Search Results (194)

The capsid protein plays a crucial role in the viral life cycle. By interacting with the viral genome, it facilitates the assembly of the nucleocapsid, ultimately leading to the formation of the viral particle. Therefore, interfering with or disrupting the interaction between the capsid protein and viral genome can effectively inhibit viral replication and infection. This review focuses on elucidating the binding mechanisms between the capsid protein and the viral genome, as well as their potential applications as therapeutic targets. In particular, it summarizes the research progress on small-molecule drugs targeting the capsid–genome binding sites of dengue virus, HBV, and SARS-CoV-2. Notably, this review provides a detailed discussion on the mechanisms by which these small-molecule inhibitors interfere with the capsid–genome interaction, aiming to offer inspiration for the future development of novel antiviral drugs targeting the capsid–genome binding. Full article

The impact of hepatitis B virus (HBV) diversity and evolution on disease progression is not well-understood. This study aims to compare intra-individual viral evolution in two groups of chronic hepatitis B (CHB) patients, using antiviral treatment initiation as a measure of lack of immunological control. From the Danish Database for Hepatitis B and C (DANHEP), 25 CHB patients were included; 14 with antiviral treatment initiation (TI group), and 11 without (NTI group). For each patient, three serial plasma samples taken before potential treatment initiation were selected. HBV DNA was amplified by PCR and analyzed by next-generation sequencing. HBV DNA and alanine transaminase were elevated in the TI group throughout the study period. Significantly higher substitution rates in the NTI group versus the TI group were found both within the viral population and at consensus level. Putative predicted CD8⁺ T cell epitopes contained significantly more substitutions in the NTI group. Genome-wide association analysis revealed several amino acid residues in the HBV genome associated with treatment initiation. This study shows that HBV has a higher rate of substitutions in CHB patients not requiring treatment. This could be linked to host immune pressure leading to disease control. Full article

Both hepatitis B virus (HBV), an hepadnavirus with a DNA genome, and hepatitis A virus (HAV), a picornavirus, require the TRAMP-like host ZCCHC14-TENT4 complex for efficient replication. However, whereas HBV requires the nucleotidyltransferase activity of TENT4 to extend and stabilize the 3′ poly(A) tails of mRNA transcribed from its genome, the role played by TENT4 in HAV replication is uncertain. HAV proteins are synthesized directly from its genomic RNA, which possesses a 3′ poly(A) tail, with its length and composition presumably maintained by 3D^pol-catalyzed RNA transcription during its replicative cycle. Using nanopore long-read sequencing of RNA from infected cells, we confirm here that the length of the HAV 3′ poly(A) tail is not altered by treating infected cells with RG7834, a small molecule TENT4 inhibitor with potent anti-HAV activity. Despite this, TENT4 catalytic activity is essential for HAV replication. Surprisingly, nanopore sequencing revealed a low abundance of HAV subgenomic RNAs (hsRNAs) that extend from the 5′ end of the genome to a site within the 5′ untranslated RNA (5′UTR) immediately downstream of a stem-loop to which the ZCCHC14-TENT4 complex is recruited. These hsRNAs are polyadenylated, and their abundance is sharply reduced by RG7834 treatment, implying they are likely products of TENT4. Similar subgenomic RNAs were not identified in poliovirus-infected cells. hsRNAs are present not only in HAV-infected cell culture but also in the liver of HAV-infected mice, where they represent 1–3% of all HAV transcripts, suggesting their physiological relevance. However, transfecting exogenous hsRNA into TENT4-depleted cells failed to rescue HAV replication, leaving the functional role of hsRNA unresolved. These findings reveal a novel picornaviral subgenomic RNA species while highlighting mechanistic differences in the manner in which HAV and HBV exploit the host ZCCHC4-TENT4 complex for their replication. Full article

The liver is an active metabolic site that generates high levels of reactive oxygen species (ROS). Oxidative stress has been implicated in the chronicity of hepatitis and hepatitis B virus (HBV) infection. This study aimed to determine the involvement of oxidative stress in HBV-infected cells and the efficacy of natural Nrf2 activators. The intracellular HBV pregenomic RNA copy number relative to total RNA was measured by RT-PCR, and various protein expressions associated with oxidative stress were analyzed by a Western blot analysis. The results showed that the Nrf2, HO-1, Akt, and Bcl-xL proteins were decreased by the continuous infection, indicating that HBV-positive cells were exposed to oxidative stress. The present study evaluated the anti-HBV infection effects of the Nrf2 activator fucoxanthin (Fx), a marine carotenoid from edible biological resources, including the comparative natural Nrf2 activator pteryxin (Ptx). These Nrf2 activators suppressed the HBV pregenomic RNA production in the HBV-infected cells, thus increasing the expression of the proteins of Nrf2 and HO-1. In the persistently infected cells transfected with the HBV genome, the Bcl-xL and Keap1 proteins, which contribute to suppressing the HBx protein involved in the HBV replication, were overexpressed. In particular, the activity of these protein expressions was marked at low concentrations of Fx. This suggests that natural Nrf2 activators may play a significant role in the HBV infection and could be a valuable source for further development through the functional utilization of food resources. 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

