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Background: Hepatitis B virus (HBV) infection is still a serious threat to global health and can lead to a variety of liver diseases, including acute and chronic hepatitis, liver cirrhosis, liver failure, hepatocellular carcinoma (HCC), and so on. At present, there are mainly two kinds of drugs for the treatment of hepatitis B at home and abroad: interferon (IFN) and nucleoside/nucleotide analogs (NAs). In recent years, natural compounds have been considered an important source for the development of new anti-HBV drugs due to their complex structure, diverse components, high efficiency, and low toxicity. Many studies have demonstrated that Solamargine has significant anticancer activity, but the antiviral effect is rarely studied. This study aimed to verify the anti-HBV effect of Solamargine and to explore the specific mechanism., Method: The relative expression of HBV pregenomic RNA (pgRNA) was detected by reverse transcription real-time fluorescence quantitative PCR (RT-qPCR). Northern blot and western blot were used to detect the relative expression of HBV pgRNA and target protein. PCR was used in the construction of HBV pg-promoter, ENII/BCP, and a series of gene deletion mutant fluorescent reporter vectors. The fluorescence relative expression of each mutant was detected by Renilla luciferase assay., Results: By binding to MZF1 (Myeloid zinc finger protein 1, MZF1), Solamargine inhibits HBV core promoter activity, reduces pregenomic RNA level, and inhibits HBV, achieving antiviral effects.

Members of the tripartite motif (TRIM) protein family strongly induced by interferons (IFNs) are parts of the innate immune system with antiviral activity. However, it is still unclear which TRIMs could play important roles in hepatitis B virus (HBV) inhibition. Here, we identified that TRIM56 expression responded in IFN-treated HepG2-NTCP cells and HBV-infected liver tissues, which was a potent IFN-inducible inhibitor of HBV replication. Mechanistically, TRIM56 suppressed HBV replication via its Ring and C-terminal domain. C-terminal domain was essential for TRIM56 translocating from cytoplasm to nucleus during HBV infection. Further analysis revealed that TRIM56's Ring domain targeted IκBα for ubiquitination. This modification induced phosphorylation of p65, which subsequently inhibited HBV core promoter activity, resulting in the inhibition of HBV replication. The p65 was found to be necessary for NF-κB signal pathway to inhibit HBV replication. We verified our findings using HepG2-NTCP and primary human hepatocytes. Our findings reveal that TRIM56 is a critical antiviral immune effector and exerts an anti-HBV activity via NF-κB signal pathway, which is essential for inhibiting transcription of HBV covalently closed circular DNA., Competing Interests: Declaration of competing interest None of the authors has any conflict of interest., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Epigallocatechin gallate (EGCG), a major polyphenol in green tea, exhibits diverse biological activities. Previous studies show that EGCG could effectively suppress HBV gene expression and replication, but the role of EGCG in HBV replication and its underlying mechanisms, especially the signaling pathways involved, remain unclear. In this study we investigated the mechanisms underlying EGCG inhibition on HBV replication with a focus on the signaling pathways. We showed that EGCG (12.5-50 μM) dose-dependently inhibited HBV gene expression and replication in HepG2.2.15 cells. Similar results were observed in HBV mice receiving EGCG (25 mg· kg -1 · d -1 , ip) for 5 days. In HepG2.2.15 cells, we showed that EGCG (12.5-50 μM) significantly activate ERK1/2 MAPK signaling, slightly activate p38 MAPK and JAK2/STAT3 signaling, while had no significant effect on the activation of JNK MAPK, PI3K/AKT/mTOR and NF-κB signaling. By using specific inhibitors of these signaling pathways, we demonstrated that ERK1/2 signaling pathway, but not other signaling pathways, was involved in EGCG-mediated inhibition of HBV transcription and replication. Furthermore, we showed that EGCG treatment dose-dependently decreased the expression of hepatocyte nuclear factor 4α (HNF4α) both at the mRNA and protein levels, which could be reversed by pretreatment with the ERK1/2 inhibitor PD98059 (20 μM). Moreover, we revealed that EGCG treatment dose-dependently inhibited the activity of HBV core promoter and the following HBV replication. In summary, our results demonstrate that EGCG inhibits HBV gene expression and replication, which involves ERK1/2-mediated downregulation of HNF4α.These data reveal a novel mechanism for EGCG to inhibit HBV gene expression and replication.

