Research Article: T-Cell Memory Responses Elicited by Yellow Fever Vaccine are Targeted to Overlapping Epitopes Containing Multiple HLA-I and -II Binding Motifs

Date Published: January 31, 2013

Publisher: Public Library of Science

Author(s): Andréa Barbosa de Melo, Eduardo J. M. Nascimento, Ulisses Braga-Neto, Rafael Dhalia, Ana Maria Silva, Mathias Oelke, Jonathan P. Schneck, John Sidney, Alessandro Sette, Silvia M. L. Montenegro, Ernesto T. A. Marques, Alan L. Rothman.

Abstract: The yellow fever vaccines (YF-17D-204 and 17DD) are considered to be among the safest vaccines and the presence of neutralizing antibodies is correlated with protection, although other immune effector mechanisms are known to be involved. T-cell responses are known to play an important role modulating antibody production and the killing of infected cells. However, little is known about the repertoire of T-cell responses elicited by the YF-17DD vaccine in humans. In this report, a library of 653 partially overlapping 15-mer peptides covering the envelope (Env) and nonstructural (NS) proteins 1 to 5 of the vaccine was utilized to perform a comprehensive analysis of the virus-specific CD4+ and CD8+ T-cell responses. The T-cell responses were screened ex-vivo by IFN-γ ELISPOT assays using blood samples from 220 YF-17DD vaccinees collected two months to four years after immunization. Each peptide was tested in 75 to 208 separate individuals of the cohort. The screening identified sixteen immunodominant antigens that elicited activation of circulating memory T-cells in 10% to 33% of the individuals. Biochemical in-vitro binding assays and immunogenetic and immunogenicity studies indicated that each of the sixteen immunogenic 15-mer peptides contained two or more partially overlapping epitopes that could bind with high affinity to molecules of different HLAs. The prevalence of the immunogenicity of a peptide in the cohort was correlated with the diversity of HLA-II alleles that they could bind. These findings suggest that overlapping of HLA binding motifs within a peptide enhances its T-cell immunogenicity and the prevalence of the response in the population. In summary, the results suggests that in addition to factors of the innate immunity, “promiscuous” T-cell antigens might contribute to the high efficacy of the yellow fever vaccines.

Partial Text: The yellow fever (YF) vaccines (YF-17D-204 and 17DD) are considered to be among the most effective vaccines [1], [2]. Antibody and T-cell responses are believed to mediate protection [3], [4], [5], and recent studies have also implicated the innate immune responses as one of the critical elements for developing the immune responses [6]. However, the immune adaptive mechanisms that make this vaccine so highly effective are unclear. T-cell immune responses against YF wild type virus and other flaviviruses, such as dengue and West Nile virus [7], [8], are considered to be important for development of neutralizing antibodies, and activation of CD4+ helper T-cells and CD8+ cytotoxic T lymphocytes (CTLs) against YF wild type virus has been reported [6], [9], [10]. The CTL responses appear 14 days after vaccination and these cells differentiate into long-lived memory T-cells after a few months [11]; however, only a few YF wild type virus T-cell epitopes have been described in humans [12], [13].

This study presents the screening of 653 peptides of the YF-17DD Env, NS1, NS2, NS3, NS4 and NS5 proteins in the context of a cohort of healthy adults immunized with the YF-17DD vaccine. Considering the two rounds of peptide screening, each peptide was tested in at least 75 individuals, and in the case of the Env protein each peptide was tested in 208 volunteers. The screening allowed the identification of 16 T-cell peptides that were immunogenic in 10% or more of the individuals tested. Only a few YF-17D epitopes have been characterized previously [12], [13], [24]. Overall, among the 16 immunogenic peptides identified herein, 14 contain new human T-cell antigens (Env57–71, Env65–79, Env73–87, Env337–351, Env345–359, Env361–375, NS2b97–111, NS3137–151, NS4a197–211, NS5341–355, NS5345–359, NS5465–479, NS5469–483, NS5481–495). Two peptides, NS2b113–127[12] and the NS4b77–91[24], contained epitopes previously described in humans, while the Env57–71 and NS2b113–127 peptides contain epitopes that have been described in the murine H2d background. The murine epitope (Env57–71) was reported to be able to stimulate both CD8+ and CD4+ T-cells to secrete IFN-γ in YF-17DD vaccine immunized H2 d mice and also HLA A02, B07 and A24 transgenic mice [29], [30]. These highly prevalent immunogenic peptides were shown to contain multiple HLA binding motifs and that the degree of prevalence of its immunogenicity was correlated with the HLA promiscuity. Previous studies have shown some degree of correlation between predicted binding affinity and immunogenicity [30]. However, additional studies are required to determine the precise breath and differences in functionalities of these immunogenic peptides in different HLA contexts.