been productive. However, it has been a major challenge to use standard pharmacologic approaches to target loss-of-function mutations of tumor suppressor genes. Novel approaches, including synthetic lethality and collateral vulnerability screens, are now being developed to target gene defects in p53, PTEN, and BRCA1/2. Here, we review and summarize the recent findings in cancer genomics, drug development, and molecular cancer biology, which show promise in targeting tumor suppressors in cancer therapeutics.

  14. Anti-cancer effects of traditional Korean wild vegetables in complementary and alternative medicine.

    Science.gov (United States)

    Ju, Hyun-Mok; Yu, Kwang-Won; Cho, Sung-Dae; Cheong, Sun Hee; Kwon, Ki Han

    2016-02-01

    This research study explored the anti-cancer effects of natural materials in South Korea. Although South Korea has a long history of traditional medicine, many natural materials of South Korea have not yet been introduced to the rest of the world because of language barriers and inconsistent study conditions. In the past 3 years, 56 papers introducing 56 natural materials, which have anti-cancer effects, have been published by scientists in South Korea. Further, these studies have introduced five kinds of natural materials presented in research papers that were written in Korean and are therefore virtually unknown overseas. The anti-cancer effects were confirmed by 2-3 cancer markers in the majority of the studies, with the most common targets being breast cancer cells and gastric cancer cells. These cancers have the greatest incidence in South Korea. The natural materials studied not only exhibit anti-cancer activity but also display anti-inflammatory, anti-oxidative stress, and anti-diabetic activities. They have not yet been used for the direct treatment of disease but have potential as medicinal materials for alternative and complementary medicine for the treatment of many modern diseases. Many natural materials of South Korea are already known all over the world, and with this study, we hope to further future research to learn more about these natural medicines.

  15. Targeted gene repair – in the arena

    OpenAIRE

    2003-01-01

    The development of targeted gene repair is under way and, despite some setbacks, shows promise as an alternative form of gene therapy. This approach uses synthetic DNA molecules to activate and direct the cell’s inherent DNA repair systems to correct inborn errors. The progress of this technique and its therapeutic potential are discussed in relation to the treatment of genetic diseases.

  16. Magnetic targeting strategies in gene delivery.

    Science.gov (United States)

    Delyagina, Evgenya; Li, Wenzhong; Ma, Nan; Steinhoff, Gustav

    2011-11-01

    Gene delivery is a process of the insertion of transgenes into cells with the purpose to obtain the expression of encoded protein. The therapeutic application of this process is termed gene therapy, which is becoming a promising instrument to treat genetic and acquired diseases. Although numerous methods of gene transfer have already been developed, including biological, physical and chemical approaches, the optimal strategy has to be discovered. Importantly, it should be effective, selective and safe to be translated to the clinic. Magnetic targeting has been demonstrated as an effective strategy to decrease side effects of gene transfer, while increasing the selectivity and efficiency of the applied vector. This article will focus on the latest progress in the development of different magnetic vectors, based on both viral and nonviral gene delivery agents. It will also include a description of magnetic targeting applications in stem cells and in vivo, which has gained interest in recent years due to the rapid development of technology.

  17. Targeting Herpetic Keratitis by Gene Therapy

    Directory of Open Access Journals (Sweden)

    Hossein Mostafa Elbadawy

    2012-01-01

    Full Text Available Ocular gene therapy is rapidly becoming a reality. By November 2012, approximately 28 clinical trials were approved to assess novel gene therapy agents. Viral infections such as herpetic keratitis caused by herpes simplex virus 1 (HSV-1 can cause serious complications that may lead to blindness. Recurrence of the disease is likely and cornea transplantation, therefore, might not be the ideal therapeutic solution. This paper will focus on the current situation of ocular gene therapy research against herpetic keratitis, including the use of viral and nonviral vectors, routes of delivery of therapeutic genes, new techniques, and key research strategies. Whereas the correction of inherited diseases was the initial goal of the field of gene therapy, here we discuss transgene expression, gene replacement, silencing, or clipping. Gene therapy of herpetic keratitis previously reported in the literature is screened emphasizing candidate gene therapy targets. Commonly adopted strategies are discussed to assess the relative advantages of the protective therapy using antiviral drugs and the common gene therapy against long-term HSV-1 ocular infections signs, inflammation and neovascularization. Successful gene therapy can provide innovative physiological and pharmaceutical solutions against herpetic keratitis.

  18. Selective anti-cancer agents as anti-aging drugs.

    Science.gov (United States)

    Blagosklonny, Mikhail V

    2013-12-01

    Recent groundbreaking discoveries have revealed that IGF-1, Ras, MEK, AMPK, TSC1/2, FOXO, PI3K, mTOR, S6K, and NFκB are involved in the aging process. This is remarkable because the same signaling molecules, oncoproteins and tumor suppressors, are well-known targets for cancer therapy. Furthermore, anti-cancer drugs aimed at some of these targets have been already developed. This arsenal could be potentially employed for anti-aging interventions (given that similar signaling molecules are involved in both cancer and aging). In cancer, intrinsic and acquired resistance, tumor heterogeneity, adaptation, and genetic instability of cancer cells all hinder cancer-directed therapy. But for anti-aging applications, these hurdles are irrelevant. For example, since anti-aging interventions should be aimed at normal postmitotic cells, no selection for resistance is expected. At low doses, certain agents may decelerate aging and age-related diseases. Importantly, deceleration of aging can in turn postpone cancer, which is an age-related disease.

  19. Gene targeting in adult rhesus macaque fibroblasts

    Directory of Open Access Journals (Sweden)

    Wolf Don P

    2008-03-01

    Full Text Available Abstract Background Gene targeting in nonhuman primates has the potential to produce critical animal models for translational studies related to human diseases. Successful gene targeting in fibroblasts followed by somatic cell nuclear transfer (SCNT has been achieved in several species of large mammals but not yet in primates. Our goal was to establish the protocols necessary to achieve gene targeting in primary culture of adult rhesus macaque fibroblasts as a first step in creating nonhuman primate models of genetic disease using nuclear transfer technology. Results A primary culture of adult male fibroblasts was transfected with hTERT to overcome senescence and allow long term in vitro manipulations. Successful gene targeting of the HPRT locus in rhesus macaques was achieved by electroporating S-phase synchronized cells with a construct containing a SV40 enhancer. Conclusion The cell lines reported here could be used for the production of null mutant rhesus macaque models of human genetic disease using SCNT technology. In addition, given the close evolutionary relationship and biological similarity between rhesus macaques and humans, the protocols described here may prove useful in the genetic engineering of human somatic cells.

  20. Anti-cancer activities of diospyrin, its derivatives and analogues

    KAUST Repository

    Sagar, Sunil

    2010-09-01

    Natural products have played a vital role in drug discovery and development process for cancer. Diospyrin, a plant based bisnaphthoquinonoid, has been used as a lead molecule in an effort to develop anti-cancer drugs. Several derivatives/analogues have been synthesized and screened for their pro-apoptotic/anti-cancer activities so far. Our review is focused on the pro-apoptotic/anti-cancer activities of diospyrin, its derivatives/analogues and the different mechanisms potentially involved in the bioactivity of these compounds. Particular focus has been placed on the different mechanisms (both chemical and molecular) thought to underlie the bioactivity of these compounds. A brief bioinformatics analysis at the end of the article provides novel insights into the new potential mechanisms and pathways by which these compounds might exert their effects and lead to a better realization of the full therapeutic potential of these compounds as anti-cancer drugs. © 2010 Elsevier Masson SAS. All rights reserved.

  1. The anti-cancer activity of noscapine: a review.

    Science.gov (United States)

    Mahmoudian, Massoud; Rahimi-Moghaddam, Parvaneh

    2009-01-01

    Noscapine is an isoqiunoline alkaloid found in opium latex. Unlike most other alkaloids obtained from opium latex, noscapine is not sedative and has been used as antitussive drug in various countries. Recently, it has been introduced as an anti-mitotic agent. This drug can be used orally. When the resistance to other anti-cancer drugs such as paclitaxel manifests, noscapine might be effective. Therefore, noscapine and its analogs have great potential as novel anti-cancer agents.

  2. The anti-cancerous drug doxorubicin decreases the c-di-GMP content in Pseudomonas aeruginosa but promotes biofilm formation

    DEFF Research Database (Denmark)

    Groizeleau, Julie; Rybtke, Morten; Andersen, Jens Bo

    2016-01-01

    for their potential c-di-GMP-lowering effect using a recently developed c-di-GMP biosensor strain. Our screen identified the anti-cancerous drug doxorubicin as a potent c-di-GMP inhibitor. In addition, the drug decreased the transcription of many biofilm-related genes. However, despite its effect on the c...

  3. Potential Anti-cancer Activity of Furanodiene

    Institute of Scientific and Technical Information of China (English)

    Zhen-zhen Ba; Yan-ping Zheng; Hui Zhang; Xiu-yan Sun; Dong-hai Lin

    2009-01-01

    Objective: To study the anti-tumor activities of furanodiene (C15H20O), a primary sesquiterpene compound isolated from the essential oil of the rhizome of Curcuma wenyujin YH Chen et C. Ling(Wen Ezhu), in vitro and in vivo.Methods: In vitro MTT assay was used to further study the effects of time and dosage on anti-proliferation of furanodiene against the sensitive Hela, Hep-2,HL-60, U251 cells, based on the cytotoxic effects of furanodiene on 12 human malignant tumor cell lines with the essential oil of Wen Ezhu as control., and the half-inhibitory concentration (IC50) was observed. In vivo uterine cervix (U14) tumor cell was selected and the conventional assay method of anti-tumor activity was employed. Furanodiene liposome was administered intraperitoneally, and tumor-inhibitory rate, thymus and spleen indexes were observed.Results: The inhibitive effects on cell proliferation were shown in all of the twelve cell lines and the cytotoxic effects of furanodiene against Hela, Hep-2, HL-60, U251 cells were observed after 12 h of administration, the effect could last for at least 48 h in a dose dependent manner, and the IC50 values were 0.6, 1.7, 1.8, 7.0 μg/ml, respectively. Furanodiene was also found to show inhibitive effects on the proliferation of uterine cervix (U14) tumor induced in mice. The tumor inhibition rates were 36.09% (40 mg/kg), 41.55% (60 mg/kg), 58.29% (80 mg/kg), respectively.Conclusion: Furanodiene is one of primary anti-cancer active components in the essential oil of Wen Ezhu, and also a very effective agent against uterine cervix cancer, and has protection effect on the immune function.

  4. Glycan changes: cancer metastasis and anti-cancer vaccines

    Indian Academy of Sciences (India)

    Min Li; Lujun Song; Xinyu Qin

    2010-12-01

    Complex carbohydrates, which are major components of the cell membrane, perform important functions in cell–cell and cell–extracellular matrix interactions, as well as in signal transduction. They comprise three kinds of biomolecules: glycoproteins, proteoglycans and glycosphingolipids. Recent studies have also shown that glycan changes in malignant cells take a variety of forms and mediate key pathophysiological events during the various stages of tumour progression. Glycosylation changes are universal hallmarks of malignant transformation and tumour progression in human cancer, which take place on the whole cells or some specific molecules. Accordingly, those changes make them prominent candidates for cancer biomarkers in the meantime. This review mainly focuses on the correlation between glycosylation and the metastasis potential of tumour cells from comprehensive aspects to further address the vital roles of glycans in oncogenesising. Moreover, utilizing these glycosylation changes to ward off tumour metastasis by means of anti-adhesion approach or devising anti-cancer vaccine is one of promising targets of future study.

  5. Nanosuspension for the delivery of a poorly soluble anti-cancer kinase inhibitor.

    Science.gov (United States)

    Danhier, Fabienne; Ucakar, Bernard; Vanderhaegen, Marie-Lyse; Brewster, Marcus E; Arien, Tina; Préat, Véronique

    2014-09-01

    We hypothesized that nanosuspensions could be promising for the delivery of the poorly water soluble anti-cancer multi-targeted kinase inhibitor, MTKi-327. Hence, the aims of this work were (i) to evaluate the MTKi-327 nanosuspension for parenteral and oral administrations and (ii) to compare this nanosuspension with other nanocarriers in terms of anti-cancer efficacy and pharmacokinetics. Therefore, four formulations of MTKi-327 were studied: (i) PEGylated PLGA-based nanoparticles, (ii) self-assembling PEG₇₅₀-p-(CL-co-TMC) polymeric micelles, (iii) nanosuspensions of MTKi-327; and (iv) Captisol solution (pH=3.5). All the nano-formulations presented a size below 200 nm. Injections of the highest possible dose of the three nano-formulations did not induce any side effects in mice. In contrast, the maximum tolerated dose of the control Captisol solution was 20-fold lower than its highest possible dose. The highest regrowth delay of A-431-tumor-bearing nude mice was obtained with MTKi-327 nanosuspension, administered intravenously, at a dose of 650 mg/kg. After intravenous and oral administration, the AUC₀₋∞ of MTKi-327 nanosuspension was 2.4-fold greater than that of the Captisol solution. Nanosuspension may be considered as an effective anti-cancer MTKi-327 delivery method due to (i) the higher MTKi-327 maximum tolerated dose, (ii) the possible intravenous injection of MTKi-327, (iii) its ability to enhance the administered dose and (iv) its higher efficacy.

  6. Recombinant fungal entomopathogen RNAi target insect gene.

    Science.gov (United States)

    Hu, Qiongbo; Wu, Wei

    2016-11-01

    RNA interference (RNAi) technology is considered as an alternative for control of pests. However, RNAi has not been used in field conditions yet, since delivering exogenous ds/siRNA to target pests is very difficult. The laboratory methods of introducing the ds/siRNA into insects through feeding, micro feeding / dripping and injecting cannot be used in fields. Transgenic crop is perhaps the most effective application of RNAi for pest control, but it needs long-time basic researches in order to reduce the cost and evaluate the safety. Therefore, transgenic microbe is maybe a better choice. Entomopathogenic fungi generally invade the host insects through cuticle like chemical insecticides contact insect to control sucking sap pests. Isaria fumosorosea is a common fungal entomopathogen in whitefly, Bemisia tabaci. We constructed a recombinant strain of I. fumosorosea expressing specific dsRNA of whitefly's TLR7 gene. It could silence the TLR7 gene and improve the virulence against whitefly. Transgenic fungal entomopathogen has shown great potential to attain the application of RNAi technology for pests control in fields. In the future, the research interests should be focused on the selection of susceptible target pests and their vital genes, and optimizing the methods for screening genes and recombinants as well.

  7. Gene expression profiling: can we identify the right target genes?

    Directory of Open Access Journals (Sweden)

    J. E. Loyd

    2008-12-01

    Full Text Available Gene expression profiling allows the simultaneous monitoring of the transcriptional behaviour of thousands of genes, which may potentially be involved in disease development. Several studies have been performed in idiopathic pulmonary fibrosis (IPF, which aim to define genetic links to the disease in an attempt to improve the current understanding of the underlying pathogenesis of the disease and target pathways for intervention. Expression profiling has shown a clear difference in gene expression between IPF and normal lung tissue, and has identified a wide range of candidate genes, including those known to encode for proteins involved in extracellular matrix formation and degradation, growth factors and chemokines. Recently, familial pulmonary fibrosis cohorts have been examined in an attempt to detect specific genetic mutations associated with IPF. To date, these studies have identified families in which IPF is associated with mutations in the gene encoding surfactant protein C, or with mutations in genes encoding components of telomerase. Although rare and clearly not responsible for the disease in all individuals, the nature of these mutations highlight the importance of the alveolar epithelium in disease pathogenesis and demonstrate the potential for gene expression profiling in helping to advance the current understanding of idiopathic pulmonary fibrosis.

  8. Listeria monocytogenes as a vector for anti-cancer therapies.

    LENUS (Irish Health Repository)

    Tangney, Mark

    2012-01-31

    The intracellular pathogen Listeria monocytogenes represents a promising therapeutic vector for the delivery of DNA, RNA or protein to cancer cells or to prime immune responses against tumour-specific antigens. A number of biological properties make L. monocytogenes a promising platform for development as a vector for either gene therapy or as an anti-cancer vaccine vector. L. monocytogenes is particularly efficient in mediating internalization into host cells. Once inside cells, the bacterium produces specific virulence factors which lyse the vaculolar membrane and allow escape into the cytoplasm. Once in the cytosol, L. monocytogenes is capable of actin-based motility and cell-to-cell spread without an extracellular phase. The cytoplasmic location of L. monocytogenes is significant as this potentiates entry of antigens into the MHC Class I antigen processing pathway leading to priming of specific CD8(+) T cell responses. The cytoplasmic location is also beneficial for the delivery of DNA (bactofection) by L. monocytogenes whilst cell-to-cell spread may facilitate access of the vector to cells throughout the tumour. Several preclinical studies have demonstrated the ability of L. monocytogenes for intracellular gene or protein delivery in vitro and in vivo, and this vector has also displayed safety and efficacy in clinical trial. Here, we review the features of the L. monocytogenes host-pathogen interaction that make this bacterium such an attractive candidate with which to induce appropriate therapeutic responses. We focus primarily upon work that has led to attenuation of the pathogen, demonstrated DNA, RNA or protein delivery to tumour cells as well as research that shows the efficacy of L. monocytogenes as a vector for tumour-specific vaccine delivery.

  9. Anti-cancer natural products isolated from chinese medicinal herbs

    Directory of Open Access Journals (Sweden)

    Wu Guosheng

    2011-07-01

    Full Text Available Abstract In recent years, a number of natural products isolated from Chinese herbs have been found to inhibit proliferation, induce apoptosis, suppress angiogenesis, retard metastasis and enhance chemotherapy, exhibiting anti-cancer potential both in vitro and in vivo. This article summarizes recent advances in in vitro and in vivo research on the anti-cancer effects and related mechanisms of some promising natural products. These natural products are also reviewed for their therapeutic potentials, including flavonoids (gambogic acid, curcumin, wogonin and silibinin, alkaloids (berberine, terpenes (artemisinin, β-elemene, oridonin, triptolide, and ursolic acid, quinones (shikonin and emodin and saponins (ginsenoside Rg3, which are isolated from Chinese medicinal herbs. In particular, the discovery of the new use of artemisinin derivatives as excellent anti-cancer drugs is also reviewed.

  10. AAV-Based Targeting Gene Therapy

    Directory of Open Access Journals (Sweden)

    Wenfang Shi

    2008-01-01

    Full Text Available Since the first parvovirus serotype AAV2 was isolated from human and used as a vector for gene therapy application, there have been significant progresses in AAV vector development. AAV vectors have been extensively investigated in gene therapy for a broad application. AAV vectors have been considered as the first choice of vector due to efficient infectivity, stable expression and non-pathogenicity. However, the untoward events in AAV mediated in vivo gene therapy studies proposed the new challenges for their further applications. Deep understanding of the viral life cycle, viral structure and replication, infection mechanism and efficiency of AAV DNA integration, in terms of contributing viral, host-cell factors and circumstances would promote to evaluate the advantages and disadvantages and provide more insightful information for the possible clinical applications. In this review, main effort will be focused on the recent progresses in gene delivery to the target cells via receptor-ligand interaction and DNA specific integration regulation. Furthermore AAV receptor and virus particle intracellular trafficking are also discussed.

  11. Hydrofocusing Bioreactor Produces Anti-Cancer Alkaloids

    Science.gov (United States)

    Gonda, Steve R.; Valluri, Jagan V.

    2011-01-01

    microgravitation of an HFB do not need to maintain the same surface forces as in normal Earth gravitation, they can divert more energy sources to growth and differentiation and, perhaps, to biosynthesis of greater quantities of desired medicinal compounds. Because one can adjust the HFB to vary effective gravitation, one can also test the effects of intermediate levels of gravitation on biosynthesis of various products. The potential utility of this methodology for producing drugs was demonstrated in experiments in which sandalwood and Madagascar periwinkle cells were grown in an HFB. The conditions in the HFB were chosen to induce the cells to form into aggregate cultures that produced anti-cancer indole alkaloids in amounts greater than do comparable numbers of cells of the same species cultured according to previously known methodologies. The observations made in these experiments were interpreted as suggesting that the aggregation of the cells might be responsible for the enhancement of production of alkaloids.

  12. Polyamine analogues targeting epigenetic gene regulation.

    Science.gov (United States)

    Huang, Yi; Marton, Laurence J; Woster, Patrick M; Casero, Robert A

    2009-11-04

    Over the past three decades the metabolism and functions of the polyamines have been actively pursued as targets for antineoplastic therapy. Interactions between cationic polyamines and negatively charged nucleic acids play a pivotal role in DNA stabilization and RNA processing that may affect gene expression, translation and protein activity. Our growing understanding of the unique roles that the polyamines play in chromatin regulation, and the discovery of novel proteins homologous with specific regulatory enzymes in polyamine metabolism, have led to our interest in exploring chromatin remodelling enzymes as potential therapeutic targets for specific polyamine analogues. One of our initial efforts focused on utilizing the strong affinity that the polyamines have for chromatin to create a backbone structure, which could be combined with active-site-directed inhibitor moieties of HDACs (histone deacetylases). Specific PAHAs (polyaminohydroxamic acids) and PABAs (polyaminobenzamides) polyamine analogues have demonstrated potent inhibition of the HDACs, re-expression of p21 and significant inhibition of tumour growth. A second means of targeting the chromatin-remodelling enzymes with polyamine analogues was facilitated by the recent identification of flavin-dependent LSD1 (lysine-specific demethylase 1). The existence of this enzyme demonstrated that histone lysine methylation is a dynamic process similar to other histone post-translational modifications. LSD1 specifically catalyses demethylation of mono- and di-methyl Lys4 of histone 3, key positive chromatin marks associated with transcriptional activation. Structural and catalytic similarities between LSD1 and polyamine oxidases facilitated the identification of biguanide, bisguanidine and oligoamine polyamine analogues that are potent inhibitors of LSD1. Cellular inhibition of LSD1 by these unique compounds led to the re-activation of multiple epigenetically silenced genes important in tumorigenesis. The use of

  13. Synergistic anti-cancer effect of phenformin and oxamate.

    Directory of Open Access Journals (Sweden)

    W Keith Miskimins

    Full Text Available Phenformin (phenethylbiguanide; an anti-diabetic agent plus oxamate [lactate dehydrogenase (LDH inhibitor] was tested as a potential anti-cancer therapeutic combination. In in vitro studies, phenformin was more potent than metformin, another biguanide, recently recognized to have anti-cancer effects, in promoting cancer cell death in the range of 25 times to 15 million times in various cancer cell lines. The anti-cancer effect of phenformin was related to complex I inhibition in the mitochondria and subsequent overproduction of reactive oxygen species (ROS. Addition of oxamate inhibited LDH activity and lactate production by cells, which is a major side effect of biguanides, and induced more rapid cancer cell death by decreasing ATP production and accelerating ROS production. Phenformin plus oxamate was more effective than phenformin combined with LDH knockdown. In a syngeneic mouse model, phenformin with oxamate increased tumor apoptosis, reduced tumor size and (18F-fluorodeoxyglucose (FDG uptake on positron emission tomography/computed tomography compared to control. We conclude that phenformin is more cytotoxic towards cancer cells than metformin. Furthermore, phenformin and oxamate have synergistic anti-cancer effects through simultaneous inhibition of complex I in the mitochondria and LDH in the cytosol, respectively.

  14. Development of a Novel Anti-HIF-1α Screening System Coupled with Biochemical and Biological Validation for Rapidly Selecting Potent Anti-Cancer Compounds.

    Science.gov (United States)

    Lu, Yi; Madu, Chikezie; Masters, Jordan; Lu, Andrew; Li, Liyuan

    2014-01-01

    Breast cancer (BCa) is the most diagnosed cancer and the second leading cause of cancer death in the American women. Adaptation to the hypoxic environment seen in solid tumors is critical for tumor cell survival and growth. The activation of hypoxia inducible factor-1 alpha (HIF-1α), an important master transcriptional factor that is induced and stabilized by intratumoral hypoxia, stimulates a group of HIF-1α-regulated genes including vascular endothelial growth factor (VEGF), leading tumor cells towards malignant progression. Therefore, a promising therapeutic approach to cancer treatment is to target HIF-1α. The goal of this project was to develop and validate a screening system coupled with secondary screen/validation process that has the capability to screen large numbers of potential anti-cancer small-molecule compounds based on their anti-HIF-1α activities. Breast cancer MDA-231 cells were used as the model to select potent anti-HIF-1α compounds by their abilities to inhibit transactivation of a VEGF promoter fused to a luciferase reporter gene under hypoxia. Positive compounds were then validated by a series of assays that confirm compounds' anti-HIF-1α activities including measurement of HIF-1α downstream VEGF gene expression and angiogenic ability of BCa cells. Results of our pilot screening demonstrate that this prototype screening coupled with validation system can effectively select highly potent anti-HIF-1α agents from the compound library, suggesting that this prototype screen system has the potential to be developed into a high-throughput screen (HTS) coupled with automated validation process for the screening and identification of novel and effective anti-cancer drugs based on anti-HIF-1α mechanism.

  15. In silico identification of anti-cancer compounds and plants from traditional Chinese medicine database

    Science.gov (United States)

    Dai, Shao-Xing; Li, Wen-Xing; Han, Fei-Fei; Guo, Yi-Cheng; Zheng, Jun-Juan; Liu, Jia-Qian; Wang, Qian; Gao, Yue-Dong; Li, Gong-Hua; Huang, Jing-Fei

    2016-05-01

    There is a constant demand to develop new, effective, and affordable anti-cancer drugs. The traditional Chinese medicine (TCM) is a valuable and alternative resource for identifying novel anti-cancer agents. In this study, we aim to identify the anti-cancer compounds and plants from the TCM database by using cheminformatics. We first predicted 5278 anti-cancer compounds from TCM database. The top 346 compounds were highly potent active in the 60 cell lines test. Similarity analysis revealed that 75% of the 5278 compounds are highly similar to the approved anti-cancer drugs. Based on the predicted anti-cancer compounds, we identified 57 anti-cancer plants by activity enrichment. The identified plants are widely distributed in 46 genera and 28 families, which broadens the scope of the anti-cancer drug screening. Finally, we constructed a network of predicted anti-cancer plants and approved drugs based on the above results. The network highlighted the supportive role of the predicted plant in the development of anti-cancer drug and suggested different molecular anti-cancer mechanisms of the plants. Our study suggests that the predicted compounds and plants from TCM database offer an attractive starting point and a broader scope to mine for potential anti-cancer agents.

  16. Gene expression-targeted isoflavone therapy.

    Science.gov (United States)

    Węgrzyn, Alicja

    2012-04-01

    Lysosomal storage diseases (LSD) form a group of inherited metabolic disorders caused by dysfunction of one of the lysosomal proteins, resulting in the accumulation of certain compounds. Although these disorders are among first genetic diseases for which specific treatments were proposed, there are still serious unsolved problems that require development of novel therapeutic procedures. An example is neuronopathy, which develops in most of LSD and cannot be treated efficiently by currently approved therapies. Recently, a new potential therapy, called gene expression-targeted isoflavone therapy (GET IT), has been proposed for a group of LSD named mucopolysaccharidoses (MPS), in which storage of incompletely degraded glycosaminoglycans (GAGs) results in severe symptoms of virtually all tissues and organs, including central nervous system. The idea of this therapy is to inhibit synthesis of GAGs by modulating expression of genes coding for enzymes involved in synthesis of these compounds. Such a modulation is possible by using isoflavones, particularly genistein, which interfere with a signal transduction process necessary for stimulation of expression of certain genes. Results of in vitro experiments and studies on animal models indicated a high efficiency of GET IT, including correction of behavior of affected mice. However, clinical trials, performed with soy isoflavone extracts, revealed only limited efficacy. This caused a controversy about GET IT as a potential, effective treatment of patients suffering from MPS, especially neuronopathic forms of these diseases. It this critical review, I present possible molecular mechanisms of therapeutic action of isoflavones (particularly genistein) and suggest that efficacy of GET IT might be sufficiently high when using relatively high doses of synthetic genistein (which was employed in experiments on cell cultures and mouse models) rather than low doses of soy isoflavone extracts (which were used in clinical trials). This

  17. Imidazoles and benzimidazoles as tubulin-modulators for anti-cancer therapy.

    Science.gov (United States)

    Torres, Fernando C; García-Rubiño, M Eugenia; Lozano-López, César; Kawano, Daniel F; Eifler-Lima, Vera L; von Poser, Gilsane L; Campos, Joaquín M

    2015-01-01

    Imidazoles and benzimidazoles are privileged heterocyclic bioactive compounds used with success in the clinical practice of innumerous diseases. Although there are many advancements in cancer therapy, microtubules remain as one of the few macromolecular targets validated for planning active anti-cancer compounds, and the design of drugs that modulate microtubule dynamics in unknown sites of tubulin is one of the goals of the medicinal chemistry. The discussion of the role of new and commercially available imidazole and benzimidazole derivatives as tubulin modulators is scattered throughout scientific literature, and indicates that these compounds have a tubulin modulation mechanism different from that of tubulin modulators clinically available, such as paclitaxel, docetaxel, vincristine and vinblastine. In fact, recent literature indicates that these derivatives inhibit microtubule formation binding to the colchicine site, present good pharmacokinetic properties and are capable of overcoming multidrug resistance in many cell lines. The understanding of the mechanisms involved in the imidazoles/benzimidazoles modulation of microtubule dynamics is very important to develop new strategies to overcome the resistance to anti-cancer drugs and to discover new biomarkers and targets for cancer chemotherapy.

  18. Teratogens as anti-cancer drugs.

    Science.gov (United States)

    Blagosklonny, Mikhail V

    2005-11-01

    Most anticancer drugs are teratogens, merely because they target vital cellular functions. Conversely, some plants produce agents that intentionally target embryonic signaling pathways, precisely to cause birth defects if pregnant animals eat such plants. Cyclopamine, a teratogen produced by a flowering plant, inhibits the Hh/Gli pathway, causing developmental defects such as cyclopia (one eye in the middle of the face). In theory, selective teratogens may suppress cancer cells that reactivate embryonic pathways, while sparing most normal cells. I discuss the potential (and limits) of teratogens in cancer therapy, linking diverse topics from morning sickness of pregnancy, embryonic pathways and poisonous plants to the mechanism of action of anticancer teratogens and their combinations with less selective cytotoxic agents.

  19. The mechanism of gene targeting in human somatic cells.

    Directory of Open Access Journals (Sweden)

    Yinan Kan

    2014-04-01

    Full Text Available Gene targeting in human somatic cells is of importance because it can be used to either delineate the loss-of-function phenotype of a gene or correct a mutated gene back to wild-type. Both of these outcomes require a form of DNA double-strand break (DSB repair known as homologous recombination (HR. The mechanism of HR leading to gene targeting, however, is not well understood in human cells. Here, we demonstrate that a two-end, ends-out HR intermediate is valid for human gene targeting. Furthermore, the resolution step of this intermediate occurs via the classic DSB repair model of HR while synthesis-dependent strand annealing and Holliday Junction dissolution are, at best, minor pathways. Moreover, and in contrast to other systems, the positions of Holliday Junction resolution are evenly distributed along the homology arms of the targeting vector. Most unexpectedly, we demonstrate that when a meganuclease is used to introduce a chromosomal DSB to augment gene targeting, the mechanism of gene targeting is inverted to an ends-in process. Finally, we demonstrate that the anti-recombination activity of mismatch repair is a significant impediment to gene targeting. These observations significantly advance our understanding of HR and gene targeting in human cells.

  20. The mechanism of gene targeting in human somatic cells.

    Science.gov (United States)

    Kan, Yinan; Ruis, Brian; Lin, Sherry; Hendrickson, Eric A

    2014-04-01

    Gene targeting in human somatic cells is of importance because it can be used to either delineate the loss-of-function phenotype of a gene or correct a mutated gene back to wild-type. Both of these outcomes require a form of DNA double-strand break (DSB) repair known as homologous recombination (HR). The mechanism of HR leading to gene targeting, however, is not well understood in human cells. Here, we demonstrate that a two-end, ends-out HR intermediate is valid for human gene targeting. Furthermore, the resolution step of this intermediate occurs via the classic DSB repair model of HR while synthesis-dependent strand annealing and Holliday Junction dissolution are, at best, minor pathways. Moreover, and in contrast to other systems, the positions of Holliday Junction resolution are evenly distributed along the homology arms of the targeting vector. Most unexpectedly, we demonstrate that when a meganuclease is used to introduce a chromosomal DSB to augment gene targeting, the mechanism of gene targeting is inverted to an ends-in process. Finally, we demonstrate that the anti-recombination activity of mismatch repair is a significant impediment to gene targeting. These observations significantly advance our understanding of HR and gene targeting in human cells.

  1. Microfluidics: Emerging prospects for anti-cancer drug screening.

    Science.gov (United States)

    Wlodkowic, Donald; Darzynkiewicz, Zbigniew

    2010-11-10

    Cancer constitutes a heterogenic cellular system with a high level of spatio-temporal complexity. Recent discoveries by systems biologists have provided emerging evidence that cellular responses to anti-cancer modalities are stochastic in nature. To uncover the intricacies of cell-to-cell variability and its relevance to cancer therapy, new analytical screening technologies are needed. The last decade has brought forth spectacular innovations in the field of cytometry and single cell cytomics, opening new avenues for systems oncology and high-throughput real-time drug screening routines. The up-and-coming microfluidic Lab-on-a-Chip (LOC) technology and micro-total analysis systems (μTAS) are arguably the most promising platforms to address the inherent complexity of cellular systems with massive experimental parallelization and 4D analysis on a single cell level. The vast miniaturization of LOC systems and multiplexing enables innovative strategies to reduce drug screening expenditures while increasing throughput and content of information from a given sample. Small cell numbers and operational reagent volumes are sufficient for microfluidic analyzers and, as such, they enable next generation high-throughput and high-content screening of anti-cancer drugs on patient-derived specimens. Herein we highlight the selected advancements in this emerging field of bioengineering, and provide a snapshot of developments with relevance to anti-cancer drug screening routines.

  2. Association Between hTERT rs2736100 Polymorphism and Sensitivity to Anti-cancer Agents

    Directory of Open Access Journals (Sweden)

    Julie eKim

    2013-08-01

    Full Text Available Background: The rs2736100 single nucleotide polymorphism (SNP is located in the intron 2 of human telomerase reverse transcriptase (hTERT gene. Recent genome-wide association studies (GWAS have consistently supported the strong association between this SNP and risk for multiple cancers. Given the important role of the hTERT gene and this SNP in cancer biology, we hypothesize that rs2736100 may also confer susceptibility to anti-cancer drug sensitivity. In this study we aim to investigate the correlation between the rs2736100 genotype and the responsiveness to anti-cancer agents in the NCI-60 cancer cell panel. Methods and Materials: The hTERT rs2736100 was genotyped in the NCI-60 cancer cell lines. The relative telomere length of each cell line was quantified using real-time PCR. The genotype was then correlated with publically available drug sensitivity data of two agents with telomerase-inhibition activity: Geldanamycin (HSP90 inhibitor and RHPS4/BRACO19 (G-quadruplex stabilizer as well as additional 110 commonly used agents with established mechanism of action. The association between rs2736100 and mutation status of TP53 gene was also tested.Results: The C allele of the SNP was significantly correlated with increased sensitivity to RHPS4/BRACO19 with an additive effect (r=-0.35, p=0.009 but not with Geldanamycin. The same allele was also significantly associated with sensitivity to antimitotic agents compared to other agents (p=0.003. The highest correlation was observed between the SNP and paclitaxel (r=-0.36, p=0.005. The telomere length was neither associated with rs2736100 nor with sensitivity to anti-cancer agents. The C allele of rs2736100 was significantly associated with increased mutation rate in TP53 gene (p=0.004.Conclusion: Our data suggested that the cancer risk allele of hTERT rs2736100 polymorphism may also affect the cancer cell response to both TERT inhibitor and anti-mitotic agents, which might be attributed to the elevated

  3. Gene Targeting Without DSB Induction Is Inefficient in Barley.

    Science.gov (United States)

    Horvath, Mihaly; Steinbiss, Hans-Henning; Reiss, Bernd

    2016-01-01

    Double strand-break (DSB) induction allowed efficient gene targeting in barley (Hordeum vulgare), but little is known about efficiencies in its absence. To obtain such data, an assay system based on the acetolactate synthase (ALS) gene was established, a target gene which had been used previously in rice and Arabidopsis thaliana. Expression of recombinases RAD51 and RAD54 had been shown to improve gene targeting in A. thaliana and positive-negative (P-N) selection allows the routine production of targeted mutants without DSB induction in rice. We implemented these approaches in barley and analysed gene targeting with the ALS gene in wild type and RAD51 and RAD54 transgenic lines. In addition, P-N selection was tested. In contrast to the high gene targeting efficiencies obtained in the absence of DSB induction in A. thaliana or rice, not one single gene targeting event was obtained in barley. These data suggest that gene targeting efficiencies are very low in barley and can substantially differ between different plants, even at the same target locus. They also suggest that the amount of labour and time would become unreasonably high to use these methods as a tool in routine applications. This is particularly true since DSB induction offers efficient alternatives. Barley, unlike rice and A. thaliana has a large, complex genome, suggesting that genome size or complexity could be the reason for the low efficiencies. We discuss to what extent transformation methods, genome size or genome complexity could contribute to the striking differences in the gene targeting efficiencies between barley, rice and A. thaliana.

  4. Targeted gene knockout in chickens mediated by TALENs

    OpenAIRE

    Park, Tae Sub; Lee, Hong Jo; Kim, Ki Hyun; Kim, Jin-Soo; Han, Jae Yong

    2014-01-01

    Targeted gene knockout by editing specific loci in genome has revolutionized the field of functional genomics. Transcription activator-like effector nucleases (TALENs) are representative next-generation platforms for customized genomic editing in transgenic animals, as well as cultured cells in vitro. In this study, in combination with chicken primordial germ cell line with germ-line transmission capacity, we generated the ovalbumin gene knockout chickens by TALEN-mediated gene targeting. Our...

  5. Molecular pathways: targeting ETS gene fusions in cancer.

    Science.gov (United States)

    Feng, Felix Y; Brenner, J Chad; Hussain, Maha; Chinnaiyan, Arul M

    2014-09-01

    Rearrangements, or gene fusions, involving the ETS family of transcription factors are common driving events in both prostate cancer and Ewing sarcoma. These rearrangements result in pathogenic expression of the ETS genes and trigger activation of transcriptional programs enriched for invasion and other oncogenic features. Although ETS gene fusions represent intriguing therapeutic targets, transcription factors, such as those comprising the ETS family, have been notoriously difficult to target. Recently, preclinical studies have demonstrated an association between ETS gene fusions and components of the DNA damage response pathway, such as PARP1, the catalytic subunit of DNA protein kinase (DNAPK), and histone deactylase 1 (HDAC1), and have suggested that ETS fusions may confer sensitivity to inhibitors of these DNA repair proteins. In this review, we discuss the role of ETS fusions in cancer, the preclinical rationale for targeting ETS fusions with inhibitors of PARP1, DNAPK, and HDAC1, as well as ongoing clinical trials targeting ETS gene fusions.

  6. Characterisation of genome-wide PLZF/RARA target genes.

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    Salvatore Spicuglia

    Full Text Available The PLZF/RARA fusion protein generated by the t(11;17(q23;q21 translocation in acute promyelocytic leukaemia (APL is believed to act as an oncogenic transcriptional regulator recruiting epigenetic factors to genes important for its transforming potential. However, molecular mechanisms associated with PLZF/RARA-dependent leukaemogenesis still remain unclear.We searched for specific PLZF/RARA target genes by ChIP-on-chip in the haematopoietic cell line U937 conditionally expressing PLZF/RARA. By comparing bound regions found in U937 cells expressing endogenous PLZF with PLZF/RARA-induced U937 cells, we isolated specific PLZF/RARA target gene promoters. We next analysed gene expression profiles of our identified target genes in PLZF/RARA APL patients and analysed DNA sequences and epigenetic modification at PLZF/RARA binding sites. We identify 413 specific PLZF/RARA target genes including a number encoding transcription factors involved in the regulation of haematopoiesis. Among these genes, 22 were significantly down regulated in primary PLZF/RARA APL cells. In addition, repressed PLZF/RARA target genes were associated with increased levels of H3K27me3 and decreased levels of H3K9K14ac. Finally, sequence analysis of PLZF/RARA bound sequences reveals the presence of both consensus and degenerated RAREs as well as enrichment for tissue-specific transcription factor motifs, highlighting the complexity of targeting fusion protein to chromatin. Our study suggests that PLZF/RARA directly targets genes important for haematopoietic development and supports the notion that PLZF/RARA acts mainly as an epigenetic regulator of its direct target genes.

  7. Gene therapy of cancer and development of therapeutic target gene

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Chang Min; Kwon, Hee Chung

    1998-04-01

    We applied HSV-tk/GCV strategy to orthotopic rat hepatoma model and showed anticancer effects of hepatoma. The increased expression of Lac Z gene after adenovirus-mediated gene delivery throughout hepatic artery was thought that is increased the possibility of gene therapy for curing hepatoma. With the construction of kGLP-laboratory, it is possible to produce a good quantity and quality of adenovirus in lage-scale production and purification of adenovirus vector. Also, the analysis of hepatoma related genes by PCR-LOH could be used for the diagnosis of patients and the development of therapeutic gene.

  8. Profiling the anti-protozoal activity of anti-cancer HDAC inhibitors against Plasmodium and Trypanosoma parasites

    Directory of Open Access Journals (Sweden)

    Jessica A. Engel

    2015-12-01

    Full Text Available Histone deacetylase (HDAC enzymes work together with histone acetyltransferases (HATs to reversibly acetylate both histone and non-histone proteins. As a result, these enzymes are involved in regulating chromatin structure and gene expression as well as other important cellular processes. HDACs are validated drug targets for some types of cancer, with four HDAC inhibitors clinically approved. However, they are also showing promise as novel drug targets for other indications, including malaria and other parasitic diseases. In this study the in vitro activity of four anti-cancer HDAC inhibitors was examined against parasites that cause malaria and trypanosomiasis. Three of these inhibitors, suberoylanilide hydroxamic acid (SAHA; vorinostat®, romidepsin (Istodax® and belinostat (Beleodaq®, are clinically approved for the treatment of T-cell lymphoma, while the fourth, panobinostat, has recently been approved for combination therapy use in certain patients with multiple myeloma. All HDAC inhibitors were found to inhibit the growth of asexual-stage Plasmodium falciparum malaria parasites in the nanomolar range (IC50 10–200 nM, while only romidepsin was active at sub-μM concentrations against bloodstream form Trypanosoma brucei brucei parasites (IC50 35 nM. The compounds were found to have some selectivity for malaria parasites compared with mammalian cells, but were not selective for trypanosome parasites versus mammalian cells. All compounds caused hyperacetylation of histone and non-histone proteins in P. falciparum asexual stage parasites and inhibited deacetylase activity in P. falciparum nuclear extracts in addition to recombinant PfHDAC1 activity. P. falciparum histone hyperacetylation data indicate that HDAC inhibitors may differentially affect the acetylation profiles of histone H3 and H4.

  9. Profiling the anti-protozoal activity of anti-cancer HDAC inhibitors against Plasmodium and Trypanosoma parasites.

    Science.gov (United States)

    Engel, Jessica A; Jones, Amy J; Avery, Vicky M; Sumanadasa, Subathdrage D M; Ng, Susanna S; Fairlie, David P; Adams, Tina S; Andrews, Katherine T

    2015-12-01

    Histone deacetylase (HDAC) enzymes work together with histone acetyltransferases (HATs) to reversibly acetylate both histone and non-histone proteins. As a result, these enzymes are involved in regulating chromatin structure and gene expression as well as other important cellular processes. HDACs are validated drug targets for some types of cancer, with four HDAC inhibitors clinically approved. However, they are also showing promise as novel drug targets for other indications, including malaria and other parasitic diseases. In this study the in vitro activity of four anti-cancer HDAC inhibitors was examined against parasites that cause malaria and trypanosomiasis. Three of these inhibitors, suberoylanilide hydroxamic acid (SAHA; vorinostat(®)), romidepsin (Istodax(®)) and belinostat (Beleodaq(®)), are clinically approved for the treatment of T-cell lymphoma, while the fourth, panobinostat, has recently been approved for combination therapy use in certain patients with multiple myeloma. All HDAC inhibitors were found to inhibit the growth of asexual-stage Plasmodium falciparum malaria parasites in the nanomolar range (IC50 10-200 nM), while only romidepsin was active at sub-μM concentrations against bloodstream form Trypanosoma brucei brucei parasites (IC50 35 nM). The compounds were found to have some selectivity for malaria parasites compared with mammalian cells, but were not selective for trypanosome parasites versus mammalian cells. All compounds caused hyperacetylation of histone and non-histone proteins in P. falciparum asexual stage parasites and inhibited deacetylase activity in P. falciparum nuclear extracts in addition to recombinant PfHDAC1 activity. P. falciparum histone hyperacetylation data indicate that HDAC inhibitors may differentially affect the acetylation profiles of histone H3 and H4.

  10. Significance of Cancer Stem Cells in Anti-Cancer Therapies

    Science.gov (United States)

    Botelho, Mónica; Alves, Helena

    2017-01-01

    Stem cells are the focus of cutting edge research interest because of their competence both to self-renew and proliferate, and to differentiate into a variety of tissues, offering enticing prospects of growing replacement organs in vitro, among other possible therapeutic implications. It is conceivable that cancer stem cells share a number of biological hallmarks that are different from their normal-tissue counterparts and that these might be taken advantage of for therapeutic benefits. In this review we discuss the significance of cancer stem cells in diagnosis and prognosis of cancer as well as in the development of new strategies for anti-cancer drug design.

  11. Triterpenoids of Marine Origin as Anti-Cancer Agents

    Directory of Open Access Journals (Sweden)

    Yong-Xin Li

    2013-07-01

    Full Text Available Triterpenoids are the most abundant secondary metabolites present in marine organisms, such as marine sponges, sea cucumbers, marine algae and marine-derived fungi. A large number of triterpenoids are known to exhibit cytotoxicity against a variety of tumor cells, as well as anticancer efficacy in preclinical animal models. In this review efforts have been taken to review the structural features and the potential use of triterpenoids of marine origin to be used in the pharmaceutical industry as potential anti-cancer drug leads.

  12. An in vivo C. elegans model system for screening EGFR-inhibiting anti-cancer drugs.

    Directory of Open Access Journals (Sweden)

    Young-Ki Bae

    Full Text Available The epidermal growth factor receptor (EGFR is a well-established target for cancer treatment. EGFR tyrosine kinase (TK inhibitors, such as gefinitib and erlotinib, have been developed as anti-cancer drugs. Although non-small cell lung carcinoma with an activating EGFR mutation, L858R, responds well to gefinitib and erlotinib, tumors with a doubly mutated EGFR, T790M-L858R, acquire resistance to these drugs. The C. elegans EGFR homolog LET-23 and its downstream signaling pathway have been studied extensively to provide insight into regulatory mechanisms conserved from C. elegans to humans. To develop an in vivo screening system for potential cancer drugs targeting specific EGFR mutants, we expressed three LET-23 chimeras in which the TK domain was replaced with either the human wild-type TK domain (LET-23::hEGFR-TK, a TK domain with the L858R mutation (LET-23::hEGFR-TK[L858R], or a TK domain with the T790M-L858R mutations (LET-23::hEGFR-TK[T790M-L858R] in C. elegans vulval cells using the let-23 promoter. The wild-type hEGFR-TK chimeric protein rescued the let-23 mutant phenotype, and the activating mutant hEGFR-TK chimeras induced a multivulva (Muv phenotype in a wild-type C. elegans background. The anti-cancer drugs gefitinib and erlotinib suppressed the Muv phenotype in LET-23::hEGFR-TK[L858R]-expressing transgenic animals, but not in LET-23::hEGFR-TK[T790M-L858R] transgenic animals. As a pilot screen, 8,960 small chemicals were tested for Muv suppression, and AG1478 (an EGFR-TK inhibitor and U0126 (a MEK inhibitor were identified as potential inhibitors of EGFR-mediated biological function. In conclusion, transgenic C. elegans expressing chimeric LET-23::hEGFR-TK proteins are a model system that can be used in mutation-specific screens for new anti-cancer drugs.

  13. Synthetic Small Molecule Inhibitors of Hh Signaling As Anti-Cancer Chemotherapeutics

    Science.gov (United States)

    Maschinot, C.A.; Pace, J.R.; Hadden, M.K.

    2016-01-01

    The hedgehog (Hh) pathway is a developmental signaling pathway that is essential to the proper embryonic development of many vertebrate systems. Dysregulation of Hh signaling has been implicated as a causative factor in the development and progression of several forms of human cancer. As such, the development of small molecule inhibitors of Hh signaling as potential anti-cancer chemotherapeutics has been a major area of research interest in both academics and industry over the past ten years. Through these efforts, synthetic small molecules that target multiple components of the Hh pathway have been identified and advanced to preclinical or clinical development. The goal of this review is to provide an update on the current status of several synthetic small molecule Hh pathway inhibitors and explore the potential of several recently disclosed inhibitory scaffolds. PMID:26310919

  14. Trypanocidal activity of the proteasome inhibitor and anti-cancer drug bortezomib

    Directory of Open Access Journals (Sweden)

    Wang Xia

    2009-07-01

    Full Text Available Abstract The proteasome inhibitor and anti-cancer drug bortezomib was tested for in vitro activity against bloodstream forms of Trypanosoma brucei. The concentrations of bortezomib required to reduce the growth rate by 50% and to kill all trypanosomes were 3.3 nM and 10 nM, respectively. In addition, bortezomib was 10 times more toxic to trypanosomes than to human HL-60 cells. Moreover, exposure of trypanosomes to 10 nM bortezomib for 16 h was enough to kill 90% of the parasites following incubation in fresh medium. However, proteasomal peptidase activities of trypanosomes exposed to bortezomib were only inhibited by 10% and 30% indicating that the proteasome is not the main target of the drug. The results suggest that bortezomib may be useful as drug for the treatment of human African trypanosomiasis.

  15. Turning tumor-promoting copper into an anti-cancer weapon via high-throughput chemistry.

    Science.gov (United States)

    Wang, F; Jiao, P; Qi, M; Frezza, M; Dou, Q P; Yan, B

    2010-01-01

    Copper is an essential element for multiple biological processes. Its concentration is elevated to a very high level in cancer tissues for promoting cancer development through processes such as angiogenesis. Organic chelators of copper can passively reduce cellular copper and serve the role as inhibitors of angiogenesis. However, they can also actively attack cellular targets such as proteasome, which plays a critical role in cancer development and survival. The discovery of such molecules initially relied on a step by step synthesis followed by biological assays. Today high-throughput chemistry and high-throughput screening have significantly expedited the copper-binding molecules discovery to turn "cancer-promoting" copper into anti-cancer agents.

  16. Peroxisome proliferator-activated receptor alpha target genes.

    Science.gov (United States)

    Rakhshandehroo, Maryam; Knoch, Bianca; Müller, Michael; Kersten, Sander

    2010-01-01

    The peroxisome proliferator-activated receptor alpha (PPARα) is a ligand-activated transcription factor involved in the regulation of a variety of processes, ranging from inflammation and immunity to nutrient metabolism and energy homeostasis. PPARα serves as a molecular target for hypolipidemic fibrates drugs which bind the receptor with high affinity. Furthermore, PPARα binds and is activated by numerous fatty acids and fatty acid-derived compounds. PPARα governs biological processes by altering the expression of a large number of target genes. Accordingly, the specific role of PPARα is directly related to the biological function of its target genes. Here, we present an overview of the involvement of PPARα in lipid metabolism and other pathways through a detailed analysis of the different known or putative PPARα target genes. The emphasis is on gene regulation by PPARα in liver although many of the results likely apply to other organs and tissues as well.

  17. Peroxisome Proliferator-Activated Receptor Alpha Target Genes

    Directory of Open Access Journals (Sweden)

    Maryam Rakhshandehroo

    2010-01-01

    Full Text Available The peroxisome proliferator-activated receptor alpha (PPARα is a ligand-activated transcription factor involved in the regulation of a variety of processes, ranging from inflammation and immunity to nutrient metabolism and energy homeostasis. PPARα serves as a molecular target for hypolipidemic fibrates drugs which bind the receptor with high affinity. Furthermore, PPARα binds and is activated by numerous fatty acids and fatty acid-derived compounds. PPARα governs biological processes by altering the expression of a large number of target genes. Accordingly, the specific role of PPARα is directly related to the biological function of its target genes. Here, we present an overview of the involvement of PPARα in lipid metabolism and other pathways through a detailed analysis of the different known or putative PPARα target genes. The emphasis is on gene regulation by PPARα in liver although many of the results likely apply to other organs and tissues as well.

  18. Targeted gene mutation in Phytophthora spp.

    NARCIS (Netherlands)

    Lamour, K.H.; Finley, L.; Hurtado-Gonzales, O.; Gobena, D.; Tierney, M.; Meijer, H.J.G.

    2006-01-01

    The genus Phytophthora belongs to the oomycetes and is composed of plant pathogens. Currently, there are no strategies to mutate specific genes for members of this genus. Whole genome sequences are available or being prepared for Phytophthora sojae, P. ramorum, P. infestans, and P. capsici and the d

  19. Paraptosis in the anti-cancer arsenal of natural products.

    Science.gov (United States)

    Lee, Dongjoo; Kim, In Young; Saha, Sharmistha; Choi, Kyeong Sook

    2016-06-01

    Given the problems with malignant cancer cells showing innate and acquired resistance to apoptosis, we need alternative means to induce cell death in cancer. Paraptosis is a type of programmed cell death that is characterized by dilation of the endoplasmic reticulum (ER) and/or mitochondria. Although relatively little is known regarding the molecular basis of paraptosis, the underlying mechanism clearly differs from that of apoptosis. Recent studies have shown that various natural products, including curcumin, celastrol, 15d-PGJ2, ophiobolin A, and paclitaxel, demonstrate anti-cancer effects by inducing the paraptosis-associated cell death, which was commonly characterized by vacuolation derived from the ER. Perturbation of cellular proteostasis due to proteasomal inhibition and disruption of sulfhydryl homeostasis, generation of reactive oxygen species, and/or imbalanced homeostasis of ions (e.g., Ca(2+) and K(+)) appear to contribute to the accumulation of misfolded protein and proteotoxicity in this process. Given the pathophysiological importance of paraptosis and the debate regarding the importance of apoptosis in solid tumor, we need to collect the available knowledge regarding paraptosis and suggest future directions in the field. Here, we review the morphological and biochemical features of paraptosis, the natural products that induce paraptosis-associated cell death, their proposed mechanisms, and the significance of paraptosis as a potential anti-cancer strategy. Such work and future clarifications should enable the development of new strategies for preventing cancer and/or combating malignant cancer.

  20. Genome-wide identification of KANADI1 target genes.

    Directory of Open Access Journals (Sweden)

    Paz Merelo

    Full Text Available Plant organ development and polarity establishment is mediated by the action of several transcription factors. Among these, the KANADI (KAN subclade of the GARP protein family plays important roles in polarity-associated processes during embryo, shoot and root patterning. In this study, we have identified a set of potential direct target genes of KAN1 through a combination of chromatin immunoprecipitation/DNA sequencing (ChIP-Seq and genome-wide transcriptional profiling using tiling arrays. Target genes are over-represented for genes involved in the regulation of organ development as well as in the response to auxin. KAN1 affects directly the expression of several genes previously shown to be important in the establishment of polarity during lateral organ and vascular tissue development. We also show that KAN1 controls through its target genes auxin effects on organ development at different levels: transport and its regulation, and signaling. In addition, KAN1 regulates genes involved in the response to abscisic acid, jasmonic acid, brassinosteroids, ethylene, cytokinins and gibberellins. The role of KAN1 in organ polarity is antagonized by HD-ZIPIII transcription factors, including REVOLUTA (REV. A comparison of their target genes reveals that the REV/KAN1 module acts in organ patterning through opposite regulation of shared targets. Evidence of mutual repression between closely related family members is also shown.

  1. Targeting Strategies in Cancer Gene Therapy%肿瘤基因治疗的靶向策略

    Institute of Scientific and Technical Information of China (English)

    王晋慧; 刘新垣

    2003-01-01

    对肿瘤组织的靶向性可以提高基因治疗的效果,避免对正常组织的损伤,并且能降低作为载体的微生物对机体的危害.对于瘤内注射的给药方法,靶向性似乎显得不是特别重要,但是如果要系统给药,靶向性是很关键的一个问题.靶向基因治疗肿瘤可以通过靶向基因导入和靶向基因表达来实现.近年来,在靶向基因导入方面的研究有很多进展,例如,用双亲性的桥连分子协助腺病毒和逆转录病毒靶向转导;在各种病毒载体的衣壳蛋白中插入靶向性的小肽或较大的多肽靶向结构域;增殖病毒作为一种很有前途的抗肿瘤制剂可有效地靶向杀伤肿瘤细胞.受体介导的DNA或DNA-脂质体复合物的靶向系统和其他一些靶向性的有疗效的载体,如细菌,也处于研究中.其中的一些载体已经进入临床实验.为了实现基因的靶向可调控表达,组织或肿瘤特异性的启动子和人工合成的可调控表达系统被用来调控治疗基因的表达.反义核酸、核酶以及脱氧核酶(DNAzyme)被用来靶向抑制与肿瘤发生密切相关基因的表达.%Targeting to the tumor tissues can improve the therapeutic effect of gene transfer by preventing damage of healthy tissues and decreasing the risk of germ line transduction. Although targeting seems not important for intratumoral gene delivery, it becomes crucial when systemic gene transfer is performed. Targeted gene therapy of malignancies can be achieved through targeted gene delivery or targeted gene transcription. Recent advances in targeted delivery include the successful use of bifunctional crosslinkers to target adenoviral and retroviral vectors, inserting short targeting peptides and larger polypeptide-binding domains into the coat proteins of a number of different viral vectors, and replication-competent vectors which have been shown to be promise as anti-cancer agents. Some other non-viral therapeutic agents, including

  2. IMG-ABC: new features for bacterial secondary metabolism analysis and targeted biosynthetic gene cluster discovery in thousands of microbial genomes

    Science.gov (United States)

    Hadjithomas, Michalis; Chen, I-Min A.; Chu, Ken; Huang, Jinghua; Ratner, Anna; Palaniappan, Krishna; Andersen, Evan; Markowitz, Victor; Kyrpides, Nikos C.; Ivanova, Natalia N.

    2017-01-01

    Secondary metabolites produced by microbes have diverse biological functions, which makes them a great potential source of biotechnologically relevant compounds with antimicrobial, anti-cancer and other activities. The proteins needed to synthesize these natural products are often encoded by clusters of co-located genes called biosynthetic gene clusters (BCs). In order to advance the exploration of microbial secondary metabolism, we developed the largest publically available database of experimentally verified and predicted BCs, the Integrated Microbial Genomes Atlas of Biosynthetic gene Clusters (IMG-ABC) (https://img.jgi.doe.gov/abc/). Here, we describe an update of IMG-ABC, which includes ClusterScout, a tool for targeted identification of custom biosynthetic gene clusters across 40 000 isolate microbial genomes, and a new search capability to query more than 700 000 BCs from isolate genomes for clusters with similar Pfam composition. Additional features enable fast exploration and analysis of BCs through two new interactive visualization features, a BC function heatmap and a BC similarity network graph. These new tools and features add to the value of IMG-ABC's vast body of BC data, facilitating their in-depth analysis and accelerating secondary metabolite discovery. PMID:27903896

  3. TargetMine, an integrated data warehouse for candidate gene prioritisation and target discovery.

    Directory of Open Access Journals (Sweden)

    Yi-An Chen

    Full Text Available Prioritising candidate genes for further experimental characterisation is a non-trivial challenge in drug discovery and biomedical research in general. An integrated approach that combines results from multiple data types is best suited for optimal target selection. We developed TargetMine, a data warehouse for efficient target prioritisation. TargetMine utilises the InterMine framework, with new data models such as protein-DNA interactions integrated in a novel way. It enables complicated searches that are difficult to perform with existing tools and it also offers integration of custom annotations and in-house experimental data. We proposed an objective protocol for target prioritisation using TargetMine and set up a benchmarking procedure to evaluate its performance. The results show that the protocol can identify known disease-associated genes with high precision and coverage. A demonstration version of TargetMine is available at http://targetmine.nibio.go.jp/.

  4. Epigenetic Editing : targeted rewriting of epigenetic marks to modulate expression of selected target genes

    NARCIS (Netherlands)

    de Groote, Marloes L.; Verschure, Pernette J.; Rots, Marianne G.

    2012-01-01

    Despite significant advances made in epigenetic research in recent decades, many questions remain unresolved, especially concerning cause and consequence of epigenetic marks with respect to gene expression modulation (GEM). Technologies allowing the targeting of epigenetic enzymes to predetermined D

  5. Epigenetic Editing: targeted rewriting of epigenetic marks to modulate expression of selected target genes.

    NARCIS (Netherlands)

    de Groote, M.L.; Verschure, P.J.; Rots, M.G.

    2012-01-01

    Despite significant advances made in epigenetic research in recent decades, many questions remain unresolved, especially concerning cause and consequence of epigenetic marks with respect to gene expression modulation (GEM). Technologies allowing the targeting of epigenetic enzymes to predetermined D

  6. The hair follicle as a target for gene therapy.

    Science.gov (United States)

    Gupta, S; Domashenko, A; Cotsarelis, G

    2001-01-01

    The hair follicle possesses progenitor cells for continued hair follicle cycling and for epidermal keratinocytes, melanocytes and Langerhans cells. These different cell types can be targeted by topical gene delivery to mouse skin. Using a combination of liposomes and DNA, we demonstrated the feasibility of targeting hair follicle cells in human scalp xenografts as well. We defined liposome composition and stage of the hair cycle as important parameters influencing transfection of human hair follicles. Transfection occurred only during anagen onset. Considerations and obstacles for using gene therapy to treat alopecias and skin disease are discussed. A theoretical framework for future gene therapy treatments for cutaneous and systemic disorders is presented.

  7. Viroreplicative Gene Therapy Targeted to Prostate Cancer

    Science.gov (United States)

    2010-08-01

    drug 5- fluorouracil ( 5FU ), as RCR vectors using this suicide gene have moved forward to Phase I clinical trials for the treatment of patients...mutations (T5.0002). The specific enzyme activity was measured by a calibrated HPLC assay to detect 5FU , the conversion product of the 5FC prodrug...in protein extracts from infected cells harvested 5 days post-infection at MOI = 0.1, and is expressed as nmol 5FU produced per min per mg protein

  8. Anti-Cancer Effects of Xanthones from Pericarps of Mangosteen

    Directory of Open Access Journals (Sweden)

    Yoshinori Nozawa

    2008-03-01

    Full Text Available Mangosteen, Garcinia mangostana Linn, is a tree found in South East Asia, and its pericarps have been used as traditional medicine. Phytochemical studies have shown that they contain a variety of secondary metabolites, such as oxygenated and prenylated xanthones. Recent studies revealed that these xanthones exhibited a variety of biological activities containing anti-inflammatory, anti-bacterial, and anti-cancer effects. We previously investigated the anti-proliferative effects of four prenylated xanthones from the pericarps; α-mangostin, β-mangostin, γ-mangostin, and methoxy-β-mangostin in various human cancer cells. These xanthones are different in the number of hydroxyl and methoxy groups. Except for methoxy-β-mangostin, the other three xanthones strongly inhibited cell growth at low concentrations from 5 to 20 μM in human colon cancer DLD-1 cells. Our recent study focused on the mechanism of α-mangostin-induced growth inhibition in DLD-1 cells. It was shown that the anti-proliferative effects of the xanthones were associated with cell-cycle arrest by affecting the expression of cyclins, cdc2, and p27; G1 arrest by α- mangostin and β-mangostin, and S arrest by γ-mangostin. α-Mangostin found to induce apoptosis through the activation of intrinsic pathway following the down-regulation of signaling cascades involving MAP kinases and the serine/threonine kinase Akt. Synergistic effects by the combined treatment of α-mangostin and anti-cancer drug 5-FU was to be noted. α-Mangostin was found to have a cancer preventive effect in rat carcinogenesis bioassay and the extract from pericarps, which contains mainly α-mangostin and γ- mangostin, exhibited an enhancement of NK cell activity in a mouse model. These findings could provide a relevant basis for the development of xanthones as an agent for cancer prevention and the combination therapy with

  9. Advancement in research of anti-cancer effects of toad venom (ChanSu) and perspectives

    Institute of Scientific and Technical Information of China (English)

    Miao Liu; Li-Xing Feng; Li-Hong Hu; Xuan Liu; De-An Guo

    2015-01-01

    Toad venom, called as ChanSu in China, is a widely used traditional Chinese medicine (TCM) whose active components are mainly bufadienolides. ChanSu could exhibit cardiotonic, anti-microbial, anti-inflammatory and, most importantly, anti-cancer effects. In the present review, reports about the in vitro, in vivo and clinical anti-cancer effects of ChanSu or its representative component, bufalin, were summarized. And, reported anti-cancer mechanisms of cardenolides, structure analogues of bufadienolides, were also introduced. Based on the results got from research of ChanSu/bufalin and the results from cardenolides, possible signal network related to the anti-cancer effects of ChanSu/bufalin was predicted. Furthermore, future potential use of ChanSu in anti-cancer therapy was discussed.

