preS1 Domain Research Articles

Phylogeographic Genetic Diversity in the White Sucker Hepatitis B Virus across the Great Lakes Region and Alberta, Canada.

Hepatitis B viruses belong to a family of circular, double-stranded DNA viruses that infect a range of organisms, with host responses that vary from mild infection to chronic infection and cancer. The white sucker hepatitis B virus (WSHBV) was first described in the white sucker (Catostomus commersonii), a freshwater teleost, and belongs to the genus Parahepadnavirus. At present, the host range of WSHBV and its impact on fish health are unknown, and neither genetic diversity nor association with fish health have been studied in any parahepadnavirus. Given the relevance of genomic diversity to disease outcome for the orthohepadnaviruses, we sought to characterize genomic variation in WSHBV and determine how it is structured among watersheds. We identified WSHBV-positive white sucker inhabiting tributaries of Lake Michigan, Lake Superior, Lake Erie (USA), and Lake Athabasca (Canada). Copy number in plasma and in liver tissue was estimated via qPCR. Templates from 27 virus-positive fish were amplified and sequenced using a primer-specific, circular long-range amplification method coupled with amplicon sequencing on the Illumina MiSeq. Phylogenetic analysis of the WSHBV genome identified phylogeographical clustering reminiscent of that observed with human hepatitis B virus genotypes. Notably, most non-synonymous substitutions were found to cluster in the pre-S/spacer overlap region, which is relevant for both viral entry and replication. The observed predominance of p1/s3 mutations in this region is indicative of adaptive change in the polymerase open reading frame (ORF), while, at the same time, the surface ORF is under purifying selection. Although the levels of variation we observed do not meet the criteria used to define sub/genotypes of human and avian hepadnaviruses, we identified geographically associated genome variation in the pre-S and spacer domain sufficient to define five WSHBV haplotypes. This study of WSHBV genetic diversity should facilitate the development of molecular markers for future identification of genotypes and provide evidence in future investigations of possible differential disease outcomes.

Thiol-ene enabled preparation of S-lipidated anti-HBV peptides.

Despite significant efforts made towards treatments for Hepatitis B virus (HBV), a long-term curative treatment has thus far eluded scientists. Recently, the Sodium Taurocholate Co-Transporting Polypeptide (NTCP) receptor has been identified as the entry pathway of HBV into hepatocytes. Myrcludex B, an N-terminally myristoylated 47-mer peptide mimic of the preS1 domain of the Hepatitis B virion, was identified as a potent protein-protein interaction (PPI) inhibitor blocking HBV fusion (IC50 = 140 pM). Herein we report an optimised chemical synthesis of Myrcludex B and a series of novel analogues. Employing a small modification to the Cysteine Lipidation of a Peptide or Amino acid (CLipPA) thiol-ene reaction, a library of S-lipidated Myrcludex B and truncated (21-mer) analogues were prepared, providing novel chemical space to probe for the discovery of novel anti-HBV peptides. The S-lipidated analogues showed an equivalent or a slight decrease (∼2-fold) in binding effectiveness to NTCP expressing hepatocytes compared to Myrcludex B. Three S-lipidated analogues were highly potent HBV inhibitors (IC50 0.97-3.32 nM). These results demonstrate that incorporation of heteroatoms into the lipid 'anchor' is tolerated by this antiviral scaffold and to the best of our knowledge constitutes the first report of potent S-lipidated antiviral peptides. Interestingly, despite only moderate reductions in binding effectiveness, truncated analogues possessed dramatically reduced inhibitory activity thus providing new insights into the structure activity relationship of these hitherto unreported antiviral S-lipopeptides.

Association of the Hepatitis B Virus Large Surface Protein with Viral Infectivity and Endoplasmic Reticulum Stress-mediated Liver Carcinogenesis.

