Entecavir combined with furin inhibitor simultaneously reduces hepatitis B virus replication and e antigen secretion.

BACKGROUND: The antiviral therapy of chronic hepatitis B virus (HBV) infection pursues the dual goals, virological response (undetectable serum HBV DNA) and hepatitis B e antigen (HBeAg) serological response (serum HBeAg loss/seroconversion). It is relatively difficult, however, to realize the serological response, especially for nucleotide/nucleoside analogs. Furin, a proprotein convertase, is involved in HBeAg maturation. The suppression of furin using inhibitors accordingly reduces HBeAg secretion, but possibly enhances HBV replication. For these reasons, the strategy based on the combination of nucleoside analog entecavir (ETV) and furin inhibitors to inhibit HBV replication and HBeAg secretion simultaneously were studied here. METHODS: The suppression of furin was performed using inhibitors decanoyl-RVKR-chloromethylketone (CMK) and hexa-D-arginine (D6R) or the expression of furin inhibitory prosegment. The influence of furin suppression on HBV replication and the effect of CMK combined with nucleoside analog entecavir (ETV) on HBV replication and HBeAg secretion was investigated in HepG2.2.15 cells. HBeAg level in media was detected using enzyme-linked immunosorbent assay. Intracellular viral antigens and HBV DNA were detected using Western and Southern blotting analyses, respectively. RESULTS: CMK, D6R and the expression of inhibitory prosegment all significantly reduced HBeAg secretion, but only CMK enhance HBV replication. Concordantly, only CMK post-transcriptionally accumulated cytosolic HBV replication-essential hepatitis B core antigen (HBcAg). The HBcAg-accumulating effect of CMK was further found to be resulted from its redundant inhibitory effect on the trypsin-like activity of cellular proteasomes that are responsible for HBcAg degradation. Moreover, the viral replication-enhancing effect of CMK was abrogated by ETV and ETV combined with CMK reduced HBV replication and HBeAg secretion simultaneously. CONCLUSION: The suppression of furin itself does not enhance HBV replication. Nucleotide/nucleoside analogs combined with furin inhibitors may be a potential easy way to realize the dual goals of the antiviral therapy for chronic hepatitis B in the future.

Therapeutic potential of furin inhibitors for the chronic infection of hepatitis B virus.

BACKGROUND & AIMS: Hepatitis B e antigen (HBeAg) is essential for the development of chronic hepatitis B virus (HBV) infection. Furin, a proprotein convertase, plays a key role in processing of HBeAg precursor into maturated HBeAg. For these reasons, the therapeutic potential of furin inhibition for chronic HBV infection was studied. METHODS: The effects of furin inhibitor I (decanoyl-RVKR-chloromethylketone, CMK) and furin inhibitor II (hexa-D-arginine, D6R) on HBeAg secretion, the destination of unprocessed precursor and cellular secretory functions were comparatively investigated. RESULTS: CMK and D6R significantly decreased the supernatant level of HBeAg and increased the intracellular level of HBeAg precursor in HepG2.2.15 cells in vitro. The accumulated HBeAg precursor was not found to be retro-transported into the cytosol to inhibit HBV replication as expected, but was found to be expressed on the cell surface, where it may be more convenient to mediate host immune responses. Furthermore, these inhibitors at effective concentrations were not found to interfere with the maturations of albumin and prothrombin. Compared with CMK, D6R was suboptimal in effectiveness; however, D6R neither enhanced HBV replication through the accumulation of cytosolic HBcAg nor did it cause severe cell damage in an elongated safety analyses. CONCLUSION: Furin inhibitors CMK and D6R reduce HBeAg secretion and increase cell surface expression of the HBeAg precursor in HepG2.2.15 cells. Novel furin inhibitors or modified forms of D6R may promote the reduction of immune tolerance and the elimination of infected hepatocytes in patients with chronic HBV infection.