TRAIL-induced apoptosis in prostate cancer

TRAIL 诱导前列腺癌细胞凋亡

• 批准号: 6863702
• 负责人: CHRISTINA VOELKEL-JOHNSON
• 金额: $ 23万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-04-01 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6863702
• 关键词: adeno associated virus group; apoptosis; athymic mouse; cell cycle; cell population study; combination cancer therapy; confocal scanning microscopy; cysteine endopeptidases; doxorubicin; enzyme inhibitors; flow cytometry; gene delivery system; gene expression; gene therapy; genetic regulation; genetically modified animals; immunoprecipitation; prostate neoplasms; protein localization; protein protein interaction; transfection /expression vector; tumor necrosis factor alpha; ubiquitin; western blottings

DESCRIPTION (provided by applicant): Prostate cancer is a significant health problem among American men. Curative treatments for advanced disease are currently not available and novel treatment approaches are needed. Recombinant forms of TRAIL (tumor necrosis factor related apoptosis inducing factor) have been considered as novel anti-tumor agents but prostate cancer cells are relative resistant to soluble TRAIL. Resistance is overcome by sub-toxic doses of doxorubicin which downregulate the anti-apoptotic protein c-FLIP or by an adenovirus expressing membrane TRAIL. Neither approach is selective for malignant cells and will require restriction of either doxorubicin or TRAIL to a specific target population. It is our hypothesis that combination of doxorubicin with tissue-specific expression of TRAIL from the prostate-specific promotor probasin will be an effective therapeutic approach against prostate cancer. To support this hypothesis and identify underlying mechanisms of TRAIL resistance in prostate cells, the following specific aims will be investigated: (1) To test the hypothesis that the mechanism of doxorubicin-mediated downregulation of c-FLIP involves cell cycle dependent ubiquitination. Cells that are sorted into G0/G1, S and G2/M populations will be analyzed for levels of c-FLIP, ubiquitination, and TRAIL susceptibility. (2) To test the hypothesis that the TRAIL expressing adenovirus (AdTRAIL) overcomes resistance by permitting intracellular interaction of ligand and agonistic TRAIL receptors resulting in formation of an intracellular death inducing signaling complex. The role of c-FLIP in providing resistance to AdTRAIL will be investigated in PC3 cells stably overexpressing long and short isoforms of the protein. Intracellular location of TRAIL receptors and interaction with ligand, caspases and c-FLIP will be analyzed by confocal microscopy. Protein interactions are also determined by immunoprecipitation/immunoblotting. (3) To test the hypothesis that combination of chemotherapy and prostate-specific gene therapy is an effective approach against prostate cancer. The effectiveness of specificity of an adenovirus-expressing TRAIL from the prostate-specific promotor ARR2PB will be tested in cells with a functional androgen receptor and non-prostate cells. The apoptotic potential of virus alone or in combination with doxorubicin will be determined in vitro. The toxicity and effectiveness of AdARR2PBTRAIL alone or in combination with doxorubicin will also be tested in subcutaneous xenografts in athymic mice.

描述（由申请人提供）：前列腺癌是美国男性的重大健康问题。目前尚无治疗晚期疾病的治疗方法，需要新的治疗方法。重组形式的TRAIL形式（肿瘤坏死因子相关的凋亡因子）被认为是新型的抗肿瘤剂，但前列腺癌细胞相对抗可溶性步道。亚毒素的亚毒素剂量可以克服耐药性，该剂量下调抗凋亡蛋白C翼或通过表达膜径的腺病毒病毒。两种方法都不是针对恶性细胞的选择性，并且需要限制阿霉素或对特定目标群体的轨迹。我们的假设是，阿霉素与前列腺特异性启动子探针的TRAIL的组织特异性表达的结合将是针对前列腺癌的有效治疗方法。为了支持这一假设并确定前列腺细胞中耐trail耐药性的潜在机制，将研究以下特定目的：（1）检验以下假设：阿霉素介导的C翼下调C涉及细胞周期依赖性的泛素化。将分类为G0/G1，S和G2/M种群的细胞，分析C翼，泛素化和TRAIL敏感性的水平。 （2）检验以下假设：表达腺病毒（Adtrail）通过允许配体的细胞内相互作用和激动性径向受体的受体来克服抗药性，从而导致形成细胞内死亡的诱导信号传导复合物。在PC3细胞中稳定过表达蛋白质的长期和短同工型的PC3细胞将研究C翼在提供对adtrail的抗性中的作用。路径受体的细胞内位置以及与配体，胱天蛋白酶和C叶片的相互作用将通过共聚焦显微镜分析。蛋白质相互作用也通过免疫沉淀/免疫印迹决定。 （3）检验化学疗法和前列腺特异性基因治疗的组合是对前列腺癌的有效方法。来自前列腺特异性启动子ARR2PB的表达腺病毒径的特异性的有效性将在具有功能性雄激素受体和非前端细胞的细胞中进行测试。单独或与阿霉素结合的病毒的凋亡潜力将在体外确定。 ADARR2PBRAIL或与阿霉素结合的毒性和有效性也将在无胸腺小鼠的皮下注射移植物中进行测试。