Hepatitis B virus (HBV) causes serious health issues worldwide. Despite this, current treatment options for HBV have many drawbacks. Strategies to safely and specifically target HBV replication and survival at the transcriptional level within host cells are needed to combat current drawbacks in treatment. In this study, we designed a novel artificial transcription factor (ATF) with suppressive function to target and bind to the HBV core promoter, a component that plays a central role in the viral life cycle. ATF has attached specifically to the intended target site by using electrophoretic mobility shift assays (EMSA). We tested whether targeting this suppressive ATF had any effect on HBV gene expression by transfection factor, western blotting, and real-time PCR. In the presence of ATF, viral mRNA and DNA levels were significantly decreased within HepG2.2.15 cells compared to control cells. The HBV-derived protein expression of HBV-e antigen (HBeAg) and HBV-c antigen (HBcAg) was also significantly inhibited. These results show that ATF treatment targeting the HBV core protein promoter has an antiviral effect and inhibits HBV infection in host cells. These results further suggest that the design of new artificial transcription factors may be valuable antiviral therapies to treat HBV patients., (© 2020 by the Association of Clinical Scientists, Inc.)

Hepatitis B virus (HBV) infection remains a global health problem, causing nearly one million deaths annually. Forkhead box O (FoxO) transcription factors play important roles in modulating diverse physiological processes. Recent studies show that FoxOs are involved in antiviral responses. In present investigation, we found that HBV induced significant down-regulation of FoxO4 protein, while had little effect on the expression of FoxO1 and FoxO3. Further study showed that FoxO4 displayed inhibitory effect on HBV transcription and replication both in vitro and in vivo. Mechanistically, it was found that FoxO4 exerted its anti-HBV activity by targeting HBV core promoter. Further, FoxO4 was revealed to inhibit HBV core promoter activity via downregulating hepatocyte nuclear factor-4α (HNF4α), and ERK signaling was required for FoxO4-mediated suppression of HNF4α and HBV core promoter activity. Together, these data indicated that FoxO4 displayed anti-HBV activity by suppressing HNF4α expression via activation of ERK pathway, and targeting FoxO4 might present as a novel therapeutic strategy against HBV infection., (Copyright © 2019 The Authors. Published by Elsevier B.V. All rights reserved.)

Objectives: We aimed to study the dynamic changes of hepatitis B virus (HBV) core promoter/precore (CP/preC) sequences during antiviral treatment and their associations with virological responses., Materials and Methods: The baseline and 12-week CP/preC sequences (nts 1655-2014) were obtained from 52 chronic hepatitis B patients with positive hepatitis B e antigen (HBeAg), who received a 104-week lamivudine and adefovir dipivoxil combination therapy. The mutations within the CP/preC were analyzed against genotype specific reference sequences. The nucleotide change rates in individuals during therapy were analyzed in a pairwise comparison manner., Results: There was no significant difference of the mutation rate at each nucleotide site between baseline and week 12 of treatment (P>0.05). The mutation rates of A1762T/G1764A and G1896A were found to decrease from 46.2% (24/52) at baseline to 36.5% (19/52) at week 12 (P=0.426) and from 28.8% (15/52) to 21.2% (11/52) (P=0.497), respectively. The nucleotide change rates varied from 0.0% - 7.8% in individuals [0.0% in Group 1 (N=26); 0.3% - 7.8% in Group 2 (N=26)] during the first 12-week treatment. HBV DNA levels in Group 2 were significantly lower than those in Group 1 throughout therapy (P<0.01) (e.g., 1.5±1.3log 10 IU/ml vs. 2.6±1.0log 10 IU/ml at week 104, P=0.001). At week 104 the rates of HBV DNA undetectable and HBeAg loss in Group 2 were significantly higher than those in Group 1 (P<0.05). Along with the increased nucleotide change rates, the rate of HBV DNA undetectable at week 104 tended to increase (odds ratio=0.323, 95% confidence interval=0.138-0.758, P<0.001)., Conclusion: Our findings suggested that the nucleotide changes within HBV CP/preC region during the first 12-week treatment might be associated with a better virological response., (Copyright © 2017. Published by Elsevier B.V.)

The core promoter (CP) of the viral genome plays an important role for hepatitis B virus (HBV) replication as it directs initiation of transcription for the synthesis of both the precore and pregenomic (pg) RNAs. The CP consists of the upper regulatory region and the basal core promoter (BCP). The CP overlaps with the 3'-end of the X open reading frames and the 5'-end of the precore region, and contains cis-acting elements that can independently direct transcription of the precore mRNA and pgRNA. Its transcription regulation is under strict control of viral and cellular factors. Even though this regulatory region exhibits high sequence conservation, when variations appear, they may contribute to the persistence of HBV within the host, leading to chronic infection and cirrhosis, and eventually, hepatocellular carcinoma. Among CP sequence variations, those occurring at BCP may dysregulate viral gene expression with emphasis in the hepatitis B e antigen, and contribute to disease progression. In this review these molecular aspects and pathologic topics of core promoter are deeply evaluated.