  10. Cancer gene therapy targeting angiogenesis: An updated review

    Institute of Scientific and Technical Information of China (English)

    Ching-Chiu Liu; Zan Shen; Hsiang-Fu Kung; Marie CM Lin

    2006-01-01

    Since the relationship between angiogenesis and tumor growth was established by Folkman in 1971,scientists have made efforts exploring the possibilities in treating cancer by targeting angiogenesis. Inhibition of angiogenesis growth factors and administration of angiogenesis inhibitors are the basics of antiangiogenesis therapy. Transfer of anti-angiogenesis genes has Received attention recently not only because of the advancement of recombinant vectors, but also because of the localized and sustained expression of therapeutic gene product inside the tumor after gene transfer. This review provides the up-to-date information about the strategies and the vectors studied in the field of anti-angiogenesis cancer gene therapy.

  11. Positive-negative-selection-mediated gene targeting in rice

    Directory of Open Access Journals (Sweden)

    Zenpei eShimatani

    2015-01-01

    Full Text Available Gene targeting (GT refers to the designed modification of genomic sequence(s through homologous recombination (HR. GT is a powerful tool both for the study of gene function and for molecular breeding. However, in transformation of higher plants, non-homologous end joining (NHEJ occurs overwhelmingly in somatic cells, masking HR-mediated GT. Positive-negative selection (PNS is an approach for finding HR-mediated GT events because it can eliminate NHEJ effectively by expression of a negative-selection marker gene. In rice—a major crop worldwide—reproducible PNS-mediated GT of endogenous genes has now been successfully achieved. The procedure is based on strong PNS using diphtheria toxin A-fragment as a negative marker, and has succeeded in the directed modification of several endogenous rice genes in various ways. In addition to gene knock-outs and knock-ins, a nucleotide substitution in a target gene was also achieved recently. This review presents a summary of the development of the rice PNS system, highlighting its advantages. Different types of gene modification and gene editing aimed at developing new plant breeding technology (NPBT based on PNS are discussed.

  12. Different Polycomb group complexes regulate common target genes in Arabidopsis.

    Science.gov (United States)

    Makarevich, Grigory; Leroy, Olivier; Akinci, Umut; Schubert, Daniel; Clarenz, Oliver; Goodrich, Justin; Grossniklaus, Ueli; Köhler, Claudia

    2006-09-01

    Polycomb group (PcG) proteins convey epigenetic inheritance of repressed transcriptional states. Although the mechanism of the action of PcG is not completely understood, methylation of histone H3 lysine 27 (H3K27) is important in establishing PcG-mediated transcriptional repression. We show that the plant PcG target gene PHERES1 is regulated by histone trimethylation on H3K27 residues mediated by at least two different PcG complexes in plants, containing the SET domain proteins MEDEA or CURLY LEAF/SWINGER. Furthermore, we identify FUSCA3 as a potential PcG target gene and show that FUSCA3 is regulated by MEDEA and CURLY LEAF/SWINGER. We propose that different PcG complexes regulate a common set of target genes during the different stages of plant development.

  13. Hypoxia-regulated target genes implicated in tumor metastasis

    Directory of Open Access Journals (Sweden)

    Tsai Ya-Ping

    2012-12-01

    Full Text Available Abstract Hypoxia is an important microenvironmental factor that induces cancer metastasis. Hypoxia/hypoxia-inducible factor-1α (HIF-1α regulates many important steps of the metastatic processes, especially epithelial-mesenchymal transition (EMT that is one of the crucial mechanisms to cause early stage of tumor metastasis. To have a better understanding of the mechanism of hypoxia-regulated metastasis, various hypoxia/HIF-1α-regulated target genes are categorized into different classes including transcription factors, histone modifiers, enzymes, receptors, kinases, small GTPases, transporters, adhesion molecules, surface molecules, membrane proteins, and microRNAs. Different roles of these target genes are described with regards to their relationship to hypoxia-induced metastasis. We hope that this review will provide a framework for further exploration of hypoxia/HIF-1α-regulated target genes and a comprehensive view of the metastatic picture induced by hypoxia.

  14. Targeting of AID-mediated sequence diversification to immunoglobulin genes.

    Science.gov (United States)

    Kothapalli, Naga Rama; Fugmann, Sebastian D

    2011-04-01

    Activation-induced cytidine deaminase (AID) is a key enzyme for antibody-mediated immune responses. Antibodies are encoded by the immunoglobulin genes and AID acts as a transcription-dependent DNA mutator on these genes to improve antibody affinity and effector functions. An emerging theme in field is that many transcribed genes are potential targets of AID, presenting an obvious danger to genomic integrity. Thus there are mechanisms in place to ensure that mutagenic outcomes of AID activity are specifically restricted to the immunoglobulin loci. Cis-regulatory targeting elements mediate this effect and their mode of action is probably a combination of immunoglobulin gene specific activation of AID and a perversion of faithful DNA repair towards error-prone outcomes.

  15. Pancreatic Cancer Gene Therapy: From Molecular Targets to Delivery Systems

    Energy Technology Data Exchange (ETDEWEB)

    Fillat, Cristina, E-mail: cristina.fillat@crg.es; Jose, Anabel; Ros, Xavier Bofill-De; Mato-Berciano, Ana; Maliandi, Maria Victoria; Sobrevals, Luciano [Programa Gens i Malaltia, Centre de Regulació Genòmica-CRG, UPF, Parc de Recerca Biomedica de Barcelona-PRBB and Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), Barcelona (Spain)

    2011-01-18

    The continuous identification of molecular changes deregulating critical pathways in pancreatic tumor cells provides us with a large number of novel candidates to engineer gene-targeted approaches for pancreatic cancer treatment. Targets—both protein coding and non-coding—are being exploited in gene therapy to influence the deregulated pathways to facilitate cytotoxicity, enhance the immune response or sensitize to current treatments. Delivery vehicles based on viral or non-viral systems as well as cellular vectors with tumor homing characteristics are a critical part of the design of gene therapy strategies. The different behavior of tumoral versus non-tumoral cells inspires vector engineering with the generation of tumor selective products that can prevent potential toxic-associated effects. In the current review, a detailed analysis of the different targets, the delivery vectors, the preclinical approaches and a descriptive update on the conducted clinical trials are presented. Moreover, future possibilities in pancreatic cancer treatment by gene therapy strategies are discussed.

  16. Transcriptionally regulated, prostate-targeted gene therapy for prostate cancer.

    Science.gov (United States)

    Lu, Yi

    2009-07-02

    Prostate cancer is the most frequently diagnosed cancer and the second leading cause of cancer deaths in American males today. Novel and effective treatment such as gene therapy is greatly desired. The early viral based gene therapy uses tissue-nonspecific promoters, which causes unintended toxicity to other normal tissues. In this chapter, we will review the transcriptionally regulated gene therapy strategy for prostate cancer treatment. We will describe the development of transcriptionally regulated prostate cancer gene therapy in the following areas: (1) Comparison of different routes for best viral delivery to the prostate; (2) Study of transcriptionally regulated, prostate-targeted viral vectors: specificity and activity of the transgene under several different prostate-specific promoters were compared in vitro and in vivo; (3) Selection of therapeutic transgenes and strategies for prostate cancer gene therapy (4) Oncolytic virotherapy for prostate cancer. In addition, the current challenges and future directions in this field are also discussed.

  17. Gene Targeting and Expression Modulation by Peptide Nucleic Acids (PNA)

    DEFF Research Database (Denmark)

    Nielsen, Peter E

    2010-01-01

    Peptide nucleic acids (PNA) are artificial structural mimics of nucleic acids capable of sequence specific hybridization to both RNA and DNA. Thus they have obvious potential as gene targeting agents for drug discovery approaches. An overview with emphasis on recent progress on RNA "interference"...

  18. E2F target genes: unraveling the biology

    DEFF Research Database (Denmark)

    Bracken, Adrian P; Ciro, Marco; Cocito, Andrea

    2004-01-01

    The E2F transcription factors are downstream effectors of the retinoblastoma protein (pRB) pathway and are required for the timely regulation of numerous genes essential for DNA replication and cell cycle progression. Several laboratories have used genome-wide approaches to discover novel target ...

  19. Bacteriophages and medical oncology: targeted gene therapy of cancer.

    Science.gov (United States)

    Bakhshinejad, Babak; Karimi, Marzieh; Sadeghizadeh, Majid

    2014-08-01

    Targeted gene therapy of cancer is of paramount importance in medical oncology. Bacteriophages, viruses that specifically infect bacterial cells, offer a variety of potential applications in biomedicine. Their genetic flexibility to go under a variety of surface modifications serves as a basis for phage display methodology. These surface manipulations allow bacteriophages to be exploited for targeted delivery of therapeutic genes. Moreover, the excellent safety profile of these viruses paves the way for their potential use as cancer gene therapy platforms. The merge of phage display and combinatorial technology has led to the emergence of phage libraries turning phage display into a high throughput technology. Random peptide libraries, as one of the most frequently used phage libraries, provide a rich source of clinically useful peptide ligands. Peptides are known as a promising category of pharmaceutical agents in medical oncology that present advantages such as inexpensive synthesis, efficient tissue penetration and the lack of immunogenicity. Phage peptide libraries can be screened, through biopanning, against various targets including cancer cells and tissues that results in obtaining cancer-homing ligands. Cancer-specific peptides isolated from phage libraries show huge promise to be utilized for targeting of various gene therapy vectors towards malignant cells. Beyond doubt, bacteriophages will play a more impressive role in the future of medical oncology.

  20. Recombinant adenovirus vectors with knobless fibers for targeted gene transfer

    NARCIS (Netherlands)

    van Beusechem, VW; van Rijswijk, ALCT; van Es, HHG; Haisma, HJ; Pinedo, HM; Gerritsen, WR

    2000-01-01

    Adenoviral vector systems for gene therapy can be much improved by targeting vectors to specific cell types. This requires both the complete ablation of native adenovirus tropism and the introduction of a novel binding affinity in the viral capsid. We reasoned that these requirements could be fulfil

  1. [The hair follicle as a target for gene therapy].

    Science.gov (United States)

    Cotsarelis, G

    2002-05-01

    The hair follicle possesses progenitor cells required for continuous hair follicle cycling and for epidermal keratinocytes, melanocytes and Langerhans cells. These different cell types can be the target of topical gene delivery in the skin of the mouse. Using a combination of liposomes and DNA, we demonstrate the feasibility of targeting hair follicle cells in human scalp xenografts. We consider liposome composition and stage of the hair cycle as important parameters influencing transfection of human hair follicles. Transfection is possible only during the early anagen phase. Factors and obstacles for the use of gene therapy in treating alopecia and skin diseases are discussed. A theoretical framework for future treatment of cutaneous and systemic disorders using gene therapy is presented.

  2. DEPTOR-related mTOR suppression is involved in metformin's anti-cancer action in human liver cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Obara, Akio; Fujita, Yoshihito; Abudukadier, Abulizi; Fukushima, Toru; Oguri, Yasuo; Ogura, Masahito; Harashima, Shin-ichi; Hosokawa, Masaya; Inagaki, Nobuya, E-mail: inagaki@metab.kuhp.kyoto-u.ac.jp

    2015-05-15

    Metformin, one of the most commonly used drugs for patients with type 2 diabetes, recently has received much attention regarding its anti-cancer action. It is thought that the suppression of mTOR signaling is involved in metformin's anti-cancer action. Although liver cancer is one of the most responsive types of cancer for reduction of incidence by metformin, the molecular mechanism of the suppression of mTOR in liver remains unknown. In this study, we investigated the mechanism of the suppressing effect of metformin on mTOR signaling and cell proliferation using human liver cancer cells. Metformin suppressed phosphorylation of p70-S6 kinase, and ribosome protein S6, downstream targets of mTOR, and suppressed cell proliferation. We found that DEPTOR, an endogenous substrate of mTOR suppression, is involved in the suppressing effect of metformin on mTOR signaling and cell proliferation in human liver cancer cells. Metformin increases the protein levels of DEPTOR, intensifies binding to mTOR, and exerts a suppressing effect on mTOR signaling. This increasing effect of DEPTOR by metformin is regulated by the proteasome degradation system; the suppressing effect of metformin on mTOR signaling and cell proliferation is in a DEPTOR-dependent manner. Furthermore, metformin exerts a suppressing effect on proteasome activity, DEPTOR-related mTOR signaling, and cell proliferation in an AMPK-dependent manner. We conclude that DEPTOR-related mTOR suppression is involved in metformin's anti-cancer action in liver, and could be a novel target for anti-cancer therapy. - Highlights: • We elucidated a novel pathway of metformin's anti-cancer action in HCC cells. • DEPTOR is involved in the suppressing effect of metformin on mTOR signaling. • Metformin increases DEPTOR protein levels via suppression of proteasome activity. • DEPTOR-related mTOR suppression is involved in metformin's anti-cancer action.

  3. Integrative analysis of RUNX1 downstream pathways and target genes

    Directory of Open Access Journals (Sweden)

    Liu Marjorie

    2008-07-01

    Full Text Available Abstract Background The RUNX1 transcription factor gene is frequently mutated in sporadic myeloid and lymphoid leukemia through translocation, point mutation or amplification. It is also responsible for a familial platelet disorder with predisposition to acute myeloid leukemia (FPD-AML. The disruption of the largely unknown biological pathways controlled by RUNX1 is likely to be responsible for the development of leukemia. We have used multiple microarray platforms and bioinformatic techniques to help identify these biological pathways to aid in the understanding of why RUNX1 mutations lead to leukemia. Results Here we report genes regulated either directly or indirectly by RUNX1 based on the study of gene expression profiles generated from 3 different human and mouse platforms. The platforms used were global gene expression profiling of: 1 cell lines with RUNX1 mutations from FPD-AML patients, 2 over-expression of RUNX1 and CBFβ, and 3 Runx1 knockout mouse embryos using either cDNA or Affymetrix microarrays. We observe that our datasets (lists of differentially expressed genes significantly correlate with published microarray data from sporadic AML patients with mutations in either RUNX1 or its cofactor, CBFβ. A number of biological processes were identified among the differentially expressed genes and functional assays suggest that heterozygous RUNX1 point mutations in patients with FPD-AML impair cell proliferation, microtubule dynamics and possibly genetic stability. In addition, analysis of the regulatory regions of the differentially expressed genes has for the first time systematically identified numerous potential novel RUNX1 target genes. Conclusion This work is the first large-scale study attempting to identify the genetic networks regulated by RUNX1, a master regulator in the development of the hematopoietic system and leukemia. The biological pathways and target genes controlled by RUNX1 will have considerable importance in disease

  4. Perspectives in Engineered Mesenchymal Stem/Stromal Cells Based Anti- Cancer Drug Delivery Systems.

    Science.gov (United States)

    Ackova, Darinka Gjorgieva; Kanjevac, Tatjana; Rimondini, Lia; Bosnakovski, Darko

    2016-01-01

    Understanding and apprehension of the characteristics and circumstances in which mesenchymal stem cells (MSCs) affect and make alterations (enhance or reduce) to the growth of tumors and metastasis spread is pivotal, not only for reaching the possibility to employ MSCs as drug delivery systems, but also for making forward movement in the existing knowledge of involvement of major factors (tumor microenvironment, soluble signaling molecules, etc.) in the process of carcinogenesis. This capability is reliable because MSCs present a great basis for engineering and constructions of new systems to target cancers, intended to secrete therapeutic proteins in the tumor region, or for delivering of oncolytic viruses' directly at the tumor site (targeted chemotherapy with enzyme prodrug conversion or induction of tumor cell apoptosis). MSCs as a crucial segment of the tumor surroundings and their confirmed tumor tropism, are assumed to be an open gateway for the design of promising drug delivery systems. The presented paper reviews current publications in this fieldwork, searches out the most recent patents that were published after 2012 (WO2014066122, US20140017787, WO2015100268, US20150086515), and tries to present the current progress and future prospective on the design and development in anti-cancer drug delivery systems based on MSCs.

  5. SWCNT-Polymer Nanocomplexes for Anti-Cancer Drug Delivery

    Science.gov (United States)

    Withey, Paul; Momin, Zoya; Bommoju, Anvesh; Hoang, Trung; Rashid, Bazlur

    2015-03-01

    Utilization of single-walled carbon nanotubes (SWCNTs) as more effective drug-delivery agents are being considered due to their ability to easily cross cell membranes, while their high aspect ratio and large surface area provide multiple attachment sites for biocompatible drug complexes. However, excessive bundling of pristine SWCNTs caused by strong attractive Van der Walls forces between CNT sidewalls is a major obstacle. We have successfully dispersed SWCNTs with both polyvinyl alcohol and Pluronic biocompatible polymers, and attached anti-cancer drugs Camptothecin (CPT) and Doxorubicin to form non-covalent CNT-polymer-drug conjugates in aqueous solution. Polymeric dispersion of SWCNTs by both polymers is confirmed by clearly identifiable near-infrared (NIR) fluorescence emission peaks of individual (7,5) and (7,6) nanotubes, and drug attachment to form complete complexes verified by UV-Vis spectroscopy. These complexes, with varying SWCNT and drug concentrations, were tested for effectiveness by exposing them to a line of human embryonic kidney cancer cells and analyzed for cell viability. Preliminary results indicate significant improvement in drug effectiveness on the cancer cells, with more successful internalization due to unaltered SWCNTs as the drug carriers. Supported by the UHCL Faculty Research Support Fund.

  6. Annotating Cancer Variants and Anti-Cancer Therapeutics in Reactome

    Energy Technology Data Exchange (ETDEWEB)

    Milacic, Marija; Haw, Robin, E-mail: robin.haw@oicr.on.ca; Rothfels, Karen; Wu, Guanming [Informatics and Bio-computing Platform, Ontario Institute for Cancer Research, Toronto, ON, M5G0A3 (Canada); Croft, David; Hermjakob, Henning [European Bioinformatics Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, CB10 1SD (United Kingdom); D’Eustachio, Peter [Department of Biochemistry, NYU School of Medicine, New York, NY 10016 (United States); Stein, Lincoln [Informatics and Bio-computing Platform, Ontario Institute for Cancer Research, Toronto, ON, M5G0A3 (Canada)

    2012-11-08

    Reactome describes biological pathways as chemical reactions that closely mirror the actual physical interactions that occur in the cell. Recent extensions of our data model accommodate the annotation of cancer and other disease processes. First, we have extended our class of protein modifications to accommodate annotation of changes in amino acid sequence and the formation of fusion proteins to describe the proteins involved in disease processes. Second, we have added a disease attribute to reaction, pathway, and physical entity classes that uses disease ontology terms. To support the graphical representation of “cancer” pathways, we have adapted our Pathway Browser to display disease variants and events in a way that allows comparison with the wild type pathway, and shows connections between perturbations in cancer and other biological pathways. The curation of pathways associated with cancer, coupled with our efforts to create other disease-specific pathways, will interoperate with our existing pathway and network analysis tools. Using the Epidermal Growth Factor Receptor (EGFR) signaling pathway as an example, we show how Reactome annotates and presents the altered biological behavior of EGFR variants due to their altered kinase and ligand-binding properties, and the mode of action and specificity of anti-cancer therapeutics.

  7. From T cell "exhaustion" to anti-cancer immunity.

    Science.gov (United States)

    Verdeil, Grégory; Fuertes Marraco, Silvia A; Murray, Timothy; Speiser, Daniel E

    2016-01-01

    The immune system has the potential to protect from malignant diseases for extended periods of time. Unfortunately, spontaneous immune responses are often inefficient. Significant effort is required to develop reliable, broadly applicable immunotherapies for cancer patients. A major innovation was transplantation with hematopoietic stem cells from genetically distinct donors for patients with hematologic malignancies. In this setting, donor T cells induce long-term remission by keeping cancer cells in check through powerful allogeneic graft-versus-leukemia effects. More recently, a long awaited breakthrough for patients with solid tissue cancers was achieved, by means of therapeutic blockade of T cell inhibitory receptors. In untreated cancer patients, T cells are dysfunctional and remain in a state of T cell "exhaustion". Nonetheless, they often retain a high potential for successful defense against cancer, indicating that many T cells are not entirely and irreversibly exhausted but can be mobilized to become highly functional. Novel antibody therapies that block inhibitory receptors can lead to strong activation of anti-tumor T cells, mediating clinically significant anti-cancer immunity for many years. Here we review these new treatments and the current knowledge on tumor antigen-specific T cells.

  8. Identification of novel androgen receptor target genes in prostate cancer

    Directory of Open Access Journals (Sweden)

    Gerald William L

    2007-06-01

    Full Text Available Abstract Background The androgen receptor (AR plays critical roles in both androgen-dependent and castrate-resistant prostate cancer (PCa. However, little is known about AR target genes that mediate the receptor's roles in disease progression. Results Using Chromatin Immunoprecipitation (ChIP Display, we discovered 19 novel loci occupied by the AR in castrate resistant C4-2B PCa cells. Only four of the 19 AR-occupied regions were within 10-kb 5'-flanking regulatory sequences. Three were located up to 4-kb 3' of the nearest gene, eight were intragenic and four were in gene deserts. Whereas the AR occupied the same loci in C4-2B (castrate resistant and LNCaP (androgen-dependent PCa cells, differences between the two cell lines were observed in the response of nearby genes to androgens. Among the genes strongly stimulated by DHT in C4-2B cells – D-dopachrome tautomerase (DDT, Protein kinase C delta (PRKCD, Glutathione S- transferase theta 2 (GSTT2, Transient receptor potential cation channel subfamily V member 3 (TRPV3, and Pyrroline-5-carboxylate reductase 1 (PYCR1 – most were less strongly or hardly stimulated in LNCaP cells. Another AR target gene, ornithine aminotransferase (OAT, was AR-stimulated in a ligand-independent manner, since it was repressed by AR siRNA knockdown, but not stimulated by DHT. We also present evidence for in vivo AR-mediated regulation of several genes identified by ChIP Display. For example, PRKCD and PYCR1, which may contribute to PCa cell growth and survival, are expressed in PCa biopsies from primary tumors before and after ablation and in metastatic lesions in a manner consistent with AR-mediated stimulation. Conclusion AR genomic occupancy is similar between LNCaP and C4-2B cells and is not biased towards 5' gene flanking sequences. The AR transcriptionally regulates less than half the genes nearby AR-occupied regions, usually but not always, in a ligand-dependent manner. Most are stimulated and a few are

  9. Salinomycin: a novel anti-cancer agent with known anti-coccidial activities.

    Science.gov (United States)

    Zhou, Shuang; Wang, Fengfei; Wong, Eric T; Fonkem, Ekokobe; Hsieh, Tze-Chen; Wu, Joseph M; Wu, Erxi

    2013-01-01

    Salinomycin, traditionally used as an anti-coccidial drug, has recently been shown to possess anti-cancer and anti-cancer stem cell (CSC) effects, as well as activities to overcome multi-drug resistance based on studies using human cancer cell lines, xenograft mice, and in case reports involving cancer patients in pilot clinical trials. Therefore, salinomycin may be considered as a promising novel anti-cancer agent despite its largely unknown mechanism of action. This review summarizes the pharmacologic effects of salinomycin and presents possible mechanisms by which salinomycin exerts its anti-tumorigenic activities. Recent advances and potential complications that might limit the utilization of salinomycin as an anti-cancer and anti-CSC agent are also presented and discussed.

  10. RFMirTarget: predicting human microRNA target genes with a random forest classifier.

    Directory of Open Access Journals (Sweden)

    Mariana R Mendoza

    Full Text Available MicroRNAs are key regulators of eukaryotic gene expression whose fundamental role has already been identified in many cell pathways. The correct identification of miRNAs targets is still a major challenge in bioinformatics and has motivated the development of several computational methods to overcome inherent limitations of experimental analysis. Indeed, the best results reported so far in terms of specificity and sensitivity are associated to machine learning-based methods for microRNA-target prediction. Following this trend, in the current paper we discuss and explore a microRNA-target prediction method based on a random forest classifier, namely RFMirTarget. Despite its well-known robustness regarding general classifying tasks, to the best of our knowledge, random forest have not been deeply explored for the specific context of predicting microRNAs targets. Our framework first analyzes alignments between candidate microRNA-target pairs and extracts a set of structural, thermodynamics, alignment, seed and position-based features, upon which classification is performed. Experiments have shown that RFMirTarget outperforms several well-known classifiers with statistical significance, and that its performance is not impaired by the class imbalance problem or features correlation. Moreover, comparing it against other algorithms for microRNA target prediction using independent test data sets from TarBase and starBase, we observe a very promising performance, with higher sensitivity in relation to other methods. Finally, tests performed with RFMirTarget show the benefits of feature selection even for a classifier with embedded feature importance analysis, and the consistency between relevant features identified and important biological properties for effective microRNA-target gene alignment.

  11. C/EBPδ gene targets in human keratinocytes.

    Directory of Open Access Journals (Sweden)

    Serena Borrelli

    Full Text Available C/EBPs are a family of B-Zip transcription factors--TFs--involved in the regulation of differentiation in several tissues. The two most studied members--C/EBPα and C/EBPβ--play important roles in skin homeostasis and their ablation reveals cells with stem cells signatures. Much less is known about C/EBPδ which is highly expressed in the granular layer of interfollicular epidermis and is a direct target of p63, the master regular of multilayered epithelia. We identified C/EBPδ target genes in human primary keratinocytes by ChIP on chip and profiling of cells functionally inactivated with siRNA. Categorization suggests a role in differentiation and control of cell-cycle, particularly of G2/M genes. Among positively controlled targets are numerous genes involved in barrier function. Functional inactivation of C/EBPδ as well as overexpressions of two TF targets--MafB and SOX2--affect expression of markers of keratinocyte differentiation. We performed IHC on skin tumor tissue arrays: expression of C/EBPδ is lost in Basal Cell Carcinomas, but a majority of Squamous Cell Carcinomas showed elevated levels of the protein. Our data indicate that C/EBPδ plays a role in late stages of keratinocyte differentiation.

  12. Identification of novel Notch target genes in T cell leukaemia

    Directory of Open Access Journals (Sweden)

    Warrander Fiona

    2009-06-01

    Full Text Available Abstract Background Dysregulated Notch signalling is believed to play an important role in the development and maintenance of T cell leukaemia. At a cellular level, Notch signalling promotes proliferation and inhibits apoptosis of T cell acute lymphoblastic leukaemia (T-ALL cells. In this study we aimed to identify novel transcriptional targets of Notch signalling in the T-ALL cell line, Jurkat. Results RNA was prepared from Jurkat cells retrovirally transduced with an empty vector (GFP-alone or vectors containing constitutively active forms of Notch (N1ΔE or N3ΔE, and used for Affymetrix microarray analysis. A subset of genes found to be regulated by Notch was chosen for real-time PCR validation and in some cases, validation at the protein level, using several Notch-transduced T-ALL and non-T-ALL leukaemic cell lines. As expected, several known transcriptional target of Notch, such as HES1 and Deltex, were found to be overexpressed in Notch-transduced cells, however, many novel transcriptional targets of Notch signalling were identified using this approach. These included the T cell costimulatory molecule CD28, the anti-apoptotic protein GIMAP5, and inhibitor of DNA binding 1 (1D1. Conclusion The identification of such downstream Notch target genes provides insights into the mechanisms of Notch function in T cell leukaemia, and may help identify novel therapeutic targets in this disease.

  13. A Novel Gene Delivery System Targeting Urokinase Receptor

    Institute of Scientific and Technical Information of China (English)

    Xing-Hui SUN; Li TAN; Chun-Yang LI; Chang TONG; Jin FAN; Ping LI; Yun-Song ZHU

    2004-01-01

    Recombinant proteins that combine different functions required for cell targeting and intracellular delivery of DNA present an attractive approach for the development of nonviral gene delivery vectors. Here, we described a novel protein termed ATF-lys10 which facilitated cell-specific gene transfer via receptor-mediated endocytosis. ATF-lys 10 was composed of the amino-terminal fragment of urokinase and ten lysines at the carboxyl terminus. Bacterially expressed ATF-lys 10 protein existed in soluble form, and had antigenicity of human urokinase. Purified ATF-lys 10 specifically bound to uPAR-expressing cells and formed protein-DNA complexes with plasmid pGL3-control. After neutralization of excess negative charge with poly-L-lysine, these complexes served as a specific gene delivery vector for uPAR-expressing cells. Lysosomotropic compounds, such as chloroquine, drastically increased the ATF-lysl0 mediated gene delivery efficiency. Our results suggest that the recombinant protein ATF-lys 10 with the properties of DNA binding and tumor cell targeting represents a promising method for gene transfer and expression in tumor cells.

  14. RNA Interference Targeting Leptin Gene Effect on Hepatic Stellate Cells

    Institute of Scientific and Technical Information of China (English)

    XUE Xiulan; LIN Jusheng; SONG Yuhu; SUN Xuemei; ZHOU Hejun

    2005-01-01

    To construct the specific siRNA expression vectors and investigate their effect on leptin and collagen I in HSC, which provide a new approach to the prevent and treat hepatic fibrosis. The five siRNAs against leptin gene were transcript synthesized intracellularly by expression templates of plasmid vector psiRNA-hH1neo. The recombinant leptin siRNA plasmid vectors could express in eukaryocyte , and then to evaluate them by using enzyme cutting and sequencing. The recombinant plasmids were transfected into HSCs using Lipofectamine methods respectively. The cells were selected after growing in DMEM containing 300 μg/mL G418 for about 4 weeks. Gene expression of leptin and collagen I were showed by Western blot analysis and reverse transcription polymerase chain reaction (RT-PCR). Identification by enzyme cutting and sequencing showed that the leptin siRNA expression vectors were constructed successfully, and leptin siRNA could inhibit the leptin and collagen I gene expression effectively. It was concluded that RNA interference-mediated silencing of leptin gene diminished leptin and collagen I gene expression in HSCs. Furthermore, attenuated the extracellular matrix over-deposition at the same time. Leptin gene is ideal targets of gene therapy for liver fibrosis.

  15. Double-strand breaks at the target locus stimulate gene targeting in embryonic stem cells.

    Science.gov (United States)

    Smih, F; Rouet, P; Romanienko, P J; Jasin, M

    1995-01-01

    Double-strand breaks (DSBs) are recombinogenic lesions in chromosomal DNA in yeast, Drosophila and Caenorhabditis elegans. Recent studies in mammalian cells utilizing the I-Scel endonuclease have demonstrated that in some immortalized cell lines DSBs in chromosomal DNA are also recombinogenic. We have now tested embryonic stem (ES) cells, a non-transformed mouse cell line frequently used in gene targeting studies. We find that a DSB introduced by I-Scel stimulates gene targeting at a selectable neo locus at least 50-fold. The enhanced level of targeting is achieved by transient expression of the I-Scel endonuclease. In 97% of targeted clones a single base pair polymorphism in the transfected homologous fragment was incorporated into the target locus. Analysis of the targeted locus demonstrated that most of the homologous recombination events were 'two-sided', in contrast to previous studies in 3T3 cells in which 'one-sided' homologous events predominated. Thus ES cells may be more faithful in incorporating homologous fragments into their genome than other cells in culture. Images PMID:8559659

  16. Technology whitespaces India should focus: a comparative anti-cancer patent rational analysis of Indian and international public funded universities.

    Science.gov (United States)

    Dara, Ajay; Sangamwar, Abhay T

    2015-01-01

    The article reported an in-depth comparative technology analysis of 1708 Anti-cancer patents from top 20 international universities and leading 10 Indian public funded organization and research institutes. The study segregated pioneer universities vs. technologies used in the field of Anticancer research at a level of drug discovery, development, diagnosis and treatment, which are illustrated in the form of novel substantive patent landscape maps. The reported competitive intelligent maps identified genetics, composition and synthetic compounds as dominating technologies; followed by natural extracts, combination and drug delivery systems as upcoming technologies. The least number of patents were reported by surgical apparatus, targeted therapy and animal models. In addition, the study analysed the key technologies followed by Indian universities in comparison to the international universities, to identify the overlooked technologies by the Indian public funded institutes. In an ever changing competitive world, it is essential for every university to have their own research plan and thrust areas; but at the same time, it is equally important for any organisation to have an idea of their competitor's research plan as well. So, the article suggested Indian institutes to focus on the latest emerging Anti-cancer technology trends, which are in practice by the international universities. Concurrently, this study may be a landmark indication for Indian public funded universities and institutes, calling for a U-turn from their traditional approaches.

  17. Reproducible gene targeting in recalcitrant Escherichia coli isolates

    Directory of Open Access Journals (Sweden)

    De Greve Henri

    2011-06-01

    Full Text Available Abstract Background A number of allele replacement methods can be used to mutate bacterial genes. For instance, the Red recombinase system of phage Lambda has been used very efficiently to inactivate chromosomal genes in E. coli K-12, through recombination between regions of homology. However, this method does not work reproducibly in some clinical E. coli isolates. Findings The procedure was modified by using longer homologous regions (85 bp and 500-600 bp, to inactivate genes in the uropathogenic E. coli strain UTI89. An lrhA regulator mutant, and deletions of the lac operon as well as the complete type 1 fimbrial gene cluster, were obtained reproducibly. The modified method is also functional in other recalcitrant E. coli, like the avian pathogenic E. coli strain APEC1. The lrhA regulator and lac operon deletion mutants of APEC1 were successfully constructed in the same way as the UTI89 mutants. In other avian pathogenic E. coli strains (APEC3E, APEC11A and APEC16A it was very difficult or impossible to construct these mutants, with the original Red recombinase-based method, with a Red recombinase-based method using longer (85 bp homologous regions or with our modified protocol, using 500 - 600 bp homologous regions. Conclusions The method using 500-600 bp homologous regions can be used reliably in some clinical isolates, to delete single genes or entire operons by homologous recombination. However, it does not invariably show a greater efficiency in obtaining mutants, when compared to the original Red-mediated gene targeting method or to the gene targeting method with 85 bp homologous regions. Therefore the length of the homology regions is not the only limiting factor for the construction of mutants in these recalcitrant strains.

  18. Mannan-Modified PLGA Nanoparticles for Targeted Gene Delivery

    Directory of Open Access Journals (Sweden)

    Fansheng Kong

    2012-01-01

    Full Text Available The studies of targeted gene delivery nanocarriers have gained increasing attention during the past decades. In this study, mannan modified DNA loaded bioadhesive PLGA nanoparticles (MAN-DNA-NPs were investigated for targeted gene delivery to the Kupffer cells (KCs. Bioadhesive PLGA nanoparticles were prepared and subsequently bound with pEGFP. Following the coupling of the mannan-based PE-grafted ligands (MAN-PE with the DNA-NPs, the MAN-DNA-NPs were delivered intravenously to rats. The transfection efficiency was determined from the isolated KCs and flow cytometry was applied for the quantitation of gene expression after 48 h post transfection. The size of the MAN-DNA-NPs was found to be around 190 nm and the Zeta potential was determined to be −15.46mV. The pEGFP binding capacity of MAN-DNA-NPs was (88.9±5.8% and the in vitro release profiles of the MAN-DNA-NPs follow the Higuchi model. When compared with non-modified DNA-NPs and Lipofectamine 2000-DNA, MAN-DNA-NPs produced the highest gene expressions, especially in vivo. The in vivo data from flow cytometry analysis showed that MAN-DNA-NPs displayed a remarkably higher transfection efficiency (39% than non-modified DNA-NPs (25% and Lipofectamine 2000-DNA (23% in KCs. The results illustrate that MAN-DNA-NPs have the ability to target liver KCs and could function as promising active targeting drug delivery vectors.

  19. Tapping natural reservoirs of homing endonucleases for targeted gene modification

    OpenAIRE

    2011-01-01

    Homing endonucleases mobilize their own genes by generating double-strand breaks at individual target sites within potential host DNA. Because of their high specificity, these proteins are used for “genome editing” in higher eukaryotes. However, alteration of homing endonuclease specificity is quite challenging. Here we describe the identification and phylogenetic analysis of over 200 naturally occurring LAGLIDADG homing endonucleases (LHEs). Biochemical and structural characterization of end...

  20. Targeted gene repair: the ups and downs of a promising gene therapy approach.

    Science.gov (United States)

    de Semir, David; Aran, Josep M

    2006-08-01

    As a novel form of molecular medicine based on direct actions over the genes, targeted gene repair has raised consideration recently above classical gene therapy strategies based on genetic augmentation or complementation. Targeted gene repair relies on the local induction of the cell's endogenous DNA repair mechanisms to attain a therapeutic gene conversion event within the genome of the diseased cell. Successful repair has been achieved both in vitro and in vivo with a variety of corrective molecules ranging from oligonucleotides (chimeraplasts, modified single-stranded oligonucleotides, triplex-forming oligonucleotides), to small DNA fragments (small fragment homologous replacement (SFHR)), and even viral vectors (AAV-based). However, controversy on the consistency and lack of reproducibility of early experiments regarding frequencies and persistence of targeted gene repair, particularly for chimeraplasty, has flecked the field. Nevertheless, several hurdles such as inefficient nuclear uptake of the corrective molecules, and misleading assessment of targeted repair frequencies have been identified and are being addressed. One of the key bottlenecks for exploiting the overall potential of the different targeted gene repair modalities is the lack of a detailed knowledge of their mechanisms of action at the molecular level. Several studies are now focusing on the assessment of the specific repair pathway(s) involved (homologous recombination, mismatch repair, etc.), devising additional strategies to increase their activity (using chemotherapeutic drugs, chimeric nucleases, etc.), and assessing the influence of the cell cycle in the regulation of the repair process. Until therapeutic correction frequencies for single gene disorders are reached both in cellular and animal models, precision and undesired side effects of this promising gene therapy approach will not be thoroughly evaluated.

  1. Construction of gene targeting vectors from lambda KOS genomic libraries.

    Science.gov (United States)

    Wattler, S; Kelly, M; Nehls, M

    1999-06-01

    We describe a highly redundant murine genomic library in a new lambda phage, lambda knockout shuttle (lambda KOS) that facilitates the very rapid construction of replacement-type gene targeting vectors. The library consists of 94 individually amplified subpools, each containing an average of 40,000 independent genomic clones. The subpools are arrayed into a 96-well format that allows a PCR-based efficient recovery of independent genomic clones. The lambda KOS vector backbone permits the CRE-mediated conversion into high-copy number pKOS plasmids, wherein the genomic inserts are automatically flanked by negative-selection cassettes. The lambda KOS vector system exploits the yeast homologous recombination machinery to simplify the construction of replacement-type gene targeting vectors independent of restriction sites within the genomic insert. We outline procedures that allow the generation of simple and more sophisticated conditional gene targeting vectors within 3-4 weeks, beginning with the screening of the lambda KOS genomic library.

  2. Treating psoriasis by targeting its susceptibility gene Rel.

    Science.gov (United States)

    Fan, Tingting; Wang, Shaowen; Yu, Linjiang; Yi, Huqiang; Liu, Ruiling; Geng, Wenwen; Wan, Xiaochun; Ma, Yifan; Cai, Lintao; Chen, Youhai H; Ruan, Qingguo

    2016-04-01

    Psoriasis is a chronic inflammatory disorder of the skin. Accumulating evidence indicates that the Rel gene, a member of the NF-κB family, is a risk factor for the disease. We sought to investigate whether psoriasis can be prevented by directly targeting the Rel gene transcript, i.e., the c-Rel mRNA. Using chemically-modified c-Rel specific siRNA (siRel) and poly(ethylene glycol)-b-poly(l-lysine)-b-poly(l-leucine) (PEG-PLL-PLLeu) micelles, we successfully knocked down the expression of c-Rel, and showed that the expression of cytokine IL-23, a direct target of c-Rel that can drive the development of IL-17-producing T cells, was markedly inhibited. More importantly, treating mice with siRel not only prevented but also ameliorated imiquimod (IMQ)-induced psoriasis. Mechanistic studies showed that siRel treatment down-regulated the expression of multiple inflammatory cytokines. Taken together, these results indicate that the susceptibility gene Rel can be targeted to treat and prevent psoriasis.

  3. HDAC Inhibitors as Novel Anti-Cancer Therapeutics.

    Science.gov (United States)

    De Souza, Cristabelle; Chatterji, Biswa Prasun

    2015-01-01

    Malignant growth of cells is a condition characterized by unchecked cellular proliferation, genetic instability and epigenetic dysregulation. Up-regulated HDAC (Histone Deacetylase) enzyme activity is associated with a closed chromatin assembly and subsequent gene repression, forming a characteristic feature of malignantly transformed cells. Novel therapeutics are now targeting the zinc containing HDAC enzymes for treating various types of cancers. Recently, a spate of drugs acting via HDAC inhibition have been undergoing clinical trials and several patents present exciting molecules like PCI-24781 (Abexinostat), ITF- 2357 (Givinostat); MS-275 (Entinostat), MGCD 0103 (Mocetinostat), LBH-589 (Panobinostat), FK228 (Romidepsin), PXD-101 (Belinostat) and Valproic Acid to be used as alternatives or adjuvants to traditional chemotherapeutics. However, only three HDAC inhibitors have acquired FDA approval till date. Recently, PXD-101 obtained FDA approval for the treatment of Refractory or Relapsed Peripheral T cell lymphoma. The current article reviews patents that have introduced novel molecules that are HDAC isoform specific, superior to first generation HDAC inhibitors like SAHA (Suberoylanilide Hydroxamic Acid) and TSA (Trichostatin A) and can be modified structurally to reduce toxic side effects and increase specificity. These molecules can combine the best characteristics of an ideal HDAC inhibiting drug either as monotherapy or in combinatorial therapy for cancer treatment thus, indicating promise to be included in the next generation of target specific HDAC inhibiting drugs.

  4. A novel gene delivery system targeting cells expressing VEGF receptors

    Institute of Scientific and Technical Information of China (English)

    LIJUNMIN; JINGCHULUO; 等

    1999-01-01

    Two ligand oligopeptides GV1 and GV2 were designed according to the putative binding region of VEGF to its receptors.GV1,GV2 and endosome releasing oligopeptide HA20 were conjugated with poly-L-lysine or protamine and the resulting conjugates could interact with DNA in a noncovalent bond to form a complex.Using pSV2-β-galactosidase as a reporter gene,it has been demonstrated that exogenous gene was transferred into bovine aortic arch-derived endothelial cells (ABAE) and human malignant melanoma cell lines (A375) in vitro.In vivo experiments,exogenous gene was transferred into tumor vascular endothelial cells and tumor cells of subcutaneously transplanted human colon cancer LOVO,human malignant melanoma A375 and human hepatoma graft in nude mice.This system could also target gene to intrahepatically transplanted human hepatoma injected via portal vein in nude mice.These results are correlated with the relevant receptors(flt-1,flk-1/KDR) expression on the targeted cells and tissues.

  5. Liver-targeted gene therapy: Approaches and challenges.

    Science.gov (United States)

    Aravalli, Rajagopal N; Belcher, John D; Steer, Clifford J

    2015-06-01

    The liver plays a major role in many inherited and acquired genetic disorders. It is also the site for the treatment of certain inborn errors of metabolism that do not directly cause injury to the liver. The advancement of nucleic acid-based therapies for liver maladies has been severely limited because of the myriad untoward side effects and methodological limitations. To address these issues, research efforts in recent years have been intensified toward the development of targeted gene approaches using novel genetic tools, such as zinc-finger nucleases, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats as well as various nonviral vectors such as Sleeping Beauty transposons, PiggyBac transposons, and PhiC31 integrase. Although each of these methods uses a distinct mechanism of gene modification, all of them are dependent on the efficient delivery of DNA and RNA molecules into the cell. This review provides an overview of current and emerging therapeutic strategies for liver-targeted gene therapy and gene repair.

  6. Apoptosis as a target for gene therapy in rheumatoid arthritis

    Directory of Open Access Journals (Sweden)

    Gabriel Adrián Rabinovich

    2000-01-01

    Full Text Available Rheumatoid arthritis (RA is characterized by chronic inflammation of the synovial joints resulting from hyperplasia of synovial fibroblasts and infiltration of lymphocytes, macrophages and plasma cells, all of which manifest signs of activation. All these cells proliferate abnormally, invade bone and cartilage, produce an elevated amount of pro-inflammatory cytokines, metalloproteinases and trigger osteoclast formation and activation. Some of the pathophysiological consequences of the disease may be explained by the inadequate apoptosis, which may promote the survival of autoreactive T cells, macrophages or synovial fibroblasts. Although RA does not result from single genetic mutations, elucidation of the molecular mechanisms implicated in joint destruction has revealed novel targets for gene therapy. Gene transfer strategies include inhibition of pro-inflammatory cytokines, blockade of cartilage-degrading metalloproteinases, inhibition of synovial cell activation and manipulation of the Th1-Th2 cytokine balance. Recent findings have iluminated the idea that induction of apoptosis in the rheumatoid joint can be also used to gain therapeutic advantage in the disease. In the present review we will discuss different strategies used for gene transfer in RA and chronic inflammation. Particularly, we will highlight the importance of programmed cell death as a novel target for gene therapy using endogenous biological mediators, such as galectin-1, a beta-galactoside-binding protein that induces apoptosis of activated T cells and immature thymocytes.

  7. Pancreatic Cancer Gene Therapy: From Molecular Targets to Delivery Systems

    Directory of Open Access Journals (Sweden)

    Maria Victoria Maliandi

    2011-01-01

    Full Text Available The continuous identification of molecular changes deregulating critical pathways in pancreatic tumor cells provides us with a large number of novel candidates to engineer gene-targeted approaches for pancreatic cancer treatment. Targets—both protein coding and non-coding—are being exploited in gene therapy to influence the deregulated pathways to facilitate cytotoxicity, enhance the immune response or sensitize to current treatments. Delivery vehicles based on viral or non-viral systems as well as cellular vectors with tumor homing characteristics are a critical part of the design of gene therapy strategies. The different behavior of tumoral versus non-tumoral cells inspires vector engineering with the generation of tumor selective products that can prevent potential toxic-associated effects. In the current review, a detailed analysis of the different targets, the delivery vectors, the preclinical approaches and a descriptive update on the conducted clinical trials are presented. Moreover, future possibilities in pancreatic cancer treatment by gene therapy strategies are discussed.

  8. Specifically targeted gene therapy for small-cell lung cancer

    DEFF Research Database (Denmark)

    Christensen, C.L.; Zandi, R.; Gjetting, T.

    2009-01-01

    Small-cell lung cancer (SCLC) is a highly malignant disease with poor prognosis. Hence, there is great demand for new therapies that can replace or supplement the current available treatment regimes. Gene therapy constitutes a promising strategy and relies on the principle of introducing exogenous....... This review describes and discusses the current status of the application of gene therapy in relation to SCLC Udgivelsesdato: 2009/4...... DNA into malignant cells causing them to die. Since SCLC is a highly disseminated malignancy, the gene therapeutic agent must be administered systemically, obligating a high level of targeting of tumor tissue and the use of delivery vehicles designed for systemic circulation of the therapeutic DNA...

  9. Assessment of antimicrobial (host defense) peptides as anti-cancer agents.

    Science.gov (United States)

    Douglas, Susan; Hoskin, David W; Hilchie, Ashley L

    2014-01-01

    Cationic antimicrobial (host defense) peptides (CAPs) are able to kill microorganisms and cancer cells, leading to their consideration as novel candidate therapeutic agents in human medicine. CAPs can physically associate with anionic membrane structures, such as those found on cancer cells, causing pore formation, intracellular disturbances, and leakage of cell contents. In contrast, normal cells are less negatively-charged and are typically not susceptible to CAP-mediated cell death. Because the interaction of CAPs with cells is based on charge properties rather than cell proliferation, both rapidly dividing and quiescent cancer cells, as well as multidrug-resistant cancer cells, are targeted by CAPs, making CAPS potentially valuable as anti-cancer agents. CAPs often exist as families of peptides with slightly different amino acid sequences. In addition, libraries of synthetic peptide variants based on naturally occurring CAP templates can be generated in order to improve upon their action. High-throughput screens are needed to quickly and efficiently assess the suitability of each CAP variant. Here we present the methods for assessing CAP-mediated cytotoxicity against cancer cells (suspension and adherent) and untransformed cells (measured using the tritiated thymidine-release or MTT assay), and for discriminating between cell death caused by necrosis (measured using lactate dehydrogenase- or (51)Cr-release assays), or apoptosis and necrosis (single-stranded DNA content measured by flow cytometry). In addition the clonogenic assay, which assesses the ability of single transformed cells to multiply and produce colonies, is described.

  10. Fucoxanthin: A Marine Carotenoid Exerting Anti-Cancer Effects by Affecting Multiple Mechanisms

    Directory of Open Access Journals (Sweden)

    Sangeetha Ravi Kumar

    2013-12-01

    Full Text Available Fucoxanthin is a marine carotenoid exhibiting several health benefits. The anti-cancer effect of fucoxanthin and its deacetylated metabolite, fucoxanthinol, is well documented. In view of its potent anti-carcinogenic activity, the need to understand the underlying mechanisms has gained prominence. Towards achieving this goal, several researchers have carried out studies in various cell lines and in vivo and have deciphered that fucoxanthin exerts its anti-proliferative and cancer preventing influence via different molecules and pathways including the Bcl-2 proteins, MAPK, NFκB, Caspases, GADD45, and several other molecules that are involved in either cell cycle arrest, apoptosis, or metastasis. Thus, in addition to decreasing the frequency of occurrence and growth of tumours, fucoxanthin has a cytotoxic effect on cancer cells. Some studies show that this effect is selective, i.e., fucoxanthin has the capability to target cancer cells only, leaving normal physiological cells unaffected/less affected. Hence, fucoxanthin and its metabolites show great promise as chemotherapeutic agents in cancer.

  11. Current advances in mathematical modeling of anti-cancer drug penetration into tumor tissues

    Directory of Open Access Journals (Sweden)

    MunJu eKim

    2013-11-01

    Full Text Available Delivery of anti-cancer drugs to tumor tissues, including their interstitial transport and cellular uptake, is a complex process involving various biochemical, mechanical, and biophysical factors. Mathematical modeling provides a means through which to understand this complexity better, as well as to examine interactions between contributing components in a systematic way via computational simulations and quantitative analyses. In this review, we present the current state of mathematical modeling approaches that address phenomena related to drug delivery. We describe how various types of models were used to predict spatio-temporal distributions of drugs within the tumor tissue, to simulate different ways to overcome barriers to drug transport, or to optimize treatment schedules. Finally, we discuss how integration of mathematical modeling with experimental or clinical data can provide better tools to understand the drug delivery process, in particular to examine the specific tissue- or compound-related factors that limit drug penetration through tumors. Such tools will be important in designing new chemotherapy targets and optimal treatment strategies, as well as in developing non-invasive diagnosis to monitor treatment response and detect tumor recurrence.

  12. Advances in individual markers of interferon in anti-cancer therapy

    Institute of Scientific and Technical Information of China (English)

    Chi Pan; Chenjing Zhang; Jianjin Huang

    2013-01-01

    Interferon (IFN) is a cytokine with various biological functions, including antivirus, immunoregulation and anti-tumor. It has been wildly used in many anti-cancer therapies, including malignant melanoma, hepatocellular carcinoma, ad-vanced renal-cell carcinoma, non-Hodgkin's lymphoma, chronic myelogenous leukemia and AIDS-related Kaposi's sarcoma. However, its effective dose is always very high, which may bring some serious side effects, nevertheless, not all patients can benefit from the IFN therapy. So a problem we have faced is that how to improve the efficiency and sensitivity of IFN? To solve this problem, many studies have been launched to find the effective prognostic factors and individual biomarkers for guiding the treatment better. In addition, further clarifying the anti-tumor mechanisms of IFN is benefit for explaining how the biomark-ers predict prognosis of patients. In recent studies, many IFN associated genes and proteins predicting sensitivity of IFN therapy have been found, which may associate with the progression of cancer, such as IFN regulatory factor (IRF), IFNAR2 mRNA, microRNA, IFITM-1. Some factors in peripheral blood are easier to detect and have the potential to been popularized in clinical practice, such as CD8high CD57+ lymphocyte levels in malignant melanoma, serum IFNAR2 mRNA in mCRC. This review briefly summarized the advances of antitumorally individual markers of IFN.

  13. 4',6-Dihydroxy-4-methoxyisoaurone inhibits TNF-α-induced NF-κB activation and expressions of NF-κB-regulated target gene products.

    Science.gov (United States)

    Ma, Juan; Mi, Chunliu; Wang, Ke Si; Lee, Jung Joon; Jin, Xuejun

    2016-02-01

    The nuclear factor-κB (NF-κB) transcription factors control many physiological processes including inflammation, apoptosis, and angiogenesis. In our search for NF-κB inhibitors from natural resources, we identified 4',6-dihydroxy-4-methoxyisoaurone (ISOA) as an inhibitor of NF-κB activation from the seeds of Trichosanthes kirilowii. However, the mechanism by which ISOA inhibits NF-κB activation is not fully understood. In the present study, we demonstrated the effect of ISOA on NF-κB activation in TNF-α-stimulated HeLa cells. This compound suppressed NF-κB activation through the inhibition of IκB kinase (IKK) activation. ISOA also has an influence on upstream signaling of IKK through the inhibition of expression of adaptor proteins, TNF receptor-associated factor 2 (TRAF2) and receptor interacting protein 1 (RIP1). Consequently, ISOA blocked the phosphorylation and degradation of the inhibitor of NF-κB alpha (IκBα), and subsequent phosphorylation and nuclear translocation of p65. The suppression of NF-κB activation by ISOA led to the down-regulation of target genes involved in inflammation, proliferation, as well as potentiation of TNF-α-induced apoptosis. Taken together, this study extends our understanding on the mechanisms underlying the anti-inflammatory and anti-cancer activities of ISOA. Our findings provide new insight into the molecular mechanisms and a potential application of ISOA for inflammatory diseases as well as certain cancers.

  14. Construction of RNAi lentiviral vector targeting mouse Islet-1 gene

    Directory of Open Access Journals (Sweden)

    Shen-shen ZHI

    2011-02-01

    Full Text Available Objective To construct and select RNAi lentiviral vectors that can silence mouse Islet-1 gene effectively.Methods Three groups of RNAi-target of mouse Islet-1 gene were designed,and corresponding shRNA oligo(sh1,sh2 and sh3 were synthesized,and then they were respectively inserted to the PLVTHM vector that had been digested by endonuclease.Agarose gel electrophoresis and sequencing were used to select and indentify the positive clones.The positive clones were extracted and then mixed with E.coli to amplify positive clones.The amplified clones were then infected into 293T along with the other 3 helper plasmids to produce lentiviral vector.After the construction of the lentiviral vector,plaque formation test was performed to determine the titer of lentiviral vector.The lentiviral vectors were then infected into C3H10T1/2 cells.The transfect efficiency of the lentiviral vectors was determined with flow cytometry with detection of green fluorescent protein(GFP.Q-PCR was employed to detect the RNAi efficiency of the lentiviral vectors.Results Agarose gel electrophoresis analysis showed that the clones with right gene at the target size were successfully established;gene sequencing showed that the right DNA fragments had been inserted;plaque formation test showed that the titer of the virus solution was 3.87×108TU/ml;the transfect efficiency of the lentiviral vector infected into C3H10T1/2 cells was 90.36%.All the 3 groups of shRNA targets(sh1,sh2 and sh3 showed an inhibitory effect on Islet-1 gene,and the sh1 showed the highest inhibitory effect(76.8%,as compared with that of normal cells(P < 0.05.Conclusion The RNAi lentiviral vector that can effectively silence the mouse Islet-1 gene has been constructed successfully,which may lay a foundation for further investigation of Islet-1 gene.

  15. Sgs1 and Exo1 suppress targeted chromosome duplication during ends-in and ends-out gene targeting.

    Science.gov (United States)

    Štafa, Anamarija; Miklenić, Marina; Zunar, Bojan; Lisnić, Berislav; Symington, Lorraine S; Svetec, Ivan-Krešimir

    2014-10-01

    Gene targeting is extremely efficient in the yeast Saccharomyces cerevisiae. It is performed by transformation with a linear, non-replicative DNA fragment carrying a selectable marker and containing ends homologous to the particular locus in a genome. However, even in S. cerevisiae, transformation can result in unwanted (aberrant) integration events, the frequency and spectra of which are quite different for ends-out and ends-in transformation assays. It has been observed that gene replacement (ends-out gene targeting) can result in illegitimate integration, integration of the transforming DNA fragment next to the target sequence and duplication of a targeted chromosome. By contrast, plasmid integration (ends-in gene targeting) is often associated with multiple targeted integration events but illegitimate integration is extremely rare and a targeted chromosome duplication has not been reported. Here we systematically investigated the influence of design of the ends-out assay on the success of targeted genetic modification. We have determined transformation efficiency, fidelity of gene targeting and spectra of all aberrant events in several ends-out gene targeting assays designed to insert, delete or replace a particular sequence in the targeted region of the yeast genome. Furthermore, we have demonstrated for the first time that targeted chromosome duplications occur even during ends-in gene targeting. Most importantly, the whole chromosome duplication is POL32 dependent pointing to break-induced replication (BIR) as the underlying mechanism. Moreover, the occurrence of duplication of the targeted chromosome was strikingly increased in the exo1Δ sgs1Δ double mutant but not in the respective single mutants demonstrating that the Exo1 and Sgs1 proteins independently suppress whole chromosome duplication during gene targeting.

  16. Engineering zinc finger protein transcription factors to downregulate the epithelial glycoprotein-2 promoter as a novel anti-cancer treatment.

    Science.gov (United States)

    Gommans, Willemijn M; McLaughlin, Pamela M J; Lindhout, Beatrice I; Segal, David J; Wiegman, D J; Haisma, Hidde J; van der Zaal, Bert J; Rots, Marianne G

    2007-05-01

    Zinc finger protein transcription factors (ZFP-TFs) are emerging as powerful novel tools for the treatment of many different diseases. ZFPs are DNA-binding motifs and consist of modular zinc finger domains. Each domain can be engineered to recognize a specific DNA triplet, and stitching six domains together results in the recognition of a gene-specific sequence. Inhibition of gene expression can be achieved by fusing a repressor domain to these DNA-binding motifs. In this study, we engineered ZFP-TFs to downregulate the activity of the epithelial glycoprotein-2 (EGP-2) promoter. The protein EGP-2 is overexpressed in a wide variety of cancer types and EGP-2 downregulation has been shown to result in a decreased oncogenic potential of tumor cells. Therefore, downregulation of EGP-2 expression by ZFP-TFs provides a novel anti-cancer therapeutic. Using a straightforward strategy, we engineered a 3-ZFP that could bind a 9 bp sequence within the EGP-2 promoter. After the addition of a repressor domain, this 3-ZFP-TF could efficiently downregulate EGP-2 promoter activity by 60%. To demonstrate the flexibility of this technology, we coupled an activation domain to the engineered ZFP, resulting in a nearly 200% increase in EGP-2 promoter activity. To inhibit the endogenous EGP-2 promoter, we engineered 6-ZFP-TFs. Although none of the constructed ZFP-TFs could convincingly modulate the endogenous promoter, efficient and specific inhibition of the exogenous promoter was observed. Overall, ZFP-TFs are versatile bi-directional modulators of gene expression and downregulation of EGP-2 promoter activity using ZFP-TFs can ultimately result in a novel anti-cancer treatment.

  17. Insulators target active genes to transcription factories and polycomb-repressed genes to polycomb bodies.

    Directory of Open Access Journals (Sweden)

    Hua-Bing Li

    2013-04-01

    Full Text Available Polycomb bodies are foci of Polycomb proteins in which different Polycomb target genes are thought to co-localize in the nucleus, looping out from their chromosomal context. We have shown previously that insulators, not Polycomb response elements (PREs, mediate associations among Polycomb Group (PcG targets to form Polycomb bodies. Here we use live imaging and 3C interactions to show that transgenes containing PREs and endogenous PcG-regulated genes are targeted by insulator proteins to different nuclear structures depending on their state of activity. When two genes are repressed, they co-localize in Polycomb bodies. When both are active, they are targeted to transcription factories in a fashion dependent on Trithorax and enhancer specificity as well as the insulator protein CTCF. In the absence of CTCF, assembly of Polycomb bodies is essentially reduced to those representing genomic clusters of Polycomb target genes. The critical role of Trithorax suggests that stable association with a specialized transcription factory underlies the cellular memory of the active state.

  18. Anti-EGFR immunonanoparticles containing IL12 and salmosin genes for targeted cancer gene therapy.

    Science.gov (United States)

    Kim, Jung Seok; Kang, Seong Jae; Jeong, Hwa Yeon; Kim, Min Woo; Park, Sang Il; Lee, Yeon Kyung; Kim, Hong Sung; Kim, Keun Sik; Park, Yong Serk

    2016-09-01

    Tumor-directed gene delivery is of major interest in the field of cancer gene therapy. Varied functionalizations of non-viral vectors have been suggested to enhance tumor targetability. In the present study, we prepared two different types of anti-EGF receptor (EGFR) immunonanoparticles containing pDNA, neutrally charged liposomes and cationic lipoplexes, for tumor-directed transfection of cancer therapeutic genes. Even though both anti-EGFR immunonanoparticles had a high binding affinity to the EGFR-positive cancer cells, the anti-EGFR immunolipoplex formulation exhibited approximately 100-fold higher transfection to the target cells than anti-EGFR immunoliposomes. The lipoplex formulation also showed a higher transfection to SK-OV-3 tumor xenografts in mice. Thus, IL12 and/or salmosin genes were loaded in the anti-EGFR immunolipoplexes and intravenously administered to mice carrying SK-OV-3 tumors. Co-transfection of IL12 and salmosin genes using anti-EGFR immunolipoplexes significantly reduced tumor growth and pulmonary metastasis. Furthermore, combinatorial treatment with doxorubicin synergistically inhibited tumor growth. These results suggest that anti-EGFR immunolipoplexes containing pDNA encoding therapeutic genes could be utilized as a gene-transfer modality for cancer gene therapy.

  19. Specific genetic modifications of domestic animals by gene targeting and animal cloning.

    Science.gov (United States)

    Wang, Bin; Zhou, Jiangfeng

    2003-11-13

    The technology of gene targeting through homologous recombination has been extremely useful for elucidating gene functions in mice. The application of this technology was thought impossible in the large livestock species until the successful creation of the first mammalian clone "Dolly" the sheep. The combination of the technologies for gene targeting of somatic cells with those of animal cloning made it possible to introduce specific genetic mutations into domestic animals. In this review, the principles of gene targeting in somatic cells and the challenges of nuclear transfer using gene-targeted cells are discussed. The relevance of gene targeting in domestic animals for applications in bio-medicine and agriculture are also examined.

