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

Abstract

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.

Fig. 1

Polymorphic amino acids shown on the AMA-1 crystal structure. Polymorphisms are based on sequence data from P. falciparum infections acquired at a vaccine testing site in Mali, West Africa. (A) Polymorphic residues are numbered and highlighted. Yellow and blue residues are dimorphic, orange residues are trimorphic, and red residues have four to six possible amino acids. Residues highlighted in green and blue make up the hydrophobic pocket hypothesized to be a binding site between AMA-1 and the rest of the erythrocyte invasion machinery, with blue indicating polymorphic residues within the pocket (11, 12). (B) Conserved residues in AMA-1 domains I, II, and III are highlighted in light pink, light blue, and light orange, respectively. Polymorphic residues in domain I are highlighted in dark brown (c1), red (c1 and c1L), purple (c2), dark pink (c3), and light brown (not incorporated in a cluster). Polymorphic residues in domains II and III are highlighted in dark blue and dark orange, respectively. Light gray residues are not part of any of the three major domains, and dark gray residues are polymorphisms within the interdomain region.

Fig. 2

Prevalence of AMA-1 domain I c1L haplotypes in Bandiagara, Mali. Haplotypes based on all polymorphic sites in the AMA-1 ectodomain are shown on the left, and putative serotypes defined by the eight polymorphic sites in domain I c1L (196, 197, 199, 200, 201, 204, 206, and 207) are shown on the right. The c1L haplotypes are indicated by the one-letter amino acid abbreviation at each of the eight polymorphic sites, with haplotypes corresponding to specific strains of P. falciparum indicated in parentheses. The areas of the circles correspond to haplotype prevalence. Abbreviations for the amino acid residues are as follows: D, Asp; E, Glu; F, Phe; G, Gly; H, His; I, Ile; K, Lys; L, Leu; M, Met; N, Asn; P, Pro; Q, Gln; R, Arg; S, Ser; and Y, Tyr.

Fig. 3

Prediction of clinical symptoms by amino acid changes at individual polymorphic sites in AMA-1. Plot of variable importance where the mean decrease in the accuracy of prediction of the outcome (a measure of variable importance) is plotted on the x axis and predictive variables are listed on the y axis. Change at each polymorphic site is represented by the amino acid position number. The variable age group represents the age group of the study participant. The random forest comprised 1 × 10⁵ individual tree-based statistical models, with eight variables tried at each split. The overall error rate in predicting the outcome was 37.4%.

Fig. 4

Global distribution of AMA-1 domain I c1L haplotypes. Prevalence of the 10 most prevalent domain I c1L haplotypes in a data set of 1121 AMA-1 sequences, including AMA-1 sequences available in GenBank. Haplotypes are indicated by the one-letter amino acid abbreviation at each of the eight polymorphic sites in c1L: 196, 197, 199, 200, 201, 204, 206, and 207. Haplotypes corresponding to specific strains of P. falciparum are indicated in parentheses. Pie charts are shown only for regions with 50 or more available sequences. (A) South America, n = 58,7 haplotypes. (B) Mali, n = 570, 27 haplotypes. (C) Nigeria, n = 51, 16 haplotypes. (D) India, n = 99, 26 haplotypes. (E) Thailand, n = 71, 9 haplotypes. (F) Papua New Guinea, n = 184, 16 haplotypes.