Hepatitis B virus (HBV) is a well-known hepadnavirus with a double-stranded circular DNA genome. Although HBV was first described approximately 50 years ago, the precise mechanisms of HBV infection and effective therapeutic strategies remain unclear. Here, we focus on summarizing the complicated mechanisms of HBV replication and infection, as well as genomic factors and epigenetic regulation. Additionally, we discuss in vivo models of HBV, as well as diagnosis, prevention and therapeutic drugs for HBV. Together, the data in this 50-year review may provide new clues to elucidate molecular mechanisms of HBV pathogenesis and shed new light on the future HBV therapies., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Our understanding of the interactions between HBV and its host cells is still quite limited. Spliceosome associated factor 1 (SART1) has recently been found to restrict HCV. Thus, we aimed to dissect its role in HBV infection. SART1 was knocked down by RNA interference and over-expressed by lentiviral or adeno-associated virus (AAV) vectors in HBV-infected cell cultures and in vivo in HBV-infected mice. Luciferase reporter assays were used to determine viral or host factor promoter activities, and chromatin immunoprecipitation (ChIP) was used to investigate protein-DNA interactions. In HBV-infected cell cultures, downregulation of SART1 did not affect covalently closed circular HBV DNA but resulted in markedly enhanced HBV RNA, antigen expression and progeny virus production. On the other hand, HBV transcription and replication were significantly inhibited by overexpression of SART1. Similar results were observed in AAV-HBV-infected mice persistently replicating HBV. Inhibition of Janus kinases had no effect on SART1-mediated inhibition of HBV replication. HBV promoter assays revealed that SART1 reduced HBV core promoter activity. By screening known HBV transcription factors, we found that SART1 specifically suppressed the expression of hepatocyte nuclear factor 4α (HNF4α). Luciferase reporter and ChIP assays demonstrated a direct downregulation of HNF4α expression by association of SART1 with the HNF4α proximal P1 promoter element. We identify SART1 as a novel host factor suppressing HBV cccDNA transcription. Besides its effect on interferon-stimulated genes, SART1 exerts an anti-HBV activity by suppressing HNF4α expression, which is essential for transcription of HBV cccDNA. Hepatitis B virus (HBV) infects hepatocytes and persists in the form of covalently closed circular DNA (cccDNA), which remains a major obstacle to successful antiviral treatment. In this study, using various HBV models, we demonstrate that the protein SART1 restricts HBV cccDNA transcription by suppressing a key transcription factor, HNF4α. [Display omitted] • SART1 is an antiviral host factor that restricts HBV replication. • SART1 suppresses HNF4α expression. • SART1 associates with the HNF4α proximal P1 promoter element. [ABSTRACT FROM AUTHOR]

Hepatitis B virus (HBV) is a major public health threat and anti-HBV drugs are limited to nucleos(t)ide analogs (NAs) and pegylated interferon alpha (Peg-IFNα). Toward identifying an effective compound for HBV treatment is important to suppress and eradicate HBV. In this study, we explored the anti-viral effect of Sirtuin 6 (SIRT6) inhibitor, OSS_128167, in HBV transcription and replication. Firstly, we found that OSS_128167 could decrease the level of HBV core deoxyribonucleic acid (DNA) and 3.5-Kb ribonucleic acid (RNA) in vitro. Furthermore, the level of HBV DNA and 3.5-Kb RNA were also markedly suppressed by OSS_128167 administration in HBV transgenic mice. In addition, we found that depletion of SIRT6 inhibited HBV transcription and replication in HepG2.2.15 and HBV-infected HepG2-sodium taurocholate cotransporting polypeptide cells, whereas overexpression of SIRT6 enhanced HBV transcription and replication. Importantly, the positive effect of SIRT6 overexpression on HBV transcription could be blocked by OSS_128167 treatment. Further mechanism studies showed that HBV core promoter was significantly activated by SIRT6 through upregulating peroxisome proliferator-activated receptors α (PPARα) expression. And ectopical expression of SIRT6 or PPARα relieved the restriction of HBV transcription mediated by OSS_128167. In summary, our results showed that OSS_128167 might serve as a potential antiviral agent for HBV therapy and SIRT6 played a pivotal role in HBV transcription and replication.

