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422
Eliminating HBV Virus By Disrupting the
Synthesis of HBV Covalently Closed Circular
DNA
Yong Liao1, Di Mu2 and Lingyu Jiang2, (1)Key Laboratory
of Molecular Biology for Infectious Diseases (Ministry
of Education), Institute for Viral Hepatitis, Department
of Infectious Diseases, the Second Affiliated Hospital,
Chongqing Medical University, Chongqing, PR China., (2)
Institute for Viral Hepatitis, Chongqing Medical University,
Chongqing, PR China
Background: Currently, there are two types of anti-viral
drugs, namely interferon (IFN) and nucleotide/nucleoside
analogues (NAs), that are effective in terms of repression
of HBV viral replication in patients with chronic infection of
hepatitis B virus (HBV). However, outcomes in patients who
had received these anti-viral therapies, either IFN or NAs
alone or in combination of both, are far from satisfactory,
mainly due to the persistence of cccDNA in the nucleus of
infected hepatocytes. Therefore, novel strategies and more
effective therapeutic drugs are urgently needed in order to
improve the clinic outcomes. Here, we report compounds that
can specifically disrupt the synthesis of the viral replication
template, HBV cccDNA. Methods: A panel of compounds
that have well-characterized and highly specific targets
for the de novo synthesis of nucleotides and amino acids
were screened against cell lines with stable expression of
a replicative HBV virus and its template cccDNA, such as
HepG2.215 and tetracycline-inducible replication of HBV
virus, HepAD38 cells. Levels of intrahepatic HBV cccDNA and
HBV DNA in hepatocytes and supernatants were absolutely
quantified by a droplet digital PCR assay developed recently
by us (Mu D, et al, Biotechnology Letter, 2015; Mu D., et
al, Scientific Report, 2017). For animal model evaluation
in vivo, mice were hydrodynamically injected i.v. with 10
ug pAAV-HBV1.2 plasmid. Selected compounds that have
demonstrated efficacy in vitro were then administered, either
intra-peritoneally or intra-nasal-gastrically, depends on the
formulation of the compound to be tested. Levels of serum
HBV DNA and other parameters related to HBV infection,
such HBsAg, HBeAg, anti-HBsAb, Anti-HBeAb, etc, and
liver functions were monitored regularly during the antiviral
therapy in the animals. IFN or Tenofovir was tested in
parallel as a positive control for our study. Results: In the cell
culture screening, we observed that a few compounds were
highly effective and dramatically reduced the copy numbers
of HBV DNA and/or intrahepatic cccDNA within a couple of
weeks, but exposure to interferon failed to affect intrahepatic
HBV cccDNA within the same time frame. Some of these
compounds had also achieved a complete elimination of
serum and intrahepatic HBV DNA in the animal models.
Meanwhile, Tenofovir showed modest effects in repression of
HBV DNA replication in vivo, but none of the animals that had
received Tenofovir reached a complete elimination of serum
and intrahepatic HBV DNA within 12 weeks. Conclusion:
We demonstrated here that certain compounds were able
to disrupt the synthesis of HBV cccDNA in cell lines with
expression of a replicative HBV virus and animal models with
chronic HBV infection, an efficacy that cannot be achieved by
either interferon or Tenofovir in the parallel experiments |
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发表于 2018-10-20 20:53
