CIRCUMVENTION OF CISPLATIN RESISTANCE

虽然铂类抗肿瘤药物(如顺铂和卡铂)在多种癌症的治疗中发挥了关键作用，但在大多数患者中，这是一种主要的阻碍复发的因素，因为他们的肿瘤细胞开始产生耐药性，因此无法接受铂类药物的后续挑战。这一局限性导致了一种协调一致的化学努力，以确定对耐药肿瘤有效的铂类似物。事实上，几个这样的类似物(例如奥沙利铂和JM216)已经进入临床试验。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和它们在抵抗性疾病环境中的合理临床应用至关重要。虽然药物积聚减少、细胞内谷胱甘肽增加和DNA加合物修复增加通常被认为是顺铂耐药的关键机制，但它们并不能为类似物逆转耐药性提供令人满意的解释。最近，我们已经证明，顺铂对DNA损伤导致的P53活性丧失是这种铂复合体耐药的一个基本机制。P53有可能通过另一种途径被激活。因此，提出的特定目的是为了研究类似物诱导潜伏的P53的潜力。我们将在DNA水平上研究P53功能对类似物生化药理的影响，研究结构多样化的类似物对DNA损伤的识别，并描述P53的调控。为了达到特定的目的，我们将使用标准的转染方法来调节P53的诱导和/或功能，并利用生化、药理学和其他分子技术来研究这种调节在逆转耐药中的作用。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、单抗/多克隆抗体检测和流式细胞术。我们的研究很可能会确定类似物如何逆转顺铂耐药性，这可能会为设计临床方案以更合理地治疗耐药临床癌症提供基础。

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.