Development of Cdc14 inhibitors as novel antifungal agents against diverse fungal pathogens - PROJECT SUMMARY Invasive fungal diseases are a growing global health problem, currently estimated to contribute to the death of 3.8 million people each year. Primarily affecting the immune-compromised and those with chronic underlying health problems, fungal diseases are challenging to treat and exhibit very high mortality rates. There are few effective treatment options and pathogens are developing resistance to our limited antifungal drugs. Therefore, development of new antifungal agents directed towards novel targets with unique modes of action is a high priority. The overall objective of the proposed project is to develop novel antifungal lead compounds directed towards a recently validated new drug target, the Cdc14 protein phosphatase. Cdc14 is required for virulence of a variety of fungal species, including the most common human pathogen, Candida albicans. It’s structure and enzyme specificity are highly conserved across the fungal kingdom, and are unique among phosphatases. The ability to design chemical compounds that selectively inhibit Cdc14 phosphatases was recently demonstrated by this group, including an inhibitor that exhibits biological activity in C. albicans cells and can reduce virulence in a mouse model of invasive candidiasis. The proposed project will build on these findings to further optimize Cdc14 inhibitor compounds to have enhanced affinity, selectivity, bioavailability, and drug-like properties and to develop and use suitable models for pre-clinical testing. In the initial R21 phase, three independent Aims will be pursued. In Aim 1, the initial Cdc14 inhibitor will undergo lead optimization and SAR analysis based on structural modeling and recent success with the human orthologs to improve affinity and selectivity. This aim will involve organic synthesis coupled with in vitro enzyme assays. The most-improved compounds will be screened for cellular toxicity and improved in vivo efficacy against C. albicans. In Aim 2, the in vivo mechanism of action of the compounds in C. albicans cells will be characterized to test if Cdc14 is the relevant target responsible for the observed biological activity. In Aim 3, the importance of Cdc14 for virulence of two other fungal pathogens, Candida auris and Candida glabrata, will be tested using mouse infection models to begin addressing the potential breadth of action of Cdc14 inhibitors against diverse fungal diseases. In the subsequent R33 phase, the following aims will be pursued. In Aim 1, the optimization and SAR data from the R21 phase will be used to design and synthesize additional generations of Cdc14 inhibitor derivatives intended to maximize affinity, specificity and drug-like properties. In Aim 2, affinity and selectivity of the new compounds will be compared in biochemical assays, and mouse toxicity and pharmacokinetics measured. Finally, in Aim 3, efficacy of the most promising compounds will be assessed in cultured cells and mouse models of invasive candidiasis caused by C. albicans, C. auris, and C. glabrata. The R33 work will involve iterative rounds of optimization guided by output from the biochemical and in vivo assays, eventually resulting in lead compounds with promising characteristics for full pre-clinical and future clinical testing.
