Abstract 11164: Precise Genomic Correction of RBM20 Mutations Rescues Dilated Cardiomyopathy

Abstract

Introduction: Despite current medical advancements, effective treatment for familial cardiomyopathies remains challenging. Mutations in RNA binding motif protein 20 ( RBM20 ) are a common cause of dilated cardiomyopathy (DCM). Many RBM20 mutations cluster within the arginine/serine rich (RS-rich) domain, which mediates nuclear localization. These mutations induce RBM20 mis-localization and abnormal alternative splicing of cardiac genes. Importantly, the formation of aberrant ribonucleoprotein (RNP) granules in the cytoplasm of cardiomyocytes may contribute to DCM. Hypothesis: Precise genomic editing using base editing (BE) or prime editing (PE) corrects the mutations and rescues the pathophysiology of RBM20-related DCM. Methods: We used adenine base editing (ABE) or PE to correct the pathogenic p.R634Q (c.1901 G>A) and p.R636S (c.1906 C>A) mutations in the RS-rich domain of RBM20 in human isogenic induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM). Additionally, using a humanized Rbm20 mutant mouse (R636Q/R636Q), we assessed in vivo ABE correction for DCM treatment. Results: Correcting RBM20 R634Q human iPSC by ABE using ABEmax-VRQR-SpCas9 and sgRNA components, we showed 89% efficiency of A to G editing, recovery of alternative splicing of cardiac genes, restoration of nuclear localization of RBM20 and elimination of RNP granule formation. Additionally, we developed a PE strategy to correct the RBM20 R636S mutation in iPSCs and observed A to C editing at 40% efficiency. To evaluate the potential of ABE for DCM treatment, we systemically delivered ABE components by adeno-associated virus in postnatal Rbm20 mutant mice (R636Q/R636Q). Uncorrected R636Q/R636Q mice exhibited severe heart failure and premature death by 3-months of age. In contrast, R636Q/R636Q mice receiving systemic ABE components showed a substantial improvement of LV function and an increase in life span. As seen by RNA-seq analysis, ABE correction rescued the transcriptional profile of treated R636Q/R636Q mice, while untreated mice showed altered cardiac gene expression. Conclusions: Precise gene editing technologies provide an innovative approach to correct disease-causing genomic mutations in cardiovascular diseases.