  20. Production of cloned pigs with targeted attenuation of gene expression.

    Directory of Open Access Journals (Sweden)

    Vilceu Bordignon

    Full Text Available The objective of this study was to demonstrate that RNA interference (RNAi and somatic cell nuclear transfer (SCNT technologies can be used to attenuate the expression of specific genes in tissues of swine, a large animal species. Apolipoprotein E (apoE, a secreted glycoprotein known for its major role in lipid and lipoprotein metabolism and transport, was selected as the target gene for this study. Three synthetic small interfering RNAs (siRNA targeting the porcine apoE mRNA were tested in porcine granulosa cells in primary culture and reduced apoE mRNA abundance ranging from 45-82% compared to control cells. The most effective sequence was selected for cloning into a short hairpin RNA (shRNA expression vector under the control of RNA polymerase III (U6 promoter. Stably transfected fetal porcine fibroblast cells were generated and used to produce embryos with in vitro matured porcine oocytes, which were then transferred into the uterus of surrogate gilts. Seven live and one stillborn piglet were born from three gilts that became pregnant. Integration of the shRNA expression vector into the genome of clone piglets was confirmed by PCR and expression of the GFP transgene linked to the expression vector. Analysis showed that apoE protein levels in the liver and plasma of the clone pigs bearing the shRNA expression vector targeting the apoE mRNA was significantly reduced compared to control pigs cloned from non-transfected fibroblasts of the same cell line. These results demonstrate the feasibility of applying RNAi and SCNT technologies for introducing stable genetic modifications in somatic cells for eventual attenuation of gene expression in vivo in large animal species.

  1. Targeted Gene Capture by Hybridization to Illuminate Ecosystem Functioning.

    Science.gov (United States)

    Ribière, Céline; Beugnot, Réjane; Parisot, Nicolas; Gasc, Cyrielle; Defois, Clémence; Denonfoux, Jérémie; Boucher, Delphine; Peyretaillade, Eric; Peyret, Pierre

    2016-01-01

    Microbial communities are extremely abundant and diverse on earth surface and play key role in the ecosystem functioning. Thus, although next-generation sequencing (NGS) technologies have greatly improved knowledge on microbial diversity, it is necessary to reduce the biological complexity to better understand the microorganism functions. To achieve this goal, we describe a promising approach, based on the solution hybrid selection (SHS) method for the selective enrichment in a target-specific biomarker from metagenomic and metatranscriptomic samples. The success of this method strongly depends on the determination of sensitive, specific, and explorative probes to assess the complete targeted gene repertoire. Indeed, in this method, RNA probes were used to capture large DNA or RNA fragments harboring biomarkers of interest that potentially allow to link structure and function of communities of interest.

  2. Identification of targetable FGFR gene fusions in diverse cancers.

    Science.gov (United States)

    Wu, Yi-Mi; Su, Fengyun; Kalyana-Sundaram, Shanker; Khazanov, Nickolay; Ateeq, Bushra; Cao, Xuhong; Lonigro, Robert J; Vats, Pankaj; Wang, Rui; Lin, Su-Fang; Cheng, Ann-Joy; Kunju, Lakshmi P; Siddiqui, Javed; Tomlins, Scott A; Wyngaard, Peter; Sadis, Seth; Roychowdhury, Sameek; Hussain, Maha H; Feng, Felix Y; Zalupski, Mark M; Talpaz, Moshe; Pienta, Kenneth J; Rhodes, Daniel R; Robinson, Dan R; Chinnaiyan, Arul M

    2013-06-01

    Through a prospective clinical sequencing program for advanced cancers, four index cases were identified which harbor gene rearrangements of FGFR2, including patients with cholangiocarcinoma, breast cancer, and prostate cancer. After extending our assessment of FGFR rearrangements across multiple tumor cohorts, we identified additional FGFR fusions with intact kinase domains in lung squamous cell cancer, bladder cancer, thyroid cancer, oral cancer, glioblastoma, and head and neck squamous cell cancer. All FGFR fusion partners tested exhibit oligomerization capability, suggesting a shared mode of kinase activation. Overexpression of FGFR fusion proteins induced cell proliferation. Two bladder cancer cell lines that harbor FGFR3 fusion proteins exhibited enhanced susceptibility to pharmacologic inhibition in vitro and in vivo. Because of the combinatorial possibilities of FGFR family fusion to a variety of oligomerization partners, clinical sequencing efforts, which incorporate transcriptome analysis for gene fusions, are poised to identify rare, targetable FGFR fusions across diverse cancer types.

  3. Research progress of gene target therapy for refractory epilepsy

    Directory of Open Access Journals (Sweden)

    Xing-hua TANG

    2014-12-01

    Full Text Available Nowadays, the strategies of gene therapy for the treatment of refractory epilepsy (RE mainly include modulating neurotransmitter systems, neuropeptide Y (NPY and neurotrophic factors. Among them, the hot target spots include γ-aminobutyric acid (GABA and its receptor, N-methyl-D-aspartate (NMDA and its receptor, galanin, NPY and neurotrophic factors. This paper reviews the chief research results, and advantages and disadvantages of studies, and provides evidence for the treatment of refractory epilepsy. doi: 10.3969/j.issn.1672-6731.2014.12.004

  4. Modularly assembled designer TAL effector nucleases for targeted gene knockout and gene replacement in eukaryotes.

    Science.gov (United States)

    Li, Ting; Huang, Sheng; Zhao, Xuefeng; Wright, David A; Carpenter, Susan; Spalding, Martin H; Weeks, Donald P; Yang, Bing

    2011-08-01

    Recent studies indicate that the DNA recognition domain of transcription activator-like (TAL) effectors can be combined with the nuclease domain of FokI restriction enzyme to produce TAL effector nucleases (TALENs) that, in pairs, bind adjacent DNA target sites and produce double-strand breaks between the target sequences, stimulating non-homologous end-joining and homologous recombination. Here, we exploit the four prevalent TAL repeats and their DNA recognition cipher to develop a 'modular assembly' method for rapid production of designer TALENs (dTALENs) that recognize unique DNA sequence up to 23 bases in any gene. We have used this approach to engineer 10 dTALENs to target specific loci in native yeast chromosomal genes. All dTALENs produced high rates of site-specific gene disruptions and created strains with expected mutant phenotypes. Moreover, dTALENs stimulated high rates (up to 34%) of gene replacement by homologous recombination. Finally, dTALENs caused no detectable cytotoxicity and minimal levels of undesired genetic mutations in the treated yeast strains. These studies expand the realm of verified TALEN activity from cultured human cells to an intact eukaryotic organism and suggest that low-cost, highly dependable dTALENs can assume a significant role for gene modifications of value in human and animal health, agriculture and industry.

  5. Modularly assembled designer TAL effector nucleases for targeted gene knockout and gene replacement in eukaryotes

    Energy Technology Data Exchange (ETDEWEB)

    Li, T; Huang, S; Zhao, XF; Wright, DA; Carpenter, S; Spalding, MH; Weeks, DP; Yang, B

    2011-08-08

    Recent studies indicate that the DNA recognition domain of transcription activator-like (TAL) effectors can be combined with the nuclease domain of FokI restriction enzyme to produce TAL effector nucleases (TALENs) that, in pairs, bind adjacent DNA target sites and produce double-strand breaks between the target sequences, stimulating non-homologous end-joining and homologous recombination. Here, we exploit the four prevalent TAL repeats and their DNA recognition cipher to develop a 'modular assembly' method for rapid production of designer TALENs (dTALENs) that recognize unique DNA sequence up to 23 bases in any gene. We have used this approach to engineer 10 dTALENs to target specific loci in native yeast chromosomal genes. All dTALENs produced high rates of site-specific gene disruptions and created strains with expected mutant phenotypes. Moreover, dTALENs stimulated high rates (up to 34%) of gene replacement by homologous recombination. Finally, dTALENs caused no detectable cytotoxicity and minimal levels of undesired genetic mutations in the treated yeast strains. These studies expand the realm of verified TALEN activity from cultured human cells to an intact eukaryotic organism and suggest that low-cost, highly dependable dTALENs can assume a significant role for gene modifications of value in human and animal health, agriculture and industry.

  6. Diverging Novobiocin Anti-Cancer Activity from Neuroprotective Activity through Modification of the Amide Tail.

    Science.gov (United States)

    Ghosh, Suman; Liu, Yang; Garg, Gaurav; Anyika, Mercy; McPherson, Nolan T; Ma, Jiacheng; Dobrowsky, Rick T; Blagg, Brian S J

    2016-08-11

    Novobiocin is a natural product that binds the Hsp90 C-terminus and manifests Hsp90 inhibitory activity. Structural investigations on novobiocin led to the development of both anti-cancer and neuroprotective agents. The varied pharmacological activity manifested by these novobiocin analogs prompted the investigation of structure-function studies to identify these contradictory effects, which revealed that modifications to the amide side chain produce either anti-cancer or neuroprotective activity. Compounds that exhibit neuroprotective activity contain a short alkyl or cycloalkyl amide side chain. In contrast, anti-cancer agents contain five or more carbons, disrupt interactions between Hsp90α and Aha1, and induce the degradation of Hsp90-dependent client proteins.

  7. [Targeted modification of CCR5 gene in rabbits by TALEN].

    Science.gov (United States)

    Tang, Chengcheng; Zhang, Quanjun; Li, Xiaoping; Fan, Nana; Yang, Yi; Quan, Longquan; Lai, Liangxue

    2014-04-01

    The lack of suitable animal model for HIV-1 infection has become a bottleneck for the development of AIDS vaccines and drugs. Wild-type rabbits can be infected by HIV-1 persistently and HIV-1 can be efficiently replicated resulting in syncytia in rabbit cell line co-expressing human CD4 and CCR5.Therefore, a rabbit highly expressing human CD4 and CCR5 may be an ideal animal model for AIDS disease study. In the present report, by using the efficient gene targeting technology, transcription activator-like effector nuclease (TALEN), we explored the feasibility of generating a HIV-1 model by knocking in human CD4 and CCR5 into rabbit genome. First we constructed two TALEN vectors targeting rabbit CCR5 gene and a vector with homologous arms. TALEN mRNAs and donor DNA were then co-injected into fertilized oocytes. After 3?5 days, 24 embryos were collected and used to conduct mutation analysis with PCR and sequencing. All the 24 embryos were detected with CCR5 knockouts and 5 were human CD4 and CCR5 knockins. Our results laid a foundation for establishing a new animal model for the study of AIDS.

  8. Application of next generation sequencing to human gene fusion detection: computational tools, features and perspectives.

    Science.gov (United States)

    Wang, Qingguo; Xia, Junfeng; Jia, Peilin; Pao, William; Zhao, Zhongming

    2013-07-01

    Gene fusions are important genomic events in human cancer because their fusion gene products can drive the development of cancer and thus are potential prognostic tools or therapeutic targets in anti-cancer treatment. Major advancements have been made in computational approaches for fusion gene discovery over the past 3 years due to improvements and widespread applications of high-throughput next generation sequencing (NGS) technologies. To identify fusions from NGS data, existing methods typically leverage the strengths of both sequencing technologies and computational strategies. In this article, we review the NGS and computational features of existing methods for fusion gene detection and suggest directions for future development.

  9. Long-term cultivation of colorectal carcinoma cells with anti-cancer drugs induces drug resistance and telomere elongation: an in vitro study

    Directory of Open Access Journals (Sweden)

    Mochizuki Hidetaka

    2001-08-01

    Full Text Available Abstract Background The role of telomerase activation in the expression and/or maintenance of drug resistance is not clearly understood. Therefore, we investigated the relationships, among the telomerase activity, telomere length and the expression of multidrug resistance genes in colorectal cancer cell lines cultivated with anti-cancer drugs. Methods LoVo and DLD-1 cells were continuously grown in the presence of both CDDP and 5-FU for up to 100 days. Cell proliferation, telomerase activity, telomere length and the expression of multidrug resistance genes were serially monitored as the PDL increased. Results The expression of multidrug resistance genes tended to increase as the PDL increased. However, an abnormal aneuploid clone was not detected as far as the cells were monitored by a DNA histogram analysis. Tumor cells showing resistance to anti-cancer drugs revealed a higher cell proliferation rate. The telomere length gradually increased with a progressive PDL. The telomerase activity reached a maximum level at 15 PDL in LoVo cells and at 27 PDL in DLD-1 cells. An increase in the mRNA expression of the telomerase components, especially in hTERT and in hTR, was observed at the same PDLs. Conclusions These results suggest that a high telomerase activity and an elongation of telomeres both appear to help maintain and/or increase drug resistance in colorectal cancer cells. Cancer cells with long telomeres and a high proliferative activity may thus be able to better survive exposure to anti-cancer drugs. This is presumably due to an increased chromosome stability and a strong expression of both mdr-1 and MRP genes.

  10. Multifunctional liposomes for enhanced anti-cancer therapy

    Science.gov (United States)

    Falcao, Claudio Borges

    2011-12-01

    with half of the concentration needed for G3139 alone in CL to reduce the cell viability by 40%. Also, it was found greater apoptotic signal in cells treated with CLs containing D-(KLAKLAK)2/G3139 complexes than CLs with G3139 only. In vivo, D-(KLAKLAK) 2/G3139 complexes in CL significantly inhibited tumor growth compared to the saline treated group, through apoptosis induction. However, the mechanism involved in cell death by apoptosis seems to be independent of reduction of bcl-2 protein levels. PEG2000 at 1% mol could significantly reduce activity of PCL formulation towards B16(F10) cells compared to CLs. After pre-incubation at pH 7.4, PCL and pH-PCL had decreased activity compared to CL towards B16(F10) cells. After pre-incubation at pH 5.0, while CL and PCL had the same activity with the cells as in neutral pH, pH-PCL formulation had its PEG cleaved and its cytotoxicity was restored against the melanoma cells. Thus, D-(KLAKLAK)2/G3139 complexes in CL had enhanced anti-cancer therapy, through apoptosis, than G3139 alone in CL in vitro and in vivo. In vitro, PCL and pH-PCL particles obtained can have a prolonged blood residence time, and, once a tumor tissue is reached, pH-PCL can have its cytotoxicity restored because of hydrolysis of cleavable PEG at a lowered pH.

  11. Colorimetric biosensing of targeted gene sequence using dual nanoparticle platforms

    Directory of Open Access Journals (Sweden)

    Thavanathan J

    2015-04-01

    Full Text Available Jeevan Thavanathan,1 Nay Ming Huang,1 Kwai Lin Thong2 1Low Dimension Material Research Center, Department of Physics, 2Institute of Biological Sciences, Faculty of Science, University of Malaya, Kuala Lumpur, Malaysia Abstract: We have developed a colorimetric biosensor using a dual platform of gold nanoparticles and graphene oxide sheets for the detection of Salmonella enterica. The presence of the invA gene in S. enterica causes a change in color of the biosensor from its original pinkish-red to a light purplish solution. This occurs through the aggregation of the primary gold nanoparticles–conjugated DNA probe onto the surface of the secondary graphene oxide–conjugated DNA probe through DNA hybridization with the targeted DNA sequence. Spectrophotometry analysis showed a shift in wavelength from 525 nm to 600 nm with 1 µM of DNA target. Specificity testing revealed that the biosensor was able to detect various serovars of the S. enterica while no color change was observed with the other bacterial species. Sensitivity testing revealed the limit of detection was at 1 nM of DNA target. This proves the effectiveness of the biosensor in the detection of S. enterica through DNA hybridization. Keywords: biosensor, DNA hybridization, DNA probe, gold nanoparticles, graphene oxide, Salmonella enterica

  12. AAC as a Potential Target Gene to Control Verticillium dahliae

    Directory of Open Access Journals (Sweden)

    Xiaofeng Su

    2017-01-01

    Full Text Available Verticillium dahliae invades the roots of host plants and causes vascular wilt, which seriously diminishes the yield of cotton and other important crops. The protein AAC (ADP, ATP carrier is responsible for transferring ATP from the mitochondria into the cytoplasm. When V. dahliae protoplasts were transformed with short interfering RNAs (siRNAs targeting the VdAAC gene, fungal growth and sporulation were significantly inhibited. To further confirm a role for VdAAC in fungal development, we generated knockout mutants (ΔVdACC. Compared with wild-type V. dahliae (Vd wt, ΔVdAAC was impaired in germination and virulence; these impairments were rescued in the complementary strains (ΔVdAAC-C. Moreover, when an RNAi construct of VdAAC under the control of the 35S promoter was used to transform Nicotiana benthamiana, the expression of VdAAC was downregulated in the transgenic seedlings, and they had elevated resistance against V. dahliae. The results of this study suggest that VdAAC contributes to fungal development, virulence and is a promising candidate gene to control V. dahliae. In addition, RNAi is a highly efficient way to silence fungal genes and provides a novel strategy to improve disease resistance in plants.

  13. Targeted Gene Therapy of Cancer: Second Amendment toward Holistic Therapy

    Directory of Open Access Journals (Sweden)

    Jaleh Barar

    2013-02-01

    Full Text Available It seems solid tumors are developing smart organs with specialized cells creating specified bio-territory, the so called “tumor microenvironment (TME”, in which there is reciprocal crosstalk among cancer cells, immune system cells and stromal cells. TME as an intricate milieu also consists of cancer stem cells (CSCs that can resist against chemotherapies. In solid tumors, metabolism and vascularization appears to be aberrant and tumor interstitial fluid (TIF functions as physiologic barrier. Thus, chemotherapy, immunotherapy and gene therapy often fail to provide cogent clinical outcomes. It looms that it is the time to accept the fact that initiation of cancer could be generation of another form of life that involves a cluster of thousands of genes, while we have failed to observe all aspects of it. Hence, the current treatment modalities need to be re-visited to cover all key aspects of disease using combination therapy based on the condition of patients. Perhaps personalized cluster of genes need to be simultaneously targeted.

  14. Targeted Gene Therapy of Cancer: Second Amendment toward Holistic Therapy.

    Science.gov (United States)

    Barar, Jaleh; Omidi, Yadollah

    2013-01-01

    It seems solid tumors are developing smart organs with specialized cells creating specified bio-territory, the so called "tumor microenvironment (TME)", in which there is reciprocal crosstalk among cancer cells, immune system cells and stromal cells. TME as an intricate milieu also consists of cancer stem cells (CSCs) that can resist against chemotherapies. In solid tumors, metabolism and vascularization appears to be aberrant and tumor interstitial fluid (TIF) functions as physiologic barrier. Thus, chemotherapy, immunotherapy and gene therapy often fail to provide cogent clinical outcomes. It looms that it is the time to accept the fact that initiation of cancer could be generation of another form of life that involves a cluster of thousands of genes, while we have failed to observe all aspects of it. Hence, the current treatment modalities need to be re-visited to cover all key aspects of disease using combination therapy based on the condition of patients. Perhaps personalized cluster of genes need to be simultaneously targeted.

  15. Systematic targeted integration to study Albumin gene control elements.

    Directory of Open Access Journals (Sweden)

    Sanchari Bhattacharyya

    Full Text Available To study transcriptional regulation by distant enhancers, we devised a system of easily modified reporter plasmids for integration into single-copy targeting cassettes in clones of HuH7, a human hepatocellular carcinoma. The plasmid constructs tested transcriptional function of a 35-kb region that contained the rat albumin gene and its upstream flanking region. Expression of integrants was analyzed in two orientations, and compared to transient expression of non-integrated plasmids. Enhancers were studied in their natural positions relative to the promoter and localized by deletion. All constructs were also analyzed by transient transfection assays. In addition to the known albumin gene enhancer (E1 at -10 kb, we demonstrated two new enhancers, E2 at -13, and E4 at +1.2 kb. All three enhancers functioned in both transient assays and integrated constructs. However, chromosomal integration demonstrated several differences from transient expression. For example, analysis of E2 showed that enhancer function within the chromosome required a larger gene region than in transient assays. Another conserved region, E3 at -0.7 kb, functioned as an enhancer in transient assays but inhibited the function of E1 and E2 when chromosomally integrated. The enhancers did not show additive or synergistic behavior,an effect consistent with competition for the promoter or inhibitory interactions among enhancers. Growth arrest by serum starvation strongly stimulated the function of some integrated enhancers, consistent with the expected disruption of enhancer-promoter looping during the cell cycle.

  16. Investigation of gene expression profiles in coronary heart disease and functional analysis of target gene

    Institute of Scientific and Technical Information of China (English)

    YIN HuiJun; MA Xiaoduan; JIANG YueRong; SHI DaZhuo; CHEN KeJi

    2009-01-01

    The research outlined here includes constitution of the differential gene expression profile by means of oligonucleotide gene microarray and functional analysis of the target gene for coronary heart disease (CHD). In a microarray screening experiment, the predominance of inflammation-and immune-related genes is presented in the expression profile of 107 differential genes based on the analysis of gene ontology and gene pathway. IL-8, an inflammatory factor, is identified as one of the genes that were markedly up-regulated in CHD. The plasma level of IL-8 is significantly raised in patients with CHD (n = 30) compared with healthy controls (n = 40), which underscores the clinical relevance of the in vitro finding. The further functional analysis shows that IL-8 affects platelet aggregation percentage, ex-pression of CD62p and platelet aggregation morphology in 12 healthy volunteers to some extent. These findings suggest the relevance of inflammation and immune responses to CHD at the DNA level. Moreover, IL-8 may be involved in the pathogenesis of CHD through the pathway of platelet activation.

  17. [Targeted Therapy and Immunotherapy for Non-small Cell Lung Cancer 
with Brain Metastasis].

    Science.gov (United States)

    Song, Qi; Jiao, Shunchang; Li, Fang

    2016-08-20

    Brain metastasis, a common complication of non-small cell lung cancer (NSCLC) with an incidence rate of 30%-50%, significantly affects the patients' quality of life. The prognosis of patients of NSCLC with brain metastasis is extremely poor, the average median survival is only 1 m-2 m without treatment. The targeted therapy based on lung cancer driven gene is a new treatment. Besides, the immunotherapy which can enhance the effect of anti-cancer by simulating the immune system is a new approach. The combination of targeted therapy and immunotherapy can greatly benefit patients in clinical work.

  18. Targeted Therapy and Immunotherapy for Non-small Cell Lung Cancer 
with Brain Metastasis

    Directory of Open Access Journals (Sweden)

    Qi SONG

    2016-08-01

    Full Text Available Brain metastasis, a common complication of non-small cell lung cancer (NSCLC with an incidence rate of 30%-50%, significantly affects the patients’ quality of life. The prognosis of patients of NSCLC with brain metastasis is extremely poor, the average median survival is only 1 m-2 m without treatment. The targeted therapy based on lung cancer driven gene is a new treatment. Besides, the immunotherapy which can enhance the effect of anti-cancer by simulating the immune system is a new approach. The combination of targeted therapy and immunotherapy can greatly benefit patients in clinical work.

  19. Transcription factors and target genes of pre-TCR signaling.

    Science.gov (United States)

    López-Rodríguez, Cristina; Aramburu, Jose; Berga-Bolaños, Rosa

    2015-06-01

    Almost 30 years ago pioneering work by the laboratories of Harald von Boehmer and Susumo Tonegawa provided the first indications that developing thymocytes could assemble a functional TCRβ chain-containing receptor complex, the pre-TCR, before TCRα expression. The discovery and study of the pre-TCR complex revealed paradigms of signaling pathways in control of cell survival and proliferation, and culminated in the recognition of the multifunctional nature of this receptor. As a receptor integrated in a dynamic developmental process, the pre-TCR must be viewed not only in the light of the biological outcomes it promotes, but also in context with those molecular processes that drive its expression in thymocytes. This review article focuses on transcription factors and target genes activated by the pre-TCR to drive its different outcomes.

  20. Il2rg gene-targeted severe combined immunodeficiency pigs.

    Science.gov (United States)

    Suzuki, Shunichi; Iwamoto, Masaki; Saito, Yoriko; Fuchimoto, Daiichiro; Sembon, Shoichiro; Suzuki, Misae; Mikawa, Satoshi; Hashimoto, Michiko; Aoki, Yuki; Najima, Yuho; Takagi, Shinsuke; Suzuki, Nahoko; Suzuki, Emi; Kubo, Masanori; Mimuro, Jun; Kashiwakura, Yuji; Madoiwa, Seiji; Sakata, Yoichi; Perry, Anthony C F; Ishikawa, Fumihiko; Onishi, Akira

    2012-06-14

    A porcine model of severe combined immunodeficiency (SCID) promises to facilitate human cancer studies, the humanization of tissue for xenotransplantation, and the evaluation of stem cells for clinical therapy, but SCID pigs have not been described. We report here the generation and preliminary evaluation of a porcine SCID model. Fibroblasts containing a targeted disruption of the X-linked interleukin-2 receptor gamma chain gene, Il2rg, were used as donors to generate cloned pigs by serial nuclear transfer. Germline transmission of the Il2rg deletion produced healthy Il2rg(+/-) females, while Il2rg(-/Y) males were athymic and exhibited markedly impaired immunoglobulin and T and NK cell production, robustly recapitulating human SCID. Following allogeneic bone marrow transplantation, donor cells stably integrated in Il2rg(-/Y) heterozygotes and reconstituted the Il2rg(-/Y) lymphoid lineage. The SCID pigs described here represent a step toward the comprehensive evaluation of preclinical cellular regenerative strategies.

  1. Rationale for stimulator of interferon genes-targeted cancer immunotherapy.

    Science.gov (United States)

    Rivera Vargas, Thaiz; Benoit-Lizon, Isis; Apetoh, Lionel

    2017-02-17

    The efficacy of checkpoint inhibitor therapy illustrates that cancer immunotherapy, which aims to foster the host immune response against cancer to achieve durable anticancer responses, can be successfully implemented in a routine clinical practice. However, a substantial proportion of patients does not benefit from this treatment, underscoring the need to identify alternative strategies to defeat cancer. Despite the demonstration in the 1990's that the detection of danger signals, including the nucleic acids DNA and RNA, by dendritic cells (DCs) in a cancer setting is essential for eliciting host defence, the molecular sensors responsible for recognising these danger signals and eliciting anticancer immune responses remain incompletely characterised, possibly explaining the disappointing results obtained so far upon the clinical implementation of DC-based cancer vaccines. In 2008, STING (stimulator of interferon genes), was identified as a protein that is indispensable for the recognition of cytosolic DNA. The central role of STING in controlling anticancer immune responses was exemplified by observations that spontaneous and radiation-induced adaptive anticancer immunity was reduced in the absence of STING, illustrating the potential of STING-targeting for cancer immunotherapy. Here, we will discuss the relevance of manipulating the STING signalling pathway for cancer treatment and integrating STING-targeting based strategies into combinatorial therapies to obtain long-lasting anticancer immune responses.

  2. Challenges and strategies in anti-cancer nanomedicine development: An industry perspective

    NARCIS (Netherlands)

    Hare, J.I.; Lammers, T.G.G.M.; Ashford, M.B.; Puri, S.; Storm, G.; Barry, S.T.

    2017-01-01

    Successfully translating anti-cancer nanomedicines from pre-clinical proof of concept to demonstration of therapeutic value in the clinic is challenging. Having made significant advances with drug delivery technologies, we must learn from other areas of oncology drug development, where patient strat

  3. Plant derived substances with anti-cancer activity: from folklore to practice.

    Science.gov (United States)

    Fridlender, Marcelo; Kapulnik, Yoram; Koltai, Hinanit

    2015-01-01

    Plants have had an essential role in the folklore of ancient cultures. In addition to the use as food and spices, plants have also been utilized as medicines for over 5000 years. It is estimated that 70-95% of the population in developing countries continues to use traditional medicines even today. A new trend, that involved the isolation of plant active compounds begun during the early nineteenth century. This trend led to the discovery of different active compounds that are derived from plants. In the last decades, more and more new materials derived from plants have been authorized and subscribed as medicines, including those with anti-cancer activity. Cancer is among the leading causes of morbidity and mortality worldwide. The number of new cases is expected to rise by about 70% over the next two decades. Thus, there is a real need for new efficient anti-cancer drugs with reduced side effects, and plants are a promising source for such entities. Here we focus on some plant-derived substances exhibiting anti-cancer and chemoprevention activity, their mode of action and bioavailability. These include paclitaxel, curcumin, and cannabinoids. In addition, development and use of their synthetic analogs, and those of strigolactones, are discussed. Also discussed are commercial considerations and future prospects for development of plant derived substances with anti-cancer activity.

  4. Anti-cancer efficacy of silybin derivatives -- a structure-activity relationship.

    Directory of Open Access Journals (Sweden)

    Chapla Agarwal

    Full Text Available Silybin or silibinin, a flavonolignan isolated from Milk thistle seeds, is one of the popular dietary supplements and has been extensively studied for its antioxidant, hepatoprotective and anti-cancer properties. We have envisioned that potency of silybin could be further enhanced through suitable modification/s in its chemical structure. Accordingly, here, we synthesized and characterized a series of silybin derivatives namely 2,3-dehydrosilybin (DHS, 7-O-methylsilybin (7OM, 7-O-galloylsilybin (7OG, 7,23-disulphatesilybin (DSS, 7-O-palmitoylsilybin (7OP, and 23-O-palmitoylsilybin (23OP; and compared their anti-cancer efficacy using human bladder cancer HTB9, colon cancer HCT116 and prostate carcinoma PC3 cells. In all the 3 cell lines, DHS, 7OM and 7OG demonstrated better growth inhibitory effects and compared to silybin, while other silybin derivatives showed lesser or no efficacy. Next, we prepared the optical isomers (A and B of silybin, DHS, 7OM and 7OG, and compared their anti-cancer efficacy. Isomers of these three silybin derivatives also showed better efficacy compared with respective silybin isomers, but in each, there was no clear cut silybin A versus B isomer activity preference. Further studies in HTB cells found that DHS, 7OM and 7OG exert better apoptotic activity than silibinin. Clonogenic assays in HTB9 cells further confirmed that both the racemic mixtures as well as pure optical isomers of DHS, 7OM and 7OG were more effective than silybin. Overall, these results clearly suggest that the anti-cancer efficacy of silybin could be significantly enhanced through structural modifications, and identify strong anti-cancer efficacy of silybin derivatives, namely DHS, 7OM, and 7OG, signifying that their efficacy and toxicity should be evaluated in relevant pre-clinical cancer models in rodents.

  5. Screening for Anti-Cancer Compounds in Marine Organisms in Oman

    Directory of Open Access Journals (Sweden)

    Sergey Dobretsov

    2016-05-01

    Full Text Available Objectives: Marine organisms are a rich source of bioactive molecules with potential applications in medicine, biotechnology and industry; however, few bioactive compounds have been isolated from organisms inhabiting the Arabian Gulf and the Gulf of Oman. This study aimed to isolate and screen the anti-cancer activity of compounds and extracts from 40 natural products of marine organisms collected from the Gulf of Oman. Methods: This study was carried out between January 2012 and December 2014 at the Sultan Qaboos University, Muscat, Oman. Fungi, bacteria, sponges, algae, soft corals, tunicates, bryozoans, mangrove tree samples and sea cucumbers were collected from seawater at Marina Bandar Al-Rowdha and Bandar Al-Khayran in Oman. Bacteria and fungi were isolated using a marine broth and organisms were extracted with methanol and ethyl acetate. Compounds were identified from spectroscopic data. The anti-cancer activity of the compounds and extracts was tested in a Michigan Cancer Foundation (MCF-7 cell line breast adenocarcinoma model. Results: Eight pure compounds and 32 extracts were investigated. Of these, 22.5% showed strong or medium anti-cancer activity, with malformin A, kuanoniamine D, hymenialdisine and gallic acid showing the greatest activity, as well as the soft coral Sarcophyton sp. extract. Treatment of MCF-7 cells at different concentrations of Sarcophyton sp. extracts indicated the induction of concentration-dependent cell death. Ultrastructural analysis highlighted the presence of nuclear fragmentation, membrane protrusion, blebbing and chromatic segregation at the nuclear membrane, which are typical characteristics of cell death by apoptosis induction. Conclusion: Some Omani marine organisms showed high anti-cancer potential. The efficacy, specificity and molecular mechanisms of anti-cancer compounds from Omani marine organisms on various cancer models should be investigated in future in vitro and in vivo studies.

  6. Evolutionary relationships of Aurora kinases: Implications for model organism studies and the development of anti-cancer drugs

    Directory of Open Access Journals (Sweden)

    Patrick Denis R

    2004-10-01

    Full Text Available Abstract Background As key regulators of mitotic chromosome segregation, the Aurora family of serine/threonine kinases play an important role in cell division. Abnormalities in Aurora kinases have been strongly linked with cancer, which has lead to the recent development of new classes of anti-cancer drugs that specifically target the ATP-binding domain of these kinases. From an evolutionary perspective, the species distribution of the Aurora kinase family is complex. Mammals uniquely have three Aurora kinases, Aurora-A, Aurora-B, and Aurora-C, while for other metazoans, including the frog, fruitfly and nematode, only Aurora-A and Aurora-B kinases are known. The fungi have a single Aurora-like homolog. Based on the tacit assumption of orthology to human counterparts, model organism studies have been central to the functional characterization of Aurora kinases. However, the ortholog and paralog relationships of these kinases across various species have not been rigorously examined. Here, we present comprehensive evolutionary analyses of the Aurora kinase family. Results Phylogenetic trees suggest that all three vertebrate Auroras evolved from a single urochordate ancestor. Specifically, Aurora-A is an orthologous lineage in cold-blooded vertebrates and mammals, while structurally similar Aurora-B and Aurora-C evolved more recently in mammals from a duplication of an ancestral Aurora-B/C gene found in cold-blooded vertebrates. All so-called Aurora-A and Aurora-B kinases of non-chordates are ancestral to the clade of chordate Auroras and, therefore, are not strictly orthologous to vertebrate counterparts. Comparisons of human Aurora-B and Aurora-C sequences to the resolved 3D structure of human Aurora-A lends further support to the evolutionary scenario that vertebrate Aurora-B and Aurora-C are closely related paralogs. Of the 26 residues lining the ATP-binding active site, only three were variant and all were specific to Aurora-A. Conclusions In

  7. Id-1 gene and gene products as therapeutic targets for treatment of breast cancer and other types of carcinoma

    Science.gov (United States)

    Desprez, Pierre-Yves; Campisi, Judith

    2014-08-19

    A method for treatment of breast cancer and other types of cancer. The method comprises targeting and modulating Id-1 gene expression, if any, for the Id-1 gene, or gene products in breast or other epithelial cancers in a patient by delivering products that modulate Id-1 gene expression. When expressed, Id-1 gene is a prognostic indicator that cancer cells are invasive and metastatic.

  8. Gene targeting, genome editing: from Dolly to editors.

    Science.gov (United States)

    Tan, Wenfang; Proudfoot, Chris; Lillico, Simon G; Whitelaw, C Bruce A

    2016-06-01

    One of the most powerful strategies to investigate biology we have as scientists, is the ability to transfer genetic material in a controlled and deliberate manner between organisms. When applied to livestock, applications worthy of commercial venture can be devised. Although initial methods used to generate transgenic livestock resulted in random transgene insertion, the development of SCNT technology enabled homologous recombination gene targeting strategies to be used in livestock. Much has been accomplished using this approach. However, now we have the ability to change a specific base in the genome without leaving any other DNA mark, with no need for a transgene. With the advent of the genome editors this is now possible and like other significant technological leaps, the result is an even greater diversity of possible applications. Indeed, in merely 5 years, these 'molecular scissors' have enabled the production of more than 300 differently edited pigs, cattle, sheep and goats. The advent of genome editors has brought genetic engineering of livestock to a position where industry, the public and politicians are all eager to see real use of genetically engineered livestock to address societal needs. Since the first transgenic livestock reported just over three decades ago the field of livestock biotechnology has come a long way-but the most exciting period is just starting.

  9.   Co-factors necessary for PPAR mediated transactivation of endogenous target genes

    DEFF Research Database (Denmark)

    Grøntved, Lars; Nielsen, Ronni; Stunnenberg, Henk

    of endogenous target gene in different cell types are elusive. To mutually compare the ability of the PPAR subtypes to activate endogenous target genes in a given cell, PPARa, PPARb/d and PPARg2 were HA tagged and rapidly, equally and synchronously expressed using adenoviral delivery. Within a few hours after...... adenoviral delivery the PPARs establish transcriptional active complexes on genomic target loci and launch immediate activation even of silent target genes. Direct comparison of the PPAR subtypes in a given cell line reveals that they selectively occupy genomic target promoters and in correlation show...

  10. Moringa oleifera as an Anti-Cancer Agent against Breast and Colorectal Cancer Cell Lines.

    Directory of Open Access Journals (Sweden)

    Abdulrahman Khazim Al-Asmari

    Full Text Available In this study we investigated the anti-cancer effect of Moringa oleifera leaves, bark and seed extracts. When tested against MDA-MB-231 and HCT-8 cancer cell lines, the extracts of leaves and bark showed remarkable anti-cancer properties while surprisingly, seed extracts exhibited hardly any such properties. Cell survival was significantly low in both cells lines when treated with leaves and bark extracts. Furthermore, a striking reduction (about 70-90% in colony formation as well as cell motility was observed upon treatment with leaves and bark. Additionally, apoptosis assay performed on these treated breast and colorectal cancer lines showed a remarkable increase in the number of apoptotic cells; with a 7 fold increase in MD-MB-231 to an increase of several fold in colorectal cancer cell lines. However, no significant apoptotic cells were detected upon seeds extract treatment. Moreover, the cell cycle distribution showed a G2/M enrichment (about 2-3 fold indicating that these extracts effectively arrest the cell progression at the G2/M phase. The GC-MS analyses of these extracts revealed numerous known anti-cancer compounds, namely eugenol, isopropyl isothiocynate, D-allose, and hexadeconoic acid ethyl ester, all of which possess long chain hydrocarbons, sugar moiety and an aromatic ring. This suggests that the anti-cancer properties of Moringa oleifera could be attributed to the bioactive compounds present in the extracts from this plant. This is a novel study because no report has yet been cited on the effectiveness of Moringa extracts obtained in the locally grown environment as an anti-cancer agent against breast and colorectal cancers. Our study is the first of its kind to evaluate the anti-malignant properties of Moringa not only in leaves but also in bark. These findings suggest that both the leaf and bark extracts of Moringa collected from the Saudi Arabian region possess anti-cancer activity that can be used to develop new drugs for

  11. Moringa oleifera as an Anti-Cancer Agent against Breast and Colorectal Cancer Cell Lines.

    Science.gov (United States)

    Al-Asmari, Abdulrahman Khazim; Albalawi, Sulaiman Mansour; Athar, Md Tanwir; Khan, Abdul Quaiyoom; Al-Shahrani, Hamoud; Islam, Mozaffarul

    2015-01-01

    In this study we investigated the anti-cancer effect of Moringa oleifera leaves, bark and seed extracts. When tested against MDA-MB-231 and HCT-8 cancer cell lines, the extracts of leaves and bark showed remarkable anti-cancer properties while surprisingly, seed extracts exhibited hardly any such properties. Cell survival was significantly low in both cells lines when treated with leaves and bark extracts. Furthermore, a striking reduction (about 70-90%) in colony formation as well as cell motility was observed upon treatment with leaves and bark. Additionally, apoptosis assay performed on these treated breast and colorectal cancer lines showed a remarkable increase in the number of apoptotic cells; with a 7 fold increase in MD-MB-231 to an increase of several fold in colorectal cancer cell lines. However, no significant apoptotic cells were detected upon seeds extract treatment. Moreover, the cell cycle distribution showed a G2/M enrichment (about 2-3 fold) indicating that these extracts effectively arrest the cell progression at the G2/M phase. The GC-MS analyses of these extracts revealed numerous known anti-cancer compounds, namely eugenol, isopropyl isothiocynate, D-allose, and hexadeconoic acid ethyl ester, all of which possess long chain hydrocarbons, sugar moiety and an aromatic ring. This suggests that the anti-cancer properties of Moringa oleifera could be attributed to the bioactive compounds present in the extracts from this plant. This is a novel study because no report has yet been cited on the effectiveness of Moringa extracts obtained in the locally grown environment as an anti-cancer agent against breast and colorectal cancers. Our study is the first of its kind to evaluate the anti-malignant properties of Moringa not only in leaves but also in bark. These findings suggest that both the leaf and bark extracts of Moringa collected from the Saudi Arabian region possess anti-cancer activity that can be used to develop new drugs for treatment of breast

  12. Specific genetic modifications of domestic animals by gene targeting and animal cloning

    Directory of Open Access Journals (Sweden)

    Zhou Jiangfeng

    2003-11-01

    Full Text Available Abstract The technology of gene targeting through homologous recombination has been extremely useful for elucidating gene functions in mice. The application of this technology was thought impossible in the large livestock species until the successful creation of the first mammalian clone "Dolly" the sheep. The combination of the technologies for gene targeting of somatic cells with those of animal cloning made it possible to introduce specific genetic mutations into domestic animals. In this review, the principles of gene targeting in somatic cells and the challenges of nuclear transfer using gene-targeted cells are discussed. The relevance of gene targeting in domestic animals for applications in bio-medicine and agriculture are also examined.

  13. Surface functionalization of liposomes with proteins and carbohydrates for use in anti-cancer applications

    Science.gov (United States)

    Platt, Virginia M.

    Liposomes can be used to exploit the altered biology of cancer thereby increasing delivery of liposome-associated anti-cancer drugs. In this dissertation, I explore methods that utilize the unique cancer expression of the polymeric glycosaminoglycan hyaluronan (HA) and the HA receptor CD44 to target liposomes to tumors, using liposomes functionalized with proteins or oligosaccharides on their surface. To make it easier to prepare protein-functionalized liposomes, a non-covalent protein/liposome association method based upon metal chelation/his 6 interaction was devised and characterized. I evaluated non-covalent attachment of the prodrug converting enzyme yeast cytosine deaminase, the far-red fluorescent protein mKate, two antigens ovalbumin and the membrane proximal region of an HIV GAG and hyaluronidase, a HA-degrading enzyme. In Chapter 2, I describe the synthesis of hyaluronan-oligosaccharide (HA-O) lipid conjugates and their incorporation into liposomes to target CD44-overexpressing cancer cells. HA-O ligands of defined-length, up to 10 monosaccharides, were attached to lipids via various linkers by reductive amination. The HA-lipids were easily incorporated into liposomes but did not mediate binding of liposomes to CD44 overexpressing cells. In Chapter 3, I evaluate the capacity of tris-NTA-Ni-lipids incorporated within a liposome bilayer to associate with his6-tagged proteins. Tris-NTA-lipids of differing structures and avidities were used to associate yeast cytosine deaminase and mKate to the surface of liposomes. Two tris-NTA-lipids and a mono-NTA lipid associated his-tagged proteins to a 1:1 molar ratio in solution. The proteins remained active while associated with the liposome surface. When challenged in vitro with fetal calf serum, tris-NTA-containing liposomes retained his-tagged proteins longer than mono-NTA. However, the tris-NTA/his6 interaction was found to be in a dynamic state; free yeast cytosine deaminase rapidly competed with pre-bound m

  14. Controlled release of an anti-cancer drug from DNA structured nano-films

    Science.gov (United States)

    Cho, Younghyun; Lee, Jong Bum; Hong, Jinkee

    2014-02-01

    We demonstrate the generation of systemically releasable anti-cancer drugs from multilayer nanofilms. Nanofilms designed to drug release profiles in programmable fashion are promising new and alternative way for drug delivery. For the nanofilm structure, we synthesized various unique 3-dimensional anti cancer drug incorporated DNA origami structures (hairpin, Y, and X shaped) and assembled with peptide via layer-by-layer (LbL) deposition method. The key to the successful application of these nanofilms requires a novel approach of the influence of DNA architecture for the drug release from functional nano-sized surface. Herein, we have taken first steps in building and controlling the drug incorporated DNA origami based multilayered nanostructure. Our finding highlights the novel and unique drug release character of LbL systems in serum condition taken full advantages of DNA origami structure. This multilayer thin film dramatically affects not only the release profiles but also the structure stability in protein rich serum condition.

  15. A Journey Under the Sea: The Quest for Marine Anti-Cancer Alkaloids

    Directory of Open Access Journals (Sweden)

    Nadine Darwiche

    2011-11-01

    Full Text Available The alarming increase in the global cancer death toll has fueled the quest for new effective anti-tumor drugs thorough biological screening of both terrestrial and marine organisms. Several plant-derived alkaloids are leading drugs in the treatment of different types of cancer and many are now being tested in various phases of clinical trials. Recently, marine-derived alkaloids, isolated from aquatic fungi, cyanobacteria, sponges, algae, and tunicates, have been found to also exhibit various anti-cancer activities including anti-angiogenic, anti-proliferative, inhibition of topoisomerase activities and tubulin polymerization, and induction of apoptosis and cytotoxicity. Two tunicate-derived alkaloids, aplidin and trabectedin, offer promising drug profiles, and are currently in phase II clinical trials against several solid and hematologic tumors. This review sheds light on the rich array of anti-cancer alkaloids in the marine ecosystem and introduces the most investigated compounds and their mechanisms of action.

  16. Hedgehog Signaling Inhibitors as Anti-Cancer Agents in Osteosarcoma

    Energy Technology Data Exchange (ETDEWEB)

    Ram Kumar, Ram Mohan, E-mail: rkumar@research.balgrist.ch; Fuchs, Bruno [Laboratory for Orthopaedic Research, Balgrist University Hospital, Sarcoma Center-UZH University of Zurich, Zurich 8008 (Switzerland)

    2015-05-13

    Osteosarcoma is a rare type of cancer associated with a poor clinical outcome. Even though the pathologic characteristics of OS are well established, much remains to be understood, particularly at the molecular signaling level. The molecular mechanisms of osteosarcoma progression and metastases have not yet been fully elucidated and several evolutionary signaling pathways have been found to be linked with osteosarcoma pathogenesis, especially the hedgehog signaling (Hh) pathway. The present review will outline the importance and targeting the hedgehog signaling (Hh) pathway in osteosarcoma tumor biology. Available data also suggest that aberrant Hh signaling has pro-migratory effects and leads to the development of osteoblastic osteosarcoma. Activation of Hh signaling has been observed in osteosarcoma cell lines and also in primary human osteosarcoma specimens. Emerging data suggests that interference with Hh signal transduction by inhibitors may reduce osteosarcoma cell proliferation and tumor growth thereby preventing osteosarcomagenesis. From this perspective, we outline the current state of Hh pathway inhibitors in osteosarcoma. In summary, targeting Hh signaling by inhibitors promise to increase the efficacy of osteosarcoma treatment and improve patient outcome.

  17. Cancer immunology - development of novel anti-cancer therapies.

    Science.gov (United States)

    Rothschild, Sacha I; Thommen, Daniela S; Moersig, Wolfgang; Müller, Philipp; Zippelius, Alfred

    2015-01-01

    The vast majority of tumours are characterised by high frequencies of genetic and epigenetic alterations resulting in tumour-specific antigens, which may, in principle, be recognised by cytotoxic T cells. Though early clinical immunotherapy trials have yielded mixed results with ambiguous clinical benefit, cancer immunotherapy is now attracting increasing attention as a viable therapeutic option, mainly in melanoma and lung cancer, but increasingly also in other malignancies. In particular, recent therapeutic efforts targeting inhibitory receptors on T cells to overcome tumour-induced immune dysfunction have the potential to reshape current treatment standards in oncology. The clinical development has been pioneered by the antibody ipilimumab, which blocks cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) and has demonstrated survival benefit in two randomised landmark trials in melanoma. Capitalising on this success, the research on the clinical implication of T cell checkpoint inhibition has been boosted. Early clinical trials have demonstrated meaningful response rates, sustained clinical benefits with encouraging survival rates and good tolerability of next-generation checkpoint inhibitors, including programmed death-1 (PD-1) and programmed death ligand 1 (PD-L1) inhibitors, across multiple cancer types. Attractive perspectives include the concurrent blockade of immunological (non-redundant) checkpoints, which has recently been demonstrated using combinations of immune checkpoint modulators themselves or with other therapies, such as chemotherapy, targeted therapy or radiotherapy. This article summarises the mechanism of action and subsequent clinical studies of immune checkpoint antibodies in oncology with a particular focus on melanoma and lung cancer.

  18. Computer-aided discovery of biological activity spectra for anti-aging and anti-cancer olive oil oleuropeins.

    Science.gov (United States)

    Corominas-Faja, Bruna; Santangelo, Elvira; Cuyàs, Elisabet; Micol, Vicente; Joven, Jorge; Ariza, Xavier; Segura-Carretero, Antonio; García, Jordi; Menendez, Javier A

    2014-09-01

    Aging is associated with common conditions, including cancer, diabetes, cardiovascular disease, and Alzheimer's disease. The type of multi-targeted pharmacological approach necessary to address a complex multifaceted disease such as aging might take advantage of pleiotropic natural polyphenols affecting a wide variety of biological processes. We have recently postulated that the secoiridoids oleuropein aglycone (OA) and decarboxymethyl oleuropein aglycone (DOA), two complex polyphenols present in health-promoting extra virgin olive oil (EVOO), might constitute a new family of plant-produced gerosuppressant agents. This paper describes an analysis of the biological activity spectra (BAS) of OA and DOA using PASS (Prediction of Activity Spectra for Substances) software. PASS can predict thousands of biological activities, as the BAS of a compound is an intrinsic property that is largely dependent on the compound's structure and reflects pharmacological effects, physiological and biochemical mechanisms of action, and specific toxicities. Using Pharmaexpert, a tool that analyzes the PASS-predicted BAS of substances based on thousands of "mechanism-effect" and "effect-mechanism" relationships, we illuminate hypothesis-generating pharmacological effects, mechanisms of action, and targets that might underlie the anti-aging/anti-cancer activities of the gerosuppressant EVOO oleuropeins.

  19. The substance P/NK-1 receptor system: NK-1 receptor antagonists as anti-cancer drugs

    Indian Academy of Sciences (India)

    Miguel Muñoz; Rafael Coveñas; Francisco Esteban; Maximino Redondo

    2015-06-01

    The substance P (SP)/neurokinin (NK)-1 receptor system plays an important role in cancer. SP promotes the proliferation of tumour cells, angiogenesis and the migration of tumour cells. We review the involvement of SP, the NK-1 receptor and NK-1 receptor antagonists in cancer. Tumour cells overexpress NK-1 receptors, which are involved in their viability. This overexpression suggests the possibility of specific treatment against tumour cells using NK-1 receptor antagonists, thus promoting a considerable decrease in the side effects of the treatment. This strategy opens up new approaches for cancer treatment, since these antagonists, after binding to their molecular target, induce the death of tumour cells by apoptosis, exert an antiangiogenic action and inhibit the migration of tumour cells. The use of NK-1 receptor antagonists such as aprepitant (used in clinical practice) as antitumour agents could be a promising innovation. The value of aprepitant as an antitumour agent could be determined faster than for less well-known compounds because many studies addressing its safety and characterization have already been completed. The NK-1 receptor may be a promising target in the treatment of cancer; NK-1 receptor antagonists could act as specific drugs against tumour cells; and these antagonists could be new candidate anti-cancer drugs.

  20. Glycyrrhetinic Acid and Its Derivatives: Anti-Cancer and Cancer Chemopreventive Properties, Mechanisms of Action and Structure- Cytotoxic Activity Relationship.

    Science.gov (United States)

    Roohbakhsh, Ali; Iranshahy, Milad; Iranshahi, Mehrdad

    2016-01-01

    The anti-cancer properties of liquorice have been attributed, at least in part, to glycyrrhizin (GL). However, GL is not directly absorbed through the gastrointestinal tract. It is hydrolyzed to 18-β-glycyrrhetinic acid (GA), the pharmacologically active metabolite, by human intestinal microflora. GA exhibits remarkable cytotoxic and anti-tumor properties. The pro-apoptotic targets and mechanisms of action of GA have been extensively studied over the past decade. In addition, GA is an inexpensive and available triterpene with functional groups (COOH and OH) in its structure, which make it an attractive lead compound for medicinal chemists to prepare a large number of analogues. To date, more than 400 cytotoxic derivatives have been prepared on the basis of GA scaffold, including 128 cytotoxic derivatives with IC50 values less than 30 µM. Researchers have also succeeded in synthesizing very potent cytotoxic derivatives with IC50s ≤ 1 µM. Studies have shown that the introduction of a double bound at the C1-C2 position combined with an electronegative functional group, such as CN, CF3 or iodine at C2 position, and the oxidation of the hydroxyl group of C3 to the carbonyl group, significantly increased cytotoxicity. This review describes the cytotoxic and anti-tumor properties of GA and its derivatives, targets and mechanisms of action and provides insight into the structure-activity relationship of GA derivatives.

  1. Computer-aided discovery of biological activity spectra for anti-aging and anti-cancer olive oil oleuropeins

    Science.gov (United States)

    Corominas-Faja, Bruna; Santangelo, Elvira; Cuyàs, Elisabet; Micol, Vicente; Joven, Jorge; Ariza, Xavier; Segura-Carretero, Antonio; García, Jordi; Menendez, Javier A.

    2014-01-01

    Aging is associated with common conditions, including cancer, diabetes, cardiovascular disease, and Alzheimer's disease. The type of multi-targeted pharmacological approach necessary to address a complex multifaceteddisease such as aging might take advantage of pleiotropic natural polyphenols affecting a wide variety of biological processes. We have recently postulated that the secoiridoids oleuropein aglycone (OA) and decarboxymethyl oleuropein aglycone (DOA), two complex polyphenols present in health-promoting extra virgin olive oil (EVOO), might constitute anew family of plant-produced gerosuppressant agents. This paper describes an analysis of the biological activity spectra (BAS) of OA and DOA using PASS (Prediction of Activity Spectra for Substances) software. PASS can predict thousands of biological activities, as the BAS of a compound is an intrinsic property that is largely dependent on the compound's structure and reflects pharmacological effects, physiological and biochemical mechanisms of action, and specific toxicities. Using Pharmaexpert, a tool that analyzes the PASS-predicted BAS of substances based on thousands of “mechanism-effect” and “effect-mechanism” relationships, we illuminate hypothesis-generating pharmacological effects, mechanisms of action, and targets that might underlie the anti-aging/anti-cancer activities of the gerosuppressant EVOO oleuropeins. PMID:25324469

  2. Protocatechualdehyde possesses anti-cancer activity through downregulating cyclin D1 and HDAC2 in human colorectal cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Jeong, Jin Boo [Department of Nutrition and Food Science, University of Maryland, College Park, MD 20742 (United States); Lee, Seong-Ho, E-mail: slee2000@umd.edu [Department of Nutrition and Food Science, University of Maryland, College Park, MD 20742 (United States)

    2013-01-04

    Highlights: Black-Right-Pointing-Pointer Protocatechualdehyde (PCA) suppressed cell proliferation and induced apoptosis in human colorectal cancer cells. Black-Right-Pointing-Pointer PCA enhanced transcriptional downregulation of cyclin D1 gene. Black-Right-Pointing-Pointer PCA suppressed HDAC2 expression and activity. Black-Right-Pointing-Pointer These findings suggest that anti-cancer activity of PCA may be mediated by reducing HDAC2-derived cyclin D1 expression. -- Abstract: Protocatechualdehyde (PCA) is a naturally occurring polyphenol found in barley, green cavendish bananas, and grapevine leaves. Although a few studies reported growth-inhibitory activity of PCA in breast and leukemia cancer cells, the underlying mechanisms are still poorly understood. Thus, we performed in vitro study to investigate if treatment of PCA affects cell proliferation and apoptosis in human colorectal cancer cells and define potential mechanisms by which PCA mediates growth arrest and apoptosis of cancer cells. Exposure of PCA to human colorectal cancer cells (HCT116 and SW480 cells) suppressed cell growth and induced apoptosis in dose-dependent manner. PCA decreased cyclin D1 expression in protein and mRNA level and suppressed luciferase activity of cyclin D1 promoter, indicating transcriptional downregulation of cyclin D1 gene by PCA. We also observed that PCA treatment attenuated enzyme activity of histone deacetylase (HDAC) and reduced expression of HDAC2, but not HDAC1. These findings suggest that cell growth inhibition and apoptosis by PCA may be a result of HDAC2-mediated cyclin D1 suppression.

  3. Nanotech revolution for the anti-cancer drug delivery through blood-brain barrier.

    Science.gov (United States)

    Caraglia, M; De Rosa, G; Salzano, G; Santini, D; Lamberti, M; Sperlongano, P; Lombardi, A; Abbruzzese, A; Addeo, R

    2012-03-01

    Nanotechnology-based drug delivery was born as a chance for pharmaceutical weapons to be delivered in the body sites where drug action is required. Specifically, the incorporation of anti-cancer agents in nanodevices of 100-300 nm allows their delivery in tissues that have a fenestrated vasculature and a reduced lymphatic drainage. These two features are typical of neoplastic tissues and, therefore, allow the accumulation of nanostructured devices in tumours. An important issue of anti-cancer pharmacological strategies is the overcoming of anatomical barriers such as the bloodbrain- barrier (BBB) that protects brain from toxicological injuries but, at the same time, makes impossible for most of the pharmacological agents with anti-cancer activity to reach tumour cells placed in the brain and derived from either primary tumours or metastases. In fact, only highly lipophilic molecules can passively diffuse through BBB to reach central nervous system (CNS). Another possibility is to use nanotechnological approaches as powerful tools to across BBB, by both prolonging the plasma half-life of the drugs and crossing fenestrations of BBB damaged by brain metastases. Moreover, modifications of nanocarrier surface with specific endogenous or exogenous ligands can promote the crossing of intact BBB as in the case of primary brain tumours. This aim can be achieved through the binding of the nanodevices to carriers or receptors expressed by the endothelial cells of BBB and that can favour the internalization of the nanostructured devices delivering anti-cancer drugs. This review summarizes the most meaningful advances in the field of nanotechnologies for brain delivery of drugs.

  4. Screening for Anti-Cancer Compounds in Marine Organisms in Oman

    OpenAIRE

    Sergey Dobretsov; Yahya Tamimi; Al-Kindi, Mohamed A.; Ikram Burney

    2016-01-01

    Objectives: Marine organisms are a rich source of bioactive molecules with potential applications in medicine, biotechnology and industry; however, few bioactive compounds have been isolated from organisms inhabiting the Arabian Gulf and the Gulf of Oman. This study aimed to isolate and screen the anti-cancer activity of compounds and extracts from 40 natural products of marine organisms collected from the Gulf of Oman. Methods: This study was carried out between January 2012 and December ...

  5. Advances in identification and application of tumor antigen inducing anti-cancer responses

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    @@ Tumor antigen is one of the important bases of tumor immunotherapy[1]. With the discovery of novel tumor antigens, interest in specific immunotherapy for treatment of malignancies has increased substantially. Nowadays more and more scientists paid close attention to various tumor antigens with their roles or/and applications in anti-cancer immune responses, immune tolerance, tumor markers, tumor immunotherapy and so on. Here we discussed the classification of tumor antigens and summarized the technologies of identification and application of tumor antigens.

  6. Quantification of cell viability and rapid screening anti-cancer drug utilizing nanomechanical fluctuation.

    Science.gov (United States)

    Wu, Shangquan; Liu, Xiaoli; Zhou, Xiarong; Liang, Xin M; Gao, Dayong; Liu, Hong; Zhao, Gang; Zhang, Qingchuan; Wu, Xiaoping

    2016-03-15

    Cancer is a serious threat to human health. Although numerous anti-cancer drugs are available clinically, many have shown toxic side effects due to poor tumor-selectivity, and reduced effectiveness due to cancers rapid development of resistance to treatment. The development of new highly efficient and practical methods to quantify cell viability and its change under drug treatment is thus of significant importance in both understanding of anti-cancer mechanism and anti-cancer drug screening. Here, we present an approach of utilizing a nanomechanical fluctuation based highly sensitive microcantilever sensor, which is capable of characterizing the viability of cells and quantitatively screening (within tens of minutes) their responses to a drug with the obvious advantages of a rapid, label-free, quantitative, noninvasive, real-time and in-situ assay. The microcantilever sensor operated in fluctuation mode was used in evaluating the paclitaxel effectiveness on breast cancer cell line MCF-7. This study demonstrated that the nanomechanical fluctuations of the microcantilever sensor are sensitive enough to detect the dynamic variation in cellular force which is provided by the cytoskeleton, using cell metabolism as its energy source, and the dynamic instability of microtubules plays an important role in the generation of the force. We propose that cell viability consists of two parts: biological viability and mechanical viability. Our experimental results suggest that paclitaxel has little effect on biological viability, but has a significant effect on mechanical viability. This new method provides a new concept and strategy for the evaluation of cell viability and the screening of anti-cancer drugs.

  7. Comparison of quantitative PCR assays for Escherichia coli targeting ribosomal RNA and single copy genes

    Science.gov (United States)

    Aims: Compare specificity and sensitivity of quantitative PCR (qPCR) assays targeting single and multi-copy gene regions of Escherichia coli. Methods and Results: A previously reported assay targeting the uidA gene (uidA405) was used as the basis for comparing the taxono...

  8. Anti-cancer vaccination by transdermal delivery of antigen peptide-loaded nanogels via iontophoresis.

    Science.gov (United States)

    Toyoda, Mao; Hama, Susumu; Ikeda, Yutaka; Nagasaki, Yukio; Kogure, Kentaro

    2015-04-10

    Transdermal vaccination with cancer antigens is expected to become a useful anti-cancer therapy. However, it is difficult to accumulate enough antigen in the epidermis for effective exposure to Langerhans cells because of diffusion into the skin and muscle. Carriers, such as liposomes and nanoparticles, may be useful for the prevention of antigen diffusion. Iontophoresis, via application of a small electric current, is a noninvasive and efficient technology for transdermal drug delivery. Previously, we succeeded in the iontophoretic transdermal delivery of liposomes encapsulating insulin, and accumulation of polymer-based nanoparticle nanogels in the stratum corneum of the skin. Therefore, in the present study, we examined the use of iontophoresis with cancer antigen gp-100 peptide KVPRNQDWL-loaded nanogels for anti-cancer vaccination. Iontophoresis resulted in the accumulation of gp-100 peptide and nanogels in the epidermis, and subsequent increase in the number of Langerhans cells in the epidermis. Moreover, tumor growth was significantly suppressed by iontophoresis of the antigen peptide-loaded nanogels. Thus, iontophoresis of the antigen peptide-loaded nanogels may serve as an effective transdermal delivery system for anti-cancer vaccination.

  9. Anti-cancer Activities of Ginseng Extract Fermented with Phellinus linteus.

    Science.gov (United States)

    Lee, Jong-Jin; Kwon, Ho-Kyun; Jung, In-Ho; Cho, Yong-Baik; Kim, Kyu-Joong; Kim, Jong-Lae

    2009-03-01

    In the present study, the anti-cancer effects of ginseng fermented with Phellinus linteus (GFPL) extract were examined through in vitro and in vivo assays. GFPL was produced by co-cultivating ginseng and Phellinus linteus together. Ginsenoside Rg3, Rh1 and Rh2 are important mediators of anti-angiogenesis and their levels in GFPL were enriched 24, 19 and 16 times, respectively, more than that of ginseng itself through the fermentation. GFPL exhibited distinct anti-cancer effects, including growth inhibition of the human lung carcinoma cell line A549, and promotion of immune activation by stimulating nitric oxide (NO) production in Raw 264.7 cells. Further evidence supporting anti-cancer effects of GFPL was its significant prolongment of the survival of B16F10 cancer cell-implanted mice. These results suggest that the GFPL may be a candidate for cancer prevention and treatment through immune activation and anti-angiogenic effects by enriching Rg3, Rh1 and Rh2.

  10. Readability Comparison of Pro- and Anti-Cancer Screening Online Messages in Japan

    Science.gov (United States)

    Okuhara, Tsuyoshi; Ishikawa, Hirono; Okada, Masahumi; Kato, Mio; Kiuchi, Takahiro

    2016-12-01

    Background: Cancer screening rates are lower in Japan than those in western countries. Health professionals publish procancer screening messages on the internet to encourage audiences to undergo cancer screening. However, the information provided is often difficult to read for lay persons. Further, anti-cancer screening activists warn against cancer screening with messages on the Internet. We aimed to assess and compare the readability of pro- and anti-cancer screening online messages in Japan using a measure of readability. Methods: We conducted web searches at the beginning of September 2016 using two major Japanese search engines (Google.jp and Yahoo!.jp). The included websites were classified as “anti”, “pro”, or “neutral” depending on the claims, and “health professional” or “non-health professional” depending on the writers. Readability was determined using a validated measure of Japanese readability. Statistical analysis was conducted using two-way ANOVA. Results: In the total 159 websites analyzed, anti-cancer screening online messages were generally easier to read than pro-cancer screening online messages, Messages written by health professionals were more difficult to read than those written by non-health professionals. Claim × writer interaction was not significant. Conclusion: When health professionals prepare pro-cancer screening materials for publication online, we recommend they check for readability using readability assessment tools and improve text for easy comprehension when necessary.