Background: Hepatitis delta virus (HDV) infection represents the most severe form of viral hepatitis and is a significant global health challenge. Bulevirtide (BLV) is a novel therapeutic treatment that has resulted in variable response rates in HBV/HDV-coinfected patients. We evaluated clinical, virological, and polymorphic factors for the purpose of predicting BLV treatment success. Methods: Thirty HBV/HDV-coinfected patients received BLV monotherapy (2 mg/day) for 24 to 48 weeks. Baseline (BL) serum samples were collected to assess clinical parameters and virological markers (HDV RNA, HBV DNA, HBsAg, HBcrAg, anti-HBc IgG) at treatment weeks 24 (TW24) and 48 (TW48). Additionally, full-genome HDV sequencing and a phylogenetic analysis were performed. Finally, analyses of the HDAg protein sequence and HDV RNA secondary structure were conducted to evaluate potential associations with treatment response. Results: A significant reduction in HDV RNA levels was observed at TW48, with a virological response (HDV RNA undetectable or ≥2 Log decline from BL) achieved by 58% of patients. Median BL levels of anti-HBc IgG were significantly different between virological responders (39.3 COI; interquartile range [IQR] 31.6–47.1) and virological non-responders (244.7 COI; IQR 127.0–299.4) (p = 0.0001). HDV genotype 1e was predominant across the cohort, and no specific HDAg polymorphisms predicted the response. However, secondary structure analysis of HDV RNA revealed that a specific pattern of internal loops in the region 63–100 nucleotides downstream of the editing site may influence treatment response by impacting editing efficacy. Conclusions: This study revealed key factors influencing BLV efficacy in HBV/HDV coinfection. Lower baseline anti-HBc IgG levels strongly correlated with a positive virological response, suggesting that the liver’s inflammatory state affects treatment success. Additionally, the analysis of HDV RNA secondary structure in patients receiving BLV treatment revealed a higher editing efficiency in virological responders, highlighting areas for further research. Full article

Information on circulating HBV (sub-)genotype, variants, and hepatitis D virus (HDV) coinfection, which vary by geographical area, is crucial for the efficient control and management of HBV. We investigated the genomic characteristics of HBV (with a prevalence of 8.1%) and the prevalence of HDV in Nigeria. We utilised 777 HBV-positive samples and epidemiological data from the two-stage sampled population-based, nationally representative Nigeria HIV/AIDS Indicator and Impact Survey conducted in 2018. We assessed 732 HBV DNA-extracted samples with detectable viral loads (VLs) for (sub-)genotypes and variants by whole-genome pre-amplification, nested PCR of the s-and pol-gene, and BigDye Terminator sequencing. We conducted HDV serology. In total, 19 out of the 36 + 1 states in Nigeria had a high prevalence of HBV (≥8%), with the highest prevalence (10.4%) in the north-central geopolitical zone. Up to 33.2% (95% CI 30.0–36.6) of the participants had detectable VLs of ≥300 copies/mL. The predominant circulating HBV genotype was E with 98.4% (95% CI 97.1–99.1), followed by A with 1.6% (95% CI 0.9–2.9). Drug-resistant associated variants and immune escape variants were detected in 9.3% and 0.4%, respectively. The seroprevalence of HDV was 7.34% (95% CI 5.5–9.2). Nigeria has subtype E as the major genotype with many variants. Full article

Hepatitis B virus (HBV) is a major global health concern, affecting millions of people worldwide. HBV is part of the hepadnaviridae family and one of the primary causes of acute and chronic liver infections, leading to conditions such as cirrhosis and hepatocellular carcinoma (HCC). Understanding the intracellular transport and genome repair mechanisms of HBV is crucial for developing new drugs, which—in combination with immune modulators—may contribute to potential cures. This review will explore the current knowledge of HBV intracytoplasmic and nuclear transport, as well as genome repair processes, while drawing comparisons to other viruses with nuclear replication. Full article

