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

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.

• Authors: Xiao Wang, Avanthi Raghavan, Tao Chen, Lyon Qiao, Yongxian Zhang, Qiurong Ding, Kiran Musunuru
• Journal: Arteriosclerosis, thrombosis, and vascular biology (Arterioscler Thromb Vasc Biol) Vol. 36 Issue 5 Pg. 783-6 (05 2016) ISSN: 1524-4636 [Electronic] United States
• PMID: 26941020 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)
• Copyright: © 2016 American Heart Association, Inc.

Chemical References

• CRISPR-Associated Proteins
• DNA-Binding Proteins
• Il2rg protein, mouse
• Interleukin Receptor Common gamma Subunit
• Rag2 protein, mouse
• Hydrolases
• PCSK9 protein, human
• Proprotein Convertase 9
• fumarylacetoacetase

Topics

• Animals
• CRISPR-Associated Proteins (genetics, metabolism)
• CRISPR-Cas Systems
• Clustered Regularly Interspaced Short Palindromic Repeats
• DNA-Binding Proteins (deficiency, genetics)
• Down-Regulation
• Gene Editing (methods)
• Gene Targeting (methods)
• Genotype
• Hepatocytes (enzymology, transplantation)
• Humans
• Hydrolases (deficiency, genetics)
• Interleukin Receptor Common gamma Subunit (deficiency, genetics)
• Mice
• Mice, Inbred C57BL
• Mice, Knockout
• Mutation
• Proprotein Convertase 9 (biosynthesis, blood, genetics)