Comprehensive Analysis of Hepatitis B Virus Promoter Region Mutations.
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ABSTRACT:
SUBMITTER: Meier-Stephenson V
PROVIDER: S-EPMC6265984 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Hepatitis B virus (HBV) is a highly variable DNA virus due to its unique life cycle, which involves an error-prone reverse transcriptase. The high substitution rate drives the evolution of HBV by generating genetic variants upon which selection operates. HBV mutants with clinical implications have been documented worldwide, indicating the potential for spreading and developing their own epidemiology. However, the prevalence of such mutants among the different HBV genotypes and subgenotypes has not been systematically analyzed. In the current study, we performed large-scale analysis of 6,479 full-length HBV genome sequences from genotypes A-H, with the aim of gaining comprehensive insights into the relationships of relevant mutations associated with immune escape, antiviral resistance and hepatocellular carcinoma (HCC) development with HBV (sub)genotypes and geographic regions. Immune escape mutations were detected in 10.7% of the sequences, the most common being I/T126S (1.8%), G145R (1.2%), M133T (1.2%), and Q129R (1.0%). HBV genotype B showed the highest rate of escape mutations (14.7%) while genotype H had no mutations (P < 0.001). HCC-associated mutations were detected in 33.7% of the sequences, with significantly higher frequency of C1653T, T1753V and A1762T/G1764A in genotype G than C (P < 0.001). The overall frequencies of lamivudine-, telbivudine-, adefovir-, and entecavir-resistant mutants were 7.3, 7.2, 0.5, and 0.2%, respectively, while only 0.05% showed reduced susceptibility to tenofovir. In particular, the highest frequency of lamivudine-resistant mutations was observed in genotype G and the lowest frequency in genotype E (32.5 and 0.3%; P < 0.001). The prevalence of HBV mutants was also biased by geographic location, with North America identified as one of the regions with the highest rates of immune escape, antiviral resistance, and HCC-associated mutants. The collective findings were discussed in light of natural selection and the known characteristics of HBV (sub)genotypes. Our data provide relevant information on the prevalence of clinically relevant HBV mutations, which may contribute to further improvement of diagnostic procedures, immunization programs, therapeutic protocols, and disease prognosis.
| S-EPMC7767914 | biostudies-literature
Project description:AimTo identify the distribution of hepatitis B virus (HBV) subgenotype and basal core promoter (BCP) mutations among patients with HBV-associated liver disease in Indonesia.MethodsPatients with chronic hepatitis (CH, n = 61), liver cirrhosis (LC, n = 62), and hepatocellular carcinoma (HCC, n = 48) were included in this study. HBV subgenotype was identified based on S or preS gene sequence, and mutations in the HBx gene including the overlapping BCP region were examined by direct sequencing.ResultsHBV genotype B (subgenotypes B2, B3, B4, B5 and B7) the major genotype in the samples, accounted for 75.4%, 71.0% and 75.0% of CH, LC and HCC patients, respectively, while the genotype C (subgenotypes C1, C2 and C3) was detected in 24.6%, 29.0%, and 25.0% of CH, LC, and HCC patients, respectively. Subgenotypes B3 (84.9%) and C1 (82.2%) were the main subgenotype in HBV genotype B and C, respectively. Serotype adw2 (84.9%) and adrq+ (89.4%) were the most prevalent in HBV genotype B and C, respectively. Double mutation (A1762T/G1764A) in the BCP was significantly higher in LC (59.7%) and HCC (54.2%) than in CH (19.7%), suggesting that this mutation was associated with severity of liver disease. The T1753V was also higher in LC (46.8%), but lower in HCC (22.9%) and CH (18.0%), suggesting that this mutation may be an indicator of cirrhosis.ConclusionHBV genotype B/B3 and C/C1 are the major genotypes in Indonesia. Mutations in BCP, such as A1762T/G1764A and T1753V, might have an association with manifestations of liver disease.
