A recent study has shown a higher frequency
of an A3B deletion allele in persistent HBV
carriers and hepatocellular carcinoma patients
relative to healthy controls (25). This finding was
further supported by the moderate deamination
of cccDNA even in the absence of treatment, and
by the observation that knockdown of A3B in the
absence of any treatment increased cccDNA levels. Although deregulated expression of A3A and
A3B has been shown to correlate with genomic
DNA mutations (39, 40), we did not detect any
alterations of genomic DNA using analyses of AP
sites, 3D-PCR analysis, and deep sequencing of
a set of human genes.
Our data indicate that cccDNA degradation is
possible and can be induced without side effects
on the infected host cell. An important task will
be the testing of combinations of nucleoside or
nucleotide analogs with novel antiviral strategies
[e.g., LTbR agonists or adoptive T cell therapy
(41)] to activate A3A or A3B to cure hepatitis B.
References and Notes
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Fig. 6. Interaction of A3A,
HBV core protein (HBc),
and cccDNA. (A) Chromatin
was performed using lysates
of HepG2-H1.3 cells transfected
with A3A-expressing plasmid
or HBV-infected dHepaRG
cells treated with IFN-a for
3 days. IPs using antibodies
against histone H3, A3A, HBc,
and control rabbit IgG (RIgG)
were analyzed by qPCR for
cccDNA. (B) Interaction between HBc and A3A was assessed by proximity ligation
assay (PLA) in HBV-infected,
IFN-a–treated dHepaRG cells.
PLA spots were quantified in
single cells by software-based
spot counting. Data were analyzed by one-way analysis
of variance. **P < 0.01,
***P < 0.001. (C) Serial HBV
core deletion mutants (left)
were fused to cyan fluorescent
protein (CFP), and interaction
with A3A-YFP was assessed
by fluorescence-activated cell
sorting and FRET in HuH7.5
hepatoma cells (right). Cells
cotransfected with CFP and
yellow fluorescent protein (YFP) served as controls to exclude false
positive FRET and subtract background signals. A CFP-YFP fusion construct was used as positive control. Data are means T SD of FRET-positive
cells from three or four independent experiments. Black boxes indicate shared regions of HBc mutants giving a FRET signal. (D) Model of
cccDNA degradation induced by IFN-a treatment or LTbR activation.
IFNAR, type I IFN receptor.