The Use of Hyperthermia and Copper Chelation to Overcome Cisplatin Resistance

描述(申请人提供)：抗药性，以及正常的组织毒性，仍然是成功治疗癌症的主要障碍。以这些耐药机制为靶点将导致肿瘤细胞死亡增加，肿瘤体积缩小，并可能导致患者总体生存。本研究的目的是了解高温(HT)如何改变细胞耐药机制，并利用这些能力来增加药物对肿瘤部位的输送。本申请中概述的工作将侧重于针对药物转运的低效和固有的细胞耐药机制。具体地说，我们建议将羟色胺与铜络合剂和顺铂包裹的温敏脂质体(TSL)的许多治疗优点结合起来，以靶向和治疗膀胱癌患者的肌肉侵袭性疾病。我们团队最近开发了一种封装顺铂的TSL。将该药物制剂与羟色胺联合使用是一种方式，它将克服药物传输的低效，并减少对其他组织的不良毒性。众所周知，羟色胺与包括顺铂在内的多种化疗药物具有协同作用，但这种相互作用的分子机制尚不清楚。铜转运蛋白1(CTR1)是细胞内顺铂摄取的主要调节因子。我们的初步数据显示，在野生型细胞中，HT增加顺铂的摄取和铂-DNA加合物的形成，但在CTR1-/-细胞中则不增加，这表明CTR1参与了这种协同作用。我们研究的目标之一是使用铜络合剂来创造铜缺乏的环境，以提高细胞表面CTR1的表达水平。在顺铂存在的情况下，CTR1膜表达增加将导致药物摄取、蓄积和细胞毒性增加。通过铜螯合增加CTR1的表达来提高顺铂的摄取和疗效还有待研究，可能被证明是一种具有临床适用性和潜力的有效治疗方式。我们假设，利用基于TSL的系统与CTR1操作相结合将增加顺铂在肿瘤细胞中的输送和摄取，并将被证明是一种有效的抗癌策略。我们将用两个具体的目标来解释我们的假设。在体外，我们计划阐明CTR1在顺铂和羟色胺协同作用中所起的作用，并利用这一知识来提高药物疗效。在体内，我们将检测顺铂-TSL和HT联合的药代动力学和抗肿瘤效果，并应用CTR1研究获得的信息来改进这种联合治疗，以增加整体抗癌效果。由于顺铂和铜络合剂目前都是FDA批准的药物，我们建议的治疗方案应该有一个相对简单的过渡到临床。 公共卫生相关性：肌肉侵袭性疾病(MID)约占新诊断的膀胱癌患者的33%，10-15%的浅表性疾病患者最终将进展为MID。大约50%的MID患者发生转移，这反过来又显着降低了存活率。目前对MID的标准治疗包括新辅助化疗和含有顺铂的方案，然后是根治性膀胱切除术。利用区域热疗(HT)、顺铂温敏脂质体(TSL)和铜络合的新组合，我们的目标是开发一种治疗MID的策略，既能对患者的生存产生积极影响，又能通过保留膀胱来提高患者的生活质量。羟色胺是一种耐受性良好的有效辅助治疗方法，可增加药物输送和药物靶向，特别是在脂质体治疗的情况下，作为TSL的局部药物释放触发因素。顺铂与羟色胺具有协同作用，导致细胞毒性增加。我们提出的工作旨在开发和优化一种独特的治疗组合，以克服药物疗效的三个障碍：药物传递、细胞耐药性和正常组织毒性。我们将使用新型顺铂-TSL联合HT来增加肿瘤药物的释放，降低正常组织的毒性，并使用铜螯合和HT来操纵顺铂细胞转运体CTR1，以增加顺铂在细胞内的蓄积和细胞毒性。

DESCRIPTION (provided by applicant): Drug resistance, along with normal tissue toxicity, remains a major obstacle for the successful treatment of cancer. Targeting these resistance mechanisms will result in increased tumor cell death, decreased tumor size, and may lead to overall patient survival. The purpose of this study is to understand how hyperthermia (HT) alters cellular resistance mechanisms and to exploit these abilities to increase drug delivery to the tumor site. The work outlined in this application will focus on targeting inefficiencies in drug transport and inherent cellular resistance mechanisms. Specifically, we propose to combine the many therapeutic benefits of HT with copper chelators and cisplatin-encapsulated thermosensitive liposomes (TSL) to target and treat muscle invasive disease in bladder cancer patients. Our group has recently developed a TSL that encapsulates cisplatin. Using this drug formulation in combination with HT is a modality that will overcome inefficiencies in drug transport, and reduce the undesirable toxicity to other tissues. HT is known to interact synergistically with multiple chemotherapeutic drugs, including cisplatin, but the molecular mechanisms underlying this interaction remain unclear. Copper transporter protein 1 (Ctr1) is a major regulator of cellular cisplatin uptake. Our preliminary data show that HT increases cisplatin uptake and platinum-DNA adduct formation in wild type but not in Ctr1-/- cells indicating the involvement of Ctr1 in this synergism. One goal of our study is to use copper chelators to create an environment of copper deficiency in order to increase Ctr1 expression levels on the cell surface. Increased Ctr1 membrane expression in the presence of cisplatin will lead to increased drug uptake, accumulation, and cytotoxicity. Enhancing cisplatin uptake and efficacy by increasing Ctr1 expression via copper chelation has yet to be studied and could prove to be an effective treatment modality with clinical applicability and potential. We hypothesize that utilizing the combination of a TSL-based system paired with Ctr1 manipulation will increase cisplatin delivery and uptake in tumor cells and will prove to be an effective anti-cancer strategy. We will address our hypothesis with two Specific Aims. In vitro, we plan to elucidate the role Ctr1 plays in the synergistic interaction between cisplatin and HT and use this knowledge to increase drug efficacy. In vivo, we will examine the pharmacokinetics and anti-tumor effect of the cisplatin-TSL and HT combination and apply the information derived from the Ctr1 studies to improve this combinational treatment to increase the overall anti- cancer effect. Since cisplatin and copper chelators are currently both FDA approved drugs, our proposed treatment regime should have a relatively straightforward transition into the clinic. PUBLIC HEALTH RELEVANCE: Muscle invasive disease (MID) accounts for approximately 33% of the newly diagnosed bladder cancer patients, and 10-15% of patients with superficial disease will eventually progress to MID. Approximately 50% of MID patients develop metastases which in turn significantly decreases survival rates. Current standard of care for MID involves neo-adjuvant chemotherapy with a cisplatin containing regimen followed by radical cystectomy. Using a novel combination of regional hyperthermia (HT), cisplatin-thermosensitive liposome (TSL), and copper chelation, our goal is to develop a treatment strategy for MID that will both positively impact patient survival and improve patient quality of life by allowing for bladder preservation. HT is a well-tolerated, useful adjuvant treatment that increases drug delivery and drug targeting, particularly in the case of liposomal therapeutics by acting as a focal drug release trigger for TSL. Cisplatin synergistically interacts with HT resulting in increased cytotoxicity. Our proposed work seeks to develop and optimize a unique combination of treatments that will overcome three barriers of drug efficacy: drug delivery, cellular drug resistance, and normal tissue toxicity. We will use the novel cisplatin-TSL combined with HT to increase tumor drug delivery and decrease normal tissue toxicity and copper chelation and HT to manipulate the cisplatin cellular transporter Ctr1 to increase cellular cisplatin accumulation and cytotoxicity.