Efficacy of two alternate vaccines based on Plasmodium falciparum merozoite surface protein 1 in an Aotus challenge trial

Abstract

In an attempt to produce a more defined, clinical-grade version of a vaccine based on Plasmodium falciparum merozoite surface protein 1 (MSP1), we evaluated the efficacy of two recombinant forms of MSP1 in an Aotus nancymai challenge model system. One recombinant vaccine, bvMSP1(42), based on the 42-kDa C-terminal portion of MSP1, was expressed as a secreted protein in baculovirus-infected insect cells. A highly pure baculovirus product could be reproducibly expressed and purified at yields in excess of 8 mg of pure protein per liter of culture. This protein, when tested for efficacy in the Aotus challenge model, gave significant protection, with only one of seven monkeys requiring treatment for uncontrolled parasitemia after challenge with P. falciparum. The second recombinant protein, P30P2MSP1(19), has been used in previous studies and is based on the smaller, C-terminal 19-kDa portion of MSP1 expressed in Saccharomyces cerevisiae. Substantial changes were made in its production process to optimize expression. The optimum form of this vaccine antigen (as judged by in vitro and in vivo indicators) was then evaluated, along with bvMSP1(42), for efficacy in the A. nancymai system. The new formulation of P30P3MSP1(19) performed significantly worse than bvMSP1(42) and appeared to be less efficacious than we have found in the past, with four of seven monkeys in the vaccinated group requiring treatment for uncontrolled parasitemia. With both antigens, protection was seen only when high antibody levels were obtained by formulation of the vaccines in Freund's adjuvant. Vaccine formulation in an alternate adjuvant, MF59, resulted in significantly lower antibody titers and no protection.

Figures

FIG. 1

Different forms of P30P2MSP119 produced by various fermentation conditions (standard, 5b, 6b, and 8b). The products of the four different fermentations were purified identically, and equal amounts were run in nonreducing SDS-PAGE. A, B, and C indicate the migration positions of the three known major polypeptides derived from the sequence of P30P2MSP119 (A, NISQ…; B, FIGITEVENISQ…; and C, EVENISQ… ). The band migrating above band A is a longer protein with an inconsistent starting point. The material migrating below band C is known to be misfolded P30P2MSP119 (32a). The sizes of the molecular mass standards are shown on the left in kilodaltons.

FIG. 2

Examples of competitive ELISAs performed with sera from rabbits immunized with P30P2MSP119. Sera from four rabbits immunized with P30P2MSP119 (Q-KNG allele) were mixed with various concentrations of a competitor MSP119 of either the Q-KNG allele (open circles) or the E-TSR allele (closed circles) prior to an ELISA with the Q-KNG allele as the coating antigen. Rabbits 8075 and 8077 were immunized with P30P2MSP119 produced under condition 5b, rabbit 8089 was immunized with P30P2MSP119 from condition 8b, and rabbit 8076 was immunized with P30P2MSP119 from standard conditions.

FIG. 3

Purity of the recombinant bvMSP142 protein. Coomassie blue-stained SDS-PAGE analysis of purified bvMSP142. Lane 1, molecular weight markers; lane 2, purified bvMSP142 run under nonreducing conditions; lane 3, purified bvMSP142 run under reducing conditions. Purity estimates obtained by laser scanning densitometry were 97.4 and 94.4% for lanes 2 and 3, respectively.

FIG. 4

Antibody titers of individual monkeys prior to challenge. Shown on the x axis is the immunogen-adjuvant combination for that group of monkeys (42/CFA, bvMSP142 in CFA; 19/MF59, P30P2MSP119 in MF59, etc). (A and B) ELISA titers are recorded as the inverse of the serum dilution corresponding to an optical density at 405 nm of 0.5. (C) IFA titers are recorded as the inverse of the serum dilution corresponding to a reading above the background. ELISA titers were measured against two capture antigens, bvMSP142 (A) and P30P2MSP119 (B). Open circles represent animals which required treatment for parasitemia during challenge; closed circles represent animals which controlled their parasitemia but required treatment for anemia; closed triangles indicate animals that self-cured without treatment. Significant differences determined by Student's t test between animals immunized with the same antigen but with either Freund's adjuvant or MF59 are shown.

FIG. 5

Course of the daily parasitemia in individual monkeys. Monkeys were challenged on day 0 with 104 P. falciparum FVO parasites. Parasitemia was determined by counting 2,000 red blood cells on Giemsa-stained thin smears. The broken line is for ease of reference between graphs. Also indicated are the treatment times for uncontrolled parasitemia greater than 4% (P), for hematocrits below 25% (H), and for self-curing animals (∗).