Hepatitis B virus (HBV) is a major global health issue, with an estimated 254 million people living with chronic HBV infection worldwide as of 2022. Chronic HBV infection is the leading cause of cirrhosis and liver cancer. Current treatment with nucleos(t)ide analogs is effective in the suppression of viral activity but generally requires lifelong treatment. They fail to eradicate the HBV viral reservoir, called covalently closed circular DNA (cccDNA), which replicates in the nucleus of liver cells. The cccDNA serves as the sole template for viral replication, as it generates the pregenomic RNA (pgRNA) necessary for producing new viral genomes. This stable form of viral DNA can reactivate the virus when treatment is stopped. HBV cccDNA is therefore one of the main challenges in curing chronic HBV infections. By targeting steps such as cccDNA formation, capsid assembly, or particle secretion, researchers continue to seek ways to interfere with HBV replication and to reduce its persistence, ultimately to eradicate HBV as a global health problem. This review provides an overview of what is currently known about cccDNA formation and biogenesis and the ongoing efforts to target and eradicate it to cure chronic HBV infections. Full article

This study utilized a genome-wide association study (GWAS) to investigate the genetic variations linked to the risk of hepatitis B virus (HBV) reactivation in patients who have undergone liver transplantation (LT), aiming to enhance understanding and improve clinical outcomes. Genotyping performed on a selected patients from the Korean Organ Transplantation Registry (KOTRY) data using high-throughput platforms with the Axiom Korea Biobank array 1.1. The discovery cohort included 21 patients who experienced HBV reactivation (cases) and 888 patients without HBV reactivation (controls) following LT. The replication cohort consisted of 5 patients with HBV reactivation (cases) and 312 patients without HBV reactivation (controls) after LT. Additive logistic regression analysis was conducted using PLINK software ver 1.9, with adjustments for age and gender. The GWAS findings from the discovery cohort were validated using the replication cohort. The GWAS identified several single-nucleotide polymorphisms (SNPs) in the RGL1, CDCA7L, and AQP9 genes that were significantly linked to HBV reactivation after LT, with genome-wide significance thresholds set at p < 10⁻⁷. Down-regulation of RGL1 cDNAs was observed in primary duck hepatocytes infected with duck HBV. Overexpression of CDCA7L was found to promote hepatocellular carcinoma cell proliferation and colony formation, whereas knocking down CDCA7L inhibited these processes. Additionally, the absence of AQP9 triggered immune and inflammatory responses, leading to mild and scattered liver cell pyroptosis, accompanied by compensatory liver cell proliferation. This study provides critical insights into the genetic factors influencing HBV reactivation after LT, identifying significant associations with SNPs in RGL1, CDCA7L, and AQP9. These findings hold promise for developing predictive biomarkers and personalized management strategies to improve outcomes for HBV-infected LT recipients. Full article

Persistence is a strategy used by many viruses to evade eradication by the immune system, ensuring their permanence and transmission within the host and optimizing viral fitness. During persistence, viruses can trigger various phenomena, including target organ damage, mainly due to an inflammatory state induced by infection, as well as cell proliferation and/or immortalization. In addition to immune evasion and chronic inflammation, factors contributing to viral persistence include low-level viral replication, the accumulation of viral mutants, and, most importantly, maintenance of the viral genome and reliance on viral oncoprotein production. This review focuses on the process of genome integration, which may occur at different stages of infection (e.g., HBV), during the chronic phase of infection (e.g., HPV, EBV), or as an essential part of the viral life cycle, as seen in retroviruses (HIV, HTLV-1). It also explores the close relationship between integration, persistence, and oncogenesis. Several models have been proposed to describe the genome integration process, including non-homologous recombination, looping, and microhomology models. Integration can occur either randomly or at specific genomic sites, often leading to genome destabilization. In some cases, integration results in the loss of genomic regions or impairs the regulation of oncogene and/or oncosuppressor expression, contributing to tumor development. Full article

