Bactericidal antibody responses induced by meningococcal recombinant chimeric factor H-binding protein vaccines

Abstract

Factor H-binding protein (fHbp) is a novel meningococcal vaccine candidate that elicits serum antibodies that activate classical complement pathway bacteriolysis and also inhibit binding of the complement down-regulatory protein, factor H, to the bacterial surface. One limitation of fHbp as a vaccine candidate is antigenic variability, since antibodies to fHbp in the variant 1 (v.1) antigenic group do not protect against strains expressing v.2 or v.3 proteins, and vice versa. We have identified amino acid residues of epitopes recognized by bactericidal anti-fHbp monoclonal antibodies prepared against fHbp from each of the variant groups. One epitope expressed by nearly all v.1 proteins mapped to the B domain, while epitopes expressed by fHbp v.2 or v.3 mapped to the C domain. The results provided the rationale for engineering chimeric fHbp molecules containing the A domain (which is conserved across all variant groups), a portion of the B domain of a v.1 protein, and the carboxyl-terminal portion of the B domain and the C domain of a v.2 protein. By enzyme-linked immunosorbent assay, the resulting recombinant chimeric proteins expressed epitopes from all three variant groups. In mice, the chimeric vaccines elicited serum antibodies with bactericidal activity against a panel of genetically diverse strains expressing fHbp v.1, v.2, or v.3. The data demonstrate the feasibility of preparing a meningococcal vaccine from a single recombinant protein that elicits broad bactericidal activity, including group B strains, which account for 50 percent of cases of meningococcal disease and for which there currently is no broadly protective vaccine.

FIG. 1.

Schematic model of fHbp chimera I. The A domain (not shown) and N-terminal portion of the B domain (right, dark gray) are encoded by the fHbp gene from strain MC58 and express the v.1 epitopes recognized by anti-fHbp MAbs, JAR 3 and JAR 5. The carboxy-terminal portion of the B domain and the C domain (left, light gray) are encoded by the gene from strain 8047 and contain epitopes recognized by JAR 10, 11, 13, and 36 (v.2 and v.3). The junction point between the two portions of the chimera is residue G136, which immediately precedes the alpha-helix. Chimera II differs from chimera I in having the A174K substitution, which eliminated the JAR 11 epitope and introduced the JAR 32 and JAR 35 epitopes. The coordinates were from the nuclear magnetic resonance solution structure of the combined B and C domains of fHbp v.1 (7). The amino terminus (residue 109) and carboxyl terminus (residue 255) are labeled N and C, respectively.

FIG. 2.

Concentration-dependent binding of anti-fHbp MAbs to epitopes on wild-type and chimeric fHbps. The chimera I and II vaccines are identical except for a single amino acid substitution in chimera II (A174K). (A) Binding of MAbs prepared against fHbp v.1. JAR 1 and 5 are specific for v.1 proteins, while JAR 4 cross-reacts with v.1, v.2, and, to a lesser extent, v.3 (44). (B) Binding of MAbs prepared against fHbp v.2. JAR 10 cross-reacts with fHbp v.1, v.2, and v.3 (4), and JAR 11 and 13 cross-react with v.2 and v.3. (C) Binding of MAbs prepared against fHbp v.3. JAR 32 and 36 cross-react with v.2 and v.3.

FIG. 3.

IgG anti-fHbp antibody responses (1/GMT ± 2 SE) of mice (n = 5 per group), measured by ELISA using rfHbp v.1 (gene from strain MC58, top panel) or fHbp v.2 (gene from strain 8047, bottom panel) as the antigen on the plates. Mice immunized with wild-type rfHbp v.1 had a higher anti-rfHbp v.1 reciprocal GMT than mice immunized with fHbp v.2 (P = 0.097). Mice immunized with rfHbp v.2 had a higher anti-fHbp v.2 reciprocal GMT than mice immunized with fHbp v.1 (P = 0.005). There were no significant differences between the respective anti-v.1 antibody responses of mice given the chimera I or II vaccine and those of control mice immunized with the wild-type v.1 protein, and there were no significant differences between the respective anti-v.2 antibody responses of mice given the chimera I or II vaccine and those of control mice immunized with the wild-type v.2 protein.

FIG. 4.

Serum bactericidal antibody responses (1/GMT ± 2 SE) of mice (n = 5 per group) immunized with chimeric rfHbp vaccines as measured against strains expressing fHbp in the v.1 antigenic group. Strain H44/76 expresses fHbp v.1 with an amino acid sequence identical to that of the fHbp v.1 control vaccine. The remaining strains express subvariants of fHbp v.1 (Table 2).

FIG. 5.

Serum bactericidal antibody responses (1/GMT ± 2 SE) of mice (n = 5 per group) immunized with rfHbp vaccines against strains expressing fHbp in the v.2 or v.3 antigenic group. Strain 8047 expresses fHbp v.2 identical to that of control rfHbp v.2 vaccine. The remaining strains express subvariants of fHbp v.2 or v.3 (Table 2). The data are grouped by JAR 11 (left panel) or JAR 32 (right panel) reactivity. The chimera I and II vaccines are identical except that chimera I is JAR 11 positive and JAR 32 negative, whereas chimera II is JAR 11 negative and JAR 32 positive (see text).