Human immunome, bioinformatic analyses using HLA supermotifs and the parasite genome, binding assays, studies of human T cell responses, and immunization of HLA-A*1101 transgenic mice including novel adjuvants provide a foundation for HLA-A03 restricted CD8+T cell epitope based, adjuvanted vaccine protective against Toxoplasma gondii

Abstract

BackgroundToxoplasmosis causes loss of life, cognitive and motor function, and sight. A vaccine is greatly needed to prevent this disease. The purpose of this study was to use an immmunosense approach to develop a foundation for development of vaccines to protect humans with the HLA-A03 supertype. Three peptides had been identified with high binding scores for HLA-A03 supertypes using bioinformatic algorhythms, high measured binding affinity for HLA-A03 supertype molecules, and ability to elicit IFN-γ production by human HLA-A03 supertype peripheral blood CD8+ T cells from seropositive but not seronegative persons.ResultsHerein, when these peptides were administered with the universal CD4+T cell epitope PADRE (AKFVAAWTLKAAA) and formulated as lipopeptides, or administered with GLA-SE either alone, or with Pam2Cys added, we found we successfully created preparations that induced IFN-γ and reduced parasite burden in HLA-A*1101(an HLA-A03 supertype allele) transgenic mice. GLA-SE is a novel emulsified synthetic TLR4 ligand that is known to facilitate development of T Helper 1 cell (TH1) responses. Then, so our peptides would include those expressed in tachyzoites, bradyzoites and sporozoites from both Type I and II parasites, we used our approaches which had identified the initial peptides. We identified additional peptides using bioinformatics, binding affinity assays, and study of responses of HLA-A03 human cells. Lastly, we found that immunization of HLA-A*1101 transgenic mice with all the pooled peptides administered with PADRE, GLA-SE, and Pam2Cys is an effective way to elicit IFN-γ producing CD8+ splenic T cells and protection. Immunizations included the following peptides together: KSFKDILPK (SAG1224-232); AMLTAFFLR (GRA6164-172); RSFKDLLKK (GRA7134-142); STFWPCLLR (SAG2C13-21); SSAYVFSVK(SPA250-258); and AVVSLLRLLK(SPA89-98). This immunization elicited robust protection, measured as reduced parasite burden using a luciferase transfected parasite, luciferin, this novel, HLA transgenic mouse model, and imaging with a Xenogen camera.ConclusionsToxoplasma gondii peptides elicit HLA-A03 restricted, IFN-γ producing, CD8+ T cells in humans and mice. These peptides administered with adjuvants reduce parasite burden in HLA-A*1101 transgenic mice. This work provides a foundation for immunosense based vaccines. It also defines novel adjuvants for newly identified peptides for vaccines to prevent toxoplasmosis in those with HLA-A03 supertype alleles.

Adjuvanted multi-epitope vaccines protect HLA-A*11:01 transgenic mice against Toxoplasma gondii

“…In these linked and single-peptide lipopeptides, responses are much less or absent (3,5). Other arrangements of sequence in DNA vaccines encoding peptides in DNA vaccine or peptides in a nonionic surfactant vesicle (NISV) also elicited less robust response or were or absent (data not shown).…”

“…Our recent work has focused on development of an epitope-based vaccine to prevent toxoplamosis to enhance host immunity by stimulating efficacious responses of CD4 + helper T lymphocytes (HTLs) and CD8 + IFN-γ-producing T lymphocytes (3-6) for humans. These CD8 + T cells recognize the octamer/nonamer peptides presented by HLA supermotif major histocompatibility complex class I (MHC-I) molecules on antigen presenting cells (APCs) in humans (3)(4)(5)(6). Considerable effort has been made to identify promising vaccine candidate antigens for Toxoplasma gondii…”

“…Using bioinformatic analyses, multiple predicted CD8 + epitopes of tachyzoite-and bradyzoite-specific antigens have recently entered our vaccine development pipeline (3)(4)(5)(6). Candidate amino acid sequences are from surface antigens (SAG1 and SUSA1) and secreted proteins, including dense granule proteins (GRA2, GRA3, GRA6, GRA7) and rhoptry proteins (ROP2, ROP16, ROP18).…”

“…Candidate amino acid sequences are from surface antigens (SAG1 and SUSA1) and secreted proteins, including dense granule proteins (GRA2, GRA3, GRA6, GRA7) and rhoptry proteins (ROP2, ROP16, ROP18). We reported in our previous studies CD8 + epitopes derived from these antigens that can be presented by HLA-A02, HLA-A03, and HLA-B*07 supermotif human MHC molecules (3)(4)(5)(6). These CD8 + T cell-restricted peptides -administered with a universal helper T cell epitope, PADRE, and adjuvants -elicit IFN-γ and reduce parasite burden in HLA-B*07:02, HLA-A*11:01, and HLA-A*02:01 transgenic mice.…”

Identification and bioinformatic analysis of a putative calcium-dependent protein kinase (CDPK6) from Toxoplasma gondii

“…Analyses of the structural contents of an unknown protein to predict its immunity by bioinformatics contributed to the development of a new vaccine candidate antigen against T. gondii (Rosenberg et al, 2009;Cong et al, 2010;Bai et al, 2012). The secondary structures, such as the random coils exposed on the surface of the protein, generally make a significant impact on the epitopes.…”