Genetic determinants associated with pemetrexed response and toxicity

描述（由申请人提供）：培美曲塞是一种多靶点抗叶酸药物，最近被批准用于非小细胞肺癌（NSCLC）的二线治疗。在临床可达到的浓度下，最显著的相互作用是与胸苷酸合成酶（TYMS），尽管已知该药物至少靶向其他两种叶酸依赖酶。本提案的目的是通过采用全基因组、全面的方法来验证遗传多态性显著影响培美曲塞相关毒性和反应易感性的假设，从而确定影响培美曲塞相关毒性和反应的其他遗传变异。为此，将对培美曲塞的细胞毒性和细胞凋亡进行表型分析，这些细胞系来自于大型参考人类多态研究中心（Centre d’Etude du Polymorphisme）的家谱和HapMap（高加索、约鲁班、亚洲）样本。基因型信息在公共领域是可用的，这使我们能够应用家族遗传策略，包括连锁分析，来检查基因组中含有对培美曲塞诱导的细胞毒性和细胞凋亡易感性的遗传变异的区域。此外，将使用来自非洲、欧洲和亚洲后裔的国际HapMap样本，比较培美曲塞诱导的细胞毒性的个体间和种族间差异。我们实验室生成的外显子表达阵列将被纳入鉴定基因变异，这些变异通过影响与培美曲塞敏感性相关的基线基因表达来起作用。通过这种全基因组方法鉴定的遗传变异将在lcl的单独样本集中以及通过定量PCR和siRNA进行验证。我们将评估TYMS、GARFT、DHFR中新的snp和已知的候选变异，以确定培美曲塞与厄洛替尼作为NSCLC二线治疗的组间（NCCTG、CALGB、ECOG、SWOG、NCI-C）研究中与反应或毒性相关的基因型。我们的长期目标是确定一种“药理学特征”，以预测培美曲塞反应机会降低或不良反应风险增加的患者。具体目标是：1.；目的：探讨培美曲塞对细胞毒性和细胞凋亡敏感性的遗传影响。2. 检查培美曲塞治疗后的基线和时间依赖性表达模式。3. 通过国际HapMap项目提供的数据，在欧洲人（CEU）、非洲人（YRI）和亚洲人（CHB， JPT）的细胞系中确定与培美曲塞细胞毒性和基因表达相关的snp。4. 在一项多中心试验中，确定TYMS、GARFT和DHFR的共同多态性以及在特定目标1- 3中确定的候选多态性是否与培美曲塞治疗晚期NSCLC患者的反应和/或毒性相关。公共卫生相关性：本研究的目的是确定使用培美曲塞（一种化疗药物）治疗后存在毒性或缺乏反应风险的肺癌患者。该项目旨在利用细胞系建立一个模型，以更好地了解基因变异如何解释患者对药物反应的差异。此外，我们将在培美曲塞治疗的肺癌患者中测试我们的模型结果。最终，我们希望找到培美曲塞反应和毒性的预测标记。

DESCRIPTION (provided by applicant): Pemetrexed is a multitargeted antifolate recently approved for second line therapy of non-small cell lung cancer (NSCLC). The most significant interaction at clinically achievable concentrations is with thymidylate synthase (TYMS), although the drug is known to target at least two other folate dependent enzymes. The goal of this proposal is to identify additional genetic variants that influence pemetrexed-associated toxicity and response by employing a genome-wide, comprehensive approach to test the hypothesis that genetic polymorphisms significantly influence susceptibility to toxicities and response associated with pemetrexed. To this end, lymphoblastoid cell lines, derived from large reference Centre d' Etude du Polymorphisme Humain pedigrees and HapMap (Caucasian, Yoruban, Asian) samples, will be phenotyped for pemetrexed cytotoxicity and apoptosis. Genotypic information is available in the public domain enabling us to apply familial genetic strategies, including linkage analysis, to examine regions in the genome harboring the genetic variants responsible for susceptibility to pemetrexed-induced cytotoxicity and apoptosis. In addition, inter-individual and inter-ethnic variation in pemetrexed-induced cytotoxicity will be compared using the International HapMap samples from populations of African, European and Asian descent. Exon expression array generated in our laboratory will be incorporated to identify genetic variants acting through their effect on baseline gene expression that associate with sensitivity to pemetrexed. Genetic variants identified through this whole genome approach will be validated in a separate sample set of LCLs and by quantitative PCR and siRNA. We will evaluate both novel SNPs and known candidate variants in TYMS, GARFT, DHFR to determine genotypes associated with response or toxicity in an intergroup (NCCTG, CALGB, ECOG, SWOG, NCI-C) study of pemetrexed compared to erlotinib as second line therapy for NSCLC. Our long-term goal is to identify a "pharmacogenetic signature" that will predict patients with a decreased chance for response or an increased risk for adverse reactions to pemetrexed. Specific aims are: 1. To determine the genetic contribution of susceptibility to the cytotoxic and apoptotic effects of pemetrexed. 2. To examine baseline and time dependent patterns of expression following pemetrexed treatment. 3. Identify SNPs, through data available from the International HapMap project, associated with both pemetrexed cytotoxicity and gene expression in cell lines derived from Europeans (CEU), Africans (YRI) and Asians (CHB, JPT). 4. To determine if a common polymorphism in TYMS, GARFT, and DHFR, as well as candidate polymorphisms identified in specific aims 1- 3 correlate with response and/or toxicity in advanced NSCLC patients treated with pemetrexed in a multicenter trial. PUBLIC HEALTH RELEVANCE: The objective of this research is to identify lung cancer patients at risk for toxicities or lack of response following treatment with pemetrexed, a chemotherapeutic agent. The project aims to build a model using cell lines to better understand how genetic variation explains differences in patient response to drug. In addition, we will test the results of our model in lung cancer patients treated with pemetrexed. Ultimately, we hope to find predictive markers of pemetrexed response and toxicity.