The small molecule CBR-5884 inhibits theCandida albicansphosphatidylserine synthase

Abstract

Systemic infections byCandida spp.are associated with high mortality rates, partly due to limitations in current antifungals, highlighting the need for novel drugs and drug targets. The fungal phosphatidylserine synthase, Cho1, fromCandida albicansis a logical antifungal drug target due to its importance in virulence, absence in the host and conservation among fungal pathogens. Inhibitors of Cho1 could serve as lead compounds for drug development, so we developed a target-based screen for inhibitors of purified Cho1. This enzyme condenses serine and cytidyldiphosphate-diacylglycerol (CDP-DAG) into phosphatidylserine (PS) and releases cytidylmonophosphate (CMP). Accordingly, we developed anin vitronucleotidase-coupled malachite green-based high throughput assay for purifiedC. albicansCho1 that monitors CMP production as a proxy for PS synthesis. Over 7,300 molecules curated from repurposing chemical libraries were interrogated in primary and dose-responsivity assays using this platform. The screen had a promising average Z’ score of ∼0.8, and seven compounds were identified that inhibit Cho1. Three of these, ebselen, LOC14, and CBR-5884, exhibited inhibitory effects againstC. albicanscells, with fungicidal inhibition by ebselen and fungistatic inhibition by LOC14 and CBR-5884. Only CBR-5884 showed evidence of disruptingin vivoCho1 function by inducing phenotypes consistent with thecho1ΔΔ mutant, including a reduction of cellular PS levels. Kinetics curves and computational docking indicate that CBR-5884 competes with serine for binding to Cho1 with aKiof 1,550 ± 245.6 nM. Thus, this compound has the potential for development into an antifungal compound.

Innovations in Antifungal Drug Discovery among Cell Envelope Synthesis Enzymes through Structural Insights

“…The other screen identified bleomycin, but it was also found that this compound affects phospholipid-associated processes rather than targeting C. albicans PS synthase directly [ 245 ]. Recently, Zhou et al identified a molecule, CBR-5884 ( Table 3 ), that inhibits both purified PS synthase and its function in vivo, with a K i of 1550 ± 245.6 nM [ 121 ]. This molecule acts as a competitive inhibitor for serine, thus having the potential for further development.…”