  11. Molecular mechanisms of anti-cancer action of garlic compounds in neuroblastoma.

    Science.gov (United States)

    Karmakar, Surajit; Choudhury, Subhasree Roy; Banik, Naren L; Ray, Swapan K

    2011-05-01

    The medicinal properties of garlic (Allium sativum) have been well known and widely used since historical times. Garlic compounds have received increasing attention during the last few years due to their cancer chemopreventive properties. The anti-cancer activity of garlic-derived organosulfur compounds (OSCs) are extensively reported in many cancers but only a few in the pediatric tumor neuroblastoma, which warrants exploration of new therapy for its management. There are some recent reports suggesting that garlic-derived OSCs cause cell cycle arrest, generate reactive oxygen species (ROS), activate stress kinases, and also stimulate the mitochondrial pathway for apoptosis in malignant neuroblastoma. The comprehensive mechanisms of anti-cancer action of OSCs still remain unclear and require more studies in neuroblastoma. This review is designed to highlight the molecular mechanisms of anti-cancer actions of garlic-derived OSCs in neuroblastoma and as well as in several other cancers. Further studies should be conducted to establish the clinical expediency of garlic-derived OSCs for treatment of malignant neuroblastoma in humans.

  12. Control of target gene specificity during metamorphosis by the steroid response gene E93.

    Science.gov (United States)

    Mou, Xiaochun; Duncan, Dianne M; Baehrecke, Eric H; Duncan, Ian

    2012-02-21

    Hormonal control of sexual maturation is a common feature in animal development. A particularly dramatic example is the metamorphosis of insects, in which pulses of the steroid hormone ecdysone drive the wholesale transformation of the larva into an adult. The mechanisms responsible for this transformation are not well understood. Work in Drosophila indicates that the larval and adult forms are patterned by the same underlying sets of developmental regulators, but it is not understood how the same regulators pattern two distinct forms. Recent studies indicate that this ability is facilitated by a global change in the responsiveness of target genes during metamorphosis. Here we show that this shift is controlled in part by the ecdysone-induced transcription factor E93. Although long considered a dedicated regulator of larval cell death, we find that E93 is expressed widely in adult cells at the pupal stage and is required for many patterning processes at this time. To understand the role of E93 in adult patterning, we focused on a simple E93-dependent process, the induction of the Dll gene within bract cells of the pupal leg by EGF receptor signaling. In this system, we show that E93 functions to cause Dll to become responsive to EGF receptor signaling. We demonstrate that E93 is both necessary and sufficient for directing this switch. E93 likely controls the responsiveness of many other target genes because it is required broadly for patterning during metamorphosis. The wide conservation of E93 orthologs suggests that similar mechanisms control life-cycle transitions in other organisms, including vertebrates.

  13. Advances of Driver Gene and Targeted Therapy of Non-small Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Dan ZHANG

    2014-10-01

    Full Text Available Lung cancer is the leading cause of cancer-related mortality in the worldwide. The discovery of drive gene makes tumor treatment is no longer "one-size-fits-all". Targeted therapy to change the present situation of cancer drugs become "bullet" with eyes, the effect is visible and bring a revolution in the treatment of lung cancer. The diver gene and targeted therapy have became the new cedule of non-small cell lung cancer (NSCLC. Society of Clinical Oncology (ASCO has showed 11 kinds of diver genes. Here, we review the functional and structural characteristics and the targeted therapy in the 11 kinds of driver gene mutations.

  14. [Advances of driver gene and targeted therapy of non-small cell lung cancer].

    Science.gov (United States)

    Zhang, Dan; Huang, Yan; Wang, Hongyang

    2014-10-20

    Lung cancer is the leading cause of cancer-related mortality in the worldwide. The discovery of drive gene makes tumor treatment is no longer "one-size-fits-all". Targeted therapy to change the present situation of cancer drugs become "bullet" with eyes, the effect is visible and bring a revolution in the treatment of lung cancer. The diver gene and targeted therapy have became the new cedule of non-small cell lung cancer (NSCLC). Society of Clinical Oncology (ASCO) has showed 11 kinds of diver genes. Here, we review the functional and structural characteristics and the targeted therapy in the 11 kinds of driver gene mutations.

  15. The latest advances of experimental research on targeted gene therapy for prostate cancer

    Institute of Scientific and Technical Information of China (English)

    Dongliang Pan; Lianchao Jin; Xianghua Zhang

    2013-01-01

    The absence of ef ective therapies for castration-resistant prostate cancer (CRPC) establishes the need to de-velop novel therapeutic modality, such as targeted gene therapy, which is ideal for the treatment of CRPC. But its application has been limited due to lack of favorable gene vector and the reduction of“bystander ef ect”. Consequently, scientists al over the world focus their main experimental research on the fol owing four aspects:targeted gene, vector, transfer means and comprehensive therapy. In this paper, we reviewed the latest advances of experimental research on targeted gene therapy for prostate cancer .

  16. Rapid and Cost-Effective Gene Targeting in Rat Embryonic Stem Cells by TALENs

    Institute of Scientific and Technical Information of China (English)

    Chang Tong; Guanyi Huang; Charles Ashton; Hongping Wu; Hexin Yan; Qi-Long Ying

    2012-01-01

    The rat is the preferred animal model in many areas of biomedical research and drug development.Genetic manipulation in rats has lagged behind that in mice due to the lack of efficient gene targeting tools.Previously,we generated a knockout rat via conventional homologous recombination in rat embryonic stem (ES) cells.Here,we show that efficient gene targeting in rat ES cells can be achieved quickly through transcription activator-like effector nuclease (TALEN)-mediated DNA double-strand breaks.Using the Golden Gate cloning technique,we constructed a pair of TALEN targeting vectors for the gene of interest in 5 days.After gene transfection,the targeted rat ES cell colonies were isolated,screened,and confirmed by PCR without the need of drug selection.Our results suggest that TALEN-mediated gene targeting is a superior means of establishing genetically modified rat ES cell lines with high efficiency and short turnaround time.

  17. Targeted Therapy and Immunotherapy for Non-small Cell Lung Cancer 
with Brain Metastasis

    OpenAIRE

    Song, Qi; Jiao, Shunchang; Li, Fang

    2016-01-01

    Brain metastasis, a common complication of non-small cell lung cancer (NSCLC) with an incidence rate of 30%-50%, significantly affects the patients’ quality of life. The prognosis of patients of NSCLC with brain metastasis is extremely poor, the average median survival is only 1 m-2 m without treatment. The targeted therapy based on lung cancer driven gene is a new treatment. Besides, the immunotherapy which can enhance the effect of anti-cancer by simulating the immune system is a new approa...

  18. Using PCR to Target Misconceptions about Gene Expression

    Directory of Open Access Journals (Sweden)

    Leslie K. Wright

    2013-02-01

    Full Text Available We present a PCR-based laboratory exercise that can be used with first- or second-year biology students to help overcome common misconceptions about gene expression. Biology students typically do not have a clear understanding of the difference between genes (DNA and gene expression (mRNA/protein and often believe that genes exist in an organism or cell only when they are expressed. This laboratory exercise allows students to carry out a PCR-based experiment designed to challenge their misunderstanding of the difference between genes and gene expression. Students first transform E. coli with an inducible GFP gene containing plasmid and observe induced and un-induced colonies. The following exercise creates cognitive dissonance when actual PCR results contradict their initial (incorrect predictions of the presence of the GFP gene in transformed cells. Field testing of this laboratory exercise resulted in learning gains on both knowledge and application questions on concepts related to genes and gene expression.

  19. An approach for the identification of targets specific to bone metastasis using cancer genes interactome and gene ontology analysis.

    Directory of Open Access Journals (Sweden)

    Shikha Vashisht

    Full Text Available Metastasis is one of the most enigmatic aspects of cancer pathogenesis and is a major cause of cancer-associated mortality. Secondary bone cancer (SBC is a complex disease caused by metastasis of tumor cells from their primary site and is characterized by intricate interplay of molecular interactions. Identification of targets for multifactorial diseases such as SBC, the most frequent complication of breast and prostate cancers, is a challenge. Towards achieving our aim of identification of targets specific to SBC, we constructed a 'Cancer Genes Network', a representative protein interactome of cancer genes. Using graph theoretical methods, we obtained a set of key genes that are relevant for generic mechanisms of cancers and have a role in biological essentiality. We also compiled a curated dataset of 391 SBC genes from published literature which serves as a basis of ontological correlates of secondary bone cancer. Building on these results, we implement a strategy based on generic cancer genes, SBC genes and gene ontology enrichment method, to obtain a set of targets that are specific to bone metastasis. Through this study, we present an approach for probing one of the major complications in cancers, namely, metastasis. The results on genes that play generic roles in cancer phenotype, obtained by network analysis of 'Cancer Genes Network', have broader implications in understanding the role of molecular regulators in mechanisms of cancers. Specifically, our study provides a set of potential targets that are of ontological and regulatory relevance to secondary bone cancer.

  20. Efficient four fragment cloning for the construction of vectors for targeted gene replacement in filamentous fungi

    DEFF Research Database (Denmark)

    Frandsen, Rasmus John Normand; Andersson, Jens A.; Kristensen, Matilde Bylov;

    2008-01-01

    technique that allows single step cloning of the two required homologous recombination sequences into different sites of a recipient vector. The advantages are: A simple experimental design, free choice of target sequence, few procedures and user convenience. The vectors are intented for Agrobacterium...... with an average efficiency of 84% for gene replacement and 80% for targeted overexpression. Conclusion: The new vectors designed for USER Friendly cloning provided a fast reliable method to construct vectors for targeted gene manipulations in fungi....

  1. Human recombinant RNASET2: A potential anti-cancer drug

    Science.gov (United States)

    Roiz, Levava; Smirnoff, Patricia; Lewin, Iris; Shoseyov, Oded; Schwartz, Betty

    2016-01-01

    The roles of cell motility and angiogenetic processes in metastatic spread and tumor aggressiveness are well established and must be simultaneously targeted to maximize antitumor drug potency. This work evaluated the antitumorigenic capacities of human recombinant RNASET2 (hrRNASET2), a homologue of the Aspergillus niger T2RNase ACTIBIND, which has been shown to display both antitumorigenic and antiangiogenic activities. hrRNASET2 disrupted intracellular actin filament and actin-rich extracellular extrusion organization in both CT29 colon cancer and A375SM melanoma cells and induced a significant dose-dependent inhibition of A375SM cell migration. hrRNASET2 also induced full arrest of angiogenin-induced tube formation and brought to a three-fold lower relative HT29 colorectal and A375SM melanoma tumor volume, when compared to Avastin-treated animals. In parallel, mean blood vessel counts were 36.9% lower in hrRNASET2-vs. Avastin-treated mice and survival rates of hrRNASET2-treated mice were 50% at 73 days post-treatment, while the median survival time for untreated animals was 22 days. Moreover, a 60-day hrRNASET2 treatment period reduced mean A375SM lung metastasis foci counts by three-fold when compared to untreated animals. Taken together, the combined antiangiogenic and antitumorigenic capacities of hrRNASET2, seemingly arising from its direct interaction with intercellular and extracellular matrices, render it an attractive anticancer therapy candidate. PMID:27014725

  2. Human recombinant RNASET2: A potential anti-cancer drug.

    Science.gov (United States)

    Roiz, Levava; Smirnoff, Patricia; Lewin, Iris; Shoseyov, Oded; Schwartz, Betty

    2016-01-01

    The roles of cell motility and angiogenetic processes in metastatic spread and tumor aggressiveness are well established and must be simultaneously targeted to maximize antitumor drug potency. This work evaluated the antitumorigenic capacities of human recombinant RNASET2 (hrRNASET2), a homologue of the Aspergillus niger T2RNase ACTIBIND, which has been shown to display both antitumorigenic and antiangiogenic activities. hrRNASET2 disrupted intracellular actin filament and actin-rich extracellular extrusion organization in both CT29 colon cancer and A375SM melanoma cells and induced a significant dose-dependent inhibition of A375SM cell migration. hrRNASET2 also induced full arrest of angiogenin-induced tube formation and brought to a three-fold lower relative HT29 colorectal and A375SM melanoma tumor volume, when compared to Avastin-treated animals. In parallel, mean blood vessel counts were 36.9% lower in hrRNASET2-vs. Avastin-treated mice and survival rates of hrRNASET2-treated mice were 50% at 73 days post-treatment, while the median survival time for untreated animals was 22 days. Moreover, a 60-day hrRNASET2 treatment period reduced mean A375SM lung metastasis foci counts by three-fold when compared to untreated animals. Taken together, the combined antiangiogenic and antitumorigenic capacities of hrRNASET2, seemingly arising from its direct interaction with intercellular and extracellular matrices, render it an attractive anticancer therapy candidate.

  3. Anti cancer effects of curcumin: cycle of life and death

    Directory of Open Access Journals (Sweden)

    Das Tanya

    2008-10-01

    Full Text Available Abstract Increasing knowledge on the cell cycle deregulations in cancers has promoted the introduction of phytochemicals, which can either modulate signaling pathways leading to cell cycle regulation or directly alter cell cycle regulatory molecules, in cancer therapy. Most human malignancies are driven by chromosomal translocations or other genetic alterations that directly affect the function of critical cell cycle proteins such as cyclins as well as tumor suppressors, e.g., p53. In this respect, cell cycle regulation and its modulation by curcumin are gaining widespread attention in recent years. Extensive research has addressed the chemotherapeutic potential of curcumin (diferuloylmethane, a relatively non-toxic plant derived polyphenol. The mechanisms implicated are diverse and appear to involve a combination of cell signaling pathways at multiple levels. In the present review we discuss how alterations in the cell cycle control contribute to the malignant transformation and provide an overview of how curcumin targets cell cycle regulatory molecules to assert anti-proliferative and/or apoptotic effects in cancer cells. The purpose of the current article is to present an appraisal of the current level of knowledge regarding the potential of curcumin as an agent for the chemoprevention of cancer via an understanding of its mechanism of action at the level of cell cycle regulation. Taken together, this review seeks to summarize the unique properties of curcumin that may be exploited for successful clinical cancer prevention.

  4. Curcumin AntiCancer Studies in Pancreatic Cancer

    Science.gov (United States)

    Bimonte, Sabrina; Barbieri, Antonio; Leongito, Maddalena; Piccirillo, Mauro; Giudice, Aldo; Pivonello, Claudia; de Angelis, Cristina; Granata, Vincenza; Palaia, Raffaele; Izzo, Francesco

    2016-01-01

    Pancreatic cancer (PC) is one of the deadliest cancers worldwide. Surgical resection remains the only curative therapeutic treatment for this disease, although only the minority of patients can be resected due to late diagnosis. Systemic gemcitabine-based chemotherapy plus nab-paclitaxel are used as the gold-standard therapy for patients with advanced PC; although this treatment is associated with a better overall survival compared to the old treatment, many side effects and poor results are still present. Therefore, new alternative therapies have been considered for treatment of advanced PC. Several preclinical studies have demonstrated that curcumin, a naturally occurring polyphenolic compound, has anticancer effects against different types of cancer, including PC, by modulating many molecular targets. Regarding PC, in vitro studies have shown potent cytotoxic effects of curcumin on different PC cell lines including MiaPaCa-2, Panc-1, AsPC-1, and BxPC-3. In addition, in vivo studies on PC models have shown that the anti-proliferative effects of curcumin are caused by the inhibition of oxidative stress and angiogenesis and are due to the induction of apoptosis. On the basis of these results, several researchers tested the anticancer effects of curcumin in clinical trials, trying to overcome the poor bioavailability of this agent by developing new bioavailable forms of curcumin. In this article, we review the results of pre-clinical and clinical studies on the effects of curcumin in the treatment of PC. PMID:27438851

  5. Curcumin AntiCancer Studies in Pancreatic Cancer

    Directory of Open Access Journals (Sweden)

    Sabrina Bimonte

    2016-07-01

    Full Text Available Pancreatic cancer (PC is one of the deadliest cancers worldwide. Surgical resection remains the only curative therapeutic treatment for this disease, although only the minority of patients can be resected due to late diagnosis. Systemic gemcitabine-based chemotherapy plus nab-paclitaxel are used as the gold-standard therapy for patients with advanced PC; although this treatment is associated with a better overall survival compared to the old treatment, many side effects and poor results are still present. Therefore, new alternative therapies have been considered for treatment of advanced PC. Several preclinical studies have demonstrated that curcumin, a naturally occurring polyphenolic compound, has anticancer effects against different types of cancer, including PC, by modulating many molecular targets. Regarding PC, in vitro studies have shown potent cytotoxic effects of curcumin on different PC cell lines including MiaPaCa-2, Panc-1, AsPC-1, and BxPC-3. In addition, in vivo studies on PC models have shown that the anti-proliferative effects of curcumin are caused by the inhibition of oxidative stress and angiogenesis and are due to the induction of apoptosis. On the basis of these results, several researchers tested the anticancer effects of curcumin in clinical trials, trying to overcome the poor bioavailability of this agent by developing new bioavailable forms of curcumin. In this article, we review the results of pre-clinical and clinical studies on the effects of curcumin in the treatment of PC.

  6. Multiple Mechanisms of Anti-Cancer Effects Exerted by Astaxanthin

    Directory of Open Access Journals (Sweden)

    Li Zhang

    2015-07-01

    Full Text Available Astaxanthin (ATX is a xanthophyll carotenoid which has been approved by the United States Food and Drug Administration (USFDA as food colorant in animal and fish feed. It is widely found in algae and aquatic animals and has powerful anti-oxidative activity. Previous studies have revealed that ATX, with its anti-oxidative property, is beneficial as a therapeutic agent for various diseases without any side effects or toxicity. In addition, ATX also shows preclinical anti-tumor efficacy both in vivo and in vitro in various cancer models. Several researches have deciphered that ATX exerts its anti-proliferative, anti-apoptosis and anti-invasion influence via different molecules and pathways including signal transducer and activator of transcription 3 (STAT3, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB and peroxisome proliferator-activated receptor gamma (PPARγ. Hence, ATX shows great promise as chemotherapeutic agents in cancer. Here, we review the rapidly advancing field of ATX in cancer therapy as well as some molecular targets of ATX.

  7. Applications of Gene Targeting Technology to Mental Retardation and Developmental Disability Research

    Science.gov (United States)

    Pimenta, Aurea F.; Levitt, Pat

    2005-01-01

    The human and mouse genome projects elucidated the sequence and position map of innumerous genes expressed in the central nervous system (CNS), advancing our ability to manipulate these sequences and create models to investigate regulation of gene expression and function. In this article, we reviewed gene targeting methodologies with emphasis on…

  8. Identification of therapeutic targets for Alzheimer's disease via differentially expressed gene and weighted gene co-expression network analyses.

    Science.gov (United States)

    Jia, Yujie; Nie, Kun; Li, Jing; Liang, Xinyue; Zhang, Xuezhu

    2016-11-01

    In order to investigate the pathogenic targets and associated biological process of Alzheimer's disease in the present study, mRNA expression profiles (GSE28146) and microRNA (miRNA) expression profiles (GSE16759) were downloaded from the Gene Expression Omnibus database. In GSE28146, eight control samples, and Alzheimer's disease samples comprising seven incipient, eight moderate, seven severe Alzheimer's disease samples, were included. The Affy package in R was used for background correction and normalization of the raw microarray data. The differentially expressed genes (DEGs) and differentially expressed miRNAs were identified using the Limma package. In addition, mRNAs were clustered using weighted gene correlation network analysis, and modules found to be significantly associated with the stages of Alzheimer's disease were screened out. The Database for Annotation, Visualization, and Integrated Discovery was used to perform Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses. The target genes of the differentially expressed miRNAs were identified using the miRWalk database. Compared with the control samples, 175,59 genes and 90 DEGs were identified in the incipient, moderate and severe Alzheimer's disease samples, respectively. A module, which contained 1,592 genes was found to be closely associated with the stage of Alzheimer's disease and biological processes. In addition, pathways associated with Alzheimer's disease and other neurological diseases were found to be enriched in those genes. A total of 139 overlapped genes were identified between those genes and the DEGs in the three groups. From the miRNA expression profiles, 189 miRNAs were found differentially expressed in the samples from patients with Alzheimer's disease and 1,647 target genes were obtained. In addition, five overlapped genes were identified between those 1,647 target genes and the 139 genes, and these genes may be important pathogenic targets for Alzheimer

  9. New Anti-cancer Progress Scored in Vascular-targeting Treatment

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    @@ A research team headed by YAN Xiyun at the CAS Institute of Biophysics (IBP) has scored encouraging progress in developing a vasculartargeting therapy against cancers.The cancer is the No.1 killer disease in today's world. Three conventional approaches have been developed by physicians to deal with it,namely, the surgical removal, chemotherapy and radiotherapy.

  10. PPARgene: A Database of Experimentally Verified and Computationally Predicted PPAR Target Genes.

    Science.gov (United States)

    Fang, Li; Zhang, Man; Li, Yanhui; Liu, Yan; Cui, Qinghua; Wang, Nanping

    2016-01-01

    The peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors of the nuclear receptor superfamily. Upon ligand binding, PPARs activate target gene transcription and regulate a variety of important physiological processes such as lipid metabolism, inflammation, and wound healing. Here, we describe the first database of PPAR target genes, PPARgene. Among the 225 experimentally verified PPAR target genes, 83 are for PPARα, 83 are for PPARβ/δ, and 104 are for PPARγ. Detailed information including tissue types, species, and reference PubMed IDs was also provided. In addition, we developed a machine learning method to predict novel PPAR target genes by integrating in silico PPAR-responsive element (PPRE) analysis with high throughput gene expression data. Fivefold cross validation showed that the performance of this prediction method was significantly improved compared to the in silico PPRE analysis method. The prediction tool is also implemented in the PPARgene database.

  11. Identification of novel gene targets and functions of p21-activated kinase 1 during DNA damage by gene expression profiling.

    Directory of Open Access Journals (Sweden)

    Mona Motwani

    Full Text Available P21-activated kinase 1 (PAK1, a serine/threonine protein kinase, modulates many cellular processes by phosphorylating its downstream substrates. In addition to its role in the cytoplasm, PAK1 also affects gene transcription due to its nuclear localization and association with chromatin. It is now recognized that PAK1 kinase activity and its nuclear translocation are rapidly stimulated by ionizing radiation (IR, and that PAK1 activation is a component of the DNA damage response. Owing to the role of PAK1 in the cell survival, its association with the chromatin, and now, stimulation by ionizing radiation, we hypothesize that PAK1 may be contributing to modulation of genes with roles in cellular processes that might be important in the DNA damage response. The purpose of this study was to identify new PAK1 targets in response to ionizing radiation with putative role in the DNA damage response. We examined the effect of IR on the gene expression patterns in the murine embryonic fibroblasts with or without Pak1 using microarray technology. Differentially expressed transcripts were identified using Gene Spring GX 10.0.2. Pathway, network, functional analyses and gene family classification were carried out using Kyoto Encyclopedia of Genes and Genomes (KEGG, Ingenuity Pathway, Gene Ontology and PANTHER respectively. Selective targets of PAK1 were validated by RT-qPCR. For the first time, we provide a genome-wide analysis of PAK1 and identify its targets with potential roles in the DNA damage response. Gene Ontology analysis identified genes in the IR-stimulated cells that were involved in cell cycle arrest and cell death. Pathway analysis revealed p53 pathway being most influenced by IR responsive, PAK1 targets. Gene family of transcription factors was over represented and gene networks involved in DNA replication, repair and cellular signaling were identified. In brief, this study identifies novel PAK1 dependent IR responsive genes which reveal new

  12. Design, synthesis, and evaluation of asymmetric EF24 analogues as potential anti-cancer agents for lung cancer.

    Science.gov (United States)

    Wu, Jianzhang; Wu, Shoubiao; Shi, Lingyi; Zhang, Shanshan; Ren, Jiye; Yao, Song; Yun, Di; Huang, Lili; Wang, Jiabing; Li, Wulan; Wu, Xiaoping; Qiu, Peihong; Liang, Guang

    2017-01-05

    The nuclear factor-kappa B (NF-κB) signaling pathway has been targeted for the therapy of various cancers, including lung cancer. EF24 was considered as a potent inhibitor of NF-κB signaling pathway. In this study, a series of asymmetric EF24 analogues were synthesized and evaluated for their anti-cancer activity against three lung cancer cell lines (A549, LLC, H1650). Most of the compounds exhibited good anti-tumor activity. Among them, compound 81 showed greater cytotoxicity than EF24. Compound 81 also possessed a potent anti-migration and anti-proliferative ability against A549 cells in a concentration-dependent manner. Moreover, compound 81 induced lung cancer cells death by inhibiting NF-κB signaling pathway, and activated the JNK-mitochondrial apoptotic pathway by increasing reactive oxygen species (ROS) generation resulting in apoptosis. In summary, compound 81 is a valuable candidate for anti-lung cancer therapy.

  13. Perifosine as a potential novel anti-cancer agent inhibits EGFR/MET-AKT axis in malignant pleural mesothelioma.

    Directory of Open Access Journals (Sweden)

    Giulia Pinton

    Full Text Available BACKGROUND: PI3K/AKT signalling pathway is aberrantly active and plays a critical role for cell cycle progression of human malignant pleural mesothelioma (MMe cells. AKT is one of the important cellular targets of perifosine, a novel bio-available alkylphospholipid that has displayed significant anti-proliferative activity in vitro and in vivo in several human tumour model systems and is currently being tested in clinical trials. METHODS: We tested Perifosine activity on human mesothelial cells and different mesothelioma cell lines, in order to provide evidence of its efficacy as single agent and combined therapy. RESULTS: We demonstrate here that perifosine, currently being evaluated as an anti-cancer agent in phase 1 and 2 clinical trials, caused a dose-dependent reduction of AKT activation, at concentrations causing MMe cell growth arrest. In this study we firstly describe that MMe cells express aside from AKT1 also AKT3 and that either the myristoylated, constitutively active, forms of the two proteins, abrogated perifosine-mediated cell growth inhibition. Moreover, we describe here a novel mechanism of perifosine that interferes, upstream of AKT, affecting EGFR and MET phosphorylation. Finally, we demonstrate a significant increase in cell toxicity when MMe cells were treated with perifosine in combination with cisplatin. CONCLUSIONS: This study provides a novel mechanism of action of perifosine, directly inhibiting EGFR/MET-AKT1/3 axis, providing a rationale for a novel translational approach to the treatment of MMe.

  14. Targeted therapy in non-small cell lung cancer

    Institute of Scientific and Technical Information of China (English)

    Shou-Ching Tang

    2004-01-01

    @@ 1 Introduction Recent progress in molecular biology has enabled us to better understand the molecular mechanism underlying pathogenesis of human malignancy including lung cancer. Sequencing of human genome has identified many oncogenes and tumor suppressor genes,giving us a better understanding of the molecular events leading to the formation, progression, metastasis, and the development of drug resistance in human lung cancer. In addition, many signal transduction pathways have been discovered that play important roles in lung cancer. Novel strategy of anti-cancer drug development now involves the identification and development of targeted therapy that interrupts one or more than one pathways or cross-talk among different signal transduction pathways. In addition, efforts are underway that combine the traditional cytotoxic (non-targeted) agents with the biological (targeted) therapy to increase the response rate and survival in patients with lung cancer, especially advanced non-small cell lung cancer (NSCLC).

  15. Identification of target genes of synovial sarcoma-associated fusion oncoprotein using human pluripotent stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Hayakawa, Kazuo [Department of Tissue Regeneration, Institute for Frontier Medical Sciences, Kyoto University, Kyoto (Japan); Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application, Kyoto University, Kyoto (Japan); Department of Orthopaedic Surgery, Graduate School of Medical Sciences, Nagoya City University, Nagoya (Japan); Ikeya, Makoto [Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application, Kyoto University, Kyoto (Japan); Fukuta, Makoto [Department of Tissue Regeneration, Institute for Frontier Medical Sciences, Kyoto University, Kyoto (Japan); Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application, Kyoto University, Kyoto (Japan); Department of Orthopaedic Surgery, Graduate School of Medical Sciences, Nagoya City University, Nagoya (Japan); Woltjen, Knut [Department of Reprogramming Sciences, Center for iPS Cell Research and Application, Kyoto University, Kyoto (Japan); Tamaki, Sakura; Takahara, Naoko; Kato, Tomohisa; Sato, Shingo [Department of Tissue Regeneration, Institute for Frontier Medical Sciences, Kyoto University, Kyoto (Japan); Otsuka, Takanobu [Department of Orthopaedic Surgery, Graduate School of Medical Sciences, Nagoya City University, Nagoya (Japan); Toguchida, Junya, E-mail: togjun@frontier.kyoto-u.ac.jp [Department of Tissue Regeneration, Institute for Frontier Medical Sciences, Kyoto University, Kyoto (Japan); Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application, Kyoto University, Kyoto (Japan); Department of Orthopaedic Surgery, Graduate School of Medicine, Kyoto University, Kyoto (Japan)

    2013-03-22

    Highlights: ► We tried to identify targets of synovial sarcoma (SS)-associated SYT–SSX fusion gene. ► We established pluripotent stem cell (PSC) lines with inducible SYT–SSX gene. ► SYT–SSX responsive genes were identified by the induction of SYT–SSX in PSC. ► SS-related genes were selected from database by in silico analyses. ► 51 genes were finally identified among SS-related genes as targets of SYT–SSX in PSC. -- Abstract: Synovial sarcoma (SS) is a malignant soft tissue tumor harboring chromosomal translocation t(X; 18)(p11.2; q11.2), which produces SS-specific fusion gene, SYT–SSX. Although precise function of SYT–SSX remains to be investigated, accumulating evidences suggest its role in gene regulation via epigenetic mechanisms, and the product of SYT–SSX target genes may serve as biomarkers of SS. Lack of knowledge about the cell-of-origin of SS, however, has placed obstacle in the way of target identification. Here we report a novel approach to identify SYT–SSX2 target genes using human pluripotent stem cells (hPSCs) containing a doxycycline-inducible SYT–SSX2 gene. SYT–SSX2 was efficiently induced both at mRNA and protein levels within three hours after doxycycline administration, while no morphological change of hPSCs was observed until 24 h. Serial microarray analyses identified genes of which the expression level changed more than twofold within 24 h. Surprisingly, the majority (297/312, 95.2%) were up-regulated genes and a result inconsistent with the current concept of SYT–SSX as a transcriptional repressor. Comparing these genes with SS-related genes which were selected by a series of in silico analyses, 49 and 2 genes were finally identified as candidates of up- and down-regulated target of SYT–SSX, respectively. Association of these genes with SYT–SSX in SS cells was confirmed by knockdown experiments. Expression profiles of SS-related genes in hPSCs and human mesenchymal stem cells (hMSCs) were strikingly

  16. Gene targeting in melanoma therapy: exploiting of surface markers and specific promoters

    Directory of Open Access Journals (Sweden)

    Sverdlov E. D.

    2012-01-01

    Full Text Available One of the problems of gene therapy of melanoma is effective expression of therapeutic gene in tumor cells and their metastases but not in normal cells. In this review, we will consider a two-step approach to a highly specific gene therapy. At the first step, therapeutic genes are delivered specifically to tumor cells using cell surface markers of melanoma cells as targets. At the second step, a specific expression of the therapeutic genes in tumor cells is ensured. Surface markers of melanoma cells were analyzed as potential targets for therapeutic treatment. Criteria for choosing the most promising targets are proposed. The use of specific melanoma promoters allows to further increase the specificity of treatment via transcriptional control of therapeutic gene expression in melanoma cells.

  17. Peptide targeting of adenoviral vectors to augment tumor gene transfer.

    Science.gov (United States)

    Ballard, E N; Trinh, V T; Hogg, R T; Gerard, R D

    2012-07-01

    Adenovirus serotype 5 remains one of the most promising vectors for delivering genetic material to cancer cells for imaging or therapy, but optimization of these agents to selectively promote tumor cell infection is needed to further their clinical development. Peptide sequences that bind to specific cell surface receptors have been inserted into adenoviral capsid proteins to improve tumor targeting, often in the background of mutations designed to ablate normal ligand:receptor interactions and thereby reduce off target effects and toxicities in non-target tissues. Different tumor types also express highly variable complements of cell surface receptors, so a customized targeting strategy using a particular peptide in the context of specific adenoviral mutations may be needed to achieve optimal efficacy. To further investigate peptide targeting strategies in adenoviral vectors, we used a set of peptide motifs originally isolated using phage display technology that evince tumor specificity in vivo. To demonstrate their abilities as targeting motifs, we genetically incorporated these peptides into a surface loop of the fiber capsid protein to construct targeted adenovirus vectors. We then systematically evaluated the ability of these peptide targeted vectors to infect several tumor cell types, both in vitro and in vivo, in a variety of mutational backgrounds designed to reduce CAR and/or HSG-mediated binding. Results from this study support previous observations that peptide insertions in the HI loop of the fiber knob domain are generally ineffective when used in combination with HSG detargeting mutations. The evidence also suggests that this strategy can attenuate other fiber knob interactions, such as CAR-mediated binding, and reduce overall viral infectivity. The insertion of peptides into fiber proved more effective for targeting tumor cell types expressing low levels of CAR receptor, as this strategy can partially compensate for the very low infectivity of wild

  18. Gene-carried hepatoma targeting complex induced high gene transfection efficiency with low toxicity and significant antitumor activity

    Directory of Open Access Journals (Sweden)

    Zhao QQ

    2012-06-01

    Full Text Available Qing-Qing Zhao,1,2 Yu-Lan Hu,1 Yang Zhou,3 Ni Li,1 Min Han,1 Gu-Ping Tang,4 Feng Qiu,2 Yasuhiko Tabata,5 Jian-Qing Gao,11Institute of Pharmaceutics, Zhejiang University, Hangzhou, China; 2Department of Pharmacy, The First Affiliated Hospital of Chongqing Medical University, Chongqing, China; 3Institute of Biochemistry, Iowa State University, Ames, IA, USA; 4Institute of Chemical Biology and Pharmaceutical Chemistry, Zhejiang University, Hangzhou, China; 5Institute for Frontier Medical Sciences, Kyoto University, Kyoto, JapanBackground: The success of gene transfection is largely dependent on the development of a vehicle or vector that can efficiently deliver a gene to cells with minimal toxicity.Methods: A liver cancer-targeted specific peptide (FQHPSF sequence was successfully synthesized and linked with chitosan-linked polyethylenimine (CP to form a new targeted gene delivery vector called CPT (CP/peptide. The structure of CPT was confirmed by 1H nuclear magnetic resonance spectroscopy and ultraviolet spectrophotometry. The particle size of CPT/DNA complexes was measured using laser diffraction spectrometry and the cytotoxicity of the copolymer was evaluated by methylthiazol tetrazolium method. The transfection efficiency evaluation of the CP copolymer was performed using luciferase activity assay. Cellular internalization of the CP/DNA complex was observed under confocal laser scanning microscopy. The targeting specificity of the polymer coupled to peptide was measured by competitive inhibition transfection study. The liver targeting specificity of the CPT copolymer in vivo was demonstrated by combining the copolymer with a therapeutic gene, interleukin-12, and assessed by its abilities in suppressing the growth of ascites tumor in mouse model.Results: The results showed that the liver cancer-targeted specific peptide was successfully synthesized and linked with CP to form a new targeted gene delivery vector called CPT. The composition of CPT

  19. Flux variability scanning based on enforced objective flux for identifying gene amplification targets

    Directory of Open Access Journals (Sweden)

    Park Jong

    2012-08-01

    Full Text Available Abstract Background In order to reduce time and efforts to develop microbial strains with better capability of producing desired bioproducts, genome-scale metabolic simulations have proven useful in identifying gene knockout and amplification targets. Constraints-based flux analysis has successfully been employed for such simulation, but is limited in its ability to properly describe the complex nature of biological systems. Gene knockout simulations are relatively straightforward to implement, simply by constraining the flux values of the target reaction to zero, but the identification of reliable gene amplification targets is rather difficult. Here, we report a new algorithm which incorporates physiological data into a model to improve the model’s prediction capabilities and to capitalize on the relationships between genes and metabolic fluxes. Results We developed an algorithm, flux variability scanning based on enforced objective flux (FVSEOF with grouping reaction (GR constraints, in an effort to identify gene amplification targets by considering reactions that co-carry flux values based on physiological omics data via “GR constraints”. This method scans changes in the variabilities of metabolic fluxes in response to an artificially enforced objective flux of product formation. The gene amplification targets predicted using this method were validated by comparing the predicted effects with the previous experimental results obtained for the production of shikimic acid and putrescine in Escherichia coli. Moreover, new gene amplification targets for further enhancing putrescine production were validated through experiments involving the overexpression of each identified targeted gene under condition-controlled batch cultivation. Conclusions FVSEOF with GR constraints allows identification of gene amplification targets for metabolic engineering of microbial strains in order to enhance the production of desired bioproducts. The algorithm

  20. Gene targeting using homologous recombination in embryonic stem cells: The future for behavior genetics?

    Directory of Open Access Journals (Sweden)

    Robert eGerlai

    2016-04-01

    Full Text Available Gene targeting with homologous recombination in embryonic stem cells created a revolution in the analysis of the function of genes in behavioral brain research. The technology allowed unprecedented precision with which one could manipulate genes and study the effect of this manipulation on the central nervous system. With gene targeting, the uncertainty inherent in psychopharmacology regarding whether a particular compound would act only through a specific target was removed. Thus, gene targeting became highly popular. However, with this popularity came the realization that like other methods, gene targeting also suffered from some technical and principal problems. For example, two decades ago, issues about compensatory changes and about genetic linkage were raised. Since then, the technology developed, and its utility has been better delineated. This review will discuss the pros and cons of the technique along with these advancements from the perspective of the neuroscientist user. It will also compare and contrast methods that may represent novel alternatives to the homologous recombination based gene targeting approach, including the TALEN and the CRISPR/Cas9 systems. The goal of the review is not to provide detailed recipes, but to attempt to present a short summary of these approaches a behavioral geneticist or neuroscientist may consider for the analysis of brain function and behavior.

  1. High efficiency TALENs enable F0 functional analysis by targeted gene disruption in Xenopus laevis embryos

    Directory of Open Access Journals (Sweden)

    Ken-ichi T. Suzuki

    2013-03-01

    Recently, gene editing with transcription activator-like effector nucleases (TALENs has been used in the life sciences. TALENs can be easily customized to recognize a specific DNA sequence and efficiently introduce double-strand breaks at the targeted genomic locus. Subsequent non-homologous end-joining repair leads to targeted gene disruption by base insertion, deletion, or both. Here, to readily evaluate the efficacy of TALENs in Xenopus laevis embryos, we performed the targeted gene disruption of tyrosinase (tyr and pax6 genes that are involved in pigmentation and eye formation, respectively. We constructed TALENs targeting tyr and pax6 and injected their mRNAs into fertilized eggs at the one-cell stage. Expectedly, introduction of tyr TALEN mRNA resulted in drastic loss of pigmentation with high efficiency. Similarly, for pax6, TALENs led to deformed eyes in the injected embryos. We confirmed mutations of the target alleles by restriction enzyme digestion and sequence analyses of genomic PCR products. Surprisingly, not only biallelic but also paralogous, gene disruption was observed. Our results demonstrate that targeted gene disruption by TALENs provides a method comparable to antisense morpholinos in analyzing gene function in Xenopus F0 embryos, but also applies beyond embryogenesis to any life stage.

  2. Stable gene replacement in barley by targeted double-strand break induction.

    Science.gov (United States)

    Watanabe, Koichi; Breier, Ulrike; Hensel, Götz; Kumlehn, Jochen; Schubert, Ingo; Reiss, Bernd

    2016-03-01

    Gene targeting is becoming an important tool for precision genome engineering in plants. During gene replacement, a variant of gene targeting, transformed DNA integrates into the genome by homologous recombination (HR) to replace resident sequences. We have analysed gene targeting in barley (Hordeum vulgare) using a model system based on double-strand break (DSB) induction by the meganuclease I-SceI and a transgenic, artificial target locus. In the plants we obtained, the donor construct was inserted at the target locus by homology-directed DNA integration in at least two transformants obtained in a single experiment and was stably inherited as a single Mendelian trait. Both events were produced by one-sided integration. Our data suggest that gene replacement can be achieved in barley with a frequency suitable for routine application. The use of a codon-optimized nuclease and co-transfer of the nuclease gene together with the donor construct are probably the components important for efficient gene targeting. Such an approach, employing the recently developed synthetic nucleases/nickases that allow DSB induction at almost any sequence of a genome of interest, sets the stage for precision genome engineering as a routine tool even for important crops such as barley.

  3. Gene Targeting Using Homologous Recombination in Embryonic Stem Cells: The Future for Behavior Genetics?

    Science.gov (United States)

    Gerlai, Robert

    2016-01-01

    Gene targeting with homologous recombination in embryonic stem cells created a revolution in the analysis of the function of genes in behavioral brain research. The technology allowed unprecedented precision with which one could manipulate genes and study the effect of this manipulation on the central nervous system. With gene targeting, the uncertainty inherent in psychopharmacology regarding whether a particular compound would act only through a specific target was removed. Thus, gene targeting became highly popular. However, with this popularity came the realization that like other methods, gene targeting also suffered from some technical and principal problems. For example, two decades ago, issues about compensatory changes and about genetic linkage were raised. Since then, the technology developed, and its utility has been better delineated. This review will discuss the pros and cons of the technique along with these advancements from the perspective of the neuroscientist user. It will also compare and contrast methods that may represent novel alternatives to the homologous recombination based gene targeting approach, including the TALEN and the CRISPR/Cas9 systems. The goal of the review is not to provide detailed recipes, but to attempt to present a short summary of these approaches a behavioral geneticist or neuroscientist may consider for the analysis of brain function and behavior.

  4. Target gene analyses of 39 amelogenesis imperfecta kindreds.

    Science.gov (United States)

    Chan, Hui-Chen; Estrella, Ninna M R P; Milkovich, Rachel N; Kim, Jung-Wook; Simmer, James P; Hu, Jan C-C

    2011-12-01

    Previously, mutational analyses identified six disease-causing mutations in 24 amelogenesis imperfecta (AI) kindreds. We have since expanded the number of AI kindreds to 39, and performed mutation analyses covering the coding exons and adjoining intron sequences for the six proven AI candidate genes [amelogenin (AMELX), enamelin (ENAM), family with sequence similarity 83, member H (FAM83H), WD repeat containing domain 72 (WDR72), enamelysin (MMP20), and kallikrein-related peptidase 4 (KLK4)] and for ameloblastin (AMBN) (a suspected candidate gene). All four of the X-linked AI families (100%) had disease-causing mutations in AMELX, suggesting that AMELX is the only gene involved in the aetiology of X-linked AI. Eighteen families showed an autosomal-dominant pattern of inheritance. Disease-causing mutations were identified in 12 (67%): eight in FAM83H, and four in ENAM. No FAM83H coding-region or splice-junction mutations were identified in three probands with autosomal-dominant hypocalcification AI (ADHCAI), suggesting that a second gene may contribute to the aetiology of ADHCAI. Six families showed an autosomal-recessive pattern of inheritance, and disease-causing mutations were identified in three (50%): two in MMP20, and one in WDR72. No disease-causing mutations were found in 11 families with only one affected member. We conclude that mutation analyses of the current candidate genes for AI have about a 50% chance of identifying the disease-causing mutation in a given kindred.

  5. Characterization of three loci for homologous gene targeting and transgene expression.

    Science.gov (United States)

    Eyquem, Justin; Poirot, Laurent; Galetto, Roman; Scharenberg, Andrew M; Smith, Julianne

    2013-08-01

    Integrative gene transfer is widely used for bioproduction, drug screening, and therapeutic applications but usual viral methods lead to random and multicopy insertions, contribute to unstable transgene expression and can disturb endogenous gene expression. Homologous targeting of an expression cassette using rare-cutting endonucleases is a potential solution; however the number of studied loci remains limited. Furthermore, the behavior and performance of various types of gene cassettes following gene targeting is poorly defined. Here we have evaluated three loci for gene targeting, including one locus compatible with the proposed Safe Harbor criteria for human translational applications. Using optimized conditions for homologous gene targeting, reporter genes under the control of different promoters were efficiently inserted at each locus in both sense and antisense orientations. Sustainable expression was achieved at all three loci without detectable disturbance of flanking gene expression. However, the promoter, the integration locus and the cassette orientation have a strong impact on transgene expression. Finally, single targeted integrations exhibited greatly improved transgene expression stability versus multicopy or random integration. Taken together, our data suggest a potential set of loci for site-specific transgene integration, suitable for a variety of biotechnological applications.

  6. Apoptosis and the target genes of microRNA-21

    Institute of Scientific and Technical Information of China (English)

    Lindsey E. Becker Buscaglia; Yong Li

    2011-01-01

    MicroRNA-21 (miR-21) is frequently up-regulated in cancer and the majodty of its reported targets are tumor suppressors. Through functional suppression, miR-21 is implicated in practically every walk of oncogenic life: the promotion of cell proliferation, invasion and metastasis, genome instability and mutation, inflammation, replicative immortalization, abnormal metabolism, angiogenesis, and evading apoptosis, immune destruction, and growth suppressors. In particular, miR-21 is strongly involved in apoptosis. In this article, we reviewed the experimentally validated targets of miR-21 and found that two thirds are linked to intrinsic and/or extrinsic pathways of cellular apoptosis. This suggests that miR-21 is an oncogene which plays a key role in resisting programmed cell death in cancer cells and that targeting apoptosis is a viable therapeutic option against cancers expressing miR-21.

  7. Identification of Multiple Cryptococcal Fungicidal Drug Targets by Combined Gene Dosing and Drug Affinity Responsive Target Stability Screening

    Directory of Open Access Journals (Sweden)

    Yoon-Dong Park

    2016-08-01

    Full Text Available Cryptococcus neoformans is a pathogenic fungus that is responsible for up to half a million cases of meningitis globally, especially in immunocompromised individuals. Common fungistatic drugs, such as fluconazole, are less toxic for patients but have low efficacy for initial therapy of the disease. Effective therapy against the disease is provided by the fungicidal drug amphotericin B; however, due to its high toxicity and the difficulty in administering its intravenous formulation, it is imperative to find new therapies targeting the fungus. The antiparasitic drug bithionol has been recently identified as having potent fungicidal activity. In this study, we used a combined gene dosing and drug affinity responsive target stability (GD-DARTS screen as well as protein modeling to identify a common drug binding site of bithionol within multiple NAD-dependent dehydrogenase drug targets. This combination genetic and proteomic method thus provides a powerful method for identifying novel fungicidal drug targets for further development.

  8. Evaluation of anti-oxidant and anti-cancer properties of Dendropanax morbifera Léveille.

    Science.gov (United States)

    Hyun, Tae Kyung; Kim, Myeong-ok; Lee, Hyunkyoung; Kim, Younjoo; Kim, Euikyung; Kim, Ju-Sung

    2013-12-01

    Dendropanax morbifera Léveille, an endemic species in Korea, is best known as a tree that produces a resinous sap. Although D. morbifera is used in folk medicine, its biological activities are poorly understood. In this study, the methanolic extracts of D. morbifera branches, debarked stems, bark, and two different stages of leaves were evaluated for anti-oxidant activity and anti-cancer potential. The debarked stem extract exhibited strong 1,1-diphenyl-2-picrylhydrazyl (DPPH) scavenging activity and reducing power compared with other samples. In addition, the cytotoxic activities of these extracts were investigated in human tumour cell lines. The results suggested that the extracts of debarked stems, green leaves, and yellow leaves were the potent source of anti-cancer compounds, particularly in Huh-7 cells. Furthermore, treatment with the extracts of debarked stems, green leaves, and yellow leaves caused an increase of apoptotic or senescent cells in Huh-7 cells. Twenty-four hour treatment with debarked stems extract resulted in the strong induction of p53 and p16, whereas both leaf extracts inhibited the activation of ERK. The debarked stems and green leaf extracts reduced Akt levels in Huh-7 cells, indicating that D. morbifera extracts caused the activation of p16 and p53 pathways. This, together with the inhibition of Akt or ERK signalling, resulted in suppression of Huh-7 cell proliferation. These results suggest that methanolic leaf and debarked stems extracts are a source of anti-oxidant and anti-cancer compounds, and could be developed as a botanical drug.

  9. Study of Malformin C, a Fungal Source Cyclic Pentapeptide, as an Anti-Cancer Drug.

    Directory of Open Access Journals (Sweden)

    Jing Wang

    Full Text Available Malformin C, a fungal cyclic pentapeptide, has been claimed to have anti-cancer potential, but no in vivo study was available to substantiate this property. Therefore, we conducted in vitro and in vivo experiments to investigate its anti-cancer effects and toxicity. Our studies showed Malformin C inhibited Colon 38 and HCT 116 cell growth dose-dependently with an IC50 of 0.27±0.07μM and 0.18±0.023μM respectively. This inhibition was explicated by Malformin C's effect on G2/M arrest. Moreover, we observed up-regulated expression of phospho-histone H2A.X, p53, cleaved CASPASE 3 and LC3 after Malformin C treatment, while the apoptosis assay indicated an increased population of necrotic and late apoptotic cells. In vivo, the pathological study exhibited the acute toxicity of Malformin C at lethal dosage in BDF1 mice might be caused by an acute yet subtle inflammatory response, consistent with elevated IL-6 in the plasma cytokine assay. Further anti-tumor and toxicity experiments proved that 0.3mg/kg injected weekly was the best therapeutic dosage of Malformin C in Colon 38 xenografted BDF1 mice, whereas 0.1mg/kg every other day showed no effect with higher resistance, and 0.9mg/kg per week either led to fatal toxicity in seven-week old mice or displayed no advantage over 0.3mg/kg group in nine-week old mice. Overall, we conclude that Malformin C arrests Colon 38 cells in G2/M phase and induces multiple forms of cell death through necrosis, apoptosis and autophagy. Malformin C has potent cell growth inhibition activity, but the therapeutic index is too low to be an anti-cancer drug.

  10. New development and application of ultrasound targeted microbubble destruction in gene therapy and drug delivery.

    Science.gov (United States)

    Chen, Zhi-Yi; Yang, Feng; Lin, Yan; Zhang, Jin-Shan; Qiu, Ri-Xiang; Jiang, Lan; Zhou, Xing-Xing; Yu, Jiang-Xiu

    2013-08-01

    Ultrasound is a common used technique for clinical imaging. In recent years, with the advances in preparation technology of microbubbles and the innovations in ultrasound imaging, ultrasound is no longer confined to detection of tissue perfusion, but extends to specific ultrasound molecular imaging and target therapy gradually. With the development of research, ultrasound molecular imaging and target therapy have made great progresses. Targeted microbubbles for molecular imaging are achieved by binding target molecules, specific antibody or ligand to the surface of microbubbles to obtain specific imaging by attaching to target tissues. Meanwhile, it can also achieve targeting gene therapy or drug delivery by ultrasound targeted microbubble destruction (UTMD) mediating genes or drugs to specific target sites. UTMD has a number of advantages, such as target-specific, highly effective, non-invasivity, relatively low-cost and no radiation, and has broad application prospects, which is regarded as one hot spot in medical studies. We reviewed the new development and application of UTMD in gene therapy and drug delivery in this paper. With further development of technology and research, the gene or drug delivery system and related methods will be widely used in application and researches.

  11. RNA-guided genome editing for target gene mutations in wheat.

    Science.gov (United States)

    Upadhyay, Santosh Kumar; Kumar, Jitesh; Alok, Anshu; Tuli, Rakesh

    2013-12-09

    The clustered, regularly interspaced, short palindromic repeats (CRISPR) and CRISPR-associated protein (Cas) system has been used as an efficient tool for genome editing. We report the application of CRISPR-Cas-mediated genome editing to wheat (Triticum aestivum), the most important food crop plant with a very large and complex genome. The mutations were targeted in the inositol oxygenase (inox) and phytoene desaturase (pds) genes using cell suspension culture of wheat and in the pds gene in leaves of Nicotiana benthamiana. The expression of chimeric guide RNAs (cgRNA) targeting single and multiple sites resulted in indel mutations in all the tested samples. The expression of Cas9 or sgRNA alone did not cause any mutation. The expression of duplex cgRNA with Cas9 targeting two sites in the same gene resulted in deletion of DNA fragment between the targeted sequences. Multiplexing the cgRNA could target two genes at one time. Target specificity analysis of cgRNA showed that mismatches at the 3' end of the target site abolished the cleavage activity completely. The mismatches at the 5' end reduced cleavage, suggesting that the off target effects can be abolished in vivo by selecting target sites with unique sequences at 3' end. This approach provides a powerful method for genome engineering in plants.

  12. Restricted mobility of specific functional groups reduces anti-cancer drug activity in healthy cells

    DEFF Research Database (Denmark)

    Longo Martins, Murillo; Ignazzi, Rosanna; Eckert, Juergen

    2016-01-01

    The most common cancer treatments currently available are radio- and chemo-therapy. These therapies have, however, drawbacks, such as, the reduction in quality of life and the low efficiency of radiotherapy in cases of multiple metastases. To lessen these effects, we have encapsulated an anti-cancer...... drug into a biocompatible matrix. In-vitro assays indicate that this bio-nanocomposite is able to interact and cause morphological changes in cancer cells. Meanwhile, no alterations were observed in monocytes and fibroblasts, indicating that this system might carry the drug in living organisms...

  13. Gene-Specific Demethylation as Targeted Therapy in MDS

    Science.gov (United States)

    2016-07-01

    selected based on the prediction that they 6 would form a triplex structure with the locus being targeted. The second approach utilized a CRISPR ...MsgRNA. This type of screening should allow us to select the most effective and strongest guides to utilize with the inducible Crispr /dCas9 system and

  14. Peroxisome Proliferator-Activated Receptor Alpha Target Genes

    NARCIS (Netherlands)

    Rakhshandehroo, M.; Knoch, B.; Müller, M.R.; Kersten, A.H.

    2010-01-01

    The peroxisome proliferator-activated receptor alpha (PPAR alpha) is a ligand-activated transcription factor involved in the regulation of a variety of processes, ranging from inflammation and immunity to nutrient metabolism and energy homeostasis. PPAR alpha serves as a molecular target for hypolip

  15. Systems Pharmacology‐Based Discovery of Natural Products for Precision Oncology Through Targeting Cancer Mutated Genes

    Science.gov (United States)

    Fang, J; Cai, C; Wang, Q; Lin, P

    2017-01-01

    Massive cancer genomics data have facilitated the rapid revolution of a novel oncology drug discovery paradigm through targeting clinically relevant driver genes or mutations for the development of precision oncology. Natural products with polypharmacological profiles have been demonstrated as promising agents for the development of novel cancer therapies. In this study, we developed an integrated systems pharmacology framework that facilitated identifying potential natural products that target mutated genes across 15 cancer types or subtypes in the realm of precision medicine. High performance was achieved for our systems pharmacology framework. In case studies, we computationally identified novel anticancer indications for several US Food and Drug Administration‐approved or clinically investigational natural products (e.g., resveratrol, quercetin, genistein, and fisetin) through targeting significantly mutated genes in multiple cancer types. In summary, this study provides a powerful tool for the development of molecularly targeted cancer therapies through targeting the clinically actionable alterations by exploiting the systems pharmacology of natural products. PMID:28294568

  16. Efficient four fragment cloning for the construction of vectors for targeted gene replacement in filamentous fungi

    DEFF Research Database (Denmark)

    Frandsen, Rasmus John Normand; Andersson, Jens A.; Kristensen, Matilde Bylov;

    2008-01-01

    of coding sequences with fluorescent markers such as GFP are essential for this process. Construction of vectors for these experiments depends on the directional cloning of two homologous recombination sequences on each side of a selection marker gene. Results: Here, we present a USER Friendly cloning based...... tumefaciens and protoplast based transformation technologies. The system has been tested by the construction of vectors for targeted replacement of 17 genes and overexpression of 12 genes in Fusarium graminearum. The results show that four fragment vectors can be constructed in a single cloning step...... with an average efficiency of 84% for gene replacement and 80% for targeted overexpression. Conclusion: The new vectors designed for USER Friendly cloning provided a fast reliable method to construct vectors for targeted gene manipulations in fungi....

  17. Gene targeting in embryonic stem cells, II: conditional technologies

    Science.gov (United States)

    Genome modification via transgenesis has allowed researchers to link genotype and phenotype as an alternative approach to the characterization of random mutations through evolution. The synergy of technologies from the fields of embryonic stem (ES) cells, gene knockouts, and protein-mediated recombi...

  18. Parallel evolution of domesticated Caenorhabditis species targets pheromone receptor genes.

    Science.gov (United States)

    McGrath, Patrick T; Xu, Yifan; Ailion, Michael; Garrison, Jennifer L; Butcher, Rebecca A; Bargmann, Cornelia I

    2011-08-17

    Evolution can follow predictable genetic trajectories, indicating that discrete environmental shifts can select for reproducible genetic changes. Conspecific individuals are an important feature of an animal's environment, and a potential source of selective pressures. Here we show that adaptation of two Caenorhabditis species to growth at high density, a feature common to domestic environments, occurs by reproducible genetic changes to pheromone receptor genes. Chemical communication through pheromones that accumulate during high-density growth causes young nematode larvae to enter the long-lived but non-reproductive dauer stage. Two strains of Caenorhabditis elegans grown at high density have independently acquired multigenic resistance to pheromone-induced dauer formation. In each strain, resistance to the pheromone ascaroside C3 results from a deletion that disrupts the adjacent chemoreceptor genes serpentine receptor class g (srg)-36 and -37. Through misexpression experiments, we show that these genes encode redundant G-protein-coupled receptors for ascaroside C3. Multigenic resistance to dauer formation has also arisen in high-density cultures of a different nematode species, Caenorhabditis briggsae, resulting in part from deletion of an srg gene paralogous to srg-36 and srg-37. These results demonstrate rapid remodelling of the chemoreceptor repertoire as an adaptation to specific environments, and indicate that parallel changes to a common genetic substrate can affect life-history traits across species.

  19. Molecular Subtyping of Primary Prostate Cancer Reveals Specific and Shared Target Genes of Different ETS Rearrangements

    Directory of Open Access Journals (Sweden)

    Paula Paulo

    2012-07-01

    Full Text Available This work aimed to evaluate whether ETS transcription factors frequently involved in rearrangements in prostate carcinomas (PCa, namely ERG and ETV1, regulate specific or shared target genes. We performed differential expression analysis on nine normal prostate tissues and 50 PCa enriched for different ETS rearrangements using exon-level expression microarrays, followed by in vitro validation using cell line models. We found specific deregulation of 57 genes in ERG-positive PCa and 15 genes in ETV1-positive PCa, whereas deregulation of 27 genes was shared in both tumor subtypes. We further showed that the expression of seven tumor-associated ERG target genes (PLA1A, CACNA1D, ATP8A2, HLA-DMB, PDE3B, TDRD1, and TMBIM1 and two tumor-associated ETV1 target genes (FKBP10 and GLYATL2 was significantly affected by specific ETS silencing in VCaP and LNCaP cell line models, respectively, whereas the expression of three candidate ERG and ETV1 shared targets (GRPR, KCNH8, and TMEM45B was significantly affected by silencing of either ETS. Interestingly, we demonstrate that the expression of TDRD1, the topmost overexpressed gene of our list of ERG-specific candidate targets, is inversely correlated with the methylation levels of a CpG island found at -66 bp of the transcription start site in PCa and that TDRD1 expression is regulated by direct binding of ERG to the CpG island in VCaP cells. We conclude that ETS transcription factors regulate specific and shared target genes and that TDRD1, FKBP10, and GRPR are promising therapeutic targets and can serve as diagnostic markers for molecular subtypes of PCa harboring specific fusion gene rearrangements.

  20. Synthesis and evaluation of multi-wall carbon nanotube–paclitaxel complex as an anti-cancer agent

    Science.gov (United States)

    Ghasemvand, Fariba; Biazar, Esmaeil; Tavakolifard, Sara; Khaledian, Mohammad; Rahmanzadeh, Saeid; Momenzadeh, Daruosh; Afroosheh, Roshanak; Zarkalami, Faezeh; Shabannezhad, Marjan; Hesami Tackallou, Saeed; Massoudi, Nilofar; Heidari Keshel, Saeed

    2016-01-01

    Aim: The aim of this study was to design multi-walled carbon nanotubes (MWCNTs) loaded with paclitaxel (PTX) anti-cancer drug and investigate its anti-cancerous efficacy of human gastric cancer. Background: Carbon nanotubes (CNTs) represent a novel nano-materials applied in various fields such as drug delivery due to their unique chemical properties and high drug loading. Patients and methods: In this study, multi-walled carbon nanotubes (MWCNTs) pre-functionalized covalently with a paclitaxel (PTX) as an anti-cancer drug and evaluated by different analyses including, scanning electron microscope (SEM), particle size analyzer and cellular analyses. Results: A well conjugated of anti-cancer drug on the carbon nanotube surfaces was shown. This study demonstrates that the MWCN-PTX complex is a potentially useful system for delivery of anti-cancer drugs. The flow cytometry, CFU and MTT assay results have disclosed that MWCNT/PTXs might promote apoptosis in MKN-45 gastric adenocarcinoma cell line. Conclusion: According to results, our simple method can be designed a candidate material for chemotherapy. It has presented a few bio-related applications including, their successful use as a nano-carriers for drug transport. PMID:27458512

  1. Bacteria as vectors for gene therapy of cancer.

    LENUS (Irish Health Repository)

    Baban, Chwanrow K

    2012-01-31

    Anti-cancer therapy faces major challenges, particularly in terms of specificity of treatment. The ideal therapy would eradicate tumor cells selectively with minimum side effects on normal tissue. Gene or cell therapies have emerged as realistic prospects for the treatment of cancer, and involve the delivery of genetic information to a tumor to facilitate the production of therapeutic proteins. However, there is still much to be done before an efficient and safe gene medicine is achieved, primarily developing the means of targeting genes to tumors safely and efficiently. An emerging family of vectors involves bacteria of various genera. It has been shown that bacteria are naturally capable of homing to tumors when systemically administered resulting in high levels of replication locally. Furthermore, invasive species can deliver heterologous genes intra-cellularly for tumor cell expression. Here, we review the use of bacteria as vehicles for gene therapy of cancer, detailing the mechanisms of action and successes at preclinical and clinical levels.

  2. Identification of conserved microRNAs and their target genes in tomato (Lycopersicon esculentum).

    Science.gov (United States)

    Yin, Zujun; Li, Chunhe; Han, Xiulan; Shen, Fafu

    2008-05-15

    MicroRNAs (miRNAs) are a class of non-coding RNAs that have important gene regulation roles in various organisms. To date, a total of 1279 plant miRNAs have been deposited in the miRNA miRBase database (Release 10.1). Many of them are conserved during the evolution of land plants suggesting that the well-conserved miRNAs may also retain homologous target interactions. Recently, little is known about the experimental or computational identification of conserved miRNAs and their target genes in tomato. Here, using a computational homology search approach, 21 conserved miRNAs were detected in the Expressed Sequence Tags (EST) and Genomic Survey Sequence (GSS) databases. Following this, 57 potential target genes were predicted by searching the mRNA database. Most of the target mRNAs appeared to be involved in plant growth and development. Our findings verified that the well-conserved tomato miRNAs have retained homologous target interactions amongst divergent plant species. Some miRNAs express diverse combinations in different cell types and have been shown to regulate cell-specific target genes coordinately. We believe that the targeting propensity for genes in different biological processes can be explained largely by their protein connectivity.

