Genomic Prediction of Doxorubicin-Induced Cardiotoxicity

项目摘要 应用多柔比星治疗约60%有转移实体的儿童癌症患者 肿瘤(肉瘤)、母细胞瘤、白血病和淋巴瘤。阿霉素的治疗因其复杂性而变得复杂 公认的心脏毒性副作用，影响大约16%的儿科患者，可导致心脏 衰竭需要心脏移植，限制了阿霉素的临床应用。尽管50多年来 目前，在这一领域的研究中，预测或预防心脏毒性的潜力仍然很小。那里 显然需要新颖和创新的方法来克服这一障碍。候选基因关联 研究和全基因组关联研究发现了许多单核苷酸多态 (SNPs)与阿霉素诱导的心脏毒性(DIC)统计相关，但实验验证 其中，由于分离和体外培养人心肌细胞的困难，这些SNPs尚不可行。 在我们最近的工作中，我们显示了患者特异性的人类诱导多能干细胞来源 心肌细胞(HiPSC-CM)是患者发生DIC可能性的有效预测因子，证实 这是DIC第一次有基因组基础。尽管Gwas已被证明是一种强大的方法 告知这些基因组基础，它检测相关性而不是因果关系，识别出的SNPs通常失败 将在随后的研究中复制。在这里，我们假设HiPSC-CMS可以用于三种不同的 研究与DIC相关的遗传变异的方式：第一，发现新的预测性SNPs；第二， 验证SNPs；第三，检查调节途径并确定特定的基因型别 心脏保护方法学。在目标1中，我们将招募100名接触过 阿霉素，并评估患者来源的HiPSC-CM对阿霉素的体外反应，以验证我们以前的 在具有不同生物协变量的大型儿科队列中的发现，以验证该工具的力量。在目标2中，我们 将使用这100个患者特定的株系来识别药物反应差异表达数量性状基因座 (DeQTL)，评估生物协变量，如剂量、年龄、性别、SF和癌症诊断 加在一起。然后我们将通过基因组编辑来验证这些变异，并机械地检查路径 引起DIC的易感性集中在在心肌病、心肌保护、 和阿霉素代谢。在目标3中，我们将询问&gt；40现有DIC SNP的严密性和重复性 研究中，使用CRISPR/Cas9编辑控制等基因HiPSC系中的感兴趣基因，然后评估响应 HPSC-CM对阿霉素的作用。然后，我们将使用上面的发现来发现/重新利用基因组信息 预防DIC的心脏保护药物是一种针对特定基因的药物。总之，这项工作将为我们提供 患者经历DIC并为临床提供1个完全经过人类验证的SNP数据的遗传学基础 应用和2，新的心脏保护药物，以减轻DIC。

Project Summary The anthracycline doxorubicin used in approximately 60% of pediatric cancer patients with metastatic solid tumors (sarcomas), blastomas, leukemia, and lymphoma. Treatments using doxorubicin are complicated by its well-established cardiotoxic side effect, which affects approximately 16% of pediatric patients, can lead to heart failure requiring heart transplant, and limits doxorubicin’s clinical utilization. Despite more than 50 years of research in this field, there is still, at present, little potential for either predicting or preventing cardiotoxicity. There is an obvious need for novel and innovative approaches to overcome this hurdle. Candidate gene association studies and genome–wide association studies (GWAS) have identified many single nucleotide polymorphisms (SNPs) that are statistically correlated with doxorubicin–induced cardiotoxicity (DIC), yet experimental validation of these SNPs has not been feasible due to the difficulty in isolating and culturing human cardiomyocytes in vitro. In our recent work, we showed that patient–specific human induced pluripotent stem cell–derived cardiomyocytes (hiPSC–CM) are efficient predictors of a patient’s likelihood of developing DIC, confirming for the first time that there is a genomic basis to DIC. Although GWAS has proven to be a powerful methodology for informing such genomic bases, it detects correlation rather than causation, and identified SNPs commonly fail to be replicated in subsequent studies. Here, we hypothesize that hiPSC-CMs can be utilized in three different modalities to study genetic variants associated with DIC: firstly, to discover novel predictive SNPs; secondly, to validate SNPs; and thirdly, to examine the modulated pathways and determine genotype-specific cardioprotective methodologies. In Aim 1, we will recruit 100 pediatric cancer patients who were exposed to doxorubicin and assess the response of patient-derived hiPSC-CM to doxorubicin in vitro to validate our previous findings in a large pediatric cohort with diverse biological covariates to verify the power of this tool. In Aim 2, we will use these 100 patient-specific lines to identify drug response differential expression quantitative trait loci (deQTL), assessing biological covariates such as dose, age, sex, SF, and cancer diagnosis both individually and combined. We will then validate these variants with genome editing, and mechanistically examine pathways causative to DIC susceptibility concentrating on genes with known roles in cardiomyopathy, cardioprotection, and doxorubicin metabolism. In Aim 3, we will interrogate the rigor and reproducibility of >40 existing DIC SNP studies, using CRISPR/Cas9 to edit the gene of interest in control isogenic hiPSC lines then assess the response of hiPSC-CM to doxorubicin. We will then use the discoveries above to discover/repurpose genome-informed cardioprotective drugs to prevent DIC in a genotype-specific manner. In summary, this work will deliver us the genetic rationale for why patients experience DIC and provide 1, fully human validated SNP data for clinical application, and 2, novel cardioprotective drugs to attenuate DIC.