Hepatitis B is the most prevalent viral hepatitis worldwide, affecting approximately one-third of the world’s population. Among HBV factors, the surface protein is the most sensitive biomarker for viral infection, given that it is expressed at high levels in all viral infection phases. The large HBV surface protein (LHBs) contains the integral pre-S1 domain, which binds to the HBV receptor sodium taurocholate co transporting polypeptide on the hepatocyte to facilitate viral entry. The accumulation of viral LHBs and its prevalent pre-S mutants in chronic HBV carriers triggers a sustained endoplasmic reticulum (ER) overload response, leading to ER stress-mediated cell proliferation, metabolic switching and genomic instability, which are associated with pro-oncogenic effects. Ground glass hepatocytes identified in HBV-related hepatocellular carcinoma (HCC) patients harbor pre-S deletion variants that largely accumulate in the ER lumen due to mutation-induced protein misfolding and are associated with increased risks of cancer recurrence and metastasis. Moreover, in contrast to the major HBs, which is decreased in tumors to a greater extent than it is in peritumorous regions, LHBs is continuously expressed during tumorigenesis, indicating that LHBs serves as a promising biomarker for HCC in people with CHB. Continuing efforts to delineate the molecular mechanisms by which LHBs regulates pathological changes in CHB patients are important for establishing a correlation between LHBs biomarkers and HCC development.

Quantification, epitope mapping and genotype cross-reactivity of hepatitis B preS-specific antibodies in subjects vaccinated with different dosage regimens of BM32.

BackgroundChronic hepatitis B virus (HBV) infections are a global health problem. There is a need for therapeutic strategies blocking continuous infection of liver cells. The grass pollen allergy vaccine BM32 containing the preS domain of the large HBV surface protein (LHBs) as immunogenic carrier induced IgG antibodies in human subjects inhibiting HBV infection in vitro. Aim of this study was the quantification, epitope mapping and investigation of HBV genotype cross-reactivity of preS-specific antibodies in subjects treated with different dosage regimens of BM32MethodsHundred twenty eight grass pollen allergic patients received in a double-blind, placebo-controlled trial five monthly injections of placebo (aluminum hydroxide, n= 34) or different courses of BM32 (2 placebo + 3 BM32, n= 33; 1 placebo + 4 BM32, n= 30; 5 BM32, n= 31). Recombinant Escherichia coli-expressed preS was purified. Overlapping peptides spanning preS and the receptor-binding sites from consensus sequences of genotypes A–H were synthesized and purified. Isotype (IgM, IgG, IgA, IgE) and IgG subclass (IgG1-IgG4) responses to preS and peptides were determined by ELISA at baseline, one and four months after the last injection. IgG1 and IgG4 subclass concentrations specific for preS and the receptor-binding site were measured by quantitative ELISA.FindingsFive monthly injections induced the highest levels of preS-specific IgG consisting mainly of IgG1 and IgG4, with a sum of median preS-specific IgG1 and IgG4 concentrations of >135 μg/ml reaching up to 1.8 mg/ml. More than 20% of preS-specific IgG was directed against the receptor-binding site. BM32-induced IgG cross-reacted with the receptor-binding domains from all eight HBV genotypes A-H.InterpretationBM32 induces high levels of IgG1 and IgG4 antibodies against the receptor binding sites of all eight HBV genotypes and hence might be suitable for therapeutic HBV vaccination.FundingThis study was supported by the PhD program IAI (KPW01212FW), by Viravaxx AG and by the Danube-ARC funded by the Government of Lower Austria. Rudolf Valenta is a recipient of a Megagrant of the Government of the Russian Federation, grant No 14.W03.31.0024.

Dual-targeting nanoparticle vaccine elicits a therapeutic antibody response against chronic hepatitis B.

Chronic hepatitis B is caused by prolonged infection with the hepatitis B virus (HBV), which can substantially increase the risk of developing liver disease. Despite the development of preventive vaccines against HBV, a therapeutic vaccine inducing an effective antibody response still remains elusive. The preS1 domain of the large HBV surface protein is the major viral attachment site on hepatocytes and thus offers a therapeutic target; however, its poor immunogenicity limits clinical translation. Here, we design a ferritin nanoparticle vaccine that can deliver preS1 to specific myeloid cells, including SIGNR1+ dendritic cells (which activate T follicular helper cells) and lymphatic sinus-associated SIGNR1+ macrophages (which can activate B cells). This nanoparticle vaccine induces a high-level and persistent anti-preS1 response that results in efficient viral clearance and partial serological conversion in a chronic HBV mouse model, offering a promising translatable vaccination strategy for the functional cure of chronic hepatitis B.

