Tobacco Smoking is Positively Associated with Hepatocellular Carcinoma Due to the p53 Mutation at Serine 249

Abstract Background The major risk factors for hepatocellular carcinoma (HCC) are known to be hepatitis b virus (HBV) infection and aflatoxin B1(AFB1) exposure. However, there is still controversy about whether smoking is related to HCC.Methods A binary unconditional logistic regression was used for the data about a total of 300 cases and 612 controls. The approach of functional analysis of separated alleles in yeast (FASAY) was applied to analyze p53 status in HCC group. The relationship between p53 mutation at Serine 249 (p53-RS) and smoking was assessed. Quantitative reverse-transcription PCR (qRT-PCR) was employed for the evaluation to transcriptional activity of p53 and p53-RS. Results Smoking was linked to the risk of HCC with an increased dose-response effect. Moreover, among subjects without alcohol drinker, the risks of HCC were significantly increased for smokers between HCC and controls. Besides, there was an increase in the number of HCC in smokers compared to non-smokers after exclusion of HBV and/or HCV infection. Also, a significant difference was observed about p53-RS between smokers and nonsmokers. Furthermore, the p53-RS transcriptional activity was significantly increased in tumor tissues.Conclusions It demonstrated that tobacco smoking is positively and independently associated with HCC, which may be attributed to p53-RS and its gain of function.

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p53 Mutation at Serine 249 and Its Gain of Function Are Highly Related to Hepatocellular Carcinoma after Smoking Exposure

Public Health Genomics ◽

10.1159/000516598 ◽

2021 ◽

pp. 1-11

Author(s):

Huai Wang ◽

Lu Chen ◽

Tong Zhou ◽

Zhongwei Zhang ◽

Canwei Zeng

Keyword(s):

Hepatocellular Carcinoma ◽

Transcriptional Activity ◽

Direct Sequencing ◽

Close Correlation ◽

P53 Mutation ◽

Gain Of Function ◽

Reverse Transcription Pcr ◽

Smoking Exposure ◽

Significant Difference ◽

Dose Response Effect

Background: It has been convincingly suggested that a close correlation exists between the incidence of hepatocellular carcinoma (HCC) and cigarette smoking. However, the underlying effect of smoking on HCC is not clear. Methods: A binary unconditional logistic regression was used for the data on a total of 300 cases and 612 controls. The approach of functional analysis of separated alleles in yeast and direct sequencing of TP53 mutations were applied to analyze the p53 status in the HCC group. The relationship between p53 mutation at serine 249 (p53-RS) and smoking was assessed. Quantitative reverse transcription PCR was employed for the evaluation to transcriptional activity of p53 and p53-RS. Results: Smoking was linked to the risk of HCC with an increased dose-response effect. Moreover, among subjects who did not drink, the risks of HCC were significantly increased for smokers between HCC and controls. Besides, there was an increase in the number of HCC in smokers compared to nonsmokers after exclusion of HBV and/or HCV infection. Also, a significant difference was observed in the incidence of p53-RS between smokers and nonsmokers the HCC group. Furthermore, the p53-RS transcriptional activity was significantly increased in tumor tissues. Conclusions: It strongly demonstrated that tobacco smoking is positively and independently associated with HCC, which may be attributed to p53-RS and its gain of function.

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Persistence of intrahepatic hepatitis B virus DNA integration in patients developing hepatocellular carcinoma after hepatitis B surface antigen seroclearance

Clinical and Molecular Hepatology ◽

10.3350/cmh.2020.0115 ◽

2021 ◽

Vol 27 (1) ◽

pp. 207-218

Author(s):

Jeong Won Jang ◽

Jin Seoub Kim ◽

Hye Seon Kim ◽

Kwon Yong Tak ◽

Heechul Nam ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Surface Antigen ◽

