The Activity Evaluation And Mechanism Exploration Of Novel Nonnucleoside Anti-HBV Agents

Hepatitis B virus(HBV) infection is a most common chronic viral infection in the world. Several antiviral agents, such as(Pegylated-) interferon-α and nucleos(t)ide analogs are approved by FDA for chronic HBV infection management. But the cost and need for essentially life-long treatment are considerable challenges. And none of these current treatments can eradicate the intracellular virus. Given these issues, there is still an unmet medical need for an efficient HBV cure.Here we first identified a novel pyridazinone derivative, compound 3711, as a nonnucleosidic hepatitis B virus(HBV) inhibitor in cell model system. 3711 decreased extracellular HBV DNA levels by 50%(50% inhibitory concentration [IC50]) at 1.5 ± 0.2 μM and intracellular DNA levels at 1.9 ± 0.1 μM, which demonstrated antiviral activity at levels far below those associated with toxicity. Both the 3TC/ETV dually resistant L180M/M204 I mutant and the adefovir(ADV)-resistant A181T/N236 T mutant were as susceptible to 3711 as wild-type HBV. 3711 treatment induced the formation of genome-free capsids, a portion of which migrated faster on 1.8% native agarose gel. The induced genome-free capsids sedimented more slowly in isopycnic Cs Cl gradient centrifugation without significant morphological changes. 3711 treatment decreased levels of HBV DNA contained in both secreted enveloped virion and naked virus particles in supernatant. 3711 could interfere with capsid formation of the core protein(Cp) assembly domain. A Cp V124 W mutant, which strengthens capsid interdimer interactions, recapitulated the effect of 3711 on capsid assembly. Moreover, we are still working on the structural optimazation of pyridazinone derivatives, and we have identified compound 5045 with better solubility and compounds 5165, 5174 with better anti-HBV activity.Here we also compare the mechanism of 3711 with a drug candidate, Isothiafludine(NZ-4), as HBV inhibitors. Although both compounds inhibit HBV replication by inducing genome-free capsid formation in cell culture system, NZ-4’s activity was dependent on the presence of the ARD I of core protein.In conclusion, we first identify pyridazinone derivatives as HBV inhibitors, which act by interfering with HBV capsid formation. Pyridazinone derivatives can not only be used as tool drugs to investigate HBV capsid formation process, but also may provide a new opportunity to combat chronic HBV infection. NZ-4, a drug candidate whose phase I clinical trials is now being evaluated in China, is currently the only compound that inhibit HBV replication depending on the presence of the ARD I of core protein. The unique mechanism of NZ-4 might make it a possible option for HBV cure in the future.