The novel HBx mutation F30V correlates with hepatocellular carcinoma in vivo, reduces hepatitis B virus replicative efficiency and enhances anti-apoptotic activity of HBx N terminus in vitro

Abstract

Objective: We aimed to investigate HBx genetic elements correlated with hepatitis B virus (HBV) -related hepatocellular carcinoma (HCC) and their impact on (a) HBV replicative efficiency, (b) HBx binding to circular covalently closed DNA (cccDNA), (c) apoptosis and cell-cycle progression, and (d) HBx structural stability. Methods: This study included 123 individuals chronically infected with HBV: 27 with HCC (77.9% (21/27) genotype D; 22.1% (6/27) genotype A) and 96 without HCC (75% (72/96) genotype D; 25.0% (24/96) genotype A). HepG2 cells were transfected by wild-type or mutated linear HBV genome to assess pregenomic RNA (pgRNA) and core-associated HBV-DNA levels, HBx-binding onto cccDNA by chromatin immunoprecipitation-based quantitative assay, and rate of apoptosis and cell-cycle progression by cytofluorimetry. Results: F30V was the only HBx mutation correlated with HCC (18.5% (5/27) in HCC patients versus 1.0% (1/96) in non-HCC patients, p 0.002); a result confirmed by multivariate analysis. In vitro, F30V determined a 40% and 60% reduction in pgRNA and core-associated HBV-DNA compared with wild-type (p <0.05), in parallel with a significant decrease of HBx binding to cccDNA and decreased HBx stability. F30V also decreased the percentage of apoptotic cells compared with wild-type (14.8 ± 6.8% versus 19.1 ± 10.1%, p <0.01, without affecting cell-cycle progression) and increased the probability of HBx-Ser-31 being phosphorylated by PI3K-Akt kinase (known to promote anti-apoptotic activity). Conclusions: F30V was closely correlated with HBV-induced HCC in vivo, reduced HBV replicative efficiency by affecting HBx-binding to cccDNA and increased anti-apoptotic HBx activity in vitro. This suggests that F30V (although hampering HBV's replicative capacity) may promote hepatocyte survival, so potentially allowing persistent production of viral progeny and initiating HBV-driven

Sophisticated viral quasispecies with a genotype-related pattern of mutations in the hepatitis B X gene of HBeAg-ve chronically infected patients

“…The HBV QS may evolve differently depending on the genotype, and thereby, differentially influence disease progression and therapy outcome 27 , 28 . Specific mutations in the HBsAg C-terminal domain of genotype D HBV have been associated with viremia < 2000 IU/mL 29 , and certain HBx mutations highly associated with HCC have been reported specifically in genotypes C and D 30 , 31 . Here, we detected a pattern of aa changes (A12S/P33S/P46S/T36G-D) that was highly represented in low-replicative groups, mainly genotype D CI haplotypes.…”

Modern views on the role of X gene of the hepatitis B virus (Hepadnaviridae: Orthohepadnavirus: Hepatitis B virus) in the pathogenesis of the infection it causes

“…Особый интерес представляет обнаружение R. Salpini и соавт. [15] аминокислотной замены F30V в высококонсервативной области N-концевого домена белка Х. Авторы связывают её наличие с угнетением апоптоза повреждённых клеточных элементов, что создаёт условия для развития ГЦК посредством сохранения и накопления клеток с повреждениями в геноме. Тем не менее охарактеризованная замена не связана с ускорением клеточного цикла.…”

“…Как показали результаты многих выполненных к настоящему моменту исследований, феномен канцерогенеза при ВГВ-инфекции обусловлен свойствами и взаимодействиями белков, кодируемых генами вируса, главным образом -взаимодействием белка Х с генами-мишенями и белками клетки организма хозяина. Определение роли гена Х и кодируемого им одноимённого белка в патологическом процессе, инициированном ВГВ, -одно из ведущих исследовательских направлений, реализуемых в различных странах [9][10][11][12][13][14][15][16][17]. Известные на сегодняшний день аминокислотные замены в Х-белке детерминируют появление новых воздействий вируса на гепатоциты, не свойственных его дикому варианту, таких как инициация канцерогенеза, геномная нестабильность, формирование раковых стволовых клеток, активация репликации вируса иммунодефицита человека (Retroviridae: Orthoretrovirinae: Lentivirus: Human immunodeficiency virus) (ВИЧ), ускоренное развитие канцерогенеза при микст-инфекции ХГВ и хронического вирусного гепатита С (ХГС), а также суперинфекция вирусом гепатита D (дельта) (ВГD) [9,10,13,[16][17][18].…”

Association of Hepatitis B Virus DNA Level and Follow-up Interval With Hepatocellular Carcinoma Recurrence

“…Although F30V hampers HBV replication, the variation may be associated with hepatocyte survival, potentially allowing persistent production of viral progeny and initiating HBV-driven hepatocarcinogenesis. 29 ameliorating HBV-induced hepatitis, and promoting the recovery of rescued liver tissue, which provide more chances for patients to receive local and systemic treatment during recurrence, prolonging OS. 31,32 In patients with high levels of active HBV replication, antiviral treatment may be associated with elimination of the virus after antitumor management and with improved OS.…”

“…This study has limitations. The lack of comprehensive analysis of HDQTI, AFP, circulating tumor cells, 29 and imaging characteristics to estimate HCC recurrence is a limitation of this study. In addition, the total data set should ideally have been divided into training and validation data sets to identify the best cutoff for the HDQTI and to assess the performance of the selected cutoff, respectively, which was not done in the current study.…”