A Diversity-Covering Approach to Immunization with Plasmodium falciparum Apical Membrane Antigen 1 Induces Broader Allelic Recognition and Growth Inhibition Responses in Rabbits

Abstract

Plasmodium falciparum apical membrane antigen 1 (PfAMA1), a candidate malaria vaccine, is polymorphic. This polymorphism is believed to be generated predominantly under immune selection pressure and, as a result, may compromise attempts at vaccination. Alignment of 355 PfAMA1 sequences shows that around 10% of the 622 amino acid residues can vary between alleles and that linkages between polymorphic residues occur. Using this analysis, we have designed three diversity-covering (DiCo) PfAMA1 sequences that take account of these linkages and, when taken together, on average incorporate 97% of amino acid variability observed. For each of the three DiCo sequences, a synthetic gene was constructed and used to transform the methylotrophic yeast Pichia pastoris, allowing recombinant expression. All three DiCo proteins were reactive with the reduction-sensitive monoclonal antibody 4G2, suggesting the DiCo sequences had conformations similar to those of naturally occurring PfAMA1. Rabbits were immunized with FVO strain PfAMA1 or with the DiCo proteins either individually or as a mixture. Antibody titers and the ability to inhibit parasite growth in vitro were determined. Animals immunized with the DiCo mix performed similarly to animals immunized with FVO AMA1 when measured against FCR3 strain parasites but outperformed animals immunized with FVO AMA1 when assessed against other strains. The levels of growth inhibition (ϳ70%) induced by the mix of three DiCo proteins were comparable for FVO, 3D7, and HB3, suggesting that a considerable degree of diversity in AMA1 is adequately covered. This suggests that vaccines based upon the DiCo mix approach provide a broader functional immunity than immunization with a single allele.

Simulation of B Cell Affinity Maturation Explains Enhanced Antibody Cross-Reactivity Induced by the Polyvalent Malaria Vaccine AMA1

“…Researchers have explored two approaches to tackle the issue of antigenic variation in AMA1: polyvalent formulations using multiple strains of AMA1 (47,48) and chimeric variants of AMA1 (51,68), which incorporate a subset of high-frequency polymorphisms into a small set of constructs. In both cases, allelic coverage is the guiding principle, meaning that the vaccine should incorporate as broad a range of polymorphic variation as is possible (43).…”

Assessment of Immune Interference, Antagonism, and Diversion following Human Immunization with Biallelic Blood-Stage Malaria Viral-Vectored Vaccines and Controlled Malaria Infection

“…Attempts to address this issue of antigenic polymorphism have involved the development of multivalent vaccine formulations containing multiple allelic variants of the MSP1 or AMA1 target Ag (28)(29)(30) or artificial diversity covering consensus sequences (31). Similarly, both of the viral-vectored vaccine transgene inserts had been previously designed to address the issues surrounding target Ag polymorphism by encoding biallelic vaccine inserts for AMA1 (32,33) and MSP1 (34,35).…”

Molecular vaccines for malaria

“…29 The Biomedical Primate Research Center (BPRC) in The Netherlands has synthesized three "diversity covering" versions of AMA1 encompassing 97% of the amino acid variability observed in nature and conducted preliminary studies in rabbits showing the induction of crossinhibitory antibodies. 30 Similar studies are underway at the La Trobe University and at the Walter and Eliza Hall Institute of Medical Research in Australia.…”