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Identification of CD4+ T cell epitopes in C. burnetii antigens targeted by antibody responses.

ABSTRACT: Coxiella burnetii is an obligate intracellular gram-negative bacterium that causes acute Q fever and chronic infections in humans. A killed, whole cell vaccine is efficacious, but vaccination can result in severe local or systemic adverse reactions. Although T cell responses are considered pivotal for vaccine derived protective immunity, the epitope targets of CD4(+) T cell responses in C. burnetii vaccination have not been elucidated. Since mapping CD4(+) epitopes in a genome with over 2,000 ORFs is resource intensive, we focused on 7 antigens that were known to be targeted by antibody responses. 117 candidate peptides were selected from these antigens based on bioinformatics predictions of binding to the murine MHC class II molecule H-2 IA(b). We screened these peptides for recognition by IFN-γ producing CD4(+) T cell in phase I C. burnetii whole cell vaccine (PI-WCV) vaccinated C57BL/6 mice and identified 8 distinct epitopes from four different proteins. The identified epitope targets account for 8% of the total vaccination induced IFN-γ producing CD4(+) T cells. Given that less than 0.4% of the antigens contained in C. burnetii were screened, this suggests that prioritizing antigens targeted by antibody responses is an efficient strategy to identify at least a subset of CD4(+) targets in large pathogens. Finally, we examined the nature of linkage between CD4(+) T cell and antibody responses in PI-WCV vaccinated mice. We found a surprisingly non-uniform pattern in the help provided by epitope specific CD4(+) T cells for antibody production, which can be specific for the epitope source antigen as well as non-specific. This suggests that a complete map of CD4(+) response targets in PI-WCV vaccinated mice will likely include antigens against which no antibody responses are made.

SUBMITTER: Chen C

PROVIDER: S-EPMC3057979 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Reports have shown that activation of tumor-specific CD4(+) helper T (Th) cells is crucial for effective anti-tumor immunity and identification of Th-cell epitopes is critical for peptide vaccine-based cancer immunotherapy. Although computer algorithms are available to predict peptides with high binding affinity to a specific HLA class II molecule, the ability of those peptides to induce Th-cell responses must be evaluated. We have established HLA-DR4 (HLA-DRA*01:01/HLA-DRB1*04:05) transgenic mice (Tgm), since this HLA-DR allele is most frequent (13.6%) in Japanese population, to evaluate HLA-DR4-restricted Th-cell responses to tumor-associated antigen (TAA)-derived peptides predicted to bind to HLA-DR4. To avoid weak binding between mouse CD4 and HLA-DR4, Tgm were designed to express chimeric HLA-DR4/I-E(d), where I-E(d) ?1 and ?1 domains were replaced with those from HLA-DR4. Th cells isolated from Tgm immunized with adjuvant and HLA-DR4-binding cytomegalovirus-derived peptide proliferated when stimulated with peptide-pulsed HLA-DR4-transduced mouse L cells, indicating chimeric HLA-DR4/I-E(d) has equivalent antigen presenting capacity to HLA-DR4. Immunization with CDCA155-78 peptide, a computer algorithm-predicted HLA-DR4-binding peptide derived from TAA CDCA1, successfully induced Th-cell responses in Tgm, while immunization of HLA-DR4-binding Wilms' tumor 1 antigen-derived peptide with identical amino acid sequence to mouse ortholog failed. This was overcome by using peptide-pulsed syngeneic bone marrow-derived dendritic cells (BM-DC) followed by immunization with peptide/CFA booster. BM-DC-based immunization of KIF20A494-517 peptide from another TAA KIF20A, with an almost identical HLA-binding core amino acid sequence to mouse ortholog, successfully induced Th-cell responses in Tgm. Notably, both CDCA155-78 and KIF20A494-517 peptides induced human Th-cell responses in PBMCs from HLA-DR4-positive donors. Finally, an HLA-DR4 binding DEPDC1191-213 peptide from a new TAA DEPDC1 overexpressed in bladder cancer induced strong Th-cell responses both in Tgm and in PBMCs from an HLA-DR4-positive donor. Thus, the HLA-DR4 Tgm combined with computer algorithm was useful for preliminary screening of candidate peptides for vaccination.

Project description:Coxiella burnetii, the causative agent of Q fever, is a Gram-negative intracellular bacterium transmitted via aerosol. Regulatory approval of the Australian whole-cell vaccine Q-VAX® in the US and Europe is hindered by reactogenicity in previously exposed individuals. The aim of this study was to identify and rationally select C. burnetii epitopes for design of a safe, effective, and less reactogenic T-cell targeted human Q fever vaccine. Immunoinformatic methods were used to predict 65 HLA class I epitopes and 50 promiscuous HLA class II C. burnetii epitope clusters, which are conserved across strains of C. burnetii. HLA binding assays confirmed 89% of class I and 75% of class II predictions, and 11 HLA class II epitopes elicited IFN? responses following heterologous DNA/DNA/peptide/peptide prime-boost immunizations of HLA-DR3 transgenic mice. Human immune responses to the predicted epitopes were characterized in individuals naturally exposed to C. burnetii during the 2007-2010 Dutch Q fever outbreak. Subjects were divided into three groups: controls with no immunological evidence of previous infection and individuals with responses to heat-killed C. burnetii in a whole blood IFN? release assay (IGRA) who remained asymptomatic or who experienced clinical Q fever during the outbreak. Recall responses to C. burnetii epitopes were assessed by cultured IFN? ELISpot. While HLA class I epitope responses were sparse in this cohort, we identified 21 HLA class II epitopes that recalled T-cell IFN? responses in 10-28% of IGRA+ subjects. IGRA+ individuals with past asymptomatic and symptomatic C. burnetii infection showed a comparable response pattern and cumulative peptide response which correlated with IGRA responses. None of the peptides elicited reactogenicity in a C. burnetii exposure-primed guinea pig model. These data demonstrate that a substantial proportion of immunoinformatically identified HLA class II epitopes show long-lived immunoreactivity in naturally infected individuals, making them desirable candidates for a novel human multi-epitope Q fever vaccine.

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Identification of CD4+ T cell epitopes in C. burnetii antigens targeted by antibody responses.

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