Hepatitis B Surface Antigen ◽

Next Generation Sequencing Technology ◽

Tumor Tissues ◽

Hbsag Seroclearance ◽

B Virus ◽

Integration Sites

Background/Aims: The role of hepatitis B virus (HBV) integration into the host genome in hepatocarcinogenesis following hepatitis B surface antigen (HBsAg) seroclearance remains unknown. Our study aimed to investigate and characterize HBV integration events in chronic hepatitis B (CHB) patients who developed hepatocellular carcinoma (HCC) after HBsAg seroclearance. Methods: Using probe-based HBV capturing followed by next-generation sequencing technology, HBV integration was examined in 10 samples (seven tumors and three non-tumor tissues) from seven chronic carriers who developed HCC after HBsAg loss. Genomic locations and patterns of HBV integration were investigated. Results: HBV integration was observed in six patients (85.7%) and eight (80.0%) of 10 tested samples. HBV integration breakpoints were detected in all of the non-tumor (3/3, 100%) and five of the seven (71.4%) tumor samples, with an average number of breakpoints of 4.00 and 2.43, respectively. Despite the lower total number of tumoral integration breakpoints, HBV integration sites in the tumors were more enriched within the genic area. In contrast, non-tumor tissues more often showed intergenic integration. Regarding functions of the affected genes, tumoral genes with HBV integration were mostly associated with carcinogenesis. At enrollment, patients who did not remain under regular HCC surveillance after HBsAg seroclearance had a large HCC, while those on regular surveillance had a small HCC. Conclusions: The biological functions of HBV integration are almost comparable between HBsAg-positive and HBsAgserocleared HCCs, with continuing pro-oncogenic effects of HBV integration. Thus, ongoing HCC surveillance and clinical management should continue even after HBsAg seroclearance in patients with CHB.

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Overexpression of LGR-5 as a Predictor of Poor Outcome in Patients with Hepatocellular Carcinoma

International Journal of Environmental Research and Public Health ◽

10.3390/ijerph16101836 ◽

2019 ◽

Vol 16 (10) ◽

pp. 1836 ◽

Cited By ~ 3

Author(s):

Chih-Jan Ko ◽

Chia-Jung Li ◽

Meng-Yu Wu ◽

Pei-Yi Chu

Keyword(s):

Hepatocellular Carcinoma ◽

Tumor Development ◽

Normal Liver ◽

Leucine Rich Repeat ◽

Tumor Tissues ◽

B Virus ◽

Cell Signal Transduction ◽

Poor Outcomes ◽

Positive Groups ◽

G Protein Coupled

Hepatocarcinogenesis and distant metastasis pose major challenges for physicians. They are regulated by several genes, such as AKT, JUK, Wnt, and P53, and their expression activates several important processes such as cell proliferation, migration, motility, and interaction in the microenvironment. The leucine-rich repeat-containing G-protein-coupled receptor 5 (LGR-5) is a novel biomarker, particularly in stem cells, and is involved in embryogenesis, tumor development, and tumor cell signal transduction. Here, we investigated LGR-5 expression using immunohistochemistry and analyzed the correlation between clinical features and prognosis in patients with hepatocellular carcinoma (HCC). We found that LGR-5 expression was higher in tumor tissues than in normal liver tissues, and that high LGR-5 expression possibly favored poor outcomes in HCC, especially in well/moderate differentiation grade, hepatitis C virus (HCV)-negative, and hepatitis B virus (HBV)-positive groups. Thus, the LGR-5 marker is suggested to be a routine biomarker for poor prognosis, thereby providing a platform for anti-LGR-5-targeted therapy in the future.

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Quantification of Pregenomic RNA and Covalently Closed Circular DNA in Hepatitis B Virus-Related Hepatocellular Carcinoma

International Journal of Hepatology ◽

10.1155/2013/849290 ◽

2013 ◽

Vol 2013 ◽

pp. 1-9 ◽

Cited By ~ 17

Author(s):

Fugui Bai ◽

Yoshihiko Yano ◽

Takumi Fukumoto ◽

Atsushi Takebe ◽

Motofumi Tanaka ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Circular Dna ◽