Hepatitis B virus (HBV) is a major public health threat and anti-HBV drugs are limited to nucleos(t)ide analogs (NAs) and pegylated interferon alpha (Peg-IFNα). Toward identifying an effective compound for HBV treatment is important to suppress and eradicate HBV. In this study, we explored the anti-viral effect of Sirtuin 6 (SIRT6) inhibitor, OSS_128167, in HBV transcription and replication. Firstly, we found that OSS_128167 could decrease the level of HBV core deoxyribonucleic acid (DNA) and 3.5-Kb ribonucleic acid (RNA) in vitro. Furthermore, the level of HBV DNA and 3.5-Kb RNA were also markedly suppressed by OSS_128167 administration in HBV transgenic mice. In addition, we found that depletion of SIRT6 inhibited HBV transcription and replication in HepG2.2.15 and HBV-infected HepG2-sodium taurocholate cotransporting polypeptide cells, whereas overexpression of SIRT6 enhanced HBV transcription and replication. Importantly, the positive effect of SIRT6 overexpression on HBV transcription could be blocked by OSS_128167 treatment. Further mechanism studies showed that HBV core promoter was significantly activated by SIRT6 through upregulating peroxisome proliferator-activated receptors α (PPARα) expression. And ectopical expression of SIRT6 or PPARα relieved the restriction of HBV transcription mediated by OSS_128167. In summary, our results showed that OSS_128167 might serve as a potential antiviral agent for HBV therapy and SIRT6 played a pivotal role in HBV transcription and replication. [ABSTRACT FROM AUTHOR]

The objective of this study was to examine hepatitis B virus (HBV) subgenotypes and mutations in enhancer II, basal core promoter, and precore regions of HBV in relation to risks of liver cirrhosis (LC) and hepatocellular carcinoma (HCC) in Southeast China. A case-control study was performed, including chronic hepatitis B (CHB; n=125), LC (n=120), and HCC (n=136). HBV was genotyped by multiplex polymerase chain reaction and subgenotyped by restriction fragment length polymorphism. HBV mutations were measured by DNA sequencing. HBV genotype C (68.2%) predominated and genotype B (30.2%) was the second most common. Of these, C2 (67.5%) was the most prevalent subgenotype, and B2 (30.2%) ranked second. Thirteen mutations with a frequency >5% were detected. Seven mutation patterns (C1653T, G1719T, G1730C, T1753C, A1762T, G1764A, and G1799C) were associated with C2, and four patterns (C1810T, A1846T, G1862T, and G1896A) were associated with B2. Six patterns (C1653T, G1730C, T1753C, A1762T, G1764A, and G1799C) were obviously associated with LC, and 10 patterns (C1653T, G1730C, T1753C, A1762T, G1764A, G1799C, C1810T, A1846T, G1862T, and G1896A) were significantly associated with HCC compared with CHB. Four patterns (C1810T, A1846T, G1862T, and G1896A) were significantly associated with HCC compared with LC. Multivariate regression analyses showed that HBV subgenotype C2 and C2-associated mutation patterns (C1653T, T1753C, A1762T, and G1764A) were independent risk factors for LC when CHB was the control, and that B2-associated mutation patterns (C1810T, A1846T, G1862T, and G1896A) were independent risk factors for HCC when LC was the control.

Mutations in the precore and core promoter regions of hepatitis B 'e' antigen (HBeAg) are implicated in HBeAg-negative chronic hepatitis B virus (HBV) infection (CHB). The objective of the current study was to investigate novel mutant patterns that lead to HBeAg-negative CHB. The . PreX-X genomic region from the sera of HBV-infected patients was amplified, and analysis of the sequences displayed a unique deletion region, 234 nucleotides in length, which was observed in 54 clones and named core promoter deletion (CPD). CPD may have an important role in the cause of HBeAg-negative CHB. In addition, a novel deletion mutation in the X gene was observed in patients with CHB. This deletion mutant codes a 76-amino-acid X factor instead of the X protein. In the present study, a new mutation pattern was discovered that may contribute to the cause of HBeAg-negative CHB, and therefore it is worthy of future studies. [ABSTRACT FROM AUTHOR]

Advances in molecular biology technology in the last two decades have allowed detailed study of the viral mutations and genomic heterogeneity of hepatitis B virus (HBV). The first mutant discovered was precore stop codon mutation. It was reported in HBeAg-negative patients and initially thought to associate with fulminant hepatitis. Subsequent studies have suggested that it is merely one of the mechanisms of losing HBeAg by the virus. Another mutation that can downregulate the production of HBeAg is the basal core promoter mutation, which is located in the X gene upstream of the precore region. Based on the configuration of codon 15 and the stability of the epsilon of the precore region, these two mutants will be differentially selected during the course of HBeAg seroconversion. The most common HBV genotypes in South-East Asia are genotype B and C HBV. The higher hepatocellular carcinoma (HCC) risk of genotype C HBV has been confirmed by longitudinal studies in Hong Kong and Taiwan. One possible carcinogenic mechanism is its association with basal core promoter mutation, which has also been found to be a risk factor of HCC. Within genotype C HBV, subgenotype Cs is predominant in South-East Asia and subgenotype Ce is predominant in East Asia. Subgenotype Ce HBV has been found to have the highest risk of HCC as compared with subgenotype Cs or genotype B HBV. The understanding of the carcinogenic mechanisms of these HBV strains may shed light into future therapeutics in the prevention and treatment of HBV-related HCC. [ABSTRACT FROM AUTHOR]