| S-EPMC2731954 | biostudies-literature
Project description:Telomerase reverse transcriptase (TERT) promoter mutations are among the most frequent noncoding somatic mutations in multiple cancers, including hepatocellular carcinoma (HCC). The clinical and pathological implications of TERT promoter mutations in hepatitis B virus (HBV)-associated HCC have not been resolved. To investigate TERT promoter mutations, protein expression, and their clinical-pathological implications, we sequenced the TERT promoter region for hotspot mutations in HCC tissues and performed immunostaining for TERT protein expression from HBV-associated HCC in Chinese patients. Of 276 HCC tumor DNA samples sequenced, 85 (31%) carried TERT promoter mutations. TERT promoter mutations were more frequent in those with low α-fetoprotein (AFP) serum levels (p = 0.03), advanced age (p = 0.04), and in those lacking HCC family history (p = 0.02), but were not correlated with HCC stages and grades. TERT protein levels were higher in HCC (n = 28) compared to normal liver tissues (n = 8) (p =0.001), but did not differ between mutated and non-mutated tumor tissues. In conclusion, TERT promoter mutations are common somatic mutations in HCC of Han Chinese with HBV infection. Detection of TERT promoter mutations in those with low levels of AFP may aid diagnosis of HCC with atypical presentation.
| S-EPMC5053691 | biostudies-literature
Project description:ObjectivesThe study aimed at detecting the prevailing hepatitis B virus (HBV) genotypes and the presence of clinically relevant mutations in the precore/core gene of the HBV DNA, among patients with chronic infection in South-eastern, Nigeria.MethodsA total of 72 participants with chronic HBV infection were enrolled into the study. Plasma samples from those with detectable HBV DNA were subjected to nested Polymerase Chain Reaction amplification using the precore/core specific primers. This resulted to the successful amplification and sequencing of the HBV precore/core region DNA from 16 participants. Mutation analysis on the precore/core region detected the presence of certain HBV precore/core gene mutations. Genotyping was carried out by phylogenetic analysis.ResultsThe precore region mutation at nucleotide position 1896, which is a G to A change resulting to a nonsense mutation, was detected in 6.25% of the participants. Other HBV precore region mutations that were detected include: G1899A, T1846A, G1862C, G1888A, T1821C, C1826T, A1827C, A1850T, C1858T, precore start codon Kozak sequence mutations and some novel core region mutations such as G/A1951T and G1957A. Genotyping revealed the existence of HBV genotype/subgenotype A1 (87.5%) and D (12.5%) among the participants. There was no significant difference in the occurrence of specific precore/core mutations among the HBV/hepatitis C virus dually infected and HBV mono-infected participants.ConclusionThe data suggest the likelihood of a more severe outcome of hepatitis caused by HBV in South-eastern Nigeria due to the occurrence of a variety of precore/core mutation, which resulted to HBeAg-negative chronic HBV infection among the participants.
| S-EPMC7934135 | biostudies-literature
Project description:ObjectivesMutations in hepatitis B virus (HBV) X region (HBx) play important roles in hepatocarcinogenesis while the results remain controversial. We sought to clarify potential hepatocellular carcinoma (HCC)-characteristic mutations in HBx from HBV genotype C-infected patients and the distribution of those mutations in different disease phases and genotypes.MethodsHBx sequences downloaded from an online global HBV database were screened and then classified into Non-HCC or HCC group by diagnosis information. Patients' data of patient age, gender, country or area, and viral genotype were also extracted. Logistic regression was performed to evaluate the effects of mutations on HCC risk.Results1) Full length HBx sequences (HCC: 161; Non-HCC: 954) originated from 1115 human sera across 29 countries/areas were extracted from the downloaded 5956 HBx sequences. Genotype C occupied 40.6% of Non-HCC (387/954) and 89.4% of HCC (144/161). 2) Sixteen nucleotide positions showed significantly different distributions between genotype C HCC and Non-HCC groups. 3) Logistic regression showed that mutations A1383C (OR: 2.32, 95% CI: 1.34-4.01), R1479C/T (OR: 1.96, 95% CI: 1.05-3.64; OR: 5.15, 95% CI: 2.53-10.48), C1485T (OR: 2.40, 95% CI: 1.41-4.08), C1631T (OR: 4.09, 95% CI: 1.41-11.85), C1653T (OR: 2.58, 95% CI: 1.59-4.19), G1719T (OR: 2.11, 95% CI: 1.19-3.73), and T1800C (OR: 23.59, 95% CI: 2.25-247.65) were independent risk factors for genotype C HBV-related HCC, presenting different trends among individual disease phases. 4) Several genotype C HCC risk mutations pre-existed, even as major types, in early disease phases with other genotypes.ConclusionsMutations associated with HCC risk were mainly located in HBx transactivation domain, viral promoter, protein/miRNA binding sites, and the area for immune epitopes. Furthermore, the signatures of these mutations were unique to disease phases leading to HCC, suggesting molecular counteractions between the virus and host during hepatocarcinogenesis.