Hepatitis B virus (HBV) can cause chronic infections, significantly increasing the risk of death from cirrhosis and hepatocellular carcinoma (HCC). A key player in chronic HBV infection is covalently closed circular DNA (cccDNA), a stable episomal form of viral DNA that acts as a persistent reservoir in infected hepatocytes and drives continuous viral replication. Despite the development of several animal models, few adequately replicate cccDNA formation and maintenance, limiting our understanding of its dynamics and the evaluation of potential therapeutic interventions targeting cccDNA. In this study, we aimed to develop a mouse model to investigate cccDNA formation and maintenance. We infected C57BL/6 mice with recombinant adeno-associated virus (rAAV) carrying a 1.3-overlength HBV genome (genotype C) and collected liver tissue at various time points to assess cccDNA levels and viral replication. Our results demonstrated the successful establishment of a chronic hepatitis B mouse model using rAAV-HBV1.3, which supported persistent HBV infection with sustained cccDNA expression in hepatocytes. Serum levels of HBsAg and HBeAg were elevated for up to 12 weeks, while alanine transaminase (ALT) levels remained within the normal range, indicating limited liver damage during this period. We confirmed HBV DNA expression in hepatocytes, and importantly, cccDNA was detected using qPCR after Plasmid-Safe ATP-Dependent DNase treatment, which selectively removes non-cccDNA forms. Additionally, Southern blot analysis confirmed the presence of cccDNA isolated using the Hirt extraction method. This established model provides a valuable platform for studying the long-term maintenance of cccDNA in chronic HBV infection and offers an important tool for testing novel therapeutic strategies aimed at targeting cccDNA. Full article

Chronic Hepatitis B Virus (HBV) infection remains a significant public health concern, particularly in Africa, where the burden is substantial. HBV is an enveloped virus, classified into ten phylogenetically distinct genotypes (A–J). Tests to determine HBV genotypes are based on full-genome sequencing or reverse hybridization. In practice, both approaches have limitations. Whereas diagnostic sequencing, generally using the Sanger approach, tends to focus only on the S-gene and yields little or no information on intra-patient HBV genetic diversity, reverse hybridization detects only known genotype-specific mutations. To resolve these limitations, we developed an Oxford Nanopore Technology (ONT)-based HBV diagnostic sequencing protocol suitable for clinical virology that yields both complete genome sequences and extensive intra-patient HBV diversity data. Specifically, the protocol involves tiling-based PCR amplification of HBV sequences, library preparation using the ONT Rapid Barcoding Kit (Oxford nanopore Technologies, Oxford, OX4 4DQ, UK), ONT GridION sequencing, genotyping using genome detective software v1.132/1.133, a recombination analysis using jpHMM (26 October 2011 version) and RDP5.61 software, and drug resistance profiling using Geno2pheno v2.0 software. We prove the utility of our protocol by efficiently generating and characterizing high-quality near full-length HBV genomes from 148 residual diagnostic samples from HBV-infected patients in the Western Cape province of South Africa, providing valuable insights into the genetic diversity and epidemiology of HBV in this region of the world. Full article

Background: Despite an existing safe and effective vaccine for hepatitis B virus (HBV), it is still a major public health concern. Nowadays, several drugs are used to treat chronic hepatitis B; however, full healing remains controversial. The viral covalently closed circular DNA (cccDNA) formed by HBV forms a major challenge in its treatment, as does the ability of HBV to integrate itself into the host genome, which enables infection reactivation. Interfering RNA (RNAi) is a gene-silencing post-transcriptional mechanism which forms as a promising alternative to treat chronic hepatitis B. The aim of the present review is to assess the evolution of hepatitis B treatment approaches based on using RNA interference. Methods: Data published between 2016 and 2023 in scientific databases (PubMed, PMC, LILACS, and Bireme) were assessed. Results: In total, 76,949 articles were initially identified and quality-checked, and 226 eligible reports were analyzed in depth. The main genomic targets, delivery systems, and major HBV therapy innovations are discussed in this review. This review reinforces the therapeutic potential of RNAi and identifies the need for conducting further studies to fill the remaining gaps between bench and clinical practice. Full article

Cells have developed various mechanisms to counteract viral infections. In an evolutionary arms race, cells mobilize cellular restriction factors to fight off viruses, targeted by viral factors to facilitate their own replication. The hepatitis B virus (HBV) is a small dsDNA virus that causes acute and chronic infections of the liver. Its genome persists in the nuclei of infected hepatocytes as a covalently closed circular DNA (cccDNA) minichromosome, thus building up an episomal persistence reservoir. The chromosomal maintenance complex SMC5/6 acts as a restriction factor hindering cccDNA transcription, whereas the viral regulatory protein HBx targets SMC5/6 for proteasomal degradation, thus relieving transcriptional suppression of the HBV minichromosome. To date, no curative therapies are available for chronic HBV carriers. Knowledge of the factors regulating the cccDNA and the development of therapies involving silencing the minichromosome or specifically interfering with the HBx-SMC5/6 axis holds promise in achieving sustained viral control. Here, we summarize the current knowledge of the mechanism of SMC5/6-mediated HBV restriction. We also give an overview of SMC5/6 cellular functions and how this compares to the restriction of other DNA viruses. We further discuss the therapeutic potential of available and investigational drugs interfering with the HBx-SMC5/6 axis. Full article