SCGE Disease Models Studies Supplement: Cardioediting Ttntvs in a humanized mouse model

摘要 体细胞基因组编辑（SCGE）有显着的承诺，以改变我们的治疗工具箱的心脏 由于Titin截短变体（TTNtvs）而失败。然而，尽管有致病的知识， TTNtvs，它们的分子后果，以及SCGE技术，包括CRISPR基因组编辑和 由于缺乏使用人源化TTNtv动物模型的体内研究， 阻碍了临床发展。关键的知识差距仍然限制着TTNtv的SCGE开发 包括缺乏对TTNtv等位基因和心肌细胞比例的系统检查， 对临床获益进行校正，以及SCGE酶和递送系统的毒理学特征，包括 遗传毒性和免疫原性。我们的长期目标是为心力衰竭患者开发SCGE。在体外 评估SCGE编辑器和输送系统的有效性和安全性的研究，例如通过利用人类 工程心脏组织（EHT）是重要的，但它们不能取代动物模型研究的需要， 概括了心力衰竭的复杂病理生理学。根据我们全面的初步数据， 独特的试剂，包括在TTNtv EHT、人源化Ttntv小鼠和 心脏嗜性腺相关病毒;我们的1年研究将包括两个特定目的，以确定疗效， 1）TTNtv阅读框修复和2）TTN转录激活在人源化Ttntv小鼠中的安全性。 这些目标的执行将为两种SCGE方法提供编辑阈值、安全性和毒性， 我们希望TTNtv将成为更广泛的科学界的资源。

ABSTRACT Somatic cell genome editing (SCGE) has remarkable promise to transform our therapeutic toolbox for heart failure due to Titin truncation variants (TTNtvs). However, despite having the knowledge of the pathogenic TTNtvs, their molecular consequences, and SCGE technologies including CRISPR genome editing and transcriptional activation to target TTNtvs, the lack of in vivo studies using humanized TTNtv animal models has hindered clinical development. There remain key knowledge gaps limiting SCGE development for TTNtvs including the lack of a systematic examination of the proportion of TTNtv alleles and cardiomyocytes that need to be corrected for clinical benefit, and the toxicology profile of SCGE enzymes and delivery systems including genotoxicity and immunogenicity. Our long-term goal is to develop SCGE for heart failure patients. While in vitro studies to evaluate the efficacy and safety of SCGE editors and delivery systems such as by utilizing human engineered heart tissues (EHTs) are important, they do not replace the need for animal model studies that recapitulate the complex pathophysiology of heart failure. Guided by our comprehensive preliminary data and unique reagents including SCGE components functionally validated in TTNtv EHTs, humanized Ttntv mice and cardiotropic adeno-associated viruses; our 1-year study will include two Specific Aims to determine efficacy and safety of 1) TTNtv reading frame repair and 2) TTN transcriptional activation in a humanized Ttntv mouse. Execution of these Aims will provide editing thresholds, safety, and toxicity for two SCGE approaches to treat TTNtvs that we expect will serve as a resource for the broader scientific community.