Our research has focused on Plasmodium falciparum sporozoite (SPZ) interactions with hepatic cells as a basis for development of in vitro assays predictive of or correlative with protective anti-SPZ immunity that might be used during human vaccine trials. We have established P. falciparum SPZ invasion assays using HepG2 cells and recently the highly polarizd WIF-B cell line. 1. SPZ integrin-like molecule. Rabbit anti-a5b1 integrin antibodies, and MAb's specific for each chain, recognized P. falciparum SPZ by IFA and immuno-electron microscopy, and blocked invasion. RGD peptides that bind to a5b1 integrin also blocked SPZ invasion. We are determining whether labeled RGD peptides bind to SPZ, and recognize a molecule of similar size to integrins.2. N-glycosylation has been reported not to occur in P. falciparum. Lectin studies cofirmed that P. falciparum blood stage and SPZ lack glycosylated proteins. However, lectins bound to falciparum liver stage parasites, suggesting that glycosylation may be stage regulated. P. falciparum proteins contain putative N-glycosylation sites. We have suggested that malaria vaccine antigens expressed in eukaryotic cells might become abnormally N- glycosylated and affect antigen processing and presentation, with implications for vaccine immunogenicity.3. We previously isolated two P. falciparum gene fragments encoding novel repetitive sequences, and immunized rabbits with synthetic peptides representing these repeats. Antibodies to repeat A reacted with the surface of P. falciparum SPZ and hepatic parasites, and blocked invasion of HepG2 cells; antibodies to repeat B reacted with SPZ surface and micronemes, and weakly blocked invasion. We are further investigating the protein recognized by anti- repeat A antibodies. In addition, we are collaborating with Case Western Reserve University to study the relationship between CD4+ responses to the P. falciparum liver stage vaccine antigen LSA-1, malaria morbidity and HLA, in a defined population in Papua New Guinea by a newly funded NIAID grant to CWRU. Since the intrahepatic fate of malaria parasites may determine whether CD4+ or CD8+ responses are elicited, a new NIH grant application proposes to study the cell biology of this parasite, in collaboration with CWRU and Dr. Lippincott-Schwartz, Natl Inst Child Hlth Human Develop, NIH.
