ABSTRACT & COMMENTARY
Synopsis: The results presented here represent a considerable advance in the development of malaria vaccines, with adjuvants playing an important role as immune potentiators in inducing protection and high levels of circumsporozoite antibodies.
Source: Stoute JA, et al. A preliminary evaluation of a recombinant circumsporozoite protein vaccine against Plasmodium falciparum malaria. N Engl J Med 1997;336:86-91.
As the global malaria threat continues to expand into new areas and kill more people than it did a few decades ago, the search for an effective malaria vaccine continues. Existing vaccine candidates are poorly immunogenic and ineffective in preventing infection. A recent report from The New England Journal of Medicine presents data from the Walter Reed Army Institute of Research regarding a vaccine that may actually work due to the successful use of strong adjuvants in combination with the circumsporozoite protein.
In this study, a vaccine was created by hybridizing the circumsporozoite protein, which is the main surface antigen of sporozoites, with hepatitis B protein. The vaccine was used in combination with various potent adjuvants in order to generate adequate recognition by the immune system. Three different formulations of the vaccine, called RTS,S, were studied in an open-label trial with no placebo control. The authors assessed seroconversion and vaccine efficacy against a malaria challenge. Forty-six subjects, aged 18 to 45 years, who had not been exposed to malaria agreed to participate. Subjects were excluded if they had undergone splenectomy; had any cardiovascular, hepatic, or renal abnormalities; were allergic to any antimalarial drugs; were immunodeficient or pregnant; or had conditions that would increase the risk of an adverse outcome from malaria.
Vaccine 1 consisted of RTS,S with alum and monophosphoryl lipid A; vaccine 2 contained RTS,S in an oil-in-water emulsion; and vaccine 3 contained RTS,S with the oil-in-water emulsion, and two immune stimulantsmonophosphoryl lipid A and QS21. Each dose delivered 50 mcg of RTS,S antigen. Vaccines were administered in a three-dose series intramuscularly in the deltoid region on days 0, 4, and at approximately 28 weeks. Subjects were randomly assigned to receive Vaccine 1, 2, or 3. All 46 subjects enrolled received at least one immunization dose. Approximately one-third of subjects were assigned to receive vaccine 1, 2, or 3, respectively. Outcome was determined by serologies obtained on days 1, 2, 7, and 14 post-vaccination measured by both ELISA and IFA. Seroconversion was considered to have occurred if post-immunization antibody titers against circumsporozoite tandem repeat epitopes exceeded the mean baseline values plus 2 SD. Antibodies to hepatitis B surface antigen were also measured. Some subjects were positive for hepatitis B surface antigen on entry into the study.
Only 27 subjects received all three vaccine doses. Of these, 22 agreed to malaria challenge, with exposure to mosquitoes confirmed to contain viable P. falciparum sporozoites. This same exposure produced infection in 100% of unimmunized control subjects.
All formulations appear to be safe. No clinically relevant abnormal laboratory values were detected after administration of any dose. All initial doses were well-tolerated, causing mild discomfort at the injection site. The second doses of vaccines 2 and 3 produced more reactions such as headache, pain, malaise, feverishness, and myalgias within 24 hours after receiving the second dose. Based upon this, only one-fifth of the third dose was given to remaining subjects receiving vaccine 2 or 3. All third doses were well-tolerated.
Notably, all subjects who received two or more doses developed antibodies against the circumsporozoite protein tandem repeat epitopes. Levels peaked after the second dose, declined between second and third doses, then returned to maximum levels after the third dose. Responses to vaccines 2 and 3 were significantly greater than those to vaccine 1 (P < 0.02). ELISA and IFA antibody titers correlated well with each other on the day of challenge. Subjects who were positive for hepatitis B surface antigen before immunization had nearly maximal responses after the first dose.
Vaccine efficacy was evaluable in 22 subjects who received three doses of vaccine and who agreed to challenge with P. falciparum sporozoites. Eight subjects were given vaccine 1. Seven each received vaccines 2 and 3. All six unimmunized control subjects developed parasitemia 11-13 days after sporozoite challenge. Remarkably, only one of seven subjects given vaccine 3 became infected, for an estimated vaccine efficacy of 86%. (Relative risks of infection, 0.14; 95% confidence intervals 0.2-0.88; P < 0.005). Seven of eight subjects given vaccine 1, and five of seven subjects given vaccine 2 became infected, although length of time to the onset of malaria was significantly longer among those given vaccine 2 than among controls (P < 0.01 by log rank test). Subjects who were asymptomatic and parasite-free for 60 days after challenge remained so more than six months later. Protected subjects tended to have higher antibody titers against tandem repeat epitopes than those who developed malaria. The only subject given vaccine 3 who became infected had a poor antibody response.
