Novel HBsAg mutations correlate with hepatocellular carcinoma, hamper HBsAg secretion and promote cell proliferation in vitro

Abstract

BackgroundAn impaired HBsAg-secretion can increase HBV oncogenic-properties. Here, we investigate genetic-determinants in HBsAg correlated with HBV-induced hepatocellular carcinoma (HCC), and their impact on HBsAg-secretion and cell-proliferation.MethodsThis study included 128 chronically HBV-infected patients: 23 with HCC (73.9% D; 26.1% A HBV-genotype), and 105 without cirrhosis/HCC (72.4% D, 27.6% A) as reference-group. The impact of mutations on HBsAg-secretion was assessed by measuring the ratio [secreted/intracellular HBsAg] until day 5 post-transfection. The impact of mutations on cell-cycle advancement was assessed by flow-cytometry.ResultsTwo HBsAg mutations significantly correlated with HCC: P203Q (17.4% [4/23] in HCC vs 1.0% [1/105] in non-HCC, P=0.004); S210R (34.8% [8/23] in HCC vs 3.8% [4/105] in non-HCC, P <0.001); P203Q+S210R (17.4% [4/23] in HCC vs 0% [0/110] in non-HCC, P=0.001). Both mutations reside in trans-membrane C-terminal domain critical for HBsAg-secretion. In in-vitro experiments, P203Q, S210R and P203Q+S210R significantly reduced the ratio [secreted/intracellular HBsAg] compared to wt at each time-point analysed (P <0.05), supporting an impaired HBsAg-secretion. Furthermore, P203Q and P203Q+S210R increased the percentage of cells in S-phase compared to wt, indicating cell-cycle progression (P203Q:26±13%; P203Q+S210R:29±14%; wt:18%±9, P <0.01. Additionally, S210R increased the percentage of cells in G2/M-phase (26±8% for wt versus 33±6% for S210R, P <0.001).ConclusionsSpecific mutations in HBsAg C-terminus significantly correlate with HBV-induced HCC. They hamper HBsAg-secretion and are associated with increased cellular proliferation, supporting their involvement in HCC-development. The identification of viral genetic markers associated with HCC is critical to identify patients at higher HCC-risk that may deserve intensive liver monitoring, and/or early anti-HBV therapy.

Hepatitis B virus pre-S/S variants in liver diseases

“…Mutations in this domain can result in a stable, glycosylated, but nonsecreted chain, thus affecting the biogenesis and secretion of subviral particles[ 77 ]. Two C-terminus S mutations were found and significantly correlated with HCC: P203Q (4/23, 17.4% in HCC vs 1/105, 1.0 in non-HCC, P = 0.004); S210R (8/23, 34.8% in HCC vs 4/105, 3.8% in non-HCC, P < 0.001); P203Q + S210R (4/23, 17.4% in HCC vs 0/110, 0 in non-HCC, P = 0.001)[ 78 ]. In vitro experiments revealed that P203Q, S210R, and P203Q+S210R significantly reduced the ratio of secreted and intracellular HBsAg compared with WT at each time point analyzed ( P < 0.05); P203Q and P203Q+S210R increased the percentage of cells in S-phase compared with WT (P203Q: 26% ± 13%; P203Q+S210R: 29% ± 14%; WT: 18% ± 9%, P < 0.01); S210R increased the percentage of cells in the G2/M-phase (33% ± 6% for S210R vs 26% ± 8% for WT, P < 0.001)[ 78 ].…”

“…Two C-terminus S mutations were found and significantly correlated with HCC: P203Q (4/23, 17.4% in HCC vs 1/105, 1.0 in non-HCC, P = 0.004); S210R (8/23, 34.8% in HCC vs 4/105, 3.8% in non-HCC, P < 0.001); P203Q + S210R (4/23, 17.4% in HCC vs 0/110, 0 in non-HCC, P = 0.001)[ 78 ]. In vitro experiments revealed that P203Q, S210R, and P203Q+S210R significantly reduced the ratio of secreted and intracellular HBsAg compared with WT at each time point analyzed ( P < 0.05); P203Q and P203Q+S210R increased the percentage of cells in S-phase compared with WT (P203Q: 26% ± 13%; P203Q+S210R: 29% ± 14%; WT: 18% ± 9%, P < 0.01); S210R increased the percentage of cells in the G2/M-phase (33% ± 6% for S210R vs 26% ± 8% for WT, P < 0.001)[ 78 ]. These results show that these two C-terminus S mutations, P203Q and S210R, hamper HBsAg secretion and are associated with increased cellular proliferation, supporting their involvement in HCC development.…”

“…Type V-C-terminus S mutants influence protein folding in the ER membrane, thus impairing HBsAg release, resulting in its accumulation in specific intracellular compartments (presumably represented by the ER and Golgi apparatus) and in turn contributing to cell proliferation[ 78 ]. An in vitro study revealed that C-terminus S mutants can also activate the proliferation control[ 78 ].…”

Personalized subcutaneous administration of hepatitis B surface antibodies without nucleos(t)ide analogs for patients at risk of renal failure after liver transplantation: a prospective single center cohort study

“…This occurs in NA monotherapy . It has been demonstrated that this large protein is oncogenic . In this study, 37% of the patients were transplanted for HCC.…”

IL-18 polymorphisms contribute to hepatitis B virus-related cirrhosis and hepatocellular carcinoma susceptibility in Chinese population: a case-control study

“…Genomic mutations were significantly involved in the development of HBV-related HCC. P203Q and S210R which are mutations in HBsAg C-terminus hamper HBsAg-secretion and correlate with increased cellular proliferation and HBV-induced HCC [ 5 ]. A meta-analysis performed by Tian T et al indicated that miR-146a C>G increased HBV-related HCC risk while miR-196a-2 C>T decreased the risk of HBV-related HCC, especially in the Chinese population [ 6 ].…”