PROJECT SUMMARY Fungal diseases affect an estimated 300 million people a year and lead to approximately 1.6 million deaths. Despite the high prevalence of serious fungal diseases, no area of infectious disease drug development has made less progress in the last fifty years than mycology. Of the three primary classes of drugs used to treat invasive fungal infections (IFIs), amphotericin B was introduced in the 1950s; azoles were developed in the 1970s; and the echinocandins were approved in 2002. Unfortunately, this slow rate of progress is not because these drugs are highly effective. Indeed, the mortality rates for invasive candidiasis, invasive aspergillosis, and cryptococcal meningitis, three of the most common IFIs, remain unacceptably high at 30-40%, 50%, and >50%, respectively. Furthermore, the poor outcomes associated with IFIs are likely to worsen as the incidence of antifungal drug resistance continues to rise. Accordingly, the clear consensus in the field is that novel classes of antifungal drugs represent one of the most pressing un-met clinical needs facing infectious disease. One approach to expediting this development is to optimize the antifungal activity of drugs currently used to treat other diseases. The phenothiazine class of molecules related to antipsychotic and antiemetic medications has been shown by us and others to have antifungal properties. In preliminary studies toward optimizing these compounds, we have been able to increase their antifungal activity and decrease their affinity for neurotransmitter receptors, the prime driver of dose limiting side effects. Here, we propose a hit-to-lead development of these phenothiazines focusing on structure guided reduction in dopamine/serotonin receptor activity and reducing multi drug efflux affinity. Our goal is to advance 3-4 molecules into initial animal efficacy studies. To achieve these goals we propose the following aims: 1) medicinal chemistry-based optimization of the antifungal therapeutic index of the phenothiazine scaffold; 2) characterize the mechanism of increased anti- candidal activity of novel fluphenazine and trifluoperazine derivatives; and 3) in vivo pharmacology and efficacy testing of three lead candidates using murine infection models.