Leveraging Vulnerabilities in Copper Trafficking for Synergistic Antifungal Activity

Candida albicans is an opportunistic fungal pathogen that causes millions of infections per year, for which more efficacious treatments are needed. Observations that azole antifungals incite C. albicans to adjust a variety of metal-dependent processes led us to hypothesize that vulnerabilities in metallohomeostasis incurred by drug stress could be leveraged by compounds that interrupt metal trafficking. Here, we show that tetrathiomolybdate (TTM), a copper (Cu) chelator that interferes with Cu trafficking and use, inhibits growth of C. albicans on its own and synergizes with select azoles to enhance antifungal activity. Proteomic and biochemical experiments revealed that TTM causes differential expression and stabilization of proteins involved in fermentation and oxidative stress responses in C. albicans. The synergy between TTM and azoles was found to arise from increased expression and stability of the nitric oxide dioxygenase Yhb1, a response driven by the decreased stability and activity incurred by TTM of CuZn superoxide dismutase 1. Addition of imidazole-based antifungals highjacks this stress response by inhibiting Yhb1. This study highlights the centrality of Cu homeostasis as a regulatory hub connecting energy production, oxidative stress management, and overall cellular fitness in ways that can be pharmacologically manipulated to enhance efficacy of existing antifungal agents.白色念珠菌（Candida albicans）是一种机会致病性真菌病原体，每年引发数百万例感染，目前仍亟需更为有效的治疗手段。研究观察到唑类（azole）抗真菌药可诱导白色念珠菌调控多种金属依赖的生理过程，据此我们提出假说：药物应激所引发的金属稳态脆弱性，可被干扰金属转运的化合物加以利用。本研究证实，四硫钼酸盐（tetrathiomolybdate，TTM）作为一种可干扰铜（Cu）转运与利用的铜螯合剂，能够单独抑制白色念珠菌的生长，且可与特定唑类药物协同，增强抗真菌活性。蛋白质组学与生化实验结果显示，四硫钼酸盐会使白色念珠菌中参与发酵与氧化应激反应的蛋白质出现差异表达与稳定性改变。研究发现，四硫钼酸盐与唑类药物的协同作用，源于一氧化氮双加氧酶（nitric oxide dioxygenase）Yhb1的表达与稳定性上调，而这一响应由四硫钼酸盐对CuZn超氧化物歧化酶1（CuZn superoxide dismutase 1）的稳定性与活性的抑制所驱动。咪唑类抗真菌药物可通过抑制Yhb1劫持该应激响应通路。本研究凸显了铜稳态作为调控枢纽的核心地位，其可连接能量产生、氧化应激管理与整体细胞健康状态，且可通过药理学手段加以干预，从而增强现有抗真菌药物的临床疗效。