Inhibition of hepatitis B virus DNA replication by a thermostable interferon-γ variant.
BACKGROUND
Treatment of HBV chronic carriers using interferon (IFN)-α or nucleoside/nucleotide analogues fails to suppress viral infection. Type-II IFN-γ has been shown to inhibit HBV replication. The goal of the present work was to evaluate the antiviral efficacy against HBV of a thermostable IFN-γ variant isolated using Massive Mutagenesis and thermoresistant selection (THR) technologies.


METHODS
The thermostability of wild-type (wt) and S63C IFN-γ was determined in vitro and in vivo. Activation of the IFN-γ responsive element by wt and S63C IFN-γ was tested using a luciferase assay. HepG2.2.15 cells constitutively expressing HBV were used to analyse the antiviral activity of wt and S63C IFN-γ against HBV replication. Intracellular HBV DNA was detected by Southern blot and quantified by real-time PCR analyses.


RESULTS
S63C IFN-γ was shown to be more thermostable and had a longer half-life than wt IFN-γ. Both wt and S63C IFN-γ displayed a similar capacity to activate the IFN pathway. The treatment of HepG2.2.15 cells with wt or S63C IFN-γ induced the inhibition of HBV viral replication. After heating, S63C IFN-γ displayed better conservation of its antiviral activity against HBV when compared with wt IFN-γ.


CONCLUSIONS
These results confirm that the THR method can be used to isolate mutants with enhanced thermostability and demonstrate that a thermostable IFN-γ variant presents antiviral properties against HBV replication. This molecule could provide a new strategy to treat patients who do not respond to antiviral therapy.
Brunelle
Marie Noëlle
MN
INSERM U, Paris, France.
Saboulard
Didier
D
Massinet
Hélène
H
Lamant
Céline
C
Soussan
Patrick
P
Brezillon
Nicolas
N
Kremsdorf
Dina
D
D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology; D004261:DNA Replication; D004279:DNA, Viral / Q000096:biosynthesis; D005260:Female; D006207:Half-Life; D006367:HeLa Cells; D056945:Hep G2 Cells; D006515:Hepatitis B virus / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D006801:Humans; D007371:Interferon-gamma / Q000235:genetics / Q000494:pharmacology*; D051379:Mice; D008810:Mice, Inbred C57BL; D011994:Recombinant Proteins; D014779:Virus Replication
D000998:Antiviral Agents; D004279:DNA, Viral; D011994:Recombinant Proteins; D007371:Interferon-gamma