Elucidating anti-angiogenic tyrosine kinase inhibitor-induced vascular dysfunction

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PROJECT SUMMARY Although tyrosine kinase inhibitors (TKIs) are highly potent in treating malignancies, >60% of them have been reported to lead to adverse cardiovascular outcomes. Moreover, anti-angiogenic TKIs, such as sunitinib, preferentially induce microvascular toxicity followed by cardiac dysfunction. However, our knowledge of the underlying pathogenic mechanisms of TKI-induced vascular toxicity (TKI-VT) has been hampered partially by the limited access to human diseased vascular tissues for molecular and cellular analysis. As such, no effective strategies have been developed to prevent or treat these otherwise life-threatening cardiovascular complications. This proposed project will leverage patient-specific induced pluripotent stem cell (iPSC)-derived cardiac pericytes (PCs) and endothelial cells (ECs) to understand the molecular and cellular basis of TKI-VT and discover personalized therapies for cancer patients who are suffering TKI-induced cardiovascular disease. In Aim 1, both monoculture and vessel-on-chip (VoC) coculture systems will be employed to characterize sunitinib-induced cell type-specific cytotoxicity profiles and aberrant cellular crosstalk between iPSC-PCs and iPSC-ECs that can contribute to TKI-VT. Overlapping upregulated and downregulated differentially expressed genes (DEGs) triggered by sunitinib in both cell types will serve as the candidate genes for large-scale druggable target screens. In Aim 2, CRISPR interference/activation (CRISPRi/a) survival screens will be performed in sunitinib-treated iPSC-PCs using a customized lentiviral sgRNA library targeting the overlapping DEGs identified in Aim 1. Top 10 hit genes in CRISPRi and CRSIPRa machineries will be subjected to structure-based virtual screens (SBVS) to discover candidate compounds that can mitigate TKI-VT phenotypes in iPSC-VoCs. In Aim 3, 3D iPSC- engineered vascular tissues (EVTs) and a mouse model will be used to validate the mitigation efficacy of candidate compounds on TKI-VT in a more physiological setting. The ex vivo plasma proteome generated from human whole blood-perfused 3D iPSC-EVTs will be correlated with those identified in patients to discover reliable disease-relevant biomarkers in predicting individual cancer patients’ susceptibility to TKI-VT. The research and career development training plans during the K99 phase, under the mentorship of Drs. Wu and Ky, as well as an expert interdisciplinary advisory committee, will provide Dr. Shen with advanced knowledge in stem cell biology, vascular biology, cardio-oncology, CRISPR technology, and bioengineering. The development of CRISPRi/a and SBVS screen platforms (K99) and 3D iPSC-EVTs (R00) will enable him to conduct disease modeling and drug discovery research in cardio-oncology specifically and vascular disease in general. The new skills and experience gained during this K99/R00 career development award, combined with Dr. Shen’s prior expertise in vascular biology, will facilitate his transition to an independent career conducting basic and translational research in cardio-oncology with a particular focus on the vascular aspect.