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.
