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77
Suppression of Hepatitis B Virus Antigens
By RNA Interference Enables Therapeutic
Vaccination to Achieve Immune Control in
High-Titer Virus Carriers
Thomas Michler1, Anna Kosinska1, Dirk Grimm2, Mathias
Heikenwalder3, Stuart Milstein4, Laura Sepp-Lorenzino4,
Percy Knolle5 and Ulrike Protzer1, (1)Institute of Virology,
Technical University / Helmholtz Center Munich, (2)
Department of Infectious Diseases/Virology, (3)Division of
Chronic Inflammation and Cancer, German Cancer Research
Center (DKFZ), (4)Alnylam Pharmaceuticals, Cambridge, MA
02142, USA, (5)Institute of Molecular Immunology, University
Hospital Rechts Der Isar, Technical University of Munich
Background: Hepatitis B Virus (HBV) persistence was found
to correlate with a failure to develop an efficient virus-specific
T cell response due to high HBV antigen load. We evaluated
the capacity of stabilized, liver-targeted siRNAs to (i) suppress
HBV gene expression, (ii) allow recovery of HBV-specific Band
T cell responses spontaneously or (iii) after therapeutic
vaccination. Methods: High viremic HBV-transgenic or
C57/Bl6 mice transduced with an Adeno-Associated Virus
(AAV)-HBV vector were treated with nucleoside analogue
Entecavir (ETV), shRNA-expressing AAV (AAV-shHBV) or
N-Acetylgalactosamine (GalNAc)-conjugated siRNAs. B
and T cell immunity were monitored following therapeutic
vaccination with a HBV core (HBc) and surface (HBs) protein
prime vaccination and a Modified Vaccinia Ankara virus
(MVA)- boost immunization (TherVacB). Results: RNAi by
monthly s.c. injections of 3 mg/kg GalNAc-siRNA or 1x1011
geq AAV-shHBV i.v. suppressed serum HBsAg and HBV DNA
by 2-3 log10 and HBeAg by >1 log10. ETV reduced viremia
by 4 log10 but antigen levels remained unchanged. TherVacB
induced HBV-specific B-cell immunity and CD4 T cell
responses independent of antigen levels, but HBV-specific
CD8 T cell responses were only seen in animals with reduced
antigen levels after RNAi. Induction of CD8 T cell responses
coincided with suppression of HBV replication in the liver to
levels undetectable by Southern blot or PCR. To evaluate if
the combinatorial RNAi/vaccination therapy could achieve
long-term control of HBV replication and functional cure,
C57/Bl6 mice were transduced with 2x1011 geq AAV-HBV
to establish high-titer, persistent HBV replication. 3 monthly
doses of GalNAc-siRNAs followed by therapeutic vaccination
using TherVacB reduced HBsAg and HBeAg to levels below
the detection limit by 5 and 4 log10, respectively, and triggered
seroconversion to anti-HBs and anti-HBe. Control of HBV
replication was maintained for >5 months after the last siRNA
dose. Mild, but long-term ALT elevation was observed after
start of vaccination lasting until the siRNA effect had ceased.
This suggests that by reducing antigen expression, siRNA
therapy could mitigate killing of hepatocytes by CD8 T cells
and thereby could attenuate liver damage. Conclusion: We
developed a combinatorial RNAi / therapeutic vaccination
therapy for hepatitis B that is achieving long-term functional
HBV cure in a preclinical mouse model, suggesting potential
for clinical translation. |
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发表于 2018-10-6 19:05
