title: Permanent alteration of PCSK9 with in vivo CRISPR-Cas9 genome editing.
abstract: BACKGROUND
  Individuals with naturally occurring loss-of-function proprotein convertase subtilisin/kexin type 9 (PCSK9) mutations experience reduced low-density lipoprotein cholesterol levels and protection against cardiovascular disease.
  
  
  OBJECTIVE
  The goal of this study was to assess whether genome editing using a clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated system can efficiently introduce loss-of-function mutations into the endogenous PCSK9 gene in vivo.
  
  
  RESULTS
  We used adenovirus to express CRISPR-associated 9 and a CRISPR guide RNA targeting Pcsk9 in mouse liver, where the gene is specifically expressed. We found that <3 to 4 days of administration of the virus, the mutagenesis rate of Pcsk9 in the liver was as high as >50%. This resulted in decreased plasma PCSK9 levels, increased hepatic low-density lipoprotein receptor levels, and decreased plasma cholesterol levels (by 35-40%). No off-target mutagenesis was detected in 10 selected sites.
  
  
  CONCLUSIONS
  Genome editing with the CRISPR-CRISPR-associated 9 system disrupts the Pcsk9 gene in vivo with high efficiency and reduces blood cholesterol levels in mice. This approach may have therapeutic potential for the prevention of cardiovascular disease in humans.
authors.affiliation: From the Department of Stem Cell and Regenerative Biology, Harvard University and Harvard Stem Cell Institute, Cambridge, MA (Q.D., S.-L.N., B.S.G., S.N.R., C.A.C., K.M.); Institute for Translational Medicine and Therapeutics, Institute for Diabetes, Obesity and Metabolism, and Cardiovascular Institute, Perelman School of Medicine at the University of Pennsylvania, Philadelphia (A.S., K.M.P., D.J.R.); Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (K.M.); and Broad Institute, Cambridge, MA (C.A.C., K.M.).
authors.forename: Qiurong
authors.initials: Q
authors.lastname: Ding
authors.affiliation: From the Department of Stem Cell and Regenerative Biology, Harvard University and Harvard Stem Cell Institute, Cambridge, MA (Q.D., S.-L.N., B.S.G., S.N.R., C.A.C., K.M.); Institute for Translational Medicine and Therapeutics, Institute for Diabetes, Obesity and Metabolism, and Cardiovascular Institute, Perelman School of Medicine at the University of Pennsylvania, Philadelphia (A.S., K.M.P., D.J.R.); Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (K.M.); and Broad Institute, Cambridge, MA (C.A.C., K.M.).
authors.forename: Alanna
authors.initials: A
authors.lastname: Strong
authors.affiliation: From the Department of Stem Cell and Regenerative Biology, Harvard University and Harvard Stem Cell Institute, Cambridge, MA (Q.D., S.-L.N., B.S.G., S.N.R., C.A.C., K.M.); Institute for Translational Medicine and Therapeutics, Institute for Diabetes, Obesity and Metabolism, and Cardiovascular Institute, Perelman School of Medicine at the University of Pennsylvania, Philadelphia (A.S., K.M.P., D.J.R.); Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (K.M.); and Broad Institute, Cambridge, MA (C.A.C., K.M.).
authors.forename: Kevin M
authors.initials: KM
authors.lastname: Patel
authors.affiliation: From the Department of Stem Cell and Regenerative Biology, Harvard University and Harvard Stem Cell Institute, Cambridge, MA (Q.D., S.-L.N., B.S.G., S.N.R., C.A.C., K.M.); Institute for Translational Medicine and Therapeutics, Institute for Diabetes, Obesity and Metabolism, and Cardiovascular Institute, Perelman School of Medicine at the University of Pennsylvania, Philadelphia (A.S., K.M.P., D.J.R.); Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (K.M.); and Broad Institute, Cambridge, MA (C.A.C., K.M.).
authors.forename: Sze-Ling
authors.initials: SL
authors.lastname: Ng
authors.affiliation: From the Department of Stem Cell and Regenerative Biology, Harvard University and Harvard Stem Cell Institute, Cambridge, MA (Q.D., S.-L.N., B.S.G., S.N.R., C.A.C., K.M.); Institute for Translational Medicine and Therapeutics, Institute for Diabetes, Obesity and Metabolism, and Cardiovascular Institute, Perelman School of Medicine at the University of Pennsylvania, Philadelphia (A.S., K.M.P., D.J.R.); Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (K.M.); and Broad Institute, Cambridge, MA (C.A.C., K.M.).
