Endoplasmic reticulum stress reduces caspofungin resistance in Cryptococcus neoformans

Echinocandin antifungal drugs, such as caspofungin, inhibit β-1,3-glucan synthesis to impair cell wall formation. Caspofungin is effective in the clinic against Aspergillus and Candida species but ineffective against Cryptococcus neoformans, the fungus responsible for life-threatening meningoencephalitis in immunocompromised individuals. The mechanisms of caspofungin resistance in C. neoformans are associated with calcineurin signaling and functions that influence membrane and cell wall composition. The synergistic effects of caspofungin have been studied in combination with various inhibitors, but it is not known whether resistance can be overcome by agents that target the endoplasmic reticulum to influence protein glycosylation, as well as cell wall and membrane composition. Additionally, the role of the polysaccharide capsule of C. neoformans in blocking the access of caspofungin to the target Fks1 glucan synthase is not clear. Here, we identified a potential synergistic effect between caspofungin and the glycosylation inhibitor tunicamycin, with a significant reduction in proliferation, capsule attachment, and chitin and β-1,3-glucan levels when the drugs were used in combination. A targeted screen also identified other agents that influence ER function and that potentially display synergy with caspofungin. Overall, our analysis provides evidence that targeting the ER to impair cell wall and membrane integrity and capsule formation is an effective strategy to overcome caspofungin resistance in C. neoformans.IMPORTANCECryptococcus neoformans is the major cause of life-threatening meningoencephalitis in immunocompromised people. Unfortunately, the number of antifungal drugs available to treat cryptococcal disease is quite limited, and unlike other pathogenic fungi, C. neoformans is intrinsically resistant to the echinocandin group of drugs. These drugs target cell wall biosynthesis, and the mechanisms of cryptococcal resistance are not entirely clear. In this study, we demonstrate synergy between the echinocandin drug caspofungin and tunicamycin, an inhibitor of protein glycosylation in the endoplasmic reticulum. Combined treatment impacts cell wall and plasma membrane composition, thus providing insights into approaches to overcome resistance.