| S-EPMC4420286 | biostudies-literature
Project description:BackgroundChronic hepatitis B (CHB) infection which is associated with an increased risk of developing liver disease including cirrhosis and hepatocellular carcinoma. Viral factors that may increase the risk for HCC development include HBV DNA level, genotypes, and naturally occurring mutations such as hepatitis B virus precore (PC) (G1896A) and basal core promoter (BCP) A1762T/G1764A double mutations. HBV genotypes and subgenotypes can significantly influence HBeAg seroconversion rates, viremia levels, mutational patterns that could significantly influence the heterogeneity in clinical manifestations and even response to antiviral therapy.Method94 CHB infected individuals with detectable serum HBV DNA levels were studied. HBsAg, HBeAg, anti-HBc IgM antibody estimations were done by ELISA. HBV DNA estimation was done. The HBV genotypes were determined by TSP-PCR and 10 samples randomly selected for DNA sequencing. PC and BCP mutations were determined by DNA sequence analysis of core region.ResultOf 94 study participant samples with detectable serum HBV DNA levels, 75 were successfully genotyped and sequenced for BCP/PC region. 30/75 (40%) harbored PC and BCP mutations. The total Double mutations of BCP at A1762T/G1764A nucleotide positions, and PC mutation at G1896A nucleotide position were seen in 29.3% and 21.3%, respectively. All 75 isolates were subtype D using TSP-PCR. However, by sequencing 2/10 were subtype A, while 8 were subtype D.ConclusionOur study reinforces that D is the predominant genotype in Indian population. It reveals that Indian CHB subjects have increased prevalence of BCP & PC mutations, which possibly may lead to development of HCC.
| S-EPMC4534539 | biostudies-literature
Project description:We have determined the nucleotide sequence of the 5' noncoding (NC) region of the hepatitis C virus (HCV) genome in 44 isolates from around the world. We have identified several HCV isolates with significantly greater sequence heterogeneity than reported previously within the 5' NC region. The most distantly related isolates were only 90.1% identical. Nucleotide insertions were seen in three isolates. Analysis of the nucleotide sequence from 44 HCV isolates in this study combined with that of 37 isolates reported in the literature reveals that the 5' NC region of HCV consists of highly conserved domains interspersed with variable domains. The consensus sequence was identical to the prototype HCV sequence. Nucleotide variations were found in 45 (16%) of the 282 nucleotide positions analyzed and were primarily located in three domains of significant heterogeneity (positions -239 to -222, -167 to -118, and -100 to -72). Conversely, there were three highly conserved domains consisting of 18, 22, and 63 completely invariant nucleotides (positions -263 to -246, -199 to -178, and -65 to -3, respectively). Two nucleotide domains within the 5' NC region, conserved among all HCV isolates studied to date, shared statistically significant similarity with pestivirus 5' NC sequences, providing further evidence for a close evolutionary relationship between these two groups of viruses. Additional analysis revealed the presence of short open reading frames in all HCV isolates. Our sequence analysis of the 5' NC region of the HCV genome provides additional information about conserved elements within this region and suggests a possible functional role for the region in viral replication or gene expression. These data also have implications for selection of optimal primer sequences for the detection of HCV RNA by the PCR assay.
| S-EPMC49204 | biostudies-other
Project description:Clinical studies have associated hepatitis B virus core promoter (CP) mutations with an increased risk of hepatocellular carcinoma. The CP region overlaps with the HBV X (HBx) gene, which has been implicated in hepatocarcinogenesis. The cyclin kinase inhibitor p21WAF1/CIP1 is an important regulator of cell cycle progression and proliferation. We determined whether HBx mutants that result from mutations in the CP deregulate p21 and these processes.We constructed a series of HBx mutants with changes in the CP region that correspond to A1762T/G1764A (TA), T1753A, T1768A, or a combination of these (combo) and expressed them, along with wild-type HBx under control of its endogenous promoter, in primary human hepatocytes (PHHs) and HepG2 cells. We then analyzed the effects of CP mutations on expression and degradation of p21 and the effects on cell cycle progression and proliferation.The combo mutant decreased levels of p21 and increased cyclin E expression in PHHs and HepG2 cells. The combo mutant, but not HBx with single or double CP mutations, accelerated p21 degradation in HepG2 cells. The combo mutant increased expression of S-phase kinase-associated protein 2 (SKP2) in PHHs and Huh7 cells. Silencing of SKP2 abrogated the effects of CP mutations on p21 expression. The kinetics of p21 expression correlated with changes in cell cycle distribution. The combo mutant accelerated cell cycle progression; p21 overexpression restored G1 arrest.HBx mutants with changes that correspond to a combination of CP mutations up-regulate SKP2, which then down-regulates p21 via ubiquitin-mediated proteasomal degradation. CP mutations might increase the risk of hepatocellular carcinoma via this pathway.
