hiPSC-Cardiomyocytes to Screen Variants Predictive of Doxorubicin Cardiotoxicity

hiPSC-心肌细胞筛选预测阿霉素心脏毒性的变异体

• 批准号: 8909180
• 负责人: Daniel Bernstein
• 金额: $ 23.21万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-08 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8909180
• 关键词: Adult; Adverse effects; Affect; Anthracyclines; Antineoplastic Agents; Antineoplastic Protocols; Biological Assay; Biological Models; Calcium; Candidate Disease Gene; Cardiac; Cardiac Myocytes; Cardiotonic Agents; Cardiotoxicity; Cardiovascular Diseases; Cell Line; Cell Survival; Cell physiology; Cells; Chemotherapy-Oncologic Procedure; Child; Childhood; Childhood Cancer Treatment; Clinical; Data; Development; Disease; Dose; Doxorubicin; Drug toxicity; Echocardiography; Exposure to; Gene Mutation; Genes; Genetic; Genetic Polymorphism; Genomic approach; Genomics; Glucuronosyltransferase; Health; Heart; Heart failure; Human; In Vitro; Incidence; Lead; Life; Linkage Disequilibrium; Long-Term Survivors; Malignant Childhood Neoplasm; Malignant Neoplasms; Methods; Mitochondria; Modeling; Molecular; Nuclear Receptors; Nucleosides; Odds Ratio; Oncologist; Patients; Pharmaceutical Preparations; Pharmacogenomics; Phase; Physicians; Population; Predisposition; Probability; Production; Protocols documentation; RARG gene; Report (document); Reporting; Retinoic Acid Receptor; Risk; Survival Rate; Testing; Toxic effect; Transcription Coactivator; Translations; Validation; Variant; abstracting; analog; base; cell injury; childhood cancer survivor; clinical care; clinical practice; cohort; drug efficacy; follow-up; genetic variant; genome wide association study; high risk; improved; induced pluripotent stem cell; loss of function; malignant breast neoplasm; meetings; mortality; new technology; nuclease; oncology; pharmacogenetic testing; predictive modeling; risk variant; screening; tool; transcription factor

DESCRIPTION (provided by applicant): Doxorubicin, although one of the oldest anti-cancer agents, is highly effective in treating a wide range of cancers and is still utilized in 70% of all childhood cancer treatments. However, its utility is limited by its cardiac toxicity, occurring in p to 65% of long-term survivors of childhood cancer. Children are more susceptible to this life-threatening side effect than adults. We have found two genetic variants associated with dramatically altered risk of doxorubicin cardiotoxicity. One protective variant is in the gene SLC28A3, an anti-cancer drug transporter and one risk variant is in RARG, a nuclear receptor and transcription factor that alters expression of other genes. Although these studies represent an advance in using a patient's genetics to guide doxorubicin usage (pharmacogenomics), the true effect of these gene variants is far from proven. Additional criteria that must be met include (1) Confirmation in other patient cohorts; (2) Validation, using a model system, that the gene variant alters cardiotoxicity; (3) Validation of a mechanism for its effects (e.g. does a loss-of-function change in a drug transporter lead to decreased intracellular drug levels and decreased toxicity); and (4) Demonstration that reversion of the variant to the normal (wild-type) gene rescues the altered toxicity effect. Patient-derived hiPSC-CMs (human induced pluripotent stem cell-derived cardiomyocytes) represent a novel technology which has been applied to understanding disease mechanisms and to screening drugs for toxicity. Although hiPSC-CMs do not replicate all aspects of mature cardiomyocytes, we show that hiPSC-CMs from patients who have had doxorubicin cardiotoxicity show increased doxorubicin damage compared to cells from patients without cardiotoxicity. We hypothesize that hiPSC-CMs represent a model platform for studying the validity and mechanisms of gene variants in regulating doxorubicin cardiotoxicity. Aim 1: To develop hiPSC lines with the gene variant in SLC28A3 and examine for decreased susceptibility to doxorubicin cardiotoxicity. Cells will be derived (a) directly from patients with the gene variant; and (b) by genetically inducing the same gene alterations in a control hiPSC line. Doxorubicin toxicity will be quantified by assays of cell function and viabilit. Aim 2: To develop hiPSC lines with the candidate gene variant in RARG and examine for increased susceptibility to in vitro doxorubicin cardiotoxicity. Aim 3: To explore the mechanism(s) by which each variant alters doxorubicin cardiotoxicity. (a) Expression of each candidate gene will be increased or decreased in a control hiPSC-CM line; (b) The variant will be reverted to normal (wild-type) in hiPSC-CMs from patients with each variant (c) We will then explore the specific mechanisms by which each variant affects doxorubicin cardiotoxicity. Aim 4: To utilize our platform to validate additional high risk hits. We will duplicate the above studies or other variants, chosen by meeting a strict definition of high probability and replicability.

