CD4(+) T-cell- and gamma interferon-dependent protection against murine malaria by immunization with linear synthetic peptides from a Plasmodium yoelii 17-kilodalton hepatocyte erythrocyte protein

Abstract

Most work on protective immunity against the pre-erythrocytic stages of malaria has focused on induction of antibodies that prevent sporozoite invasion of hepatocytes, and CD8(+) T-cell responses that eliminate infected hepatocytes. We recently reported that immunization of A/J mice with an 18-amino-acid synthetic linear peptide from Plasmodium yoelii sporozoite surface protein 2 (SSP2) in TiterMax adjuvant induces sterile protection that is dependent on CD4(+) T cells and gamma interferon (IFN-gamma). We now report that immunization of inbred A/J mice and outbred CD1 mice with each of two linear synthetic peptides from the 17-kDa P. yoelii hepatocyte erythrocyte protein (HEP17) in the same adjuvant also induces protection against sporozoite challenge that is dependent on CD4(+) T cells and IFN-gamma. The SSP2 peptide and the two HEP17 peptides are recognized by B cells as well as T cells, and the protection induced by these peptides appears to be directed against the infected hepatocytes. In contrast to the peptide-induced protection, immunization of eight different strains of mice with radiation-attenuated sporozoites induces protection that is absolutely dependent on CD8(+) T cells. Data represented here demonstrate that CD4(+) T-cell-dependent protection can be induced by immunization with linear synthetic peptides. These studies therefore provide the foundation for an approach to pre-erythrocytic-stage malaria vaccine development, based on the induction of protective CD4(+) T-cell responses, which will complement efforts to induce protective antibody and CD8(+) T-cell responses.

FIG. 1

Schematic diagram of MAP4(SFPMNEESPLGFSPE)₃P2P30 and MAP4(GFSPEEMEAVASKFR)₃P2P30. Each MAP contains a central lysine core and four branched chains of three copies of a B-cell epitope (SFPMNEESPLGFSPE) or (GFSPEEMEAVASKFR) from P. yoelii HEP17 protein, conjugated to two T-helper epitopes, P2 (QYIKANSKFIGITE) and P30 (FNNFTVSFWLRVPKVSASHLE) from tetanus toxin (28).

FIG. 2

Antibody levels in sera of A/J, C57BL/6, BALB/c, and CD1 mice immunized with MAP4(SFPMNEESPLGFSPE)₃P2P30 in TiterMax. Control (cont.) sera were obtained from mice immunized with TiterMax alone. Serial dilutions of sera collected 3 weeks after the first and second immunizations and 10 days after the third immunization were analyzed by ELISA against (SFPMNEESPLGFSPE)₃ as described in Materials and Methods. Data are shown as mean ± SD of the OD readings of quadruplicate assays.

FIG. 3

Antibody isotypes in sera of A/J, C57BL/6, BALB/c, and CD1 mice immunized with MAP4(SFPMNEESPLGFSPE)₃P2P30 in TiterMax. Sera collected from mice 10 days after the third immunization (prior to challenge) were analyzed by ELISA against (SFPMNEESPLGFSPE)₃, using heavy-chain-specific goat anti-mouse immunoglobulins as detecting antibodies. Data are shown as mean ± SD of the OD readings of quadruplicate assays.

FIG. 4

Antibody isotypes in sera of A/J mice immunized with linear synthetic peptides (SFPMNEESPLGFSPE)₃ and (GFSPEEMEAVASKFR)₃ in TiterMax. Sera collected from mice 10 days after the second immunization (prior to challenge) were analyzed by ELISA against the relevant peptide, using heavy-chain-specific goat anti-mouse immunoglobulins as detecting antibodies. Data are shown as mean ± SD of the OD readings of quadruplicate assays.