Lanosterol synthase inhibition as a potential approach for re-sensitizing triazole-resistant Aspergillus fumigatus

The increasing prevalence of triazole resistance among Aspergillus fumigatus clinical isolates threatens the efficacy of this front-line treatment, highlighting the urgent need for novel therapeutic targets. In this study, we characterized three putative lanosterol synthase (also known as oxidosqualene cyclase) paralogs and identified Erg7A as the dominant enzyme. Although erg7B provided functional redundancy when overexpressed, deletion of erg7A alone created a significant ergosterol biosynthetic blockade. erg7A deficiency led to the accumulation of upstream substrates (squalene and 2,3-oxidosqualene) as well as downstream sterol intermediates, including lanosterol and eburicol. This sterol perturbation conferred triazole hypersusceptibility to a laboratory strain and restored susceptibility in resistant clinical isolates harboring cyp51A or hmg1 mutations. We further validated these findings using Ro 48-8071, a specific oxidosqualene cyclase inhibitor, and found that Ro 48-8071 exhibited potent synergy with the long-tailed triazoles, itraconazole, and posaconazole. Our findings demonstrate that targeting Erg7 sensitizes A. fumigatus to triazoles, offering a potential combination therapy against resistant isolates.IMPORTANCEAspergillus fumigatus is a mold pathogen and the major etiologic agent of invasive aspergillosis. Treatment of invasive aspergillosis is currently limited to only three classes of antifungals, the use of which is hampered by toxicity and rising antifungal resistance rates. The front-line therapeutic class, the triazoles, functions through the inhibition of ergosterol biosynthesis, and the majority of clinical triazole resistance is caused by target gene mutations impacting the sterol demethylase Cyp51. In this study, we differentiate the biological roles of three putative A. fumigatus lanosterol synthase paralogs as enzymes functioning upstream of the triazole target. We provide strong genetic and pharmacological evidence that inhibition of this upstream enzymatic step in the ergosterol biosynthetic pathway is synergistic with triazoles and can resensitize resistant clinical isolates. The development of lanosterol synthase inhibitors with fungus-specific activity is expected to provide new stand-alone and/or combination therapies to improve outcomes of invasive aspergillosis.