Clinical Studies of Multidrug Resistance Reversal

我们部门研究计划的重点是开发旨在克服癌症耐药性的治疗策略。我们的研究一直致力于将耐药性逆转策略转化为临床。在实验室的支持下，我们的临床试验设计得到了加强，这使我们能够分析临床样本并解释临床试验结果。一项重要的临床试验与p -糖蛋白的抑制有关，p -糖蛋白是一种ABC转运蛋白，通过抗癌药物的外转运介导耐药性。这些研究是与Tito Fojo博士合作进行的，评估了Pgp调节可能增加抗癌药物疗效的假设。在全球范围内进行的试验中，从失败的第一代试验开始，因为使用的药物没有足够的效力，然后是失败的第二代试验，主要集中在缬草酰胺及其伴随的抗癌药物剂量减少的需要，在治疗策略上有很多令人失望的地方。就连一项多国随机试验也因为毒性问题而提前结束，该试验将新药物tariquar与紫杉醇或长春瑞滨联合使用。必须指出的是，到目前为止，还没有令人信服的证据表明这种策略最终会提供临床益处，而且耐药性逆转模式仍然是一种假设。然而，失败的早期策略并不能否定支持Pgp拮抗剂继续发展的有力证据。该项目对于多种肿瘤类型具有高风险和潜在的高收益，因此非常适合NCI内部项目。目前的研究正在评估第三代抑制剂tariquar （XR9576）。在我们完成的vinorelbine和tariquidar的I期相互作用研究中，在CD56+细胞中观察到pgp介导的药物外排的完全抑制，单次静脉注射tariquidar后抑制持续48小时。99mTc-sestamibi成像被用作正常和肿瘤组织中药物积累改变的替代方法。超过一半的患者有可检测到的99mTc-sestamibi肿瘤摄取增加。我们开展一项新的tariquidar试验的目的是收集更多关于tariquidar与一种化疗药物联合使用的安全性的数据，并确定一种可以作为单一药物使用的组合。多西他赛被选为一种极好的Pgp底物，具有已知的疗效，可以通过增加肺癌、宫颈癌或卵巢癌的药物积累而受益。在计划多西他赛与tariquar的相互作用试验时，我们选择了一个有效但保守的剂量，多西他赛75mg /m2，每q-3周。该试验设计了药代动力学和药效学分析。为了检查他奎达是否会干扰多西他赛的清除，将对多西他赛加用和不加用他奎达的剂量进行仔细的药代动力学研究。为了限制患者在没有调节剂的情况下治疗的时间长度，研究的药代动力学部分以两次40mg /m2的多西他赛剂量进行，间隔一周。给药顺序在第1天和第8天之间随机分配。患者开始以75 mg/m2的剂量治疗，每3周，在第二个周期中联合tariquidar。除了药代动力学分析，99mTc-sestamibi研究在每个入组患者中进行，有或没有tariquidar，我们的实验室在外周单核细胞中进行CD56+罗丹明测定。该试验是开放的，并积累了没有重大毒性的患者，我们对非小细胞肺癌患者的疾病反应特别感兴趣。尽管99mTc-sestamibi研究提供了很好的概念证明，表明在tariquida后放射性核素积累增加，但这些研究的定量很差，因为它们是平面图像，背景往往压倒差异。在Peter Herscovitch博士的带领下，临床中心PET部门开发了一种用94mTc标记sestamibi用于正电子发射成像的方法，有望成为一种更定量的显像剂。该药物的临床试验是开放的，并增加了患者。我们希望定量PET成像能让我们更好地回答tariquar对患者肿瘤有多大影响的问题。除了PET-sestamibi试验外，我们还与Robert Innis博士和Karen Kurdziel博士讨论了合作，旨在使用PET制剂11c - n -去甲基-洛哌丁胺和18f -紫杉醇评估药物积累。这些PET研究为该领域的重大发展提供了机会。除了tariquidar的研究，CBA Pharma是一家开发Pgp抑制剂CBT-1的小公司。这是一种经过临床试验的口服药物。然而，该药物对Pgp的抑制作用尚未在患者中得到证实。我们已经在离体实验中证实了这种药剂的活性。在接受紫杉醇和CBT-1治疗的患者中，一项临床试验检查了Pgp抑制的替代标志物，包括循环CD56+单核细胞中sestamibi摄取的改变和罗丹明摄取的增加，该试验已提交给IRB进行最终审查。我们的实验室也对研究其他模型系统的耐药性保持兴趣。几年前，在与NCIs发育治疗项目的合作中，我们根据60个细胞系面板的细胞毒性数据进行比较分析，确定了一些对肾细胞癌具有选择性的化合物。这些化合物在我们的实验室进行了评估，并证实了肾脏选择性。其中一类化合物，二甲烷磺酸盐，一直在DTP的临床前开发中，其中一类化合物NSC-281612已被批准进行I期试验。正在准备一份协议，并将提交一份IND。I期试验的目标之一是开发生物标志物，以评估DMS化合物治疗后肿瘤细胞或替代组织中DNA损伤的存在。后一个项目的第二个方面是利用生物信息学策略评估其他耐药性机制。该项目是与耶路撒冷希伯来大学的Wilfred Stein博士和丹麦哥本哈根大学的Thomas Litman博士合作进行的。两种不同的生物信息学策略利用公开数据库中报告的临床样本中的基因表达数据，确定了细胞粘附是一种耐药性机制。这些实验与已有的文献一致，表明细胞粘附是耐药性的重要机制。有趣的是，对体外细胞粘附模型（如球体形成）的评估表明，以这种方式培养的细胞与在塑料上培养的融合细胞具有相似的基因表达谱。这表明球形培养本身不能作为内在耐药模型，应该开发其他体外模型

The focus of our sections research program is to develop therapeutic strategies aimed at overcoming drug resistance in cancer. Our research has been dedicated to the translation of drug resistance reversal strategies to the clinic. The design of our clinical trials has been enhanced by laboratory support that has allowed us to analyze clinical samples and interpret the clinical trial findings. A significant clinical trial effort has related to the inhibition of P-glycoprotein, an ABC transporter mediating resistance through outward transport of anticancer agents. These studies, which have been carried out collaboratively with Dr. Tito Fojo, evaluate the hypothesis that Pgp modulation may increase anticancer drug efficacy. In trials carried out across the globe, beginning with the failed first-generation trials that employed agents