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Abstract The HBV core protein (HBc) is an important viral protein of HBV that plays an indispensable role in the lifecycle of HBV, including capsid assembly and transport, reverse transcription and virus release. In recent years, evidence has shown that HBc may be involved in the malignant progression of HCC. Thus, HBc is an attractive target for antiviral agents and provides a new strategy for the treatment of HBV-related HCC. Here, we identified a novel anti‐HBc compound—colchicine, an alkaloid compound—that promoted selective autophagic degradation of HBc through the AMPK/mTOR/ULK1 signalling pathway. We further confirmed that colchicine promoted the selective autophagy of HBc by enhancing the binding of HBc to the autophagy receptor p62. Finally, we evaluated the effects of colchicine on HBV replication and HBc-mediated HCC metastasis in vitro and in vivo. Our research indicated that the inhibitory effects of colchicine on HBV and HBV-related HCC depend on the selective autophagic degradation of HBc. Thus, colchicine is not only a promising therapeutic strategy for chronic hepatitis B but also a new treatment for HBV-related HCC.

Abstract Background Progressive hepatitis B virus (HBV) infection can result in cirrhosis, hepatocellular cancer, and chronic hepatitis. While antiviral drugs that are now on the market are efficient in controlling HBV infection, finding a functional cure is still quite difficult. Identifying host factors involved in regulating the HBV life cycle will contribute to the development of new antiviral strategies. Zinc finger proteins have a significant function in HBV replication, according to earlier studies. Zinc finger protein 148 (ZNF148), a zinc finger transcription factor, regulates the expression of various genes by specifically binding to GC-rich sequences within promoter regions. The function of ZNF148 in HBV replication was investigated in this study. Methods HepG2-Na+/taurocholate cotransporting polypeptide (HepG2-NTCP) cells and Huh7 cells were used to evaluate the function of ZNF148 in vitro. Northern blotting and real-time PCR were used to quantify the amount of viral RNA. Southern blotting and real-time PCR were used to quantify the amount of viral DNA. Viral protein levels were elevated, according to the Western blot results. Dual-luciferase reporter assays were used to examine the transcriptional activity of viral promoters. ZNF148’s impact on HBV in vivo was investigated using an established rcccDNA mouse model. Results ZNF148 overexpression significantly decreased the levels of HBV RNAs and HBV core DNA in HBV-infected HepG2-NTCP cells and Huh7 cells expressing prcccDNA. Silencing ZNF148 exhibited the opposite effects in both cell lines. Furthermore, ZNF148 inhibited the activity of HBV ENII/Cp and the transcriptional activity of cccDNA. Mechanistic studies revealed that ZNF148 attenuated retinoid X receptor alpha (RXRα) expression by binding to the RXRα promoter sequence. RXRα binding site mutation or RXRα overexpression abolished the suppressive effect of ZNF148 on HBV replication. The inhibitory effect of ZNF148 was also observed in the rcccDNA mouse model. Conclusions ZNF148 inhibited HBV replication by downregulating RXRα transcription. Our findings reveal that ZNF148 may be a new target for anti-HBV strategies.

Abstract Persistent transcription of HBV covalently closed circular DNA (cccDNA) is critical for chronic HBV infection. Silencing cccDNA transcription through epigenetic mechanisms offers an effective strategy to control HBV. Long non‐coding RNAs (lncRNAs), as important epigenetic regulators, have an unclear role in cccDNA transcription regulation. In this study, lncRNA sequencing (lncRNA seq) is conducted on five pairs of HBV‐positive and HBV‐negative liver tissue. Through analysis, HOXA‐AS2 (HOXA cluster antisense RNA 2) is identified as a significantly upregulated lncRNA in HBV‐infected livers. Further experiments demonstrate that HBV DNA polymerase (DNA pol) induces HOXA‐AS2 after establishing persistent high‐level HBV replication. Functional studies reveal that HOXA‐AS2 physically binds to cccDNA and significantly inhibits its transcription. Mechanistically, HOXA‐AS2 recruits the MTA1‐HDAC1/2 deacetylase complex to cccDNA minichromosome by physically interacting with metastasis associated 1 (MTA1) subunit, resulting in reduced acetylation of histone H3 at lysine 9 (H3K9ac) and lysine 27 (H3K27ac) associated with cccDNA and subsequently suppressing cccDNA transcription. Altogether, the study reveals a mechanism to self‐limit HBV replication, wherein the upregulation of lncRNA HOXA‐AS2, induced by HBV DNA pol, can epigenetically suppress cccDNA transcription.

