DEVELOPING WOODCHUCKS SUSCEPTIBLE TO HEPATITIS B VIRUS INFECTION BY MODIFYING THE VIRUS OR HOST - ABSTRACT CHRONIC INFECTION WITH HEPATITIS B VIRUS (HBV) IS A CRITICAL PUBLIC HEALTH PROBLEM AFFECTING APPROXIMATELY 296 MILLION INDIVIDUALS WORLDWIDE AND RESULTING IN 820,000 DEATHS EVERY YEAR DUE TO SEVERE LIVER DISEASE PROGRESSION. DEVELOPING NEW ANTIVIRALS FOR THE INTERVENTION OF CHRONIC HBV INFECTION AND A BETTER UNDERSTANDING OF THE UNDERLYING MECHANISMS OF HBV PERSISTENCE ARE URGENTLY NEEDED FOR INCREASING THE LOW HBV CURE RATE IN PATIENTS, WHICH IS HAMPERED BY DEFICIENCIES IN CURRENT ANIMAL MODELS SUSCEPTIBLE TO HBV INFECTION. THE EASTERN WOODCHUCK (MARMOTA MONAX), NATURALLY INFECTED WITH THE WOODCHUCK HEPATITIS VIRUS (WHV) THAT IS CLOSELY RELATED TO HBV, IS A WELL-ESTABLISHED, IMMUNOCOMPETENT AND OUTBRED ANIMAL MODEL FOR THE STUDY OF HBV INFECTION AND FOR THE PRECLINICAL EVALUATION OF NEW ANTI-HBV DRUGS. HOWEVER, SEQUENCE AND STRUCTURAL DIFFERENCES BETWEEN HBV AND WHV MAKE THE DEVELOPMENT OF HBV-SPECIFIC DRUGS RATHER CHALLENGING IN THIS MODEL. OUR PRELIMINARY IN VITRO AND IN VIVO DATA DEMONSTRATE THAT A MODIFIED HBV CONSTRUCT MAINTAINS REPLICATION COMPETENCE IN HUMAN AND WOODCHUCK HEPATIC CELL LINES AND THAT CELL CULTURE-DERIVED VIRIONS INFECT ADULT WOODCHUCKS, ALTHOUGH AT A VERY LOW LEVEL. OTHER PRELIMINARY IN VITRO DATA INDICATE THAT MODIFIED WOODCHUCK HEPATOMA CELLS SUPPORT THE ENTIRE LIFE CYCLE OF HBV. THUS, WE PROPOSE TO EXPLORE THE POSSIBILITY OF PRODUCTIVELY INFECTING WOODCHUCKS WITH HBV BY EITHER A MODIFIED VIRUS IN AIM 1 OR A MODIFIED HOST IN AIM 2 FOR CONTINUED DEVELOPMENT OF THE WOODCHUCK AS AN ANIMAL MODEL FOR CHRONIC HBV INFECTION. UNDER AIM 1, THIS WILL BE ACHIEVED BY ADDITIONAL PASSAGE OF HBV- CONTAINING WOODCHUCK SERUM FROM OUR PRELIMINARY STUDY IN ADULT WOODCHUCKS. ANIMALS WILL BE EITHER IMMUNOCOMPETENT TO ALLOW CONTINUED VIRAL ADAPTION TO THE WOODCHUCK IMMUNE SYSTEM, AS WELL AS HEPATOCYTES, OR IMMUNOSUPPRESSED TO PROVIDE MORE TIME AND OPPORTUNITIES FOR VIRAL EVOLUTION AND CONTINUED HOST ADAPTION IN THE ABSENCE OF IMMUNE PRESSURE. SEQUENCE ANALYSIS OF ADAPTED/EVOLVED HBV IN SERUM AND LIVER OF THESE ANIMALS WILL IDENTIFY POSSIBLE MUTATIONS IN VIRAL PROMOTER/ENHANCER ELEMENTS AND THE CORE GENE CRITICAL FOR WOODCHUCK ADAPTION, AND THUS WILL ALLOW TO ENGINEER ADDITIONAL HBV CONSTRUCTS WITH INCREASED INFECTION EFFICIENCY FOR SUBSEQUENT EVALUATION IN ADULT WOODCHUCKS. THE BEST PERFORMING HBV, EITHER ADAPTED/EVOLVED OR ENGINEERED, WILL THEN BE USED FOR THE INFECTION OF NEONATAL WOODCHUCKS TO ALLOW THE ESTABLISHMENT OF CHRONIC HBV INFECTION AND LIVER DISEASE PROGRESSION, INCLUDING DEVELOPMENT OF HEPATOCELLULAR CARCINOMA, AS IT IS TYPICALLY OBSERVED IN HUMANS. UNDER AIM 2, THIS WILL BE ACCOMPLISHED BY INFECTION OF ADULT WOODCHUCKS WITH WILDTYPE OR ENGINEERED HBV AFTER RENDERING THE HOST'S LIVER CELLS PERMISSIVE TO THE VIRUS. THESE ANIMALS WILL BE EITHER IMMUNOCOMPETENT FOR PROOF-OF-CONCEPT OF TRANSIENT, RESOLVING HBV INFECTION OR IMMUNOSUPPRESSED TO ENABLE CHRONIC HBV INFECTION IN THE ABSENCE OF AN IMMUNE RESPONSE. SUCCESSFUL COMPLETION OF THE PROPOSED RESEARCH WILL HAVE A SIGNIFICANT IMPACT ON DEVELOPING NEW ANTIVIRALS IN WOODCHUCKS WITH CHRONIC HBV INFECTION.