Development of two monoclonal antibodies against Plasmodium falciparum sporozoite surface protein 2 and mapping of B-cell epitopes

Abstract

The Plasmodium yoelii sporozoite surface protein 2 (PySSP2) is the target of protective cellular immunity. Cytotoxic T cells specific for the Plasmodium falciparum analog PfSSP2, also known as thrombospondin-related anonymous protein (TRAP), are induced in human volunteers immunized with irradiated sporozoites. PfSSP2 is an important candidate antigen for a multicomponent malaria vaccine. We generated and characterized three monoclonal antibodies (MAbs) specific for PfSSP2/TRAP. The MAbs PfSSP2.1 (immunoglobulin G1 [IgG1]), PfSSP2.2 (IgG2a), and PfSSP2.3 (IgM) were species specific and identified three distinct B-cell epitopes containing sequences DRYI, CHPSDGKC, and TRPHGR, respectively. PfSSP2.1 partially inhibited P. falciparum liver-stage parasite development in human hepatocyte cultures (42 and 86% in two experiments at 100 g/ml). Mice immunized with vaccinia virus expressing full-length PfSSP2 protein produced antibodies to (DRYIPYSP) 3 , and humans living in malaria-endemic areas (Indonesia and Kenya), who have lifelong exposure and partial clinical immunity to malaria, had antibodies to both (DRYIPYSP) 3 and (CHPSDGKCN) 2. Mice immunized with multiple antigen peptides MAP4 (DRYIPYSP) 3 P2P30 and MAP4 (CHPSDGKCN) 3 P2P30 in TiterMax developed antibodies to sporozoites that partially inhibited sporozoite invasion of human hepatoma cells (39 to 71% at a serum dilution of 1:50 in three different experiments). The modest inhibitory activities of the MAbs and the polyclonal antibodies to PfSSP2/TRAP epitopes do not suggest that a single-component vaccine designed to induce antibodies against PfSSP2/TRAP will be protective. Nonetheless, the MAbs directed against PfSSP2, and the peptides recognized by these MAbs, will be essential reagents in the development of PfSSP2/TRAP as a component of a multivalent P. falciparum human malaria vaccine.

Genetic diversity of Plasmodium falciparum merozoite surface protein‐1 (block 2), glutamate‐rich protein and sexual stage antigen Pfs25 from Chandigarh, North India

“…To date, numerous studies have been conducted in different geographical regions on pfs25 polymorphism in natural isolates but showed limited or no polymorphism . Furthermore, it has been reported that in the midgut of mosquito vector, the use of monoclonal antibodies against the native pfs25 completely blocks the development of P. falciparum oocysts .…”

Plasmodium falciparum: binding studies of peptide derived from the sporozoite surface protein 2 to Hep G2 cells

“…These peptide sequences are conserved; only the 3271 peptide T residue 161 shows genetic variability (11). Part of the sequence of 3279 ( 201 FLVG CHPSDGK GNLY 215 ) has been reported as being a B epitope (underlined), recognized by sera from malaria immune humans living in endemic areas (12). However, this epitope is not recognized by sera from Balb/C mice inoculated with the Pf SSP2 (vP1155) recombinant protein (12).…”

“…Part of the sequence of 3279 ( 201 FLVG CHPSDGK GNLY 215 ) has been reported as being a B epitope (underlined), recognized by sera from malaria immune humans living in endemic areas (12). However, this epitope is not recognized by sera from Balb/C mice inoculated with the Pf SSP2 (vP1155) recombinant protein (12). In contrast, mice immunized with multiple antigen peptides (MAPs) containing this epitope generate antibodies partially inhibiting in vitro sporozoite invasion of Hep G2 cells (12).…”

“…However, this epitope is not recognized by sera from Balb/C mice inoculated with the Pf SSP2 (vP1155) recombinant protein (12). In contrast, mice immunized with multiple antigen peptides (MAPs) containing this epitope generate antibodies partially inhibiting in vitro sporozoite invasion of Hep G2 cells (12). The HABP 3271 sequence ( 161 T D GIPDSIQDSLKES 175 ) has been involved in sporozoite invasion of mosquito salivary glands, as the D residue 162 mutation dramatically affects this invasion.…”

“…A substitution in an A‐domain Pf TRAP‐conserved residue ( 150 RENANQLVVILT D GIPDSIQ 169 ) or deletion of Pf SSP2/TRAP region II plus reduces the ability of the sporozoite to invade mosquito salivary glands (11). Pf SSP2/TRAP has several B epitopes in its sequence that are recognized by species‐specific monoclonal antibodies partially inhibiting P. falciparum sporozoite invasion (12). Our work was directed at the identification of Pf SSP2/TRAP adhesion sequences specifically interacting with the hepatocyte.…”