Structure-Guided Approach to Identify Potential Inhibitors of Large Envelope Protein to Prevent Hepatitis B Virus Infection.

Hepatitis B virus (HBV) infection is one of the major causes of liver diseases, which can lead to hepatocellular carcinoma. The role of HBV envelope proteins is crucial in viral morphogenesis, infection, and propagation. Thus, blocking the pleiotropic functions of these proteins especially the PreS1 and PreS2 domains of the large surface protein (LHBs) is a promising strategy for designing efficient antivirals against HBV infection. Unfortunately, the structure of the LHBs protein has not been elucidated yet, and it seems that any structure-based drug discovery is critically dependent on this. To find effective inhibitors of LHBs, we have modeled and validated its three-dimensional structure and subsequently performed a virtual high-throughput screening against the ZINC database using RASPD and ParDOCK tools. We have identified four compounds, ZINC11882026, ZINC19741044, ZINC00653293, and ZINC15000762, showing appreciable binding affinity with the LHBs protein. The drug likeness was further validated using ADME screening and toxicity analysis. Interestingly, three of the four compounds showed the formation of hydrogen bonds with amino acid residues lying in the capsid binding region of the PreS1 domain of LHBs, suggesting the possibility of inhibiting the viral assembly and maturation process. The identification of potential lead molecules will help to discover more potent inhibitors with significant antiviral activities.

Formation of semi-enveloped particles as a unique feature of a hepatitis B virus PreS1 deletion mutant.

Naturally occurring variants with deletions or mutations in the C-terminal PreS1 domain from hepatitis B virus (HBV) chronically infected patients have been shown to promote HBsAg retention, inhibit HBsAg secretion and change the extracellular appearance of PreS1-containing HBV particles (filaments and virions). To study the impact of N-terminal deletion in preS1 domain on viral secretion and morphogenesis. An HBV mutant with 15 amino acids (aa 25-39) deletion in N-terminal preS1 was isolated. Intracellular and extracellular HBsAg were quantified by Western blot. Subcellular HBsAg distribution was analysed by confocal laser scanning microscopy. The viral morphology was characterised by sucrose density gradient ultracentrifugation, Western blot, electron microscopy, HBV mixed ELISA and HBV particle gel essay. Expression of this mutant genome released higher amounts of HBsAg in the form of shorter filaments. A significant fraction of semi-enveloped virions was observed in the supernatant that has been unprecedented so far. Stepwise insertion of aa 25-31, aa 32-39 and aa 25-39 increased the length of filaments. The rescue of aa 25-31 and aa 25-39 drastically reduced the amounts of extracellular HBsAg and semi-enveloped virions, while such effects could not be observed after insertion of aa 32-39, arguing against a simple spacer function of this region. The deletion and rescued mutants do not differ in subcellular HBsAg distribution and colocalisation with ER, Golgi and multivesicular bodies markers arguing against differences in release pathways. N-terminal PreS1-domain (aa 25-31) determines HBsAg secretion and triggers proper assembly of PreS1-containing particles.

Variability in the immunogenic preS region of Pakistani hepatitis B virus isolates.

This paper reports the genetic variability of hepatitis B virus in Pakistan population. The worldwide prevalence of hepatitis B virus is estimated to be around 350 million that causes significant mortality especially in developing countries like Pakistan. In this study, genetic diversity of HBV was checked by using preS domain of HBV. About seventy-five samples were selected for study. Among these samples nine samples showed positive results after PCR and gel analysis. These nine samples were named SBS001-SBS008. After gel purification these samples were ligated in T/A cloning vector and transformed with E. coli DH5α. After successful cloning and positive restriction analysis these samples were subjected to DNA sequencing. Sequencing results showed that eight samples (SBS002-SBS008) have a deletion of 33 nucleotides at N-terminal that is characteristics of genotype D while SBS001 belongs to genotype C. Silent mutations and amino acid changes were also searched in this highly variable region of genome. Based upon this study it was concluded that genotype D is the most common genotype in Pakistan.