Expression Levels ◽

Covalently Closed Circular Dna ◽

Occult Hbv ◽

Significant Difference ◽

B Virus ◽

Liver Tissues

Pregenomic RNA (pgRNA) is generated from covalently closed circular DNA (cccDNA) and plays important roles in viral genome amplification and replication. Hepatic pgRNA and cccDNA expression levels indicate viral persistence and replication activity. This study was aimed to measure hepatic pgRNA and cccDNA expression levels in various states of hepatitis B virus (HBV) infection. Thirty-eight hepatocellular carcinoma (HCC) patients, including 14 positive for hepatitis B surface antigen (HBsAg) and 24 negative for HBsAg but positive for anti-hepatitis B core (anti-HBc) antibody, were enrolled in this study. In HBsAg-negative but anti-HBc-positive group, HBV-DNA was detected in 20 of 24 (83%) noncancerous liver tissues for at least two genomic regions based on polymerase chain reaction (PCR) analysis. pgRNA and cccDNA expression levels in occult HBV-infected patients were significantly lower than those in HBsAg-positive patients (P<0.001). pgRNA and cccDNA in cancerous tissues were also detected without significant difference from those in noncancerous tissues. In conclusion, cccDNA and pgRNA are detected and represented HBV replication not only in noncancerous but also in cancerous liver tissues. In addition, the replication is shown in not only patients with HBsAg-positive but also occult HBV-infected patients, suggesting the contribution to HCC development.

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Hepatitis B virus, tobacco smoking and ethanol consumption in the etiology of hepatocellular carcinoma

International Journal of Cancer ◽

10.1002/ijc.2910390109 ◽

1987 ◽

Vol 39 (1) ◽

pp. 45-49 ◽

Cited By ~ 73

Author(s):

Dimitrios Trichopoulos ◽

Anastasia Tzonou ◽

Evangelia Kaklamani ◽

Xenophon Zavitsanos ◽

Yvonne Koumantaki ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Tobacco Smoking ◽

Ethanol Consumption ◽

B Virus

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Original Article. Hepatitis B Virus S Promoter Deletion in Hepatocellular Carcinoma

Infection International ◽

10.1515/ii-2017-0003 ◽

2012 ◽

Vol 1 (1) ◽

pp. 14-24 ◽

Cited By ~ 2

Author(s):

Su-zhen Jiang ◽

Jia-jia Zheng ◽

Xiang-Mei Chen ◽

Ting Zhang ◽

Qiang Xu ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Tumor Tissue ◽

Direct Sequencing ◽

Start Codon ◽

Promoter Deletion ◽

Tumor Tissues ◽

B Virus

Abstract Objective To identify the difference and significance of dominant types of hepatitis B virus (HBV) pre-S mutation between liver tumor tissues and paired adjacent non-tumor tissues and to test if the mutations were tumor tissue specific. Methods HBV DNA isolated from 34 paired intratumoral and peritumoral tissues of hepatocellular carcinoma (HCC) patients were screened by PCR and direct sequencing. All patients carried HBV with genotype C, except for one B/C heterozygote. The expression, localization and excretion of LHBs mutant carrying pre-S deletions were characterized in vitro. The expression of endoplasmic reticulum (ER) GRP78 mRNA was assayed. Results Four patterns of pre-S mutations were identified: pre-S1 in-frame deletion involving the first start codon; pre-S2 in-frame deletion; pre-S2 start codon mutation with or without in-frame deletion; and S promoter in-frame deletion (ΔSP). The first two types were evenly found in both tumor and non-tumor tissues. They were rarely present as dominant strains. The last two types were frequently found in the dominant strains in tumor tissues. The overall prevalence of HBV carrying ΔSP was 17.64% (6/34) in tumor tissues, but none were dominant in nontumor tissues. HBV carrying ΔSP was unable to produce S protein in vitro. Immunocytofluorescence assay showed that the ΔSP LHBs mutant aggregated in the cytoplasm, accumulating mainly in the ER. Transient transfection and expression of ΔSP mutant caused GRP78 up-regulation in vitro. Conclusions HBV S promoter deletion was found dominantly in HCC tumor tissue. The aggregation of mutant large surface proteins in the ER possibly involved in HBV-related HCC.

