Safety and immunogenicity of an AMA1 malaria vaccine in Malian children: results of a phase 1 randomized controlled trial.

BACKGROUND: The objective was to evaluate the safety and immunogenicity of the AMA1-based malaria vaccine FMP2.1/AS02(A) in children exposed to seasonal falciparum malaria. METHODOLOGY/PRINCIPAL FINDINGS: A Phase 1 double blind randomized controlled dose escalation trial was conducted in Bandiagara, Mali, West Africa, a rural town with intense seasonal transmission of Plasmodium falciparum malaria. The malaria vaccine FMP2.1/AS02(A) is a recombinant protein (FMP2.1) based on apical membrane antigen 1 (AMA1) from the 3D7 clone of P. falciparum, formulated in the Adjuvant System AS02(A). The comparator vaccine was a cell-culture rabies virus vaccine (RabAvert). One hundred healthy Malian children aged 1-6 years were recruited into 3 cohorts and randomized to receive either 10 microg FMP2.1 in 0.1 mL AS02(A), or 25 microg FMP2.1 in 0.25 mL AS02(A), or 50 microg FMP2.1 50 microg in 0.5 mL AS02(A), or rabies vaccine. Three doses of vaccine were given at 0, 1 and 2 months, and children were followed for 1 year. Solicited symptoms were assessed for 7 days and unsolicited symptoms for 30 days after each vaccination. Serious adverse events were assessed throughout the study. Transient local pain and swelling were common and more frequent in all malaria vaccine dosage groups than in the comparator group, but were acceptable to parents of participants. Levels of anti-AMA1 antibodies measured by ELISA increased significantly (at least 100-fold compared to baseline) in all 3 malaria vaccine groups, and remained high during the year of follow up. CONCLUSION/SIGNIFICANCE: The FMP2.1/AS02(A) vaccine had a good safety profile, was well-tolerated, and induced high and sustained antibody levels in malaria-exposed children. This malaria vaccine is being evaluated in a Phase 2 efficacy trial in children at this site. TRIAL REGISTRATION: ClinicalTrials.gov NCT00358332 [NCT00358332].

Extreme polymorphism in a vaccine antigen and risk of clinical malaria: implications for vaccine development.

Vaccines directed against the blood stages of Plasmodium falciparum malaria are intended to prevent the parasite from invading and replicating within host cells. No blood-stage malaria vaccine has shown clinical efficacy in humans. Most malaria vaccine antigens are parasite surface proteins that have evolved extensive genetic diversity, and this diversity could allow malaria parasites to escape vaccine-induced immunity. We examined the extent and within-host dynamics of genetic diversity in the blood-stage malaria vaccine antigen apical membrane antigen-1 in a longitudinal study in Mali. Two hundred and fourteen unique apical membrane antigen-1 haplotypes were identified among 506 human infections, and amino acid changes near a putative invasion machinery binding site were strongly associated with the development of clinical symptoms, suggesting that these residues may be important to consider in designing polyvalent apical membrane antigen-1 vaccines and in assessing vaccine efficacy in field trials. This extreme diversity may pose a serious obstacle to an effective polyvalent recombinant subunit apical membrane antigen-1 vaccine.

Safety and immunogenicity of an AMA-1 malaria vaccine in Malian adults: results of a phase 1 randomized controlled trial.

