Search Results (83)

Hepatocellular carcinoma (HCC), the most common primary liver malignancy worldwide, is strongly associated with chronic Hepatitis B Virus (HBV) infection, a significant risk factor. The ubiquitin–proteasome system, central to protein degradation, cellular homeostasis, and cell cycle regulation, has been implicated in the pathogenesis of several cancers, including HCC. Despite this, the specific expression patterns of proteasomal subunits during HBV infection and HBV-induced HCC, as well as the association between mRNA expression of proteasomal subunits and proteasomal activity, remain poorly defined. To address this critical knowledge gap, we analyzed mRNA expression profiles of proteasomal subunits in HBV-infected humanized mouse models to uncover HBV-specific molecular alterations. Our findings revealed that the chymotrypsin-like activity (β5) subunit of the proteasome (PSMB5) is consistently overexpressed following HBV infection. Functional studies demonstrated that β5 deficiency decreases MHC I levels on the cell surface and leads to the accumulation of ubiquitinated proteins, establishing a direct link between β5 overexpression and increased proteasomal activity. Concordantly, HBV-infected patient livers—regardless of HCC status—displayed elevated β5 mRNA/protein levels and enhanced chymotrypsin-like activity. Additionally, analysis of Protein Atlas data revealed that elevated β5 mRNA expression correlates with poor clinical prognosis in HCC patients. In summary, this study highlights how HBV infection induces significant alterations in proteasome function by elevating β5 expression and activity in human and mouse livers. These findings underscore the critical role of proteasomal dysregulation in HBV-associated liver pathology and provide new insights into its involvement in HCC development. Understanding the interplay between HBV infection and proteasome dynamics offers a valuable avenue for the identification of novel therapeutic targets and biomarkers in HCC. Full article

Background: Chronic hepatitis B (CHB) remains being a major public health threat, and currently existing CHB therapies have limited efficacy and side effects. We have recently developed a vaccine termed VVX001 based on a recombinant fusion protein consisting of the preS domain of the large surface protein of hepatitis B virus (HBV) fused to grass pollen allergen peptides. VVX001 has been shown to induce preS-specific antibodies in grass pollen allergic patients, and sera of immunized subjects inhibited HBV infection in vitro. Methods: In this study we investigated if immunization with VVX001 can induce preS-specific antibodies in CHB using the adeno-associated virus (AAV)-HBV murine model of CHB. Six groups of C57BL/6 female mice (n = 6) were transduced with AAV-HBV or AAV-Empty, and after six weeks, they were immunized five times with 20 µg of aluminum hydroxide-adsorbed VVX001 or preS or vehicle (Alum alone). Serum samples were taken continuously. Two weeks after the last immunization, spleen and liver mononuclear cells were collected. Serum reactivity to preS and preS-derived peptides was assessed by ELISA. B-cell responses were measured by ELISPOT assay, and intrahepatic lymphocyte (ILH) counts were determined by FACS. HBV DNA, HBsAg, HBeAg, ALT, and AST were assessed using commercial kits. Results: Our results show that VVX001 induces preS-specific IgG antibodies that cross-react with different HBV genotypes A-H and are directed against the sodium taurocholate co-transporting polypeptide (NTCP) receptor binding site of preS both in mice with and without HBV. Actively immunized AAV-HBV-treated mice had a higher number of intrahepatic lymphocytes than vehicle-vaccinated and mock-transduced animals. Conclusions: These findings encourage performing further trials to study the potential of VVX001 for therapeutic vaccination against CHB. Full article

Liver cancer has high incidence and mortality rates worldwide, with hepatocellular carcinoma (HCC) being the main histological subtype, accounting for 90% of primary liver cancers. The high mutation rate of viruses combined with endoplasmic reticulum stress may lead to the occurrence of cancer. Hepatitis B virus (HBV) infection is one of the most important pathogenic factors of HCC. The carcinogenic mechanisms of HBV have been widely studied. Among these mechanisms, immune escape and vaccine escape caused by mutations in the HBV S gene have been reported in numerous studies of patients with chronic hepatitis B. In addition, pre-S1/S2 mutations and surface protein truncation mutations may activate multiple signaling pathways. This activation leads to the abnormal proliferation and differentiation of hepatocytes, thereby contributing to the development of HCC. This review aims to integrate the existing literature, summarize the common mutations in the HBV S gene region, and explore the related pathogenic mechanisms. Full article