| S-EPMC3186859 | biostudies-literature
Project description:Hepatitis C virus (HCV) strain JFH-1, which belongs to genotype 2a, replicates autonomously in cultured cells, whereas another genotype 2a strain, J6CF, does not. Previously, we found that replacement of the NS3 helicase and NS5B-to-3'X regions of J6CF with those of JFH-1 confers J6CF replication competence. In this study, we aimed to identify the minimum modifications within these genomic regions needed to establish replication-competent J6CF. We previously identified 4 mutations in the NS5B-to-3'X region that could be used instead of replacement of this region to confer J6CF replication competence. Here, we induced cell culture-adaptive mutations in J6CF by the long-term culture of J6CF/JFH-1 chimeras composed of JFH-1 NS5B-to-3'X or individual parts of this but not the NS3 helicase region. After 2 months of culture, efficient HCV replication and infectious virus production in chimeric RNA-transfected cells were observed, and several amino acid mutations in NS4A were identified in replicating HCV genomes. The introduction of NS4A mutations into the J6CF/JFH-1 chimeras enhanced viral replication and infectious virus production. Immunofluorescence microscopy demonstrated that some of these mutations altered the subcellular localization of the coexpressed NS3 protein and affected the interaction between NS3 and NS4A. Finally, introduction of the most effective NS4A mutation, A1680E, into J6CF contributed to its replication competence in cultured cells when introduced in conjunction with four previously identified adaptive mutations in the NS5B-to-3'X region. In conclusion, we identified an adaptive mutation in NS4A that confers J6CF replication competence when introduced in conjunction with 4 mutations in NS5B-to-3'X and established a replication-competent J6CF strain with minimum essential modifications in cultured cells. The HCV cell culture system using the JFH-1 strain and HuH-7 cells can be used to assess the complete HCV life cycle in cultured cells. This cell culture system has been used to develop direct-acting antivirals against HCV, and the ability to use various HCV strains within this system is important for future studies. In this study, we aimed to establish a novel HCV cell culture system using another HCV genotype 2a strain, J6CF, which replicates in chimpanzees but not in cultured cells. We identified an effective cell culture-adaptive mutation in NS4A and established a replication-competent J6CF strain in cultured cells with minimum essential modifications. The described strategy can be used in establishing a novel HCV cell culture system, and the replication-competent J6CF clone composed of the minimum essential modifications needed for cell culture adaptation will be valuable as another representative of genotype 2a strains.
| S-EPMC5286874 | biostudies-literature
Project description:BackgroundMuch evidence has demonstrated the influence of Hepatitis B virus (HBV) mutations on the clinical course of HBV infection. As large (L) protein plays a crucial role for viral entry, we hypothesized that mutations in the pre-S1 promoter region might affect the expression of L protein and subsequently change the biological characters of virus.MethodsPatients infected with genotype C HBV were enrolled for analysis. HBV DNA sequences were inserted into a TA cloning vector and analyzed. To evaluate the effects of mutations in the pre-S1 promoter region, promoter activity and the expression of mRNA and L protein were analyzed using HepG2 cells.ResultsIn total, 35 patients were enrolled and 13 patients (37.1%) had a single base substitution in the pre-S1 promoter region; the most frequent substitution was a G-to-A substitution at the 2765th base (G2765A) in the Sp1 region. The HBV viral load showed a negative correlation with the substitution ratio of the Sp1 region or G2765A (r = - 0.493 and - 0.473, respectively). Among those with a viral load ≤5.0 log IU/ml, patients with the G2765A substitution showed a significantly lower HBV viral load than those with the wild-type sequence. HepG2 cells transfected with the G2765A substitution vector showed reduced luciferase activity of the pre-S1 promoter, as well as reduced expression of pre-S1 mRNA and L protein. Furthermore, the G2765A substitution greatly reduced the L protein expression level of vector-produced virus particles.ConclusionG2765A substitution in the pre-S1 promoter reduced the expression of L protein and resulted in a low viral load and less severe disease in chronic HBV infections.
| S-EPMC6498540 | biostudies-literature