BACKGROUND: The objective was to evaluate the safety, reactogenicity and immunogenicity of the AMA-1-based blood-stage malaria vaccine FMP2.1/AS02A in adults exposed to seasonal malaria. METHODOLOGY/PRINCIPAL FINDINGS: A phase 1 double blind randomized controlled dose escalation trial was conducted in Bandiagara, Mali, West Africa, a rural town with intense seasonal transmission of Plasmodium falciparum malaria. The malaria vaccine FMP2.1/AS02A is a recombinant protein (FMP2.1) based on apical membrane antigen-1 (AMA-1) from the 3D7 clone of P. falciparum, adjuvanted with AS02A. The comparator vaccine was a cell-culture rabies virus vaccine (RabAvert). Sixty healthy, malaria-experienced adults aged 18-55 y were recruited into 2 cohorts and randomized to receive either a half dose or full dose of the malaria vaccine (FMP2.1 25 microg/AS02A 0.25 mL or FMP2.1 50 microg/AS02A 0.5 mL) or rabies vaccine given in 3 doses at 0, 1 and 2 mo, and were followed for 1 y. Solicited symptoms were assessed for 7 d and unsolicited symptoms for 30 d after each vaccination. Serious adverse events were assessed throughout the study. Titers of anti-AMA-1 antibodies were measured by ELISA and P. falciparum growth inhibition assays were performed on sera collected at pre- and post-vaccination time points. Transient local pain and swelling were common and more frequent in both malaria vaccine dosage groups than in the comparator group. Anti-AMA-1 antibodies increased significantly in both malaria vaccine groups, peaking at nearly 5-fold and more than 6-fold higher than baseline in the half-dose and full-dose groups, respectively. CONCLUSION/SIGNIFICANCE: The FMP2.1/AS02A vaccine had a good safety profile, was well-tolerated, and was highly immunogenic in malaria-exposed adults. This malaria vaccine is being evaluated in Phase 1 and 2 trials in children at this site.

Short report: rare Plasmodium falciparum merozoite surface protein 1 19-kda (msp-1(19)) haplotypes identified in Mali using high-throughput genotyping methods.

Genetic diversity in malaria vaccine antigens may compromise malaria vaccine efficacy, so it is important to understand this diversity and the processes that generate it. By applying new high-throughput genotyping methods to a large sample of infections from Mali (N = 1369), seven new 19-kDa merozoite surface protein 1 (MSP-1(19)) haplotypes were identified. Herein we report the sequences of these new haplotypes and discuss their possible origins. Although they are present in < 1% of the samples examined, the existence of these rare haplotypes reveals a greater degree of diversity at this locus than previously reported and highlights the potential for Plasmodium to evolve under selective pressure from the immune system and from such interventions as vaccines and drugs.

Dynamics of polymorphism in a malaria vaccine antigen at a vaccine-testing site in Mali.

BACKGROUND: Malaria vaccines based on the 19-kDa region of merozoite surface protein 1 (MSP-1(19)) derived from the 3D7 strain of Plasmodium falciparum are being tested in clinical trials in Africa. Knowledge of the distribution and natural dynamics of vaccine antigen polymorphisms in populations in which malaria vaccines will be tested will guide vaccine design and permit distinction between natural fluctuations in genetic diversity and vaccine-induced selection. METHODS AND FINDINGS: Using pyrosequencing, six single-nucleotide polymorphisms in the nucleotide sequence encoding MSP-1(19) were genotyped from 1,363 malaria infections experienced by 100 children who participated in a prospective cohort study in Mali from 1999 to 2001. The frequencies of 14 MSP-1(19) haplotypes were compared over the course of the malaria transmission season for all three years, in three age groups, and in consecutive infections within individuals. While the frequency of individual MSP-1(19) haplotypes fluctuated, haplotypes corresponding to FVO and FUP strains of P. falciparum (MSP-1(19) haplotypes QKSNGL and EKSNGL, respectively) were most prevalent during three consecutive years and in all age groups with overall prevalences of 46% (95% confidence interval [CI] 44%-49%) and 36% (95% CI 34%-39%), respectively. The 3D7 haplotype had a lower overall prevalence of 16% (95% CI 14%-18%). Multiplicity of infection based on MSP-1(19) was higher at the beginning of the transmission season and in the oldest individuals (aged > or =11 y). Three MSP-1(19) haplotypes had a reduced frequency in symptomatic infections compared to asymptomatic infections. Analyses of the dynamics of MSP-1(19) polymorphisms in consecutive infections implicate three polymorphisms (at positions 1691, 1700, and 1701) as being particularly important in determining allele specificity of anti-MSP-1(19) immunity. CONCLUSIONS: Parasites with MSP-1(19) haplotypes different from that of the leading vaccine strain were consistently the most prevalent at a vaccine trial site. If immunity elicited by an MSP-1-based vaccine is allele-specific, a vaccine based on either the FVO or FUP strain might have better initial efficacy at this site. This study, to our knowledge the largest of its kind to date, provides molecular information needed to interpret population responses to MSP-1-based vaccines and suggests that certain MSP-1(19) polymorphisms may be relevant to cross-protective immunity.

