Isolation of a monoclonal antibody from a malaria patient-derived phage display library recognising the Block 2 region of Plasmodium falciparum merozoite surface protein-1

Abstract

Antigens associated with different life-cycle stages of Plasmodium falciparum (P. falciparum) are being investigated as potential malaria vaccines. However, the difficulties of obtaining immunological reagents have hampered detailed characterisation of these antigens and human antibody responses to different antigenic variants. Phage display antibodies offer an alternative method for the production of high affinity single chain variable fragment (scFv) derivatives of human antibodies of ‘natural host’ origin [1]. These cloned recombinant human B-cell derived antibody fragments, containing a single antigen binding site, are derived from the natural induction of the immune response of the host to the parasite and thus, by-pass the need for animal immunisation. Phage display scFv antibodies are assembled as antibody fragments in the periplasmic space of Escherichia coli in a reducing environment similar to that found in eukaryotic assembly pathways [2,3]. The abundant merozoite surface protein-1 (MSP-1) of P. falciparum is a major blood-stage malaria vaccine candidate. Antibodies to MSP-1 of P. falciparum are involved in the protection of animals against experimental P. falciparum infections [4,5] and MSP-1 has been successfully used as a protective immunogen in animal model vaccination trials [6–8]. Most studies have used the conserved C-terminal end of the protein (MSP-119) [9,10]. Less is known about the function and immunogenicity of the other regions of MSP-1. Block 2 (Bl2), the most variable region of this polymorphic molecule, is at the N-terminus of the protein. All MSP-1 variants can be classified into three types referred to as, K1, MAD20 and RO33 types, based on Block 2 amino acid sequence [11]. K1-type and MAD20-type Block 2 regions are characterised by the presence of internal trior hexa-peptide repeats flanked by type-specific sequences. The Block 2 region of RO33-type variants contains a non-repetitive sequence that varies little between isolates [12]. Recently, a strikingly positive correlation between protection against clinical malaria and the presence of anti-Block 2 (Bl2) antibodies has been demonstrated in a prospective study of disease resistance and susceptibility in the Gambia [13]. MAD 20 Block 2-type specific human or mouse monoclonal antibodies do not exist. To obtain such a reagent, we have isolated phage antibodies that bind a Abbre6iations: E. coli, Escherichia coli ; GST, glutathione-S-transferase; MAD20/Bl2, MAD20 Block 2; MSP-1, merozoite surface protein-1; P. falciparum, Plasmodium falciparum ; scFv, single chain variable fragment. Note : Nucleotide sequence data reported in this paper have been submitted to the DDJB, EMBL and GenBankTM databases under the accession numbers AF240360, AF240361 and AF240362. * Corresponding author. Tel.: +44-131-6508655; fax: +44-1316508655. E-mail address: d.e.arnot@ed.ac.uk (D.E. Arnot).