Taking berberine (BBR) as the lead, 23 new BBR derivatives were synthesized and examined for their antiviral activities against four different genotype enterovirus 71 (EV71) strains with a cytopathic effect (CPE) assay. Structure-activity relationship (SAR) studies indicated that introduction of a suitable substituent at the 9-position might be beneficial for potency. Among them, compound 2d exhibited most potent activities with IC₅₀ values of 7.12–14.8 μM, similar to that of BBR. The effect of 2d was further confirmed in a dose-dependent manner both in RNA and protein level. The mechanism revealed that 2d could inhibit the activation of MEK/ERK signaling pathway. Meanwhile, it could suppress the EV71-induced autophagy by activating AKT and inhibiting the phosphorylation of JNK and PI3KIII proteins. We consider BBR derivatives to be a new family of anti-EV71 agents through targeting host components, with an advantage of broad-spectrum anti-EV71 potency.

Chronic hepatitis induced by hepatitis B virus (HBV) infection is a serious public health problem, leading to hepatic cirrhosis and liver cancer. Although the currently approved medications can reliably decrease the virus load and prevent the development of hepatic diseases, they fail to induce durable off-drug control of HBV replication in the majority of patients. The roots of Isatis indigotica Fortune ex Lindl., a traditional Chinese medicine, were frequently used for the prevention of viral disease in China. In the present study, (−)-lariciresinol ((−)-LRSL), isolated from the roots of Isatis indigotica Fortune ex Lindl., was found to inhibit HBV DNA replication of both wild-type and nucleos(t)ide analogues (NUCs)-resistant strains in vitro. Mechanism studies revealed that (−)-LRSL could block RNA production after treatment, followed by viral proteins, and then viral particles and DNA. Promoter reporter assays and RNA decaying dynamic experiments indicated that (−)-LRSL mediated HBV RNA reduction was mainly due to transcriptional inhibition rather than degradation. Moreover, (−)-LRSL in a dose-dependent manner also inhibited other animal hepadnaviruses, including woodchuck hepatitis virus (WHV) and duck hepatitis B virus (DHBV). Combining the analysis of RNA-seq, we further found that the decrease in HBV transcriptional activity by (−)-LRSL may be related to hepatocyte nuclear factor 1α (HNF1α). Taken together, (−)-LRSL represents a novel chemical entity that inhibits HBV replication by regulating HNF1α mediated HBV transcription, which may provide a new perspective for HBV therapeutics.

Chronic hepatitis B virus (HBV) infection remains a global public health challenge, and the currently approved medications can not achieve a cure. Synthetic triterpenoids have shown promising therapeutic potential for liver pathologies. In our search for novel antiviral agents against HBV, we found that two triterpenoids, 2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid (CDDO) and CDDO-ethyl amide (CDDO-EA), significantly inhibited HBV DNA replication. Further mechanistic investigation indicated that these two compounds did not significantly alter the levels of total HBV pgRNA, but dramatically reduced extracellular pgRNA and intracellular encapsidated pgRNA in a dose-dependent manner. Western blot analysis indicated minimal effects on core protein expression. Interestingly, using a particle gel assay, we observed that CDDO and CDDO-EA promoted the formation of empty capsids with no alteration in electrophoretic mobility. Moreover, we demonstrated that both compounds modulated the phosphorylation status of the core protein. Further cellular thermal shift assay (CETSA), surface plasmon resonance (SPR) assay, and molecular docking analyses collectively suggested that CDDO and CDDO-EA could bind directly to the dimer–dimer interfaces of HBV core protein. Finally, a synergistic effect was observed between CDDO-EA and lamivudine in reducing intracellular and extracellular HBV DNA levels. Our findings indicate that triterpenoids CDDO and CDDO-EA are new mechanistically type of HBV capsid assembly modulators and warranted for further development as lead compounds against HBV.