Abstract
Neglected tropical diseases (NTDs) encompass a broad spectrum of infectious diseases predominantly found in tropical and subtropical regions. The limitations of current therapies underscore the critical demand for novel antileishmanial agents. In this investigation, we designed, synthesized, and evaluated ten hybrid compounds (5, 8a-e, and 12a-d) integrating a 7-chloroquinoline scaffold with thiadiazole and thiazole moieties, assessing their in vitro efficacy against Leishmania major. These hybrids exhibited potent activity against the promastigote stage, displaying IC(50) values between 0.52 and 3.97 μM, outperforming miltefosine (IC(50) = 7.83 μM). Additionally, they demonstrated strong inhibition of the intracellular amastigote form, with IC(50) values ranging from 0.76 to 5.62 μM, compared to miltefosine's 8.07 μM. Notably, compound 5 emerged as a highly effective antileishmanial agent against both parasitic stages, while maintaining a favorable safety profile. Mechanistic studies revealed that compound 5 acts via an antifolate mechanism, selectively inhibiting key enzymes in the folate pathway: pteridine reductase 1 (PTR1) and dihydrofolate reductase-thymidylate synthase (DHFR-TS). Molecular docking and 100 ns molecular dynamics (MD) simulations demonstrated that the quinoline core occupies a hydrophobic pocket formed by residues Phe113, Leu188, Leu226, and Leu229, engaging in stable hydrophobic interactions and π-π stacking with Phe113. Furthermore, the quinoline scaffold and hydrazinecarbodithioate moiety formed hydrogen bonds with Tyr194, Gly225, and His241, reinforcing binding stability. Our findings introduce a promising new class of antileishmanial agents that disrupt the folate biosynthesis pathway, offering significant therapeutic potential for combating leishmaniasis.