Redefining an epitope of a malaria vaccine candidate, with antibodies against the N-terminal MSA-2 antigen of Plasmodium harboring non-natural peptide bonds

Abstract

The aim of obtaining novel vaccine candidates against malaria and other transmissible diseases can be partly based on selecting non-polymorphic peptides from relevant antigens of pathogens, which have to be then precisely modified for inducing a protective immunity against the disease. Bearing in mind the high degree of the MSA-221–40 peptide primary structure’s genetic conservation among malaria species, and its crucial role in the high RBC binding ability of Plasmodium falciparum (the main agent causing malaria), structurally defined probes based on non-natural peptide-bond isosteres were thus designed. Thus, two peptide mimetics were obtained (so-called reduced amide pseudopeptides), in which naturally made amide bonds of the 30FIN32-binding motif of MSA-2 were replaced with ψ–[CH2–NH] methylene amide isostere bonds, one between the F–I and the second between I–N amino acid pairs, respectively, coded as ψ-128 ψ-130. These peptide mimetics were used to produce poly- and monoclonal antibodies in Aotus monkeys and BALB/c mice. Parent reactive mice-derived IgM isotype cell clones were induced to Ig isotype switching to IgG sub-classes by controlled in vitro immunization experiments. These mature isotype immunoglobulins revealed a novel epitope in the MSA-225–32 antigen and two polypeptides of rodent malaria species. Also, these antibodies’ functional activity against malaria was tested by in vitro assays, demonstrating high efficacy in controlling infection and evidencing neutralizing capacity for the rodent in vivo malaria infection. The neutralizing effect of antibodies induced by site-directed designed peptide mimetics on Plasmodium’s biological development make these pseudopeptides a valuable tool for future development of immunoprophylactic strategies for controlling malarial infection.

The Next Vaccine Generation Against Malaria: Structurally Modulated Plasmodium Antigens

“…Similarly, CD profile for the L-native sequence compared with that of the Retro-inverso analogue (green line), behave as mirror images of each other bearing in mind that both were made with only l-amino acids, but having opposite peptide backbone orientation. [42][43][44][45][46][47][48][49][50][51][52][53][54][55][56][57][58][59][60][61] peptide. The native sequence is represented in capital letters for l-amino acids, D-enantiomer represented by lowercase letters purple highlighted, Retro-sequence is constituted by d-amino acids in lowercase letters blue highlighted and the so-named Retro-inverso peptidomimetic is confirmed by l-amino acids green highlighted.…”

“…The native sequence is represented in capital letters for l-amino acids, D-enantiomer represented by lowercase letters purple highlighted, Retro-sequence is constituted by d-amino acids in lowercase letters blue highlighted and the so-named Retro-inverso peptidomimetic is confirmed by l-amino acids green highlighted. (B) Circular dichroism secondary structure patterns for PfMSP1 [42][43][44][45][46][47][48][49][50][51][52][53][54][55][56][57][58][59][60][61] peptido-mimetics. Used color code was black line for the native sequence, purple for its D-enantiomer, blue line for the Retro-analogue and green line for the Retroinverso peptido-mimetic.…”

“…Used color code was black line for the native sequence, purple for its D-enantiomer, blue line for the Retro-analogue and green line for the Retroinverso peptido-mimetic. Competition ELISA was used for the PfMSP1 [42][43][44][45][46][47][48][49][50][51][52][53][54][55][56][57][58][59][60][61] peptide mapping with monoclonal antibodies coded Ig-αψ-437( 52 V-L 53 ) and Ig-αψ-439( 51 M-V 52 ) as shown in (C) and (D), respectively. IC50 μM values represent the average of data obtained in triplicate.…”

“…In order to test the unique recognition of an antibody stimulated by a reduced amide peptidomimetic in which the oxygen atom of the carbonyl group (-R-CO-NH-R′-) of a relevant peptide-bond was replaced with two hydrogen atoms to lead an analogue being the reduced form of it and herein named reduced amide peptido-mimetic ψ(-R-CH 2 -NH-R′-). Thus, two monoclonal immunoglobulins (mAb) were produced, one directed to PfMSP1 [42][43][44][45][46][47][48][49][50][51][52][53][54][55][56][57][58][59][60][61] modified between the -52 V-L 53 -and another directed to the -51 M-V 52 -amino acid pairs, respectively, which herein are named as Igα-ψ-VL 52-53 and Igα-ψ-MV 51-52 . Both mAbs possess inhibitory capacities in both in vitro and in vivo malaria infections by P. falciparum as well as P. yoelii and P. berghei as elsewhere published [42].…”

“…Both mAbs possess inhibitory capacities in both in vitro and in vivo malaria infections by P. falciparum as well as P. yoelii and P. berghei as elsewhere published [42]. Also our experiments allowed to a fine mapping of the MSP-1 Nterminus portion using monoclonal and polyclonal antibodies induced by reduced amide peptido-mimetics of the PfMSP1 [42][43][44][45][46][47][48][49][50][51][52][53][54][55][56][57][58][59][60][61] peptide and so we identified the antigen epitope whose amino acid sequence is 51 MVLNEGTSGTA 61 as elsewhere reported [44].…”