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: 9330916
• 负责人: Paul W. Burridge
• 金额: $ 24.63万
• 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/9330916
• 关键词: ABCB1 gene; ASBT protein; Acute; Adult; Adverse effects; Advisory Committees; Affect; Aftercare; Alpha Cell; American Society of Clinical Oncology; Animal Model; Animals; Anthracyclines; Binding; Biological Assay; 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; Heart; Heart failure; Hemochromatosis; Human; Image; 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; Population Study; Predisposition; Prevention; Probability; Program Development; RARG gene; Research Personnel; Research Project Grants; Respiration; Risk; Role; Sampling; Scientist; Signal Transduction; Single Nucleotide Polymorphism; Somatic Cell; 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; 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)：本提案描述了一个为期五年的职业发展计划，为候选人保罗·伯里奇博士作为一名独立调查员的职业生涯做准备。该项目将以布里奇博士作为干细胞生物学家的背景为基础，提供分子心脏病学和药物基因组学方面的专业知识。导师是医学/心脏病学教授、斯坦福大学斯坦福心血管研究所所长Joseph Wu博士。被推荐的导师是一位在干细胞生物学方面拥有丰富专业知识的内科科学家，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有组织的指导、与咨询委员会的补充会议、正式的课程作业、具有挑衅性的研究项目和职业过渡计划。阿霉素是一种久负盛名的高效化疗药物，通常用于治疗淋巴瘤、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：阿霉素对心肌细胞产生毒性，对心脏造成损害。心脏毒性可从无症状的左心室射血分数(LVEF)降低到高度症状性(III级至I级)心力衰竭。阿霉素引起的急性心脏毒性在~11%的患者中发生，而在多达36%的患者中观察到长期的心脏毒性副作用，这种副作用可在治疗后长达10年出现。目前，我们无法预测哪些患者会出现心脏毒性，目前，肿瘤学家在决定阿霉素的剂量之前，不会评估患者特有的基因组数据。现有的减少阿霉素引起的心脏毒性(DIC)的策略包括(I)减少剂量，潜在地降低化疗效果，(Ii)开发心脏毒性较小的蒽环类类似物，或(Iii)与心脏保护剂如右氧氟沙星联合治疗，尽管这尚未被证明有效，目前也没有得到美国临床肿瘤学会临床试验以外的认可。在填补我们对心脏毒性机制和如何最好地预防的认识上的重大空白的一个主要障碍是，由于无法获得成人心肌细胞患者的样本，以及在体外分离和维护心肌细胞的困难，没有良好的人体模型。动物模型受到动物和人类心肌细胞之间显著功能差异的限制。这一障碍现在已经被人类诱导多能干细胞(HiPSCs)的产生的最新进展所克服，在这种细胞中，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保持。然后，这些多能细胞可以有效地分化为心肌细胞，并在 细节。在初步研究中，伯里奇博士开发并验证了一套工具，用于评估HiPSC来源的心肌细胞(hiPSC-CMS)的DIC。伯里奇博士已经证实，来自患有DIC患者的hiPSC-CMS在体外可以准确地概括易感表型。通过对我们的合作者和其他人在患者中进行的单核苷酸多态(SNP)研究的荟萃分析，布里奇博士发现了两个基因中的SNP，这两个基因被预测与DIC高度相关(P=10-9或10-5)。然而，在任何SNP可以用于临床实践之前，其有效性必须通过研究来证实。 将SNP与DIC机制联系起来。在这项建议中，伯里奇博士打算使用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的表征。在K99阶段，伯里奇博士将产生具有两个最高概率候选SNP的HiPSC系(目标1)。然后，Burbridge博士将使用在初步研究中建立的分析方法来评估这些SNPs对DIC易感性的影响，并评估等基因HiPSC系的基因改造，以过度表达或敲除已确定的全部基因，以确认每个基因变异的机制(目标2)。在R00阶段，Burbridge博士将扩大这项工作，以验证另外15个高风险SNP命中(Aim 3)，最终开发一个高通量平台，以患者特有的方式筛选新型蒽环类类似物和心脏保护剂的心脏毒性(Aim 4)。这项建议的总体目标是使用患者特有的HiPSC-CMS来帮助阐明这些SNP影响心脏毒性的机制。伯里奇博士的最终目标是利用这些信息开发新的治疗方式，以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有的HiPSC进行研究提供基础 研究其他具有心脏毒性的化疗药物的机制，如酪氨酸激酶抑制剂，最终由Burbridge博士作为独立研究员进行。