Leveraging synthetic genetic array screening to identify therapeutic targets and inhibitors for combatting azole resistance in <i>Candida glabrata</i>

Fungal infections affect over 6.55 million individuals annually, with <i>Candida glabrata</i> (recently reclassified as <i>Nakaseomyces glabratus</i>) being the second most common cause of invasive candidiasis and candidemia, conditions that carry a high mortality rate of approximately 63.6%. <i>C. glabrata</i> exhibits significant genomic plasticity, enabling rapid adaptation to antifungal treatments. This ability is largely driven by gain-of-function (GOF) mutations in the <i>CgPDR1</i> gene, which enhance the activity of efflux pumps and confer azole resistance. Over 80 unique <i>CgPDR1</i> mutations have been identified, leading to diverse resistance profiles, from single-azole resistance to pan-azole resistance. These mutations not only increase resistance but also enhance the virulence of <i>C. glabrata</i>. To combat multi-drug-resistant (MDR) strains, which show resistance to azoles, echinocandins, and polyenes, novel multi-target therapeutic approaches are essential. This study employs synthetic genetic array (SGA) screening to explore the genetic interactions underlying <i>CgPDR1</i>⁺-mediated azole resistance, using the clinical allele R592S. By generating genome-wide double mutant strains, we identified gene deletions that cause synthetic sick (SS) and synthetic lethal (SL) interactions, providing potential therapeutic targets. <i>In silico</i> screening identified inhibitors of these targets, and one was validated <i>in vitro</i>. Our findings support the potential of SGA screening to identify antifungal adjuvants that can slow the emergence of azole resistance in <i>C. glabrata</i>.IMPORTANCE<i>Candida glabrata</i> is a significant cause of life-threatening fungal infections, especially concerning due to its rapid development of resistance to commonly used antifungal drugs like azoles. The growing prevalence of multi-drug-resistant <i>C. glabrata</i> strains, coupled with their high mortality rate, underscores the urgency of identifying new treatment strategies. This study investigates the genetic mechanisms underlying drug resistance, specifically focusing on mutations in the <i>CgPDR1</i> gene, which increase azole resistance. By leveraging a novel genetic screening method, this work identifies genes that interact with drug-resistant mutations, offering potential new therapeutic targets. Additionally, an inhibitor identified through <i>in silico</i> screening shows promise in delaying azole resistance when tested <i>in vitro</i>. The findings highlight the potential of multi-target therapies to combat drug resistance, paving the way for more effective treatment options for <i>C. glabrata</i> infections and addressing an urgent clinical need.