Background & Aims: Current antiviral drugs, including nucleoside analogs and interferon, fail to eliminate the HBV covalently closed circular DNA (cccDNA), which serves as a transcript template in infected hepatocytes. Silencing the HBV X protein, which plays a crucial role in cccDNA transcription, is a promising approach to inhibit HBV replication. Therefore, the identification of novel compounds that can inhibit HBx-mediated cccDNA transcription is critical. Methods: Initially, a compound library consisting of 715 monomers derived from traditional Chinese medicines known for their liver-protecting properties was established. Then, MTT assays were used to determine the cytotoxicity of each compound. The effect of candidates on Flag-HBx expression was examined by real-time PCR and western blotting in Flag-HBx transfected HepG2-NTCP cells. Ultimately, the antiviral effect of gambogic acid (GA) on HBV was observed in HBV-infected HepG2-NTCP cells. Mechanistically, the functional role of DTX1 in GA-induced HBV inhibition was examined using RNA-seq. Finally, the antiviral effect of GA was estimated in vivo. Results: Gambogic acid (GA), a natural bioactive compound with a myriad of biological activities, markedly reduced Flag-HBx expression. Potent and dose-dependent reductions in extracellular HBV RNAs, HBV DNA, HBsAg, HBeAg and HBc protein were discovered three days after GA treatment in HBV-infected cells, accompanied by the absence of significant cytotoxicity. Furthermore, our research revealed that GA exhibited a dose-dependent inhibition of HBx expression, which is a pleiotropic protein required for HBV infection in vivo. We explored the mechanisms underlying GA-mediated inhibition of HBV and confirmed that this inhibition is accomplished by upregulating the expression of the DTX1 gene and boosting the Notch signaling pathway. Finally, the inhibitory effect of GA on HBV replication was tested in vivo using a mouse model of hepatitis B virus recombinant cccDNA. Conclusions: Herein, we discovered GA, which is a natural bioactive compound that targets HBx to inhibit hepatitis B virus replication by activating the DTX1-Notch signaling pathway.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths globally. Tumor metastasis is one of the major causes of high mortality of HCC. Identifying underlying key factors contributing to invasion and metastasis is critical to understand the molecular mechanisms of HCC metastasis. Here, we identified RNA binding protein L23 (RPL23) as a tumor metastasis driver in HCC. RPL23 was significantly upregulated in HCC tissues compared to adjacent normal tissues, and closely related to poor clinical outcomes in HCC patients. RPL23 depletion inhibited HCC cell proliferation, migration and invasion, and distant metastasis. Mechanistically, RPL23 directly associated with 3’UTR of MMP9, therefore positively regulated MMP9 expression. In conclusion, we identified that RPL23 might play an important role in HCC metastasis in an MMP9-dependent manner and be a potential therapeutic target for HCC tumorigenesis and metastasis.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths globally. Tumor metastasis is one of the major causes of high mortality of HCC. Identifying underlying key factors contributing to invasion and metastasis is critical to understand the molecular mechanisms of HCC metastasis. Here, we identified RNA binding protein L23 (RPL23) as a tumor metastasis driver in HCC. RPL23 was significantly upregulated in HCC tissues compared to adjacent normal tissues, and closely related to poor clinical outcomes in HCC patients. RPL23 depletion inhibited HCC cell proliferation, migration and invasion, and distant metastasis. Mechanistically, RPL23 directly associated with 3'UTR of MMP9, therefore positively regulated MMP9 expression. In conclusion, we identified that RPL23 might play an important role in HCC metastasis in an MMP9-dependent manner and be a potential therapeutic target for HCC tumorigenesis and metastasis. [ABSTRACT FROM AUTHOR]