  3. Establishment of Smad2 conditional gene targeting mice based on the Cre-LoxP system

    Institute of Scientific and Technical Information of China (English)

    ZHOU; Jiang(周江); CHENG; Xuan(程萱); SUN; Yanxun(孙彦洵); HUANG; Peitang(黄培堂); HUANG; Cuifen(黄翠芬); YANG; Xiao(杨晓)

    2002-01-01

    Smads is a new gene family in transforming growth factor-β(TGF-β) signaling pathway. Smad2 mutated in multiple human tumors and may be a candidate tumor suppressor gene. Targeted disruption of murine Smad2 gene resulted in embryonic lethality at E6.5. To study the function of Smad2 in vertebrate organgenesis and tumorigenesis, we constructed the Smad2 conditional targeting vector in which two LoxP sequences were placed to flank the sequences encoding the C terminal functional domain of Smad2. The validity of the LoxP sites in the targeting construct was tested in E. coli that express the Cre recombinase constitutively. The vector was electroporated into ES cells and 3 targeted ES cell clones were obtained by Southern blot screening. Targeted ES cells were introduced into C57BL/6J blastocysts by microinjection to generate germ-line chimeras. Genotyping analysis showed that 2 progeny among these chimeras carried the Smad2 conditional targeted allele. The establishment of Smad2 conditional gene targeting mouse has laid a solid foundation for producing the tissue specific Smad2 gene knockout mice.

  4. Identification of novel endogenous antisense transcripts by DNA microarray analysis targeting complementary strand of annotated genes

    Directory of Open Access Journals (Sweden)

    Kohama Chihiro

    2009-08-01

    Full Text Available Abstract Background Recent transcriptomic analyses in mammals have uncovered the widespread occurrence of endogenous antisense transcripts, termed natural antisense transcripts (NATs. NATs are transcribed from the opposite strand of the gene locus and are thought to control sense gene expression, but the mechanism of such regulation is as yet unknown. Although several thousand potential sense-antisense pairs have been identified in mammals, examples of functionally characterized NATs remain limited. To identify NAT candidates suitable for further functional analyses, we performed DNA microarray-based NAT screening using mouse adult normal tissues and mammary tumors to target not only the sense orientation but also the complementary strand of the annotated genes. Results First, we designed microarray probes to target the complementary strand of genes for which an antisense counterpart had been identified only in human public cDNA sources, but not in the mouse. We observed a prominent expression signal from 66.1% of 635 target genes, and 58 genes of these showed tissue-specific expression. Expression analyses of selected examples (Acaa1b and Aard confirmed their dynamic transcription in vivo. Although interspecies conservation of NAT expression was previously investigated by the presence of cDNA sources in both species, our results suggest that there are more examples of human-mouse conserved NATs that could not be identified by cDNA sources. We also designed probes to target the complementary strand of well-characterized genes, including oncogenes, and compared the expression of these genes between mammary cancerous tissues and non-pathological tissues. We found that antisense expression of 95 genes of 404 well-annotated genes was markedly altered in tumor tissue compared with that in normal tissue and that 19 of these genes also exhibited changes in sense gene expression. These results highlight the importance of NAT expression in the regulation

  5. Cancer-targeted BikDD gene therapy elicits protective antitumor immunity against lung cancer.

    Science.gov (United States)

    Sher, Yuh-Pyng; Liu, Shih-Jen; Chang, Chun-Mien; Lien, Shu-Pei; Chen, Chien-Hua; Han, Zhenbo; Li, Long-Yuan; Chen, Jin-Shing; Wu, Cheng-Wen; Hung, Mien-Chie

    2011-04-01

    Targeted cancer-specific gene therapy is a promising strategy for treating metastatic lung cancer, which is a leading cause of lung cancer-related deaths. Previously, we developed a cancer-targeted gene therapy expression system with high tumor specificity and strong activity that selectively induced lung cancer cell killing without affecting normal cells in immunocompromised mice. Here, we found this cancer-targeted gene therapy, SV-BikDD, composed of the survivin promoter in the VP16-GAL4-WPRE integrated systemic amplifier system to drive the apoptotic gene BikDD, not only caused cytotoxic effects in cancer cells but also elicited a cancer-specific cytotoxic T lymphocyte response to synergistically increase the therapeutic effect and further develop an effective systemic antitumoral immunity against rechallenges of tumorigenic dose of parental tumor cells inoculated at distant sites in immunocompetent mice. In addition, this cancer-targeted gene therapy does not elicit an immune response against normal tissues, but CMV-BikDD treatment does. The therapeutic vector could also induce proinflammatory cytokines to activate innate immunity and provide some benefits in antitumor gene therapy. Thus, this study provides a promising strategy with benefit of antitumoral immune response worthy of further development in clinical trials for treating lung cancer via cancer-targeted gene therapy.

  6. Gene Regulatory Mechanisms Underlying the Spatial and Temporal Regulation of Target-Dependent Gene Expression in Drosophila Neurons.

    Directory of Open Access Journals (Sweden)

    Anthony J E Berndt

    2015-12-01

    Full Text Available Neuronal differentiation often requires target-derived signals from the cells they innervate. These signals typically activate neural subtype-specific genes, but the gene regulatory mechanisms remain largely unknown. Highly restricted expression of the FMRFa neuropeptide in Drosophila Tv4 neurons requires target-derived BMP signaling and a transcription factor code that includes Apterous. Using integrase transgenesis of enhancer reporters, we functionally dissected the Tv4-enhancer of FMRFa within its native cellular context. We identified two essential but discrete cis-elements, a BMP-response element (BMP-RE that binds BMP-activated pMad, and a homeodomain-response element (HD-RE that binds Apterous. These cis-elements have low activity and must be combined for Tv4-enhancer activity. Such combinatorial activity is often a mechanism for restricting expression to the intersection of cis-element spatiotemporal activities. However, concatemers of the HD-RE and BMP-RE cis-elements were found to independently generate the same spatiotemporal expression as the Tv4-enhancer. Thus, the Tv4-enhancer atypically combines two low-activity cis-elements that confer the same output from distinct inputs. The activation of target-dependent genes is assumed to 'wait' for target contact. We tested this directly, and unexpectedly found that premature BMP activity could not induce early FMRFa expression; also, we show that the BMP-insensitive HD-RE cis-element is activated at the time of target contact. This led us to uncover a role for the nuclear receptor, seven up (svp, as a repressor of FMRFa induction prior to target contact. Svp is normally downregulated immediately prior to target contact, and we found that maintaining Svp expression prevents cis-element activation, whereas reducing svp gene dosage prematurely activates cis-element activity. We conclude that the target-dependent FMRFa gene is repressed prior to target contact, and that target-derived BMP

  7. Expression of RNA-interference/antisense transgenes by the cognate promoters of target genes is a better gene-silencing strategy to study gene functions in rice.

    Science.gov (United States)

    Li, Jing; Jiang, Dagang; Zhou, Hai; Li, Feng; Yang, Jiawei; Hong, Laifa; Fu, Xiao; Li, Zhibin; Liu, Zhenlan; Li, Jianming; Zhuang, Chuxiong

    2011-03-03

    Antisense and RNA interference (RNAi)-mediated gene silencing systems are powerful reverse genetic methods for studying gene function. Most RNAi and antisense experiments used constitutive promoters to drive the expression of RNAi/antisense transgenes; however, several reports showed that constitutive promoters were not expressed in all cell types in cereal plants, suggesting that the constitutive promoter systems are not effective for silencing gene expression in certain tissues/organs. To develop an alternative method that complements the constitutive promoter systems, we constructed RNAi and/or antisense transgenes for four rice genes using a constitutive promoter or a cognate promoter of a selected rice target gene and generated many independent transgenic lines. Genetic, molecular, and phenotypic analyses of these RNAi/antisense transgenic rice plants, in comparison to previously-reported transgenic lines that silenced similar genes, revealed that expression of the cognate promoter-driven RNAi/antisense transgenes resulted in novel growth/developmental defects that were not observed in transgenic lines expressing constitutive promoter-driven gene-silencing transgenes of the same target genes. Our results strongly suggested that expression of RNAi/antisense transgenes by cognate promoters of target genes is a better gene-silencing approach to discovery gene function in rice.

  8. Expression of RNA-interference/antisense transgenes by the cognate promoters of target genes is a better gene-silencing strategy to study gene functions in rice.

    Directory of Open Access Journals (Sweden)

    Jing Li

    Full Text Available Antisense and RNA interference (RNAi-mediated gene silencing systems are powerful reverse genetic methods for studying gene function. Most RNAi and antisense experiments used constitutive promoters to drive the expression of RNAi/antisense transgenes; however, several reports showed that constitutive promoters were not expressed in all cell types in cereal plants, suggesting that the constitutive promoter systems are not effective for silencing gene expression in certain tissues/organs. To develop an alternative method that complements the constitutive promoter systems, we constructed RNAi and/or antisense transgenes for four rice genes using a constitutive promoter or a cognate promoter of a selected rice target gene and generated many independent transgenic lines. Genetic, molecular, and phenotypic analyses of these RNAi/antisense transgenic rice plants, in comparison to previously-reported transgenic lines that silenced similar genes, revealed that expression of the cognate promoter-driven RNAi/antisense transgenes resulted in novel growth/developmental defects that were not observed in transgenic lines expressing constitutive promoter-driven gene-silencing transgenes of the same target genes. Our results strongly suggested that expression of RNAi/antisense transgenes by cognate promoters of target genes is a better gene-silencing approach to discovery gene function in rice.

  9. Peroxisome Proliferator-Activated Receptor Alpha Target Genes

    OpenAIRE

    Maryam Rakhshandehroo; Bianca Knoch; Michael Müller; Sander Kersten

    2010-01-01

    The peroxisome proliferator-activated receptor alpha (PPAR alpha) is a ligand-activated transcription factor involved in the regulation of a variety of processes, ranging from inflammation and immunity to nutrient metabolism and energy homeostasis. PPAR alpha serves as a molecular target for hypolipidemic fibrates drugs which bind the receptor with high affinity. Furthermore, PPAR alpha binds and is activated by numerous fatty acids and fatty acid-derived compounds. PPAR alpha governs biologi...

  10. Expression of androgen receptor target genes in skeletal muscle

    OpenAIRE

    2014-01-01

    We aimed to determine the mechanisms of the anabolic actions of androgens in skeletal muscle by investigating potential androgen receptor (AR)-regulated genes in in vitro and in vivo models. The expression of the myogenic regulatory factor myogenin was significantly decreased in skeletal muscle from testosterone-treated orchidectomized male mice compared to control orchidectomized males, and was increased in muscle from male AR knockout mice that lacked DNA binding activity (ARΔZF2 ) versus w...

  11. Genome-wide identification of structural variants in genes encoding drug targets

    DEFF Research Database (Denmark)

    Rasmussen, Henrik Berg; Dahmcke, Christina Mackeprang

    2012-01-01

    The objective of the present study was to identify structural variants of drug target-encoding genes on a genome-wide scale. We also aimed at identifying drugs that are potentially amenable for individualization of treatments based on knowledge about structural variation in the genes encoding the...

  12. RYP1 gene as a target for molecular diagnosis of histoplasmosis.

    Science.gov (United States)

    Brilhante, Raimunda Sâmia Nogueira; Guedes, Glaucia Morgana de Melo; Riello, Giovanna Barbosa; Ribeiro, Joyce Fonteles; Alencar, Lucas Pereira; Bandeira, Silviane Praciano; Castelo-Branco, Débora Souza Collares Maia; Oliveira, Jonathas Sales; Freire, Janaína Maria Maia; Mesquita, Jacó Ricarte Lima de; Camargo, Zoilo Pires de; Cordeiro, Rossana de Aguiar; Rocha, Marcos Fábio Gadelha; Sidrim, José Júlio Costa

    2016-11-01

    This study analyzed the RYP1 gene as a target for the molecular diagnosis of histoplasmosis. This assay detected fungal DNA in 13/13 blood samples from HIV/AIDS-patients with histoplasmosis. Therefore, the detection of RYP1 gene in whole blood sample is a quick and sensitive test to diagnose histoplasmosis.

  13. Therapeutic brain cancer targeting by gene therapy and immunomodulation : a translational study

    NARCIS (Netherlands)

    Stathopoulos, A.

    2012-01-01

    The hypothesis pertinent to this thesis is that glioma tumours can be therapeutically targeted by gene and/or immunotherapy in order to eliminate or delay tumour recurrence leading to significant morbidity and mortality. In our gene therapeutic approach, described in Chapter 2, we observed that chro

  14. Demystifying the secret mission of enhancers: linking distal regulatory elements to target genes.

    Science.gov (United States)

    Yao, Lijing; Berman, Benjamin P; Farnham, Peggy J

    2015-01-01

    Enhancers are short regulatory sequences bound by sequence-specific transcription factors and play a major role in the spatiotemporal specificity of gene expression patterns in development and disease. While it is now possible to identify enhancer regions genomewide in both cultured cells and primary tissues using epigenomic approaches, it has been more challenging to develop methods to understand the function of individual enhancers because enhancers are located far from the gene(s) that they regulate. However, it is essential to identify target genes of enhancers not only so that we can understand the role of enhancers in disease but also because this information will assist in the development of future therapeutic options. After reviewing models of enhancer function, we discuss recent methods for identifying target genes of enhancers. First, we describe chromatin structure-based approaches for directly mapping interactions between enhancers and promoters. Second, we describe the use of correlation-based approaches to link enhancer state with the activity of nearby promoters and/or gene expression. Third, we describe how to test the function of specific enhancers experimentally by perturbing enhancer-target relationships using high-throughput reporter assays and genome editing. Finally, we conclude by discussing as yet unanswered questions concerning how enhancers function, how target genes can be identified, and how to distinguish direct from indirect changes in gene expression mediated by individual enhancers.

  15. Finding Quantitative Trait Loci Genes with Collaborative Targeted Maximum Likelihood Learning.

    Science.gov (United States)

    Wang, Hui; Rose, Sherri; van der Laan, Mark J

    2011-07-01

    Quantitative trait loci mapping is focused on identifying the positions and effect of genes underlying an an observed trait. We present a collaborative targeted maximum likelihood estimator in a semi-parametric model using a newly proposed 2-part super learning algorithm to find quantitative trait loci genes in listeria data. Results are compared to the parametric composite interval mapping approach.

  16. Generation of CRISPR/Cas9-mediated gene-targeted pigs via somatic cell nuclear transfer.

    Science.gov (United States)

    Zhou, Xiaoqing; Xin, Jige; Fan, Nana; Zou, Qingjian; Huang, Jiao; Ouyang, Zhen; Zhao, Yu; Zhao, Bentian; Liu, Zhaoming; Lai, Sisi; Yi, Xiaoling; Guo, Lin; Esteban, Miguel A; Zeng, Yangzhi; Yang, Huaqiang; Lai, Liangxue

    2015-03-01

    The domestic pig has been widely used as an important large animal model. Precise and efficient genetic modification in pig provides a great promise in biomedical research. Recently, clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated (Cas) system has been successfully used to produce many gene-targeted animals. However, these animals have been generated by co-injection of Cas9 mRNA and single-guide RNA (sgRNA) into one-cell stage embryos, which mostly resulted in mosaicism of the modification. One or two rounds of further breeding should be performed to obtain homozygotes with identical genotype and phenotype. To address this issue, gene-targeted somatic cells can be used as donor for somatic cell nuclear transfer (SCNT) to produce gene-targeted animals with single and identical mutations. In this study, we applied Cas9/sgRNAs to effectively direct gene editing in porcine fetal fibroblasts and then mutant cell colonies were used as donor to generate homozygous gene-targeted pigs through single round of SCNT. As a result, we successfully obtained 15 tyrosinase (TYR) biallelic mutant pigs and 20 PARK2 and PINK1 double-gene knockout (KO) pigs. They were all homozygous and no off-target mutagenesis was detected by comprehensive analysis. TYR (-/-) pigs showed typical albinism and the expression of parkin and PINK1 were depleted in PARK2 (-/-)/PINK1 (-/-) pigs. The results demonstrated that single- or double-gene targeted pigs can be effectively achieved by using the CRISPR/Cas9 system combined with SCNT without mosaic mutation and detectable off-target effects. This gene-editing system provides an efficient, rapid, and less costly manner to generate genetically modified pigs or other large animals.

  17. Double-stranded Let-7 mimics, potential candidates for cancer gene therapy.

    Science.gov (United States)

    Wang, Qi-zhao; Lv, Ying-hui; Gong, Yu-hua; Li, Zhao-fa; Xu, William; Diao, Yong; Xu, Ruian

    2012-03-01

    MicroRNAs (miRNAs), a class of small, single-stranded endogenous RNAs, act as post-transcriptional regulators of gene expression. The ability of one single miRNA regulating multiple functionally related mRNAs makes it a new potential candidate for cancer gene therapy. Let-7s miRNAs have been demonstrated as tumor-suppressor genes in various types of cancers, providing one choice of gene therapy by replenishing this miRNA. In the present studies, we demonstrate that the chemically synthesized, double-stranded Let-7 mimics can inhibit the growth and migration and induce the cell cycle arrest of lung cancer cell lines in vitro. Let-7 mimics silence gene expression by binding to the 3' UTR of targeting mRNAs. Mutation of seed sequence significantly depresses the gene silencing activity of Let-7 mimics. Our results also demonstrate that it is possible to increase the activity of Let-7s through mutating the sequence within the 3'end of the antisense strand. Directly, co-transfection Let-7 mimics with active siRNAs impairs the anti-cancer activities of Let-7 mimics. However, a 3-h interval between the introduction of Let-7 mimics and a kind of siRNA avoids the competition and enhances the anti-cancer activities of Let-7 mimics. Taken together, these results have revealed that Let-7s mimics are potential candidates for cancer gene therapy.

  18. Targeted inhibition of tumor growth and angiogenesis

    NARCIS (Netherlands)

    van der Meel, R.

    2013-01-01

    Two main strategies have been pursued for the development of an effective and targeted anti-cancer treatment. The first strategy comprised the generation of a targeted nanomedicine for the inhibition of tumor cell proliferation by blocking growth factor receptor pathways. The epidermal growth factor

  19. Transgenic gene knock-outs: functional genomics and therapeutic target selection.

    Science.gov (United States)

    Harris, S; Foord, S M

    2000-11-01

    The completion of the first draft of the human genome presents both a tremendous opportunity and enormous challenge to the pharmaceutical industry since the whole community, with few exceptions, will soon have access to the same pool of candidate gene sequences from which to select future therapeutic targets. The commercial imperative to select and pursue therapeutically relevant genes from within the overall content of the genome will be particularly intense for those gene families that currently represent the chemically tractable or 'drugable' gene targets. As a consequence the emphasis within exploratory research has shifted towards the evaluation and adoption of technology platforms that can add additional value to the gene selection process, either through functional studies or direct/indirect measures of disease alignment e.g., genetics, differential gene expression, proteomics, tissue distribution, comparative species data etc. The selection of biological targets for the development of potential new medicines relies, in part, on the quality of the in vivo biological data that correlates a particular molecular target with the underlying pathophysiology of a disease. Within the pharmaceutical industry, studies employing transgenic animals and, in particular, animals with specific gene deletions are playing an increasingly important role in the therapeutic target gene selection, drug candidate selection and product development phases of the overall drug discovery process. The potential of phenotypic information from gene knock-outs to contribute to a high-throughput target selection/validation strategy has hitherto been limited by the resources required to rapidly generate and characterise a large number of knock-out transgenics in a timely fashion. The offerings of several companies that provide an opportunity to overcome these hurdles, albeit at a cost, are assessed with respect to the strategic business needs of the pharmaceutical industry.

  20. Gene Dosage Analysis in a Clinical Environment: Gene-Targeted Microarrays as the Platform-of-Choice

    Directory of Open Access Journals (Sweden)

    Donald R. Love

    2013-03-01

    Full Text Available The role of gene deletion and duplication in the aetiology of disease has become increasingly evident over the last decade. In addition to the classical deletion/duplication disorders diagnosed using molecular techniques, such as Duchenne Muscular Dystrophy and Charcot-Marie-Tooth Neuropathy Type 1A, the significance of partial or whole gene deletions in the pathogenesis of a large number single-gene disorders is becoming more apparent. A variety of dosage analysis methods are available to the diagnostic laboratory but the widespread application of many of these techniques is limited by the expense of the kits/reagents and restrictive targeting to a particular gene or portion of a gene. These limitations are particularly important in the context of a small diagnostic laboratory with modest sample throughput. We have developed a gene-targeted, custom-designed comparative genomic hybridisation (CGH array that allows twelve clinical samples to be interrogated simultaneously for exonic deletions/duplications within any gene (or panel of genes on the array. We report here on the use of the array in the analysis of a series of clinical samples processed by our laboratory over a twelve-month period. The array has proven itself to be robust, flexible and highly suited to the diagnostic environment.

  1. Autophagy inhibits cell death induced by the anti-cancer drug morusin

    Science.gov (United States)

    Cho, Sang Woo; Na, Wooju; Choi, Minji; Kang, Shin Jung; Lee, Seok-Geun; Choi, Cheol Yong

    2017-01-01

    Autophagy is a cellular process by which damaged organelles and dysfunctional proteins are degraded. Morusin is an anti-cancer drug isolated from the root bark of Morus alba. Morusin induces apoptosis in human prostate cancer cells by reducing STAT3 activity. In this study, we examined whether morusin induces autophagy and also examined the effects of autophagy on the morusin-induced apoptosis. Morusin induces LC3-II accumulation and ULK1 activation in HeLa cells. In addition, we found that induction of ULK1 Ser317 phosphorylation and reduction of ULK1 Ser757 phosphorylation occurred simultaneously during morusin-induced autophagy. Consistently, morusin induces autophagy by activation of AMPK and inhibition of mTOR activity. Next, we investigated the role of autophagy in morusin-induced apoptosis. Inhibition of autophagy by treating cells with the 3-methyladenine (3-MA) autophagic inhibitor induces high levels of morusin-mediated apoptosis, while treatment of cells with morusin alone induces moderate levels of apoptosis. Cell survival was greatly reduced when cells were treated with morusin and 3-MA. Taken together, morusin induces autophagy, which is an impediment for morusin-induced apoptosis, suggesting combined treatment of morusin with an autophagic inhibitor would increase the efficacy of morusin as an anti-cancer drug.

  2. Anti-cancer effects of Kochia scoparia fruit in human breast cancer cells

    Directory of Open Access Journals (Sweden)

    Hye-Yeon Han

    2014-01-01

    Full Text Available Background: The fruit of Kochia scoparia Scharder is widely used as a medicinal ingredient for the treatment of dysuria and skin diseases in China, Japan and Korea. Especially, K. scoparia had been used for breast masses and chest and flank pain. Objective: To investigate the anti-cancer effect of K. scoparia on breast cancer. Materials and Methods: We investigated the anti-cancer effects of K. scoparia, methanol extract (MEKS in vitro. We examined the effects of MEKS on the proliferation rate, cell cycle arrest, reactive oxygen species (ROS generation and activation of apoptosis-associated proteins in MDA-MB-231, human breast cancer cells. Results: MTT assay results demonstrated that MEKS decreased the proliferation rates of MDA-MB-231 cells in a dose-dependent manner with an IC 50 value of 36.2 μg/ml. MEKS at 25 μg/ml significantly increased the sub-G1 DNA contents of MDA-MB-231 cells to 44.7%, versus untreated cells. In addition, MEKS induced apoptosis by increasing the levels of apoptosis-associated proteins such as cleaved caspase 3, cleaved caspase 8, cleaved caspase 9 and cleaved Poly (ADP-ribose polymerase (PARP. Conclusion: These results suggest that MEKS inhibits cell proliferation and induces apoptosis in breast cancer cells and that MEKS may have potential chemotherapeutic value for the treatment of human breast cancer.

  3. Application of an Efficient Gene Targeting System Linking Secondary Metabolites to their Biosynthetic Genes in Aspergillus terreus

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Chun-Jun; Knox, Benjamin P.; Sanchez, James F.; Chiang, Yi-Ming; Bruno, Kenneth S.; Wang, Clay C.

    2013-07-19

    Nonribosomal peptides (NRPs) are natural products biosynthesized by NRP synthetases. A kusA-, pyrG- mutant strain of Aspergillusterreus NIH 2624 was developed that greatly facilitated the gene targeting efficiency in this organism. Application of this tool allowed us to link four major types of NRP related secondary metabolites to their responsible genes in A. terreus. In addition, an NRP related melanin synthetase was also identified in this species.

  4. Glucagon-like peptide-2 (GLP-2) response to enteral intake in children during anti-cancer treatment

    DEFF Research Database (Denmark)

    Andreassen, B U; Paerregaard, A; Schmiegelow, K

    2005-01-01

    BACKGROUND: Intestinal dysfunction is frequent in cancer and during anti-cancer treatment. Glucagon-like peptide-2 (GLP-2) is secreted in a nutrition-dependent manner from the intestinal enteroendocrine L-cells. It accelerates crypt cell proliferation and nutrient absorption, inhibits enterocyte...... apoptosis and decreases mucosal permeability. Lack of GLP-2 may increase the risk of malabsorption and intestinal bacterial translocation. The aim of this study is to evaluate meal stimulated secretion of GLP-2 in children with cancer undergoing anti-cancer treatment. METHODS: Plasma-GLP-2 analysis after...... an overnight fast and 1 hour after intake of a mixed test meal. Data on gastrointestinal toxicity, blood neutrophile counts and food records were included in the analysis. RESULTS: Forty-four meal stimulation tests were performed in 25 children (median age, 6.0 years; range, 2.5-19) during anti-cancer...

  5. Study on Wusan Granule Anti-tumor Related Target Gene Screened by Cdna Microarray

    Institute of Scientific and Technical Information of China (English)

    YOU Zi-li; SHI Jin-ping; CHEN Hai-hong

    2006-01-01

    To screen Wusan Granule anti-tumor related target gene using cDNA microarray technique, both mRNA from Lewis lung carcinoma tissues treated by Wusan Granule and untreated control are reversibly transcribed to prepare cDNA probes which are labeled by Cy5 and Cy3. Then, the probes are hybridized to the mice cDNA microarray type MGEC-20S. After hybridization, the cDNA microarray is scanned by ScanArray 3 000 scanner and the data is analyzed by ImaGene 3 software to screen the differentially expressed genes. There are 45 differentially expressed genes including 18 known genes and 27 unknown genes between the two groups, and among them, 20 elevated genes and 25 reduced genes are identified. Additionally, the genes related to invasion and metastasis of malignant carcinomas are down-regulated and the genes related to apoptosis are up-regulated. The cDNA microarray technique is a high-throughput approach to screen the Wusan Granule anti-tumor related target genes, which allow us to explore the molecular biological mechanism on a genomic scale.

  6. Simple and Efficient Targeting of Multiple Genes Through CRISPR-Cas9 in Physcomitrella patens

    Directory of Open Access Journals (Sweden)

    Mauricio Lopez-Obando

    2016-11-01

    Full Text Available Powerful genome editing technologies are needed for efficient gene function analysis. The CRISPR-Cas9 system has been adapted as an efficient gene-knock-out technology in a variety of species. However, in a number of situations, knocking out or modifying a single gene is not sufficient; this is particularly true for genes belonging to a common family, or for genes showing redundant functions. Like many plants, the model organism Physcomitrella patens has experienced multiple events of polyploidization during evolution that has resulted in a number of families of duplicated genes. Here, we report a robust CRISPR-Cas9 system, based on the codelivery of a CAS9 expressing cassette, multiple sgRNA vectors, and a cassette for transient transformation selection, for gene knock-out in multiple gene families. We demonstrate that CRISPR-Cas9-mediated targeting of five different genes allows the selection of a quintuple mutant, and all possible subcombinations of mutants, in one experiment, with no mutations detected in potential off-target sequences. Furthermore, we confirmed the observation that the presence of repeats in the vicinity of the cutting region favors deletion due to the alternative end joining pathway, for which induced frameshift mutations can be potentially predicted. Because the number of multiple gene families in Physcomitrella is substantial, this tool opens new perspectives to study the role of expanded gene families in the colonization of land by plants.

  7. Targeting New Candidate Genes by Small Molecules Approaching Neurodegenerative Diseases.

    Science.gov (United States)

    Fan, Hueng-Chuen; Chi, Ching-Shiang; Cheng, Shin-Nan; Lee, Hsiu-Fen; Tsai, Jeng-Dau; Lin, Shinn-Zong; Harn, Horng-Jyh

    2015-12-25

    Neurodegenerative diseases (NDs) are among the most feared of the disorders that afflict humankind for the lack of specific diagnostic tests and effective treatments. Understanding the molecular, cellular, biochemical changes of NDs may hold therapeutic promise against debilitating central nerve system (CNS) disorders. In the present review, we summarized the clinical presentations and biology backgrounds of NDs, including Parkinson's disease (PD), Huntington's disease (HD), and Alzheimer's disease (AD) and explored the role of molecular mechanisms, including dys-regulation of epigenetic control mechanisms, Ataxia-telangiectasia-mutated protein kinase (ATM), and neuroinflammation in the pathogenesis of NDs. Targeting these mechanisms may hold therapeutic promise against these devastating diseases.

  8. Generating Targeted Gene Knockout Lines in Physcomitrella patens to Study Evolution of Stress-Responsive Mechanisms

    Science.gov (United States)

    Maronova, Monika; Kalyna, Maria

    2016-01-01

    The moss Physcomitrella patens possesses highly efficient homologous recombination allowing targeted gene manipulations and displays many features of the early land plants including high tolerance to abiotic stresses. It is therefore an invaluable model organism for studies of gene functions and comparative studies of evolution of stress responses in plants. Here, we describe a method for generating targeted gene knockout lines in P. patens using a polyethylene glycol-mediated transformation of protoplasts including basic in vitro growth, propagation, and maintenance techniques. PMID:26867627

  9. Targeting of human aFGF gene into silkworm,Bombyx mori L. through homologous recombination

    Institute of Scientific and Technical Information of China (English)

    吴小锋; 曹翠平

    2004-01-01

    The long-arm and short-arm genes of fibroin light chain (L-chain) of silkworm, Bombyx Mori L., and the gene of human acidic fibroblast growth factor were cloned respectively and subsequently inserted into a transfer vector pVL 1392 used as a tool to target the L-chain region of the silkworm genome. Genomic DNA from their offsprings was extracted and the expected targeting was detected using polymerase chain reaction and DNA sequencing, as well as protein analysis. The results showed that positive events occurred and that the FGF gene was integrated into the L-chain locus through homologous recombination.

  10. Targeting of human aFGF gene into silkworm, Bombyx mori L.Through homologous recombination

    Institute of Scientific and Technical Information of China (English)

    吴小锋; 曹翠平

    2004-01-01

    The long-arm and short-arm genes of fibroin light chain (L-chain) of silkworm, Bombyx Mori L., and the gene of human acidic fibroblast growth factor were cloned respectively and subsequently inserted into a transfer vector pVL1392 used as a tool to target the L-chain region of the silkworm genome. Genomic DNA from their offsprings was extracted and the expected targeting was detected using polymerase chain reaction and DNA sequencing, as well as protein analysis. The results showed that positive events occurred and that the FGF gene was integrated into the L-chain locus through homologous recombination.

  11. Identification and Regulation of c-Myb Target Genes in MCF-7 Cells

    Directory of Open Access Journals (Sweden)

    O'Rourke John P

    2011-01-01

    Full Text Available Abstract Background The c-Myb transcription factor regulates differentiation and proliferation in hematopoietic cells, stem cells and epithelial cells. Although oncogenic versions of c-Myb were first associated with leukemias, over expression or rearrangement of the c-myb gene is common in several types of solid tumors, including breast cancers. Expression of the c-myb gene in human breast cancer cells is dependent on estrogen stimulation, but little is known about the activities of the c-Myb protein or what genes it regulates in estrogen-stimulated cells. Methods We used chromatin immunoprecipitation coupled with whole genome promoter tiling microarrays to identify endogenous c-Myb target genes in human MCF-7 breast cancer cells and characterized the activity of c-Myb at a panel of target genes during different stages of estrogen deprivation and stimulation. Results By using different antibodies and different growth conditions, the c-Myb protein was found associated with over 10,000 promoters in MCF-7 cells, including many genes that encode cell cycle regulators or transcription factors and more than 60 genes that encode microRNAs. Several previously identified c-Myb target genes were identified, including CCNB1, MYC and CXCR4 and novel targets such as JUN, KLF4, NANOG and SND1. By studying a panel of these targets to validate the results, we found that estradiol stimulation triggered the association of c-Myb with promoters and that association correlated with increased target gene expression. We studied one target gene, CXCR4, in detail, showing that c-Myb associated with the CXCR4 gene promoter and activated a CXCR4 reporter gene in transfection assays. Conclusions Our results show that c-Myb associates with a surprisingly large number of promoters in human cells. The results also suggest that estradiol stimulation leads to large-scale, genome-wide changes in c-Myb activity and subsequent changes in gene expression in human breast cancer

  12. Capsid modification of adeno-associated virus and tumor targeting gene therapy

    Institute of Scientific and Technical Information of China (English)

    XU ZengHu; ZHOU XiuMei; SHI WenFang; QIAN QiJun

    2008-01-01

    Targeting is critical for successful tumor gene therapy. The adeno-associated virus (AAV) has aroused wide concern due to its excellent advantages over other viral vectors in gene therapy. AAV has a broad infection spectrum, which also results in poor specificity towards tissues or cells and low transduction efficiency. Therefore, it is imperative to improve target and transduction efficiency in AAV-mediated gene therapy. Up to now, researchers have developed many strategies to modify AAV capsids for improving targeting or retargeting only desired cells. These strategies include not only traditional chemical modification, phage display technology, modification of AAV capsid genome, chimeric vectors and so on, but also many novel strategies involved in marker rescue strategy, direct evolution of capsid proteins, direct display random peptides on AAV capsid, AAVP (AAV-Phage), and etc. This review will summarize the advances of researches on the capsid modification of AAV to target malignant cells.

  13. Expression of androgen receptor target genes in skeletal muscle

    Directory of Open Access Journals (Sweden)

    Kesha Rana

    2014-10-01

    Full Text Available We aimed to determine the mechanisms of the anabolic actions of androgens in skeletal muscle by investigating potential androgen receptor (AR-regulated genes in in vitro and in vivo models. The expression of the myogenic regulatory factor myogenin was significantly decreased in skeletal muscle from testosterone-treated orchidectomized male mice compared to control orchidectomized males, and was increased in muscle from male AR knockout mice that lacked DNA binding activity (ARΔZF2 versus wildtype mice, demonstrating that myogenin is repressed by the androgen/AR pathway. The ubiquitin ligase Fbxo32 was repressed by 12 h dihydrotestosterone treatment in human skeletal muscle cell myoblasts, and c-Myc expression was decreased in testosterone-treated orchidectomized male muscle compared to control orchidectomized male muscle, and increased in AR∆ZF2 muscle. The expression of a group of genes that regulate the transition from myoblast proliferation to differentiation, Tceal7 , p57 Kip2, Igf2 and calcineurin Aa, was increased in AR∆ZF2 muscle, and the expression of all but p57 Kip2 was also decreased in testosterone-treated orchidectomized male muscle compared to control orchidectomized male muscle. We conclude that in males, androgens act via the AR in part to promote peak muscle mass by maintaining myoblasts in the proliferative state and delaying the transition to differentiation during muscle growth and development, and by suppressing ubiquitin ligase-mediated atrophy pathways to preserve muscle mass in adult muscle.

  14. Expression of androgen receptor target genes in skeletal muscle

    Institute of Scientific and Technical Information of China (English)

    Kesha Rana; Nicole KL Lee; Jeffrey D Zajac; Helen E MacLean

    2014-01-01

    We aimed to determine the mechanisms of the anabolic actions of androgens in skeletal muscle by investigating potential androgen receptor(AR)‑regulated genes ininvitroandinvivomodels. The expression of the myogenic regulatory factormyogenin was signiifcantly decreased in skeletal muscle from testosterone‑treated orchidectomized male mice compared to control orchidectomized males, and was increased in muscle from male AR knockout mice that lacked DNA binding activity(ARΔZF2) versus wildtype mice, demonstrating thatmyogenin is repressed by the androgen/AR pathway. The ubiquitin ligaseFbxo32 was repressed by 12h dihydrotestosterone treatment in human skeletal muscle cell myoblasts, andc‑Myc expression was decreased in testosterone‑treated orchidectomized male muscle compared to control orchidectomized male muscle, and increased in AR∆ZF2 muscle. The expression of a group of genes that regulate the transition from myoblast proliferation to differentiation, Tceal7, p57Kip2, Igf2 andcalcineurin Aa, was increased in AR∆ZF2 muscle, and the expression of all butp57Kip2was also decreased in testosterone‑treated orchidectomized male muscle compared to control orchidectomized male muscle. We conclude that in males, androgens act via the AR in part to promote peak muscle mass by maintaining myoblasts in the proliferative state and delaying the transition to differentiation during muscle growth and development, and by suppressing ubiquitin ligase‑mediated atrophy pathways to preserve muscle mass in adult muscle.

  15. Expression of androgen receptor target genes in skeletal muscle.

    Science.gov (United States)

    Rana, Kesha; Lee, Nicole K L; Zajac, Jeffrey D; MacLean, Helen E

    2014-01-01

    We aimed to determine the mechanisms of the anabolic actions of androgens in skeletal muscle by investigating potential androgen receptor (AR)-regulated genes in in vitro and in vivo models. The expression of the myogenic regulatory factor myogenin was significantly decreased in skeletal muscle from testosterone-treated orchidectomized male mice compared to control orchidectomized males, and was increased in muscle from male AR knockout mice that lacked DNA binding activity (AR(ΔZF2)) versus wildtype mice, demonstrating that myogenin is repressed by the androgen/AR pathway. The ubiquitin ligase Fbxo32 was repressed by 12 h dihydrotestosterone treatment in human skeletal muscle cell myoblasts, and c-Myc expression was decreased in testosterone-treated orchidectomized male muscle compared to control orchidectomized male muscle, and increased in AR(∆ZF2) muscle. The expression of a group of genes that regulate the transition from myoblast proliferation to differentiation, Tceal7 , p57(Kip2), Igf2 and calcineurin Aa, was increased in AR(∆ZF2) muscle, and the expression of all but p57(Kip2) was also decreased in testosterone-treated orchidectomized male muscle compared to control orchidectomized male muscle. We conclude that in males, androgens act via the AR in part to promote peak muscle mass by maintaining myoblasts in the proliferative state and delaying the transition to differentiation during muscle growth and development, and by suppressing ubiquitin ligase-mediated atrophy pathways to preserve muscle mass in adult muscle.

  16. Nonimmunoglobulin target loci of activation-induced cytidine deaminase (AID) share unique features with immunoglobulin genes.

    Science.gov (United States)

    Kato, Lucia; Begum, Nasim A; Burroughs, A Maxwell; Doi, Tomomitsu; Kawai, Jun; Daub, Carsten O; Kawaguchi, Takahisa; Matsuda, Fumihiko; Hayashizaki, Yoshihide; Honjo, Tasuku

    2012-02-14

    Activation-induced cytidine deaminase (AID) is required for both somatic hypermutation and class-switch recombination in activated B cells. AID is also known to target nonimmunoglobulin genes and introduce mutations or chromosomal translocations, eventually causing tumors. To identify as-yet-unknown AID targets, we screened early AID-induced DNA breaks by using two independent genome-wide approaches. Along with known AID targets, this screen identified a set of unique genes (SNHG3, MALAT1, BCL7A, and CUX1) and confirmed that these loci accumulated mutations as frequently as Ig locus after AID activation. Moreover, these genes share three important characteristics with the Ig gene: translocations in tumors, repetitive sequences, and the epigenetic modification of chromatin by H3K4 trimethylation in the vicinity of cleavage sites.

  17. Optimal in silico target gene deletion through nonlinear programming for genetic engineering.

    Directory of Open Access Journals (Sweden)

    Chung-Chien Hong

    Full Text Available BACKGROUND: Optimal selection of multiple regulatory genes, known as targets, for deletion to enhance or suppress the activities of downstream genes or metabolites is an important problem in genetic engineering. Such problems become more feasible to address in silico due to the availability of more realistic dynamical system models of gene regulatory and metabolic networks. The goal of the computational problem is to search for a subset of genes to knock out so that the activity of a downstream gene or a metabolite is optimized. METHODOLOGY/PRINCIPAL FINDINGS: Based on discrete dynamical system modeling of gene regulatory networks, an integer programming problem is formulated for the optimal in silico target gene deletion problem. In the first result, the integer programming problem is proved to be NP-hard and equivalent to a nonlinear programming problem. In the second result, a heuristic algorithm, called GKONP, is designed to approximate the optimal solution, involving an approach to prune insignificant terms in the objective function, and the parallel differential evolution algorithm. In the third result, the effectiveness of the GKONP algorithm is demonstrated by applying it to a discrete dynamical system model of the yeast pheromone pathways. The empirical accuracy and time efficiency are assessed in comparison to an optimal, but exhaustive search strategy. SIGNIFICANCE: Although the in silico target gene deletion problem has enormous potential applications in genetic engineering, one must overcome the computational challenge due to its NP-hardness. The presented solution, which has been demonstrated to approximate the optimal solution in a practical amount of time, is among the few that address the computational challenge. In the experiment on the yeast pheromone pathways, the identified best subset of genes for deletion showed advantage over genes that were selected empirically. Once validated in vivo, the optimal target genes are

  18. Focal DNA copy number changes in neuroblastoma target MYCN regulated genes.

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    Candy Kumps

    Full Text Available Neuroblastoma is an embryonic tumor arising from immature sympathetic nervous system cells. Recurrent genomic alterations include MYCN and ALK amplification as well as recurrent patterns of gains and losses of whole or large partial chromosome segments. A recent whole genome sequencing effort yielded no frequently recurring mutations in genes other than those affecting ALK. However, the study further stresses the importance of DNA copy number alterations in this disease, in particular for genes implicated in neuritogenesis. Here we provide additional evidence for the importance of focal DNA copy number gains and losses, which are predominantly observed in MYCN amplified tumors. A focal 5 kb gain encompassing the MYCN regulated miR-17~92 cluster as sole gene was detected in a neuroblastoma cell line and further analyses of the array CGH data set demonstrated enrichment for other MYCN target genes in focal gains and amplifications. Next we applied an integrated genomics analysis to prioritize MYCN down regulated genes mediated by MYCN driven miRNAs within regions of focal heterozygous or homozygous deletion. We identified RGS5, a negative regulator of G-protein signaling implicated in vascular normalization, invasion and metastasis, targeted by a focal homozygous deletion, as a new MYCN target gene, down regulated through MYCN activated miRNAs. In addition, we expand the miR-17~92 regulatory network controlling TGFß signaling in neuroblastoma with the ring finger protein 11 encoding gene RNF11, which was previously shown to be targeted by the miR-17~92 member miR-19b. Taken together, our data indicate that focal DNA copy number imbalances in neuroblastoma (1 target genes that are implicated in MYCN signaling, possibly selected to reinforce MYCN oncogene addiction and (2 serve as a resource for identifying new molecular targets for treatment.

  19. Targeted cancer gene therapy : the flexibility of adenoviral gene therapy vectors

    NARCIS (Netherlands)

    Rots, MG; Curiel, DT; Gerritsen, WR; Haisma, HJ

    2003-01-01

    Recombinant adenoviral vectors are promising reagents for therapeutic interventions in humans, including gene therapy for biologically complex diseases like cancer and cardiovascular diseases. In this regard, the major advantage of adenoviral vectors is their superior in vivo gene transfer efficienc

  20. Rational design, synthesis, and biological evaluation of third generation α-noscapine analogues as potent tubulin binding anti-cancer agents.

    Science.gov (United States)

    Manchukonda, Naresh Kumar; Naik, Pradeep Kumar; Santoshi, Seneha; Lopus, Manu; Joseph, Silja; Sridhar, Balasubramanian; Kantevari, Srinivas

    2013-01-01

    Systematic screening based on structural similarity of drugs such as colchicine and podophyllotoxin led to identification of noscapine, a microtubule-targeted agent that attenuates the dynamic instability of microtubules without affecting the total polymer mass of microtubules. We report a new generation of noscapine derivatives as potential tubulin binding anti-cancer agents. Molecular modeling experiments of these derivatives 5a, 6a-j yielded better docking score (-7.252 to -5.402 kCal/mol) than the parent compound, noscapine (-5.505 kCal/mol) and its existing derivatives (-5.563 to -6.412 kCal/mol). Free energy (ΔG bind ) calculations based on the linear interaction energy (LIE) empirical equation utilizing Surface Generalized Born (SGB) continuum solvent model predicted the tubulin-binding affinities for the derivatives 5a, 6a-j (ranging from -4.923 to -6.189 kCal/mol). Compound 6f showed highest binding affinity to tubulin (-6.189 kCal/mol). The experimental evaluation of these compounds corroborated with theoretical studies. N-(3-brormobenzyl) noscapine (6f) binds tubulin with highest binding affinity (KD, 38 ± 4.0 µM), which is ~ 4.0 times higher than that of the parent compound, noscapine (KD, 144 ± 1.0 µM) and is also more potent than that of the first generation clinical candidate EM011, 9-bromonoscapine (KD, 54 ± 9.1 µM). All these compounds exhibited substantial cytotoxicity toward cancer cells, with IC50 values ranging from 6.7 µM to 72.9 µM; compound 6f showed prominent anti-cancer efficacy with IC50 values ranging from 6.7 µM to 26.9 µM in cancer cells of different tissues of origin. These compounds perturbed DNA synthesis, delayed the cell cycle progression at G2/M phase, and induced apoptotic cell death in cancer cells. Collectively, the study reported here identified potent, third generation noscapinoids as new anti-cancer agents.

  1. Rational design, synthesis, and biological evaluation of third generation α-noscapine analogues as potent tubulin binding anti-cancer agents.

    Directory of Open Access Journals (Sweden)

    Naresh Kumar Manchukonda

    Full Text Available Systematic screening based on structural similarity of drugs such as colchicine and podophyllotoxin led to identification of noscapine, a microtubule-targeted agent that attenuates the dynamic instability of microtubules without affecting the total polymer mass of microtubules. We report a new generation of noscapine derivatives as potential tubulin binding anti-cancer agents. Molecular modeling experiments of these derivatives 5a, 6a-j yielded better docking score (-7.252 to -5.402 kCal/mol than the parent compound, noscapine (-5.505 kCal/mol and its existing derivatives (-5.563 to -6.412 kCal/mol. Free energy (ΔG bind calculations based on the linear interaction energy (LIE empirical equation utilizing Surface Generalized Born (SGB continuum solvent model predicted the tubulin-binding affinities for the derivatives 5a, 6a-j (ranging from -4.923 to -6.189 kCal/mol. Compound 6f showed highest binding affinity to tubulin (-6.189 kCal/mol. The experimental evaluation of these compounds corroborated with theoretical studies. N-(3-brormobenzyl noscapine (6f binds tubulin with highest binding affinity (KD, 38 ± 4.0 µM, which is ~ 4.0 times higher than that of the parent compound, noscapine (KD, 144 ± 1.0 µM and is also more potent than that of the first generation clinical candidate EM011, 9-bromonoscapine (KD, 54 ± 9.1 µM. All these compounds exhibited substantial cytotoxicity toward cancer cells, with IC50 values ranging from 6.7 µM to 72.9 µM; compound 6f showed prominent anti-cancer efficacy with IC50 values ranging from 6.7 µM to 26.9 µM in cancer cells of different tissues of origin. These compounds perturbed DNA synthesis, delayed the cell cycle progression at G2/M phase, and induced apoptotic cell death in cancer cells. Collectively, the study reported here identified potent, third generation noscapinoids as new anti-cancer agents.

  2. Targeting New Candidate Genes by Small Molecules Approaching Neurodegenerative Diseases

    Directory of Open Access Journals (Sweden)

    Hueng-Chuen Fan

    2015-12-01

    Full Text Available Neurodegenerative diseases (NDs are among the most feared of the disorders that afflict humankind for the lack of specific diagnostic tests and effective treatments. Understanding the molecular, cellular, biochemical changes of NDs may hold therapeutic promise against debilitating central nerve system (CNS disorders. In the present review, we summarized the clinical presentations and biology backgrounds of NDs, including Parkinson’s disease (PD, Huntington’s disease (HD, and Alzheimer’s disease (AD and explored the role of molecular mechanisms, including dys-regulation of epigenetic control mechanisms, Ataxia-telangiectasia-mutated protein kinase (ATM, and neuroinflammation in the pathogenesis of NDs. Targeting these mechanisms may hold therapeutic promise against these devastating diseases.

  3. Transfection and mutagenesis of target genes in mosquito cells by locked nucleic acid-modified oligonucleotides.

    Science.gov (United States)

    Pakpour, Nazzy; Cheung, Kong Wai; Souvannaseng, Lattha; Concordet, Jean-Paul; Luckhart, Shirley

    2010-12-26

    Plasmodium parasites, the causative agent of malaria, are transmitted through the bites of infected Anopheles mosquitoes resulting in over 250 million new infections each year. Despite decades of research, there is still no vaccine against malaria, highlighting the need for novel control strategies. One innovative approach is the use of genetically modified mosquitoes to effectively control malaria parasite transmission. Deliberate alterations of cell signaling pathways in the mosquito, via targeted mutagenesis, have been found to regulate parasite development (1). From these studies, we can begin to identify potential gene targets for transformation. Targeted mutagenesis has traditionally relied upon the homologous recombination between a target gene and a large DNA molecule. However, the construction and use of such complex DNA molecules for generation of stably transformed cell lines is costly, time consuming and often inefficient. Therefore, a strategy using locked nucleic acid-modified oligonucleotides (LNA-ONs) provides a useful alternative for introducing artificial single nucleotide substitutions into episomal and chromosomal DNA gene targets (reviewed in (2)). LNA-ON-mediated targeted mutagenesis has been used to introduce point mutations into genes of interest in cultured cells of both yeast and mice (3,4). We show here that LNA-ONs can be used to introduce a single nucleotide change in a transfected episomal target that results in a switch from blue fluorescent protein (BFP) expression to green fluorescent protein (GFP) expression in both Anopheles gambiae and Anopheles stephensi cells. This conversion demonstrates for the first time that effective mutagenesis of target genes in mosquito cells can be mediated by LNA-ONs and suggests that this technique may be applicable to mutagenesis of chromosomal targets in vitro and in vivo.

  4. Efficient gene targeting of the Rosa26 locus in mouse zygotes using TALE nucleases.

    Science.gov (United States)

    Kasparek, Petr; Krausova, Michaela; Haneckova, Radka; Kriz, Vitezslav; Zbodakova, Olga; Korinek, Vladimir; Sedlacek, Radislav

    2014-11-03

    Gene targeting in mice mainly employs homologous recombination (HR) in embryonic stem (ES) cells. Although it is a standard way for production of genetically modified mice, the procedure is laborious and time-consuming. This study describes targeting of the mouse Rosa26 locus by transcription activator-like effector nucleases (TALENs). We employed TALEN-assisted HR in zygotes to introduce constructs encoding TurboRFP and TagBFP fluorescent proteins into the first intron of the Rosa26 gene, and in this way generated two transgenic mice. We also demonstrated that these Rosa26-specific TALENs exhibit high targeting efficiency superior to that of zinc-finger nucleases (ZFNs) specific for the same targeting sequence. Moreover, we devised a reporter assay to assess TALENs activity and specificity to improve the quality of TALEN-assisted targeting.

  5. Targeted Editing of Myostatin Gene in Sheep by Transcription Activator-like Effector Nucleases.

    Science.gov (United States)

    Zhao, Xinxia; Ni, Wei; Chen, Chuangfu; Sai, Wujiafu; Qiao, Jun; Sheng, Jingliang; Zhang, Hui; Li, Guozhong; Wang, Dawei; Hu, Shengwei

    2016-03-01

    Myostatin (MSTN) is a secreted growth factor expressed in skeletal muscle and adipose tissue that negatively regulates skeletal muscle mass. Gene knockout of MSTN can result in increasing muscle mass in sheep. The objectives were to investigate whether myostatin gene can be edited in sheep by transcription activator-like effector nucleases (TALENs) in tandem with single-stranded DNA oligonucleotides (ssODNs). We designed a pair of TALENs to target a highly conserved sequence in the coding region of the sheep MSTN gene. The activity of the TALENs was verified by using luciferase single-strand annealing reporter assay in HEK 293T cell line. Co-transfection of TALENs and ssODNs oligonucleotides induced precise gene editing of myostatin gene in sheep primary fibroblasts. MSTN gene-edited cells were successfully used as nuclear donors for generating cloned embryos. TALENs combined with ssDNA oligonucleotides provide a useful approach for precise gene modification in livestock animals.

  6. [Melanoma: surface markers as the first point of targeted delivery of therapeutic genes in multilevel gene therapy].

    Science.gov (United States)

    Pleshkan, V V; Zinov'eva, M V; Sverdlov, E D

    2011-01-01

    Melanoma is one of the most malignant tumors, aggressively metastasizing by lymphatic and hematogenous routes. Due to the resistance of melanoma cells to many types of chemotherapy, this disease causes high mortality rate. High hopes are pinned on gene therapeutic approaches to melanoma treatment. At present, one of the main problems of the efficient use of the post-genomic generation therapeutic means is the lack of optimal techniques of delivery of foreign genetic material to the patient's target cells. Surface specific markers of melanoma cells can be considered as promising therapeutic targets. This review describes currently known melanoma specific receptors and its stem cells, as well as contains data on melanoma antigens presented on the cell surface by major histocompatibility complex proteins. The ability of surface proteins to internalize might be successfully used for the development of methods of targeted delivery of gene therapeutic constructs. In conclusion, a concept of multilevel gene therapy and the possible role therein of surface determinants as targets of gene systems delivery to the tumor are discussed.

  7. Fusion genes in solid tumors:an emerging target for cancer diagnosis and treatment

    Institute of Scientific and Technical Information of China (English)

    Brittany C. Parker; Wei Zhang

    2013-01-01

    Studies over the past decades have uncovered fusion genes, a class of oncogenes that provide immense diagnostic and therapeutic advantages because of their tumor-specific expression. Originally associated with hemotologic cancers, fusion genes have recently been discovered in a wide array of solid tumors, including sarcomas, carcinomas, and tumors of the central nervous system. Fusion genes are attractive as both therapeutic targets and diagnostic tools due to their inherent expression in tumor tissue alone. Therefore, the discovery and elucidation of fusion genes in various cancer types may provide more effective therapies in the future for cancer patients.

  8. Cell type-specific anti-cancer properties of valproic acid: independent effects on HDAC activity and Erk1/2 phosphorylation

    DEFF Research Database (Denmark)

    Gotfryd, Kamil; Skladchikova, Galina; Lepekhin, Eugene E

    2010-01-01

    ABSTRACT: BACKGROUND: The anti-epileptic drug valproic acid (VPA) has attracted attention as an anti-cancer agent. Methods: The present study investigated effects of VPA exposure on histone deacetylase (HDAC) inhibition, cell growth, cell speed, and the degree of Erk1/2 phosphorylation in 10 cell....../2 phosphorylation are also important for the anti-cancer properties of VPA....

  9. Novel Hematopoietic Target Genes in the NRF2-Mediated Transcriptional Pathway

    Directory of Open Access Journals (Sweden)

    Michelle R. Campbell

    2013-01-01

    Full Text Available Nuclear factor- (erythroid-derived 2 like 2 (NFE2L2, NRF2 is a key transcriptional activator of the antioxidant response pathway and is closely related to erythroid transcription factor NFE2. Under oxidative stress, NRF2 heterodimerizes with small Maf proteins and binds cis-acting enhancer sequences found near oxidative stress response genes. Using the dietary isothiocyanate sulforaphane (SFN to activate NRF2, chromatin immunoprecipitation sequencing (ChIP-seq identified several hundred novel NRF2-mediated targets beyond its role in oxidative stress. Activated NRF2 bound the antioxidant response element (ARE in promoters of several known and novel target genes involved in iron homeostasis and heme metabolism, including known targets FTL and FTH1, as well as novel binding in the globin locus control region. Five novel NRF2 target genes were chosen for followup: AMBP, ABCB6, FECH, HRG-1 (SLC48A1, and TBXAS1. SFN-induced gene expression in erythroid K562 and lymphoid cells were compared for each target gene. NRF2 silencing showed reduced expression in lymphoid, lung, and hepatic cells. Furthermore, stable knockdown of NRF2 negative regulator KEAP1 in K562 cells resulted in increased NQO1, AMBP, and TBXAS1 expression. NFE2 binding sites in K562 cells revealed similar binding profiles as lymphoid NRF2 sites in all potential NRF2 candidates supporting a role for NRF2 in heme metabolism and erythropoiesis.

  10. W::Neo: a novel dual-selection marker for high efficiency gene targeting in Drosophila.

    Directory of Open Access Journals (Sweden)

    Wenke Zhou

    Full Text Available We have recently developed a so-called genomic engineering approach that allows for directed, efficient and versatile modifications of Drosophila genome by combining the homologous recombination (HR-based gene targeting with site-specific DNA integration. In genomic engineering and several similar approaches, a "founder" knock-out line must be generated first through HR-based gene targeting, which can still be a potentially time and resource intensive process. To significantly improve the efficiency and success rate of HR-based gene targeting in Drosophila, we have generated a new dual-selection marker termed W::Neo, which is a direct fusion between proteins of eye color marker White (W and neomycin resistance (Neo. In HR-based gene targeting experiments, mutants carrying W::Neo as the selection marker can be enriched as much as fifty times by taking advantage of the antibiotic selection in Drosophila larvae. We have successfully carried out three independent gene targeting experiments using the W::Neo to generate genomic engineering founder knock-out lines in Drosophila.

  11. Precision genome editing in plants via gene targeting and piggyBac-mediated marker excision.

    Science.gov (United States)

    Nishizawa-Yokoi, Ayako; Endo, Masaki; Ohtsuki, Namie; Saika, Hiroaki; Toki, Seiichi

    2015-01-01

    Precise genome engineering via homologous recombination (HR)-mediated gene targeting (GT) has become an essential tool in molecular breeding as well as in basic plant science. As HR-mediated GT is an extremely rare event, positive-negative selection has been used extensively in flowering plants to isolate cells in which GT has occurred. In order to utilize GT as a methodology for precision mutagenesis, the positive selectable marker gene should be completely eliminated from the GT locus. Here, we introduce targeted point mutations conferring resistance to herbicide into the rice acetolactate synthase (ALS) gene via GT with subsequent marker excision by piggyBac transposition. Almost all regenerated plants expressing piggyBac transposase contained exclusively targeted point mutations without concomitant re-integration of the transposon, resulting in these progeny showing a herbicide bispyribac sodium (BS)-tolerant phenotype. This approach was also applied successfully to the editing of a microRNA targeting site in the rice cleistogamy 1 gene. Therefore, our approach provides a general strategy for the targeted modification of endogenous genes in plants.

  12. Preparation of RGD-modified Long Circulating Liposome Loading Matrine, and its in vitro Anti-cancer Effects

    Directory of Open Access Journals (Sweden)

    Xiao-yan Liu, Li-ming Ruan, Wei-wei Mao, Jin-Qiang Wang, You-qing Shen, Mei-hua Sui

    2010-01-01

    Full Text Available Aim: To prepare RGD-modified long circulating liposome (LCL loading matrine (RGD-M-LCL to improve the tumor-targeting and efficacy of matrine. Methods: LCL which was prepared with HSPC, cholesterol, DSPE-PEG2000 and DSPE-PEG-MAL was modified with an RGD motif confirmed by high performance liquid chromatography (HPLC. The encapsulation efficiency of RGD-M-LCL was also detected by HPLC. MTT assay was used to examine the effects of RGD-M-LCL on the proliferation of Bcap-37, HT-29 and A375 cells. The percentage of apoptotic cells and morphological changes in Bcap-37 cells treated with RGD-M-LCL were detected by Annexin-V-FITC/PI affinity assay and observed under light microscope, respectively. Results: Spherical or oval single-chamber particles of uniform sizes with little agglutination or adhesion were observed under transmission electronic microscope. The RGD motif was successfully coupled to the DSPE-PEG-MAL on liposomes, as confirmed by HPLC. An encapsulation efficiency of 83.13% was obtained when the drug-lipid molar ratio was 0.1, and the encapsulation efficiency was negatively related to the drug-lipid ratio in the range of 0.1~0.4, and to the duration of storage. We found that, compared with free matrine, RGD-M-LCL had much stronger in vitro activity, leading to anti-proliferative and pro-apoptotic effects against cancer cells (P<0.01. Conclusion: RGD-M-LCL, a novel delivery system for anti-cancer drugs, was successfully prepared, and we demonstrated that the use of this material could augment the effects of matrine on cancer cells in vitro.

  13. Yeast-based assay identifies novel Shh/Gli target genes in vertebrate development

    Directory of Open Access Journals (Sweden)

    Milla Luis A

    2012-01-01

    Full Text Available Abstract Background The increasing number of developmental events and molecular mechanisms associated with the Hedgehog (Hh pathway from Drosophila to vertebrates, suggest that gene regulation is crucial for diverse cellular responses, including target genes not yet described. Although several high-throughput, genome-wide approaches have yielded information at the genomic, transcriptional and proteomic levels, the specificity of Gli binding sites related to direct target gene activation still remain elusive. This study aims to identify novel putative targets of Gli transcription factors through a protein-DNA binding assay using yeast, and validating a subset of targets both in-vitro and in-vivo. Testing in different Hh/Gli gain- and loss-of-function scenarios we here identified known (e.g., ptc1 and novel Hh-regulated genes in zebrafish embryos. Results The combined yeast-based screening and MEME/MAST analysis were able to predict Gli transcription factor binding sites, and position mapping of these sequences upstream or in the first intron of promoters served to identify new putative target genes of Gli regulation. These candidates were validated by qPCR in combination with either the pharmacological Hh/Gli antagonist cyc or the agonist pur in Hh-responsive C3H10T1/2 cells. We also used small-hairpin RNAs against Gli proteins to evaluate targets and confirm specific Gli regulation their expression. Taking advantage of mutants that have been identified affecting different components of the Hh/Gli signaling system in the zebrafish model, we further analyzed specific novel candidates. Studying Hh function with pharmacological inhibition or activation complemented these genetic loss-of-function approaches. We provide evidence that in zebrafish embryos, Hh signaling regulates sfrp2, neo1, and c-myc expression in-vivo. Conclusion A recently described yeast-based screening allowed us to identify new Hh/Gli target genes, functionally important in

  14. Curcumin: A review of anti-cancer properties and therapeutic activity in head and neck squamous cell carcinoma

    Directory of Open Access Journals (Sweden)

    Wang Marilene B

    2011-02-01

    Full Text Available Abstract Curcumin (diferuloylmethane is a polyphenol derived from the Curcuma longa plant, commonly known as turmeric. Curcumin has been used extensively in Ayurvedic medicine for centuries, as it is nontoxic and has a variety of therapeutic properties including anti-oxidant, analgesic, anti-inflammatory and antiseptic activity. More recently curcumin has been found to possess anti-cancer activities via its effect on a variety of biological pathways involved in mutagenesis, oncogene expression, cell cycle regulation, apoptosis, tumorigenesis and metastasis. Curcumin has shown anti-proliferative effect in multiple cancers, and is an inhibitor of the transcription factor NF-κB and downstream gene products (including c-myc, Bcl-2, COX-2, NOS, Cyclin D1, TNF-α, interleukins and MMP-9. In addition, curcumin affects a variety of growth factor receptors and cell adhesion molecules involved in tumor growth, angiogenesis and metastasis. Head and neck squamous cell carcinoma (HNSCC is the sixth most common cancer worldwide and treatment protocols include disfiguring surgery, platinum-based chemotherapy and radiation, all of which may result in tremendous patient morbidity. As a result, there is significant interest in developing adjuvant chemotherapies to augment currently available treatment protocols, which may allow decreased side effects and toxicity without compromising therapeutic efficacy. Curcumin is one such potential candidate, and this review presents an overview of the current in vitro and in vivo data supporting its therapeutic activity in head and neck cancer as well as some of the challenges concerning its development as an adjuvant chemotherapeutic agent.