Purpose: To evaluate the sequence variations in the enhancer II (EnhII)/basal core promotor (BCP)/precore (PC) and X genes of hepatitis B virus (HBV) in Thai patients with hepatocellular carcinoma (HCC) by conducting a cross-sectional case-control study. Methods: As much as 60 patients with HCC and 60 patients without HCC, who were matched for sex, age, hepatitis B e antigen (HBeAg) status, and HBV genotype, were included. Viral mutations in the EnhII/BCP/PC and X regions were characterized by direct sequencing in serum samples. Results: The prevalence of T1753C/A, A1762T/G1764A and G1899A mutations were significantly higher in the HCC group compared to the non-HCC group (43.3 vs. 23.3%, P = 0.02; 88.3 vs. 53.0%, P < 0.001; and 35.0 vs. 8.3%, P = 0.001, respectively). No significant difference between groups was found with respect to G1613A, C1653T, C1766T/T1768A, A1846T/C, T1858C, and G1896A mutations. By multiple logistic regression analysis, the presence of cirrhosis, A1762T/G1764A and G1899A mutations were independently associated with the risk of HCC. Conclusion: These data suggested that A1762T/G1764A and G1899A mutations were associated with the development of HCC in Thai patients. [ABSTRACT FROM AUTHOR]

Abstract: A plasmid carrying 1.3-fold HBV genome was constructed from a HBV strain that caused five consecutive cases of fulminant hepatitis (pBFH2), and HepG2 cells were transfected with pBFH2 or its variants. The pBFH2 construct with A1762T/G1764A, G1862T, and G1896A showed the largest amount of core particle-associated intracellular HBV DNA, but no significant increase of extracellular HBV DNA in comparison with the wild construct, suggesting that these mutations might work together for retention of the replicative intermediates in the cells. The retention might relate to the localization of hepatitis B core antigen (HBcAg) in the nucleus of HepG2, which was observed by confocal fluorescence microscopy. HBcAg immunohistochemical examination of liver tissue samples obtained from the consecutive fulminant hepatitis patients showed stronger staining in the nucleus than acute hepatitis patients. In conclusion, the fulminant HBV strain caused retention of the core particles and the core particle-associated HBV DNA in the cells. [Copyright &y& Elsevier]

Abstract: Rep78, the rep gene product of adeno-associated virus (AAV), has been shown to inhibit the replication of several DNA viruses. This study investigated the effects of Rep78 on replication of Hepatitis B virus (HBV) and possible mechanisms of inhibition. We have shown that HBV DNA replication and secretion of HBsAg and HBeAg in HepG2 2.2.15 cells were inhibited by Rep78. We have also demonstrated, using in vitro transcription and luciferase assay, that Rep78 binds to the HBV core promoter (HBV CP) and inhibits HBV CP activity. Furthermore, after Rep78 and HBV core protein expression plasmids were co-transfected into HepG2 cells, the expression of HBV core protein was inhibited significantly. These results suggest that Rep78 can inhibit the replication of HBV, correlating strongly with suppression of HBV CP activity. [Copyright &y& Elsevier]

Abstract: Hepatitis B virus (HBV) clone 4B replicated much more efficiently than clone 2A of the same genotype. Introduction of its T1753C, A1762T, G1764A, and C1766T core promoter mutations into the 2A genome greatly enhanced genome replication and suppressed HBeAg expression. Here we show that these effects are mediated by transcriptional up regulation of pregenomic RNA and suppression of precore RNA. Analysis of chimeric constructs suggested that the 5′ end of the 2A core gene conferred higher level of pregenomic RNA, but less core protein and genome replication relative to the 4B sequence. Genome maturity of secreted virions was reduced by mutations present in the core protein of the 2A genome but enhanced by mutations found in the 4B core protein. The 4B core protein migrated faster than that of clone 2A. The possible links among the various phenotypes and the responsible mutations remain to be established. [Copyright &y& Elsevier]