COMMENT BY MARIA D. MILENO, MD
The bite of an infected anopheles mosquito introduces sporozoites into the bloodstream which are then carried to the liver. After invading hepatocytes, sporozoites develop into merozoites capable of infecting erythrocytes. The immune response to malaria is not entirely known. Plasmodia are superbly adapted to humans, with natural exposure rarely producing complete immunity to malaria. The potential of using the irradiated parasites to develop an effective malaria vaccine has been recognized since 1941, when Mulligan et al demonstrated protective immunity in chickens immunized with P. gallinaceum sporozoites.1,2 In the 1970s, Clyde and Reissman showed that immunization with irradiated sporozoites protect humans against malaria.3,4 While it is the only strategy that predictably protected humans, it is a cumbersome process, which requires repeated exposure to the bites of hundreds of irradiated mosquitoes over a period of 6-10 months. Findings from this work have substantiated the role of the circumsporozoite protein in the development of immunity to malaria. The circumsporozoite protein contains epitopes that react with antibodies that inhibit invasion of sporozoites into hepatocytes and induce cellular responses that kill sporozoite infected liver cells.5 Used alone as a vaccine in clinical trials, the circumsporozoite protein is poorly immunogenic.6
There are three distinct approaches to vaccine development. The developmental stages of malaria parasites provide multiple targets for effective immune responses. Vaccine targets include: 1) The sporozoites { intrahepatic forms} ; 2) asexual stages { erythrocytic phase} ; or 3) gametocytes { later stages} . Only the first two approaches would induce immune protection that inhibits parasites in vaccine recipients. The third approach would block transmission of gametocytes to mosquitoes, indirectly protecting humans. So far, only a single vaccine, Spf 66, aimed at the erythrocytic phase of the disease, has been reported to be efficacious.7 Recent field trials in malaria-endemic areas could not confirm the efficacy of this vaccine.8
In preclinical trials, vaccine 3 proved superior for inducing strong antibody responses and strong antigen-specific delayed hypersensitivity in primates and proliferative and cytolytic T-cell responses in mice. The present study demonstrates that strong adjuvants are required, yet in comparing efficacy of vaccine 3 to vaccine 2, it appears that strong antibody responses to tandem repeat epitopes alone were insufficient to confer protection. The authors suggested that effective adjuvants such as those used in vaccine 3 may provide signals required to upregulate co-stimulatory molecules on antigen-presenting cells, induce expression of molecules that permit these cells to travel to target tissues, or induce production of cytokines that mediate protection. Enhanced local release of interferon gamma was proposed as a mechanism of T cell recruitment. In summary, the fact that antisporozoite titers tended to be highest in the group that had the largest proportion of protected recipients suggests that humoral immunity has an important protective role. It is likely that T-cell mediated mechanisms also contribute to this protection.9
Interestingly, hepatitis B surface antigen responses were clearly affected by pre-existing hepatitis B immunity, which did not occur in prior studies.10 Due to small sample size, it is difficult to determine whether pre-existing hepatitis B immunity affected antisporozoite responses, although it is surmised that this may help boost responsiveness in a population that is highly endemic for hepatitis B, where P. falciparum often occurs.11
A reliable, highly protective malaria vaccine would be an optimal and cost-effective means of controlling the worldwide malaria resurgence. The results presented here represent a considerable advance in the development of malaria vaccines, with adjuvants playing an important role as immune potentiators in inducing protection as well as high levels of circumsporozoite antibodies. While malaria vaccine work has experienced its ups and downs, these findings offer hope and warrant further field testing.
References
1. Mulligan HW, Russel P, Mohan BN. Active immunization of fowls against Plasmodium gallinaceum by injections of killed homologous sporozoites. J Malar Inst India 1941;4:25-34.
2. Scheller LF, Azad AF. Maintenance of protective immunity against malaria by persistent hepatic parasites derived from irradiated sporozoites. Proc Natl Acad Sci USA 1995;92:4066-4068.
3. Clyde DF, et al. Immunization of a man against falciparum and vivax malaria by use of attenuated sporozoites. Am J Trop Med Hyg 1975;24:397-401.
4. Rieckmann KH, et al. Use of attenuated sporozoites in the immunization of human volunteers against falciparum malaria. Bull WHO 1979;57:261-265.
5. Ballou WR, et al. Immunogenicity of synthetic peptides from circumsporozoite protein of Plasmodium falciparum. Science 1985;228:996-999.
6. Hoffman SL, et al. Preerythrocytic malaria vaccine development. Molecular immunological considerations. In: Good MF, SaulAJ, eds. Malaria Vaccine Development Boca Raton, FL: CRC Press; 1993:149-167.
7. Patarroyo ME, et al. A synthetic vaccine protects humans against challenge with asexual blood stages of Plasmodium falciparum malaria. Nature 1988;332: 158-161.
8. Nosten F, et al. Randomized double blind placebo controlled trial of Spf66 malaria vaccine in children in Northwestern Thialand. Lancet 1996;348:701-707.
9. DiRosa F, Marzinger P. Long-lasting CD8 T cell memory in the absence of CD4 T cells or B cells. J Exp Med 1996;183:2153-2163.
10. Vreden SGS, et al. Phase I clinical trial of a recombinant malaria vaccine consisting of the Circumsporozoite repeat region of Plasmodium falciparum coupled to Hepatitis B surface antigen. Am J Trop Med Hyg 1991;45:533-538.
11. Gordon DM, et al. Safety, immunogenicity, and efficacy of recombinantly produced Plasmodium falciparum circumsporozoite protein-hepatitis B surface antigen subunit vaccine. J Infect Dis 1995;171:1576-1585.
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