without sufficient potency, and continuing with the failed second-generation trials centered on valspodar with its accompanying need for anticancer agent dose reduction, there has been much disappointment in therapeutic strategy. Even a multinational randomized trial combining the new agent tariquidar with paclitaxel or vinorelbine closed early for toxicity. It must be stated that there is no convincing proof to date that this strategy will eventually be shown to provide clinical benefit and the resistance reversal paradigm remains a hypothesis. However, the failed earlier strategies do not negate strong evidence supporting continued development of Pgp antagonists. The project can be viewed as high risk with potentially high gain for multiple tumor types and thus very appropriate for the NCI intramural program. Current studies are evaluating the third generation inhibitor tariquidar (XR9576). In our completed Phase I interaction study with vinorelbine and tariquidar, total inhibition of Pgp-mediated drug efflux was observed in CD56+ cells, with persistence of inhibition for 48 hours after a single intravenous dose of tariquidar. 99mTc-sestamibi imaging was employed as a surrogate for altered drug accumulation in normal and tumor tissues. More than half of the patients had detectable increases in tumor uptake of 99mTc-sestamibi. Our goal in launching a new tariquidar trial was to gather more data regarding the safety of tariquidar in combination with a chemotherapeutic agent and to identify a combination that could be used as a single agent. Docetaxel was chosen as an excellent Pgp substrate with known efficacy that could be benefited by increasing drug accumulation in lung, cervical, or ovarian cancer. In planning an interaction trial of docetaxel with tariquidar, we selected an effective but conservative dose of docetaxel 75 mg/m2 on a q-3-week schedule. The trial is designed with both pharmacokinetic and pharmacodynamic assays. To examine whether tariquidar interferes with docetaxel clearance, careful pharmacokinetics will be performed on a dose of docetaxel administered with and without tariquidar. To limit the length of time that a patient is treated without the modulator, the pharmacokinetic portion of the study is carried out on two 40 mg/m2 docetaxel doses, one week apart. The order of administration of tariquidar is randomized between the day 1 and day 8 doses. Patients begin therapy with 75 mg/m2 q-3-weeks in combination with tariquidar in the second cycle. In addition to pharmacokinetic analysis, 99mTc-sestamibi studies are performed in each enrolled patient with and without tariquidar, and our laboratory carries out CD56+ rhodamine assays in peripheral mononuclear cells. The trial is open and accruing patients without major toxicity, and we have been particularly intrigued by the disease responses in patients with nonsmall cell lung cancer. Although the 99mTc-sestamibi studies provide good proof-of-concept showing increased radionuclide accumulation following tariquidar, the studies are poorly quantitative because they are planar images and background often overwhelms