G-quadruplexes represent promising new targets to overcome multidrug-resistant fungal infections

The growing emergence of antifungal resistance has prompted the identification of novel antifungal targets. G-quadruplexes (G4s), four-stranded secondary structures that form in DNA and RNA, have arisen as a drug target to treat bacterial, viral, and parasitic infections. Here, we provide the first demonstration that G4s form in fungi and represent a promising new target for antifungal development. We found that PhenDC3 and pyridostatin (PDS), ligands that bind to and stabilise G4s, potently inhibited the metabolism of fungal pathogens in different ways. These pathogens included the pan azole-resistant Aspergillus fumigatus isolate TR34/L98H and Candida auris. Notably, PhenDC3 could synergise with amphotericin B, was protective in an in vivo model of fungal infection, and was well-tolerated by human cells. However, continuous exposure to PhenDC3 resulted in some cross-resistance to current antifungals and this needs to be explored further. PhenDC3 could also increase the number of RNA G4s in live A. fumigatus. Further, we demonstrated the structural flexibility of DNA sequences found in cyp51A and cyp51B, with these sequences capable of forming duplexes, hairpins, G4s and i-motifs. Finally, PhenDC3, but not PDS, caused duplex DNA structures in cyp51A to transition into antiparallel G4 structures potentially associated with PhenDC3s increased antifungal potency. Taken together, G4s represent an exciting antifungal target, but a more detailed understanding of their biological roles is essential.