SCGE Disease Models Studies Supplement: Cardioediting Ttntv's in a Humanized Mouse Model

摘要 体细胞基因组编辑(SCGE)有望改变我们的心脏治疗工具箱 由于TITIN截断变量(TTNtvs)而失败。然而，尽管知道致病原因 TTNtv及其分子后果，以及包括CRISPR基因组编辑和SCGE技术在内的SCGE技术 转录激活靶向TTNtv，缺乏使用人源化TTNtv动物模型的体内研究 阻碍了临床的发展。仍然存在限制TTNtv的SCGE发展的关键知识差距 包括缺乏对TTNTV等位基因和心肌细胞所需的TTNtv等位基因比例的系统检查 为了临床利益而更正，以及SCGE酶和递送系统的毒理学概况，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。而在体外 评价SCGE编辑和传递系统的有效性和安全性的研究，如利用人类 工程心脏组织(EHTS)很重要，它们不能取代对动物模型研究的需要 重述心力衰竭的复杂病理生理学。在我们全面的初步数据和 独特的试剂包括SCGE组件在TTNtv EHTS、人源化Ttntv小鼠和 我们为期一年的研究将包括两个特定的目标，以确定疗效和 人源化Ttntv小鼠中TTNtv阅读框架修复和TTN转录激活的安全性。 这些目标的实施将为两种SCGE治疗方法提供编辑阈值、安全性和毒性 我们期待的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.