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Human cardiac microtissues with innate immune sensing to study adverse consequences of genome editing

具有先天免疫传感的人类心脏微组织用于研究基因组编辑的不良后果

基本信息

批准号：
10245264
负责人：
John Travis Hinson
金额：
$ 73.8万
依托单位：
UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-08-01 至 2023-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10245264
关键词：
3-Dimensional Adaptive Immune System Address Adverse effects Affect Alleles Architecture Arrhythmia Autologous Automobile Driving Biological Assay Biological Process Biomechanics Biomimetics CRISPR/Cas technology Calcium Cardiac Cardiac Function Study Cardiac Myocytes Cardiotoxicity Cardiovascular Diseases Cardiovascular system Cell Line Cell model Cells Cessation of life Clinical Trials Complex Computational Biology Contracts DNA sequencing Data Dilated Cardiomyopathy Disease Disease model Electrophysiology (science)Engineering Fibroblasts Gene Targeting Genes Genetic Genetic Diseases Genomics Goals Guide RNA Heart Heart failure Homeostasis Human Human Engineering Human Genetics Hypertrophic Cardiomyopathy Immunology In Vitro Inherited Innate Immune System Interferon Type I Kinetics Knowledge Mendelian disorder Methods Modeling Modification Molecular Morbidity - disease rate Mutate Other Genetics Outcome Patients Phenotype Physiology Property Public Health Reagent Research Personnel Resolution Resources Risk Risk Assessment Safety Site Structure System Testing Therapeutic Tissue Model Tissues adverse outcome clinical application clinical development connectin disease-causing mutation exhaustion experience genetic variant genome editing genome sequencing genotoxicity heart dimension/size heart function human disease human tissue immunogenicity improved in vivo induced pluripotent stem cell innate immune sensing insight macrophage mortality next generation pre-clinical precision medicine preclinical study prevent reagent testing response single-cell RNA sequencing somatic cell gene editing technology development therapeutic development tool transcriptome sequencing transcriptomics tumorigenesis

项目摘要

PROJECT SUMMARY/ABSTRACT Somatic cell genome editing (SCGE) has remarkable promise to transform our therapeutic toolbox for the treatment of human genetic disorders. However, despite having the tools available for identification and modification of human disease-causing mutations, outstanding concerns over efficacy and safety have curtailed the clinical application of SCGE broadly. Critical gaps in our current knowledge of the safety of SCGE approaches include: 1) what are the on- and off-target genome editing rates, 2) how do SCGE reagents affect human cellular and tissue function, and 3) how will the innate and adaptive immune system respond to SCGE reagents. While clinical trials are effective tools to determine efficacy and safety, they are most efficiently applied after exhaustive pre-clinical studies have optimized efficacy and safety in other systems. The goal of this proposal is to adapt biomimetic human cardiac microtissues (CMTs)--engineered from cardiomyocytes derived from human induced pluripotent stem cells (iPSCs), fibroblasts, and macrophages--to study the impact of SCGE reagents and delivery systems on a functional human tissue. Because CMTs recapitulate in vivo cardiac three-dimensional architecture, biomechanical properties, and complex multicellular interactions that are critical to cardiac tissue homeostasis and function, they are ideal for assaying cardiac functions in vitro. The CMTs have been optimized for functional assays that quantify a range of dynamic phenotypes that include orders-of-magnitude changes in tissue contractility, calcium handling, and electrophysiology that predict in vivo cardiac function. Importantly, CRISPR/Cas9 genome editors have been effectively applied to CMTs to generate monogenic disease models of common cardiovascular disorders such as dilated and hypertrophic cardiomyopathies that result in heart failure. Guided by their comprehensive preliminary data including application of next-generation DNA- and RNA- sequencing assays to CMTs, the researchers propose to pursue two Specific Aims to determine SCGE efficacy and safety: 1) interrogate CMTs by comprehensive assessment of contractility, calcium handling and electrical function in combination with single-cell transcriptomics paired with off-target genome sequencing to identify adverse consequences of SCGE reagents that target the titin-encoding gene TTN, and 2) engineer autologous CMTs assembled with two distinct classes of macrophages to study cardiac function and SCGE reagents. Execution of these Aims will provide multi-scale insights into the safety and efficacy of SCGE reagents by producing an informative testing platform and system of associated methods to identify adverse outcomes. Establishing these resources will be a pivotal step toward realizing the promise of genome editing and human precision medicine of cardiovascular and other disorders.

项目总结/文摘