Modeling the role of the genome in chemotherapy induced cardiotoxicity using iPSC

使用 iPSC 模拟基因组在化疗引起的心脏毒性中的作用

• 批准号: 9130233
• 负责人: Paul W. Burridge
• 金额: $ 24.77万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2018-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9130233
• 关键词: ABCB1 gene; ASBT protein; Acute; Adult; Adverse effects; Advisory Committees; Affect; Aftercare; American Society of Clinical Oncology; Animal Model; Animals; Anthracyclines; Binding; Biological Assay; CYBA gene; Calcium; Cardiac; Cardiac Myocytes; Cardiac development; Cardiology; Cardiotonic Agents; Cardiotoxicity; Cardiovascular Diseases; Cardiovascular system; Career Mobility; Cell Line; Cell Survival; Cells; Clinical; Clinical Trials; Cohort Studies; Complex; Complication; Data; Development; Dexrazoxane; Disease model; Dose; Doxorubicin; Drug usage; Effectiveness; Foundations; Future; Gene Mutation; Generations; Genes; Genetic; Genetic Markers; Genetic Polymorphism; Genome; Glucuronosyltransferase; Goals; HFE gene; Heart; Heart failure; Hemochromatosis; Human; In Vitro; Individual; Institutes; Knowledge; Left Ventricular Ejection Fraction; Link; Malignant Neoplasms; Malignant neoplasm of lung; Malignant neoplasm of ovary; Mass Spectrum Analysis; Measurement; Medicine; Membrane Potentials; Mentors; Mentorship; Meta-Analysis; Mitochondria; Modality; Modeling; Molecular; NADP; NADPH Oxidase; Nuclear; Odds Ratio; Oncologist; Patients; Pharmaceutical Preparations; Pharmacogenomics; Phase; Phenotype; Physicians; Pilot Projects; Population; Predisposition; Prevention; Probability; RARG gene; Research Personnel; Research Project Grants; Respiration; Risk; Role; Sampling; Scientist; Signal Transduction; Single Nucleotide Polymorphism; Somatic Cell; Staging; Stem cells; Structure; TAP1 gene; Toxic effect; Transcriptional Regulation; Translating; Treatment Protocols; Tyrosine Kinase Inhibitor; Universities; Work; analog; annexin A5; base; career; career development; catalase; cellular imaging; chemotherapeutic agent; chemotherapy; clinical practice; cohort; cytochrome b245; gene function; genetic variant; genome wide association study; genomic data; high risk; induced pluripotent stem cell; knock-down; leukemia/lymphoma; loss of function; malignant breast neoplasm; meetings; mitochondrial membrane; mutant; neutrophil; novel; novel therapeutics; overexpression; pluripotency; prevent; professor; programs; reuptake; risk variant; role model; screening; stem cell biology; tool; transcription activator-like effector nucleases

DESCRIPTION (provided by applicant): This proposal describes a five-year career development program to prepare the candidate, Dr. Paul Burridge, for a career as an independent investigator. This program will build on Dr. Burridge's background as a stem cell biologist by providing expertise in molecular cardiology and pharmacogenomics. The mentor is Dr. Joseph Wu, a Professor of Medicine/Cardiology and Director of the Stanford Cardiovascular Institute at Stanford University. The proposed mentor is a physician scientist with significant expertise in stem cell biology and is an expert in cardiovascular disease modeling. The K99 phase will consist of structured mentorship by the primary mentor, complementary meetings with the advisory committee, formal coursework, a provocative research project, and a program of career transition. Doxorubicin is a well-established and highly effective chemotherapy drug commonly used to treat multiple cancers such as lymphoma, leukemia, ovary, lung and breast cancer, but its use is limited by a serious side effect: doxorubicin causes toxicity in cardiomyocytes, causing damage to the heart. Cardiotoxicity can range from asymptomatic reductions in left ventricular ejection fraction (LVEF) to highly