  15. Genome-wide discovery of Pax7 target genes during development.

    Science.gov (United States)

    White, Robert B; Ziman, Melanie R

    2008-03-14

    Pax7 plays critical roles in development of brain, spinal cord, neural crest, and skeletal muscle. As a sequence-specific DNA-binding transcription factor, any direct functional role played by Pax7 during development is mediated through target gene selection. Thus, we have sought to identify genes targeted by Pax7 during embryonic development using an unbiased chromatin immunoprecipitation (ChIP) cloning assay to isolate cis-regulatory regions bound by Pax7 in vivo. Sequencing and genomic localization of a library of chromatin-DNA fragments bound by Pax7 has identified 34 candidate Pax7 target genes, with occupancy of a selection confirmed with independent chromatin enrichment tests (ChIP-PCR). To assess the capacity of Pax7 to regulate transcription from these loci, we have cloned alternate transcripts of Pax7 (differing significantly in their DNA binding domain) into expression vectors and transfected cultured cells with these constructs, then analyzed target gene expression levels using RT-PCR. We show that Pax7 directly occupies sites within genes encoding transcription factors Gbx1 and Eya4, the neurogenic cytokine receptor ciliary neurotrophic factor receptor, the neuronal potassium channel Kcnk2, and the signal transduction kinase Camk1d in vivo and regulates the transcriptional state of these genes in cultured cells. This analysis gives us greater insight into the direct functional role played by Pax7 during embryonic development.

  16. Global identification of target genes regulated by APETALA3 and PISTILLATA floral homeotic gene action.

    Science.gov (United States)

    Zik, Moriyah; Irish, Vivian F

    2003-01-01

    Identifying the genes regulated by the floral homeotic genes APETALA3 (AP3) and PISTILLATA (PI) is crucial for understanding the molecular mechanisms that lead to petal and stamen formation. We have used microarray analysis to conduct a broad survey of genes whose expression is affected by AP3 and PI activity. DNA microarrays consisting of 9216 Arabidopsis ESTs were screened with probes corresponding to mRNAs from different mutant and transgenic lines that misexpress AP3 and/or PI. The microarray results were further confirmed by RNA gel blot analyses. Our results suggest that AP3 and PI regulate a relatively small number of genes, implying that many genes used in petal and stamen development are not tissue specific and likely have roles in other processes as well. We recovered genes similar to previously identified petal- and stamen-expressed genes as well as genes that were not implicated previously in petal and stamen development. A very low percentage of the genes recovered encoded transcription factors. This finding suggests that AP3 and PI act relatively directly to regulate the genes required for the basic cellular processes responsible for petal and stamen morphogenesis.

  17. Anti-Cancer Effect of Angelica Sinensis on Women’s Reproductive Cancer

    Directory of Open Access Journals (Sweden)

    Hong-Hong Zhu

    2012-06-01

    Full Text Available Objective: Danggui, the root of Angelica Sinensis, has traditionally been used for the treatment of women’s reproductive disorders in China for thousands of years. This study was to determine whether Danggui have potential anti-cancer effect on women’s cancer and its potential mechanism. Methods: Danggui was extracted by ethanol. The Cell Titer 96® Aqueous Non-Radioactive Cell Proliferation Assay was used to compare the effects of Danggui on human breast (MCF-7 and 7368 and cervical (CaSki and SiHa cancer cells with its effects on normal fibroblasts (HTB-125. A revised Ames test was used to test for antimutagenicity. The standard strains of Salmonella typhimarium (TA 100 and 102 were used in the test. Methyl methane sulfonate (MMS and UV light were used as positive mutagen controls and ethanol and double distilled water (DDW as controls. The SAS statistical software was used to analyze the data. Results: Danggui was found to be much more toxic to all cancer cell lines tested than to normal fibroblasts. There was a significant negative dose-effect relationship between Danggui and cancer cell viability. Average viability of MCF-7 was 69.5%, 18.4%, 5.7%, 5.7%, and 5.0% of control for Danggui doses 0.07, 0.14, 0.21, 0.32, and 0.64 ug/ul, respectively, with a Ptrend < 0.0001. Half maximal inhibitory dose (ID50 of Danggui for cancer cell lines MCF-7, CaSki, SiHa and CRL-7368 was 0.10, 0.09, 0.10 and 0.07 ug/ul, Functional Foods in Health and Disease 2012, 2(6:242-250respectively. For the normal fibroblasts, ID50 was 0.58 ug/ul. At a dose of 0.32 ug/ul, Danggui killed over 90% of the cells in each cancer cell line, but at the same dose, only 12.3 % of the normal HTB-125 cells were killed. Revertants per plate of TA 100 decreased with the introduction of increasing doses of Danggui extracts with a Ptrend < 0.0001 when UV light was used as a mutagen. There was no difference in revertants per plate between ethanol and DDW control groups. Conclusions

  18. Analysis of target genes induced by IL-13 cytotoxin in human glioblastoma cells.

    Science.gov (United States)

    Han, Jing; Yang, Liming; Puri, Raj K

    2005-03-01

    IL-13 cytotoxin comprised of IL-13 and a mutated form of Pseudomonas exotoxin (fusion protein termed IL-13-PE38QQR) has been shown to inhibit protein synthesis leading to necrotic and apoptotic cell death in glioblastoma cells that express high levels of interleukin-13 receptors (IL-13R). To identify target genes of cell death and other cellular genes with IL-13 receptors in glioblastoma cells, we utilized the cDNA microarrays to analyze global gene expression profiles after IL-13 cytotoxin and IL-13 treatment. IL-13 cytotoxin mediated cytotoxicity to U251 cells in a dose-dependent manner. Hierarchical cluster analysis of differentially expressed genes in U251 glioma cells at different time points after IL-13 cytotoxin treatment showed three major groups, each representing a specific expression pattern. Randomly selected differentially expressed genes from each group were confirmed by RT-PCR analysis. Most down-regulated genes belong to cell adhesion, motility, angiogenesis, DNA repair, and metabolic pathways. While up-regulated genes belong to cell cycle arrest, apoptosis, signaling and various metabolic pathways. Unexpectedly, at early time points, both IL-13 and IL-13 cytotoxin induced several genes belonging to different pathways most notably IL-8, DIO2, END1, and ALDH1A3 indicating that these genes are early response genes and their products may be associated with IL-13R. In addition, IL-13 cytotoxin induced IL-13Ralpha2 mRNA expression during the treatment in glioma cells. Our results indicate that novel cellular genes are involved with IL-13 receptors and that IL-13 cytotoxin induced cell death involves various target genes in human glioblastoma cells. On going studies will determine the role of associated genes and their products in the IL-13R functions in glioma cells.

  19. Dynamic impacts of the inhibition of the molecular chaperone Hsp90 on the T-cell proteome have implications for anti-cancer therapy.

    Directory of Open Access Journals (Sweden)

    Ivo Fierro-Monti

    Full Text Available The molecular chaperone Hsp90-dependent proteome represents a complex protein network of critical biological and medical relevance. Known to associate with proteins with a broad variety of functions termed clients, Hsp90 maintains key essential and oncogenic signalling pathways. Consequently, Hsp90 inhibitors are being tested as anti-cancer drugs. Using an integrated systematic approach to analyse the effects of Hsp90 inhibition in T-cells, we quantified differential changes in the Hsp90-dependent proteome, Hsp90 interactome, and a selection of the transcriptome. Kinetic behaviours in the Hsp90-dependent proteome were assessed using a novel pulse-chase strategy (Fierro-Monti et al., accompanying article, detecting effects on both protein stability and synthesis. Global and specific dynamic impacts, including proteostatic responses, are due to direct inhibition of Hsp90 as well as indirect effects. As a result, a decrease was detected in most proteins that changed their levels, including known Hsp90 clients. Most likely, consequences of the role of Hsp90 in gene expression determined a global reduction in net de novo protein synthesis. This decrease appeared to be greater in magnitude than a concomitantly observed global increase in protein decay rates. Several novel putative Hsp90 clients were validated, and interestingly, protein families with critical functions, particularly the Hsp90 family and cofactors themselves as well as protein kinases, displayed strongly increased decay rates due to Hsp90 inhibitor treatment. Remarkably, an upsurge in survival pathways, involving molecular chaperones and several oncoproteins, and decreased levels of some tumour suppressors, have implications for anti-cancer therapy with Hsp90 inhibitors. The diversity of global effects may represent a paradigm of mechanisms that are operating to shield cells from proteotoxic stress, by promoting pro-survival and anti-proliferative functions. Data are available via

  20. Integrative Analysis of CRISPR/Cas9 Target Sites in the Human HBB Gene

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    Yumei Luo

    2015-01-01

    Full Text Available Recently, the clustered regularly interspaced short palindromic repeats (CRISPR system has emerged as a powerful customizable artificial nuclease to facilitate precise genetic correction for tissue regeneration and isogenic disease modeling. However, previous studies reported substantial off-target activities of CRISPR system in human cells, and the enormous putative off-target sites are labor-intensive to be validated experimentally, thus motivating bioinformatics methods for rational design of CRISPR system and prediction of its potential off-target effects. Here, we describe an integrative analytical process to identify specific CRISPR target sites in the human β-globin gene (HBB and predict their off-target effects. Our method includes off-target analysis in both coding and noncoding regions, which was neglected by previous studies. It was found that the CRISPR target sites in the introns have fewer off-target sites in the coding regions than those in the exons. Remarkably, target sites containing certain transcriptional factor motif have enriched binding sites of relevant transcriptional factor in their off-target sets. We also found that the intron sites have fewer SNPs, which leads to less variation of CRISPR efficiency in different individuals during clinical applications. Our studies provide a standard analytical procedure to select specific CRISPR targets for genetic correction.

  1. Integrative Analysis of CRISPR/Cas9 Target Sites in the Human HBB Gene.

    Science.gov (United States)

    Luo, Yumei; Zhu, Detu; Zhang, Zhizhuo; Chen, Yaoyong; Sun, Xiaofang

    2015-01-01

    Recently, the clustered regularly interspaced short palindromic repeats (CRISPR) system has emerged as a powerful customizable artificial nuclease to facilitate precise genetic correction for tissue regeneration and isogenic disease modeling. However, previous studies reported substantial off-target activities of CRISPR system in human cells, and the enormous putative off-target sites are labor-intensive to be validated experimentally, thus motivating bioinformatics methods for rational design of CRISPR system and prediction of its potential off-target effects. Here, we describe an integrative analytical process to identify specific CRISPR target sites in the human β-globin gene (HBB) and predict their off-target effects. Our method includes off-target analysis in both coding and noncoding regions, which was neglected by previous studies. It was found that the CRISPR target sites in the introns have fewer off-target sites in the coding regions than those in the exons. Remarkably, target sites containing certain transcriptional factor motif have enriched binding sites of relevant transcriptional factor in their off-target sets. We also found that the intron sites have fewer SNPs, which leads to less variation of CRISPR efficiency in different individuals during clinical applications. Our studies provide a standard analytical procedure to select specific CRISPR targets for genetic correction.

  2. Primer and interviews: advances in targeted gene modification. Interview by Julie C. Kiefer.

    Science.gov (United States)

    Caroll, Dana; Zhang, Bo

    2011-12-01

    Gene targeting in mice, first reported 25 years ago, has led to monumental advances in the understanding of basic biology and human disease. The ability to employ a similarly straightforward method for gene manipulation in other experimental organisms would make their already significant contributions all the more powerful. Here, we briefly outline the strengths and weaknesses of reverse genetics techniques in non-murine model organisms, ending with a more detailed description of two that promise to bring targeted gene modification to the masses: zinc finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs). Dana Caroll, a forefather of zinc finger technology, and Bo Zhang, among the first to introduce TALEN-targeted mutagenesis to zebrafish, discuss their experience with these techniques, and speculate about the future of the field.

  3. Efficient targeted gene disruption in Xenopus embryos using engineered transcription activator-like effector nucleases (TALENs).

    Science.gov (United States)

    Lei, Yong; Guo, Xiaogang; Liu, Yun; Cao, Yang; Deng, Yi; Chen, Xiongfeng; Cheng, Christopher H K; Dawid, Igor B; Chen, Yonglong; Zhao, Hui

    2012-10-23

    Transcription activator-like effector nucleases (TALENs) are an approach for directed gene disruption and have been proved to be effective in various animal models. Here, we report that TALENs can induce somatic mutations in Xenopus embryos with reliably high efficiency and that such mutations are heritable through germ-line transmission. We modified the Golden Gate method for TALEN assembly to make the product suitable for RNA transcription and microinjection into Xenopus embryos. Eight pairs of TALENs were constructed to target eight Xenopus genes, and all resulted in indel mutations with high efficiencies of up to 95.7% at the targeted loci. Furthermore, mutations induced by TALENs were highly efficiently passed through the germ line to F(1) frogs. Together with simple and reliable PCR-based approaches for detecting TALEN-induced mutations, our results indicate that TALENs are an effective tool for targeted gene editing/knockout in Xenopus.

  4. Identification and Characterization of Genes Involved in Leishmania Pathogenesis: The Potential for Drug Target Selection

    Directory of Open Access Journals (Sweden)

    Robert Duncan

    2011-01-01

    Full Text Available Identifying and characterizing Leishmania donovani genes and the proteins they encode for their role in pathogenesis can reveal the value of this approach for finding new drug targets. Effective drug targets are likely to be proteins differentially expressed or required in the amastigote life cycle stage found in the patient. Several examples and their potential for chemotherapeutic disruption are presented. A pathway nearly ubiquitous in living cells targeted by anticancer drugs, the ubiquitin system, is examined. New findings in ubiquitin and ubiquitin-like modifiers in Leishmania show how disruption of those pathways could point to additional drug targets. The programmed cell death pathway, now recognized among protozoan parasites, is reviewed for some of its components and evidence that suggests they could be targeted for antiparasitic drug therapy. Finally, the endoplasmic reticulum quality control system is involved in secretion of many virulence factors. How disruptions in this pathway reduce virulence as evidence for potential drug targets is presented.

  5. Mapping of HNF4alpha target genes in intestinal epithelial cells

    DEFF Research Database (Denmark)

    Boyd, Mette; Bressendorff, Simon; Moller, Jette;

    2009-01-01

    . The HNF4alpha ChIP-chip data was matched with gene expression and histone H3 acetylation status of the promoters in order to identify HNF4alpha binding to actively transcribed genes with an open chromatin structure. RESULTS: 1,541 genes were identified as potential HNF4alpha targets, many of which have...... not previously been described as being regulated by HNF4alpha. The 1,541 genes contributed significantly to gene ontology (GO) pathways categorized by lipid and amino acid transport and metabolism. An analysis of the homeodomain transcription factor Cdx-2 (CDX2), the disaccharidase trehalase (TREH...... a transcription factor network also including HNF1alpha, all of which are transcription factors involved in intestinal development and gene expression....

  6. Binding Sites of miR-1273 Family on the mRNA of Target Genes

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    Anatoly Ivashchenko

    2014-01-01

    Full Text Available This study examined binding sites of 2,578 miRNAs in the mRNAs of 12,175 human genes using the MirTarget program. It found that the miRNAs of miR-1273 family have between 33 and 1,074 mRNA target genes, with a free hybridization energy of 90% or more of its maximum value. The miR-1273 family consists of miR-1273a, miR-1273c, miR-1273d, miR-1273e, miR-1273f, miR-1273g-3p, miR-1273g-5p, miR-1273h-3p, and miR-1273h-5p. Unique miRNAs (miR-1273e, miR-1273f, and miR-1273g-3p have more than 400 target genes. We established 99 mRNA nucleotide sequences that contain arranged binding sites for the miR-1273 family. High conservation of each miRNA binding site in the mRNA of the target genes was found. The arranged binding sites of the miR-1273 family are located in the 5′UTR, CDS, or 3′UTR of many mRNAs. Five repeating sites containing some of the miR-1273 family’s binding sites were found in the 3′UTR of several target genes. The oligonucleotide sequences of miR-1273 binding sites located in CDSs code for homologous amino acid sequences in the proteins of target genes. The biological role of unique miRNAs was also discussed.

  7. Extracellular control of intracellular drug release for enhanced safety of anti-cancer chemotherapy

    Science.gov (United States)

    Zhu, Qian; Qi, Haixia; Long, Ziyan; Liu, Shang; Huang, Zhen; Zhang, Junfeng; Wang, Chunming; Dong, Lei

    2016-06-01

    The difficulty of controlling drug release at an intracellular level remains a key challenge for maximising drug safety and efficacy. We demonstrate herein a new, efficient and convenient approach to extracellularly control the intracellular release of doxorubicin (DOX), by designing a delivery system that harnesses the interactions between the system and a particular set of cellular machinery. By simply adding a small-molecule chemical into the cell medium, we could lower the release rate of DOX in the cytosol, and thereby increase its accumulation in the nuclei while decreasing its presence at mitochondria. Delivery of DOX with this system effectively prevented DOX-induced mitochondria damage that is the main mechanism of its toxicity, while exerting the maximum efficacy of this anti-cancer chemotherapeutic agent. The present study sheds light on the design of drug delivery systems for extracellular control of intracellular drug delivery, with immediate therapeutic implications.

  8. Gaojushen:a novel anti-cancer drug prepared from SEC superantigen

    Institute of Scientific and Technical Information of China (English)

    陈廷祚

    2005-01-01

    @@ 1 Clinical observations Gaojushen is a novel anti-cancer drug developed by Xiehe Bio-pharmaceutical Company,Shenyang, China. It is prepared and processed from the filtrate of Staphylococcus aureus culture. The active component contained in it has been shown to be a SEC superantigen that is a metabolite of the culture.This superantigen is marked by its ability to stimulate T cells at a high frequency, thereby giving rise to potent cell-mediated immunological responses and producing a large variety of cytokines with the final rsult of apoptosis of tumor cells. The drug was approved for trial prodoction in 1994 by the Center of the State Evaluation and Review of New Drugs,China,and was licenced for marketing by 1996 after finishing the phase III clinical trial.

  9. Anti-cancer potential of banana flower extract: An in vitro study

    Directory of Open Access Journals (Sweden)

    Varalakshmi Kilingar Nadumane

    2014-12-01

    Full Text Available Banana (Musa paradisiaca flower is rich in phytochemicals (vitamins, flavonoids, proteins and has antioxidant properties. The anti-cancer activity of banana flower extract has been evaluated on the cervical cancer cell line HeLa. The antiproliferative effects were evaluated by MTT assay. The extract was further purified by TLC and characterized by LC-MS method. The ethanol extract had significant cytotoxicity to HeLa cells with an IC50 of 20 µg/mL. By thin layer chromatography we could isolate three fractions out of which fraction 2 had exhibited maximum anti-proliferative effects with an IC50 value of <10 µg/mL. By LC-MS analysis, bioactive fraction was found to have an m/z value of 224.2 indicating it as a novel one.

  10. NOVEL HYDROXAMIC ACIDS HAVING HISTONE DEACETYLASE INHIBITING ACTIVITY AND ANTI-CANCER COMPOSITION COMPRISING THE SAME AS AN ACTIVE INGREDIENT

    DEFF Research Database (Denmark)

    2013-01-01

    The present invention relates to a pharmaceutical composition for anticancer including novel hydroxamic acid with histone deacetylase inhibiting activity as an active ingredient. Hydroxamic acid compound of the present invention has inhibitory activity of histone deacetylase (HDAC) and shows cyto...... cytotoxicity to a variety of cancer cells, being useful in strong anti-cancer drug....

  11. Liquid Chromatography - Triple Quadrupole Mass Spectrometry : The gold standard for quantitative bioanalysis of anti-cancer agents

    NARCIS (Netherlands)

    Vainchtein, L.D.

    2008-01-01

    To understand the pharmacologic mechanisms of action, efficacy and toxicity of any anti-cancer drug it is important to know how the compound is transformed in the body: either into active metabolites or inactive and toxic (degradation) products. This information may lead to the success or failure of

  12. Comparison of the cancer gene targeting and biochemical selectivities of all targeted kinase inhibitors approved for clinical use.

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    Joost C M Uitdehaag

    Full Text Available The anti-proliferative activities of all twenty-five targeted kinase inhibitor drugs that are in clinical use were measured in two large assay panels: (1 a panel of proliferation assays of forty-four human cancer cell lines from diverse tumour tissue origins; and (2 a panel of more than 300 kinase enzyme activity assays. This study provides a head-on comparison of all kinase inhibitor drugs in use (status Nov. 2013, and for six of these drugs, the first kinome profiling data in the public domain. Correlation of drug activities with cancer gene mutations revealed novel drug sensitivity markers, suggesting that cancers dependent on mutant CTNNB1 will respond to trametinib and other MEK inhibitors, and cancers dependent on SMAD4 to small molecule EGFR inhibitor drugs. Comparison of cellular targeting efficacies reveals the most targeted inhibitors for EGFR, ABL1 and BRAF(V600E-driven cell growth, and demonstrates that the best targeted agents combine high biochemical potency with good selectivity. For ABL1 inhibitors, we computationally deduce optimized kinase profiles for use in a next generation of drugs. Our study shows the power of combining biochemical and cellular profiling data in the evaluation of kinase inhibitor drug action.

  13. Characterization of Rad51 from apicomplexan parasite Toxoplasma gondii: an implication for inefficient gene targeting.

    Science.gov (United States)

    Achanta, Sita Swati; Varunan, Shalu M; Bhattacharyya, Sunanda; Bhattacharyya, Mrinal Kanti

    2012-01-01

    Repairing double strand breaks (DSBs) is absolutely essential for the survival of obligate intracellular parasite Toxoplasma gondii. Thus, DSB repair mechanisms could be excellent targets for chemotherapeutic interventions. Recent genetic and bioinformatics analyses confirm the presence of both homologous recombination (HR) as well as non homologous end joining (NHEJ) proteins in this lower eukaryote. In order to get mechanistic insights into the HR mediated DSB repair pathway in this parasite, we have characterized the key protein involved in homologous recombination, namely TgRad51, at the biochemical and genetic levels. We have purified recombinant TgRad51 protein to 99% homogeneity and have characterized it biochemically. The ATP hydrolysis activity of TgRad51 shows a higher K(M) and much lower k(cat) compared to bacterial RecA or Rad51 from other related protozoan parasites. Taking yeast as a surrogate model system we have shown that TgRad51 is less efficient in gene conversion mechanism. Further, we have found that TgRad51 mediated gene integration is more prone towards random genetic loci rather than targeted locus. We hypothesize that compromised ATPase activity of TgRad51 is responsible for inefficient gene targeting and poor gene conversion efficiency in this protozoan parasite. With increase in homologous flanking regions almost three fold increments in targeted gene integration is observed, which is similar to the trend found with ScRad51. Our findings not only help us in understanding the reason behind inefficient gene targeting in T. gondii but also could be exploited to facilitate high throughput knockout as well as epitope tagging of Toxoplasma genes.

  14. Characterization of Rad51 from apicomplexan parasite Toxoplasma gondii: an implication for inefficient gene targeting.

    Directory of Open Access Journals (Sweden)

    Sita Swati Achanta

    Full Text Available Repairing double strand breaks (DSBs is absolutely essential for the survival of obligate intracellular parasite Toxoplasma gondii. Thus, DSB repair mechanisms could be excellent targets for chemotherapeutic interventions. Recent genetic and bioinformatics analyses confirm the presence of both homologous recombination (HR as well as non homologous end joining (NHEJ proteins in this lower eukaryote. In order to get mechanistic insights into the HR mediated DSB repair pathway in this parasite, we have characterized the key protein involved in homologous recombination, namely TgRad51, at the biochemical and genetic levels. We have purified recombinant TgRad51 protein to 99% homogeneity and have characterized it biochemically. The ATP hydrolysis activity of TgRad51 shows a higher K(M and much lower k(cat compared to bacterial RecA or Rad51 from other related protozoan parasites. Taking yeast as a surrogate model system we have shown that TgRad51 is less efficient in gene conversion mechanism. Further, we have found that TgRad51 mediated gene integration is more prone towards random genetic loci rather than targeted locus. We hypothesize that compromised ATPase activity of TgRad51 is responsible for inefficient gene targeting and poor gene conversion efficiency in this protozoan parasite. With increase in homologous flanking regions almost three fold increments in targeted gene integration is observed, which is similar to the trend found with ScRad51. Our findings not only help us in understanding the reason behind inefficient gene targeting in T. gondii but also could be exploited to facilitate high throughput knockout as well as epitope tagging of Toxoplasma genes.

  15. Targeted gene correction using psoralen, chlorambucil and camptothecin conjugates of triplex forming peptide nucleic acid (PNA)

    DEFF Research Database (Denmark)

    Birkedal, Henrik; Nielsen, Peter E

    2011-01-01

    Gene correction activation effects of a small series of triplex forming peptide nucleic acid (PNA) covalently conjugated to the DNA interacting ligands psoralen, chlorambucil and camptothecin targeted proximal to a stop codon mutation in an EGFP reporter gene were studied. A 15-mer homopyrimidine...... interstrand crosslinked adducts with dsDNA dramatically decreased the frequency of targeted repair/correction. The PNA conjugates were also studied in mammalian cell lines upon transfection of PNA bound EGFP reporter vector and scoring repair of the EGFP gene by FACS analysis of functional EGFP expression...... suggest that simple triplex forming PNAs have little effect on proximal gene correctional events whereas PNA conjugates capable of forming DNA adducts and interstrand crosslinks are strong inhibitors. Most interestingly the PNA conjugated to the topoisomerase inhibitor, camptothecin enhanced repair...

  16. NFAT targets signaling molecules to gene promoters in pancreatic β-cells.

    Science.gov (United States)

    Lawrence, Michael C; Borenstein-Auerbach, Nofit; McGlynn, Kathleen; Kunnathodi, Faisal; Shahbazov, Rauf; Syed, Ilham; Kanak, Mazhar; Takita, Morihito; Levy, Marlon F; Naziruddin, Bashoo

    2015-02-01

    Nuclear factor of activated T cells (NFAT) is activated by calcineurin in response to calcium signals derived by metabolic and inflammatory stress to regulate genes in pancreatic islets. Here, we show that NFAT targets MAPKs, histone acetyltransferase p300, and histone deacetylases (HDACs) to gene promoters to differentially regulate insulin and TNF-α genes. NFAT and ERK associated with the insulin gene promoter in response to glucagon-like peptide 1, whereas NFAT formed complexes with p38 MAPK (p38) and Jun N-terminal kinase (JNK) upon promoters of the TNF-α gene in response to IL-1β. Translocation of NFAT and MAPKs to gene promoters was calcineurin/NFAT dependent, and complex stability required MAPK activity. Knocking down NFATc2 expression, eliminating NFAT DNA binding sites, or interfering with NFAT nuclear import prevented association of MAPKs with gene promoters. Inhibiting p38 and JNK activity increased NFAT-ERK association with promoters, which repressed TNF-α and enhanced insulin gene expression. Moreover, inhibiting p38 and JNK induced a switch from NFAT-p38/JNK-histone acetyltransferase p300 to NFAT-ERK-HDAC3 complex formation upon the TNF-α promoter, which resulted in gene repression. Histone acetyltransferase/HDAC exchange was reversed on the insulin gene by p38/JNK inhibition in the presence of glucagon-like peptide 1, which enhanced gene expression. Overall, these data indicate that NFAT directs signaling enzymes to gene promoters in islets, which contribute to protein-DNA complex stability and promoter regulation. Furthermore, the data suggest that TNF-α can be repressed and insulin production can be enhanced by selectively targeting signaling components of NFAT-MAPK transcriptional/signaling complex formation in pancreatic β-cells. These findings have therapeutic potential for suppressing islet inflammation while preserving islet function in diabetes and islet transplantation.

  17. AKTIVITAS ANTI KANKER SENYAWA-SENYAWA KITOOLIGOMER [Anti Cancer Activity of Chitooligomers

    Directory of Open Access Journals (Sweden)

    Dahrul Syah2

    2006-04-01

    Full Text Available The chitin obtained from the crab industries can be used as a source for production of chitooligomers which has an important biological activity. The aims of this research was to evaluate anti cancer activity of the chitooligomers obtained from enzymatic hydrolysis using chitosanase from thermophilic bacterium Bacillus licheniformis MB2 isolated from Tompaso Manado. Media for producing the enzyme contained colloidal chitosan 1% and the enzyme was harvested after seven days of incubation at 550C. The heat stable protein enzyme was coagulated with 80% saturated ammonium sulphate and purificated using hydrophobic interaction chromatography with butyl sepharose gel. Enzyme of 0.005, 0.0085, 0.10 dan 0,17 IU/mg chitosan on soluble chitosan 1% substrate with 85% degree of deacylation were used to produce chitooligomers through incubation for one and three hours. The reaction products were analyzed (and fractionated using HPLC. The effect of this samples on cancer cells was evaluated using K562 cells (chronic myelogenous leukemia and investigated after being treated with MTT (3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide. In general, hydrolysates and fractionated chitooligomers showed better anti cancer activity than the 2- Bromo deoxy uridine used as positive control at similiar concentration (17 ?g/ml. Both of hydrolysates and fractionated chitooligomers (trimer to hexamer inhibited proliferation of human K562 cancer cells line in vitro about 20.57% and 15.68% respectively.The apoptosis phenomena was found on K562 cells treated with chitooligomer hydrolysate which can be examined by Hoechts staining fluorescent method. Chitooligomers hydrolysate showed anti metastatic potential, the chitooligomers were found also as potent protease inhibitor.

  18. Gene Regulatory Scenarios of Primary 1,25-Dihydroxyvitamin D3 Target Genes in a Human Myeloid Leukemia Cell Line

    Directory of Open Access Journals (Sweden)

    Moray J. Campbell

    2013-10-01

    Full Text Available Genome- and transcriptome-wide data has significantly increased the amount of available information about primary 1,25-dihydroxyvitamin D3 (1,25(OH2D3 target genes in cancer cell models, such as human THP-1 myelomonocytic leukemia cells. In this study, we investigated the genes G0S2, CDKN1A and MYC as master examples of primary vitamin D receptor (VDR targets being involved in the control of cellular proliferation. The chromosomal domains of G0S2 and CDKN1A are 140–170 kb in size and contain one and three VDR binding sites, respectively. This is rather compact compared to the MYC locus that is 15 times larger and accommodates four VDR binding sites. All eight VDR binding sites were studied by chromatin immunoprecipitation in THP-1 cells. Interestingly, the site closest to the transcription start site of the down-regulated MYC gene showed 1,25(OH2D3-dependent reduction of VDR binding and is not associated with open chromatin. Four of the other seven VDR binding regions contain a typical DR3-type VDR binding sequence, three of which are also occupied with VDR in macrophage-like cells. In conclusion, the three examples suggest that each VDR target gene has an individual regulatory scenario. However, some general components of these scenarios may be useful for the development of new therapy regimens.

  19. An efficient strategy for gene targeting and phenotypic assessment in the Plasmodium yoelii rodent malaria model.

    Science.gov (United States)

    Mikolajczak, Sebastian A; Aly, Ahmed S I; Dumpit, Ronald F; Vaughan, Ashley M; Kappe, Stefan H I

    2008-04-01

    In this report, we describe a cloning procedure for gene replacement by double homologous recombination in Plasmodium yoelii, which requires only one digestion and ligation step. This significantly shortens the time required to complete the production of the targeting vector. Furthermore, for more efficient phenotypic evaluation of the gene knockout parasites, we have also introduced a fluorescent protein cassette into the targeting vector. This allows for a more rapid assessment of parasite growth in all of its developmental stages. In addition, the introduction of the fluorescent marker via the replacement strategy confers the stable integration of the marker.

  20. Genome-wide mapping of Polycomb target genes unravels their roles in cell fate transitions

    DEFF Research Database (Denmark)

    Bracken, Adrian P; Dietrich, Nikolaj; Pasini, Diego;

    2006-01-01

    The Polycomb group (PcG) proteins form chromatin-modifying complexes that are essential for embryonic development and stem cell renewal and are commonly deregulated in cancer. Here, we identify their target genes using genome-wide location analysis in human embryonic fibroblasts. We find that Pol......The Polycomb group (PcG) proteins form chromatin-modifying complexes that are essential for embryonic development and stem cell renewal and are commonly deregulated in cancer. Here, we identify their target genes using genome-wide location analysis in human embryonic fibroblasts. We find...

  1. Cooperative interactions between CBP and TORC2 confer selectivity to CREB target gene expression

    DEFF Research Database (Denmark)

    Ravnskjær, Kim; Kester, Henri; Liu, Yi;

    2007-01-01

    , but have minimal effects on CRE-dependent transcription. Here, we show that the latent cytoplasmic coactivator TORC2 mediates target gene activation in response to cAMP signaling by associating with CBP/p300 and increasing its recruitment to a subset of CREB target genes. TORC2 is not activated in response...... to stress signals, however; and in its absence, P-CREB is unable to stimulate CRE-dependent transcription, due to a block in CBP recruitment. The effect of TORC2 on CBP/p300 promoter occupancy appears pivotal because a gain of function mutant CREB polypeptide with increased affinity for CBP restored CRE...

  2. Integrated functional, gene expression and genomic analysis for the identification of cancer targets.

    Directory of Open Access Journals (Sweden)

    Elizabeth Iorns

    Full Text Available The majority of new drug approvals for cancer are based on existing therapeutic targets. One approach to the identification of novel targets is to perform high-throughput RNA interference (RNAi cellular viability screens. We describe a novel approach combining RNAi screening in multiple cell lines with gene expression and genomic profiling to identify novel cancer targets. We performed parallel RNAi screens in multiple cancer cell lines to identify genes that are essential for viability in some cell lines but not others, suggesting that these genes constitute key drivers of cellular survival in specific cancer cells. This approach was verified by the identification of PIK3CA, silencing of which was selectively lethal to the MCF7 cell line, which harbours an activating oncogenic PIK3CA mutation. We combined our functional RNAi approach with gene expression and genomic analysis, allowing the identification of several novel kinases, including WEE1, that are essential for viability only in cell lines that have an elevated level of expression of this kinase. Furthermore, we identified a subset of breast tumours that highly express WEE1 suggesting that WEE1 could be a novel therapeutic target in breast cancer. In conclusion, this strategy represents a novel and effective strategy for the identification of functionally important therapeutic targets in cancer.

  3. Enteropeptidase: a gene associated with a starvation human phenotype and a novel target for obesity treatment.

    Directory of Open Access Journals (Sweden)

    Sandrine Braud

    Full Text Available BACKGROUND: Obesity research focuses essentially on gene targets associated with the obese phenotype. None of these targets have yet provided a viable drug therapy. Focusing instead on genes that are involved in energy absorption and that are associated with a "human starvation phenotype", we have identified enteropeptidase (EP, a gene associated with congenital enteropeptidase deficiency, as a novel target for obesity treatment. The advantages of this target are that the gene is expressed exclusively in the brush border of the intestine; it is peripheral and not redundant. METHODOLOGY/PRINCIPAL FINDINGS: Potent and selective EP inhibitors were designed around a boroarginine or borolysine motif. Oral administration of these compounds to mice restricted the bioavailability of dietary energy, and in a long-term treatment it significantly diminished the rate of increase in body weight, despite ad libitum food intake. No adverse reactions of the type seen with lipase inhibitors, such as diarrhea or steatorrhea, were observed. This validates EP as a novel, druggable target for obesity treatment. CONCLUSIONS: In vivo testing of novel boroarginine or borolysine-based EP inhibitors validates a novel approach to the treatment of obesity.

  4. Identifying gene targets for the metabolic engineering of lycopene biosynthesis in Escherichia coli.

    Science.gov (United States)

    Alper, Hal; Jin, Yong-Su; Moxley, J F; Stephanopoulos, G

    2005-05-01

    The identification of genetic targets that are effective in bringing about a desired phenotype change is still an open problem. While random gene knockouts have yielded improved strains in certain cases, it is also important to seek the guidance of cell-wide stoichiometric constraints in identifying promising gene knockout targets. To investigate these issues, we undertook a genome-wide stoichiometric flux balance analysis as an aid in discovering putative genes impacting network properties and cellular phenotype. Specifically, we calculated metabolic fluxes such as to optimize growth and then scanned the genome for single and multiple gene knockouts that yield improved product yield while maintaining acceptable overall growth rate. For the particular case of lycopene biosynthesis in Escherichia coli, we identified such targets that we subsequently tested experimentally by constructing the corresponding single, double and triple gene knockouts. While such strains are suggested (by the stoichiometric calculations) to increase precursor availability, this beneficial effect may be further impacted by kinetic and regulatory effects not captured by the stoichiometric model. For the case of lycopene biosynthesis, the so identified knockout targets yielded a triple knockout construct that exhibited a nearly 40% increase over an engineered, high producing parental strain.

  5. Androgen Receptor-Target Genes in African American Prostate Cancer Disparities

    Directory of Open Access Journals (Sweden)

    Bi-Dar Wang

    2013-01-01

    Full Text Available The incidence and mortality rates of prostate cancer (PCa are higher in African American (AA compared to Caucasian American (CA men. To elucidate the molecular mechanisms underlying PCa disparities, we employed an integrative approach combining gene expression profiling and pathway and promoter analyses to investigate differential transcriptomes and deregulated signaling pathways in AA versus CA cancers. A comparison of AA and CA PCa specimens identified 1,188 differentially expressed genes. Interestingly, these transcriptional differences were overrepresented in signaling pathways that converged on the androgen receptor (AR, suggesting that the AR may be a unifying oncogenic theme in AA PCa. Gene promoter analysis revealed that 382 out of 1,188 genes contained cis-acting AR-binding sequences. Chromatin immunoprecipitation confirmed STAT1, RHOA, ITGB5, MAPKAPK2, CSNK2A,1 and PIK3CB genes as novel AR targets in PCa disparities. Moreover, functional screens revealed that androgen-stimulated AR binding and upregulation of RHOA, ITGB5, and PIK3CB genes were associated with increased invasive activity of AA PCa cells, as siRNA-mediated knockdown of each gene caused a loss of androgen-stimulated invasion. In summation, our findings demonstrate that transcriptional changes have preferentially occurred in multiple signaling pathways converging (“transcriptional convergence” on AR signaling, thereby contributing to AR-target gene activation and PCa aggressiveness in AAs.

  6. Multifunctional Virus-Nanoshell Assembly for Targeted Hyperthermia and Viral Gene Therapy for Breast Cancer

    Science.gov (United States)

    2012-06-01

    cancer cells in synergy with gene therapy. We proposed to develop virus- nanoshell assemblies by attaching adeno-associated virus (AAV) to gold... nanoshells (Au NS) through chemical bonds. We have successfully completed majority of tasks 1 and 2 of our Statement of Work. Specifically, we have...therapy, virus, Au nanoshell Multifunctional Virus- Nanoshell Assembly for Targeted Hyperthermia and Viral Gene Therapy for Breast Cancer Dr. Fang Wei

  7. Targeted Editing of Myostatin Gene in Sheep by Transcription Activator-like Effector Nucleases

    OpenAIRE

    Zhao, Xinxia; Ni, Wei; Chen, Chuangfu; Sai, Wujiafu; Qiao, Jun; Sheng, Jingliang; Zhang, Hui; Li, Guozhong; Wang, Dawei; Hu, Shengwei

    2016-01-01

    Myostatin (MSTN) is a secreted growth factor expressed in skeletal muscle and adipose tissue that negatively regulates skeletal muscle mass. Gene knockout of MSTN can result in increasing muscle mass in sheep. The objectives were to investigate whether myostatin gene can be edited in sheep by transcription activator-like effector nucleases (TALENs) in tandem with single-stranded DNA oligonucleotides (ssODNs). We designed a pair of TALENs to target a highly conserved sequence in the coding reg...

  8. Validation and target gene screening of hsa-miR-205 in lung squamous cell carcinoma

    Institute of Scientific and Technical Information of China (English)

    Huang Wei; Jin Yi; Yuan Yunfeng; Bai Chunxue; Wu Ying; Zhu Hongguang; Lu Shaohua

    2014-01-01

    Background Lung cancers are classified as squamous cell carcinoma (SQ),adenocarcinoma (AC) and small cell lung carcinoma (SCLC).SQ is the major subtype of lung cancer.Currently,there are no targeted therapies for SQ due to lack of understanding its driving oncogenes.In this study,we validated an SQ specific biomarker hsa-miR-205 in Chinese patients with lung cancer and screened its candidate target genes for further functional studies to enrich knowledge in SQ target therapies.Methods Quantitative reverse-transcription PCR (quantitative RT-PCR)was performed on 197 macro-dissected (cancerous cells >75%) surgical lung tissues (45 SQ,44 AC,54 SCLC and 54 adjacent normal tissues) to validate the expression profiles of miR-205.Furthermore,the targets of this microRNA were predicted through the gateway miRecords and mapped to lung cancer-associated pathways using the KEGG (Kyoto Encyclopedia of Genes and Genomes) database.Then quantitative RT-PCR was performed on an independent cohort of 44 snap-frozen surgical lung tissues to concurrently assess the expression profiles of miR-205 and its 52 putative targeted genes.Results MicroRNA-205 yielded high diagnostic accuracy in discriminating SQ from AC with an area under the curve (AUC) of 0.985,and discriminating SQ from SCLC with an AUC of 0.978 in formalin-fixed paraffin-embedded (FFPE)surgical lung tissues.Predicted targets of miR-205 were associated with 52 key members of lung cancer signaling pathways.Ten target genes (ACSL1,AXIN2,CACNA2D2,FOXO3,PPP1R3A,PRKAG3,RUNX1,SMAD4,STK3 and TBL1XR1) were significantly down-regulated in SQ and had a strong negative correlation with miR-205,while one target gene (CDH3) was up-regulated in SQ and exhibited a strong positive correlation with miR-205.Conclusions We confirmed the high diagnostic accuracy of miR-205 in discriminating SQ from AC and SCLC in Chinese patients.Moreover,we identified 11 significant target genes of miR-205 which could be used for further functional studies

  9. Efficient gene targeting in golden Syrian hamsters by the CRISPR/Cas9 system.

    Directory of Open Access Journals (Sweden)

    Zhiqiang Fan

    Full Text Available The golden Syrian hamster is the model of choice or the only rodent model for studying many human diseases. However, the lack of gene targeting tools in hamsters severely limits their use in biomedical research. Here, we report the first successful application of the CRISPR/Cas9 system to efficiently conduct gene targeting in hamsters. We designed five synthetic single-guide RNAs (sgRNAs--three for targeting the coding sequences for different functional domains of the hamster STAT2 protein, one for KCNQ1, and one for PPP1R12C--and demonstrated that the CRISPR/Cas9 system is highly efficient in introducing site-specific mutations in hamster somatic cells. We then developed unique pronuclear (PN and cytoplasmic injection protocols in hamsters and produced STAT2 knockout (KO hamsters by injecting the sgRNA/Cas9, either in the form of plasmid or mRNA, targeting exon 4 of hamster STAT2. Among the produced hamsters, 14.3% and 88.9% harbored germline-transmitted STAT2 mutations from plasmid and mRNA injection, respectively. Notably, 10.4% of the animals produced from mRNA injection were biallelically targeted. This is the first success in conducting site-specific gene targeting in hamsters and can serve as the foundation for developing other genetically engineered hamster models for human disease.

  10. Targeted disruption of Ataxia-telangiectasia mutated gene in miniature pigs by somatic cell nuclear transfer

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Young June; Ahn, Kwang Sung; Kim, Minjeong; Kim, Min Ju; Park, Sang-Min; Ryu, Junghyun; Ahn, Jin Seop; Heo, Soon Young; Kang, Jee Hyun; Choi, You Jung [Department of Nanobiomedical Science and BK21 PLUS NBM Global Research Center for Regenerative Medicine, Dankook University, Cheonan (Korea, Republic of); Choi, Seong-Jun [Institute of Tissue Regeneration Engineering, Dankook University, Cheonan (Korea, Republic of); Shim, Hosup, E-mail: shim@dku.edu [Department of Nanobiomedical Science and BK21 PLUS NBM Global Research Center for Regenerative Medicine, Dankook University, Cheonan (Korea, Republic of); Institute of Tissue Regeneration Engineering, Dankook University, Cheonan (Korea, Republic of); Department of Physiology, Dankook University School of Medicine, Cheonan (Korea, Republic of)

    2014-10-03

    Highlights: • ATM gene-targeted pigs were produced by somatic cell nuclear transfer. • A novel large animal model for ataxia telangiectasia was developed. • The new model may provide an alternative to the mouse model. - Abstract: Ataxia telangiectasia (A-T) is a recessive autosomal disorder associated with pleiotropic phenotypes, including progressive cerebellar degeneration, gonad atrophy, and growth retardation. Even though A-T is known to be caused by the mutations in the Ataxia telangiectasia mutated (ATM) gene, the correlation between abnormal cellular physiology caused by ATM mutations and the multiple symptoms of A-T disease has not been clearly determined. None of the existing ATM mouse models properly reflects the extent to which neurological degeneration occurs in human. In an attempt to provide a large animal model for A-T, we produced gene-targeted pigs with mutations in the ATM gene by somatic cell nuclear transfer. The disrupted allele in the ATM gene of cloned piglets was confirmed via PCR and Southern blot analysis. The ATM gene-targeted pigs generated in the present study may provide an alternative to the current mouse model for the study of mechanisms underlying A-T disorder and for the development of new therapies.

  11. RNA polymerase V targets transcriptional silencing components to promoters of protein-coding genes.

    Science.gov (United States)

    Zheng, Qi; Rowley, M Jordan; Böhmdorfer, Gudrun; Sandhu, Davinder; Gregory, Brian D; Wierzbicki, Andrzej T

    2013-01-01

    Transcriptional gene silencing controls transposons and other repetitive elements through RNA-directed DNA methylation (RdDM) and heterochromatin formation. A key component of the Arabidopsis RdDM pathway is ARGONAUTE4 (AGO4), which associates with siRNAs to mediate DNA methylation. Here, we show that AGO4 preferentially targets transposable elements embedded within promoters of protein-coding genes. This pattern of AGO4 binding cannot be simply explained by the sequences of AGO4-bound siRNAs; instead, AGO4 binding to specific gene promoters is also mediated by long non-coding RNAs (lncRNAs) produced by RNA polymerase V. lncRNA-mediated AGO4 binding to gene promoters directs asymmetric DNA methylation to these genomic regions and is involved in regulating the expression of targeted genes. Finally, AGO4 binding overlaps sites of DNA methylation affected by the biotic stress response. Based on these findings, we propose that the targets of AGO4-directed RdDM are regulatory units responsible for controlling gene expression under specific environmental conditions.

  12. Targeted gene transfer into rat facial muscles by nanosecond pulsed laser-induced stress waves

    Science.gov (United States)

    Kurita, Akihiro; Matsunobu, Takeshi; Satoh, Yasushi; Ando, Takahiro; Sato, Shunichi; Obara, Minoru; Shiotani, Akihiro

    2011-09-01

    We investigate the feasibility of using nanosecond pulsed laser-induced stress waves (LISWs) for gene transfer into rat facial muscles. LISWs are generated by irradiating a black natural rubber disk placed on the target tissue with nanosecond pulsed laser light from the second harmonics (532 nm) of a Q-switched Nd:YAG laser, which is widely used in head and neck surgery and proven to be safe. After injection of plasmid deoxyribose nucleic acid (DNA) coding for Lac Z into rat facial muscles, pulsed laser is used to irradiate the laser target on the skin surface without incision or exposure of muscles. Lac Z expression is detected by X-gal staining of excised rat facial skin and muscles. Strong Lac Z expression is observed seven days after gene transfer, and sustained for up to 14 days. Gene transfer is achieved in facial muscles several millimeters deep from the surface. Gene expression is localized to the tissue exposed to LISWs. No tissue damage from LISWs is observed. LISW is a promising nonviral target gene transfer method because of its high spatial controllability, easy applicability, and minimal invasiveness. Gene transfer using LISW to produce therapeutic proteins such as growth factors could be used to treat nerve injury and paralysis.

  13. Functional characterization of endogenous siRNA target genes in Caenorhabditis elegans

    Directory of Open Access Journals (Sweden)

    Heikkinen Liisa

    2008-06-01

    Full Text Available Abstract Background Small interfering RNA (siRNA molecules mediate sequence specific silencing in RNA interference (RNAi, a gene regulatory phenomenon observed in almost all organisms. Large scale sequencing of small RNA libraries obtained from C. elegans has revealed that a broad spectrum of siRNAs is endogenously transcribed from genomic sequences. The biological role and molecular diversity of C. elegans endogenous siRNA (endo-siRNA molecules, nonetheless, remain poorly understood. In order to gain insight into their biological function, we annotated two large libraries of endo-siRNA sequences, identified their cognate targets, and performed gene ontology analysis to identify enriched functional categories. Results Systematic trends in categorization of target genes according to the specific length of siRNA sequences were observed: 18- to 22-mer siRNAs were associated with genes required for embryonic development; 23-mers were associated uniquely with post-embryonic development; 24–26-mers were associated with phosphorus metabolism or protein modification. Moreover, we observe that some argonaute related genes associate with siRNAs with multiple reads. Sequence frequency graphs suggest that different lengths of siRNAs share similarities in overall sequence structure: the 5' end begins with G, while the body predominates with U and C. Conclusion These results suggest that the lengths of endogenous siRNA molecules are consequential to their biological functions since the gene ontology categories for their cognate mRNA targets vary depending upon their lengths.

  14. Targeted delivery of genes to endothelial cells and cell- and gene-based therapy in pulmonary vascular diseases.

    Science.gov (United States)

    Suen, Colin M; Mei, Shirley H J; Kugathasan, Lakshmi; Stewart, Duncan J

    2013-10-01

    Pulmonary arterial hypertension (PAH) is a devastating disease that, despite significant advances in medical therapies over the last several decades, continues to have an extremely poor prognosis. Gene therapy is a method to deliver therapeutic genes to replace defective or mutant genes or supplement existing cellular processes to modify disease. Over the last few decades, several viral and nonviral methods of gene therapy have been developed for preclinical PAH studies with varying degrees of efficacy. However, these gene delivery methods face challenges of immunogenicity, low transduction rates, and nonspecific targeting which have limited their translation to clinical studies. More recently, the emergence of regenerative approaches using stem and progenitor cells such as endothelial progenitor cells (EPCs) and mesenchymal stem cells (MSCs) have offered a new approach to gene therapy. Cell-based gene therapy is an approach that augments the therapeutic potential of EPCs and MSCs and may deliver on the promise of reversal of established PAH. These new regenerative approaches have shown tremendous potential in preclinical studies; however, large, rigorously designed clinical studies will be necessary to evaluate clinical efficacy and safety.

  15. A meta analysis of pancreatic microarray datasets yields new targets as cancer genes and biomarkers.

    Directory of Open Access Journals (Sweden)

    Nalin C W Goonesekere

    Full Text Available The lack of specific symptoms at early tumor stages, together with a high biological aggressiveness of the tumor contribute to the high mortality rate for pancreatic cancer (PC, which has a five year survival rate of less than 5%. Improved screening for earlier diagnosis, through the detection of diagnostic and prognostic biomarkers provides the best hope of increasing the rate of curatively resectable carcinomas. Though many serum markers have been reported to be elevated in patients with PC, so far, most of these markers have not been implemented into clinical routine due to low sensitivity or specificity. In this study, we have identified genes that are significantly upregulated in PC, through a meta-analysis of large number of microarray datasets. We demonstrate that the biological functions ascribed to these genes are clearly associated with PC and metastasis, and that that these genes exhibit a strong link to pathways involved with inflammation and the immune response. This investigation has yielded new targets for cancer genes, and potential biomarkers for pancreatic cancer. The candidate list of cancer genes includes protein kinase genes, new members of gene families currently associated with PC, as well as genes not previously linked to PC. In this study, we are also able to move towards developing a signature for hypomethylated genes, which could be useful for early detection of PC. We also show that the significantly upregulated 800+ genes in our analysis can serve as an enriched pool for tissue and serum protein biomarkers in pancreatic cancer.

  16. PKA-chromatin association at stress responsive target genes from Saccharomyces cerevisiae.

    Science.gov (United States)

    Baccarini, Leticia; Martínez-Montañés, Fernando; Rossi, Silvia; Proft, Markus; Portela, Paula

    2015-11-01

    Gene expression regulation by intracellular stimulus-activated protein kinases is essential for cell adaptation to environmental changes. There are three PKA catalytic subunits in Saccharomyces cerevisiae: Tpk1, Tpk2, and Tpk3 and one regulatory subunit: Bcy1. Previously, it has been demonstrated that Tpk1 and Tpk2 are associated with coding regions and promoters of target genes in a carbon source and oxidative stress dependent manner. Here we studied five genes, ALD6, SED1, HSP42, RPS29B, and RPL1B whose expression is regulated by saline stress. We found that PKA catalytic and regulatory subunits are associated with both coding regions and promoters of the analyzed genes in a stress dependent manner. Tpk1 and Tpk2 recruitment was completely abolished in catalytic inactive mutants. BCY1 deletion changed the binding kinetic to chromatin of each Tpk isoform and this strain displayed a deregulated gene expression in response to osmotic stress. In addition, yeast mutants with high PKA activity exhibit sustained association to target genes of chromatin-remodeling complexes such as Snf2-catalytic subunit of the SWI/SNF complex and Arp8-component of INO80 complex, leading to upregulation of gene expression during osmotic stress. Tpk1 accumulation in the nucleus was stimulated upon osmotic stress, while the nuclear localization of Tpk2 and Bcy1 showed no change. We found that each PKA subunit is transported into the nucleus by a different β-karyopherin pathway. Moreover, β-karyopherin mutant strains abolished the chromatin association of Tpk1 or Tpk2, suggesting that nuclear localization of PKA catalytic subunits is required for its association to target genes and properly gene expression.

  17. Gene Therapy of Cancerous Diseases

    OpenAIRE

    Valenčáková, A.; Dziaková, A.; Hatalová, E.

    2015-01-01

    Gene therapy of cancerous diseases provides new means of curing patients with oncologic illnesses. There are several approaches in treating cancer by gene therapy. Most commonly used methods are: cancer immunogene therapy, suicide gene therapy, application of tumor-suppressor genes, antiangiogenic therapy, mesenchymal stem cells used as vectors, gene directed enzyme/prodrug therapy and bacteria used as anti-cancer agents. Cancer gene immunotherapy uses several immunologic agents for the purp...

  18. Transcription factor-microRNA-target gene networks associated with ovarian cancer survival and recurrence.

    Science.gov (United States)

    Delfino, Kristin R; Rodriguez-Zas, Sandra L

    2013-01-01

    The identification of reliable transcriptome biomarkers requires the simultaneous consideration of regulatory and target elements including microRNAs (miRNAs), transcription factors (TFs), and target genes. A novel approach that integrates multivariate survival analysis, feature selection, and regulatory network visualization was used to identify reliable biomarkers of ovarian cancer survival and recurrence. Expression profiles of 799 miRNAs, 17,814 TFs and target genes and cohort clinical records on 272 patients diagnosed with ovarian cancer were simultaneously considered and results were validated on an independent group of 146 patients. Three miRNAs (hsa-miR-16, hsa-miR-22*, and ebv-miR-BHRF1-2*) were associated with both ovarian cancer survival and recurrence and 27 miRNAs were associated with either one hazard. Two miRNAs (hsa-miR-521 and hsa-miR-497) were cohort-dependent, while 28 were cohort-independent. This study confirmed 19 miRNAs previously associated with ovarian cancer and identified two miRNAs that have previously been associated with other cancer types. In total, the expression of 838 and 734 target genes and 12 and eight TFs were associated (FDR-adjusted P-value cancer survival and recurrence, respectively. Functional analysis highlighted the association between cellular and nucleotide metabolic processes and ovarian cancer. The more direct connections and higher centrality of the miRNAs, TFs and target genes in the survival network studied suggest that network-based approaches to prognosticate or predict ovarian cancer survival may be more effective than those for ovarian cancer recurrence. This study demonstrated the feasibility to infer reliable miRNA-TF-target gene networks associated with survival and recurrence of ovarian cancer based on the simultaneous analysis of co-expression profiles and consideration of the clinical characteristics of the patients.

  19. Transcription factor-microRNA-target gene networks associated with ovarian cancer survival and recurrence.

    Directory of Open Access Journals (Sweden)

    Kristin R Delfino

    Full Text Available The identification of reliable transcriptome biomarkers requires the simultaneous consideration of regulatory and target elements including microRNAs (miRNAs, transcription factors (TFs, and target genes. A novel approach that integrates multivariate survival analysis, feature selection, and regulatory network visualization was used to identify reliable biomarkers of ovarian cancer survival and recurrence. Expression profiles of 799 miRNAs, 17,814 TFs and target genes and cohort clinical records on 272 patients diagnosed with ovarian cancer were simultaneously considered and results were validated on an independent group of 146 patients. Three miRNAs (hsa-miR-16, hsa-miR-22*, and ebv-miR-BHRF1-2* were associated with both ovarian cancer survival and recurrence and 27 miRNAs were associated with either one hazard. Two miRNAs (hsa-miR-521 and hsa-miR-497 were cohort-dependent, while 28 were cohort-independent. This study confirmed 19 miRNAs previously associated with ovarian cancer and identified two miRNAs that have previously been associated with other cancer types. In total, the expression of 838 and 734 target genes and 12 and eight TFs were associated (FDR-adjusted P-value <0.05 with ovarian cancer survival and recurrence, respectively. Functional analysis highlighted the association between cellular and nucleotide metabolic processes and ovarian cancer. The more direct connections and higher centrality of the miRNAs, TFs and target genes in the survival network studied suggest that network-based approaches to prognosticate or predict ovarian cancer survival may be more effective than those for ovarian cancer recurrence. This study demonstrated the feasibility to infer reliable miRNA-TF-target gene networks associated with survival and recurrence of ovarian cancer based on the simultaneous analysis of co-expression profiles and consideration of the clinical characteristics of the patients.

  20. Construction of a mouse model of factor VIII deficiency by gene targeting

    Energy Technology Data Exchange (ETDEWEB)

    Bi, L.; Lawler, A.; Gearhart, J. [Univ. of Pennsylvania School of Medicine, Philadelphia, PA (United States)] [and others

    1994-09-01

    To develop a small animal model of hemophilia A for gene therapy experiments, we set out to construct a mouse model for factor VIII deficiency by gene targeting. First, we screened a mouse liver cDNA library using a human FVIII cDNA probe. We cloned a 2.6 Kb partial mouse factor VIII cDNA which extends from 800 base pairs of the 3{prime} end of exon 14 to the 5{prime} end of exon 26. A mouse genomic library made from strain 129 was then screened to obtain genomic fragments covering the exons desired for homologous recombination. Two genomic clones were obtained, and one covering exon 15 through 22 was used for gene targeting. To make gene targeting constructs, a 5.8 Kb genomic DNA fragment covering exons 15 to 19 of the mouse FVIII gene was subcloned, and the neo expression cassette was inserted into exons 16 and 17 separately by different strategies. These two constructs were named MFVIIIC-16 and MFVIIIC-17. The constructs were linearized and transfected into strain 129 mouse ES cells by electroporation. Factor VIII gene-knockout ES cell lines were selected by G-418 and screened by genomic Southern blots. Eight exon 16 targeted cell lines and five exon 17 targeted cell lines were obtained. Three cell lines from each construct were injected into blastocysts and surgically transferred into foster mothers. Multiple chimeric mice with 70-90% hair color derived from the ES-cell genotype were seen with both constructs. Germ line transmission of the ES-cell genotype has been obtained for the MFVIIIC-16 construct, and multiple hemophilia A carrier females have been identified. Factor VIII-deficient males will be conceived soon.

  1. Fast and sensitive detection of indels induced by precise gene targeting

    DEFF Research Database (Denmark)

    Yang, Zhang; Steentoft, Catharina; Hauge, Camilla

    2015-01-01

    The nuclease-based gene editing tools are rapidly transforming capabilities for altering the genome of cells and organisms with great precision and in high throughput studies. A major limitation in application of precise gene editing lies in lack of sensitive and fast methods to detect...... and characterize the induced DNA changes. Precise gene editing induces double-stranded DNA breaks that are repaired by error-prone non-homologous end joining leading to introduction of insertions and deletions (indels) at the target site. These indels are often small and difficult and laborious to detect...

  2. Silencing of six hydrophobins in Cladosporium fulvum: complexities of simultaneously targeting multiple genes.

    Science.gov (United States)

    Lacroix, Hélène; Spanu, Pietro D

    2009-01-01

    In this study, we have constructed and expressed inverted repeat chimeras from the first exons of the six known hydrophobins of the fungus Cladosporium fulvum, the causal agent of tomato leaf mold. We used quantitative PCR to measure specifically the expression levels of the hydrophobins. The targeted genes are silenced to different degrees, but we also detected clear changes in the expression levels of nontargeted genes. This work highlights the difficulties that are likely to be encountered when attempting to silence more than one gene in a multigene family.

  3. E. coli recA gene improves gene targeted homologous recombination in Mycoplasma hyorhinis.

    Science.gov (United States)

    Ishag, Hassan Z A; Xiong, Qiyan; Liu, Maojun; Feng, Zhixin; Shao, Guoqing

    2017-05-01

    Mycoplasma hyorhinis is an opportunistic pathogen of pigs. Recently, it has been shown to transform cell cultures, increasing the attention of the researchers. Studies on the pathogenesis require specific genetic tool that is not yet available for the pathogen. To address this limitation, we constructed two suicide plasmids pGEMT-tetM/LR and pGEMT-recA-tetM/LR having a tetracycline resistance marker flanked by two hemolysin gene arms. The latter plasmid encodes an E. coli recA, a gene involved in DNA recombination, repair and maintenance of DNA. Using inactivation of the hemolysin gene, which results in a detectable and measurable phenotype, we found that each plasmid can disrupt the hemolysin gene of M. hyorhinis through a double cross-over homologous recombination. However, inclusion of the E. coli recA gene in the construct resulted in 9-fold increase in the frequency of hemolysin gene mutants among the screened tetracycline resistance colonies. The resultant hemolysin mutant strain lacks the ability to lyse mouse bed blood cells (RBC) when tested in vitro (p<0.001). The host-plasmid system described in this study, has applications for the genetic manipulation of this pathogen and potentially other mycoplasmas.

  4. Mining predicted essential genes of Brugia malayi for nematode drug targets.

    Directory of Open Access Journals (Sweden)

    Sanjay Kumar

    Full Text Available We report results from the first genome-wide application of a rational drug target selection methodology to a metazoan pathogen genome, the completed draft sequence of Brugia malayi, a parasitic nematode responsible for human lymphatic filariasis. More than 1.5 billion people worldwide are at risk of contracting lymphatic filariasis and onchocerciasis, a related filarial disease. Drug treatments for filariasis have not changed significantly in over 20 years, and with the risk of resistance rising, there is an urgent need for the development of new anti-filarial drug therapies. The recent publication of the draft genomic sequence for B. malayi enables a genome-wide search for new drug targets. However, there is no functional genomics data in B. malayi to guide the selection of potential drug targets. To circumvent this problem, we have utilized the free-living model nematode Caenorhabditis elegans as a surrogate for B. malayi. Sequence comparisons between the two genomes allow us to map C. elegans orthologs to B. malayi genes. Using these orthology mappings and by incorporating the extensive genomic and functional genomic data, including genome-wide RNAi screens, that already exist for C. elegans, we identify potentially essential genes in B. malayi. Further incorporation of human host genome sequence data and a custom algorithm for prioritization enables us to collect and rank nearly 600 drug target candidates. Previously identified potential drug targets cluster near the top of our prioritized list, lending credibility to our methodology. Over-represented Gene Ontology terms, predicted InterPro domains, and RNAi phenotypes of C. elegans orthologs associated with the potential target pool are identified. By virtue of the selection procedure, the potential B. malayi drug targets highlight components of key processes in nematode biology such as central metabolism, molting and regulation of gene expression.

  5. Mining predicted essential genes of Brugia malayi for nematode drug targets.

    Science.gov (United States)

    Kumar, Sanjay; Chaudhary, Kshitiz; Foster, Jeremy M; Novelli, Jacopo F; Zhang, Yinhua; Wang, Shiliang; Spiro, David; Ghedin, Elodie; Carlow, Clotilde K S

    2007-01-01

    We report results from the first genome-wide application of a rational drug target selection methodology to a metazoan pathogen genome, the completed draft sequence of Brugia malayi, a parasitic nematode responsible for human lymphatic filariasis. More than 1.5 billion people worldwide are at risk of contracting lymphatic filariasis and onchocerciasis, a related filarial disease. Drug treatments for filariasis have not changed significantly in over 20 years, and with the risk of resistance rising, there is an urgent need for the development of new anti-filarial drug therapies. The recent publication of the draft genomic sequence for B. malayi enables a genome-wide search for new drug targets. However, there is no functional genomics data in B. malayi to guide the selection of potential drug targets. To circumvent this problem, we have utilized the free-living model nematode Caenorhabditis elegans as a surrogate for B. malayi. Sequence comparisons between the two genomes allow us to map C. elegans orthologs to B. malayi genes. Using these orthology mappings and by incorporating the extensive genomic and functional genomic data, including genome-wide RNAi screens, that already exist for C. elegans, we identify potentially essential genes in B. malayi. Further incorporation of human host genome sequence data and a custom algorithm for prioritization enables us to collect and rank nearly 600 drug target candidates. Previously identified potential drug targets cluster near the top of our prioritized list, lending credibility to our methodology. Over-represented Gene Ontology terms, predicted InterPro domains, and RNAi phenotypes of C. elegans orthologs associated with the potential target pool are identified. By virtue of the selection procedure, the potential B. malayi drug targets highlight components of key processes in nematode biology such as central metabolism, molting and regulation of gene expression.