Return of chloroquine antimalarial efficacy in Malawi.

BACKGROUND: In 1993, Malawi became the first country in Africa to replace chloroquine with the combination of sulfadoxine and pyrimethamine for the treatment of malaria. At that time, the clinical efficacy of chloroquine was less than 50%. The molecular marker of chloroquine-resistant falciparum malaria subsequently declined in prevalence and was undetectable by 2001, suggesting that chloroquine might once again be effective in Malawi. METHODS: We conducted a randomized clinical trial involving 210 children with uncomplicated Plasmodium falciparum malaria in Blantyre, Malawi. The children were treated with either chloroquine or sulfadoxine\#8211;pyrimethamine and followed for 28 days to assess the antimalarial efficacy of the drug. RESULTS: In analyses conducted according to the study protocol, treatment failure occurred in 1 of 80 participants assigned to chloroquine, as compared with 71 of 87 participants assigned to sulfadoxine\#8211;pyrimethamine. The cumulative efficacy of chloroquine was 99% (95% confidence interval [CI], 93 to 100), and the efficacy of sulfadoxine\#8211;pyrimethamine was 21% (95% CI, 13 to 30). Among children treated with chloroquine, the mean time to parasite clearance was 2.6 days (95% CI, 2.5 to 2.8) and the mean time to the resolution of fever was 10.3 hours (95% CI, 8.1 to 12.6). No unexpected adverse events related to the study drugs occurred. CONCLUSIONS: Chloroquine is again an efficacious treatment for malaria, 12 years after it was withdrawn from use in Malawi. (ClinicalTrials.gov number, NCT00125489 [ClinicalTrials.gov].).

A high-throughput method for quantifying alleles and haplotypes of the malaria vaccine candidate Plasmodium falciparum merozoite surface protein-1 19 kDa.

BACKGROUND: Malaria vaccine efficacy may be compromised if the frequency of non-target alleles increases following vaccination with a genetically polymorphic target. Methods are needed to monitor genetic diversity in polymorphic vaccine antigens, but determining which genetic variants of such antigens are present in infected individuals is complicated by the frequent occurrence of mixed infections. METHODS: Pyrosequencing was used to determine allele frequencies at each of six single nucleotide polymorphisms in the Plasmodium falciparum blood-stage vaccine antigen merozoite surface protein 1 19 kDa (MSP-119) in field samples from a vaccine-testing site in Mali. Mixtures of MSP-119 clones were created to validate a haplotype-estimating algorithm that uses maximum likelihood methods to determine the most probable combination of haplotypes given the allele frequencies for an infection and the haplotypes known to be circulating in the population. RESULTS: Fourteen unique MSP-119 haplotypes were identified among 351 genotyped infections. After adjustment to a standard curve, Pyrosequencing provided accurate and precise estimates of allele frequencies in mixed infections. The haplotype-estimating algorithm provided accurate estimates of haplotypes in mixed infections containing up to three haplotypes. Based on the MSP-119 locus, approximately 90% of the 351 infections contained two or fewer haplotypes. CONCLUSION: Pyrosequencing in conjunction with a haplotype-estimating algorithm provides accurate estimates of haplotypes present in infections with up to 3 haplotypes, and can be used to monitor genetic diversity in parasite populations prior to and following introduction of MSP-1-based malaria vaccines.