symptomatic (Class III to Class I) heart failure. Acute doxorubicin-induced cardiotoxicity occurs in ~11% of patients and long-term cardiotoxic side effects, which can manifest up to 10 years after treatment, are observed in up to 36% of patients. Currently we cannot predict which patients will develop cardiotoxicity and, at present, oncologists do not assess patient-specific genomic data before deciding on doxorubicin dose. Existing strategies for reducing doxorubicin-induced cardiotoxicity (DIC) include (i) reducing dose, potentially reducing chemotherapeutic effectiveness, (ii) development of less cardiotoxic anthracycline analogues, or (iii) co- treatment with a cardioprotective agents such as dexrazoxane, although this has not proven effective and is not currently endorsed by the American Society for Clinical Oncology outside of clinical trials. A major hurdle in filling the significant gaps in our knowledge about the mechanisms of cardiotoxicity and how best to prevent it has been that there are no good human models, due to the inaccessibility of adult human cardiomyocyte patient samples, and the difficulty in isolating and maintaining cardiomyocytes in vitro. Animal models are limited by significant functional disparities between animal and human cardiomyocytes. This hurdle has now been overcome by the recent advances in the generation of human induced pluripotent stem cells (hiPSCs) where a patient's somatic cells can be reprogrammed to pluripotency and maintained indefinitely in vitro. These pluripotent cells can then be efficiently differentiated into cardiomyocytes and further studied in detail. In preliminary studies, Dr. Burridge has developed and validated a set of tools for assessing DIC in hiPSC-derived cardiomyocytes (hiPSC-CMs). Dr. Burridge has established that hiPSC-CMs, derived from patients who have developed DIC, accurately recapitulate the susceptibility phenotype in vitro. By meta-analysis of single nucleotide polymorphism (SNP) studies in patients, generated by our collaborators and others, Dr. Burridge has identified SNPs in two genes that are predicted to be highly associated with DIC (P=10-9 or 10-5). However, before any SNP can be utilized in clinical practice, its validity must be confirmed through studies linking that SNP to a mechanism for DIC. In this proposal Dr. Burridge intends to use the hiPSC-CM model to perform detailed characterization of the function of these genes identified through SNP studies in DIC. During the K99 phase Dr. Burridge will generate hiPSC lines with the two highest probability candidate SNPs (Aim 1). Dr. Burridge will then use the assays established in the pilot study to assess the effect of these SNPs on susceptibility to DIC and also isogenic hiPSC lines genetically modified to over-express or knock- down the whole genes identified to confirm the mechanism of each gene variant (Aim 2). During the R00 phase Dr. Burridge will expand this work to validate 15 additional high-risk SNP hits (Aim 3) to ultimately develop a high-throughput platform for screening cardiotoxicity of novel anthracycline analogues and cardioprotective agents in a patient-specific manner (Aim 4). The overall aim of this proposal is to use patient-specific hiPSC-CMs to help elucidate the mechanisms through which these SNPs affect cardiotoxicity. Dr. Burridge's ultimate goal is to use this information to develop novel therapeutic modalities for the prediction and prevention of chemotherapy-induced cardiotoxicity. In addition, this work will provide a foundation for future studies using patient-specific hiPSC to study the mechanism of other chemotherapeutic agents with cardiac toxicity, e.g. tyrosine kinase inhibitors, to eventually be carried out by Dr. Burridge as an independent investigator.