Background/Aims: To investigate the mutations in hepatitis B virus (HBV) that might be related to hepatocellular carcinoma (HCC) in the high-risk area Qidong, China. Methods: DNA sequences of HBV basal core promoter (BCP) and the overlapping X gene were determined in 58 HCC and 71 chronic hepatitis (CH) patients. In addition, a consecutive series of plasma samples from 15 HCC cases were employed to compare the CP/X sequences before and after the occurrence of HCC. Results: T1762/A1764 double mutation was frequently found in Qidong patients, regardless of clinical status (65.5% in HCC and 73.2% in CH, P >0.05). Unexpectedly, the adjacent T1766/A1768 mutation significantly increased the risk of HCC (P <0.05). Moreover, the prevalence of triple mutations in BCP was significantly higher in patients with HCC than those with CH (P <0.05). The longitudinal study demonstrated that the mutations in BCP were gradually accumulated during the development of HCC. Colony formation assay showed while A1764 mutation alone did not alter the colony-inhibitory activity of HBx, double or triple mutations largely abrogated this effect. Conclusions: The complex mutation involving T1766/A1768 was closely related to HCC. The enhanced risk of HCC caused by BCP variants could be attributable partially to the aberrant activity of HBx. [Copyright &y& Elsevier]

Background/Aims: Hepatitis B virus genotype C (HBV/C) has been classified into two geographically distinct subgenotypes; HBV/C1/Cs (Southeast Asia) and HBV/C2/Ce (East Asia). Methods: Viral differences in enhancer II/core promoter and precore regions between the subgenotypes and their association with hepatocellular carcinoma (HCC) were assessed in a matched cross-sectional control study of 118 carriers (from Hong Kong) with HBV/C1/Cs (48.0 years, 81% male, 40% HBeAg+, 44% HCC) and 210 HBV/C2/Ce (172 from Japan, 38 from Hong Kong) (50.2 years, 78% male, 30% HBeAg+, 46% HCC). Results: Univariate analyses showed that mutation V1753 was predictive for HCC among HBeAg-positive-C1/Cs-carriers (P =0.0055), and T1653 among HBeAg-positive-C2/Ce-carriers (P =0.018), and T1653 or V1753 or T1762/A1764 among HBeAg-negative-C2/Ce-carriers (P <0.05). In the multivariate analysis on all HBV/C subjects, independent predictive factors for HCC were subgenotype C2/Ce (odds ratio, 4.21; 95% confidence interval, 1.07–16.23), T1653 (3.64; 1.93–6.86), V1753 (3.07; 1.66–5.65) and T1762/A1764 (2.58; 1.21–5.49) mutations, age (⩾50 years), gender (male) and HBeAg (positive). Conclusions: Our data indicate that T1653 and/or V1753 mutations in addition to T1762/A1764 are differently associated with HCC in context of HBeAg status among HBV/C1/Cs and C2/Ce-carriers. HBV/C subgenotypes have specific mutation patterns, which is probably responsible for increased carcinogenesis of HBV/C2/Ce. [Copyright &y& Elsevier]

It has been shown that clinical and virological characteristics vary among hepatitis B virus (HBV) genotypes. In this study, we measured the virus level, disease severity, and presence or absence of core promoter (CP)/pre-core (PC) mutations in 241 HBV carriers, and investigated the clinical significance of measuring the HBV genotype. In genotype C HBV carriers, the proportion of hepatitis B e antigen (HBeAg)-positive patients was significantly higher than that in genotype B HBV carriers (0 vs. 34.4%, p < 0.05), and the virus level was higher (4.9 vs. 4.05 LGE/ml). In the genotype B HBV carriers, the incidence of PC mutation was significantly higher (69 vs. 34%, p < 0.05). In the genotype C HBV carriers, the incidence of CP mutation was significantly higher (13 vs. 78%, p < 0.05). We compared patients with the wild (W)/mutant (M) pattern in the CP/PC regions to those with the M/W pattern in the CP/PC regions among the genotype C HBV carriers. Both the proportion of HBeAg-positive patients (65.8 vs. 15.4%, p < 0.05) and the alanine aminotransferase (ALT) level (48 vs. 21.5 IU, p < 0.05) were higher in the patients with the M/W pattern in the CP/PC regions, and the disease severity was deteriorated. In conclusion, genotype B HBV may more frequently induce HBe seroconversion via PC mutation compared to genotype C HBV. Among the genotype C HBV carriers, hepatitis activity and the deterioration of the disease severity were significantly inhibited in the group in which PC mutation initially occurred, in comparison to the group in which CP mutation initially occurred. Copyright © 2006 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

The genetic variability of the hepatitis B virus (HBV) encounters two compounding forces: a high viral copy number produced during active replication and the lack of proofreading activity in the HBV polymerase, resulting in a high mutational rate. A large pool of quasispecies is generated in which the fittest virus, i.e. the virus that replicates best, becomes the dominant species. Immune and antiviral selection pressures result in vaccine/immunoglobulin escape mutants and antiviral resistant variants. Viruses encoding changes associated with antiviral resistance often have reduced replication in vitro, but the accumulation of additional mutations helps restore viral fitness. These compensatory mutations may occur not only in the polymerase gene but also in other genes such as the overlapping envelope gene, the precore gene, or in regulatory regions such as the basal core promoter. In this report we aim to review the new findings that have appeared in recent months. [ABSTRACT FROM AUTHOR]

