Identification of an HLA-A*0201-restricted CD8+ T-cell epitope SSp-1 of SARS-CoV spike protein - PubMed (original) (raw)

. 2004 Jul 1;104(1):200-6.

doi: 10.1182/blood-2003-11-4072. Epub 2004 Mar 11.

Huabiao Chen, Xiaodong Jiang, Minghui Zhang, Tao Wan, Nan Li, Xiangyang Zhou, Yanfeng Wu, Feng Yang, Yizhi Yu, Xiaoning Wang, Ruifu Yang, Xuetao Cao

Affiliations

Identification of an HLA-A*0201-restricted CD8+ T-cell epitope SSp-1 of SARS-CoV spike protein

Baomei Wang et al. Blood. 2004.

Abstract

A novel coronavirus, severe acute respiratory syndrome (SARS)-associated coronavirus (SARS-CoV), has been identified as the causal agent of SARS. Spike (S) protein is a major structural glycoprotein of the SARS virus and a potential target for SARS-specific cell-mediated immune responses. A panel of S protein-derived peptides was tested for their binding affinity to HLA-A*0201 molecules. Peptides with high affinity for HLA-A*0201 were then assessed for their capacity to elicit specific immune responses mediated by cytotoxic T lymphocytes (CTLs) both in vivo, in HLA-A2.1/K(b) transgenic mice, and in vitro, from peripheral blood lymphocytes (PBLs) harvested from healthy HLA-A2.1(+) donors. SARS-CoV protein-derived peptide-1 (SSp-1 RLNEVAKNL), induced peptide-specific CTLs both in vivo (transgenic mice) and in vitro (human PBLs), which specifically released interferon-gamma (IFN-gamma) upon stimulation with SSp-1-pulsed autologous dendritic cells (DCs) or T2 cells. SSp-1-specific CTLs also lysed major histocompatibility complex (MHC)-matched tumor cell lines engineered to express S proteins. HLA-A*0201-SSp-1 tetramer staining revealed the presence of significant populations of SSp-1-specific CTLs in SSp-1-induced CD8(+) T cells. We propose that the newly identified epitope SSp-1 will help in the characterization of virus control mechanisms and immunopathology in SARS-CoV infection, and may be relevant to the development of immunotherapeutic approaches for SARS.

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Figures

Figure 1

Figure 1

Immunogenicity of SARS-S–derived peptide SSp-1 in HLA-A2.1/Kb transgenic mice. IFN-γ release by bulk CTLs from immunized mice was detected by ELISPOT assay (A) and lytic activity was tested using a standard 4-hour chromium release assay (B). (A) Bulk CTLs from SSp-1–immunized mice released IFN-γ in response to T2 cells pulsed with SSp-1 (T2/SSp-1) in a dose-dependent manner, but not those with irrelevant peptide CAP-1 (T2/CAP-1) or T2 cells alone. (B) Bulk CTLs from SSp-1–immunized mice lysed T2 cells loaded with SSp-1, but not T2 cells pulsed with the irrelevant peptide CAP-1 or T2 cells alone. Data represent means ± SD.

Figure 2

Figure 2

Specific reactivity of human CTLs induced by SSP-1–loaded DCs in vitro. SSp-1–specific CTLs were generated from the PBMCs of 7 of 11 HLA-A2.1+ healthy donors through 4 sequential rounds of stimulation with SSp-1–pulsed DCs. Resulting CTLs were tested for IFN-γ release (A-B) and SSp-1–specific lysis (C) using an ELISPOT assay and a standard 4-hour chromium release assay. Autologous DCs (A) and T2 cells (B) were pulsed with indicated concentrations of SSp-1 (DC/SSp-1, T2/SSp-1) or irrelevant peptide CAP-1 (DC/CAP-1, T2/CAP-1), and then used as stimulators in an IFN-γ release assay. (C) CTLs lysed T2 cells loaded with SSp-1, but not T2 cells loaded with the irrelevant peptide CAP-1 or T2 cells alone. Results are representative of 3 independent experiments. Data represent means ± SD.

Figure 3

Figure 3

Identification of SSp-1 as a naturally processed and presented HLA-A2.1–restricted epitope. DC-induced human CTLs against SSp-1 were tested for specific lytic activity (A) and IFN-γ release (B) in response to genetically modified cell lines. (A) CTLs lysed pAd-SARS/S–transduced HLA-A2.1+ SW480 cells (SW480/SARS/S) but did not lyse pAd-SARS/S–transduced HLA-A2.1– HT29 cells (HT29/SARS/S) or pAd-LacZ–transduced SW480 (SW480/LacZ) and HT29 cells (HT29/LacZ). (B) CTLs elicited strong IFN-γ production in response to pAd-SARS/S–transduced SW480 cells, but only very low levels were elicited by pAd-SARS/S–transduced HT29 cells or pAd-LacZ–transduced cell lines. Results are representative of 3 independent experiments. Data represent means ± SD.

Figure 4

Figure 4

Frequency of SSp-1–specific CD8+ T cells. SSp-1–specific CD8+ T-cell frequency among DC-induced human CTLs against SSp-1 (A-B) and splenocytes derived from immunized mice and restimulated for 6 days in vitro (C-D). Tetramer-binding CD8+ T cells are shown in the upper-right quadrant and are labeled with the percentage of total CD8+ T cells. SSp-1–specific CTLs generated from human PBLs were stained with control HLA-A*0201/CAP-1 tetramers (A) and HLA-A*0201/SSp-1 tetramers (B). Results are representative of 3 independent experiments. Splenocytes from Tg mice immunized with SSp-1–pulsed DCs were stained with control HLA-A*0201/CAP-1 tetramers (C) and HLA-A*0201/SSp-1 tetramers (D).

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