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.

Public Health Relevance

CRISPR/Cas9 genome editing is a powerful new technique that has the potential to permanently cure severe and refractory mongenetic disorders, such as some lipid disorders. We have recently developed xenograft (transplant) models that mimic the normal human liver and the diseased liver of familial hypercholesterolemia (refractory lipid disorder). We will use this mouse model repopulated with normal and diseased human hepatocytes to test the efficacy and safety of CRISPR/Cas9 genome editing and to evaluate its therapeutic potential in the human context.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Project (R01)
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Therapeutic Approaches to Genetic Diseases Study Section (TAG)
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Liu, Lijuan
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Baylor College of Medicine
Anatomy/Cell Biology
Schools of Medicine
United States
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