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Arterioscler Thromb Vasc Biol. 2016 May;36(5):783-6. doi: 10.1161/ATVBAHA.116.307227. Epub 2016 Mar 3.

CRISPR-Cas9 Targeting of PCSK9 in Human Hepatocytes In Vivo-Brief Report.

Author information

1
From the Department of Stem Cell and Regenerative Biology, Harvard University, and Harvard Stem Cell Institute, Cambridge, MA (X.W., A.R., T.C., L.Q., K.M.); Harvard Medical School, Boston, MA (A.R., K.M.); Key Laboratory of Nutrition and Metabolism, Institute for Nutritional Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, PR China (Y.Z., Q.D.); Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (K.M.); and Broad Institute, Cambridge, MA (K.M.).
2
From the Department of Stem Cell and Regenerative Biology, Harvard University, and Harvard Stem Cell Institute, Cambridge, MA (X.W., A.R., T.C., L.Q., K.M.); Harvard Medical School, Boston, MA (A.R., K.M.); Key Laboratory of Nutrition and Metabolism, Institute for Nutritional Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, PR China (Y.Z., Q.D.); Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (K.M.); and Broad Institute, Cambridge, MA (K.M.). kiranmusunuru@gmail.com qrding@sibs.ac.cn.

Abstract

OBJECTIVE:

Although early proof-of-concept studies of somatic in vivo genome editing of the mouse ortholog of proprotein convertase subtilisin/kexin type 9 (Pcsk9) in mice have established its therapeutic potential for the prevention of cardiovascular disease, the unique nature of genome-editing technology-permanent alteration of genomic DNA sequences-mandates that it be tested in vivo against human genes in normal human cells with human genomes to give reliable preclinical insights into the efficacy (on-target mutagenesis) and safety (lack of off-target mutagenesis) of genome-editing therapy before it can be used in patients.

APPROACH AND RESULTS:

We used a clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated (Cas) 9 genome-editing system to target the human PCSK9 gene in chimeric liver-humanized mice bearing human hepatocytes. We demonstrated high on-target mutagenesis (approaching 50%), greatly reduced blood levels of human PCSK9 protein, and minimal off-target mutagenesis.

CONCLUSIONS:

This work yields important information on the efficacy and safety of CRISPR-Cas9 therapy targeting the human PCSK9 gene in human hepatocytes in vivo, and it establishes humanized mice as a useful platform for the preclinical assessment of applications of somatic in vivo genome editing.

KEYWORDS:

gene therapy; liver; molecular biology; mutagenesis; subtilisins

PMID:
26941020
PMCID:
PMC4850082
DOI:
10.1161/ATVBAHA.116.307227
[Indexed for MEDLINE]
Free PMC Article

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