Safety and Immunogenicity of a Recombinant Nonglycosylated Erythrocyte Binding Antigen 175 Region II Malaria Vaccine in Healthy Adults Living in an Area Where Malaria Is Not Endemic

Erythrocyte binding antigen region II (EBA-175) is a conserved antigen of Plasmodium falciparum that is involved in binding of the parasite to the host''s erythrocytes. We evaluated the safety and immunogenicity of a recombinant EBA-175 vaccine with aluminum phosphate adjuvant in healthy young adults living in the United States. Eighteen subjects/group received ascending doses (5, 20, 80, or 160 μg) of the vaccine at 0, 1, and 6 months; 8 subjects received placebo. Most of the injection site and systemic reactions were mild to moderate in intensity. After 2 or 3 doses of the vaccine at any concentration, antibody levels measured by enzyme-linked immunosorbent assay were significantly higher than those for the placebo group. Sera from subjects who received 3 doses of the vaccine at any concentration inhibited the growth of erythrocyte-stage P. falciparum at low levels compared to sera from placebo recipients or preimmune sera. In conclusion, the EBA-175 vaccine with adjuvant was safe and immunogenic in malaria-naïve subjects.The morbidity and mortality associated with malaria continue to exact a heavy price on many developing nations. The World Health Organization (WHO) estimates that 3.3 billion individuals live in areas where malaria is endemic, with 247 million cases and a million deaths reported for the year 2006 (41). In 1998, the Roll Back Malaria Partnership was launched to coordinate international malaria containment efforts and included the wide dissemination of long-lasting insecticidal nets, artemisinin-based combination therapy, indoor residual spraying of insecticide, and intermittent preventive treatment in pregnancy. In 2006, the success of implementing these interventions was variable, with some countries reporting a 50% decrease in the number of malaria cases and other countries reporting stable levels of transmission (5, 8, 13, 41).Some pieces missing from the available armamentarium to fight malaria are safe and effective vaccines. Plasmodium falciparum is the species associated with the greatest morbidity and mortality. The approach to the development of a P. falciparum vaccine has paralleled the multistage nature of P. falciparum infections: preerythrocytic vaccines target the sporozoite or its antigens to achieve protection from infection; erythrocytic vaccines target the merozoite antigens to achieve protection from severe disease; and transmission-blocking vaccines utilize the gametocyte, zygote, or ookinete antigens to prevent sporogenic development in the vector (4, 11, 15, 16, 34, 35, 40). A multicomponent or multistage vaccine has also been proposed as a solution to the heterogeneity of the host''s immune response and the parasite antigens.One antigen of interest in the development of P. falciparum erythrocytic vaccines is the erythrocyte binding antigen 175 (EBA-175). EBA-175 was the first P. falciparum ligand identified to have a role in high-affinity binding of the merozoite to the host''s red blood cells (RBCs) (1, 7). EBA-175 binds to the RBC glycophorin A sialic acid residues, and the interaction constitutes a major invasion pathway. The cysteine-rich second region of EBA-175 (EBA-175-RII) is the erythrocyte binding domain of the protein and is highly conserved among laboratory and clinical P. falciparum strains (19, 37). Antibodies against EBA-175-RII have been detected among individuals living in areas where malaria is endemic, with an increasing prevalence with age and a trend of disease attenuation at high antibody levels (27, 28). Antibodies directed against EBA-175-RII are capable of blocking the merozoite invasion of RBCs (39). Interestingly, anti-EBA-175-RII antibodies not only were capable of blocking the sialic acid-dependent invasion pathways but also inhibited alternative, sialic acid-independent pathways (24, 30).Taken together, the aforementioned data suggest that EBA-175-RII is a plausible vaccine candidate, especially when incorporated into a multicomponent vaccine to overcome allelic heterogeneity and/or the use of sialic acid-independent pathways by the parasite (2, 3, 27, 29). Sim and colleagues evaluated the immunogenicity of a DNA vaccine that expressed EBA-175-RII in mice, rabbits, and monkeys and found the vaccine to be safe and immunogenic (38). The antibodies produced were shown to inhibit the merozoite invasion of RBCs, and one of three vaccinated Aotus monkeys was protected from fulminant disease upon challenge (38). Jones and colleagues evaluated the immunogenicity and efficacy of 4 doses of a DNA vaccine expressing EBA-175-RII, a recombinant EBA-175-RII vaccine with adjuvant, or a combination of DNA and EBA-175-RII protein vaccines (17). Vaccinated monkeys had lower levels of parasitemia on challenge with parasitized RBCs than did control monkeys, and 3 of 4 monkeys receiving the DNA-protein combination had mild symptoms that did not require treatment (17). The promising preclinical data have prompted the development of a recombinant vaccine for use in human clinical trials.The expression of nonglycosylated EBA-175-RII (EBA-175-RII NG; referred to as EBA-175 in the rest of the text) was initially performed in a baculovirus expression system (when expressed by P. falciparum, the protein is nonglycosylated); however, the system''s productivity was low. Expression of codon-optimized EBA-175 in the yeast Pichia pastoris resulted in high-yield production of properly folded protein (42). Preclinical studies of the recombinant vaccine in mice have shown the vaccine to have an acceptable safety profile and an immunogenicity that is enhanced by the use of adjuvants. The expectation that the vaccine may be stored at a wide range of temperatures in developing countries necessitated a choice of vaccine excipient and adjuvant that allow maximum antigen stability. A study performed by Peek and colleagues demonstrated improved antigen stability for an aluminum phosphate adjuvant (Adju-Phos) and sucrose coformulation (31).We performed a phase I clinical trial to evaluate the safety and immunogenicity of escalating doses of recombinant EBA-175 with Adju-Phos in healthy young adults residing in a region where malaria is not endemic.(The data in this report were presented in part at the Vaccine Congress Meeting, Boston, MA, 6 to 9 December, 2008.)