An Antiparasitic Compound from the Medicines for Malaria Venture Pathogen Box Promotes Tubulin Polymerization.

The few frontline antileishmanial drugs are poorly effective and toxic. To search for new drugs for this neglected tropical disease, we tested the activity of compounds in the Medicines for Malaria Venture (MMV) "Pathogen Box" against axenic amastigotes. Screening yielded six discovery antileishmanial compounds with EC values from 50 to 480 nM. Concentration-response assays demonstrated that the best hit, MMV676477, had mid-nanomolar cytocidal potency against intracellular amastigotes, , and , suggesting broad antiparasitic activity. We explored structure-activity relationships (SAR) within a small group of MMV676477 analogs and observed a wide potency range (20-5000 nM) against axenic amastigotes. Compared to MMV676477, our most potent analog, SW41, had ∼5-fold improved antileishmanial potency. Multiple lines of evidence suggest that MMV676477 selectively disrupts tubulin dynamics. Morphological studies indicated that MMV676477 and analogs affected during cell division. Differential centrifugation showed that MMV676477 promoted partitioning of cellular tubulin toward the polymeric form in parasites. Turbidity assays with purified and porcine tubulin demonstrated that MMV676477 promoted leishmanial tubulin polymerization in a concentration-dependent manner. Analogs' antiparasitic activity correlated with their ability to facilitate purified tubulin polymerization. Chemical cross-linking demonstrated binding of the MMV676477 scaffold to purified tubulin, and competition studies established a correlation between binding and antileishmanial activity. Our studies demonstrate that MMV676477 is a potent antiparasitic compound that preferentially promotes microtubule polymerization. Due to its selectivity for and broad-spectrum activity against multiple parasites, this scaffold shows promise for antiparasitic drug development.