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Evaluation of Transcriptional Efficiency of Hepatitis B Virus Covalently Closed Circular DNA by Reverse Transcription-PCR Combined with the Restriction Enzyme Digestion Method

Journal of Virology ◽

10.1128/jvi.79.3.1813-1823.2005 ◽

2005 ◽

Vol 79 (3) ◽

pp. 1813-1823 ◽

Cited By ~ 35

Author(s):

Yu-Chi Chou ◽

King-Song Jeng ◽

Mong-Liang Chen ◽

Hsiao-Hui Liu ◽

Tzu-Ling Liu ◽

...

Keyword(s):

Transcriptional Regulation ◽

Hepatitis B ◽

Transcriptional Activity ◽

Enzyme Digestion ◽

Restriction Enzyme Digestion ◽

Rt Pcr ◽

Digestion Method ◽

Circular Dna ◽

Reverse Transcription Pcr ◽

B Virus

ABSTRACT Virus persistence in chronic hepatitis B patients is due to the sustaining level of covalently closed circular DNA (cccDNA) within the nuclei of infected hepatocytes. In this study, we used a modified 1.3-fold hepatitis B virus (HBV) genome, with a BclI genetic marker embedded in the redundancy region, to examine the transcriptional activity of cccDNA and the effect of the HBx protein on transcriptional regulation. After harvesting total RNA from transfected cells or stable lines, we specifically identified and monitored the transcripts from cccDNA by using reverse transcription-PCR (RT-PCR) combined with the restriction enzyme digestion method. In this approach, we have found that (i) RT-PCR combined with detection of the BclI marker is a highly specific method for distinguishing cccDNA-derived transcripts from the original integrated viral genome, (ii) the transcriptional ability of cccDNA was less efficient than that from the integrated viral genome, and (iii) the transcriptional activity of cccDNA was significantly regulated by the HBx protein, a potential transcription activator. In conclusion, we provided a tool with which to elucidate the transcriptional regulation of cccDNA and clarified the transcriptional regulation mechanism of HBx on cccDNA. The results obtained may be helpful in the development of a clinical intervention for patients with chronic HBV infections.

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HBx and c-MYC Cooperate to Induce URI1 Expression in HBV-Related Hepatocellular Carcinoma

International Journal of Molecular Sciences ◽

10.3390/ijms20225714 ◽

2019 ◽

Vol 20 (22) ◽

pp. 5714 ◽

Cited By ~ 1

Author(s):

Tsuchiya ◽

Amisaki ◽

Takenaga ◽

Honjo ◽

Fujiwara ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Rna Polymerase Ii ◽

Gene Promoter ◽

Underlying Mechanism ◽

Luciferase Assay ◽

Tumor Tissues ◽

B Virus ◽

E Box ◽

Common Etiology ◽

Oncogenic Driver

Unconventional prefoldin RNA polymerase II subunit 5 interactor (URI1) has emerged as an oncogenic driver in hepatocellular carcinoma (HCC). Although the hepatitis B virus (HBV) represents the most common etiology of HCC worldwide, it is unknown whether URI1 plays a role in HBV-related HCC (HCC-B). In the present study, we investigated URI1 expression and its underlying mechanism in HCC-B tissues and cell lines. URI1 gene-promoter activity was determined by a luciferase assay. Human HCC-B samples were used for a chromatin immunoprecipitation assay. We found that c-MYC induced URI1 expression and activated the URI1 promoter through the E-box in the promoter region while the HBx protein significantly enhanced it. The positivity of URI1 expression was significantly higher in HCC-B tumor tissues than in non-HBV-related HCC tumor tissues, suggesting that a specific mechanism underlies URI1 expression in HCC-B. In tumor tissues from HCC-B patients, a significantly higher level of c-MYC was recruited to the E-box than in non-tumor tissues. These results suggest that HBx and c-MYC are involved in URI1 expression in HCC-B. URI1 expression may play important roles in the development and progression of HCC-B because HBx and c-MYC are well-known oncogenic factors in the virus and host, respectively.