描述（由申请人提供）：阿霉素虽然是最古老的抗癌剂之一，但在治疗广泛的癌症方面非常有效，并且仍在所有人中的70％中使用 儿童癌症治疗。但是，其效用受心脏毒性的限制，在P中占据了儿童癌的长期幸存者的65％。与成年人相比，儿童更容易受到这种威胁生命的副作用。我们发现了两种与阿霉素心脏毒性风险发生巨大改变有关的遗传变异。一种保护性变体是基因SLC28A3，一种抗癌药物转运蛋白，一个风险变体是RARG，RARG是一种改变其他基因表达的核受体和转录因子。尽管这些研究代表了使用患者的遗传学指导阿霉素使用率（药物基因组学）的进步，但这些基因变异的真正效果远非证明。必须满足的其他标准包括（1）其他患者队列中的确认； （2）使用模型系统验证基因变体会改变心脏毒性； （3）验证其作用的机制（例如，药物转运蛋白的功能丧失会导致细胞内药物水平降低并降低毒性）； （4）证明变体向正常（野生型）基因的逆转挽救了改变的毒性效应。 患者衍生的HIPSC-CMS（人类诱导的多能干细胞衍生的心肌细胞）代表了一种新技术，该技术已用于理解疾病机制和筛查毒性药物。尽管HIPSC-CMS并未复制成熟心肌细胞的所有方面，但我们表明，与没有心脏毒性的患者相比，患有阿霉素心脏毒性的患者的HIPSC-CMS显示了阿霉素心脏毒性的损害增加。我们假设HIPSC-CMS代表了研究基因变体在调节阿霉素心脏毒性中的有效性和机制的模型平台。目标1：在SLC28A3中开发具有基因变体的HIPSC系，并检查对阿霉素心脏毒性的敏感性降低。细胞将直接从具有基因变异的患者中得出（a）； （b）通过遗传诱导对照hipsc系中的相同基因改变。阿霉素的毒性将通过细胞功能和金蛋白的测定来量化。目标2：与RARG中的候选基因变体开发HIPSC线，并检查对体外阿霉素心脏毒性的敏感性增加。目标3：探索每个变体都会改变阿霉素心脏毒性的机制。 （a）每个候选基因的表达将在对照HIPSC-CM线中增加或减少； （b）在每个变体（c）的患者的HIPSC-CM中，该变体将被恢复为正常（野生型）（野生型），然后我们将探索每个变体都会影响阿霉素心脏毒性的特定机制。目标4：利用我们的平台验证额外的高风险命中。我们将复制上述研究或其他变体，这是通过符合高概率和可复制性的严格定义而选择的。

项目成果

Mitochondrial remodeling: Rearranging, recycling, and reprogramming.

• DOI: 10.1016/j.ceca.2016.04.006
• 发表时间: 2016-08
• 期刊: Cell calcium
• 影响因子: 4
• 作者: Gottlieb RA;Bernstein D
• 通讯作者: Bernstein D

Thalidomide treatment prevents chronic graft rejection after aortic transplantation in rats - an experimental study.

• DOI: 10.1111/tri.13004
• 发表时间: 2017-11
• 期刊: Transplant international : official journal of the European Society for Organ Transplantation
• 作者: Miller KK;Wang D;Hu X;Hua X;Deuse T;Neofytou E;Renne T;Velden J;Reichenspurner H;Schrepfer S;Bernstein D
• 通讯作者: Bernstein D

A coding variant in RARG confers susceptibility to anthracycline-induced cardiotoxicity in childhood cancer.

• DOI: 10.1038/ng.3374
• 发表时间: 2015-09
• 期刊: Nature genetics
• 影响因子: 30.8
• 作者: Aminkeng F;Bhavsar AP;Visscher H;Rassekh SR;Li Y;Lee JW;Brunham LR;Caron HN;van Dalen EC;Kremer LC;van der Pal HJ;Amstutz U;Rieder MJ;Bernstein D;Carleton BC;Hayden MR;Ross CJ;Canadian Pharmacogenomics Network for Drug Safety Consortium
• 通讯作者: Canadian Pharmacogenomics Network for Drug Safety Consortium

Anthracycline Cardiotoxicity: Worrisome Enough to Have You Quaking?

• DOI: 10.1161/circresaha.117.312395
• 发表时间: 2018-01-19
• 期刊: Circulation research
• 影响因子: 20.1
• 作者: Bernstein D