Identification of hepatitis C virus-specific HLA-A*1101-and A*2402-restricted CD8+ T-cell epitopes#br
1.Department of Microbiology and Immunology, Wenzhou Medical University, Wenzhou, 325035; 2.Department of Laboratory, the Second Affiliated Hospital of Wenzhou Medical University, Wenzhou, 325027; 3.Department of Laboratory Ruian People’s Hospital, Wenzhou, 325200; 4.Central Blood Station of Wenzhou, Wenzhou, 325000
ZHONG Xiaozhi1,DUAN Zhiliang2,GUO Jianglong1, et al. Identification of hepatitis C virus-specific HLA-A*1101-and A*2402-restricted CD8+ T-cell epitopes#br[J]. JOURNAL OF WEZHOU MEDICAL UNIVERSITY, 2014, 44(8): 547-554.
Abstract：Objective: To identify hepatitis C virus (HCV)-specific HLA-A*1101- and A*2402-restricted CD8+ T-cell epitopes. Methods: HCV-specific CD8+ T-cell epitopes were predicted using T epitope prediction software (SYFPEITHI) and then HCV-specific HLA-A*1101 and A*2402-restricted epitope candidates were selected and synthesized. Immortalized HLA-A*1101-positive and A*2402-positive B cell lines were established. Based on above immortalized B cell lines, competitive peptide-binding assay was used to evaluate the binding affinity of epitope candidates for HLA-A*1101 and A*2402 molecules, respectively. Epitope candidates were used to stimulate PBMCs from HLA-A*1101-positive or A*2402-positive HCV-infected patients. Enzyme-linked immunospot (ELISPOT) and intracellular cytokine staining (ICS) assay were applied to measure the frequencies of IFN-γ-secreting cells in total PBMCs and the percentages of IFN-γ+ CD8+ T cells in total CD8+ T. Results: Of five HLA-A*1101-restricted epitope candidates, NS3_609 (ITLTHPITK) and NS2_165 (VVFSDMETK) had a high affinity for HLA-A*1101 molecules. Among three HLA-A*2402-restricted epitope candidates, NS3_373 (KCDELASKL) and NS5b_382 (YYLTRDPTI) had a high affinity for HLA-A*2402 molecules. Furthermore, high levels of NS3_609- and NS2_165-specific IFN-γ-secreting CD8+ T cells were detected in HLA-A*1101-positive HCV-infected patients, whereas high levels of NS3_373- and NS5b_382-specific IFN-γ-secreting CD8+ T cells were detected in HLA-A*2402 positive HCV-infected patients. Conclusion: Our Results strongly confirmed that NS3_609 (ITLTHPITK) and NS2_165 (VVFSDMETK) are novel HCV-specific HLA-A*1101-restricted CD8+ T-cell epitopes while NS3_373 (KCDELASKL) and NS5b_382 (YYLTRDPTI) are novel HCV-specific HLA-A*2402-restricted CD8+ T-cell epitopes.
[1] Lohmann V, Koch JO, Bartenschlager R. Processing path-
ways of the hepatitis C virus proteins[J]. J Hepatol, 1996,
24(2 Suppl): 11-19.
[2] Amoroso P, Rapicetta M, Tosti ME, et al. Correlation be-
tween virus genotype and chronicity rate in acute hepatitis
C[J]. J Hepatol, 1998, 28(6): 939-944.
[3] Bowen DG, Walker CM. Adaptive immune responses in
acute and chronic hepatitis C virus infection[J]. Nature,
2005, 436(7053): 946-952.
[4] Torresi J, Bharadwaj M, Jackson DC, et al. Neutralising an-
tibody, CTL and dendritic cell responses to hepatitis C virus:
a preventive vaccine strategy[J]. Curr Drug Targets, 2004,
5(1): 41-56.
[5] Hu SP, Luan JA, Li B, et al. Genetic link between Cha-
oshan and other Chinese Han populations: Evidence From
HLA-A and HLA-B allele frequency distribution[J]. Am J
Phys Anthropol, 2007, 132(1), 140-150.
[6] Dikopoulos N, Riedl P, Schirmbeck R, et al. Novel peptide-
based vaccines efficiently prime murine “help”-indepen-
dent CD8+ T cell responses in the liver[J]. Hepatology, 2004,
40(2): 300-309.
[7] Kurokohchi K, Arima K, Nishioka M. A novel cytotoxic T-
cell epitope presented by HLA-A24 molecule in hepatitis C
virus infection[J]. J Hepatol, 2001, 34(6): 930-935.
[8] 陈俊, 段志良, 巩文词, 等. 人白细胞抗原-A*0201限制性
丙型肝炎病毒细胞毒性T淋巴细胞表位的鉴定[J]. 中华传
染病杂志, 2010, 28(11): 651-655.
