CRISPR/Cas9 Screens Implicate RARA and SPNS1 in Doxorubicin Cardiotoxicity

BACKGROUND: Doxorubicin (DOX) causes cardiotoxicity and heart failure in a significant fraction of patients, but the molecular etiology is poorly understood. OBJECTIVES: We adopted a functional genomics-based approach to probe the genetic basis for DOX-induced cardiotoxicity in an exhaustive and agnostic manner. METHODS: Genome-wide and targeted CRISPR/Cas9 screens were performed in immortalized cardiomyocytes and induced pluripotent stem cell-derived cardiomyocytes to identify genetic modifiers of DOX-induced cardiotoxicity. RESULTS: Our first screen revealed that loss of the Retinoic Acid Receptor Alpha gene (RARA) increased DOX-induced cell death. Conversely, pharmacological activation of RARA protein with tamibarotene reduced DOX-induced toxicity. RNA-Seq analysis showed that whereas DOX broadly suppressed expression of metabolic and mitochondrial genes, tamibarotene mitigated this effect. In a second screen, we interrogated processes involved in DOX uptake, transport, and efflux. Loss of lysosome homeostasis, exemplified by SPNS lysolipid transporter 1 (SPNS1) deficiency, led to DOX hyperaccumulation, suppression of autophagy, increased DNA damage, and increased cell death. In contrast, ribosome loss-of-function and nutrient deprivation significantly reduced DOX accumulation and toxicity. CONCLUSIONS: Our study identified numerous drug-gene interactions that illuminate mechanisms underlying DOX-induced cardiotoxicity and provide a technical framework for future functional genomics screens to nominate therapeutic targets and genetic biomarkers.