Background: Hepatitis B virus (HBV) is a major cause of morbidity and mortality globally, and chronic infections are associated with cirrhosis and hepatocellular carcinoma. Issues with conventional treatments and vaccines mean there is a need for new therapeutic vaccines, which must elicit a strong and sustainable immune response. Here, we evaluated the immunogenicity of dual-antigen vaccines containing hybrid surface (hy-LHBs) and core (HBc) antigens, combined with a carboxyl-vinyl polymer (CVP) as a mucoadhesive excipient, following intranasal administration in mice. Methods: Mice were intranasally administered a mixed vaccine (10 µg of hy-LHBs and 2.5 or 10 µg of HBc) with or without a CVP excipient, and they were assessed for their immune response (levels of IgGs or IgA antibodies in an ELISA, IFN-γ level in splenocytes in an ELISpot assay, and cytokine/chemokine levels in a BioPlex assay). A protein stability assay was also conducted for vaccine formulations with and without excipients. Results: Significantly enhanced IgG production was noted targeting hy-LHBs and (less markedly) HBc at 10 µg/antigen, but only a non-significant elevation was noted with the vaccine containing 2.5 µg HBc. The BioPlex assay showed a significant increase in IL-2 (#00-07, 0B), IL-12(p40)(#00), eotaxin (#00), MIP1α (#00, #00-07, 0B), and MCP-1 (#00-07, 0B) in mice that received treatment compared to those of untreated mice. The endpoint titers of IgG1 and IgG2a were measured, which were higher with CVP excipients than without. From the IgG2a/IgG1 ratio, a higher IgG1 response was induced by CVPs to hy-LHBs and a higher IgG2a response was induced to HBc. Th2-dominant phenotype to hy-LHBs was induced with CVP#00 in an ELISpot assay. The highest anti-hy-LHBs antibody titer was noted with the conventional CVP#00 excipient. Consistent with these results, a higher amount of neutralizing antibodies of HBV was induced with CVP#00 treatment and followed by #00-03 and #14-00. Conclusions: We consider that the addition of CVP excipients to vaccine formulation enhances immunogenicity and HBV antigen stability for intranasal vaccines. This effect was seen for both humoral and cell-mediated immune responses, indicating the potential of CVPs as excipients in intranasal HBV vaccines. Full article

Hepatitis B virus (HBV) is an important global public health issue. The World Health Organization (WHO) 2024 Global Hepatitis Report estimated that the global prevalence of people living with HBV infection is 254 million, with an estimated prevalence incidence of 1.2 million new HBV infections yearly. Previous studies have shown that natural compounds have antiviral inhibition potentials. In silico methods such as molecular docking, virtual screening, pharmacophore modeling, quantitative structure–activity relationship (QSAR), and molecular dynamic simulations have been successfully applied in identifying bioactive compounds with strong binding energies in HBV treatment targets. The COVID-19 pandemic necessitated the importance of repurposing already approved drugs using in silico methods. This study is aimed at unveiling the benefits of in silico techniques as a potential alternative in natural compounds’ drug discovery and repurposing for HBV therapy. Relevant articles from PubMed, Google Scholar, and Web of Science were retrieved and analyzed. Furthermore, this study comprehensively reviewed the literature containing identified bioactive compounds with strong inhibition of essential HBV proteins. Notably, hesperidin, quercetin, kaempferol, myricetin, and flavonoids have shown strong binding energies for hepatitis B surface antigen (HBsAg). The investigation reveals that in silico drug discovery methods offer an understanding of the mechanisms of action, reveal previously overlooked viral targets (including PreS1 Domain of HBsAg and cccDNA (Covalently Closed Circular DNA) regulators, and facilitate the creation of specific inhibitors. The integration of in silico, in vitro, and in vivo techniques is essential for the discovery of new drugs for HBV therapy. The insights further highlight the importance of natural compounds and in silico methods as targets in drug discovery for HBV therapy. Moreover, the combination of natural compounds, an in silico approach, and drug repurposing improves the chances of personalized and precision medicine in HBV treatment. Therefore, we recommend drug repurposing strategies that combine in vitro, in vivo, and in silico approaches to facilitate the discovery of effective HBV drugs. Full article

