Antimalarial effect of cell penetrating peptides derived from the junctional region of Plasmodium falciparum dihydrofolate reductase-thymidylate synthase

Abstract

Dihydrofolate reductase-thymidylate synthase of Plasmodium falciparum (PfDHFR-TS) is an important target of antifolate antimalarial drugs. However, drug resistant parasites are widespread in malaria endemic regions. The unique bifunctional property of PfDHFR-TS could be exploited for the design of allosteric inhibitors that interfere with the active dimer conformation. In this study, peptides were derived from the junctional region (JR) of PfDHFR-TS amino acid sequence in the α_j1 helix (JR-helix) and the DHFR domain that is necessary for interaction with α_j1 helix (JR21). Five peptides were synthesized and tested for inhibition of PfDHFR-TS enzyme by Bacterial inhibition assay (BIA) based on the growth of an E. coli DHFR and TS knockout complemented with a recombinant plasmid expressing PfDHFR-TS enzyme. Significant inhibition was observed for JR21 and JR21 conjugated to cell-penetrating octa-arginine peptide (rR8-JR21) with 50 % inhibitory concentration (IC₅₀) of 3.87 and 1.53 μM, respectively. The JR-helix and rR8-JR-helix peptides were inactive. JR21 and rR8-JR21 peptides showed similar growth inhibitory effects on P. falciparum NF54 parasites cultured in vitro. Treatment with rR8-JR21 delayed parasite development, in which an accumulation of ring stage parasites was observed after 12 h of culture. Minimal red blood cell (RBC) hemolysis was observed at the highest dose of peptide tested. The most potent peptide rR8-JR21 not only compromised the development of the P. falciparum, but also inhibited the parasite growth and has low hemolytic effect on human RBCs.

Keywords: Antimalarial peptide; Cell penetrating peptides; Dihydrofolate reductase-thymidylate synthase; Junctional region; Malaria; Plasmodium falciparum.