Monitoring of human cellular immune responses in infectious diseases : malaria vaccine associated responses and immune characterization of Buruli ulcer patients

摘要

Within the context of the establishment of a virosomal-based subunit malaria vaccine including synthetic peptides derived from Plasmodium falciparum antigens demonstrated that it is possible to improve immunogenic properties of synthetic peptides in a medicinal chemistry type optimization process. A close to natural conformation of peptides appears to be crucial for the induction of parasite-binding and inhibitory antibodies. In the case of a circumsporozoite protein (CSP) derived synthetic peptide we showed that this is achieved by conformational restriction of the peptide. In a phase Ia clinical trial of the virosomally formulated apical membrane antigen(AMA-1) -derived 49 amino acid long phosphatidylethanolamin peptide conjugate AMA49-CPE, we measured both vaccine induced humoral and cellular responses. In 50% of the volunteers a peripheral blood mononuclear cells (PBMC) lymphoproliferative response specific for AMA49-CPE was elicited. Moreover, all volunteers who developed high titers of parasite cross-reactive IgG in immunofluorescence assay (IFA) and Western blot analysis, were positive in the antisynthetic peptide lymphoproliferation. No interference was found between the magnitude of the pre-existing influenza specific T cell response and the vaccinationinduced AMA49-CPE specific humoral and cellular immune response. We demonstrate that the virosomal antigen delivery platform combined with surfaceanchored synthetic peptides is suitable to elicit specific human CD4 T cell responses. Mycobacterium ulcerans infection (Buruli ulcer) is unique among mycobacteria in that much of the pathology and local immunosuppression appears to be mediated by the production of a cytopathic toxin called mycolactone. IFN-γ ELISpot was used to compare reactivity of PBMC of Buruli ulcer patients with complex antigens such as isopentenyl pyrophosphate (IPP), purified protein derivative (PPD) and influenza antigens prior to and after surgical treatment. The results demonstrated that M. ulcerans infection-associated systemic reduction in IFN-γ response is not confined to stimulation with mycobacterial antigens. Immunosuppression reversed after surgical treatment, indicating that this T cell anergy is not the consequence of a genetic defect but rather related to M. ulcerans infection. In the affected tissue we detected mRNA coding for innate immune system markers, even though the lack of inflammatory responses is a hallmark of Buruli ulcer disease. These were distributed within theududlesion in a very focal and heterogeneous way. IL-8, IL-6, and TNF-α mRNA wasuddetectable by real time PCR in all lesions, whereas the expression of IFN-α, IL12p40,udIL-10, and IFN-γ was only found in some lesions. Correlations of the distribution ofudmRNA encoding for the activation marker CD83 and the DC subset markers CD123udand CD11c indicate that both activated plasmacytoid and myeloid DC were present inudthe lesions. These results suggest that adaptive M. ulcerans specific immuneudresponses may develop once therapeutic interventions have limited the production ofudmycolactone.