authors.forename: Bridget S
authors.initials: BS
authors.lastname: Gosis
authors.affiliation: From the Department of Stem Cell and Regenerative Biology, Harvard University and Harvard Stem Cell Institute, Cambridge, MA (Q.D., S.-L.N., B.S.G., S.N.R., C.A.C., K.M.); Institute for Translational Medicine and Therapeutics, Institute for Diabetes, Obesity and Metabolism, and Cardiovascular Institute, Perelman School of Medicine at the University of Pennsylvania, Philadelphia (A.S., K.M.P., D.J.R.); Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (K.M.); and Broad Institute, Cambridge, MA (C.A.C., K.M.).
authors.forename: Stephanie N
authors.initials: SN
authors.lastname: Regan
authors.affiliation: From the Department of Stem Cell and Regenerative Biology, Harvard University and Harvard Stem Cell Institute, Cambridge, MA (Q.D., S.-L.N., B.S.G., S.N.R., C.A.C., K.M.); Institute for Translational Medicine and Therapeutics, Institute for Diabetes, Obesity and Metabolism, and Cardiovascular Institute, Perelman School of Medicine at the University of Pennsylvania, Philadelphia (A.S., K.M.P., D.J.R.); Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (K.M.); and Broad Institute, Cambridge, MA (C.A.C., K.M.).
authors.forename: Chad A
authors.initials: CA
authors.lastname: Cowan
authors.affiliation: From the Department of Stem Cell and Regenerative Biology, Harvard University and Harvard Stem Cell Institute, Cambridge, MA (Q.D., S.-L.N., B.S.G., S.N.R., C.A.C., K.M.); Institute for Translational Medicine and Therapeutics, Institute for Diabetes, Obesity and Metabolism, and Cardiovascular Institute, Perelman School of Medicine at the University of Pennsylvania, Philadelphia (A.S., K.M.P., D.J.R.); Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (K.M.); and Broad Institute, Cambridge, MA (C.A.C., K.M.). kiranmusunuru@gmail.com rader@mail.med.upenn.edu.
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Rader
authors.affiliation: From the Department of Stem Cell and Regenerative Biology, Harvard University and Harvard Stem Cell Institute, Cambridge, MA (Q.D., S.-L.N., B.S.G., S.N.R., C.A.C., K.M.); Institute for Translational Medicine and Therapeutics, Institute for Diabetes, Obesity and Metabolism, and Cardiovascular Institute, Perelman School of Medicine at the University of Pennsylvania, Philadelphia (A.S., K.M.P., D.J.R.); Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (K.M.); and Broad Institute, Cambridge, MA (C.A.C., K.M.). kiranmusunuru@gmail.com rader@mail.med.upenn.edu.
authors.forename: Kiran
authors.initials: K
authors.lastname: Musunuru
mesh_terms: D041721:3T3-L1 Cells; D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D015415:Biomarkers / Q000097:blood; D002318:Cardiovascular Diseases / Q000201:enzymology / Q000235:genetics / Q000517:prevention & control; D002784:Cholesterol / Q000097:blood; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D005822:Genetic Vectors; D005838:Genotype; D008099:Liver / Q000201:enzymology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009154:Mutation; D010641:Phenotype; D000071449:Proprotein Convertase 9; D043484:Proprotein Convertases / Q000097:blood / Q000235:genetics*; D065840:Protective Factors; D017393:RNA Editing; D000094704:RNA, Guide, CRISPR-Cas Systems / Q000235:genetics* / Q000378:metabolism; D011973:Receptors, LDL / Q000378:metabolism; D012697:Serine Endopeptidases / Q000097:blood / Q000235:genetics*; D013997:Time Factors; D014162:Transfection
chemical_list: D015415:Biomarkers; D000094704:RNA, Guide, CRISPR-Cas Systems; D011973:Receptors, LDL; D002784:Cholesterol; C472126:Pcsk9 protein, mouse; D000071449:Proprotein Convertase 9; D043484:Proprotein Convertases; D012697:Serine Endopeptidases
keywords: coronary disease; genetic therapy; lipoproteins; molecular biology; prevention and control