TcGAPDH peptidic inhibitors derived from dinoponeratoxin M-PONTX-Dq4e with trypanocidal effect

Glyceraldehyde-3-phosphate dehydrogenase enzyme (TcGAPDH) is a key target involved on Trypanosoma cruzi, etiological agent of Chagas' disease (CD), survivor. So, novel therapeutical alternatives are necessary due to low efficacy and toxicity of available drug, benznidazole (Bz). Previously, trypanocidal effect of dinoponeratoxins, antimicrobial peptides from Dinoponera quadriceps, was assessed. Among them, M-PONTX-Dq4e peptide (30 aa) presented as promisor, more effective than Bz. However, it showed cytotoxicity to host cells and biotechnological limitations due to its length. So, molecular modifications are necessary to explore its technological potential and obtain innovative compounds. Thus, this study aims to obtain novel M-PONTX-Dq4e-derived peptides with trypanocidal effect, less cytotoxic, and short length. Initially, M-PONTX-Dq4e fragments, [4-30], [1-22], [11-20], [21-30] and [1-10], were obtained by solid-phase synthesis. Cytotoxicity on host cells (LLC-MK2), and trypanocidal effect (T. cruzi Y strain) were evaluated, being M-PONTX-Dq4e [1-10] the most promisor. M-PONTX-Dq4e [1-10] analogues [R³,R⁸,R¹⁰], [H]⁴, [R]⁴, [F]⁹, and [Q]⁴ were synthesized, and outperformed in terms of anti-T. cruzi and cytotoxicity the M-PONTX-Dq4e [1-10] fragment. [F]⁹ M-PONTX-Dq4e [1-10] was more selective for amastigote (SI > 152.1) than for trypomastigote (SI > 53.0). Analogues induced membrane damage, ROS accumulation, and mitochondrial dysfunction. TcGAPDH involvement on analogues-induced cell death was investigated through in silico and in vitro analysis. Peptides coupled with catalytic site residues, and inhibited TcGAPDH activity. So, a rational design strategy was used to develop M-PONTX-Dq4e-derived trypanocidal compounds with higher efficacy than Bz, and biotechnological value, highlighting [F]⁹ M-PONTX-Dq4e [1-10], more effective against amastigote form, responsible to DC chronic phase maintenance.