  6. Deletion of ku homologs increases gene targeting frequency in Streptomyces avermitilis.

    Science.gov (United States)

    Zhang, Xiaojuan; Chen, Wei; Zhang, Yang; Jiang, Libin; Chen, Zhi; Wen, Ying; Li, Jilun

    2012-06-01

    Streptomyces avermitilis is an industrially important soil bacterium known for production of avermectins, which are antiparasitic agents useful in animal health care, agriculture, and treatment of human infections. ku genes play a key role in the non-homologous end-joining pathway for repair of DNA double strand breaks. We identified homologs of eukaryotic ku70 and ku80 genes, termed ku1 and ku2, in S. avermitilis. Mutants with deletion of ku1, ku2, and both genes were constructed and their phenotypic changes were characterized. Deletion of ku genes had no apparent adverse effects on growth, spore formation, or avermectin production. The ku mutants, in comparison to wild-type strain, were slightly more sensitive to the DNA-damaging agent ethyl methanesulfonate, but not to UV exposure or to bleomycin. Gene targeting frequencies by homologous recombination were higher in the ku mutants than in wild-type strain. We conclude that ku-deleted strains will be useful hosts for efficient gene targeting and will facilitate functional analysis of genes in S. avermitilis and other industrially important bacterial strains.

  7. Targeted sequencing of cancer-related genes in colorectal cancer using next-generation sequencing.

    Directory of Open Access Journals (Sweden)

    Sae-Won Han

    Full Text Available Recent advance in sequencing technology has enabled comprehensive profiling of genetic alterations in cancer. We have established a targeted sequencing platform using next-generation sequencing (NGS technology for clinical use, which can provide mutation and copy number variation data. NGS was performed with paired-end library enriched with exons of 183 cancer-related genes. Normal and tumor tissue pairs of 60 colorectal adenocarcinomas were used to test feasibility. Somatic mutation and copy number alteration were analyzed. A total of 526 somatic non-synonymous sequence variations were found in 113 genes. Among these, 278 single nucleotide variations were 232 different somatic point mutations. 216 SNV were 79 known single nucleotide polymorphisms in the dbSNP. 32 indels were 28 different indel mutations. Median number of mutated gene per tumor was 4 (range 0-23. Copy number gain (>X2 fold was found in 65 genes in 40 patients, whereas copy number loss (genes in 39 patients. The most frequently altered genes (mutation and/or copy number alteration were APC in 35 patients (58%, TP53 in 34 (57%, and KRAS in 24 (40%. Altered gene list revealed ErbB signaling pathway as the most commonly involved pathway (25 patients, 42%. Targeted sequencing platform using NGS technology is feasible for clinical use and provides comprehensive genetic alteration data.

  8. Generation of TALE nickase-mediated gene-targeted cows expressing human serum albumin in mammary glands.

    Science.gov (United States)

    Luo, Yan; Wang, Yongsheng; Liu, Jun; Cui, Chenchen; Wu, Yongyan; Lan, Hui; Chen, Qi; Liu, Xu; Quan, Fusheng; Guo, Zekun; Zhang, Yong

    2016-02-08

    Targeting exogenous genes at milk protein loci via gene-targeting technology is an ideal strategy for producing large quantities of pharmaceutical proteins. Transcription-activator-like effector (TALE) nucleases (TALENs) are an efficient genome-editing tool. However, the off-target effects may lead to unintended gene mutations. In this study, we constructed TALENs and TALE nickases directed against exon 2 of the bovine β-lactoglobulin (BLG) locus. The nickases can induce a site-specific DNA single-strand break, without inducing double-strand break and nonhomologous end joining mediated gene mutation, and lower cell apoptosis rate than TALENs. After co-transfecting the bovine fetal fibroblasts with human serum albumin (HSA) gene-targeting vector and TALE nickase expression vectors, approximately 4.8% (40/835) of the cell clones contained HSA at BLG locus. Unexpectedly, one homozygous gene-targeted cell clone (1/835, 0.1%) was obtained by targeting both alleles of BLG in a single round of transfection. The recombinant protein mimicking the endogenous BLG was highly expressed and correctly folded in the mammary glands of the targeted cows, and the expression level of HSA was significantly increased in the homozygous targeted cows. Results suggested that the combination of TALE nickase-mediated gene targeting and somatic cell nuclear transfer is a feasible and safe approach in producing gene-targeted livestock.

  9. Genes involved in cell adhesion and signaling: a new repertoire of retinoic acid receptor target genes in mouse embryonic fibroblasts.

    Science.gov (United States)

    Al Tanoury, Ziad; Piskunov, Aleksandr; Andriamoratsiresy, Dina; Gaouar, Samia; Lutzing, Régis; Ye, Tao; Jost, Bernard; Keime, Céline; Rochette-Egly, Cécile

    2014-02-01

    Nuclear retinoic acid (RA) receptors (RARα, β and γ) are ligand-dependent transcription factors that regulate the expression of a battery of genes involved in cell differentiation and proliferation. They are also phosphoproteins and we previously showed the importance of their phosphorylation in their transcriptional activity. In the study reported here, we conducted a genome-wide analysis of the genes that are regulated by RARs in mouse embryonic fibroblasts (MEFs) by comparing wild-type MEFs to MEFs lacking the three RARs. We found that in the absence of RA, RARs control the expression of several gene transcripts associated with cell adhesion. Consequently the knockout MEFs are unable to adhere and to spread on substrates and they display a disrupted network of actin filaments, compared with the WT cells. In contrast, in the presence of the ligand, RARs control the expression of other genes involved in signaling and in RA metabolism. Taking advantage of rescue cell lines expressing the RARα or RARγ subtypes (either wild-type or mutated at the N-terminal phosphorylation sites) in the null background, we found that the expression of RA-target genes can be controlled either by a specific single RAR or by a combination of RAR isotypes, depending on the gene. We also selected genes that require the phosphorylation of the receptors for their regulation by RA. Our results increase the repertoire of genes that are regulated by RARs and highlight the complexity and diversity of the transcriptional programs regulated by RARs, depending on the gene.

  10. Epidermal growth factor receptor targeting enhances adenoviral vector based suicide gene therapy of osteosarcoma

    NARCIS (Netherlands)

    Witlox, M.A.; van Beusechem, V.W.; Grill, J.; Haisma, H.J.; Schaap, G.; Bras, J.; Van Diest, P.; De Gast, A.; Curiel, D.T.; Pinedo, H.M.; Gerritsen, W.R.; Wuisman, P.I.

    2002-01-01

    Background Despite improvements in the treatment of osteosarcoma (OS) there are still too many patients who cannot benefit from current treatment modalities. Therefore, new therapeutic approaches are warranted. Here we explore the efficacy of targeted adenoviral based suicide gene therapy. Methods a

  11. Nonviral Gene Targeting at rDNA Locus of Human Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Youjin Hu

    2013-01-01

    Full Text Available Background. Genetic modification, such as the addition of exogenous genes to the MSC genome, is crucial to their use as cellular vehicles. Due to the risks associated with viral vectors such as insertional mutagenesis, the safer nonviral vectors have drawn a great deal of attention. Methods. VEGF, bFGF, vitamin C, and insulin-transferrin-selenium-X were supplemented in the MSC culture medium. The cells’ proliferation and survival capacity was measured by MTT, determination of the cumulative number of cells, and a colony-forming efficiency assay. The plasmid pHr2-NL was constructed and nucleofected into MSCs. The recombinants were selected using G418 and characterized using PCR and Southern blotting. Results. BFGF is critical to MSC growth and it acted synergistically with vitamin C, VEGF, and ITS-X, causing the cells to expand significantly. The neomycin gene was targeted to the rDNA locus of human MSCs using a nonviral human ribosomal targeting vector. The recombinant MSCs retained multipotential differentiation capacity, typical levels of hMSC surface marker expression, and a normal karyotype, and none were tumorigenic in nude mice. Conclusions. Exogenous genes can be targeted to the rDNA locus of human MSCs while maintaining the characteristics of MSCs. This is the first nonviral gene targeting of hMSCs.

  12. Problem-Solving Test: Conditional Gene Targeting Using the Cre/loxP Recombination System

    Science.gov (United States)

    Szeberényi, József

    2013-01-01

    Terms to be familiar with before you start to solve the test: gene targeting, knock-out mutation, bacteriophage, complementary base-pairing, homologous recombination, deletion, transgenic organisms, promoter, polyadenylation element, transgene, DNA replication, RNA polymerase, Shine-Dalgarno sequence, restriction endonuclease, polymerase chain…

  13. The effect of COMT gene on the target precision of the athlete movement

    Directory of Open Access Journals (Sweden)

    E. V. Mikhailova

    2014-01-01

    Full Text Available The aim of the study was to find correlation between COMT gene alleles and the target precision of the athlete movement. 68 Russian competing athletes involved in boxing and volleyball, participated in the study. We found interrelation between COMT Met allele and a tall stature in the volleyball players.

  14. Interactome of Radiation-Induced microRNA-Predicted Target Genes

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    Tenzin W. Lhakhang

    2012-01-01

    Full Text Available The microRNAs (miRNAs function as global negative regulators of gene expression and have been associated with a multitude of biological processes. The dysfunction of the microRNAome has been linked to various diseases including cancer. Our laboratory recently reported modulation in the expression of miRNA in a variety of cell types exposed to ionizing radiation (IR. To further understand miRNA role in IR-induced stress pathways, we catalogued a set of common miRNAs modulated in various irradiated cell lines and generated a list of predicted target genes. Using advanced bioinformatics tools we identified cellular pathways where miRNA predicted target genes function. The miRNA-targeted genes were found to play key roles in previously identified IR stress pathways such as cell cycle, p53 pathway, TGF-beta pathway, ubiquitin-mediated proteolysis, focal adhesion pathway, MAPK signaling, thyroid cancer pathway, adherens junction, insulin signaling pathway, oocyte meiosis, regulation of actin cytoskeleton, and renal cell carcinoma pathway. Interestingly, we were able to identify novel targeted pathways that have not been identified in cellular radiation response, such as aldosterone-regulated sodium reabsorption, long-term potentiation, and neutrotrophin signaling pathways. Our analysis indicates that the miRNA interactome in irradiated cells provides a platform for comprehensive modeling of the cellular stress response to IR exposure.

  15. Topical liposome targeting of dyes, melanins, genes, and proteins selectively to hair follicles.

    Science.gov (United States)

    Hoffman, R M

    1998-01-01

    For therapeutic and cosmetic modification of hair, we have developed a hair-follicle-selective macromolecule and small molecule targeting system with topical application of phosphatidylcholine-based liposomes. Liposome-entrapped melanins, proteins, genes, and small-molecules have been selectively targeted to the hair follicle and hair shafts of mice. Liposomal delivery of these molecules is time dependent. Negligible amounts of delivered molecules enter the dermis, epidermis, or bloodstream thereby demonstrating selective follicle delivery. Naked molecules are trapped in the stratum corneum and are unable to enter the follicle. The potential of the hair-follicle liposome delivery system for therapeutic use for hair disease as well as for cosmesis has been demonstrated in 3-dimensional histoculture of hair-growing skin and mouse in vivo models. Topical liposome selective delivery to hair follicles has demonstrated the ability to color hair with melanin, the delivery of the active lac-Z gene to hair matrix cells and delivery of proteins as well. Liposome-targeting of molecules to hair follicles has also been achieved in human scalp in histoculture. Liposomes thus have high potential in selective hair follicle targeting of large and small molecules, including genes, opening the field of gene therapy and other molecular therapy of the hair process to restore hair growth, physiologically restore or alter hair pigment, and to prevent or accelerate hair loss.

  16. The feasibility of targeted selective gene therapy of the hair follicle.

    Science.gov (United States)

    Li, L; Hoffman, R M

    1995-07-01

    Loss of hair and hair colour is associated with ageing, and when it involves the scalp hair, it can be distressing to both sexes. Hair loss resulting from cancer chemotherapy is particularly distressing. However, safe, effective therapies directed to hair have only just started to be developed. The hair follicle is a complex skin appendage composed of epidermal and dermal tissue, with specialized keratinocytes, the hair matrix cells, forming the hair shaft. Specific therapy of the hair follicle depends on selective targeting of specific cells of the hair follicle. We have developed the histoculture of intact hair-growing skin on sponge-gel matrices. We have recently found in histocultured skin that liposomes can selectively target hair follicles to deliver both small and large molecules. That liposomes can target the hair follicle for delivery has been confirmed independently. Two decades ago we introduced the technique of entrapping DNA in liposomes for use in gene therapy. In this report we describe the selective targeting of the lacZ reporter gene to the hair follicles in mice after topical application of the gene entrapped in liposomes. These results demonstrate that highly selective, safe gene therapy for the hair process is feasible.

  17. A single gene target of an ETS-family transcription factor determines neuronal CO2-chemosensitivity

    DEFF Research Database (Denmark)

    Brandt, Julia P; Aziz-Zaman, Sonya; Juozaityte, Vaida;

    2012-01-01

    . We report here a mechanism that endows C. elegans neurons with the ability to detect CO(2). The ETS-5 transcription factor is necessary for the specification of CO(2)-sensing BAG neurons. Expression of a single ETS-5 target gene, gcy-9, which encodes a receptor-type guanylate cyclase, is sufficient...

  18. Targeting human microRNA genes using engineered Tal-effector nucleases (TALENs.

    Directory of Open Access Journals (Sweden)

    Ruozhen Hu

    Full Text Available MicroRNAs (miRNAs have quickly emerged as important regulators of mammalian physiology owing to their precise control over the expression of critical protein coding genes. Despite significant progress in our understanding of how miRNAs function in mice, there remains a fundamental need to be able to target and edit miRNA genes in the human genome. Here, we report a novel approach to disrupting human miRNA genes ex vivo using engineered TAL-effector (TALE proteins to function as nucleases (TALENs that specifically target and disrupt human miRNA genes. We demonstrate that functional TALEN pairs can be designed to enable disruption of miRNA seed regions, or removal of entire hairpin sequences, and use this approach to successfully target several physiologically relevant human miRNAs including miR-155*, miR-155, miR-146a and miR-125b. This technology will allow for a substantially improved capacity to study the regulation and function of miRNAs in human cells, and could be developed into a strategic means by which miRNAs can be targeted therapeutically during human disease.

  19. Fine genetic mapping of target leaf spot resistance gene cca-3 in cucumber, Cucumis sativus L

    Science.gov (United States)

    The target leaf spot (TLS) is a very important fungal disease in cucumber. In this study, we conducted fine genetic mapping of a recessively inherited resistance gene, cca-2 against TLS with 1,083 F2 plants derived from the resistant cucumber inbred line D31 and the susceptible line D5. Initial mapp...

  20. Targeting Calcium Signaling Induces Epigenetic Reactivation of Tumor Suppressor Genes in Cancer.

    Science.gov (United States)

    Raynal, Noël J-M; Lee, Justin T; Wang, Youjun; Beaudry, Annie; Madireddi, Priyanka; Garriga, Judith; Malouf, Gabriel G; Dumont, Sarah; Dettman, Elisha J; Gharibyan, Vazganush; Ahmed, Saira; Chung, Woonbok; Childers, Wayne E; Abou-Gharbia, Magid; Henry, Ryan A; Andrews, Andrew J; Jelinek, Jaroslav; Cui, Ying; Baylin, Stephen B; Gill, Donald L; Issa, Jean-Pierre J

    2016-03-15

    Targeting epigenetic pathways is a promising approach for cancer therapy. Here, we report on the unexpected finding that targeting calcium signaling can reverse epigenetic silencing of tumor suppressor genes (TSG). In a screen for drugs that reactivate silenced gene expression in colon cancer cells, we found three classical epigenetic targeted drugs (DNA methylation and histone deacetylase inhibitors) and 11 other drugs that induced methylated and silenced CpG island promoters driving a reporter gene (GFP) as well as endogenous TSGs in multiple cancer cell lines. These newly identified drugs, most prominently cardiac glycosides, did not change DNA methylation locally or histone modifications globally. Instead, all 11 drugs altered calcium signaling and triggered calcium-calmodulin kinase (CamK) activity, leading to MeCP2 nuclear exclusion. Blocking CamK activity abolished gene reactivation and cancer cell killing by these drugs, showing that triggering calcium fluxes is an essential component of their epigenetic mechanism of action. Our data identify calcium signaling as a new pathway that can be targeted to reactivate TSGs in cancer.

  1. Horizontal transfer of a eukaryotic plastid-targeted protein gene to cyanobacteria

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    Keeling Patrick J

    2007-06-01

    Full Text Available Abstract Background Horizontal or lateral transfer of genetic material between distantly related prokaryotes has been shown to play a major role in the evolution of bacterial and archaeal genomes, but exchange of genes between prokaryotes and eukaryotes is not as well understood. In particular, gene flow from eukaryotes to prokaryotes is rarely documented with strong support, which is unusual since prokaryotic genomes appear to readily accept foreign genes. Results Here, we show that abundant marine cyanobacteria in the related genera Synechococcus and Prochlorococcus acquired a key Calvin cycle/glycolytic enzyme from a eukaryote. Two non-homologous forms of fructose bisphosphate aldolase (FBA are characteristic of eukaryotes and prokaryotes respectively. However, a eukaryotic gene has been inserted immediately upstream of the ancestral prokaryotic gene in several strains (ecotypes of Synechococcus and Prochlorococcus. In one lineage this new gene has replaced the ancestral gene altogether. The eukaryotic gene is most closely related to the plastid-targeted FBA from red algae. This eukaryotic-type FBA once replaced the plastid/cyanobacterial type in photosynthetic eukaryotes, hinting at a possible functional advantage in Calvin cycle reactions. The strains that now possess this eukaryotic FBA are scattered across the tree of Synechococcus and Prochlorococcus, perhaps because the gene has been transferred multiple times among cyanobacteria, or more likely because it has been selectively retained only in certain lineages. Conclusion A gene for plastid-targeted FBA has been transferred from red algae to cyanobacteria, where it has inserted itself beside its non-homologous, functional analogue. Its current distribution in Prochlorococcus and Synechococcus is punctate, suggesting a complex history since its introduction to this group.

  2. Big genes are big mutagen targets: a connection to cancerous, spherical cells?

    Science.gov (United States)

    Parry, Michele L; Ramsamooj, Michael; Blanck, George

    2015-01-28

    We determined the most commonly mutated genes in five cancer genome atlas (TCGA) datasets. Many of these genes were extraordinarily large, as are many cancer fusion gene partners. And many of these genes had cytoskeletal related functions. We further determined that these genes were distributed into high and low frequency mutation groups largely according to overall rate of gene-occurrence in the high and low mutation frequency groups, as was also the case with common metastasis and tumor suppressor genes. Oncoproteins were selectively mutated in the low mutation frequency groups in colon and lung datasets. Thus, genes that have very large coding regions and may impact the cytoskeleton are more commonly mutated than are common metastasis and tumor suppressor genes in both high and low frequency mutation groups. These analyses raise questions related to cell shape: (i) Are cancer cells often spherical because cytoskeletal-related proteins are large mutagen targets? (ii) Is drug-resistance facilitated by relatively common mutant proteins that lead to round cells, with altered cell physiology or reduced surface to volume ratios that could reduce intra-cellular drug concentrations?

  3. DbMDR: a relational database for multidrug resistance genes as potential drug targets.

    Science.gov (United States)

    Gupta, Sanchita; Mishra, Manoj; Sen, Naresh; Parihar, Rashi; Dwivedi, Gaurav Raj; Khan, Feroz; Sharma, Ashok

    2011-10-01

    DbMDR is non-redundant reference database of multidrug resistance (MDR) genes and their orthologs acting as potential drug targets. Drug resistance is a common phenomenon of pathogens, creating a serious problem of inactivation of drugs and antibiotics resulting in occurrence of diseases. Apart from other factors, the MDR genes present in pathogens are shown to be responsible for multidrug resistance. Much of the unorganized information on MDR genes is scattered across the literature and other web resources. Thus, consolidation of such knowledge about MDR genes into one database will make the drug discovery research more efficient. Mining of text for MDR genes has resulted into a large number of publications but in scattered and unorganized form. This information was compiled into a database, which enables a user not only to look at a particular MDR gene but also to find out putative homologs based on sequence similarity, conserved domains, and motifs in proteins encoded by MDR genes more efficiently. At present, DbMDR database contains 2843 MDR genes characterized experimentally as well as functionally annotated with cross-referencing search support. The DbMDR database (http://203.190.147.116/dbmdr/) is a comprehensive resource for comparative study focused on MDR genes and metabolic pathway efflux pumps and intended to provide a platform for researchers for further research in drug resistance.

  4. Analysis of the siRNA-Mediated Gene Silencing Process Targeting Three Homologous Genes Controlling Soybean Seed Oil Quality.

    Science.gov (United States)

    Lu, Sha; Yin, Xiaoyan; Spollen, William; Zhang, Ning; Xu, Dong; Schoelz, James; Bilyeu, Kristin; Zhang, Zhanyuan J

    2015-01-01

    In the past decade, RNA silencing has gained significant attention because of its success in genomic scale research and also in the genetic improvement of crop plants. However, little is known about the molecular basis of siRNA processing in association with its target transcript. To reveal this process for improving hpRNA-mediated gene silencing in crop plants, the soybean GmFAD3 gene family was chosen as a test model. We analyzed RNAi mutant soybean lines in which three members of the GmFAD3 gene family were silenced. The silencing levels of FAD3A, FAD3B and FAD3C were correlated with the degrees of sequence homology between the inverted repeat of hpRNA and the GmFAD3 transcripts in the RNAi lines. Strikingly, transgenes in two of the three RNAi lines were heavily methylated, leading to a dramatic reduction of hpRNA-derived siRNAs. Small RNAs corresponding to the loop portion of the hairpin transcript were detected while much lower levels of siRNAs were found outside of the target region. siRNAs generated from the 318-bp inverted repeat were found to be diced much more frequently at stem sequences close to the loop and associated with the inferred cleavage sites on the target transcripts, manifesting "hot spots". The top candidate hpRNA-derived siRNA share certain sequence features with mature miRNA. This is the first comprehensive and detailed study revealing the siRNA-mediated gene silencing mechanism in crop plants using gene family GmFAD3 as a test model.

  5. miRNA gene promoters are frequent targets of aberrant DNA methylation in human breast cancer.

    Science.gov (United States)

    Vrba, Lukas; Muñoz-Rodríguez, José L; Stampfer, Martha R; Futscher, Bernard W

    2013-01-01

    miRNAs are important regulators of gene expression that are frequently deregulated in cancer, with aberrant DNA methylation being an epigenetic mechanism involved in this process. We previously identified miRNA promoter regions active in normal mammary cell types and here we analyzed which of these promoters are targets of aberrant DNA methylation in human breast cancer cell lines and breast tumor specimens. Using 5-methylcytosine immunoprecipitation coupled to miRNA tiling microarray hybridization, we performed comprehensive evaluation of DNA methylation of miRNA gene promoters in breast cancer. We found almost one third (55/167) of miRNA promoters were targets for aberrant methylation in breast cancer cell lines. Breast tumor specimens displayed DNA methylation of majority of these miRNA promoters, indicating that these changes in DNA methylation might be clinically relevant. Aberrantly methylated miRNA promoters were, similar to protein coding genes, enriched for promoters targeted by polycomb in normal cells. Detailed analysis of selected miRNA promoters revealed decreased expression of miRNA linked to increased promoter methylation for mir-31, mir-130a, let-7a-3/let-7b, mir-155, mir-137 and mir-34b/mir-34c genes. The proportion of miRNA promoters we found aberrantly methylated in breast cancer is several fold larger than that observed for protein coding genes, indicating an important role of DNA methylation in miRNA deregulation in cancer.

  6. High-efficiency and heritable gene targeting in mouse by transcription activator-like effector nucleases.

    Science.gov (United States)

    Qiu, Zhongwei; Liu, Meizhen; Chen, Zhaohua; Shao, Yanjiao; Pan, Hongjie; Wei, Gaigai; Yu, Chao; Zhang, Long; Li, Xia; Wang, Ping; Fan, Heng-Yu; Du, Bing; Liu, Bin; Liu, Mingyao; Li, Dali

    2013-06-01

    Transcription activator-like effector nucleases (TALENs) are a powerful new approach for targeted gene disruption in various animal models, but little is known about their activities in Mus musculus, the widely used mammalian model organism. Here, we report that direct injection of in vitro transcribed messenger RNA of TALEN pairs into mouse zygotes induced somatic mutations, which were stably passed to the next generation through germ-line transmission. With one TALEN pair constructed for each of 10 target genes, mutant F0 mice for each gene were obtained with the mutation rate ranged from 13 to 67% and an average of ∼40% of total healthy newborns with no significant differences between C57BL/6 and FVB/N genetic background. One TALEN pair with single mismatch to their intended target sequence in each side failed to yield any mutation. Furthermore, highly efficient germ-line transmission was obtained, as all the F0 founders tested transmitted the mutations to F1 mice. In addition, we also observed that one bi-allele mutant founder of Lepr gene, encoding Leptin receptor, had similar diabetic phenotype as db/db mouse. Together, our results suggest that TALENs are an effective genetic tool for rapid gene disruption with high efficiency and heritability in mouse with distinct genetic background.

  7. Analysis of Deregulated microRNAs and Their Target Genes in Gastric Cancer.

    Directory of Open Access Journals (Sweden)

    Simonas Juzėnas

    Full Text Available MicroRNAs (miRNAs are widely studied non-coding RNAs that modulate gene expression. MiRNAs are deregulated in different tumors including gastric cancer (GC and have potential diagnostic and prognostic implications. The aim of our study was to determine miRNA profile in GC tissues, followed by evaluation of deregulated miRNAs in plasma of GC patients. Using available databases and bioinformatics methods we also aimed to evaluate potential target genes of confirmed differentially expressed miRNA and validate these findings in GC tissues.The study included 51 GC patients and 51 controls. Initially, we screened miRNA expression profile in 13 tissue samples of GC and 12 normal gastric tissues with TaqMan low density array (TLDA. In the second stage, differentially expressed miRNAs were validated in a replication cohort using qRT-PCR in tissue and plasma samples. Subsequently, we analyzed potential target genes of deregulated miRNAs using bioinformatics approach, determined their expression in GC tissues and performed correlation analysis with targeting miRNAs.Profiling with TLDA revealed 15 deregulated miRNAs in GC tissues compared to normal gastric mucosa. Replication analysis confirmed that miR-148a-3p, miR-204-5p, miR-223-3p and miR-375 were consistently deregulated in GC tissues. Analysis of GC patients' plasma samples showed significant down-regulation of miR-148a-3p, miR-375 and up-regulation of miR-223-3p compared to healthy subjects. Further, using bioinformatic tools we identified targets of replicated miRNAs and performed disease-associated gene enrichment analysis. Ultimately, we evaluated potential target gene BCL2 and DNMT3B expression by qRT-PCR in GC tissue, which correlated with targeting miRNA expression.Our study revealed miRNA profile in GC tissues and showed that miR-148a-3p, miR-223-3p and miR-375 are deregulated in GC plasma samples, but these circulating miRNAs showed relatively weak diagnostic performance as sole biomarkers

  8. Targeting an adenoviral gene vector to cytokine-activated vascular endothelium via E-selectin.

    Science.gov (United States)

    Harari, O A; Wickham, T J; Stocker, C J; Kovesdi, I; Segal, D M; Huehns, T Y; Sarraf, C; Haskard, D O

    1999-05-01

    We have aimed at selective gene delivery to vascular endothelial cells (EC) at sites of inflammation, by targeting E-selectin, a surface adhesion molecule that is only expressed by activated EC. An anti-E-selectin mAb, 1.2B6, was complexed with the adenovirus vector AdZ.FLAG (expressing the FLAG peptide) by conjugating it to an anti-FLAG mAb. Gene transduction of cultured EC was increased 20-fold compared with AdZ.FLAG complexed with a control bsAb providing EC were activated by cytokines. The anti-E-selectin-complexed vector transduced 29 +/- 9% of intimal EC in segments of pig aorta cultured with cytokines ex vivo, compared with less than 0.1% transduced with the control construct (P < 0.05). This strategy could be developed to target endothelium in inflammation with genes capable of modifying the inflammatory response.

  9. Dose-dependent regulation of target gene expression and cell proliferation by c-Myc levels.

    Science.gov (United States)

    Schuhmacher, Marino; Eick, Dirk

    2013-01-01

    The proto-oncogene c-myc encodes a basic helix-loop-helix leucine zipper transcription factor (c-Myc). c-Myc plays a crucial role in cell growth and proliferation. Here, we examined how expression of c-Myc target genes and cell proliferation depend on variation of c-Myc protein levels. We show that proliferation rates, the number of cells in S-phase, and cell size increased in a dose-dependent manner in response to increasing c-Myc levels. Likewise, the mRNA levels of c-Myc responsive genes steadily increased with rising c-Myc levels. Strikingly, steady-state mRNA levels of c-Myc target genes did not saturate even at highest c-Myc concentrations. These characteristics predestine c-Myc levels as a cellular rheostat for the control and fine-tuning of cell proliferation and growth rates.

  10. Modeling Human Severe Combined Immunodeficiency and Correction by CRISPR/Cas9-Enhanced Gene Targeting.

    Science.gov (United States)

    Chang, Chia-Wei; Lai, Yi-Shin; Westin, Erik; Khodadadi-Jamayran, Alireza; Pawlik, Kevin M; Lamb, Lawrence S; Goldman, Frederick D; Townes, Tim M

    2015-09-08

    Mutations of the Janus family kinase JAK3 gene cause severe combined immunodeficiency (SCID). JAK3 deficiency in humans is characterized by the absence of circulating T cells and natural killer (NK) cells with normal numbers of poorly functioning B cells (T(-)B(+)NK(-)). Using SCID patient-specific induced pluripotent stem cells (iPSCs) and a T cell in vitro differentiation system, we demonstrate a complete block in early T cell development of JAK3-deficient cells. Correction of the JAK3 mutation by CRISPR/Cas9-enhanced gene targeting restores normal T cell development, including the production of mature T cell populations with a broad T cell receptor (TCR) repertoire. Whole-genome sequencing of corrected cells demonstrates no CRISPR/Cas9 off-target modifications. These studies describe an approach for the study of human lymphopoiesis and provide a foundation for gene correction therapy in humans with immunodeficiencies.

  11. Targeted DNA demethylation and activation of endogenous genes using programmable TALE-TET1 fusion proteins.

    Science.gov (United States)

    Maeder, Morgan L; Angstman, James F; Richardson, Marcy E; Linder, Samantha J; Cascio, Vincent M; Tsai, Shengdar Q; Ho, Quan H; Sander, Jeffry D; Reyon, Deepak; Bernstein, Bradley E; Costello, Joseph F; Wilkinson, Miles F; Joung, J Keith

    2013-12-01

    Genome-wide studies have defined cell type-specific patterns of DNA methylation that are important for regulating gene expression in both normal development and disease. However, determining the functional significance of specific methylation events remains challenging, owing to the lack of methods for removing such modifications in a targeted manner. Here we describe an approach for efficient targeted demethylation of specific CpGs in human cells using fusions of engineered transcription activator-like effector (TALE) repeat arrays and the TET1 hydroxylase catalytic domain. Using these TALE-TET1 fusions, we demonstrate that modification of critical methylated promoter CpG positions can lead to substantial increases in the expression of endogenous human genes. Our results delineate a strategy for understanding the functional significance of specific CpG methylation marks in the context of endogenous gene loci and validate programmable DNA demethylation reagents with potential utility for research and therapeutic applications.

  12. New target genes of MITF-induced microRNA-211 contribute to melanoma cell invasion.

    Directory of Open Access Journals (Sweden)

    Christiane Margue

    Full Text Available The non-coding microRNAs (miRNA have tissue- and disease-specific expression patterns. They down-regulate target mRNAs, which likely impacts on most fundamental cellular processes. Differential expression patterns of miRNAs are currently being exploited for identification of biomarkers for early disease diagnosis, prediction of progression for melanoma and other cancers and as promising drug targets, since they can easily be inhibited or replaced in a given cellular context. Before successfully manipulating miRNAs in clinical settings, their precise expression levels, endogenous functions and thus their target genes have to be determined. MiR-211, a melanocyte lineage-specific small non-coding miRNA, is located in an intron of TRPM1, a target gene of the microphtalmia-associated transcription factor (MITF. By transcriptionally up-regulating TRPM1, MITF, which is critical for both melanocyte differentiation and survival and for melanoma progression, indirectly drives the expression of miR-211. Expression of this miRNA is often reduced in melanoma samples. Here, we investigated functional roles of miR-211 by identifying and studying new target genes. We show that MITF-correlated miR-211 expression levels are mostly but not always reduced in a panel of 11 melanoma cell lines and in primary and metastatic melanoma compared to normal melanocytes and nevi, respectively. MiR-211 itself only marginally impacted on cell invasion and migration, while perturbation of some new miR-211 target genes, such as AP1S2, SOX11, IGFBP5, and SERINC3 significantly increased invasion. These results and the variable expression levels of miR-211 raise serious doubts on the value of miR-211 as a melanoma tumor-suppressing miRNA and/or as a biomarker for melanoma.

  13. Efficient immunoglobulin gene disruption and targeted replacement in rabbit using zinc finger nucleases.

    Directory of Open Access Journals (Sweden)

    Tatiana Flisikowska

    Full Text Available Rabbits are widely used in biomedical research, yet techniques for their precise genetic modification are lacking. We demonstrate that zinc finger nucleases (ZFNs introduced into fertilized oocytes can inactivate a chosen gene by mutagenesis and also mediate precise homologous recombination with a DNA gene-targeting vector to achieve the first gene knockout and targeted sequence replacement in rabbits. Two ZFN pairs were designed that target the rabbit immunoglobulin M (IgM locus within exons 1 and 2. ZFN mRNAs were microinjected into pronuclear stage fertilized oocytes. Founder animals carrying distinct mutated IgM alleles were identified and bred to produce offspring. Functional knockout of the immunoglobulin heavy chain locus was confirmed by serum IgM and IgG deficiency and lack of IgM(+ and IgG(+ B lymphocytes. We then tested whether ZFN expression would enable efficient targeted sequence replacement in rabbit oocytes. ZFN mRNA was co-injected with a linear DNA vector designed to replace exon 1 of the IgM locus with ∼1.9 kb of novel sequence. Double strand break induced targeted replacement occurred in up to 17% of embryos and in 18% of fetuses analyzed. Two major goals have been achieved. First, inactivation of the endogenous IgM locus, which is an essential step for the production of therapeutic human polyclonal antibodies in the rabbit. Second, establishing efficient targeted gene manipulation and homologous recombination in a refractory animal species. ZFN mediated genetic engineering in the rabbit and other mammals opens new avenues of experimentation in immunology and many other research fields.

  14. ZFN-mediated gene targeting of the Arabidopsis protoporphyrinogen oxidase gene through Agrobacterium-mediated floral dip transformation.

    Science.gov (United States)

    de Pater, Sylvia; Pinas, Johan E; Hooykaas, Paul J J; van der Zaal, Bert J

    2013-05-01

    Previously, we showed that ZFN-mediated induction of double-strand breaks (DSBs) at the intended recombination site enhanced the frequency of gene targeting (GT) at an artificial target locus using Agrobacterium-mediated floral dip transformation. Here, we designed zinc finger nucleases (ZFNs) for induction of DSBs in the natural protoporphyrinogen oxidase (PPO) gene, which can be conveniently utilized for GT experiments. Wild-type Arabidopsis plants and plants expressing the ZFNs were transformed via floral dip transformation with a repair T-DNA with an incomplete PPO gene, missing the 5' coding region but containing two mutations rendering the enzyme insensitive to the herbicide butafenacil as well as an extra KpnI site for molecular analysis of GT events. Selection on butafenacil yielded 2 GT events for the wild type with a frequency of 0.8 × 10⁻³ per transformation event and 8 GT events for the ZFNs expressing plant line with a frequency of 3.1 × 10⁻³ per transformation event. Molecular analysis using PCR and Southern blot analysis showed that 9 of the GT events were so-called true GT events, repaired via homologous recombination (HR) at the 5' and the 3' end of the gene. One plant line contained a PPO gene repaired only at the 5' end via HR. Most plant lines contained extra randomly integrated T-DNA copies. Two plant lines did not contain extra T-DNAs, and the repaired PPO genes in these lines were transmitted to the next generation in a Mendelian fashion.

  15. Update on Aurora Kinase Targeted Therapeutics in Oncology

    Science.gov (United States)

    Green, Myke R.; Woolery, Joseph E.

    2011-01-01

    Introduction Mammalian cells contain three distinct serine/threonine protein kinases with highly conserved catalytic domains, including aurora A and B kinases that are essential regulators of mitotic entry and progression. Overexpression of aurora A and/or B kinase is associated with high proliferation rates and poor prognosis, making them ideal targets for anti-cancer therapy. Disruption of mitotic machinery is a proven anti-cancer strategy employed by multiple chemotherapeutic agents. Numerous small molecule inhibitors of the aurora kinases have been discovered and tested in vivo and in vitro, with a few currently in phase II testing. Areas covered This review provides the reader with updated results from both preclinical and human studies for each of the aurora kinase inhibitors (AKI) that are currently being investigated. The paper also covers in detail the late breaking and phase I data presented for AKIs thereby allowing the reader to compare and contrast individual and classrelated effects of AKIs. Expert opinion While the successful development and approval of an AKI for anti-cancer therapy remains unresolved, pre-clinical identification of resistant mechanisms would help design better early phase clinical trials where relevant combinations may be evaluated prior to phase II testing. The authors believe that aurora kinases are important anti-cancer targets that operate in collaboration with other oncogenes intimately involved in uncontrolled tumor proliferation and by providing a unique, targeted and complimentary anti-cancer mechanism, expand the available armamentarium against cancer. PMID:21556291

  16. Global Identification of EVI1 Target Genes in Acute Myeloid Leukemia.

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    Carolyn Glass

    Full Text Available The ecotropic virus integration site 1 (EVI1 transcription factor is associated with human myeloid malignancy of poor prognosis and is overexpressed in 8-10% of adult AML and strikingly up to 27% of pediatric MLL-rearranged leukemias. For the first time, we report comprehensive genomewide EVI1 binding and whole transcriptome gene deregulation in leukemic cells using a combination of ChIP-Seq and RNA-Seq expression profiling. We found disruption of terminal myeloid differentiation and cell cycle regulation to be prominent in EVI-induced leukemogenesis. Specifically, we identified EVI1 directly binds to and downregulates the master myeloid differentiation gene Cebpe and several of its downstream gene targets critical for terminal myeloid differentiation. We also found EVI1 binds to and downregulates Serpinb2 as well as numerous genes involved in the Jak-Stat signaling pathway. Finally, we identified decreased expression of several ATP-dependent P2X purinoreceptors genes involved in apoptosis mechanisms. These findings provide a foundation for future study of potential therapeutic gene targets for EVI1-induced leukemia.

  17. An Oomycete CRN Effector Reprograms Expression of Plant HSP Genes by Targeting their Promoters.

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    Tianqiao Song

    2015-12-01

    Full Text Available Oomycete pathogens produce a large number of CRN effectors to manipulate plant immune responses and promote infection. However, their functional mechanisms are largely unknown. Here, we identified a Phytophthora sojae CRN effector PsCRN108 which contains a putative DNA-binding helix-hairpin-helix (HhH motif and acts in the plant cell nucleus. Silencing of the PsCRN108 gene reduced P. sojae virulence to soybean, while expression of the gene in Nicotiana benthamiana and Arabidopsis thaliana enhanced plant susceptibility to P. capsici. Moreover, PsCRN108 could inhibit expression of HSP genes in A. thaliana, N. benthamiana and soybean. Both the HhH motif and nuclear localization signal of this effector were required for its contribution to virulence and its suppression of HSP gene expression. Furthermore, we found that PsCRN108 targeted HSP promoters in an HSE- and HhH motif-dependent manner. PsCRN108 could inhibit the association of the HSE with the plant heat shock transcription factor AtHsfA1a, which initializes HSP gene expression in response to stress. Therefore, our data support a role for PsCRN108 as a nucleomodulin in down-regulating the expression of plant defense-related genes by directly targeting specific plant promoters.

  18. Hapten-derivatized nanoparticle targeting and imaging of gene expression by multimodality imaging systems.

    Science.gov (United States)

    Cheng, C-M; Chu, P-Y; Chuang, K-H; Roffler, S R; Kao, C-H; Tseng, W-L; Shiea, J; Chang, W-D; Su, Y-C; Chen, B-M; Wang, Y-M; Cheng, T-L

    2009-01-01

    Non-invasive gene monitoring is important for most gene therapy applications to ensure selective gene transfer to specific cells or tissues. We developed a non-invasive imaging system to assess the location and persistence of gene expression by anchoring an anti-dansyl (DNS) single-chain antibody (DNS receptor) on the cell surface to trap DNS-derivatized imaging probes. DNS hapten was covalently attached to cross-linked iron oxide (CLIO) to form a 39+/-0.5 nm DNS-CLIO nanoparticle imaging probe. DNS-CLIO specifically bound to DNS receptors but not to a control single-chain antibody receptor. DNS-CLIO (100 microM Fe) was non-toxic to both B16/DNS (DNS receptor positive) and B16/phOx (control receptor positive) cells. Magnetic resonance (MR) imaging could detect as few as 10% B16/DNS cells in a mixture in vitro. Importantly, DNS-CLIO specifically bound to a B16/DNS tumor, which markedly reduced signal intensity. Similar results were also shown with DNS quantum dots, which specifically targeted CT26/DNS cells but not control CT26/phOx cells both in vitro and in vivo. These results demonstrate that DNS nanoparticles can systemically monitor the expression of DNS receptor in vivo by feasible imaging systems. This targeting strategy may provide a valuable tool to estimate the efficacy and specificity of different gene delivery systems and optimize gene therapy protocols in the clinic.

  19. An Oomycete CRN Effector Reprograms Expression of Plant HSP Genes by Targeting their Promoters.

    Science.gov (United States)

    Song, Tianqiao; Ma, Zhenchuan; Shen, Danyu; Li, Qi; Li, Wanlin; Su, Liming; Ye, Tingyue; Zhang, Meixiang; Wang, Yuanchao; Dou, Daolong

    2015-12-01

    Oomycete pathogens produce a large number of CRN effectors to manipulate plant immune responses and promote infection. However, their functional mechanisms are largely unknown. Here, we identified a Phytophthora sojae CRN effector PsCRN108 which contains a putative DNA-binding helix-hairpin-helix (HhH) motif and acts in the plant cell nucleus. Silencing of the PsCRN108 gene reduced P. sojae virulence to soybean, while expression of the gene in Nicotiana benthamiana and Arabidopsis thaliana enhanced plant susceptibility to P. capsici. Moreover, PsCRN108 could inhibit expression of HSP genes in A. thaliana, N. benthamiana and soybean. Both the HhH motif and nuclear localization signal of this effector were required for its contribution to virulence and its suppression of HSP gene expression. Furthermore, we found that PsCRN108 targeted HSP promoters in an HSE- and HhH motif-dependent manner. PsCRN108 could inhibit the association of the HSE with the plant heat shock transcription factor AtHsfA1a, which initializes HSP gene expression in response to stress. Therefore, our data support a role for PsCRN108 as a nucleomodulin in down-regulating the expression of plant defense-related genes by directly targeting specific plant promoters.

  20. H2S donor, S-propargyl-cysteine, increases CSE in SGC-7901 and cancer-induced mice: evidence for a novel anti-cancer effect of endogenous H2S?

    Directory of Open Access Journals (Sweden)

    Kaium Ma

    Full Text Available BACKGROUND: S-propargyl-cysteine (SPRC, an H(2S donor, is a structural analogue of S-allycysteine (SAC. It was investigated for its potential anti-cancer effect on SGC-7901 gastric cancer cells and the possible mechanisms that may be involved. METHODS AND FINDINGS: SPRC treatment significantly decreased cell viability, suppressed the proliferation and migration of SPRC-7901 gastric cancer cells, was pro-apoptotic as well as caused cell cycle arrest at the G(1/S phase. In an in vivo study, intra-peritoneal injection of 50 mg/kg and 100 mg/kg of SPRC significantly reduced tumor weights and tumor volumes of gastric cancer implants in nude mice, with a tumor growth inhibition rate of 40-75%. SPRC also induced a pro-apoptotic effect in cancer tissues and elevated the expressions of p53 and Bax in tumors and cells. SPRC treatment also increased protein expression of cystathione-γ-lyase (CSE in cells and tumors, and elevated H(2S levels in cell culture media, plasma and tumoral CSE activity of gastric cancer-induced nude mice by 2, 2.3 and 1.4 fold, respectively. Most of the anti-cancer functions of SPRC on cells and tumors were significantly suppressed by PAG, an inhibitor of CSE activity. CONCLUSIONS: Taken together, the results of our study provide insights into a novel anti-cancer effect of H(2S as well as of SPRC on gastric cancer through inducing the activity of a new target, CSE.

  1. Improvement of Hydrodynamics-Based Gene Transfer of Nonviral DNA Targeted to Murine Hepatocytes

    Directory of Open Access Journals (Sweden)

    Shingo Nakamura

    2013-01-01

    Full Text Available The liver is an important organ for supporting the life of an individual. Gene transfer toward this organ has been attempted in many laboratories to date; however, there have been few reports on improved liver-targeted gene delivery by using a nonviral vector. In this study, we examined the effect of various types of gene delivery carriers on enhancing the uptake and gene expression of exogenous DNA in murine hepatocytes when a hydrodynamics-based gene delivery (HGD is performed via tail-vein injection. Mice were singly injected with a large amount of phosphate-buffered saline containing reporter plasmid DNA and/or with a gene delivery carrier. One day after the gene delivery, the animals' livers were dissected and subjected to biochemical, histochemical, and molecular biological analyses. The strongest signal from the reporter plasmid DNA was observed when the DNA was mixed with a polyethylenimine- (PEI- based reagent. Coinjection with pCRTEIL (a loxP-floxed reporter construct and pTR/NCre (a liver-specific Cre expression vector resulted in the liver-specific recombination of pCRTEIL. The combination of PEI with HGD would thus be a valuable tool for liver-specific manipulation to examine the function of a gene of interest in the liver and for creating liver disease models.

  2. Differential effects of black raspberry and strawberry extracts on BaPDE-induced activation of transcription factors and their target genes.

    Science.gov (United States)

    Li, Jingxia; Zhang, Dongyun; Stoner, Gary D; Huang, Chuanshu

    2008-04-01

    The chemopreventive properties of edible berries have been demonstrated both in vitro and in vivo, however, the specific molecular mechanisms underlying their anti-cancer effects are largely unknown. Our previous studies have shown that a methanol extract fraction of freeze-dried black raspberries inhibits benzoapyrene (BaP)-induced transformation of Syrian hamster embryo cells. This fraction also blocks activation of activator protein-1 (AP-1) and nuclear factor kappaB (NF-kappaB) induced by benzoapyrene diol-epoxide (BaPDE) in mouse epidermal JB6 Cl 41 cells. To determine if different berry types exhibit specific mechanisms for their anti-cancer effects, we compared the effects of extract fractions from both black raspberries and strawberries on BaPDE-induced activation of various signaling pathways in Cl 41 cells. Black raspberry fractions inhibited the activation of AP-1, NF-kappaB, and nuclear factor of activated T cells (NFAT) by BaPDE as well as their upstream PI-3K/Akt-p70(S6K) and mitogen-activated protein kinase pathways. In contrast, strawberry fractions inhibited NFAT activation, but did not inhibit the activation of AP-1, NF-kappaB or the PI-3K/Akt-p70(S6K) and mitogen-activated protein kinase pathways. Consistent with the effects on NFAT activation, tumor necrosis factor-alpha (TNF-alpha) induction by BaPDE was blocked by extract fractions of both black raspberries and strawberries, whereas vascular endothelial growth factor (VEGF) expression, which depends on AP-1 activation, was suppressed by black raspberry fractions but not strawberry fractions. These results suggest that black raspberry and strawberry components may target different signaling pathways in exerting their anti-carcinogenic effects.

  3. A bioinformatics tool for linking gene expression profiling results with public databases of microRNA target predictions

    Science.gov (United States)

    Creighton, Chad J.; Nagaraja, Ankur K.; Hanash, Samir M.; Matzuk, Martin M.; Gunaratne, Preethi H.

    2008-01-01

    MicroRNAs are short (∼22 nucleotides) noncoding RNAs that regulate the stability and translation of mRNA targets. A number of computational algorithms have been developed to help predict which microRNAs are likely to regulate which genes. Gene expression profiling of biological systems where microRNAs might be active can yield hundreds of differentially expressed genes. The commonly used public microRNA target prediction databases facilitate gene-by-gene searches. However, integration of microRNA–mRNA target predictions with gene expression data on a large scale using these databases is currently cumbersome and time consuming for many researchers. We have developed a desktop software application which, for a given target prediction database, retrieves all microRNA:mRNA functional pairs represented by an experimentally derived set of genes. Furthermore, for each microRNA, the software computes an enrichment statistic for overrepresentation of predicted targets within the gene set, which could help to implicate roles for specific microRNAs and microRNA-regulated genes in the system under study. Currently, the software supports searching of results from PicTar, TargetScan, and miRanda algorithms. In addition, the software can accept any user-defined set of gene-to-class associations for searching, which can include the results of other target prediction algorithms, as well as gene annotation or gene-to-pathway associations. A search (using our software) of genes transcriptionally regulated in vitro by estrogen in breast cancer uncovered numerous targeting associations for specific microRNAs—above what could be observed in randomly generated gene lists—suggesting a role for microRNAs in mediating the estrogen response. The software and Excel VBA source code are freely available at http://sigterms.sourceforge.net. PMID:18812437

  4. A bioinformatics tool for linking gene expression profiling results with public databases of microRNA target predictions.

    Science.gov (United States)

    Creighton, Chad J; Nagaraja, Ankur K; Hanash, Samir M; Matzuk, Martin M; Gunaratne, Preethi H

    2008-11-01

    MicroRNAs are short (approximately 22 nucleotides) noncoding RNAs that regulate the stability and translation of mRNA targets. A number of computational algorithms have been developed to help predict which microRNAs are likely to regulate which genes. Gene expression profiling of biological systems where microRNAs might be active can yield hundreds of differentially expressed genes. The commonly used public microRNA target prediction databases facilitate gene-by-gene searches. However, integration of microRNA-mRNA target predictions with gene expression data on a large scale using these databases is currently cumbersome and time consuming for many researchers. We have developed a desktop software application which, for a given target prediction database, retrieves all microRNA:mRNA functional pairs represented by an experimentally derived set of genes. Furthermore, for each microRNA, the software computes an enrichment statistic for overrepresentation of predicted targets within the gene set, which could help to implicate roles for specific microRNAs and microRNA-regulated genes in the system under study. Currently, the software supports searching of results from PicTar, TargetScan, and miRanda algorithms. In addition, the software can accept any user-defined set of gene-to-class associations for searching, which can include the results of other target prediction algorithms, as well as gene annotation or gene-to-pathway associations. A search (using our software) of genes transcriptionally regulated in vitro by estrogen in breast cancer uncovered numerous targeting associations for specific microRNAs-above what could be observed in randomly generated gene lists-suggesting a role for microRNAs in mediating the estrogen response. The software and Excel VBA source code are freely available at http://sigterms.sourceforge.net.

  5. Comparative Proteomic Analysis of Anti-Cancer Mechanism by Periplocin Treatment in Lung Cancer Cells

    Directory of Open Access Journals (Sweden)

    Zejun Lu

    2014-03-01

    Full Text Available Background: Periplocin is used for treatment of rheumatoid arthritis, reinforcement of bones and tendons, palpitations or shortness of breath and lower extremity edema in traditional medicine. Our previous findings suggested that periplocin could inhibit the growth of lung cancer both in vitro and in vivo. But the biological processes and molecular pathways by which periplocin induces these beneficial effects remain largely undefined. Methods: To explore the molecular mechanisms of periplocin involved in anti-cancer activity, in the present study the protein profile changes of human lung cancer cell lines A549 in response to periplocin treatment were investigated using the proteomics approaches (2-DE combined with MS/MS. Western blot was employed to verify the changed proteins. Interactions between changed proteins were analyzed by STRING. Results: 29 down-regulated protein species named GTP-binding nuclear protein Ran (RAN, Rho GDP-dissociation inhibitor 1 (ARHGDIA, eukaryotic translation initiation factor 5A-1 (EIF5A and Profilin-1(PFN1, and 10 up-regulated protein species named Heat shock cognate 71 kDa protein (HSPA8,10 kDa heat shock protein (HSPE1, and Cofilin-1(CFL-1 were identified. Among them, GTP-binding nuclear protein Ran (RAN and Rho GDP-dissociation inhibitor 1 (ARHGDIA were the most significantly changed (over tenfold. The proteasome subunit beta type-6 (PSMB6, ATP synthase ecto-α-subunit (ATP5A1, Aldehyde dehydrogenase 1 (ALDH1 and EIF5A were verified by immunoblot assays to be dramatically down-regulated. By STRING bioinformatics analysis revealing interactions and signaling networks it became apparent that the proteins changed they are primarily involved in transcription and proteolysis. Conclusion: Periplocin inhibited growth of lung cancer by down-regulating proteins, such as ATP5A1, EIF5A, ALDH1 and PSMB6. These findings may improve our understanding of the molecular mechanisms underlying the anti-cancer effects of

  6. Targeting single neuronal networks for gene expression and cell labeling in vivo.

    Science.gov (United States)

    Marshel, James H; Mori, Takuma; Nielsen, Kristina J; Callaway, Edward M

    2010-08-26

    To understand fine-scale structure and function of single mammalian neuronal networks, we developed and validated a strategy to genetically target and trace monosynaptic inputs to a single neuron in vitro and in vivo. The strategy independently targets a neuron and its presynaptic network for specific gene expression and fine-scale labeling, using single-cell electroporation of DNA to target infection and monosynaptic retrograde spread of a genetically modifiable rabies virus. The technique is highly reliable, with transsynaptic labeling occurring in every electroporated neuron infected by the virus. Targeting single neocortical neuronal networks in vivo, we found clusters of both spiny and aspiny neurons surrounding the electroporated neuron in each case, in addition to intricately labeled distal cortical and subcortical inputs. This technique, broadly applicable for probing and manipulating single neuronal networks with single-cell resolution in vivo, may help shed new light on fundamental mechanisms underlying circuit development and information processing by neuronal networks throughout the brain.

  7. Evaluating Transcription Factor Activity Changes by Scoring Unexplained Target Genes in Expression Data

    Science.gov (United States)

    Berchtold, Evi; Csaba, Gergely; Zimmer, Ralf

    2016-01-01

    Several methods predict activity changes of transcription factors (TFs) from a given regulatory network and measured expression data. But available gene regulatory networks are incomplete and contain many condition-dependent regulations that are not relevant for the specific expression measurement. It is not known which combination of active TFs is needed to cause a change in the expression of a target gene. A method to systematically evaluate the inferred activity changes is missing. We present such an evaluation strategy that indicates for how many target genes the observed expression changes can be explained by a given set of active TFs. To overcome the problem that the exact combination of active TFs needed to activate a gene is typically not known, we assume a gene to be explained if there exists any combination for which the predicted active TFs can possibly explain the observed change of the gene. We introduce the i-score (inconsistency score), which quantifies how many genes could not be explained by the set of activity changes of TFs. We observe that, even for these minimal requirements, published methods yield many unexplained target genes, i.e. large i-scores. This holds for all methods and all expression datasets we evaluated. We provide new optimization methods to calculate the best possible (minimal) i-score given the network and measured expression data. The evaluation of this optimized i-score on a large data compendium yields many unexplained target genes for almost every case. This indicates that currently available regulatory networks are still far from being complete. Both the presented Act-SAT and Act-A* methods produce optimal sets of TF activity changes, which can be used to investigate the difficult interplay of expression and network data. A web server and a command line tool to calculate our i-score and to find the active TFs associated with the minimal i-score is available from https://services.bio.ifi.lmu.de/i-score. PMID:27723775

  8. Cholinergic nicotinic receptor genes implicated in a nicotine dependence association study targeting 348 candidate genes with 3713 SNPs.

    Science.gov (United States)

    Saccone, Scott F; Hinrichs, Anthony L; Saccone, Nancy L; Chase, Gary A; Konvicka, Karel; Madden, Pamela A F; Breslau, Naomi; Johnson, Eric O; Hatsukami, Dorothy; Pomerleau, Ovide; Swan, Gary E; Goate, Alison M; Rutter, Joni; Bertelsen, Sarah; Fox, Louis; Fugman, Douglas; Martin, Nicholas G; Montgomery, Grant W; Wang, Jen C; Ballinger, Dennis G; Rice, John P; Bierut, Laura Jean

    2007-01-01

    Nicotine dependence is one of the world's leading causes of preventable death. To discover genetic variants that influence risk for nicotine dependence, we targeted over 300 candidate genes and analyzed 3713 single nucleotide polymorphisms (SNPs) in 1050 cases and 879 controls. The Fagerström test for nicotine dependence (FTND) was used to assess dependence, in which cases were required to have an FTND of 4 or more. The control criterion was strict: control subjects must have smoked at least 100 cigarettes in their lifetimes and had an FTND of 0 during the heaviest period of smoking. After correcting for multiple testing by controlling the false discovery rate, several cholinergic nicotinic receptor genes dominated the top signals. The strongest association was from an SNP representing CHRNB3, the beta3 nicotinic receptor subunit gene (P = 9.4 x 10(-5)). Biologically, the most compelling evidence for a risk variant came from a non-synonymous SNP in the alpha5 nicotinic receptor subunit gene CHRNA5 (P = 6.4 x 10(-4)). This SNP exhibited evidence of a recessive mode of inheritance, resulting in individuals having a 2-fold increase in risk of developing nicotine dependence once exposed to cigarette smoking. Other genes among the top signals were KCNJ6 and GABRA4. This study represents one of the most powerful and extensive studies of nicotine dependence to date and has found novel risk loci that require confirmation by replication studies.

  9. Targeting c-Myc-activated genes with a correlation method: Detection of global changes in large gene expression network dynamics

    Science.gov (United States)

    Remondini, D.; O'Connell, B.; Intrator, N.; Sedivy, J. M.; Neretti, N.; Castellani, G. C.; Cooper, L. N.

    2005-01-01

    This work studies the dynamics of a gene expression time series network. The network, which is obtained from the correlation of gene expressions, exhibits global dynamic properties that emerge after a cell state perturbation. The main features of this network appear to be more robust when compared with those obtained with a network obtained from a linear Markov model. In particular, the network properties strongly depend on the exact time sequence relationships between genes and are destroyed by random temporal data shuffling. We discuss in detail the problem of finding targets of the c-myc protooncogene, which encodes a transcriptional regulator whose inappropriate expression has been correlated with a wide array of malignancies. The data used for network construction are a time series of gene expression, collected by microarray analysis of a rat fibroblast cell line expressing a conditional Myc-estrogen receptor oncoprotein. We show that the correlation-based model can establish a clear relationship between network structure and the cascade of c-myc-activated genes. PMID:15867157

  10. Gene targeting in rats using transcription activator-like effector nucleases.

    Science.gov (United States)

    Ménoret, Séverine; Tesson, Laurent; Rémy, Séverine; Usal, Claire; Thépenier, Virginie; Thinard, Reynald; Ouisse, Laure-Hélène; De Cian, Anne; Giovannangeli, Carine; Concordet, Jean-Paul; Anegon, Ignacio

    2014-08-15

    The rat is a model of choice to understanding gene function and modeling human diseases. Since recent years, successful engineering technologies using gene-specific nucleases have been developed to gene edit the genome of different species, including the rat. This development has become important for the creation of new rat animals models of human diseases, analyze the role of genes and express recombinant proteins. Transcription activator-like (TALE) nucleases are designed nucleases consist of a DNA binding domain fused to a nuclease domain capable of cleaving the targeted DNA. We describe a detailed protocol for generating knockout rats via microinjection of TALE nucleases into fertilized eggs. This technology is an efficient, cost- and time-effective method for creating new rat models.

  11. Identification and characterization of a novel peptide ligand of Tie2 for targeting gene therapy

    Institute of Scientific and Technical Information of China (English)

    Xianghua Wu; Jianren Gu; Zonghai Li; Ming Yao; Huamao Wang; Sumin Qu; Xianlian Chen; Jinjun Li; Ye Sun; Yuhong Xu

    2008-01-01

    Tyrosine kinase with immunoglobulin and epidermal growth factor homology domain-2 (Tie2) has been considered as a rational target for gene therapy in solid tumors. In order to identify a novel peptide ligand of Tie2 for targeted gene therapy, we screened a phage display peptide library and identified a candidate peptide ligand NSLSNASEFRAPY(designated GA5).Binding assays and Scatchard analysis revealed that GA5 could specifically bind to Tie2 with a dissociation constant of 2.1×10-8M.In addition,we showed that GA5 was internalized into tumor cells highly expressing Tie2.In the biodistribution assay.125I-GA5 was mainly accumniated in SPC-A1 xenograft tumors that express Tie2.Ingene delivery studies,GA5-conjugated polyethylenimine vector could achieve greater transgene transduction than non-targeted vectors both in vitro and in vivo.Tumor growth inhibition was observed in SPC-A1 xenograft-bearing mice that received eight intratumoral injections of GA5 polyethylenimine/p53 complexes in 3 weeks.The difference in tumor volume between the experiment and control groups was significant(P<0.05).Our results showed that GA5 is a potentially efficient targeting element for cancer gene or molecular therapy.

  12. Regeneration of pancreatic islets in vivo by ultrasound-targeted gene therapy.

    Science.gov (United States)

    Chen, S; Shimoda, M; Wang, M-Y; Ding, J; Noguchi, H; Matsumoto, S; Grayburn, P A

    2010-11-01

    This study uses a novel approach to gene therapy in which plasmid DNA is targeted to the pancreas in vivo using ultrasound-targeted microbubble destruction (UTMD) to achieve islet regeneration. Intravenous microbubbles carrying plasmids are destroyed within the pancreatic microcirculation by ultrasound, achieving local gene expression that is further targeted to β-cells by a modified rat insulin promoter (RIP3.1). A series of genes implicated in endocrine development were delivered to rats 2 days after streptozotocin-induced diabetes. The genes, PAX4, Nkx2.2, Nkx6.1, Ngn3 and Mafa, produced α-cell hyperplasia, but no significant improvement in β-cell mass or blood glucose level 30 days after UTMD. In contrast, RIP3.1-NeuroD1 promoted islet regeneration from surviving β-cells, with normalization of glucose, insulin and C-peptide levels at 30 days. In a longer-term experiment, four of six rats had a return of diabetes at 90 days, accompanied by β-cell apoptosis on Tunel staining. Pretreatment with the JNK inhibitor SP600125 successfully blocked β-cell apoptosis and resulted in restoration of β-cell mass and normalization of blood glucose level for up to 90 days. This technique allows in vivo islet regeneration, restoration of β-cell mass and normalization of blood sugar, insulin and C-peptide in rats without viruses.