The Membrane Proximal Domain of TRPV1 and TRPV2 Channels Mediates Protein⁻Protein Interactions and Lipid Binding In Vitro.

Constitutive or regulated membrane protein trafficking is a key cell biology process. Transient receptor potential channels are somatosensory proteins in charge of detecting several physical and chemical stimuli, thus requiring fine vesicular trafficking. The membrane proximal or pre-S1 domain (MPD) is a highly conserved domain in transient receptor potential channels from the vanilloid (TRPV) subfamily. MPD shows traits corresponding to protein-protein and lipid-protein interactions, and protein regulatory regions. We have expressed MPD of TRPV1 and TRPV2 as green fluorescente protein (GFP)-fusion proteins to perform an in vitro biochemical and biophysical characterization. Pull-down experiments indicate that MPD recognizes and binds Soluble N-ethylmaleimide-sensitive factor Attachment Protein Receptors (SNARE). Synchrotron radiation scattering experiments show that this domain does not self-oligomerize. MPD interacts with phosphatidic acid (PA), a metabolite of the phospholipase D (PLD) pathway, in a specific manner as shown by lipid strips and Trp fluorescence quenching experiments. We show for the first time, to the best of our knowledge, the binding to PA of an N-terminus domain in TRPV channels. The presence of a PA binding domain in TRPV channels argues for putative PLD regulation. Findings in this study open new perspectives to understand the regulated and constitutive trafficking of TRPV channels exerted by protein-protein and lipid-protein interactions.

The Variability of Amino Acid Sequences in Hepatitis B Virus.

Hepatitis B virus (HBV) is an important human pathogen belonging to the Hepadnaviridae family, Orthohepadnavirus genus. Over 240 million people are infected with HBV worldwide. The reverse transcription during its genome replication leads to low fidelity DNA synthesis, which is the source of variability in the viral proteins. To investigate the variability quantitatively, we retrieved amino acid sequences of 5,167 records of all available HBV genotypes (A-J) from the Genbank database. The amino acid sequences encoded by the open reading frames (ORF) S/C/P/X in the HBV genome were extracted and subjected to alignment. We analyzed the variability of the lengths and the sequences of proteins as well as the frequencies of amino acids. It comprehensively characterized the variability and conservation of HBV proteins at the level of amino acids. Especially for the structural proteins, hepatitis B surface antigens (HBsAg), there are potential sites critical for virus assembly and immune recognition. Interestingly, the preS1 domains in HBsAg were variable at some positions of amino acid residues, which provides a potential mechanism of immune-escape for HBV, while the preS2 and S domains were conserved in the lengths of protein sequences. In the S domain, the cysteine residues and the secondary structures of the alpha-helix and beta-sheet were likely critical for the stable folding of all HBsAg components. Also, the preC domain and C-terminal domain of the core protein are highly conserved. However, the polymerases (HBpol) and the HBx were highly variable at the amino acid level. Our research provides a basis for understanding the conserved and important domains of HBV viral proteins, which could be potential targets for anti-virus therapy.

Oral administration of a chimeric Hepatitis B Virus S/preS1 antigen produced in lettuce triggers infection neutralizing antibodies in mice

Hepatitis B Virus (HBV) infection can be prevented by vaccination. Vaccines containing the small (S) envelope protein are currently used in universal vaccination programs and achieve protective immune response in more than 90% of recipients. However, new vaccination strategies are necessary for successful immunization of the remaining non- or low-responders. We have previously characterized a novel HBV chimeric antigen, which combines neutralization epitopes of the S and the preS1 domain of the large (L) envelope protein (genotype D). The S/preS121–47 chimera produced in mammalian cells and Nicotiana benthamiana plants, induced a significantly stronger immune response in parenterally vaccinated mice than the S protein. Here we describe the transient expression of the S/preS121–47 antigen in an edible plant, Lactuca sativa, for potential development of an oral HBV vaccine. Our study shows that oral administration of adjuvant-free Lactuca sativa expressing the S/preS121–47 antigen, three times, at 1 μg/dose, was sufficient to trigger a humoral immune response in mice. Importantly, the elicited antibodies were able to neutralize HBV infection in an NTCP-expressing infection system (HepG2-NTCP cell line) more efficiently than those induced by mice fed on Lactuca sativa expressing the S protein. These results support the S/preS121–47 antigen as a promising candidate for future development as an edible HBV vaccine.

