Evidence of Pyrimethamine and Cycloguanil Analogues as Dual Inhibitors of Trypanosoma brucei Pteridine Reductase and Dihydrofolate Reductase

Abstract

Trypanosoma and Leishmania parasites are the etiological agents of various threatening neglected tropical diseases (NTDs), including human African trypanosomiasis (HAT), Chagas disease, and various types of leishmaniasis. Recently, meaningful progresses in the treatment of HAT, due to Trypanosoma brucei (Tb), have been achieved by the introduction of fexinidazole and the combination therapy eflornithine–nifurtimox. Nevertheless, due to drug resistance issues and the exitance of animal reservoirs, the development of new NTD treatments is still required. For this purpose, we explored the combined targeting of two key folate enzymes, dihydrofolate reductase (DHFR) and pteridine reductase 1 (PTR1). We formerly showed that the TbDHFR inhibitor cycloguanil (CYC) also targets TbPTR1, although with reduced affinity. Here, we explored a small library of CYC analogues to understand how their substitution pattern affects the inhibition of both TbPTR1 and TbDHFR. Some novel structural features responsible for an improved, but preferential, ability of CYC analogues to target TbPTR1 were disclosed. Furthermore, we showed that the known drug pyrimethamine (PYR) effectively targets both enzymes, also unveiling its binding mode to TbPTR1. The structural comparison between PYR and CYC binding modes to TbPTR1 and TbDHFR provided key insights for the future design of dual inhibitors for HAT therapy.

Preliminary Structure–Activity Relationship Study of the MMV Pathogen Box Compound MMV675968 (2,4-Diaminoquinazoline) Unveils Novel Inhibitors of Trypanosoma brucei brucei

“…Although lower activity was demonstrated against T. b. brucei crude protein extract, the results nevertheless portrayed a consistent inhibition of the oxidation of the NADPH cofactor by the parent hit (MMV675968) and the two promising analogs (MMV1578467 and MMV1578445) ( Figure 7 ). The effective but low activity compared to previously reported data [ 28 , 47 ] could be tentative justified by the fact that a crude protein extract was used in the assays rather than the purified DHFR enzyme. Future in vitro/in vivo studies using recombinantly purified DHFR enzyme and appropriate animal models of trypanosomiasis will enable us to confirm our assertions.…”

“…Post docking analyses revealed that, in DHFR, all the accessible protonation states of both compounds shared similar poses within the same ligand recognition site in the binding pocket. This resulted in hydrogen bond formation between the diaminopyrimidine moiety and key DHFR ligand recognition residues [ 26 , 27 , 28 ], including the highly conserved Val32 ( Figure 3 A,B). Unlike DHFR, the TR enzyme consists of a wide binding site, where substrates and inhibitors have been shown to adopt different conformations with stacking observed for some inhibitor binding poses [ 29 , 30 ].…”

Current Treatments to Control African Trypanosomiasis and One Health Perspective

“…These periods are potentiated by unrestricted large-scale use of the drug, inaccurate dosing regimens, and use of drugs that are slowly eliminated from the body. Drug-specific selection mechanisms normally follow the primary low impact generic resistance mechanisms associated with increased stress responses [ 87 ]. When drug-specific resistance events appear, the increase in the drug concentration will only lead to a greater selective pressure resulting in higher levels of resistance.…”

Rudimentary Structure-Activity-Relationship study of the MMV Pathogen Box compound MMV675968 (2,4-diaminoquinazoline) unveils novel inhibitors of Trypanosoma brucei brucei DHFR enzyme

“…Post docking analyses revealed that, in DHFR, all the accessible protonation states of both compounds shared similar poses within the same ligand recognition site in the binding pocket. This resulted in hydrogen bond formation between the diaminopyrimidine moiety and key DHFR ligand recognition residues [35][36][37], including the highly conserved Asp54, and Val32 (Figs 3A and 3B). Unlike DHFR, the TR enzyme consists of a wide binding site, where substrates and inhibitors have been shown to adopt different conformations with stacking observed for some inhibitor binding poses [38,39].…”