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Abstract:
Severe malaria is a life threatening disease caused by Plasmodium falciparum. Glycosylphosphatidylinositol (GPI) glycolipids specific to P. falciparum are abundant on the parasite surface and are implicated as a toxin in severe malaria. Glycoconjugate vaccines that induce GPI specific antibodies are predicted to ameliorate the hyper-inflammatory response against the native GPI toxin and protect against severe malaria. Here, I disclose the immunogenic properties of designed synthetic GPI glycoconjugates. Binding specificities of polyclonal and monoclonal antibodies induced by various synthetic GPI glycoconjugates were profiled using microarrays displaying synthetic GPI epitopes. This profiling gave detailed insight about how residues of the GPI core backbone affect the specificity of the induced antibodies. Based on the derived antibody specificity profile, synthetic GPI antigens PEtN Man3 and PEtN Man4 were designed to induce antibodies recognizing the non reducing terminus of the core GPI backbone. I reasoned that antibodies against this terminal epitope, specific to P. falciparum GPI, may protect against the toxic hyper inflammatory response implicated in severe malaria. Since GPIs are inflammatory only in the context of the full glycolipid, with the reducing end masked by the lipid tail, binding to the terminal non-reducing epitope may be enough to stop inflammation. Also, these antibodies would share epitope specificities with naturally occurring GPI specific antibodies. As predicted, immunisation with both glycoconjugates reproducibly induced high antibody titres recognising the targeted terminal epitope. Monoclonal antibodies against the target epitope were also generated after immunisation. Though the current glycoconjugate formulation did not protect against neurological symptoms in the mouse model of experimental cerebral malaria, it may be protective in other malaria models if a Th2 bias is needed. Furthermore, different glycoconjugate formulations can also be investigated to balance Th1/Th2 responses. Serendipitously, I discovered that the antibodies were cross reactive with L. mexicana, so possible protective effects of immunisation against other protozoan diseases may be explored. This thesis demonstrates that reliable and effective strategies for glycoconjugate vaccine design are strengthened by glycan microarray profiling. Designed antigens can be further formulated into glycoconjugates to produce poly- and monoclonal antibodies specific to targeted epitopes.
