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药物之一，但在治疗多种癌症方面非常有效，并且仍然在 70% 的癌症中使用 儿童癌症治疗。然而，其用途受到心脏毒性的限制，大约 65% 的儿童癌症长期幸存者会出现心脏毒性。儿童比成人更容易受到这种危及生命的副作用的影响。我们发现了两种与阿霉素心脏毒性风险显着改变相关的基因变异。一种保护性变体位于基因 SLC28A3（一种抗癌药物转运蛋白）中，一种风险变体位于 RARG（一种改变其他基因表达的核受体和转录因子）中。尽管这些研究代表了利用患者遗传学指导阿霉素使用（药物基因组学）的进步，但这些基因变异的真正效果还远未得到证实。必须满足的其他标准包括 (1) 在其他患者队列中得到确认； (2) 使用模型系统验证基因变异会改变心脏毒性； (3) 验证其作用机制（例如药物转运蛋白的功能丧失变化是否会导致细胞内药物水平降低和毒性降低）； (4) 证明将变体恢复为正常（野生型）基因可以挽救改变的毒性作用。 患者来源的 hiPSC-CM（人诱导多能干细胞来源的心肌细胞）代表了一项新技术，已应用于了解疾病机制和筛选药物毒性。尽管 hiPSC-CM 不能复制成熟心肌细胞的所有方面，但我们发现，与来自没有心脏毒性的患者的细胞相比，来自患有阿霉素心脏毒性的患者的 hiPSC-CM 显示出更多的阿霉素损伤。我们假设 hiPSC-CM 代表了一个模型平台，用于研究基因变异在调节阿霉素心脏毒性方面的有效性和机制。目标 1：开发具有 SLC28A3 基因变异的 hiPSC 系，并检查对阿霉素心脏毒性的敏感性降低。细胞将来自 (a) 直接来自具有基因变异的患者； (b) 通过在对照 hiPSC 系中遗传诱导相同的基因改变。阿霉素毒性将通过细胞功能和活力测定来量化。目标 2：开发具有 RARG 候选基因变体的 hiPSC 系，并检查体外阿霉素心脏毒性的敏感性是否增加。目标 3：探索每种变体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 系中增加或减少； (b) 在来自携带每种变异的患者的 hiPSC-CM 中，该变异将恢复为正常（野生型）。 (c) 然后我们将探讨每种变异影响阿霉素心脏毒性的具体机制。目标 4：利用我们的平台来验证其他高风险命中。我们将复制上述研究或其他变体，通过满足高概率和可复制性的严格定义来选择。

项目成果

Patient-Specific Pluripotent Stem Cells in Doxorubicin Cardiotoxicity: A New Window Into Personalized Medicine.

阿霉素心脏毒性中的患者特异性多能干细胞：个性化医疗的新窗口。

• 发表时间: 2014-12-01
• 影响因子: 0.9
• 作者: Bernstein, Daniel;Burridge, Paul
• 通讯作者: Burridge, Paul

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

RARG 中的编码变异使儿童癌症对蒽环类药物诱导的心脏毒性具有易感性。

• 发表时间: 2015-09
• 影响因子: 30.8
• 作者: Aminkeng, Folefac;Bhavsar, Amit P;Visscher, Henk;Rassekh, Shahrad R;Li, Yuling;Lee, Jong W;Brunham, Liam R;Caron, Huib N;van Dalen, Elvira C;Kremer, Leontien C;van der Pal, Helena J;Amstutz, Ursula;Rieder, Michael J;Bernstein, Daniel;Carleto
• 通讯作者: Carleto

Induced Pluripotent Stem Cell-Derived Cardiomyocytes: A Platform for Testing For Drug Cardiotoxicity.

诱导多能干细胞衍生的心肌细胞：药物心脏毒性测试平台。

• 发表时间: 2017-09
• 影响因子: 0.9
• 作者: Bernstein; Daniel
• 通讯作者: Daniel

Anthracycline Cardiotoxicity: Worrisome Enough to Have You Quaking?

蒽环类药物心脏毒性：令人担忧到让您发抖吗？

• 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

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

沙利度胺治疗可预防大鼠主动脉移植后的慢性移植排斥——一项实验研究。

• 发表时间: 2017-11
• 作者: Miller, Katharine K;Wang, Dong;Hu, Xiaomeng;Hua, Xiaoqin;Deuse, Tobias;Neofytou, Evgenios;Renne, Thomas;Velden, Joachim;Reichenspurner, Hermann;Schrepfer, Sonja;Bernstein, Daniel
• 通讯作者: Bernstein, Daniel