Dual action antifungal small molecule modulates multidrug efflux and TOR signaling

Abstract

There is an urgent need for new strategies to treat invasive fungal infections, which are a leading cause of human mortality. We establish two activities of the natural product beauvericin, which potentiates the activity of the most widely deployed class of antifungal against the leading human fungal pathogens, blocks the emergence of drug resistance, and renders resistant pathogens responsive to treatment in mammalian infection models. Harnessing genome sequencing of beauvericin-resistant mutants, affinity purification of a biotinylated beauvericin analog, and biochemical and genetic assays reveals that beauvericin blocks multidrug efflux and inhibits the global regulator TORC1 kinase, thereby activating protein kinase CK2 and inhibiting the molecular chaperone Hsp90. Substitutions in the multidrug transporter Pdr5 that enable beauvericin efflux impair antifungal efflux, thereby impeding resistance to the drug combination. Thus, dual targeting of multidrug efflux and TOR signaling provides a powerful, broadly effective therapeutic strategy for fungal infectious disease that evades resistance.

Beauvericin Potentiates Azole Activity via Inhibition of Multidrug Efflux, Blocks Candida albicans Morphogenesis, and Is Effluxed via Yor1 and Circuitry Controlled by Zcf29

“…Beauvericin impedes the emergence of resistance to azoles and ameliorates the therapeutic efficacy of azoles in murine models of candidiasis (21,22). We previously established the mechanisms underpinning the synergy in that beauvericin inhibits both multidrug efflux and TORC1 kinase; the latter activity stimulates protein kinase CK2 and inhibits the molecular chaperone Hsp90 (22). In the present study, we establish that beauvericin also inhibits multidrug efflux and TOR signaling in C. albicans but that the antifungal activity is independent of protein kinase CK2.…”

“…To determine whether beauvericin potentiates azole activity via inhibition of multidrug efflux in C. albicans, we assessed the drug synergy against strains lacking the ABC transporter genes CDR1 and CDR2 both individually and in combination. Cdr1 and Cdr2 are homologs of the S. cerevisiae ABC transporter Pdr5 (30,31), which we have shown to be inhibited by beauvericin (22). The synergy between beauvericin and fluconazole was abrogated in strains lacking CDR1 but not CDR2 (Fig.…”

“…We have previously shown that beauvericin enhances the efficacy of azoles against the model yeast Saccharomyces cerevisiae and the human fungal pathogens C. albicans, C. neoformans, and A. fumigatus (22). Using Etest strips to generate a concentration gradient of the azole fluconazole on solid rich medium, we further tested the ability of beauvericin to enhance azole efficacy against intrinsically azole-resistant Candida glabrata.…”

“…We previously demonstrated that beauvericin inhibits multidrug efflux in S. cerevisiae, and a recent study suggests that beauvericin inhibits the C. albicans ABC transporters Cdr1 and Cdr2 (22,29). To determine whether beauvericin potentiates azole activity via inhibition of multidrug efflux in C. albicans, we assessed the drug synergy against strains lacking the ABC transporter genes CDR1 and CDR2 both individually and in combination.…”

“…We and others have previously shown that beauvericin potentiates azole activity against diverse fungal pathogens, including C. albicans, Aspergillus fumigatus, and Cryptococcus neoformans (20)(21)(22). Beauvericin impedes the emergence of resistance to azoles and ameliorates the therapeutic efficacy of azoles in murine models of candidiasis (21,22).…”