title: Adenovirus-Mediated Somatic Genome Editing of Pten by CRISPR/Cas9 in Mouse Liver in Spite of Cas9-Specific Immune Responses.
abstract: CRISPR/Cas9 derived from the bacterial adaptive immunity pathway is a powerful tool for genome editing, but the safety profiles of in vivo delivered Cas9 (including host immune responses to the bacterial Cas9 protein) have not been comprehensively investigated in model organisms. Nonalcoholic steatohepatitis (NASH) is a prevalent human liver disease characterized by excessive fat accumulation in the liver. In this study, we used adenovirus (Ad) vector to deliver a Streptococcus pyogenes-derived Cas9 system (SpCas9) targeting Pten, a gene involved in NASH and a negative regulator of the PI3K-AKT pathway, in mouse liver. We found that the Ad vector mediated efficient Pten gene editing even in the presence of typical Ad vector-associated immunotoxicity in the liver. Four months after vector infusion, mice receiving the Pten gene-editing Ad vector showed massive hepatomegaly and features of NASH, consistent with the phenotypes following Cre-loxP-induced Pten deficiency in mouse liver. We also detected induction of humoral immunity against SpCas9 and the potential presence of an SpCas9-specific cellular immune response. Our findings provide a strategy to model human liver diseases in mice and highlight the importance considering Cas9-specific immune responses in future translational studies involving in vivo delivery of CRISPR/Cas9.
authors.affiliation: 1 Gene Therapy Center, University of Massachusetts Medical School , Worcester, Massachusetts.
authors.forename: Dan
authors.initials: D
authors.lastname: Wang
authors.affiliation: 2 RNA Therapeutics Institute and Program in Molecular Medicine, University of Massachusetts Medical School , Worcester, Massachusetts.
authors.forename: Haiwei
authors.initials: H
authors.lastname: Mou
authors.affiliation: 1 Gene Therapy Center, University of Massachusetts Medical School , Worcester, Massachusetts.
authors.forename: Shaoyong
authors.initials: S
authors.lastname: Li
authors.affiliation: 3 Department of Bioinformatics, School of Life Science and Technology, Tongji University , Shanghai, P.R. China .
authors.forename: Yingxiang
authors.initials: Y
authors.lastname: Li
authors.affiliation: 2 RNA Therapeutics Institute and Program in Molecular Medicine, University of Massachusetts Medical School , Worcester, Massachusetts.
authors.forename: Soren
authors.initials: S
authors.lastname: Hough
authors.affiliation: 1 Gene Therapy Center, University of Massachusetts Medical School , Worcester, Massachusetts.
authors.forename: Karen
authors.initials: K
authors.lastname: Tran
authors.affiliation: 1 Gene Therapy Center, University of Massachusetts Medical School , Worcester, Massachusetts.
authors.forename: Jia
authors.initials: J
authors.lastname: Li
authors.affiliation: 4 David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology , Cambridge, Massachusetts.
authors.forename: Hao
authors.initials: H
authors.lastname: Yin
authors.affiliation: 4 David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology , Cambridge, Massachusetts.|5 Department of Chemical Engineering, Massachusetts Institute of Technology , Cambridge, Massachusetts.|6 Harvard-MIT Division of Health Sciences & Technology, Cambridge, Massachusetts.|7 Institute of Medical Engineering and Science, Massachusetts Institute of Technology , Cambridge, Massachusetts.
authors.forename: Daniel G
authors.initials: DG
authors.lastname: Anderson
authors.affiliation: 2 RNA Therapeutics Institute and Program in Molecular Medicine, University of Massachusetts Medical School , Worcester, Massachusetts.
authors.forename: Erik J
authors.initials: EJ
authors.lastname: Sontheimer
authors.affiliation: 8 Program in Bioinformatics and Integrative Biology, University of Massachusetts Medical School , Worcester, Massachusetts.
authors.forename: Zhiping
authors.initials: Z
authors.lastname: Weng
authors.affiliation: 1 Gene Therapy Center, University of Massachusetts Medical School , Worcester, Massachusetts.
authors.forename: Guangping
authors.initials: G
authors.lastname: Gao
authors.affiliation: 2 RNA Therapeutics Institute and Program in Molecular Medicine, University of Massachusetts Medical School , Worcester, Massachusetts.
authors.forename: Wen
authors.initials: W
authors.lastname: Xue
mesh_terms: D000256:Adenoviridae; D000818:Animals; D001426:Bacterial Proteins / Q000276:immunology; D000076987:CRISPR-Associated Protein 9; D064113:CRISPR-Cas Systems; D064112:Clustered Regularly Interspaced Short Palindromic Repeats / Q000276:immunology; D004195:Disease Models, Animal; D004720:Endonucleases / Q000276:immunology; D015316:Genetic Therapy / Q000009:adverse effects*; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D065626:Non-alcoholic Fatty Liver Disease / Q000628:therapy*
chemical_list: D001426:Bacterial Proteins; D000076987:CRISPR-Associated Protein 9; C000606107:Cas9 endonuclease Streptococcus pyogenes; D004720:Endonucleases