Apart from core promoter A1762T/G1764A and precore G1896A mutations, other hepatitis B virus (HBV) mutants are detected in hepatitis B e antigen (HBeAg)-negative chronic hepatitis B (CHB). The aim of this study was to determine the effects of those mutants on clinical manifestation and viral loads of genotypes B and C HBV. Seventy-nine HBeAg-negative CHB patients with hepatitis flare were enrolled in this study and their HBV precore/core region were sequenced. Serial biochemical profiles and viral loads were assessed and compared. Fifty-three patients (67%) were infected by genotype B HBV and 26 (33%) were infected by genotype C HBV. The clinical manifestation and HBV viral loads were comparable between the two groups. However, genotype B was significantly associated with precore G1896A mutation (92.5%), and more mutations within nucleotide 1809–1817 were detected in patients infected by genotype B as compared with those infected by genotype C (18.9% vs 3.8%). Most of the cases had mutations at the −2, −3 or −5 position from the precore AUG initiation codon. Triple core promoter mutations C1753T/A1762T/G1764A appeared to be linked to genotype C rather than genotype B HBV (19.2% vs 1.9%; P = 0.013). In multivariate analysis, the presence of either triple core promoter 1753/1762/1764 mutation or nucleotide 1809–1817 mutation was the only factor associated with lower HBV viral load (<70 Meq/mL) (odds ratio = 9.01; 95% CI 1.11–71.43; P = 0.04). In conclusion, minor HBV variants with mutations in the core promoter and precore region were detectable in genotypes B and C. Such HBV variants are genotype specific and related to viraemia levels. [ABSTRACT FROM AUTHOR]

Abstract: In vitro studies of HBV core promoter mutations in hepatoma cell lines suggest that some mutations in core promoter transcription factor binding sites result in reduced core promoter activity and viral replication. We sought to validate this hypothesis using clinical samples with viral load differences before and after HBeAg seroconversion. A consensus sequence for transcription factor binding sites/regulatory regions was constructed based on published studies. Serum from two time points in 33 seroconverters and 10 interferon non-responders (controls) were utilized. Genotyping, HBV DNA quantification and direct sequencing of core promoter were performed. There were 216 new mutations following HBeAg seroconversion but few in controls. Mutations or mismatches to consensus transcription factor/regulatory region sequences clustered at nucleotide positions appeared genotype-specific, non-group specific or baseline mismatches and were discounted as having significant impact on viral replication. Only a few mutations in three seroconverters (9.1%) were specific, while 39.4% had no new mutations that could be attributed to reduction in viral load following HBeAg seroconversion. In 51.5% of patients, mutations were of uncertain significance because they occurred in demonstrated non-critical clustered nucleotide positions. Core promoter mutations post-seroconversion did not correlate with in vitro induced mutations that reduced the promoter activity. [Copyright &y& Elsevier]

Background/Aims: The effects of long-term lamivudine therapy on changes in polymerase and precore/core promoter mutations are unknown. The aim of this study was to determine the changes in these regions in patients with chronic hepatitis B virus (HBV) infection treated with lamivudine for 5 years.Methods: Serum samples obtained from 16 patients at the beginning of and during therapy were polymerase chain reaction-amplified, and nucleotide sequences of HBV analyzed.Results: By the end of 5-year therapy, mutations in YMDD motif emerged in ten patients. Mutations in L526M, M550V and M550I in polymerase were found in seven, six and six patients, respectively. The L526M mutant was found at the time or after detection of M550V/I mutant in six of seven patients. At baseline, precore and core promoter mutations were found in eight and 12 of 16 patients, respectively. Mutants of precore and core promoter were replaced by wild-type virus in each of three patients infected with mutants at 1 year. However, at 5 years of treatment, precore and core promoter mutations reappeared in some patients.Conclusions: Our data showed that lamivudine initially selected from precore/core promoter mutants to wild-type virus, but precore mutation reappeared during prolonged therapy. [Copyright &y& Elsevier]

summary. The precise modulation of hepatitis B virus (HBV) gene expression is essential for replication of the virus. HBV sequences are transcribed under the control of the preC/pregenomic, S1, S2 and X promoters. With the exception of S1, all the HBV promoters lack the orthodox TATA box motifs required for the formation of the transcription initiation complex, and as such they represent a unique model of transcription initiation elements. The presence of two enhancer sequences and negative regulatoryelements in the HBV genome further augments thecontrolled synthesis of HBV- RNA. All these transcription cis -elements are embedded within protein coding regions of the genome. This feature demonstrates the remarkable ability of the virus to maximize the function of its small genome. HBV transcription control elements also display a preference for liver-specific or liver-enriched trans -factors, which contributes to the liver tropism of the virus. This review outlines the major HBV transcription regulatory elements and highlights the reliance of accurate HBV gene modulation on the complex interplay between several trans -acting factors and their corresponding cis - motifs in the viral genome. [ABSTRACT FROM AUTHOR]