Capsid assembly modulators (CAMs) are a novel class of antiviral agents in clinical development for the treatment of chronic hepatitis B. CAMs inhibit hepatitis B virus (HBV) replication by binding to a hydrophobic pocket, i.e., HAP pocket, between HBV capsid protein (Cp) dimer–dimer interfaces to misdirect its assembly into empty capsids or aberrant structures and designated as CAM-E and CAM-A, respectively. Because the emergence of CAM-resistant variants results in the failure of antiviral therapy, it is important to rationally design CAMs with a high barrier of resistance for development. To establish computational approaches for the prediction of Cp mutations that confer resistance to CAMs, we investigated the interaction of representative CAM-A and CAM-E compounds, BAY 41-4109 and JNJ-56136379, with wild-type and 35 naturally occurring mutations of Cp residues at the HAP pocket using molecular docking, prime molecular mechanics with generalized Born and surface area solvation (MM/GBSA) and molecular dynamics (MD) simulation methods. Out of nine publicly available HBV capsid or CpY132A hexamer structures in the protein database, molecular docking correctly predicted the resistance and sensitivity of more than 50% Cp mutations to JNJ-56136379 with structures 5D7Y and 5T2P-FA. MM/GBSA correctly predicted the resistance and sensitivity of more than 50% Cp mutations to BAY41-4109 with the structures 5E0I-BC and 5WRE-FA, and to JNJ-56136379 with the 5E0I-FA structure. Our work indicates that only the capsid or CpY132A hexamer structure bound with a CAM with similar chemical scaffold can be used for more accurately predicting the resistance and sensitivity of Cp mutations to a CAM molecule under investigation by molecular docking and/or MM/GBSA methods. Full article

Background/Objectives: In chronic hepatitis B infection (CHB), the hepatitis B surface antigen (HBsAg) continuously exhausts the hepatitis B surface antibody (HBsAb), which leads to the formation of immune tolerance. Accordingly, the hepatitis B virus (HBV) infection can be blocked by inhibiting the binding of the hepatitis B surface pre-S1/pre-S2 antigen to the hepatocyte receptor NTCP, but the clinical cure rate of pre-S-based vaccines for CHB is limited. Methods: In this study, we designed and prepared multivalent hepatitis B therapeutic mRNA vaccines encoding three hepatitis B surface antigen proteins (L, M, and S) at the cell membrane, verified via in vitro transfection and expression experiments. An in vivo immunization experiment in HBV transgenic (Tg) mice was first completed. Subsequently, an adeno-associated virus plasmid vector carrying the HBV1.2-fold genome (pAAV HBV1.2) model and the adeno-associated virus vector carrying HBV1.3-fold genome (rAAV HBV1.3) model were constructed and immunized with mRNA vaccines. The HBV antigen, antibodies, and HBV DNA in serum were detected. Indirect (enzyme-linked immunosorbent assay) ELISA were made to analyze the activated antigen-specific IgG in HBV Tg mice. Antigen-dependent T-cell activation experiments were carried out, as well as the acute toxicity tests in mice. Results: The L protein/pre-S antigens could be stably presented at the cell membrane with the support of the S protein (and M protein). After vaccinations, the vaccines effectively reactivated the production of high levels of HBsAb, disrupted immune tolerance, and activated the production of high-affinity antibodies against structural pre-S antigen in HBV Tg mice. The HBsAg seroconversion and serum HBV DNA clearance were achieved in two HBV mice models. Furthermore, pre-S antigen-dependent T-cell response against HBV infection was confirmed. The therapeutic vaccine also showed safety in mice. Conclusions: A novel therapeutic mRNA vaccine was developed to break through HBsAg-mediated immune tolerance and treat CHB by stably presenting the pre-S antigen at the membrane, and the vaccine has great potential for the functional cure of CHB. Full article

Since the discovery of the Australia antigen, now known as the hepatitis B surface antigen (HBsAg), significant research has been conducted to elucidate its physical, chemical, structural, and functional properties. Subviral particles (SVPs) containing HBsAg are highly immunogenic, non-infectious entities that have not only revolutionized vaccine development but also provided critical insights into HBV immune evasion and viral assembly. Recent advances in cryo-electron microscopy (cryo-EM) have uncovered the heterogeneity and dynamic nature of spherical HBV SVPs, emphasizing the essential role of lipid–protein interactions in maintaining particle stability. In this review, recent progress in understanding the molecular architecture of HBV SVPs is consolidated, focusing on their symmetry, lipid organization, and disassembly–reassembly dynamics. High-resolution structural models reveal unique lipid arrangements that stabilize hydrophobic residues, preserve antigenicity, and contribute to SVP functionality. These findings highlight the significance of hydrophobic interactions and lipid–protein dynamics in HBV SVP assembly and stability, offering valuable perspectives for optimizing SVP-based vaccine platforms and therapeutic strategies. Full article