differences. Led by Dr. Peter Herscovitch, the Clinical Center PET department developed a method to label sestamibi with 94mTc for positron emission imaging, promising a more quantitative imaging agent. A clinical trial testing this agent has is open and accruing patients. It is our hope that the quantitative PET imaging will allow us to better answer the question of how much impact tariquidar can have on patient tumors. In addition to the PET-sestamibi trial, we have discussed collaborations with Dr. Robert Innis and Dr. Karen Kurdziel aimed at evaluating drug accumulation using PET agents 11C-N-desmethyl-loperamide and 18F-paclitaxel. These PET studies offer the opportunity to move the field forward in a significant way. In addition to the tariquidar studies, our group was approached by CBA Pharma, a small company developing a Pgp inhibitor CBT-1. This is an oral agent that has been in clinical trials. However, inhibition of Pgp by this agent has not been confirmed in patients. We have confirmed activity of the agent in an ex vivo assay. A clinical trial examining surrogate markers of Pgp inhibition including altered sestamibi uptake and increased rhodamine uptake in circulating CD56+ mononuclear cells in patients treated with paclitaxel and CBT-1 has been submitted for final review by the IRB. Our laboratory also maintains an interest in studying drug resistance in other model systems. Several years ago, in collaboration with the NCIs Developmental Therapeutics Program, we identified a number of compounds with selectivity against renal cell caner, based on COMPARE analysis using cytotoxicity data in the 60 cell line panel. These compounds were evaluated in our laboratory and the renal selectivity confirmed. One class of these compounds, the dimethane sulfonates, has been continuously in preclinical development at DTP and one, NSC-281612, has been approved for Phase I testing. A protocol is in preparation, and an IND will be submitted. One of the goals in the Phase I trial will be the development of biomarkers to evaluate the presence of DNA damage in tumor cells or surrogate tissues following treatment with the DMS compound. A second aspect of this latter project is the evaluation of other mechanisms of drug resistance using Bioinformatics strategies. This project is carried out in collaboration with Dr. Wilfred Stein, Hebrew University, Jerusalem, and Dr. Thomas Litman, University of Copenhagen, Denmark. Two separate bioinformatics strategies identified cell adhesion as a mechanism of resistance, using gene expression data in clinical samples reported in publicly available databases. These experiments resonated with a literature already available suggesting that cell adhesion was an important mechanism of resistance. Interestingly, evaluation of in vitro models of cell adhesion -- such as spheroid formation-- have suggested that cells cultured in this manner have similar gene expression profiles to confluent cells cultured on plastic. This would suggest that spheroid cultures per se cannot serve as models of intrinsic drug resistance and that other in vitro models should be developed