  13. Zooplankton community analysis in the Changjiang River estuary by single-gene-targeted metagenomics

    Science.gov (United States)

    Cheng, Fangping; Wang, Minxiao; Li, Chaolun; Sun, Song

    2014-07-01

    DNA barcoding provides accurate identification of zooplankton species through all life stages. Single-gene-targeted metagenomic analysis based on DNA barcode databases can facilitate longterm monitoring of zooplankton communities. With the help of the available zooplankton databases, the zooplankton community of the Changjiang (Yangtze) River estuary was studied using a single-gene-targeted metagenomic method to estimate the species richness of this community. A total of 856 mitochondrial cytochrome oxidase subunit 1 (cox1) gene sequences were determined. The environmental barcodes were clustered into 70 molecular operational taxonomic units (MOTUs). Forty-two MOTUs matched barcoded marine organisms with more than 90% similarity and were assigned to either the species (similarity>96%) or genus level (similaritymorphological methods were identified by molecular methods, especially gelatinous zooplankton and merozooplankton that were likely sampled at different life history phases. Zooplankton community structures differed significantly among all of the samples. The MOTU spatial distributions were influenced by the ecological habits of the corresponding species. In conclusion, single-gene-targeted metagenomic analysis is a useful tool for zooplankton studies, with which specimens from all life history stages can be identified quickly and effectively with a comprehensive database.

  14. Using Pharmacogenomic Databases for Discovering Patient-Target Genes and Small Molecule Candidates to Cancer Therapy

    Science.gov (United States)

    Belizário, José E.; Sangiuliano, Beatriz A.; Perez-Sosa, Marcela; Neyra, Jennifer M.; Moreira, Dayson F.

    2016-01-01

    With multiple omics strategies being applied to several cancer genomics projects, researchers have the opportunity to develop a rational planning of targeted cancer therapy. The investigation of such numerous and diverse pharmacogenomic datasets is a complex task. It requires biological knowledge and skills on a set of tools to accurately predict signaling network and clinical outcomes. Herein, we describe Web-based in silico approaches user friendly for exploring integrative studies on cancer biology and pharmacogenomics. We briefly explain how to submit a query to cancer genome databases to predict which genes are significantly altered across several types of cancers using CBioPortal. Moreover, we describe how to identify clinically available drugs and potential small molecules for gene targeting using CellMiner. We also show how to generate a gene signature and compare gene expression profiles to investigate the complex biology behind drug response using Connectivity Map. Furthermore, we discuss on-going challenges, limitations and new directions to integrate molecular, biological and epidemiological information from oncogenomics platforms to create hypothesis-driven projects. Finally, we discuss the use of Patient-Derived Xenografts models (PDXs) for drug profiling in vivo assay. These platforms and approaches are a rational way to predict patient-targeted therapy response and to develop clinically relevant small molecules drugs.

  15. An Efficient Method for Identifying Gene Fusions by Targeted RNA Sequencing from Fresh Frozen and FFPE Samples.

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    Jonathan A Scolnick

    Full Text Available Fusion genes are known to be key drivers of tumor growth in several types of cancer. Traditionally, detecting fusion genes has been a difficult task based on fluorescent in situ hybridization to detect chromosomal abnormalities. More recently, RNA sequencing has enabled an increased pace of fusion gene identification. However, RNA-Seq is inefficient for the identification of fusion genes due to the high number of sequencing reads needed to detect the small number of fusion transcripts present in cells of interest. Here we describe a method, Single Primer Enrichment Technology (SPET, for targeted RNA sequencing that is customizable to any target genes, is simple to use, and efficiently detects gene fusions. Using SPET to target 5701 exons of 401 known cancer fusion genes for sequencing, we were able to identify known and previously unreported gene fusions from both fresh-frozen and formalin-fixed paraffin-embedded (FFPE tissue RNA in both normal tissue and cancer cells.

  16. An Efficient Method for Identifying Gene Fusions by Targeted RNA Sequencing from Fresh Frozen and FFPE Samples.

    Science.gov (United States)

    Scolnick, Jonathan A; Dimon, Michelle; Wang, I-Ching; Huelga, Stephanie C; Amorese, Douglas A

    2015-01-01

    Fusion genes are known to be key drivers of tumor growth in several types of cancer. Traditionally, detecting fusion genes has been a difficult task based on fluorescent in situ hybridization to detect chromosomal abnormalities. More recently, RNA sequencing has enabled an increased pace of fusion gene identification. However, RNA-Seq is inefficient for the identification of fusion genes due to the high number of sequencing reads needed to detect the small number of fusion transcripts present in cells of interest. Here we describe a method, Single Primer Enrichment Technology (SPET), for targeted RNA sequencing that is customizable to any target genes, is simple to use, and efficiently detects gene fusions. Using SPET to target 5701 exons of 401 known cancer fusion genes for sequencing, we were able to identify known and previously unreported gene fusions from both fresh-frozen and formalin-fixed paraffin-embedded (FFPE) tissue RNA in both normal tissue and cancer cells.

  17. Identification of estrogen target genes during zebrafish embryonic development through transcriptomic analysis.

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    Ruixin Hao

    Full Text Available Estrogen signaling is important for vertebrate embryonic development. Here we have used zebrafish (Danio rerio as a vertebrate model to analyze estrogen signaling during development. Zebrafish embryos were exposed to 1 µM 17β-estradiol (E2 or vehicle from 3 hours to 4 days post fertilization (dpf, harvested at 1, 2, 3 and 4 dpf, and subjected to RNA extraction for transcriptome analysis using microarrays. Differentially expressed genes by E2-treatment were analyzed with hierarchical clustering followed by biological process and tissue enrichment analysis. Markedly distinct sets of genes were up and down-regulated by E2 at the four different time points. Among these genes, only the well-known estrogenic marker vtg1 was co-regulated at all time points. Despite this, the biological functional categories targeted by E2 were relatively similar throughout zebrafish development. According to knowledge-based tissue enrichment, estrogen responsive genes were clustered mainly in the liver, pancreas and brain. This was in line with the developmental dynamics of estrogen-target tissues that were visualized using transgenic zebrafish containing estrogen responsive elements driving the expression of GFP (Tg(5xERE:GFP. Finally, the identified embryonic estrogen-responsive genes were compared to already published estrogen-responsive genes identified in male adult zebrafish (Gene Expression Omnibus database. The expressions of a few genes were co-regulated by E2 in both embryonic and adult zebrafish. These could potentially be used as estrogenic biomarkers for exposure to estrogens or estrogenic endocrine disruptors in zebrafish. In conclusion, our data suggests that estrogen effects on early embryonic zebrafish development are stage- and tissue- specific.

  18. IGF-IR targeted therapy: Past, present and future

    NARCIS (Netherlands)

    J.A.M.J.L. Janssen (Joseph); A.J. Varewijck (Aimee)

    2014-01-01

    textabstractThe IGF-I receptor (IGF-IR) has been studied as an anti-cancer target. However, monotherapy trials with IGF-IR targeted antibodies or with IGF-IR specific tyrosine kinase inhibitors have, overall, been very disappointing in the clinical setting. This review discusses potential reasons wh

  19. Efficient gene targeting mediated by a lentiviral vector-associated meganuclease.

    Science.gov (United States)

    Izmiryan, Araksya; Basmaciogullari, Stéphane; Henry, Adrien; Paques, Frédéric; Danos, Olivier

    2011-09-01

    Gene targeting can be achieved with lentiviral vectors delivering donor sequences along with a nuclease that creates a locus-specific double-strand break (DSB). Therapeutic applications of this system would require an appropriate control of the amount of endonuclease delivered to the target cells, and potentially toxic sustained expression must be avoided. Here, we show that the nuclease can be transferred into cells as a protein associated with a lentiviral vector particle. I-SceI, a prototypic meganuclease from yeast, was incorporated into the virions as a fusion with Vpr, an HIV accessory protein. Integration-deficient lentiviral vectors containing the donor sequences and the I-SceI fusion protein were tested in reporter cells in which targeting events were scored by the repair of a puromycin resistance gene. Molecular analysis of the targeted locus indicated a 2-fold higher frequency of the expected recombination event when the nuclease was delivered as a protein rather than encoded by a separate vector. In both systems, a proportion of clones displayed multiple integrated copies of the donor sequences, either as tandems at the targeted locus or at unrelated loci. These integration patterns were dependent upon the mode of meganuclease delivery, suggesting distinct recombination processes.

  20. Comparison of target labeling methods for use with Affymetrix GeneChips

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    Vernon Suzanne D

    2007-05-01

    Full Text Available Abstract Background Several different commercial one-cycle labeling kits are available for preparation of the target for use with the Affymetrix GeneChip platform. However, there have been no evaluations of these different kits to determine if comparable results were generated. We report on the cRNA target synthesis, labeling efficiency and hybridization results using the One-Cycle Target Labeling Assay™ (Affymetrix, the BioArray RNA Amplification and Labeling System™ (Enzo Life Sciences, and the Superscript RNA Amplification System (Invitrogen Life Technologies. Results The only notable difference between kits was in the yield of cRNA target synthesized during in vitro transcription, where the BioArray assay had to be repeated several times in order to have sufficient target. However, each kit resulted in comparable signal and detection calls when hybridized to the Affymetrix GeneChip. Conclusion These 3 one-cycle labeling kits produce comparable hybridization results. This provides users with several kit options and flexibility when using the Affymetrix system.

  1. A system for the measurement of gene targeting efficiency in human cell lines using an antibiotic resistance-GFP fusion gene.

    Science.gov (United States)

    Konishi, Yuko; Karnan, Sivasundaram; Takahashi, Miyuki; Ota, Akinobu; Damdindorj, Lkhagvasuren; Hosokawa, Yoshitaka; Konishi, Hiroyuki

    2012-09-01

    Gene targeting in a broad range of human somatic cell lines has been hampered by inefficient homologous recombination. To improve this technology and facilitate its widespread application, it is critical to first have a robust and efficient research system for measuring gene targeting efficiency. Here, using a fusion gene consisting of hygromycin B phosphotransferase and 3'-truncated enhanced GFP (HygR-5' EGFP) as a reporter gene, we created a molecular system monitoring the ratio of homologous to random integration (H/R ratio) of targeting vectors into the genome. Cell clones transduced with a reporter vector containing HygR-5' EGFP were efficiently established from two human somatic cell lines. Established HygR-5' EGFP reporter clones retained their capacity to monitor gene targeting efficiency for a longer duration than a conventional reporter system using an unfused 5' EGFP gene. With the HygR-5' EGFP reporter system, we reproduced previous findings of gene targeting frequency being up-regulated by the use of an adeno-associated viral (AAV) backbone, a promoter-trap system, or a longer homology arm in a targeting vector, suggesting that this system accurately monitors H/R ratio. Thus, our HygR-5' EGFP reporter system will assist in the development of an efficient AAV-based gene targeting technology.

  2. Differential Diagnosis of Two Chinese Families with Dyschromatoses by Targeted Gene Sequencing

    Institute of Scientific and Technical Information of China (English)

    Jia-Wei Liu; Asan; Jun Sun; Sergio Vano-Galvan; Feng-Xia Liu; Xiu-Xiu Wei; Dong-Lai Ma

    2016-01-01

    Background: The dyschromatoses are a group of disorders characterized by simultaneous hyperpigmented macules together with hypopigmented macules.Dyschromatosis universalis hereditaria (DUH) and dyschromatosis symmetrica hereditaria are two major types.While clinical and histological presentations are similar in these two diseases, genetic diagnosis is critical in the differential diagnosis of these entities.Methods: Three patients initially diagnosed with DUH were included.The gene test was carried out by targeted gene sequencing.All mutations detected on ADAR1 and ABCB6 genes were analyzed according to the frequency in control database, the mutation types, and the published evidence to determine the pathogenicity.Results: Family pedigree and clinical presentations were reported in 3 patients from two Chinese families.All patients have prominent cutaneous dyschromatoses involving the whole body without systemic complications.Different pathogenic genes in these patients with similar phenotype were identified: One novel mutation on ADAR1 (c.1325C>G) and one recurrent mutation in ABCB6 (c.1270T>C), which successfully distinguished two diseases with the similar phenotype.Conclusion: Targeted gene sequencing is an effective tool for genetic diagnosis in pigmentary skin diseases.

  3. Grainy head and its target genes in epithelial morphogenesis and wound healing.

    Science.gov (United States)

    Wang, Shenqiu; Samakovlis, Christos

    2012-01-01

    The Grainy head (Grh) family of transcription factors is characterized by a unique DNA-binding domain that binds to a conserved consensus sequence. Nematodes and flies have a single grh gene, whereas mice and humans have evolved three genes encoding Grainy head-like (Grhl) factors. We review the biological function of Grh in different animals and the mechanisms modulating its activity. grh and grhl genes play a remarkably conserved role in epithelial organ development and extracellular barrier repair after tissue damage. Recent studies in flies and vertebrates suggest that Grh factors may be primary determinants of cell adhesion and epithelial tissue formation. Grh proteins can dimerize and act as activators or repressors in different developmental contexts. In flies, tissue-specific, alternative splicing generates different Grh isoforms with different DNA-binding specificities and functions. Grh activity is also modulated by receptor tyrosine kinases: it is phosphorylated by extracellular signal regulated kinase, and this phosphorylation is selectively required for epidermal barrier repair. Two mechanisms have been proposed to explain the repressive function of Grh on target gene transcription. First, Grh can target the Polycomb silencing complex to specific response elements. Second, it can directly compete for DNA binding with transcriptional activators. Understanding the molecular mechanisms of gene regulation by Grh factors is likely to elucidate phylogenetically conserved mechanisms of epithelial cell morphogenesis and regeneration upon tissue damage.

  4. Microarray analyses of glucocorticoid and vitamin D3 target genes in differentiating cultured human podocytes.

    Directory of Open Access Journals (Sweden)

    Xiwen Cheng

    Full Text Available Glomerular podocytes are highly differentiated epithelial cells that are key components of the kidney filtration units. Podocyte damage or loss is the hallmark of nephritic diseases characterized by severe proteinuria. Recent studies implicate that hormones including glucocorticoids (ligand for glucocorticoid receptor and vitamin D3 (ligand for vitamin D receptor protect or promote repair of podocytes from injury. In order to elucidate the mechanisms underlying hormone-mediated podocyte-protecting activity from injury, we carried out microarray gene expression studies to identify the target genes and corresponding pathways in response to these hormones during podocyte differentiation. We used immortalized human cultured podocytes (HPCs as a model system and carried out in vitro differentiation assays followed by dexamethasone (Dex or vitamin D3 (VD3 treatment. Upon the induction of differentiation, multiple functional categories including cell cycle, organelle dynamics, mitochondrion, apoptosis and cytoskeleton organization were among the most significantly affected. Interestingly, while Dex and VD3 are capable of protecting podocytes from injury, they only share limited target genes and affected pathways. Compared to VD3 treatment, Dex had a broader and greater impact on gene expression profiles. In-depth analyses of Dex altered genes indicate that Dex crosstalks with a broad spectrum of signaling pathways, of which inflammatory responses, cell migration, angiogenesis, NF-κB and TGFβ pathways are predominantly altered. Together, our study provides new information and identifies several new avenues for future investigation of hormone signaling in podocytes.

  5. Transcriptional regulation of gene expression during osmotic stress responses by the mammalian target of rapamycin.

    Science.gov (United States)

    Ortells, M Carmen; Morancho, Beatriz; Drews-Elger, Katherine; Viollet, Benoit; Laderoute, Keith R; López-Rodríguez, Cristina; Aramburu, Jose

    2012-05-01

    Although stress can suppress growth and proliferation, cells can induce adaptive responses that allow them to maintain these functions under stress. While numerous studies have focused on the inhibitory effects of stress on cell growth, less is known on how growth-promoting pathways influence stress responses. We have approached this question by analyzing the effect of mammalian target of rapamycin (mTOR), a central growth controller, on the osmotic stress response. Our results showed that mammalian cells exposed to moderate hypertonicity maintained active mTOR, which was required to sustain their cell size and proliferative capacity. Moreover, mTOR regulated the induction of diverse osmostress response genes, including targets of the tonicity-responsive transcription factor NFAT5 as well as NFAT5-independent genes. Genes sensitive to mTOR-included regulators of stress responses, growth and proliferation. Among them, we identified REDD1 and REDD2, which had been previously characterized as mTOR inhibitors in other stress contexts. We observed that mTOR facilitated transcription-permissive conditions for several osmoresponsive genes by enhancing histone H4 acetylation and the recruitment of RNA polymerase II. Altogether, these results reveal a previously unappreciated role of mTOR in regulating transcriptional mechanisms that control gene expression during cellular stress responses.

  6. Targeted disruption of Ataxia-telangiectasia mutated gene in miniature pigs by somatic cell nuclear transfer.

    Science.gov (United States)

    Kim, Young June; Ahn, Kwang Sung; Kim, Minjeong; Kim, Min Ju; Park, Sang-Min; Ryu, Junghyun; Ahn, Jin Seop; Heo, Soon Young; Kang, Jee Hyun; Choi, You Jung; Choi, Seong-Jun; Shim, Hosup

    2014-10-03

    Ataxia telangiectasia (A-T) is a recessive autosomal disorder associated with pleiotropic phenotypes, including progressive cerebellar degeneration, gonad atrophy, and growth retardation. Even though A-T is known to be caused by the mutations in the Ataxia telangiectasia mutated (ATM) gene, the correlation between abnormal cellular physiology caused by ATM mutations and the multiple symptoms of A-T disease has not been clearly determined. None of the existing ATM mouse models properly reflects the extent to which neurological degeneration occurs in human. In an attempt to provide a large animal model for A-T, we produced gene-targeted pigs with mutations in the ATM gene by somatic cell nuclear transfer. The disrupted allele in the ATM gene of cloned piglets was confirmed via PCR and Southern blot analysis. The ATM gene-targeted pigs generated in the present study may provide an alternative to the current mouse model for the study of mechanisms underlying A-T disorder and for the development of new therapies.

  7. RNA sequencing analysis of human podocytes reveals glucocorticoid regulated gene networks targeting non-immune pathways

    Science.gov (United States)

    Jiang, Lulu; Hindmarch, Charles C. T.; Rogers, Mark; Campbell, Colin; Waterfall, Christy; Coghill, Jane; Mathieson, Peter W.; Welsh, Gavin I.

    2016-01-01

    Glucocorticoids are steroids that reduce inflammation and are used as immunosuppressive drugs for many diseases. They are also the mainstay for the treatment of minimal change nephropathy (MCN), which is characterised by an absence of inflammation. Their mechanisms of action remain elusive. Evidence suggests that immunomodulatory drugs can directly act on glomerular epithelial cells or ‘podocytes’, the cell type which is the main target of injury in MCN. To understand the nature of glucocorticoid effects on non-immune cell functions, we generated RNA sequencing data from human podocyte cell lines and identified the genes that are significantly regulated in dexamethasone-treated podocytes compared to vehicle-treated cells. The upregulated genes are of functional relevance to cytoskeleton-related processes, whereas the downregulated genes mostly encode pro-inflammatory cytokines and growth factors. We observed a tendency for dexamethasone-upregulated genes to be downregulated in MCN patients. Integrative analysis revealed gene networks composed of critical signaling pathways that are likely targeted by dexamethasone in podocytes. PMID:27774996

  8. CD133-targeted gene transfer into long-term repopulating hematopoietic stem cells.

    Science.gov (United States)

    Brendel, Christian; Goebel, Benjamin; Daniela, Abriss; Brugman, Martijn; Kneissl, Sabrina; Schwäble, Joachim; Kaufmann, Kerstin B; Müller-Kuller, Uta; Kunkel, Hana; Chen-Wichmann, Linping; Abel, Tobias; Serve, Hubert; Bystrykh, Leonid; Buchholz, Christian J; Grez, Manuel

    2015-01-01

    Gene therapy for hematological disorders relies on the genetic modification of CD34(+) cells, a heterogeneous cell population containing about 0.01% long-term repopulating cells. Here, we show that the lentiviral vector CD133-LV, which uses a surface marker on human primitive hematopoietic stem cells (HSCs) as entry receptor, transfers genes preferentially into cells with high engraftment capability. Transduction of unstimulated CD34(+) cells with CD133-LV resulted in gene marking of cells with competitive proliferative advantage in vitro and in immunodeficient mice. The CD133-LV-transduced population contained significantly more cells with repopulating capacity than cells transduced with vesicular stomatitis virus (VSV)-LV, a lentiviral vector pseudotyped with the vesicular stomatitis virus G protein. Upon transfer of a barcode library, CD133-LV-transduced cells sustained gene marking in vivo for a prolonged period of time with a 6.7-fold higher recovery of barcodes compared to transduced control cells. Moreover, CD133-LV-transduced cells were capable of repopulating secondary recipients. Lastly, we show that this targeting strategy can be used for transfer of a therapeutic gene into CD34(+) cells obtained from patients suffering of X-linked chronic granulomatous disease. In conclusion, direct gene transfer into CD133(+) cells allows for sustained long-term engraftment of gene corrected cells.

  9. In Vivo Anti-Cancer Mechanism of Low-Molecular-Weight Fucosylated Chondroitin Sulfate (LFCS from Sea Cucumber Cucumaria frondosa

    Directory of Open Access Journals (Sweden)

    Xiaoxiao Liu

    2016-05-01

    Full Text Available The low-molecular-weight fucosylated chondroitin sulfate (LFCS was prepared from native fucosylated chondroitin sulfate (FCS, which was extracted and isolated from sea cucumber Cucumaria frondosa, and the anti-cancer mechanism of LFCS on mouse Lewis lung carcinoma (LLC was investigated. The results showed that LFCS remarkably inhibited LLC growth and metastasis in a dose-dependent manner. LFCS induced cell cycle arrest by increasing p53/p21 expression and apoptosis through activation of caspase-3 activity in LLC cells. Meanwhile, LFCS suppressed the expression of vascular endothelial growth factor (VEGF, increased the expression of tissue inhibitor of metalloproteinase-1 (TIMP-1 and downregulated the matrix metalloproteinases (MMPs level. Furthermore, LFCS significantly suppressed the activation of ERK1/2/p38 MAPK/NF-κB pathway, which played a prime role in expression of MMPs. All of these data indicate LFCS may be used as anti-cancer drug candidates and deserve further study.

  10. Three-cohort targeted gene screening reveals a non-synonymous TRKA polymorphism associated with schizophrenia

    DEFF Research Database (Denmark)

    van Schijndel, Jessica E; van Loo, Karen M J; van Zweeden, Martine;

    2009-01-01

    Schizophrenia is a complex neurodevelopmental disorder that is thought to be induced by an interaction between predisposing genes and environmental stressors. To identify predisposing genetic factors, we performed a targeted (mostly neurodevelopmental) gene approach involving the screening of 396...... selected non-synonymous single-nucleotide polymorphisms (SNPs) in three independent Caucasian schizophrenia case-control cohorts (USA, Denmark and Norway). A meta-analysis revealed ten non-synonymous SNPs that were nominally associated with schizophrenia, nine of which have not been previously linked...... for schizophrenia....

  11. A comparison of Agrobacterium-mediated transformation and protoplast-mediated transformation with CRISPR-Cas9 and bipartite gene targeting substrates, as effective gene targeting tools for Aspergillus carbonarius.

    Science.gov (United States)

    Weyda, István; Yang, Lei; Vang, Jesper; Ahring, Birgitte K; Lübeck, Mette; Lübeck, Peter S

    2017-04-01

    In recent years, versatile genetic tools have been developed and applied to a number of filamentous fungi of industrial importance. However, the existing techniques have limitations when it comes to achieve the desired genetic modifications, especially for efficient gene targeting. In this study, we used Aspergillus carbonarius as a host strain due to its potential as a cell factory, and compared three gene targeting techniques by disrupting the ayg1 gene involved in the biosynthesis of conidial pigment in A. carbonarius. The absence of the ayg1 gene leads to phenotypic change in conidia color, which facilitated the analysis on the gene targeting frequency. The examined transformation techniques included Agrobacterium-mediated transformation (AMT) and protoplast-mediated transformation (PMT). Furthermore, the PMT for the disruption of the ayg1 gene was carried out with bipartite gene targeting fragments and the recently adapted CRISPR-Cas9 system. All three techniques were successful in generating Δayg1 mutants, but showed different efficiencies. The most efficient method for gene targeting was AMT, but further it was shown to be dependent on the choice of Agrobacterium strain. However, there are different advantages and disadvantages of all three gene targeting methods which are discussed, in order to facilitate future approaches for fungal strain improvements.

  12. Screening the yeast genome for energetic metabolism pathways involved in a phenotypic response to the anti-cancer agent 3-bromopyruvate.

    Science.gov (United States)

    Lis, Paweł; Jurkiewicz, Paweł; Cal-Bąkowska, Magdalena; Ko, Young H; Pedersen, Peter L; Goffeau, Andre; Ułaszewski, Stanisław

    2016-03-01

    In this study the detailed characteristic of the anti-cancer agent 3-bromopyruvate (3-BP) activity in the yeast Saccharomyces cerevisiae model is described, with the emphasis on its influence on energetic metabolism of the cell. It shows that 3-BP toxicity in yeast is strain-dependent and influenced by the glucose-repression system. Its toxic effect is mainly due to the rapid depletion of intracellular ATP. Moreover, lack of the Whi2p phosphatase results in strongly increased sensitivity of yeast cells to 3-BP, possibly due to the non-functional system of mitophagy of damaged mitochondria through the Ras-cAMP-PKA pathway. Single deletions of genes encoding glycolytic enzymes, the TCA cycle enzymes and mitochondrial carriers result in multiple effects after 3-BP treatment. However, it can be concluded that activity of the pentose phosphate pathway is necessary to prevent the toxicity of 3-BP, probably due to the fact that large amounts of NADPH are produced by this pathway, ensuring the reducing force needed for glutathione reduction, crucial to cope with the oxidative stress. Moreover, single deletions of genes encoding the TCA cycle enzymes and mitochondrial carriers generally cause sensitivity to 3-BP, while totally inactive mitochondrial respiration in the rho0 mutant resulted in increased resistance to 3-BP.

  13. Anti-Cancer Effect of Metabotropic Glutamate Receptor 1 Inhibition in Human Glioma U87 Cells: Involvement of PI3K/Akt/mTOR Pathway

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    Chi Zhang

    2015-01-01

    Full Text Available Background: Metabotropic glutamate receptors (mGluRs are G-protein-coupled receptors that mediate neuronal excitability and synaptic plasticity in the central nervous system, and emerging evidence suggests a role of mGluRs in the biology of cancer. Previous studies showed that mGluR1 was a potential therapeutic target for the treatment of breast cancer and melanoma, but its role in human glioma has not been determined. Methods: In the present study, we investigated the effects of mGluR1 inhibition in human glioma U87 cells using specific targeted small interfering RNA (siRNA or selective antagonists Riluzole and BAY36-7620. The anti-cancer effects of mGluR1 inhibition were measured by cell viability, lactate dehydrogenase (LDH release, TUNEL staining, cell cycle assay, cell invasion and migration assays in vitro, and also examined in a U87 xenograft model in vivo. Results: Inhibition of mGluR1 significantly decreased the cell viability but increased the LDH release in a dose-dependent fashion in U87 cells. These effects were accompanied with the induction of caspase-dependent apoptosis and G0/G1 cell cycle arrest. In addition, the results of Matrigel invasion and cell tracking assays showed that inhibition of mGluR1 apparently attenuated cell invasion and migration in U87 cells. All these anti-cancer effects were ablated by the mGluR1 agonist L-quisqualic acid. The results of western blot analysis showed that mGluR1 inhibition overtly decreased the phosphorylation of PI3K, Akt, mTOR and P70S6K, indicating the mitigated activation of PI3K/Akt/mTOR pathway. Moreover, the anti-tumor activity of mGluR1 inhibition in vivo was also demonstrated in a U87 xenograft glioma model in athymic nude mice. Conclusion: The remarkable efficiency of mGluR1 inhibition to induce cell death in U87 cells may find therapeutic application for the treatment of glioma patients.

  14. Anti-cancer activity of novel dibenzo[b,f]azepine tethered isoxazoline derivatives

    Directory of Open Access Journals (Sweden)

    Sadashiva Maralinganadoddi

    2012-10-01

    Full Text Available Background Dibenzoazepine (DB derivatives are important and valuable compounds in medicinal chemistry. The synthesis and chemotherapeutic properties of naturally occurring DBs and different heterocyclic moiety tethered DBs are reported. Herein, we report the DB-fused hybrid structure that containing isoxazolines (DBIs and their anti-cancer activity, which could throw light on the structural and functional features of new molecules. Results and Conclusion The synthesis and characterization of novel ring DB tethered isoxazoline derivatives (DBIs were carried out. After the detailed structural characterization using 2D-NMR experiments, the compounds were identified as 5-substituted isoxazolines. The effect of newly synthesized DBIs against the invasion of murine osteosarcoma (LM8G7 cells was studied. Among the tested molecules, compound 4g (5-[−3-(4-chlorophenyl-4,5-dihydroisoxazol-5-yl-methyl]-5 H-dibenzo[b,f]azepine, was found to inhibit the invasion of LM8G7 cells strongly, when compared to other structurally related compounds. Cumulatively, the compound 4g inhibited the invasion MDA-MB-231 cells completely at 10 μM. In addition to anti-invasion property the compound 4g also inhibited the migration of LM8G7 and human ovarian cancer cells (OVSAHO dose-dependently. Compound 4g inhibited the proliferation of LM8G7, OVSAHO, human breast cancer cells (MCF-7 and human melphalan-resistant multiple myeloma (RPMI8226-LR5 cells that are comparable to cisplatin and suramin.

  15. Cardio-protective and anti-cancer therapeutic potential of Nigella sativa.

    Science.gov (United States)

    Shafiq, Hammad; Ahmad, Asif; Masud, Tariq; Kaleem, Muhammad

    2014-12-01

    Nigella sativa is the miraculous plant having a lot of nutritional and medicinal benefits, and attracts large number of nutrition and pharmacological researchers. N. sativa seed composition shows that it is the blessing of nature and it contains and many bioactive compounds like thymoquinone, α-hederin, alkaloids, flavonoids, antioxidants, fatty acids many other compounds that have positive effects on curing of different diseases. Several medicinal properties of N. sativa like its anti-cancer, anti-inflammatory, anti-diabetic, antioxidant activities and many others are well acknowledged. However, this article focuses on activity of N. sativa against cardiovascular diseases and cancer. For gathering required data the authors went through vast number of articles using search engines like Science direct, ELSEVIER, Pub Med, Willey on Line Library and Google scholar and the findings were classified on the basis of relevance of the topic and were reviewed in the article. N. sativa is rich source of different biologically active compounds and is found effective in controlling number of cardiovascular diseases and various cancers both in vivo and in vitro studies.

  16. Phytochemical characterization, anti-cancer and antimicrobial activity of isolated fractions of Alysicarpus vaginalis

    Directory of Open Access Journals (Sweden)

    Ganesh Tapadiya

    2017-03-01

    Full Text Available The methanolic extract of Alysicarpus vaginalis was selected for fractionation due to its known reported biological activity. The four fractions were separated and subjected for in vitro antimitotic and anti-proliferative assays along with anti-cancer activity on two human cancers cell lines (SK-MEL-2 and Hep-G2. The antimicrobial potential of fractions had been evaluated against bacteria and fungi. From all fractions, acetone and n-butanol fractions were effective against the cell lines. They show strong inhibitory action with mitotic index 6.2 and 8.4 mg/mL and IC50 values of anti-proliferative assay in between 19.7 to 14.2 mg/mL respectively, which was found to be comparable to the standard methothrexate 5.9 mg/mL and 13.2 mg/mL respectively. In antimicrobial activity, the zone of inhibition had been observed in the range of 12-27 mm and MIC value was found in the range of 0.2-0.1 mg/mL. The acetone fraction was found to be most active against fungi, and E. coli whereas chloroform and n-butanol fractions were more effective against S. aureus and B. subtilis. The phytochemical characterization by HPLC analysis indicated the presence of important polyphenolic and steroidal compounds.

  17. Photochemical properties of a new kind of anti-cancer drug: N-glycoside compound

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Due to the nontoxicity and efficient anti-cancer activity, more and more attention has been paid to N-glycoside compounds. Laser photolysis of N-(α-D-glucopyranoside) salicyloyl hydrazine (NGSH) has been performed for the first time. The research results show that NGSH has high photosensitivity and is vulnerable to be photo-ionized via a monophotonic process with a quantum yield of 0.02, generating NGSH+· and hydrated electrons. Under the aerobic condition of cells, the hydrated electrons are very easy to combine with oxygen to generate 1O2 and O2-, both of which are powerful oxidants that can kill the cancer cells. In addition, NGSH+· can be changed into neutral radicals by deprotonation with a pKa value of 4.02 and its decay constant was determined to be 2.55×109dm3·mol-1·s-1. NGSH also can be oxidized by SO4-. with a rate constant of 1.76×109 dm3·mol-1.s-1, which further confirms the results of photoionization. All of these results suggest that this new N-glycoside compound might be useful for cancer treatment.

  18. Hydroxypropyl-β-cyclodextrin–graphene oxide conjugates: Carriers for anti-cancer drugs

    Energy Technology Data Exchange (ETDEWEB)

    Tan, Jingting; Meng, Na; Fan, Yunting; Su, Yutian; Zhang, Ming [Jiangsu Collaborative Innovation Center for Biological Functional Materials, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210023 (China); National and Local Joint Engineering Research Center of Biomedical Functional Materials, Nanjing 210023 (China); Jiangsu Key Laboratory of Biofunctional Materials, Jiangsu Engineering Research Center for Biomedical Function Materials, Nanjing 210023 (China); Xiao, Yinghong, E-mail: yhxiao@njnu.edu.cn [Jiangsu Collaborative Innovation Center for Biological Functional Materials, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210023 (China); National and Local Joint Engineering Research Center of Biomedical Functional Materials, Nanjing 210023 (China); Jiangsu Key Laboratory of Biofunctional Materials, Jiangsu Engineering Research Center for Biomedical Function Materials, Nanjing 210023 (China); Zhou, Ninglin, E-mail: zhouninglin@njnu.edu.cn [Jiangsu Collaborative Innovation Center for Biological Functional Materials, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210023 (China); National and Local Joint Engineering Research Center of Biomedical Functional Materials, Nanjing 210023 (China); Jiangsu Key Laboratory of Biofunctional Materials, Jiangsu Engineering Research Center for Biomedical Function Materials, Nanjing 210023 (China); Nanjing Zhou Ninglin Advanced Materials Technology Company Limited, Nanjing 211505 (China)

    2016-04-01

    A novel drug carrier based on hydroxypropyl-β-cyclodextrin (HP-β-CD) modified carboxylated graphene oxide (GO-COOH) was designed to incorporate anti-cancer drug paclitaxel (PTX). The formulated nanomedicines were characterized by Fourier transform infrared spectroscopy (FTIR) and atomic force microscopy (AFM). Results showed that PTX can be incorporated into GO-COO-HP-β-CD nanospheres successfully, with an average diameter of about 100 nm. The solubility and stability of PTX-loaded GO-COO-HP-β-CD nanospheres in aqueous media were greatly enhanced compared with the untreated PTX. The results of hemolysis test demonstrated that the drug-loaded nanospheres were qualified with good blood compatibility for intravenous use. In vitro anti-tumor activity was measured and results demonstrated that the incorporation of PTX into the newly developed GO-COO-HP-β-CD carrier could confer significantly improved cytotoxicity to the nanosystem against tumor cells than single application of PTX. GO-COO-HP-β-CD nanospheres may represent a promising formulation platform for a broad range of therapeutic agent, especially those with poor solubility. - Highlights: • Hydroxypropyl-β-cyclodextrin (HP-β-CD) modified carboxylated graphene oxide (GO-COOH) was designed as a drug carrier. • The prepared PTX-loaded nanospheres can be dispersed in aqueous medium stably. • The GO-COO-HP-β-CD nanospheres are safe for blood-contact applications. • This newly developed PTX-delivery system could confer significantly improved cytotoxicity against tumor cells.

  19. Mesua beccariana (Clusiaceae, A Source of Potential Anti-cancer Lead Compounds in Drug Discovery

    Directory of Open Access Journals (Sweden)

    Soek Sin Teh

    2012-09-01

    Full Text Available An investigation on biologically active secondary metabolites from the stem bark of Mesua beccariana was carried out. A new cyclodione, mesuadione (1, along with several known constituents which are beccamarin (2, 2,5-dihydroxy-1,3,4-trimethoxy anthraquinone (3, 4-methoxy-1,3,5-trihydroxyanthraquinone (4, betulinic acid (5 and stigmasterol (6 were obtained from this ongoing research. Structures of these compounds were elucidated by extensive spectroscopic methods, including 1D and 2D-NMR, GC-MS, IR and UV techniques. Preliminary tests of the in vitro cytotoxic activities of all the isolated metabolites against a panel of human cancer cell lines Raji (lymphoma, SNU-1 (gastric carcinoma, K562 (erythroleukemia cells, LS-174T (colorectal adenocarcinoma, HeLa (cervical cells, SK-MEL-28 (malignant melanoma cells, NCI-H23 (lung adenocarcinoma, IMR-32 (neuroblastoma and Hep-G2 (hepatocellular liver carcinoma were carried out using an MTT assay. Mesuadione (1, beccamarin (2, betulinic acid (5 and stigmasterol (6 displayed strong inhibition of Raji cell proliferation, while the proliferation rate of SK-MEL-28 and HeLa were strongly inhibited by stigmasterol (6 and beccamarin (2, indicating these secondary metabolites could be anti-cancer lead compounds in drug discovery.

  20. Diterpenes from rosemary (Rosmarinus officinalis): Defining their potential for anti-cancer activity.

    Science.gov (United States)

    Petiwala, Sakina M; Johnson, Jeremy J

    2015-10-28

    Recently, rosemary extracts standardized to diterpenes (e.g. carnosic acid and carnosol) have been approved by the European Union (EU) and given a GRAS (Generally Recognized as Safe) status in the United States by the Food and Drug Administration (FDA). Incorporation of rosemary into our food system and through dietary selection (e.g. Mediterranean Diet) has increased the likelihood of exposure to diterpenes in rosemary. In consideration of this, a more thorough understanding of rosemary diterpenes is needed to understand its potential for a positive impact on human health. Three agents in particular have received the most attention that includes carnosic acid, carnosol, and rosmanol with promising results of anti-cancer activity. These studies have provided evidence of diterpenes to modulate deregulated signaling pathways in different solid and blood cancers. Rosemary extracts and the phytochemicals therein appear to be well tolerated in different animal models as evidenced by the extensive studies performed for approval by the EU and the FDA as an antioxidant food preservative. This mini-review reports on the pre-clinical studies performed with carnosic acid, carnosol, and rosmanol describing their mechanism of action in different cancers.

  1. Anti-cancer evaluation of quercetin embedded PLA nanoparticles synthesized by emulsified nanoprecipitation.

    Science.gov (United States)

    Pandey, Sanjeev K; Patel, Dinesh K; Thakur, Ravi; Mishra, Durga P; Maiti, Pralay; Haldar, Chandana

    2015-04-01

    This study was carried out to synthesize quercetin (Qt) embedded poly(lactic acid) (PLA) nanoparticles (PLA-Qt) and to evaluate anti-cancer efficacy of PLA-Qt by using human breast cancer cells. PLA-Qt were synthesized by using novel emulsified nanoprecipitation technique with varying dimension of 32 ± 8 to 152 ± 9 nm of PLA-Qt with 62 ± 3% (w/w) entrapment efficiency by varying the concentration of polymer, emulsifier, drug and preparation temperature. The dimension of PLA-Qt was measured through transmission electron microscopy indicating larger particle size at higher concentration of PLA. The release rate of Qt from PLA-Qt was found to be more sustained for larger particle dimension (152 ± 9 nm) as compared to smaller particle dimension (32 ± 8 nm). Interaction between Qt and PLA was verified through spectroscopic and calorimetric methods. Delayed diffusion and stronger interaction in PLA-Qt caused the sustained delivery of Qt from the polymer matrix. In vitro cytotoxicity study indicate the killing of ∼ 50% breast cancer cells in two days at 100 μg/ml of drug concentration while the ∼ 40% destruction of cells require 5 days for PLA-Qt (46 ± 6 nm; 20mg/ml of PLA). Thus our results propose anticancer efficacy of PLA-Qt nanoparticles in terms of its sustained release kinetics revealing novel vehicle for the treatment of cancer.

  2. Anti-Cancer Effects of Protein Extracts from Calvatia lilacina, Pleurotus ostreatus and Volvariella volvacea

    Directory of Open Access Journals (Sweden)

    Jin-Yi Wu

    2011-01-01

    Full Text Available Calvatia lilacina (CL, Pleurotus ostreatus (PO and Volvariella volvacea (VV are widely distributed worldwide and commonly eaten as mushrooms. In this study, cell viabilities were evaluated for a human colorectal adenocarcinoma cell line (SW480 cells and a human monocytic leukemia cell line (THP-1 cells. Apoptotic mechanisms induced by the protein extracts of PO and VV were evaluated for SW480 cells. The viabilities of THP-1 and SW480 cells decreased in a concentration-dependent manner after 24 h of treatment with the protein extracts of CL, PO or VV. Apoptosis analysis revealed that the percentage of SW480 cells in the SubG1 phase (a marker of apoptosis was increased upon PO and VV protein-extract treatments, indicating that oligonucleosomal DNA fragmentation existed concomitantly with cellular death. The PO and VV protein extracts induced reactive oxygen species (ROS production, glutathione (GSH depletion and mitochondrial transmembrane potential (ΔΨm loss in SW480 cells. Pretreatment with N-acetylcysteine, GSH or cyclosporine A partially prevented the apoptosis induced by PO protein extracts, but not that induced by VV extracts, in SW480 cells. The protein extracts of CL, PO and VV exhibited therapeutic efficacy against human colorectal adenocarcinoma cells and human monocytic leukemia cells. The PO protein extracts induced apoptosis in SW480 cells partially through ROS production, GSH depletion and mitochondrial dysfunction. Therefore, the protein extracts of these mushrooms could be considered an important source of new anti-cancer drugs.

  3. Cardio-protective and anti-cancer therapeutic potential of Nigella sativa

    Directory of Open Access Journals (Sweden)

    Hammad Shafiq

    2015-12-01

    Full Text Available Nigella sativa is the miraculous plant having a lot of nutritional and medicinal benefits, and attracts large number of nutrition and pharmacological researchers. N. sativa seed composition shows that it is the blessing of nature and it contains and many bioactive compounds like thymoquinone, α-hederin, alkaloids, flavonoids, antioxidants, fatty acids many other compounds that have positive effects on curing of different diseases. Several medicinal properties of N. sativa like its anti-cancer, anti-inflammatory, anti-diabetic, antioxidant activities and many others are well acknowledged. However, this article focuses on activity of N. sativa against cardiovascular diseases and cancer. For gathering required data the authors went through vast number of articles using search engines like Science direct, ELSEVIER, Pub Med, Willey on Line Library and Google scholar and the findings were classified on the basis of relevance of the topic and were reviewed in the article. N. sativa is rich source of different biologically active compounds and is found effective in controlling number of cardiovascular diseases and various cancers both in vivo and in vitro studies.

  4. Synthesis, Characterization and Anti-Cancer Activity of Hydrazide Derivatives Incorporating a Quinoline Moiety

    Directory of Open Access Journals (Sweden)

    Murat Bingul

    2016-07-01

    Full Text Available Identification of the novel (E-N′-((2-chloro-7-methoxyquinolin-3-ylmethylene-3-(phenylthiopropanehydrazide scaffold 18 has led to the development of a new series of biologically active hydrazide compounds. The parent compound 18 and new quinoline derivatives 19–26 were prepared from the corresponding quinoline hydrazones and substituted carboxylic acids using EDC-mediated peptide coupling reactions. Further modification of the parent compound 18 was achieved by replacement of the quinoline moiety with other aromatic systems. All the newly synthesized compounds were evaluated for their anti-cancer activity against the SH-SY5Y and Kelly neuroblastoma cell lines, as well as the MDA-MB-231 and MCF-7 breast adenocarcinoma cell lines. Analogues 19 and 22 significantly reduced the cell viability of neuroblastoma cancer cells with micromolar potency and significant selectivity over normal cells. The quinoline hydrazide 22 also induced G1 cell cycle arrest, as well as upregulation of the p27kip1 cell cycle regulating protein.

  5. Structural characterization of a broccoli polysaccharide and evaluation of anti-cancer cell proliferation effects.

    Science.gov (United States)

    Xu, Lishan; Cao, Jingjing; Chen, Wenrong

    2015-08-01

    Broccoli is a widely consumed vegetable with abundant amount of nutrients, which bring numerous beneficial effects on human health. The structural information of water-soluble polysaccharides in broccoli was eludicated for the first time in this work. A purified polysaccharide fraction (BPCa) was obtained by column chromatography. It comprised of arabinose (Ara), galactose (Gal), rhamnose (Rha) with a molar ratio of 5.3:0.8:1.0. Nuclear magnetic resonnance spectra data revealed that α-L-1,5-Araf and α-L-1,3,5-Araf are present in the backbone, while α-L-Araf terminal was attended in side chain. α-L-1,2-Rhap was found to be linked to α-L-1,5-Araf in heteronuclear multiple bond correlation spectra. The presences of β-D-1,4-Galp and α-D-1,4-GalpA were also detected. Furthermore, BPCa showed significant anti-cancer cell proliferation activities against HepG2, Siha and MDA-MB-231 carcinoma cell lines. The results indicated that BPCa had a good potential to be applied as functional food additives.

  6. Pectenotoxin-2 from Marine Sponges: A Potential Anti-Cancer Agent—A Review

    Directory of Open Access Journals (Sweden)

    Wun-Jae Kim

    2011-11-01

    Full Text Available Pectenotoxin-2 (PTX-2, which was first identified as a cytotoxic entity in marine sponges, has been reported to display significant cytotoxicity to human cancer cells where it inhibits mitotic separation and cytokinesis through the depolymerization of actin filaments. In the late stage of endoreduplication, the effects of PTX-2 on different cancer cells involves: (i down-regulation of anti-apoptotic Bcl-2 members and IAP family proteins; (ii up-regulation of pro-apoptotic Bax protein and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL-receptor 1/receptor 2 (DR4/DR5; and (iii mitochondrial dysfunction. In addition, PTX-2 induces apoptotic effects through suppression of the nuclear factor κB (NF-κB signaling pathway in several cancer cells. Analysis of cell cycle regulatory proteins showed that PTX-2 increases phosphorylation of Cdc25c and decreases protein levels of Cdc2 and cyclin B1. Cyclin-dependent kinase (Cdk inhibitor p21 and Cdk2, which are associated with the induction of endoreduplication, were upregulated. Furthermore, it was found that PTX-2 suppressed telomerase activity through the transcriptional and post-translational suppression of hTERT. The purpose of this review was to provide an update regarding the anti-cancer mechanism of PTX-2, with a special focus on its effects on different cellular signaling cascades.

  7. Maturation of dendritic cells by pullulan promotes anti-cancer effect

    Science.gov (United States)

    Xu, Li; Zhang, LiJun; Yu, Qing; Jin, Jun-O

    2016-01-01

    Previous studies have demonstrated that pullulan, a polysaccharide purified from Aureobasidium pullulans, has immune-stimulatory effects on T and B cells. Moreover, pullulan has been used as a carrier in the delivery of the antigen (Ag) peptide to lymphoid tissues. However, the in vivo effect of pullulan on dendritic cells (DC) has not been well characterized. In this study, we assessed the effect of pullulan on DC activation and anti-cancer immunity. The results showed that the pullulan treatment up-regulated co-stimulatory molecule expression and enhanced pro-inflammatory cytokine production in bone marrow-derived DCs (BMDC) in vitro and in spleen DCs in vivo. Moreover, the combination of ovalbumin (OVA) and pullulan induced OVA antigen-specific T cell activations in vivo. In tumor-bearing mice, pullulan induced the maturation of DCs in spleen and tumor draining lymph node (drLN), and promoted the OVA-specific T cell activation and migration of the T cells into the tumor. In addition, the combination of OVA and pullulan inhibited B16-OVA tumor growth and liver metastasis. The combination of tyrosinase-related protein 2 (TRP2) peptide and pullulan treatment also suppressed B16 melanoma growth. Thus, the results demonstrated that pullulan enhanced DC maturation and function, and it acted as an adjuvant in promoting Ag-specific immune responses in mice. Thus, pullulan could be a new and useful adjuvant for use in therapeutic cancer vaccines. PMID:27341129

  8. Anti-cancer effect of Cordyceps militaris in human colorectal carcinoma RKO cells via cell cycle arrest and mitochondrial apoptosis

    OpenAIRE

    Lee, Hwan Hee; Lee, Seulki; Lee, Kanghyo; Shin, Yu Su; Kang, Hyojeung; Cho, Hyosun

    2015-01-01

    Background Cordyceps militaris has been used as a traditional medicine in Asian countries for a long time. Different types of Cordyceps extract were reported to have various pharmacological activities including an anti-cancer effect. We investigated the inhibitory effect of Cordyceps militaris ethanol extract on a human colorectal cancer-derived cell line, RKO. Methods RKO cells were treated with various concentrations of nucleosides-enriched ethanol extract of Cordyceps militaris for 48 h an...

  9. The anti-cancer drug-induced pica in rats is related to their clinical emetogenic potential.

    Science.gov (United States)

    Yamamoto, Kouichi; Nakai, Miho; Nohara, Kyoko; Yamatodani, Atsushi

    2007-01-05

    Cancer chemotherapy is frequently accompanied by severe emesis. The anti-cancer drugs are classified according to their clinical emetogenic potential. We have already found that kaolin ingestion behavior "pica" is analogous to emesis in rats. The aim of this study was to examine the effects of the clinical emetogenic potential of anti-cancer drugs on the induction of the pica in rats. Rats were housed in individual cages with free access to food and kaolin pellets and the daily food and kaolin intakes were measured for 3 days after the intraperitoneal administration of anti-cancer drugs (cisplatin, cyclophosphamide, actinomycin D, 5-fluorouracil and vincristine). The drugs with high potential for inducing emesis, such as cisplatin and cyclophosphamide, induced pica in all animals on the day of administration and the behavior lasted during the observation period. The drugs with moderate emetogenic potential, i.e. actinomycin D and 5-fluorouracil, also induced pica on the first and second day after the drug administration but the kaolin intake was less than that of the drugs with high potential. Vincristine, a drug with low emetogenic potential, slightly increased the kaolin intake in rats on the only first day of the administration. Cyclophosphamide, actinomycin D and vincristine induced anorexia and decreased their body weight during the observation period. These results suggested that the both amounts of kaolin intake and duration of behavior in the anti-cancer drug-induced pica are related to the clinical emetogenic potential of the drugs and the incidence of the anorexia is not related to their emetogenic potential.

  10. Analysis of bacterial xylose isomerase gene diversity using gene-targeted metagenomics.

    Science.gov (United States)

    Nurdiani, Dini; Ito, Michihiro; Maruyama, Toru; Terahara, Takeshi; Mori, Tetsushi; Ugawa, Shin; Takeyama, Haruko

    2015-08-01

    Bacterial xylose isomerases (XI) are promising resources for efficient biofuel production from xylose in lignocellulosic biomass. Here, we investigated xylose isomerase gene (xylA) diversity in three soil metagenomes differing in plant vegetation and geographical location, using an amplicon pyrosequencing approach and two newly-designed primer sets. A total of 158,555 reads from three metagenomic DNA replicates for each soil sample were classified into 1127 phylotypes, detected in triplicate and defined by 90% amino acid identity. The phylotype coverage was estimated to be within the range of 84.0-92.7%. The xylA gene phylotypes obtained were phylogenetically distributed across the two known xylA groups. They shared 49-100% identities with their closest-related XI sequences in GenBank. Phylotypes demonstrating analysis, suggesting soil-specific xylA genotypes and taxonomic compositions. The differences among xylA members and their compositions in the soil were strongly correlated with 16S rRNA variation between soil samples, also assessed by amplicon pyrosequencing. This is the first report of xylA diversity in environmental samples assessed by amplicon pyrosequencing. Our data provide information regarding xylA diversity in nature, and can be a basis for the screening of novel xylA genotypes for practical applications.

  11. Inhibition of human esophageal squamous cell carcinomas by targeted silencing of tumor enhancer genes: an overview.

    Science.gov (United States)

    Islamian, Jalil Pirayesh; Mohammadi, Mohsen; Baradaran, Behzad

    2014-06-01

    Esophageal cancer has been reported as the ninth most common malignancy and ranks as the sixth most frequent cause of death worldwide. Esophageal cancer treatment involves surgery, chemotherapy, radiation therapy, or combination therapy. Novel strategies are needed to boost the oncologic outcome. Recent advances in the molecular biology of esophageal cancer have documented the role of genetic alterations in tumorigenesis. Oncogenes serve a pivotal function in tumorigenesis. Targeted therapies are directed at the unique molecular signature of cancer cells for enhanced efficacy with low toxicity. RNA interference (RNAi) technology is a powerful tool for silencing endogenous or exogenous genes in mammalian cells. Related results have shown that targeting oncogenes with siRNAs, specifically the mRNA, effectively reduces tumor cell proliferation and induces apoptotic cell death. This article will briefly review studies on silencing tumor enhancer genes related to the induction of esophageal cancer.

  12. Targeted Correction and Restored Function of the CFTR Gene in Cystic Fibrosis Induced Pluripotent Stem Cells

    Directory of Open Access Journals (Sweden)

    Ana M. Crane

    2015-04-01

    Full Text Available Recently developed reprogramming and genome editing technologies make possible the derivation of corrected patient-specific pluripotent stem cell sources—potentially useful for the development of new therapeutic approaches. Starting with skin fibroblasts from patients diagnosed with cystic fibrosis, we derived and characterized induced pluripotent stem cell (iPSC lines. We then utilized zinc-finger nucleases (ZFNs, designed to target the endogenous CFTR gene, to mediate correction of the inherited genetic mutation in these patient-derived lines via homology-directed repair (HDR. We observed an exquisitely sensitive, homology-dependent preference for targeting one CFTR allele versus the other. The corrected cystic fibrosis iPSCs, when induced to differentiate in vitro, expressed the corrected CFTR gene; importantly, CFTR correction resulted in restored expression of the mature CFTR glycoprotein and restoration of CFTR chloride channel function in iPSC-derived epithelial cells.

  13. Protein interactions of the MLL PHD fingers modulate MLL target gene regulation in human cells.

    Science.gov (United States)

    Fair, K; Anderson, M; Bulanova, E; Mi, H; Tropschug, M; Diaz, M O

    2001-05-01

    The PHD fingers of the human MLL and Drosophila trx proteins have strong amino acid sequence conservation but their function is unknown. We have determined that these fingers mediate homodimerization and binding of MLL to Cyp33, a nuclear cyclophilin. These two proteins interact in vitro and in vivo in mammalian cells and colocalize at specific nuclear subdomains. Overexpression of the Cyp33 protein in leukemia cells results in altered expression of HOX genes that are targets for regulation by MLL. These alterations are suppressed by cyclosporine and are not observed in cell lines that express a mutant MLL protein without PHD fingers. These results suggest that binding of Cyp33 to MLL modulates its effects on the expression of target genes.

  14. Glycoengineering of Human Cell Lines Using Zinc Finger Nuclease Gene Targeting

    DEFF Research Database (Denmark)

    Steentoft, Catharina; Bennett, Eric Paul; Clausen, Henrik

    2013-01-01

    Lectin affinity chromatography is a powerful technique for isolation of glycoproteins carrying a specific glycan structure of interest. However, the enormous diversity of glycans present on the cell surface, as well as on individual proteins, makes it difficult to isolate an entire glycoproteome...... with one or even a series of lectins. Here we present a technique to generate cell lines with homogenous truncated O-glycans using zinc finger nuclease gene targeting. Because of their simplified O-glycoproteome, the cells have been named SimpleCells. Glycoproteins from SimpleCells can be isolated...... in a single purification step by lectin chromatography performed on a long lectin column. This protocol describes Zinc finger nuclease gene targeting of human cells to simplify the glycoproteome, as well as lectin chromatography and isolation of glycopeptides from total cell lysates of SimpleCells....

  15. Comparing zinc finger nucleases and transcription activator-like effector nucleases for gene targeting in Drosophila.

    Science.gov (United States)

    Beumer, Kelly J; Trautman, Jonathan K; Christian, Michelle; Dahlem, Timothy J; Lake, Cathleen M; Hawley, R Scott; Grunwald, David J; Voytas, Daniel F; Carroll, Dana

    2013-10-03

    Zinc-finger nucleases have proven to be successful as reagents for targeted genome manipulation in Drosophila melanogaster and many other organisms. Their utility has been limited, however, by the significant failure rate of new designs, reflecting the complexity of DNA recognition by zinc fingers. Transcription activator-like effector (TALE) DNA-binding domains depend on a simple, one-module-to-one-base-pair recognition code, and they have been very productively incorporated into nucleases (TALENs) for genome engineering. In this report we describe the design of TALENs for a number of different genes in Drosophila, and we explore several parameters of TALEN design. The rate of success with TALENs was substantially greater than for zinc-finger nucleases , and the frequency of mutagenesis was comparable. Knockout mutations were isolated in several genes in which such alleles were not previously available. TALENs are an effective tool for targeted genome manipulation in Drosophila.

  16. Potential clinical insights into microRNAs and their target genes in esophageal carcinoma.

    Science.gov (United States)

    Li, Su Q; Wang, He M; Cao, Xiu F

    2011-12-01

    Esophageal carcinoma (EC) are characterized by dysregulation of microRNAs, which play an important roles as a posttranscriptional regulators in protein synthesis, and are involved in cellular processes, such as proliferation, apoptosis, and differentiation. Recently, altered miRNAs expression has been comprehensively studied in EC by high-throughput technology. Increased understanding of miRNAs target genes and their potential regulatory mechanisms have clarified the miRNAs activities and may provide exciting opportunities for cancer diagnosis and miRNA-based genetherapy. Here, we reviewed the most recently discovered miRNA target genes, with particular emphasis on the deciphering of their possible mechanisms and the potential roles in miRNAs-based tumour therapeutics.

  17. Furanodiene enhances the anti-cancer effects of doxorubicin on ERα-negative breast cancer cells in vitro.

    Science.gov (United States)

    Zhong, Zhang-Feng; Qiang, Wen-An; Wang, Chun-Ming; Tan, Wen; Wang, Yi-Tao

    2016-03-05

    Furanodiene is a natural product isolated from Rhizoma curcumae, and exhibits broad-spectrum anti-cancer activities in vitro and in vivo. Our previous study proved that furanodiene could increase growth inhibition of steroidal agent in ERα-positive breast cancer cells, but whether furanodiene can influence ER status is not clear. In this study, we confirmed that furanodiene down-regulated the ERα protein expression level and inhibited E2-induced estrogen response element (ERE)-driven reporter plasmid activity in ERα-positive MCF-7 cells. Actually, ERα-knockdown cells were more sensitive than ERα positive cells to furanodiene on the cytotoxicity effect. So the anti-cancer effects of furanodiene and non-steroidal agent in breast cancer cells still requires further investigation. Our results showed that furanodiene exposure could enhance growth inhibitory effects of doxorubicin in ERα-negative MDA-MB-231 cells and ERα-low expression 4T1 cells. However, furanodiene did not increase the cytotoxicity of doxorubicin in ERα-positive breast cancer cells, non-tumorigenic breast epithelial cells, macrophage cells, hepatocytes cells, pheochromocytoma cells and cardiac myoblasts cells. Furanodiene enhances the anti-cancer effects of doxorubicin in ERα-negative breast cancer cells through suppressing cell viability via inducing apoptosis in mitochondria-caspases-dependent and reactive oxygen species-independent manners. These results indicate that furanodiene may be a promising and safety natural agent for cancer adjuvant therapy in the future.

  18. [Development of anti-cancer drugs under new renewed GCP--from the viewpoint of drug development company developer].

    Science.gov (United States)

    Ueno, T; Kobayashi, T; Inoue, K; Yanagi, Y; Yamada, Y

    1998-04-01

    During the past 7 years since the enforcement of Japan's first GCP in October 1990, various standards and guidelines have been introduced in Japan. On the other hand, the harmonization of GCP has been the subject of major discussion at ICH in order to allow the mutual acceptance of clinical data from different countries. In order to further improve the reliability and consistency of clinical data and the ethics of clinical trials in Japan, the new GCP was enforced in April 1997. A clinical study is conducted by the sponsor, but will only be successful with the collaboration of trial subjects, medical institutions, heads of medical institutions, investigators, subinvestigators, pharmacists, nurses, laboratory technicians, and other assisting staff. Before the full enforcement of the new GCP, we, as sponsors of clinical trials, carried out a survey of the current status of clinical trials centering on the reactions of medical institutions to the new GCP, future of clinical trials on anti-cancer drugs in Japan, and differences in time from clinical trials to registration in Japan, the United State and Europe. We sent a questionnaire by facsimile to 21 pharmaceutical companies which have developed or are developing anti-cancer drugs and obtained replies from 20 companies (95%) from August 25 to 30, 1997. This paper reports issues concerning clinical trials on anti-cancer drugs based on the results of our survey.

  19. Comparative gene expression analysis of Dtg, a novel target gene of Dpp signaling pathway in the early Drosophila melanogaster embryo.

    Science.gov (United States)

    Hodar, Christian; Zuñiga, Alejandro; Pulgar, Rodrigo; Travisany, Dante; Chacon, Carlos; Pino, Michael; Maass, Alejandro; Cambiazo, Verónica

    2014-02-10

    In the early Drosophila melanogaster embryo, Dpp, a secreted molecule that belongs to the TGF-β superfamily of growth factors, activates a set of downstream genes to subdivide the dorsal region into amnioserosa and dorsal epidermis. Here, we examined the expression pattern and transcriptional regulation of Dtg, a new target gene of Dpp signaling pathway that is required for proper amnioserosa differentiation. We showed that the expression of Dtg was controlled by Dpp and characterized a 524-bp enhancer that mediated expression in the dorsal midline, as well as, in the differentiated amnioserosa in transgenic reporter embryos. This enhancer contained a highly conserved region of 48-bp in which bioinformatic predictions and in vitro assays identified three Mad binding motifs. Mutational analysis revealed that these three motifs were necessary for proper expression of a reporter gene in transgenic embryos, suggesting that short and highly conserved genomic sequences may be indicative of functional regulatory regions in D. melanogaster genes. Dtg orthologs were not detected in basal lineages of Dipterans, which unlike D. melanogaster develop two extra-embryonic membranes, amnion and serosa, nevertheless Dtg orthologs were identified in the transcriptome of Musca domestica, in which dorsal ectoderm patterning leads to the formation of a single extra-embryonic membrane. These results suggest that Dtg was recruited as a new component of the network that controls dorsal ectoderm patterning in the lineage leading to higher Cyclorrhaphan flies, such as D. melanogaster and M. domestica.

  20. Love life, scientific anti-cancer, better life%关爱生命,科学防癌,让生活更美好

    Institute of Scientific and Technical Information of China (English)

    Yi Cheng

    2010-01-01

    @@ The Chinese Anti-cancer Association launched the16th National Tumor Prophylaxis and Treatment Week in April, 2010. In order to highlight the week's spirit of "Love life, scientific anti-cancer, better life", the Tongji Cancer Center and Thoracic Surgery Department held lectures and advisory services in the outpatient lobby of Cancer Center on April 20th, 2010. The content of lec-tures involves the prophylaxis, diagnosis and treatment of lung cancer.

  1. Cell-type independent MYC target genes reveal a primordial signature involved in biomass accumulation.

    Directory of Open Access Journals (Sweden)

    Hongkai Ji

    Full Text Available The functions of key oncogenic transcription factors independent of context have not been fully delineated despite our richer understanding of the genetic alterations in human cancers. The MYC oncogene, which produces the Myc transcription factor, is frequently altered in human cancer and is a major regulatory hub for many cancers. In this regard, we sought to unravel the primordial signature of Myc function by using high-throughput genomic approaches to identify the cell-type independent core Myc target gene signature. Using a model of human B lymphoma cells bearing inducible MYC, we identified a stringent set of direct Myc target genes via chromatin immunoprecipitation (ChIP, global nuclear run-on assay, and changes in mRNA levels. We also identified direct Myc targets in human embryonic stem cells (ESCs. We further document that a Myc core signature (MCS set of target genes is shared in mouse and human ESCs as well as in four other human cancer cell types. Remarkably, the expression of t