title: Suppression of hepatitis B virus DNA accumulation in chronically infected cells using a bacterial CRISPR/Cas RNA-guided DNA endonuclease.
abstract: Hepatitis B virus (HBV) remains a major human pathogen, with over 240 million individuals suffering from chronic HBV infections. These can persist for decades due to the lack of therapies that can effectively target the stable viral covalently closed circular (ccc) DNA molecules present in infected hepatocytes. Using lentiviral transduction of a bacterial Cas9 gene and single guide RNAs (sgRNAs) specific for HBV, we observed effective inhibition of HBV DNA production in in vitro models of both chronic and de novo HBV infection. Cas9/sgRNA combinations specific for HBV reduced total viral DNA levels by up to ~1000-fold and HBV cccDNA levels by up to ~10-fold and also mutationally inactivated the majority of the residual viral DNA. Together, these data provide proof of principle for the hypothesis that CRISPR/Cas systems have the potential to serve as effective tools for the depletion of the cccDNA pool in chronically HBV infected individuals.
authors.affiliation: Department of Molecular Genetics & Microbiology and Center for Virology, Duke University Medical Center, Durham, NC, United States.
authors.forename: Edward M
authors.initials: EM
authors.lastname: Kennedy
authors.affiliation: Center for AIDS Research, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, United States.
authors.forename: Leda C
authors.initials: LC
authors.lastname: Bassit
authors.affiliation: Infectious Diseases, F. Hoffmann-LaRoche, Inc., Basel, Switzerland.
authors.forename: Henrik
authors.initials: H
authors.lastname: Mueller
authors.affiliation: Department of Molecular Genetics & Microbiology and Center for Virology, Duke University Medical Center, Durham, NC, United States.
authors.forename: Anand V R
authors.initials: AVR
authors.lastname: Kornepati
authors.affiliation: Department of Molecular Genetics & Microbiology and Center for Virology, Duke University Medical Center, Durham, NC, United States.
authors.forename: Hal P
authors.initials: HP
authors.lastname: Bogerd
authors.affiliation: Center for AIDS Research, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, United States.
authors.forename: Ting
authors.initials: T
authors.lastname: Nie
authors.affiliation: Center for AIDS Research, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, United States.
authors.forename: Payel
authors.initials: P
authors.lastname: Chatterjee
authors.affiliation: Infectious Diseases, F. Hoffmann-LaRoche, Inc., Basel, Switzerland.
authors.forename: Hassan
authors.initials: H
authors.lastname: Javanbakht
authors.affiliation: Center for AIDS Research, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, United States. Electronic address: raymond.schinazi@emory.edu.
authors.forename: Raymond F
authors.initials: RF
authors.lastname: Schinazi
authors.affiliation: Department of Molecular Genetics & Microbiology and Center for Virology, Duke University Medical Center, Durham, NC, United States. Electronic address: bryan.cullen@duke.edu.
authors.forename: Bryan R
authors.initials: BR
authors.lastname: Cullen
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D064130:CRISPR-Associated Proteins / Q000378:metabolism; D064113:CRISPR-Cas Systems; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000378:metabolism; D015536:Down-Regulation; D018390:Gene Targeting / Q000379:methods*; D006509:Hepatitis B / Q000628:therapy / Q000821:virology*; D006515:Hepatitis B virus / Q000235:genetics* / Q000502:physiology; D006801:Humans; D013297:Streptococcus pyogenes / Q000201:enzymology / Q000235:genetics; D014779:Virus Replication
chemical_list: D001426:Bacterial Proteins; D064130:CRISPR-Associated Proteins; D004279:DNA, Viral; D003850:Deoxyribonuclease I
keywords: Antiviral; CRISPR/Cas; DNA editing; HBV; Lentiviral vector; cccDNA