Reversing antifungal resistance through disruption of ClpX

Fungal disease impacts the lives of almost a billion people across the globe. The opportunistic human fungal pathogen, Cryptococcus neoformans, causes cryptococcal meningitis in immunocompromised individuals with high fatality rates in response to limited treatment options. Moreover, the emergence of azole-resistant isolates in the clinic following prolonged treatment regimes, environmental fungicide exposure, and fungal evolution, threatens the outcome of current therapeutic options, endangering the survival of infected individuals. By quantitatively characterizing the proteomes of fluconazole-susceptible and -resistant C. neoformans strains using state-of-the-art tandem mass spectrometry, we defined ClpX, an ATP-dependent unfoldase, as a target to overcome resistance. We discovered that disruption of ClpX through deletion or inhibition re-introduces fluconazole susceptibility into the resistant strains, rendering treatment effective once again. We further explored the mechanism of susceptibility and determined interruption to heme biosynthesis and ergosterol production associated with ClpX. Our results contribute to the understanding of novel mechanisms driving fluconazole resistance and provide support for targeting proteins as a therapeutic strategy to combat resistance.