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Title: The small molecule CBR-5884 inhibits the Candida albicans phosphatidylserine synthase
ABSTRACT Systemic infections byCandidaspp. 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 averageZ’ score of ~0.8, and seven compounds were identified that inhibit Cho1. Three of these, ebselen, LOC14, and CBR-5884, exhibited antifungal 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 thecho1∆∆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. IMPORTANCEFungal phosphatidylserine synthase (Cho1) is a logical antifungal target due to its crucial role in the virulence and viability of various fungal pathogens, and since it is absent in humans, drugs targeted at Cho1 are less likely to cause toxicity in patients. Using fungal Cho1 as a model, there have been two unsuccessful attempts to discover inhibitors for Cho1 homologs in whole-cell screens prior to this study. The compounds identified in these attempts do not act directly on the protein, resulting in the absence of known Cho1 inhibitors. The significance of our research is that we developed a high-throughput target-based assay and identified the first Cho1 inhibitor, CBR-5884, which acts both on the purified protein and its function in the cell. This molecule acts as a competitive inhibitor with aKivalue of 1,550 ± 245.6 nM and, thus, has the potential for development into a new class of antifungals targeting PS synthase.  more » « less
Award ID(s):
2310263
PAR ID:
10517157
Author(s) / Creator(s):
; ; ; ; ; ; ; ; ; ; ;
Editor(s):
Alspaugh, J Andrew
Publisher / Repository:
ASM Journals
Date Published:
Journal Name:
mBio
Volume:
15
Issue:
5
ISSN:
2150-7511
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
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