Background: Approximately 10–20% of subjects vaccinated with HBsAg-based hepatitis B virus (HBV) vaccines are non-responders. BM32 is a recombinant grass pollen allergy vaccine containing the HBV-derived preS surface antigen as an immunological carrier protein. PreS includes the binding site of HBV to its receptor on hepatocytes. We investigated whether immunological non-responsiveness to HBV after repeated HBsAg-based vaccinations could be overcome by immunization with VVX001 (i.e., alum-adsorbed BM325, a component of BM32). Methods: A subject failing to develop protective HBV-specific immunity after HBsAg-based vaccination received five monthly injections of 20 µg VVX001. PreS-specific antibody responses were measured by enzyme-linked immunosorbent assay (ELISA) and micro-array technology. Serum reactivity to subviral particles of different HBV genotypes was determined by sandwich ELISA. PreS-specific T cell responses were monitored by carboxyfluorescein diacetate succinimidyl ester (CFSE) staining and subsequent flow cytometry. HBV neutralization was assessed using cultured HBV-infected HepG2 cells. Results: Vaccination with VVX001 induced a strong and sustained preS-specific antibody response composed mainly of the IgG₁ subclass. PreS-specific IgG antibodies were primarily directed to the N-terminal part of preS containing the sodium taurocholate co-transporting polypeptide (NTCP) attachment site. IgG reactivity to subviral particles as well as to the N-terminal preS-derived peptides was comparable for HBV genotypes A–H. A pronounced reactivity of CD3⁺CD4⁺ lymphocytes specific for preS after the complete injection course remaining up to one year after the last injection was found. Maximal HBV neutralization (98.4%) in vitro was achieved 1 month after the last injection, which correlated with the maximal IgG reactivity to the N-terminal part of preS. Conclusions: Our data suggest that VVX001 may be used as a preventive vaccination against HBV even in non-responders to HBsAg-based HBV vaccines. Full article

Hepatitis B virus (HBV) infection remains a major health threat with limited treatment options. One of various new antiviral strategies is based on a fusion of Staphylococcus aureus nuclease (SN) with the capsid-forming HBV core protein (HBc), termed coreSN. Through co-assembly with wild-type HBc-subunits, the fusion protein is incorporated into HBV nucleocapsids, targeting the nuclease to the encapsidated viral genome. However, coreSN expression was based on transfection of a plasmid vector. Here, we explored whether introducing protein transduction domains (PTDs) into a fluorescent coreSN model could confer cell-penetrating properties for direct protein delivery into cells. Four PTDs were inserted into two different positions of the HBc sequence, comprising the amphiphilic translocation motif (TLM) derived from the HBV surface protein PreS2 domain and three basic PTDs derived from the Tat protein of human immunodeficiency virus-1 (HIV-1), namely Tat4, NP, and NS. To directly monitor the interaction with cells, the SN in coreSN was replaced with the green fluorescent protein (GFP). The fusion proteins were expressed in E. coli, and binding to and potential uptake by human cells was examined through flow cytometry and fluorescence microscopy. The data indicate PTD-dependent interactions with the cells, with evidence of uptake in particular for the basic PTDs. Uptake was enhanced by a triplicated Simian virus 40 (SV40) large T antigen nuclear localization signal (NLS). Interestingly, the basic C terminal domain of the HBV core protein was found to function as a novel PTD. Hence, further developing cell-permeable viral capsid protein fusions appears worthwhile. Full article

Identifying correlations between immune responses elicited via HIV and non-HIV vaccines could aid the search for correlates of HIV protection and increase statistical power in HIV vaccine-efficacy trial designs. An exploratory objective of the HVTN 097 phase 1b trial was to assess whether immune responses [focusing on those supported as correlates of risk (CoR) of HIV acquisition] induced via the RV144 pox-prime HIV vaccine regimen correlated with those induced via tetanus toxoid (TT) and/or hepatitis B virus (HBV) vaccines. We measured TT-specific and HBV-specific IgG-binding antibody responses and TT-specific and HBV-specific CD4+ T-cell responses at multiple time points in HVTN 097 participants, and we assessed their correlations at peak time points with HIV vaccine (ALVAC-HIV and AIDSVAX B/E)-induced responses. Four correlations were significant [false discovery rate-adjusted p-value (FDR) ≤ 0.2]. Three of these four were with IgG-binding antibody responses to TT measured one month after TT receipt, with the strongest and most significant correlation [rho = 0.368 (95% CI: 0.096, 0.588; p = 0.008; FDR = 0.137)] being with IgG-binding antibody responses to MN gp120 gDneg (B protein boost) measured two weeks after the second ALVAC-HIV and AIDSVAX B/E boost. The fourth significant correlation [(rho = 0.361; 95% CI: 0.049, 0.609; p = 0.021; FDR = 0.137)] was between CD4+ T-cell responses to a hepatitis B surface antigen peptide pool, measured 2 weeks after the third HBV vaccination, and IgG-binding antibody responses to gp70BCaseAV1V2 (B V1V2 immune correlate), measured two weeks after the second ALVAC-HIV and AIDSVAX B/E boost. These moderate correlations imply that either vaccine, TT or HBV, could potentially provide a moderately useful immunogenicity predictor for the ALVAC-HIV and AIDSVAX B/E HIV vaccine regimen. Full article

