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.

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Rational strategies for obtaining malaria vaccine candidates should include not only a proper selection of target antigens for antibody stimulation, but also a versatile molecular design based on ordering the right pieces from the complex pathogen molecular puzzle towards more active and functional immunogens. Classical Plasmodium falciparum antigens regarded as vaccine candidates have been selected as model targets in this study. Among all possibilities we have chosen epitopes of PfCSP, STARP; MSA1 and Pf155/RESA from pre- and erythrocyte stages respectively for designing a large 82-residue chimeric immunogen. A number of options aimed at diminishing steric hindrance for synthetic procedures were assessed based on standard Fmoc chemistry such as building block orthogonal ligation; pseudo-proline and microwave-assisted procedures, therefore the large-chimeric target was produced, characterized and immunologically tested. Antigenicity and functional in vivo efficacy tests of the large-chimera formulations administered alone or as antigen mixtures have proven the stimulation of high antibody titers, showing strong correlation with protection and parasite clearance of vaccinated BALB/c mice after being lethally challenged with both P. berghei-ANKA and P. yoelii 17XL malaria strains. Besides, 3D structure features shown by the large-chimera encouraged as to propose using these rational designed large synthetic molecules as reliable vaccine candidate-presenting systems.