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.

Public Health Relevance

Doxorubicin is a highly effective chemotherapy drug the use of which is complicated by its well-established cardiotoxic side effect, with little potential for predicting which patients will be adversely affected. In this proposal, we will investigate the application of human induced pluripotent stem cell-derived cardiomyocytes to validate genetic markers of doxorubicin-induced cardiotoxicity and to establish the molecular mechanisms by which they alter the risk of this complication. This knowledge should be of value in prediction of doxorubicin- induced cardiotoxicity and this platform useful for studying means of prevention.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Transition Award (R00)
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Special Emphasis Panel (NSS)
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Adhikari, Bishow B
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Northwestern University at Chicago
Schools of Medicine
United States
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Sharma, Arun; Zhang, Yuan; Buikema, Jan W et al. (2018) Stage-specific Effects of Bioactive Lipids on Human iPSC Cardiac Differentiation and Cardiomyocyte Proliferation. Sci Rep 8:6618
Sharma, Arun; Burridge, Paul W; McKeithan, Wesley L et al. (2017) High-throughput screening of tyrosine kinase inhibitor cardiotoxicity with human induced pluripotent stem cells. Sci Transl Med 9:
Matsa, Elena; Burridge, Paul W; Yu, Kun-Hsing et al. (2016) Transcriptome Profiling of Patient-Specific Human iPSC-Cardiomyocytes Predicts Individual Drug Safety and Efficacy Responses In Vitro. Cell Stem Cell 19:311-25
Burridge, Paul W; Li, Yong Fuga; Matsa, Elena et al. (2016) Human induced pluripotent stem cell-derived cardiomyocytes recapitulate the predilection of breast cancer patients to doxorubicin-induced cardiotoxicity. Nat Med 22:547-56
Magdy, Tarek; Burmeister, Brian T; Burridge, Paul W (2016) Validating the pharmacogenomics of chemotherapy-induced cardiotoxicity: What is missing? Pharmacol Ther 168:113-125
Burridge, Paul W; Diecke, Sebastian; Matsa, Elena et al. (2016) Modeling Cardiovascular Diseases with Patient-Specific Human Pluripotent Stem Cell-Derived Cardiomyocytes. Methods Mol Biol 1353:119-30
Sharma, Arun; Li, Guang; Rajarajan, Kuppusamy et al. (2015) Derivation of highly purified cardiomyocytes from human induced pluripotent stem cells using small molecule-modulated differentiation and subsequent glucose starvation. J Vis Exp :
Burridge, Paul W; Sharma, Arun; Wu, Joseph C (2015) Genetic and Epigenetic Regulation of Human Cardiac Reprogramming and Differentiation in Regenerative Medicine. Annu Rev Genet 49:461-84
Burridge, Paul W; Holmström, Alexandra; Wu, Joseph C (2015) Chemically Defined Culture and Cardiomyocyte Differentiation of Human Pluripotent Stem Cells. Curr Protoc Hum Genet 87:21.3.1-15
Burridge, Paul W; Matsa, Elena; Shukla, Praveen et al. (2014) Chemically defined generation of human cardiomyocytes. Nat Methods 11:855-60

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