描述（由申请人提供）：该提案描述了一个为期五年的职业发展计划，旨在帮助候选人 Paul Burridge 博士为独立调查员的职业生涯做好准备。该项目将建立在伯里奇博士作为干细胞生物学家的背景之上，提供分子心脏病学和药物基因组学方面的专业知识。导师是斯坦福大学医学/心脏病学教授、斯坦福心血管研究所所长 Joseph Wu 博士。拟议的导师是一位在干细胞生物学方面拥有丰富专业知识的医师科学家，也是心血管疾病建模方面的专家。 K99 阶段将包括主要导师的结构化指导、与咨询委员会的补充会议、正式课程作业、具有挑战性的研究项目和职业转型计划。 阿霉素是一种成熟且高效的化疗药物，常用于治疗淋巴瘤、白血病、卵巢癌、肺癌和乳腺癌等多种癌症，但其使用受到严重副作用的限制：阿霉素会对心肌细胞产生毒性，导致心肌细胞损伤。心脏。心脏毒性的范围从无症状的左心室射血分数 (LVEF) 降低到高度症状的（III 级至 I 级）心力衰竭。约 11% 的患者会出现急性多柔比星引起的心脏毒性，高达 36% 的患者会出现长期心脏毒性副作用，这种副作用可能在治疗后长达 10 年才会显现。目前，我们无法预测哪些患者会出现心脏毒性，并且目前肿瘤学家在决定阿霉素剂量之前不会评估患者特异性的基因组数据。减少阿霉素引起的心脏毒性（DIC）的现有策略包括（i）减少剂量，可能降低化疗效果，（ii）开发心脏毒性较小的蒽环类类似物，或（iii）与心脏保护剂（例如右雷佐生）共同治疗，尽管这尚未证明有效，目前除临床试验外尚未得到美国临床肿瘤学会的认可。 填补我们关于心脏毒性机制以及如何最好地预防心脏毒性的知识方面的重大空白的一个主要障碍是，由于成人心肌细胞患者样本难以获得，并且分离和保存也存在困难，因此没有良好的人体模型。体外心肌细胞。动物模型受到动物和人类心肌细胞之间显着功能差异的限制。如今，人类诱导多能干细胞 (hiPSC) 生成方面的最新进展已经克服了这一障碍，其中患者的体细胞可以被重新编程为多能性，并在体外无限期地维持。然后这些多能细胞可以有效分化为心肌细胞并进一步研究 细节。 在初步研究中，Burridge 博士开发并验证了一套用于评估 hiPSC 衍生心肌细胞 (hiPSC-CM) 中 DIC 的工具。 Burridge 博士已经证实，源自 DIC 患者的 hiPSC-CM 可以准确地重现体外的易感性表型。通过对我们的合作者和其他人进行的患者单核苷酸多态性 (SNP) 研究进行荟萃分析，Burridge 博士确定了两个基因中的 S​​NP，预计与 DIC 高度相关（P=10-9 或 10-5） ）。然而，任何SNP在应用于临床之前，都必须通过研究证实其有效性 将该 SNP 与 DIC 机制联系起来。在本提案中，Burridge 博士打算使用 hiPSC-CM 模型对 DIC 中通过 SNP 研究确定的这些基因的功能进行详细表征。 在 K99 阶段，Burridge 博士将生成具有两个最高概率候选 SNP 的 hiPSC 系（目标 1）。然后，Burridge 博士将使用试点研究中建立的测定法来评估这些 SNP 对 DIC 易感性的影响，以及经过基因改造以过表达或敲低已确定的整个基因的同基因 hiPSC 系，以确认每个基因的机制。变体（目标 2）。在 R00 阶段，Burridge 博士将扩展这项工作，以验证 15 个额外的高风险 SNP 命中（目标 3），最终开发一个高通量平台，以针对患者的方式筛选新型蒽环类类似物和心脏保护剂的心脏毒性（目标4）。该提案的总体目标是使用患者特异性 hiPSC-CM 来帮助阐明这些 SNP 影响心脏毒性的机制。 Burridge 博士的最终目标是利用这些信息开发新的治疗方式来预测和预防化疗引起的心脏毒性。此外，这项工作将为未来使用患者特异性 hiPSC 的研究奠定基础 研究其他具有心脏毒性的化疗药物的机制，例如酪氨酸激酶抑制剂，最终由 Burridge 博士作为独立研究者进行。