Recent advances in gene editing technology have opened up new avenues for in vivo gene therapy, which holds great promise as a potential treatment method for dilated cardiomyopathy (DCM). The CRISPR-Cas13 system has been shown to be an effective tool for knocking down RNA expression in mammalian cells. PspCas13b, a type VI-B effector that can be packed into adeno-associated viruses and improve RNA knockdown efficiency, is a potential treatment for diseases characterized by abnormal gene expression. Using PspCas13b, we were able to efficiently and specifically knockdown the mutant transcripts in the AC16 cell line carrying the heterozygous human TNNT2R141W (hTNNT2R141W) mutation. We used adeno-associated virus vector serotype 9 to deliver PspCas13b with specific single guide RNA into the hTNNT2R141W transgenic DCM mouse model, effectively knocking down hTNNT2R141W transcript expression. PspCas13b-mediated knockdown significantly increased myofilament sensitivity to Ca2+, improved cardiac function, and reduced myocardial fibrosis in hTNNT2R141W DCM mice. These findings suggest that targeting genes through Cas13b is a promising approach for in vivo gene therapy for genetic diseases caused by aberrant gene expression. Our study provides further evidence of Cas13b’s application in genetic disease therapy and paves the way for future applicability of genetic therapies for cardiomyopathy.

中文翻译：

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Cas13b介导的RNA靶向治疗可缓解小鼠遗传性扩张型心肌病

基因编辑技术的最新进展为体内基因治疗开辟了新途径，这有望成为扩张型心肌病（DCM）的潜在治疗方法。CRISPR-Cas13系统已被证明是抑制哺乳动物细胞中RNA表达的有效工具。PspCas13b是一种VI-B型效应子，可以包装到腺相关病毒中并提高RNA敲低效率，是治疗以异常基因表达为特征的疾病的潜在疗法。使用 PspCas13b，我们能够高效、特异地敲低携带杂合人 TNNT2R141W (hTNNT2R141W) 突变的 AC16 细胞系中的突变转录本。我们使用腺相关病毒载体血清型 9 将具有特定单向导 RNA 的 PspCas13b 递送至 hTNNT2R141W 转基因 DCM 小鼠模型中，有效敲低 hTNNT2R141W 转录物表达。PspCas13b 介导的敲低显着增加了 hTNNT2R141W DCM 小鼠的肌丝对 Ca2+ 的敏感性，改善了心脏功能，并减少了心肌纤维化。这些发现表明，通过 Cas13b 靶向基因是治疗由异常基因表达引起的遗传性疾病的一种有前途的体内基因治疗方法。我们的研究为Cas13b在遗传病治疗中的应用提供了进一步的证据，并为未来心肌病基因疗法的应用铺平了道路。