Precise genomic editing of pathogenic mutations in RBM20 rescues dilated cardiomyopathy

Nov 23, 2022Science translational medicine

Accurate gene editing of harmful RBM20 mutations improves dilated heart disease

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Abstract

Adenine base editing achieved 92% efficiency in correcting the p.R634Q mutation in human heart cells.

Mutations in the RNA binding motif protein 20 are commonly linked to familial dilated cardiomyopathy.
Many of these mutations cause mis-localization of RBM20, leading to abnormal gene splicing and the formation of ribonucleoprotein granules.
Correction of the p.R634Q mutation normalized splicing of cardiac genes and restored RBM20 localization in heart cells.
A separate prime editing strategy achieved 40% efficiency in correcting the p.R636S mutation.
In mutant mice, delivery of adenine base editing components improved cardiac function and extended lifespan.
RNA sequencing showed that editing rescued the abnormal gene expression profile associated with the disease.

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Mutations in RNA binding motif protein 20 () are a common cause of familial dilated cardiomyopathy (DCM). Manymutations 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 correcthuman 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 themutation in iPSCs and observed A-to-C editing at 40% efficiency. To evaluate the potential of ABE for DCM treatment, we also createdmutant 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. RBM20RBM20RBM20RBM20Rbm20R634Q R636S R636Q

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Precise genomic editing of pathogenic mutations in RBM20 rescues dilated cardiomyopathy

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