B And T Cell Responses To Epitopes In Disulfide Bond-constrained Recombinant Pfs48/45 Protein, A Malaria Transmission-blocking Vaccine Candidate Antigen
Our overall research goal is focused on the development of a malaria transmission-blocking vaccine (TBV). The antigenic target, Pfs48/45 protein, is expressed on Plasmodium gametocytes, which are stages responsible for establishing parasite infection in the mosquito vector. The epitopes recognized by functional antibodies targeting Pfs48/45 are disulfide-bond (S-S) constrained, conformational epitopes. As Pfs48/45 protein has not been crystallized, precise location of the S-S bonds and the topology of epitopes are unknown. It has been shown previously that the ability to reduce S-S in antigens can greatly influence the epitopes presented by antigen-presenting cells (APCs) and thus influence induction of effective immune responses. Gamma-interferon-inducible lysosomal thiol reductase (GILT) is an enzyme expressed in APCs that mediates reduction of S-S bonds contained within antigens, for subsequent display of peptides on MHC molecules. Using non-reduced (NR) and reduced/alkylated (RA) Pfs48/45 antigens, we sought to investigate the role of GILT on induction of protective immunity. We hypothesized that the ability to reduce S-S bonds in Pfs48/45 will impact the generation of T cell epitopes, and thus influence helper T cell responses required for B cell stimulation and production of protective antibody. We conducted immunogenicity studies in wild type (WT) and GILT-/- (KO) mice using the two structural forms of Pfs48/45 and analyzed immune responses to full length Pfs48/45, five overlapping fragments and 39 overlapping peptides. Results indicated that generation of Pfs48/45 antibodies is not significantly impacted by the availability of GILT, however there was uniquely Th2-biased T and B cell responses in the KO mice, and a contrasting Th1 bias in WT mice. Results also revealed possible effects of GILT on induction of long-lived plasma cells and memory B cells responsible for resting and antigen-recall responses to Pfs48/45. Data presented also shows reduced immunogenicity of the RA Pfs48/45 antigen and immune responses differed in magnitude and specificity between male and female animals. Overall, we aimed to gain a better understanding of the immunological mechanisms critical to generate protective and lasting immunity against Pfs48/45. These and future studies will contribute significantly to our understanding of antigenic features of Pfs48/45 important for use as a TBV.