Structure−Activity Relationship and Studies on the Molecular Mechanism of Leishmanicidal N,C-Coupled Arylisoquinolinium Salts

Abstract

Alternative drugs against leishmaniasis are desperately needed. Antimonials, the main chemotherapeutic tool, cause serious side effects and promote chemoresistance. We previously demonstrated that representatives of N,C-linked arylisoquinolines are promising leishmanicidal drug candidates. We now performed structure-activity relationship studies varying the aryl portion of our lead substrate. The new series of compounds show an enhanced selectivity against Leishmania major in comparison to their major host cell, the macrophage. Our results suggest that the arylisoquinolinium salts decrease the macrophage infection rate acting directly on the intracellular parasites. However, the activity of the 4'-i-propyl derivative might also involve the modulation of cytokine and nitric oxide production by host macrophages. Additionally, this isoquinoline acts synergistically with amphotericin B and does not interact with drug-metabolizing cytochrome P450 enzymes involved in the metabolism of antileishmanial drugs. The results demonstrate that the newly synthesized structurally simplified N,C-coupled arylisoquinolinium salts are promising candidates to be considered as leishmanicidal pharmacophores.

A Novel Leishmania major Amastigote Assay in 96-Well Format for Rapid Drug Screening and Its Use for Discovery and Evaluation of a New Class of Leishmanicidal Quinolinium Salts

“…In our previous experiments, peritoneal macrophages isolated from naive mice had been applied to EB-AO staining to determine IC 50 s against intracellular amastigotes (12,13,16). Up to three mice were required to determine the IC 50 s of one compound for L. major amastigotes by EB-AO staining.…”

“…1). IC 50 s for all compounds against L. major promastigotes and amastigotes determined in our laboratory have been published previously (12,13,16,30).…”

“…Antiparasitic activities against WT and Luc-tg L. major promastigotes with a final cell density of 1 ϫ 10 7 per ml were tested at 27°C using the alamarBlue assay as described previously (12,13,16).…”

“…Therefore, the most interesting compounds with low half-maximal inhibitory concentrations (IC 50 s) and good selectivity indices (SI) against L. major promastigotes detected by alamarBlue assay in our laboratory were further tested against intracellular amasti-gotes. We applied ethidium bromide-acridine orange (EB-AO) staining to determine the rate of infection of peritoneal macrophages with L. major amastigotes after treatment with different concentrations of drugs of interest (12,13,16). However, EB-AO staining is very labor-intensive, time-consuming, and expensive.…”

“…These quinolinium salts are analogs of the N,Ccoupled naphthylisoquinolines (18,19) representing a subclass of the naphthylisoquinoline alkaloids (20)(21)(22). After the discovery of ancisheynine, the first cationic (since N,C-coupled) naphthylisoquinoline, in 2003 by Yang and coworkers (18), these secondary metabolites have aroused much stereochemical and synthetic interest (23)(24)(25) because of their promising biological activities, e.g., against Leishmania, Plasmodium, and Trypanosoma (16,26). Based on structure-activity relationship (SAR) studies of the naturally occurring N,C-coupled naphthylisoquinolines, several derivatives with excellent activities have been synthesized in recent years.…”