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Therapeutic potential of CRISPR/Cas9 genome engineering in humanized mouse

CRISPR/Cas9 基因组工程在人源化小鼠中的治疗潜力

基本信息

批准号：
10291691
负责人：
Karl-Dimiter Bissig
金额：
$ 54.97万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-01-20 至 2022-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10291691
关键词：
Address Animal Model Apolipoproteins B Atherosclerosis Biological Assay Blood CRISPR/Cas technology Cadaver Cell Cycle Cells Cholesterol Chromatin Clinic Code Custom DNA sequencing Data Dependovirus Disease Enzymes Essential Genes Excision Exons Expression Profiling Familial Combined Hyperlipidemia Familial Hypercholesterolemia Gene Expression Genes Genome engineering Goals Guide RNA Hepatocyte Hepatotoxicity Histologic Human Human Cell Line Human Genome Hyperlipoproteinemia Immunodeficient Mouse Introns Investigational Therapies Lipids Lipoprotein (a)Lipoproteins Liver Liver diseases Mammals Measures Mediating Mendelian disorder Metabolic Metabolic Diseases Modeling Monitor Mouse Strains Mus Mutation Nodule Nonhomologous DNA End Joining Pathologic Pathology Patients Plasma Production Proteins Refractory Reporter Resistance S-Phase Fraction Safety Serotyping Squalene Synthetase System Techniques Technology Testing Therapeutic Tissues Translating Transplantation Treatment Efficacy Very low density lipoprotein Xenograft Model Xenograft procedure apolipoprotein Lp(a+)compliance behavior design efficacy testing exome sequencing experience gene therapy genome editing human disease humanized mouse hypercholesterolemia in vivo innovation lipid disorder liver cell proliferation mouse model novel particle pre-clinical repaired therapeutic genome editing transcriptome sequencing transplant model

项目摘要

PROJECT SUMMARY Liver-directed gene editing has the potential to correct numerous severe monogenic disorders. The two major hurdles of translating this therapy to the clinic are efficacy and safety. We will address both concerns using an innovative therapeutic genome editing approach in a novel humanized mouse model. CRISPR/Cas9 genome editing technology has proven extremely efficient in introducing double strand brakes (DSB) in many cellular systems. In mammals, DSB are preferentially repaired by the error prone non-homologous end joining (NHEJ) and to a lesser extent by homology-directed repair (HDR). Here, we propose to treat refractory lipid disorders with compensatory deletion of a whole exon (exon-excision) from lipid genes that increase the cholesterol levels in blood or contribute to slower clearance of blood cholesterol. Deletion will be mediated by NHEJ after introduction of two CRISPR/Cas9 induced DSBs in the two flanking introns. This repair mechanism is very efficient in non-proliferating hepatocytes and avoids the introduction of potentially harmful mutations in the coding sequence. We recently developed a mouse strain that can be repopulated with cadaveric human hepatocytes. Since genome editing is sequence dependent, it must be tested within the context of human cells of the desired target tissue. Using mice repopulated with normal human hepatocytes, we will be able to determine the safety and efficacy of liver-directed human genome editing by CRISPR/Cas9. We will then test his approach in mice repopulated with human hepatocytes from a patient with familial hypercholesterolemia (FH). This first xenograft model for FH can be used to evaluate the therapeutic efficacy and potential compensatory adaptation for this experimental therapy.

项目总结