Benzamidine Conjugation Converts Expelled Potential Active Agents into Antifungals against Drug-Resistant Fungi

Fungal infections present a growing global public health concern, necessitating the development of novel antifungal drugs. However, many potential antifungals, particularly the expelled potential active agents (EPAAs), are often underestimated owing to their limitations in cellular entry or expulsion by efflux pumps. Herein, we identified 68 EPAAs out of 2322 candidates with activity against a Candida albicans efflux pump-deficient strain and no inhibitory activity against the wild-type strain. Using a novel conjugation strategy involving benzamidine (BM) as a mitochondrion-targeting warhead, we successfully converted EPAAs into potent antifungals against various urgent-threat azole-resistantCandida strains. Among the obtained EPAA-BM conjugates, IS-2-BM (11) exhibited excellent antifungal activities and induced negligible drug resistance. Furthermore, IS-2-BM prevented biofilm formation, eradicated mature biofilms, and exhibited excellent therapeutic effects in a murine model of systemic candidiasis. These findings provide a promising strategy for increasing the possibilities of discovering more antifungals.