IMMUNOGENICITY OF THE THIRD GENERATION HEPATITIS B VACCINE (pre-S1/pre-S2/S)

Currently, second-generation hepatitis B vaccines are widely used. They are produced in biotechnological eukaryotic yeast-based systems and contain the S-protein domain of HBsAg particle in the complete absence of pre-S1 and pre-S2 domains. At the same time, these antigens were proved to significantly influence immunogenicity, which makes the development and use of third-generation vaccines containing all antigenic determinants a long-range objective. A randomized, double-blind clinical study was conducted to compare immunogenicity of second- and third-generation vaccines Engerix-B™ and Sci-B-Vac™, respectively. Healthy subjects of both sexes aged 18 to 45 years (n = 94) who are seronegative for HBsAg, HBsAb, HBcAb at screening and who previously had not received immunobiological agents for hepatitis B prophylaxis were included in the study. Group I (n = 47) received the second-generation vaccine Engerix-B™, Group II (n = 47) — the third-generation vaccine Sci-B-Vac™. Subjects received vaccines three times — on days 1, 28 and 180 of the study. HBsAb levels, rates of seroconversion (the proportion of subjects with HBsAb levels 2.1 mIU/mL) and seroprotection (the proportion of subjects with HBsAb levels ≥ 10 mIU/mL) were assessed on days 28, 90, 180 and 210 of the study. Early seroconversion rate assessed on Day 28 was 76.60% in Group I and 93.88% in Group II (p 0.05); seroprotection rate was 51.06 and 61.22%, respectively. These differences in the proportion of subjects who achieved seroconversion on Day 28 may indicate a faster immunological response to Sci-B-Vac™ vaccine. Statistically significant differences between the level of antibodies on days 90 and 180 (p 0.05) were observed when analyzing the average values of HBsAb concentration in Groups I and II. The concentration of HBsAb on Day 90 was 378.68±60.95 mIU/mL in Group I, and 618.31±58.34 mIU/mL in Group II. On Day 180, the concentration of HBsAb reached 441.34±63.83 mIU/mL in Group I, and 757.72±55.14 mIU/mL in Group II. The significance of dependence of antibody level on sex, age and body weight was analyzed. It was revealed that the age of a vaccinated subject affects antibody level after administration of Engerix-B™ (p 0.05). The results obtained suggest that there is a rapid and strong immune response to the third-generation vaccine Sci-B-Vac™. This may indicate advantage of the vaccine containing all three recombinant proteins of hepatitis B virus envelope, which, in turn, can play a key role in clinical practice for urgent prophylaxis of hepatitis B, as well as for treatment of immunocompromised conditions.

A new strategy to identify hepatitis B virus entry inhibitors by AlphaScreen technology targeting the envelope-receptor interaction

Current anti-hepatitis B virus (HBV) agents have limited effect in curing HBV infection, and thus novel anti-HBV agents with different modes of action are in demand. In this study, we applied AlphaScreen assay to high-throughput screening of small molecules inhibiting the interaction between HBV large surface antigen (LHBs) and the HBV entry receptor, sodium taurocholate cotransporting polypeptide (NTCP). From the chemical screening, we identified that rapamycin, an immunosuppressant, strongly inhibited the LHBs-NTCP interaction. Rapamycin inhibited hepatocyte infection with HBV without significant cytotoxicity. This activity was due to impaired attachment of the LHBs preS1 domain to cell surface. Pretreatment of target cells with rapamycin remarkably reduced their susceptibility to preS1 attachment, while rapamycin pretreatment to preS1 did not affect its attachment activity, suggesting that rapamycin targets the host side. In support of this, a surface plasmon resonance analysis showed a direct interaction of rapamycin with NTCP. Consistently, rapamycin also prevented hepatitis D virus infection, whose entry into cells is also mediated by NTCP. We also identified two rapamycin derivatives, everolimus and temsirolimus, which possessed higher anti-HBV potencies than rapamycin. Thus, this is the first report for application of AlphaScreen technology that monitors a viral envelope-receptor interaction to identify viral entry inhibitors.