[9] 段志良, 张丽芳, 张琴, 等. 丙型肝炎病毒CTL表位的HLA-
A2限制性及其免疫学效应研究[J]. 中华微生物学和免疫
学杂志, 2009, 29(9): 822-826.
[10] Reche PA, Glutting JP, Reinherz EL. Prediction of MHC
class I binding peptides using profile motifs[J]. Hum Immu-
nol, 2002, 63(9): 701-709.
[11] Gomez-Nunez M, Pinilla-Ibarz J, Dao T, et al. Peptide bind-
ing motif predictive algorithms correspond with experi-
mental binding of leukemia vaccine candidate peptides to
HLA-A*0201 molecules[J]. Leuk Res, 2006, 30(10): 1293-
1298.
[12] Chang KM, Gruener NH, Southwood S, et al. Identification
of HLA-A3 and -B7-restricted CTL response to hepatitis C
virus in patients with acute and chronic hepatitis C[J]. J Im-
munol, 1999, 162(2): 1156-1164.
[13] Nakamoto Y, Kaneko S, Takizawa H, et al. Analysis of the
CD8-positive T cell response in Japanese patients with
chronic hepatitis C using HLA-A*2402 peptide tetramers[J].
J Med Virol, 2003, 70(1): 51-61.
[14] Nilsson K, Klein G. Phenotypic and cytogenitic characteris-
tics of human B lymphoid cell lines and their relevance for
the etiology of Burkitt’s lymphoma[J]. Adv Cancer Res,
1982, 37: 319-380.
[15] Petersen J, Wurzbacher SJ, Williamson NA, et al. Phosphor-
ylated self-peptides alter human leukocyte antigen class
I-restricted antigen presentation and generate tumor-specific
epitopes[J]. Proc Natl Acad Sci USA, 2009, 106(8): 2776-
2781.
[16] 文金生, 段志良, 陈俊, 等. 温州地区62例HCV感染者的基
因型特征研究[J]. 临床血液学杂志, 2009, 22(1): 70-72.
[17] Simmons CP, Dong T, Chau NV, et al. Early T-cell responses
to dengue virus epitopes in Vietnamese adults with second-
ary dengue virus infections[J]. J Virol, 2005, 79(9): 5665-
5675.
[18] Farci P, Alter HJ, Wong DC, et al. Prevention of hepati-
tis C virus infection in chimpanzees after antibody-mediated
in vitro neutralization[J]. Proc Natl Acad Sci USA, 1994,
91(16): 7792-7796.
[19] Prince AM. Challenges for development of hepatitis C virus
vaccines[J]. FEMS Microbiol Rev, 1994, 14(3): 273-277.
[20] Gruner NH, Gerlach TJ, Jung MC, et al. Association of hep-
atitis C virus-specific CD8+T cells with viral clearance in
acute hepatitis C[J]. J Infect Dis, 2000, 181(5): 1528-1536.
[21] Grakoui A, Shoukry NH, Woollard DJ, et al. HCV persis-
tence and immune evasion in the absence of memory T cell
help[J]. Science, 2003, 302(5645): 659-662.
[22] Nascimbeni M, Mizukoshi E, Bosmann M, et al. Kinetics of
CD4+ and CD8+ memory T-cell responses during hepatitis
C virus rechallenge of previously recovered chimpanzees[J].
J Virol, 2003, 77(8): 4781-4793.
[23] Shoukry NH, Grakoui A, Houghton M, et al. Memory CD8+
T cells are required for protection from persistent hepatitis C
virus infection[J]. J Exp Med, 2003, 197(12): 1645-1655.
[24] Thimme R, Oldach D, Chang MK, et al. Determinants of
viral clearance and persistence during acute hepatitis C virus
infection[J]. J Exp Med, 2001, 194(10): 1395-1406.
[25] Major ME, Mihalik K, Puig M, et al. Previously infected
and recovered chimpanzees exhibit rapid responses that con-
trol hepatitis C virus replication upon rechallenge[J]. J Virol,
2002, 76(13): 6586-6595.
[26] Duan ZL, Li Q, Wang ZB, et al. HLA-A*0201-restricted
CD8+ T-cell epitopes identified in dengue viruses[J]. Virol J,
2012, 9: 259.
[27] Wen JS, Duan ZL, Jiang LF. Identification of a dengue vi-
rus-specific HLA-A*0201-restricted CD8(+) T cell epitope
[J]. J Med Virol, 2010, 82(4): 642-648.
[28] Al-Ramadi BK, Jelonek MT, Boyd LF, et al. Lack of strict
correlation of functional sensitization with the apparent af-
finity of MHC/peptide complexes for the TCR[J]. J Immu-
nol, 1995, 155(2): 662-673.