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Prognostic role of ABO blood group and Rhesus factor in cirrhotic patients with hepatocellular carcinoma

Scientific Reports ◽

10.1038/s41598-019-55685-8 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 1

Author(s):

Alihan Oral ◽

Tolga Sahin

Keyword(s):

Hepatocellular Carcinoma ◽

Blood Group ◽

Blood Groups ◽

Blood Type ◽

Abo Blood Group ◽

Rhesus Factor ◽

Significant Difference ◽

B Virus ◽

Cirrhotic Patients

AbstractHepatocellular carcinoma (HCC) is one of the most common types of cancer worldwide. There are many factors in the etiology of HCC such as hepatitis B virus (HBV), hepatitis C virus (HCV), alcohol, obesity, smoking and aflatoxin. Many types of cancer are assumed to be associated with ABO blood group and Rhesus factor (RH). In this study we aimed to evaluate the relationship between tumor characteristics and overall survival (OS), ABO blood group and RH factor in patients with HCC. A total of 507 patients with chronic liver disease (252 patients with HCC and 255 patients without HCC) were included in the study. All demographic, clinic and laboratory (biochemical parameters and blood type) features were collected retrospectively. The mean age of the patients was 54.50 ± 9.30. There was no significant difference in both ABO groups and RH factors between the two groups. We found that vascular invasion rate of the tumor was higher in the B blood group and multicentric localization of tumor was significantly higer in patients with positive RH but there was no difference between OS in ABO and RH blood groups. In addition, the tumor was less multicentric in the AB blood group. Blood groups and RH factor can be used to predict the prognosis in cirrhotic patients with HCC.

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The Effects and Underlying Mechanisms of Hepatitis B Virus X Gene Mutants on the Development of Hepatocellular Carcinoma

10.21203/rs.3.rs-818602/v1 ◽

2021 ◽

Author(s):

Rui Pu ◽

Wenbin Liu ◽

Xinyu Zhou ◽

Xiaomei Hou ◽

Shiliang Cai ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Cdna Microarray ◽

Hela Cells ◽

Cdna Microarray Analysis ◽

Tumor Tissues ◽

B Virus ◽

Capture Sequencing ◽

Liver Tissues

Abstract Background: We aimed to elucidate the mechanism by which hepatitis B virus X (HBx) gene mutations increase the risk of hepatocellular carcinoma (HCC) and identify novel therapeutic targets.Methods: Wild-type and four HBx mutants (M1, A1762T/G1764A; M2, T1674G+T1753C+A1762T/G1764A; M3, C1653T+T1674G+A1762T/G1764A; Ct-HBx, carboxylic acid-terminal truncated HBx) were delivered into the livers of fumarylacetoacetate hydrolase-deficient mice by using the Sleeping Beauty (SB) transposon system, respectively. Seven liver tissues and seven tumor tissues of the SB mouse models were subjected to HBV-capture sequencing. Three liver tissues from WT-HBx mice, three tumor tissues from M3-HBx mice, and three tumor tissues from Ct-HBx mice were subjected to cDNA microarray analysis. HeLa cells stably expressing WT-HBx and the four HBx mutants were also subjected to cDNA microarray assay.Results: The incidence of HCC was higher in the mice injected with M3-HBx or Ct-HBx. M3-HBx had a stronger capacity of upregulating inflammatory cytokines than other HBx variants. HBV-capture sequencing showed that the HBx fragments were mainly integrated into intergenic and intron regions. No significant difference was observed in the number of insertion sites between tumors and liver tissues. Ectopic expression of the HBx mutants, especially M3-HBx and Ct-HBx, significantly increased cell proliferation and the S phase proportion of HepG2 and HeLa cells, compared to WT-HBx. Liver tissues of the SB mice and the transfected cells were subjected to cDNA microarray analysis. Plasminogen activator inhibitor-1 (PAI1) and cell division cycle 20 (CDC20) were identified as novel effectors. M3-HBx and Ct-HBx significantly upregulated the expression of PAI1 and CDC20 in HepG2 and HeLa cells as well as the livers of the SB mice. PAI1 silencing attenuated the effect of M3-HBx and Ct-HBx on the growth of HepG2 cells and greatly decreased the growth of HeLa cells with Ct-HBx. Conclusion: HBx C1653T+T1674G+A1762T/G1764A mutant and Ct-HBx promote carcinogenesis via upregulating PAI1 and CDC20. PAI1, an important player bridging the HBx mutants and HCC, should be a promising candidate as a predictive and prognostic biomarker and therapeutic target in HBV-related HCC.

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