Background and Objectives: The measurement of hepatitis B surface antigen (HBsAg) is essential for managing chronic hepatitis B virus infection (CHB). HBsAg consists of three different surface envelope proteins: large, middle, and small HB surface proteins. However, in clinical practice, it is not common to evaluate each of these HB surface proteins separately. Materials and Methods: In this study, we investigated preS1 expression using seven monoclonal antibodies (mAbs) in 68 CHB patients, as well as examining their antigenicity. Results: Although the seven mAbs had been derived from genotype (Gt) C, they could recognize preS1 with Gts A to D. The epitopes were concentrated within the aa33-47 region of preS1, and their antigenicity was significantly reduced by an aa45F substitution. We found that preS1 expression remained consistent regardless of HBsAg levels and different Gts in CHB patients, in contrast to what was observed in SHBs. Conclusions: These results suggest that the antigenic epitope is preserved among different Gts and that the expression pattern of preS1 is altered during CHB, highlighting its vital role in the HBV infection cycle. Our present results suggest preS1 is a promising therapeutic target in CHB. Full article

Liver damage can progress through different stages, resulting in cirrhosis or hepatocellular carcinoma (HCC), conditions that are often associated with viral infections. Globally, 42% and 21% of cirrhosis cases correlate with HBV and HCV, respectively. In the Americas, the prevalence ranges from 1% to 44%. The WHO has the goal to eliminate viral hepatitis, but it is important to consider occult HBV infection (OBI), a clinical condition characterized by the presence of HBV genomes despite negative surface antigen tests. This review aims to provide an overview of recent data on OBI, focusing on its role in the development of hepatic diseases and its significance in the WHO Viral Hepatitis Elimination Plan. Specific HBV gene mutations have been linked to HCC and other liver diseases. Factors related to the interactions between OBI and mutated viral proteins, which induce endoplasmic reticulum stress and oxidative DNA damage, and the potential role of HBV integration sites (such as the TERT promoter) have been identified in HCC/OBI patients. Health initiatives for OBI research in Latin American countries are crucial to achieving the WHO’s goal of eradicating viral hepatitis by 2030, given the difficulty in diagnosing OBI and its unclear association with hepatic diseases. Full article

The preS1 region of the large hepatitis B virus (HBV) surface protein is a crucial component in HBV infection; however, its impact on the development of hepatocellular carcinoma (HCC) remains unknown. This study investigated the relationship between serum preS1 levels and hepatocarcinogenesis in patients with chronic hepatitis B (CHB). The preS1 levels were measured in 531 patients with CHB without a history of HCC. Among the patients, 293 HBV carriers who had never received nucleotide/nucleoside analog (NA) therapy had their preS1 levels measured at their first visit (non-NA group), and 238 patients who had received NA therapy had their preS1 levels measured at the start of NA administration (NA group). The two groups had no significant differences in hepatitis B surface antigen (HBsAg) levels; however, the NA group’s preS1/HBsAg ratio was significantly higher. The preS1/HBsAg ratio was significantly different between patients with CHB not meeting the NA treatment criteria and patients with chronic hepatitis and cirrhosis who were eligible for NA treatment. The predictors of HCC development were analyzed, and the preS1/HBsAg ratio was identified in both groups. The preS1/HBsAg ratio could predict hepatocarcinogenesis in patients with CHB with or without NA administration. Full article

Hepatitis B surface antigen (HBsAg) is not only the biomarker of hepatitis B virus (HBV) infection and expression activity in hepatocytes, but it also contributes to viral specific T cell exhaustion and HBV persistent infection. Therefore, anti-HBV therapies targeting HBsAg to achieve HBsAg loss are key approaches for an HBV functional cure. In this study, we found that YZH-106, a rupestonic acid derivative, inhibited HBsAg secretion and viral replication. Further investigation demonstrated that YZH-106 promoted the lysosomal degradation of viral L- and M-HBs proteins. A mechanistic study using Biacore and docking analysis revealed that YZH-106 bound directly to the PreS2 domain of L- and M-HBsAg, thereby blocking their entry into the endoplasmic reticulum (ER) and promoting their degradation in cytoplasm. Our work thereby provides the basis for the design of a novel compound therapy to target HBsAg against HBV infection. Full article