To investigate the association of hepatitis B virus (HBV) with core promoter mutation (T1762A1764) or precore mutation (A1896) with the clinical course of illness, we analyzed serial core promoter and precore sequences in 22 patients with HBV-associated chronic liver disease who were followed for 12+/-4 years (mean +/- SD). Sixteen of 22 patients were positive for HBeAg at baseline, and 15 of the 16 patients seroconverted to anti-HBe during the observation period. T1762A1764 mutation was detected in 16 of 22 patients, including 11 patients positive for HBeAg, at baseline. During the follow-up period, A1896 mutation emerged in 7 of 16 patients who had the wild-type HBV or only the T1762A1764 mutation at baseline. Sustained remission of hepatitis correlated with the low level of viremia, but did not with type of mutations. These results indicate that HBV with T1762A1764 mutation tends to precede A1896 mutation during the course of infection in Japanese patients with chronic liver disease. [ABSTRACT FROM AUTHOR]

BackgroundThe core promoter of hepatitis B virus (HBV) is crucial for the viral replication and mutations may lead to the establishment of chronic infection and development of liver diseases. We analysed this region in Singaporean HBV carriers and assessed their association with viral replication and liver damage. Materials and MethodsThirty-three Singaporean HBV carriers were selected. Serological markers for HBV infection and indicators for liver functions were analysed using commercial kits. Among these patients, 17 were chronic carriers, 10 had cirrhotic livers and 6 others had hepatocellular carcinoma (HCC). The region on the HBV genome covering the entire core promoter and core gene was amplified for each patient by polymerase chain reaction. The amplified DNA fragments were sequenced and analysed. ResultsThe incidence of mutations in the core promoter or the precore gene product (stop codon at amino acid 28) was not significantly higher compared with the wild type sequences in patients with liver damage. Most mutations in either the core promoter or precore gene significantly reduced the viral replication, as indicated by HBV DNA levels. High levels of HBV DNA were found in three mutants with deletion in the same region, presumably the binding site of liver enriched factor, within the core promoter. ConclusionOur findings revealed a different mutation pattern in the core promoter in Singaporean HBV carriers. While most mutations may not be directly associated with the development of liver diseases, deletions in the core promoter could contribute to enhanced viral replication and should be studied further. [ABSTRACT FROM AUTHOR]

The core promoter (CP) of hepatitis B virus (HBV) plays a central role in HBV replication and morphogenesis, directing the transcription of both species of 3.5 kb mRNA: pregenomic (pg) RNA and precore (pre-C) mRNA. The CP overlaps the 3′ end of the X open-reading frame (ORF) and the 5′ end of the pre-C/C ORF. The major functional elements of the CP are the upper regulatory region (URR) and the basic core promoter (BCP). The BCP is sufficient for accurate initiation of both pre-C mRNA and pgRNA transcription. It contains four AT-rich regions and the initiators for pre-C mRNA and pgRNA transcription. The upstream regulatory region consists of the negative regulatory element and the core upstream regulatory sequence. Co-operative interaction of various liver-enriched and ubiquitous transcription factors is necessary for liver-specific expression from the CP. These factors bind to the CP. Sequence conservation within the CP is crucial for maintaining active viral replication, and variation may contribute to the persistence of HBV within the host, leading to chronic infection and, ultimately, hepatocarcinogenesis. The most frequently described mutations within this region are an A to T transversion at position 1762 together with a G to A transition at position 1764. This double mutant is accompanied by a reduced level of hepatitis B e antigen (HBeAg) expression. Deletions, insertions and duplications occur within the CP. [ABSTRACT FROM AUTHOR]

We examined the frequency and significance of mutations in the core promoter and precore region in 103 Japanese patients with chronic hepatitis B virus (HBV) infection. HBV DNAs from the patients' sera were amplified by polymerase chain reaction and were directly sequenced. A double mutation (T1762 A1764) in the core promoter was frequently observed in the patients regardless of HBeAg status except for asymptomatic carriers with HBeAg. Furthermore, a mutation at nucleotide 1753 from T to C or G was frequently found in anti-HBe positive patients and was often accompanied by the double mutation. The A1896 mutation was found in only about one fourth of the patients with anti-HBe. These data suggest that the patients with chronic liver diseases frequently had a double mutation regardless of HBeAg status and a mutation at nucleotide 1753 might be associated with HBeAg-negative chronic hepatitis B virus infection. [ABSTRACT FROM AUTHOR]