Analysis of hepatitis B virus preS1 variability and prevalence of the rs2296651 polymorphism in a Spanish population.

AIMTo determine the variability/conservation of the domain of hepatitis B virus (HBV) preS1 region that interacts with sodium-taurocholate cotransporting polypeptide (hereafter, NTCP-interacting domain) and the prevalence of the rs2296651 polymorphism (S267F, NTCP variant) in a Spanish population.METHODSSerum samples from 246 individuals were included and divided into 3 groups: patients with chronic HBV infection (CHB) (n = 41, 73% Caucasians), patients with resolved HBV infection (n = 100, 100% Caucasians) and an HBV-uninfected control group (n = 105, 100% Caucasians). Variability/conservation of the amino acid (aa) sequences of the NTCP-interacting domain, (aa 2-48 in viral genotype D) and a highly conserved preS1 domain associated with virion morphogenesis (aa 92-103 in viral genotype D) were analyzed by next-generation sequencing and compared in 18 CHB patients with viremia > 4 log IU/mL. The rs2296651 polymorphism was determined in all individuals in all 3 groups using an in-house real-time PCR melting curve analysis.RESULTSThe HBV preS1 NTCP-interacting domain showed a high degree of conservation among the examined viral genomes especially between aa 9 and 21 (in the genotype D consensus sequence). As compared with the virion morphogenesis domain, the NTCP-interacting domain had a smaller proportion of HBV genotype-unrelated changes comprising > 1% of the quasispecies (25.5% vs 31.8%), but a larger proportion of genotype-associated viral polymorphisms (34% vs 27.3%), according to consensus sequences from GenBank patterns of HBV genotypes A to H. Variation/conservation in both domains depended on viral genotype, with genotype C being the most highly conserved and genotype E the most variable (limited finding, only 2 genotype E included). Of note, proline residues were highly conserved in both domains, and serine residues showed changes only to threonine or tyrosine in the virion morphogenesis domain. The rs2296651 polymorphism was not detected in any participant.CONCLUSIONIn our CHB population, the NTCP-interacting domain was highly conserved, particularly the proline residues and essential amino acids related with the NTCP interaction, and the prevalence of rs2296651 was low/null.

Direct Binding between Pre-S1 and TRP-like Domains in TRPP Channels Mediates Gating and Functional Regulation by PIP2

SUMMARYTransient receptor potential (TRP) channels are regulated by diverse stimuli comprising thermal, chemical, and mechanical modalities. They are also commonly regulated by phosphatidylinositol-4,5-bisphosphate (PIP2), with underlying mechanisms largely unknown. We here revealed an intramolecular interaction of the TRPP3 N and C termini (N-C) that is functionally essential. The interaction was mediated by aromatic Trp81 in pre-S1 domain and cationic Lys568 in TRP-like domain. Structure-function analyses revealed similar N-C interaction in TRPP2 as well as TRPM8/-V1/-C4 via highly conserved tryptophan and lysine/arginine residues. PIP2 bound to cationic residues in TRPP3, including K568, thereby disrupting the N-C interaction and negatively regulating TRPP3. PIP2 had similar negative effects on TRPP2. Interestingly, we found that PIP2 facilitates the N-C interaction in TRPM8/-V1, resulting in channel potentiation. The intramolecular N-C interaction might represent a shared mechanism underlying the gating and PIP2 regulation of TRP channels.

