CIRCUMVENTION OF CISPLATIN RESISTANCE

尽管铂基抗肿瘤剂（即，顺铂和卡铂）在几种癌症的治疗中起着关键作用，这是大多数患者复发的主要障碍，这些患者由于其肿瘤细胞中耐药性的出现而未能用铂剂进行后续挑战。 这种限制导致了一致的化学努力，以确定铂类似物，有效对抗耐药肿瘤。 实际上，几种这样的类似物（例如，奥沙利铂和JM 216）已进入临床试验。 然而，我们对这些类似物的有效作用模式知之甚少或一无所知。 这些信息对于它们的成功及其在耐药疾病环境中的合理临床应用至关重要。 尽管减少的药物蓄积、增加的细胞内谷胱甘肽和增加的DNA加合物修复通常被确定为顺铂耐药的关键机制，但它们没有为类似物逆转耐药提供令人满意的解释。 最近，我们已经证明，在p53激活顺铂的DNA损伤的响应损失是耐药的基本机制，这种铂络合物。 p53可能通过替代途径被激活。 因此，所提出的特定目的旨在研究类似物诱导潜伏性p53的潜力。 我们将在DNA水平上研究p53功能对类似物的生化药理学的影响，研究结构多样的类似物对DNA损伤的识别，并描述p53的调节。 为了解决具体的目标，我们将使用标准的转染方法来调节p53的诱导和/或功能，并利用生物化学，药理学和其他分子技术来研究这种调节对逆转耐药性的影响。 具体技术将包括加合物定量、DNA链断裂、RNA/蛋白质分离、蛋白质免疫沉淀、凝胶电泳、单克隆/多克隆抗体检测和流式细胞术。 我们的研究很可能将确定类似物如何逆转顺铂耐药性，这可能为设计临床方案以在更合理的基础上治疗耐药的临床癌症提供基础。

Although platinum-based antitumor agents (i.e., cisplatin and carboplatin) play critical roles in the treatment of several cancers, a major impediment relapse in a majority of patients, who fail subsequent challenge with the platinum agent due to the onset of drug resistance in their tumor cells. This limitation has resulted in a concerted chemical effort to identify platinum analogs that are effective against resistant tumors. Indeed several such analogs (e.g., oxaliplatin and JM216) have entered clinical trials. However, we have little or no knowledge of the effective mode of action of these analogs. Such information is vital for their success and their rational clinical application in the resistant disease setting. Although reduced drug accumulation, increased intracellular glutathione, and increased DNA adduct repair are usually identified as key mechanisms of resistance of cisplatin, they have not provided satisfactory explanations for reversal of resistance by the analogs. Recently we have demonstrated that a loss in p53 activation in response to DNA damage by cisplatin is a fundamental mechanism of resistance to this platinum complex. It is possible for p53 to be activated through an alternative pathway. Thus, the proposed specific aims are designed to investigate the potential of the analogs to induce latent p53. We will examine the consequence of p53 function on the biochemical pharmacology of analogs at the DNA level, investigate recognition of DNA damage by the structurally- diverse analogs, and delineate p53 regulation. To address the specific aims, we will use standard transfection approaches to modulate p53 induction and/or function, and utilize biochemical, pharmacologic, and other molecular techniques to investigate the effect of this modulation on reversal of resistance. The specific techniques will include adduct quantitation, DNA strand breaks, RNA/protein isolation, protein immunoprecipitation, gel electrophoresis, detection by monoclonal/polyclonal antibodies, and flow cytometry. It is likely that our investigations will establish how the analogs reverse cisplatin resistance, and this may provide a basis for designing clinical protocols to treat resistant clinical cancers on a more rational basis.