Human T cell responses to a semi-conserved sequence of the malaria vaccine candidate antigen MSP-1

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Over the past decade of malaria research, there has been considerable progress in the understanding of immune mechanisms involved in conferring protection to malaria and in the identification of vaccine candidate antigens. Despite this increasing knowledge, there is still no effective malaria vaccine available. Current vaccine development concentrates on multicomponent, multi-stage subunit vaccines in combination with improved delivery systems. In addition, an ideal malaria vaccine should induce both cellular and humoral immune responses and therefore requires the incorporation of T cell- as well as B cell-epitopes. The merozoite surface protein 1 (MSP-1) of Plasmodium falciparum is one of the most promising vaccine candidate antigens. A semi-conserved region at its N-terminus eliciting protective immune responses in malaria models has been incorporated into the synthetic peptide vaccine SPf66. This sequence of MSP-1 was found to be a suitable human B cell epitope, eliciting parasite-binding antibodies. In this thesis the question has been addressed whether the same region of MSP-1 also represents a suitable T cell epitope and whether semiconserved sequences are suitable elements for epitope-focussed vaccines. In addition, residues of sequence 38-58 of MSP-1 interacting with the T cell receptor (TCR) were mapped, and the potential of pseudopeptide analogues for T cell activation was explored. Furthermore, MSP- 138-58 and SPf66 were used to test a Herpesvirus saimiri-based system for T cell cloning and to assess the potency of a new adjuvant. Human MSP-138-58-specific T cell lines and clones were generated from SPf66- vacinated volunteers. The T cell clones were CD4+, mainly of Th2 type, and exhibited a high specificity for the particular sequence variant (S 44 Q47 V52) present in the vaccine: None of the four other naturally occurring variants of the semi-conserved region of MSP-1 found in P. falciparum populations stimulated T cell proliferation or cytokine secretion, although all variants exhibited activity in HLA-DR peptide binding competition assays. Thus MSP-1 38-58 although a potent stimulator of T cells, does not appear to be a suitable vaccine epitope. Multiple genetic restriction elements were used by the T cell clones to recognize MSP- 138-58. DR- and DP-restricted clones were found to recognize overlapping, but distinct, epitopes clustered within the core region of MSP-1 38-58. Substitution of individual amino acids with alanine or glycine revealed that only about nine residues of the presented peptide are “read out” by the TCR although additional epitope-flanking regions are required for T cell stimulation as well. The contribution of the peptide backbone itself to T cell activation and HLA-DRbinding was assessed with reduced-amide pseudopeptide analogues of MSP-1. Some pseudopeptides exhibited even better stimulatory activity while others were less potent than their parent peptide. Thus the peptide backbone appears to contribute critically to MHC binding and TCR triggering. Pseudopeptides, which generally exhibit decreased protease susceptibility and a better reproduction of conformational B cell epitopes, might advantageously replace natural peptides in future vaccines. Investigations of cellular immune responses on the clonal level during clinical vaccine trials are hampered by the limited volume of available blood samples. A method to generate antigen-specific T cell clones using Herpesvirus Saimiri (HVS)-transformed autologous T cells as antigen presenting cells (APCs) was established. MSP-1 38-58 specific and SPf66- specific T cell clones were generated by using either autologous PBMCs or HVS-transformed T cells as APCs. The resulting panels of T cell clones exhibited similar characteristics and identical TCR rearrangements were found in both panels. HSV transformation is thus a useful method for detailed analysis of T cell responses in the course of clinical vaccine trials where only small amounts of blood cells are available. The immunogenicity of a synthetic peptide vaccine depends on the delivery system or adjuvant. To investigate the immunogenicity of a new formulation of SPf66 in combination with the saponin adjuvant QS-21, SPf66-specific T cell lines were generated from SPf66/QS- 21 vaccinated volunteers and compared to lines generated from SPf66/alum vaccinated persons. The T cell responses elicited by the two SPf66-formulations differed significantly: Vaccination in combination with QS-21 induced both CD4+ and CD8+ T cell responses while SPf66/alum vaccination induced predominantly CD4+ T cell responses of the Th2 subtype. QS-21 is therefore a promising candidate for the delivery of the next generation of malaria vaccines.