Effect of S267F variant of NTCP on the patients with chronic hepatitis B

Sodium taurocholate cotransporting polypeptide (NTCP) was identified as an entry receptor for hepatitis B virus (HBV) infection. The substitution of serine at position 267 of NTCP with phenylalanine (S267F) is an Asian-specific variation that hampers HBV entry in vitro. In this study, we aimed to evaluate the prevalence of S267F polymorphism in Korean patients with chronic hepatitis B (CHB) and its association with disease progression and potential viral evolution in the preS1 domain of HBV. We found that the frequency of the S267F variant of NTCP in CHB patients and controls was 2.7% and 5.7% (P = 0.031), respectively, and that those who had S267F variant were less susceptible to chronic HBV infection. The frequency of the S267F variant in CHB, cirrhosis and hepatocellular carcinoma (HCC) patients was 3.3%, 0.9%, and 3.5%, respectively. Thus, the S267F variant correlated significantly with a lower risk for cirrhosis (P = 0.036). Sequencing preS1 domain of HBV from the patients who had S267F variant revealed no significant sequence change compared to the wild type. In conclusion, the S267F variant of NTCP is clinically associated with a lower risk of chronic HBV infection and cirrhosis development, which implicates suppressing HBV entry could reduce the disease burden.

Pre-S deletions of hepatitis B virus predict recurrence of hepatocellular carcinoma after curative resection.

The relationship between hepatitis B virus (HBV) and the prognosis of hepatocellular carcinoma (HCC) after surgery remains uncertain. A retrospective cohort study was performed to evaluate the impact of pre-S deletions, T1762/A1764, and A1896 mutations on prognosis of HCC after curative resection. A total of 113 patients with positive serum HBV DNA (>200 IU/mL) who had underwent curative resection of pathologically proven HCC were recruited to determine the risk factors affecting the prognosis.The median follow-up time was 36.5 months and recurrence was detected in 67 patients (59.3%). The cumulative recurrence rates and overall survival rates at 1-, 3-, and 5-year after curative resection were 18.0%, 49.7%, 70.3%, and 93.7%, 61.0%, 42.5%, respectively. Patients with pre-S deletions showed significantly higher recurrence rates compared with those with wild type infection (HR: 1.822, P = .018), but not related with a significantly poor survival (HR: 1.388, P = .235). Subgroup analysis indicated that the patients with type III deletion had significant higher tumor recurrence rates than other deletion types (HR: 2.211, 95% confidence intervals [CI]: 1.008–4.846, P = .048). Multivariate analysis revealed that pre-S deletion, tumor size >3 cm in diameter, and the presence of microvascular invasion were independent risk factors for tumor recurrence. HBV pre-S deletions were found to be clustered primarily in the 5′ end of pre-S2 region and were more often found between amino acids 120 and 142 of the pre-S2 domain. The domains most frequently potentially involved were the transactivator domain in pre-S2 and polymerized human serum albumin binding site.Our cohort showed that pre-S deletions at the time of resection could predict tumor recurrence in HCC patients after curative resection.

Quantification of large and middle proteins of hepatitis B virus surface antigen (HBsAg) as a novel tool for the identification of inactive HBV carriers

ObjectiveAmong individuals with chronic hepatitis B, those with hepatitis B e-antigen (HBeAg)-negative chronic hepatitis (CHB) can be difficult to distinguish from those with HBeAg-negative chronic HBV infection, also referred to...

A potent human neutralizing antibody Fc-dependently reduces established HBV infections.

Hepatitis B virus (HBV) infection is a major global health problem. Currently-available therapies are ineffective in curing chronic HBV infection. HBV and its satellite hepatitis D virus (HDV) infect hepatocytes via binding of the preS1 domain of its large envelope protein to sodium taurocholate cotransporting polypeptide (NTCP). Here, we developed novel human monoclonal antibodies that block the engagement of preS1 with NTCP and neutralize HBV and HDV with high potency. One antibody, 2H5-A14, functions at picomolar level and exhibited neutralization-activity-mediated prophylactic effects. It also acts therapeutically by eliciting antibody-Fc-dependent immunological effector functions that impose durable suppression of viral infection in HBV-infected mice, resulting in reductions in the levels of the small envelope antigen and viral DNA, with no emergence of escape mutants. Our results illustrate a novel antibody-Fc-dependent approach for HBV treatment and suggest 2H5-A14 as a novel clinical candidate for HBV prevention and treatment of chronic HBV infection.

Vaccines targeting preS1 domain overcome immune tolerance in hepatitis B virus carrier mice

These results suggest that preS1 can function as a therapeutic vaccine for the control of CHB. (Hepatology 2017;66:1067-1082).