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P0529
ANTI CAPSID DRUGS HAP12 AND AT130 TARGET
HBV CORE PROTEIN NUCLEAR FUNCTIONS
L. Belloni1,2, G.A. Palumbo2, L. Lupacchini2, L. Li3, S. Reddy Chirapu4,
L. Calvo2, M.G. Finn4, U. Lopatin5, A. Zlotnick3,5, M. Levrero1.
1Center for Life Nano Science (CLNS), IIT-Sapienza University, 2Dept of
Internal Medicine – DMISM, Sapienza University, Rome, Italy; 3Dept of
Molecular & Cellular Biochemistry, Indiana University, Bloomington,
4Dept Chemistry, Georgia Institute of Technology, Atlanta, 5Assembly
Bioscience, Bloomington, United States
E-mail: [email protected]
Background and Aims: HBV Core protein (Cp) represents an
attractive new therapeutic target for HBV chronic infection. In
addition to their role in capsid assembly, pgRNA packaging and
reverse transcription, Cp has been shown to bind the nuclear
cccDNA mini-chromosome as well as a number of cellular genes
promoters. Several compounds that target Cp and HBV capsids
assembly, including the Hetero-aryl-dihydropyrimidines (HAPs) and
the phenyl-propenamide derivatives AT61 and AT130, have been
shown to inhibit HBV replication in vitro and in vivo. HAPs and
AT130 enhance the rate and the extent of Cp assembly leading
to non-functional capsids and, at high concentration, stabilize
preferentially non-capsid polymers of Cp. Here we investigated the
ability of the Core protein Assembly Modulators (CaMPs) HAP12 and
AT130 to affect both nuclear (cccDNA structure and transcription)
and cytoplasmic (capsid maturation and replication) Cp functions
as part of their antiviral activity against HBV.
Methods: HAP12 and AT130 effects on capsid-associated HBV-DNA
(TaqMan real-time PCR), cccDNA (TaqMan real-time PCR)
and pgRNA levels (quantitative real-time PCR with specific
primers), were assessed in: (a) HBV-infected NTCP-HepG2 cells;
(b) AD38 inducible HBV stable cell line. Recruitment of HBc and
histone modifications on the viral minichromosome were assessed
using the cccDNA ChIP assay in AD38 cells.
Results: CaMPs treatments started at day 6 post-infection (NTCPHepG2)
or day 6 post-induction (AD38 tet-off cells) resulted in a
very strong inhibition of HBV replication (>95%) and a significant
but incomplete reduction of the stable cccDNA pool. A strong
effect on cccDNA-dependent HBeAg production (AD38 tet-off)
and pgRNA transcription (AD38 tet-off / tet-on and NTCP-HepG2
infected cells) was also demonstrated. The ability HAP12 to target
cccDNA transcription was confirmed by the reduced cccDNA-bound
H3 histone acetylation and the descreased HBc occupancy on the
cccDNA in induced AD38 cells. Importantly, when CaMPs treatment
was started during infection, cccDNA formation/accumulation was
completely inhibited (>95%) and viral replication was blunted.
Conclusions: Anti-capsid compounds (CpAMs) have an impact on
Cp nuclear functions at multiple levels: block of new cccDNA
formation / accumulation, reduction of an established cccDNA pool
and inhibition of HBc occupancy and histone acetylation on the
cccDNA that translate into a reduced pgRNA transcription
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发表于 2015-4-22 18:41
