Precise genomic editing of pathogenic mutations in RBM20 rescues dilated cardiomyopathy

Mutations in RNA binding motif protein 20 ( RBM20 ) are a common cause of familial dilated cardiomyopathy (DCM). Many RBM20 mutations cluster within an arginine/serine-rich (RS-rich) domain, which mediates nuclear localization. These mutations induce RBM20 mis-localization to form aberrant ribonucleoprotein (RNP) granules in the cytoplasm of cardiomyocytes and abnormal alternative splicing of cardiac genes, contributing to DCM. We used adenine base editing (ABE) and prime editing (PE) to correct pathogenic p.R634Q and p.R636S mutations in the RS-rich domain in human isogenic induced pluripotent stem cell (iPSC)–derived cardiomyocytes. Using ABE to correct RBM20 R634Q human iPSCs, we achieved 92% efficiency of A-to-G editing, which normalized alternative splicing of cardiac genes, restored nuclear localization of RBM20, and eliminated RNP granule formation. In addition, 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 also created Rbm20 R636Q mutant mice. Homozygous (R636Q/R636Q) mice developed severe cardiac dysfunction, heart failure, and premature death. Systemic delivery of ABE components containing ABEmax-VRQR-SpCas9 and single-guide RNA by adeno-associated virus serotype 9 in these mice restored cardiac function as assessed by echocardiography and extended life span. As seen by RNA sequencing analysis, ABE correction rescued the cardiac transcriptional profile of treated R636Q/R636Q mice, compared to the abnormal gene expression seen in untreated mice. These findings demonstrate the potential of precise correction of genetic mutations as a promising therapeutic approach for DCM.