High-Resolution Genetics Identifies the Lipid Transfer Protein Sec14p as Target for Antifungal Ergolines

Abstract

Invasive infections by fungal pathogens cause more deaths than malaria worldwide. We found the ergoline compound NGx04 in an antifungal screen, with selectivity over mammalian cells. High-resolution chemogenomics identified the lipid transfer protein Sec14p as the target of NGx04 and compound-resistant mutations in Sec14p define compound-target interactions in the substrate binding pocket of the protein. Beyond its essential lipid transfer function in a variety of pathogenic fungi, Sec14p is also involved in secretion of virulence determinants essential for the pathogenicity of fungi such as Cryptococcus neoformans, making Sec14p an attractive antifungal target. Consistent with this dual function, we demonstrate that NGx04 inhibits the growth of two clinical isolates of C. neoformans and that NGx04-related compounds have equal and even higher potency against C. neoformans. Furthermore NGx04 analogues showed fungicidal activity against a fluconazole resistant C. neoformans strain. In summary, we present genetic evidence that NGx04 inhibits fungal Sec14p and initial data supporting NGx04 as a novel antifungal starting point.

Target Identification and Mechanism of Action of Picolinamide and Benzamide Chemotypes with Antifungal Properties

“…In summary, the structural and functional data gained by the experimental approach in this study, together with the experimental tools and data from previous studies (Filipuzzi et al, 2016; Khan et al, 2016; Nile et al, 2014), now provide the scientific community a robust roadmap to the design, synthesis, and validation of the next generation of Sec14p inhibitors aimed at treating severe fungal infections.…”

“…This report describes the identification of a series of picolinamide and benzamide-containing compounds as a novel class of small molecule inhibitors of the S. cerevisiae PtdIns/PtdCho transfer protein Sec14p. Although there are two previous reports of Sec14 inhibitors (Filipuzzi et al, 2016; Nile et al, 2014), this study presents the first Sec14p::SMI co-crystal structure. This structure, rationalized both the functional variomics and SAR results of more than 40 generated analogues, supports strong structural fungal selectivity over the mammalian homolog and represents a quantum leap for rational design of the next generation of improved Sec14p inhibitors.…”

“…Sec14p is an attractive antifungal target as it has been shown to be druggable (Filipuzzi et al, 2016; Nile et al, 2014), it executes essential functions in several pathogenic fungi and it is required for efficient secretion of pathogenicity factors (Chayakulkeeree et al, 2011). We therefore sought to delineate structure-activity relationships by producing and testing a small compound collection generated by coupling different carboxylic acids and amines via established chemical synthesis methods (El-Faham and Albericio, 2010; El-Faham et al, 2009) (Figure 2, supplementary Data S1).…”

Modified carbazoles destabilize microtubules and kill glioblastoma multiform cells

“…Specifically, these cells represent a recognized preclinical model system for testing for the efficacy of novel treatments of GBM as they are categorized in three subtypes based on their mutation profile (i.e., proneural, mesenchymal and classical) that differentially respond to treatment [16][17][18][19][20]. To test for potential liver toxicity, we treated the human liver cell line HepG2, a commonly used cell culture model system [21][22][23][24]. Therefore, we treated the PD-GBM cells in culture with the four most potent modified carbazoles (8, 20, 27 and 25) from our initial screen and found that they kill the PD-GBM cells with EC 50 values and maximal efficacy values that both mirrored their activity on T98G cells and remained within similar values irrespective of the PD-GBM subtype (Table 3).…”

Synthesis, Antifungal Activities and Molecular Docking Studies of Benzoxazole and Benzothiazole Derivatives

“…Sec14p, the major PITP, executes essential functions in several pathogenic fungi and it is required for the efficient secretion of pathogenicity factors. It is an attractive antifungal target as it has been shown to be druggable [ 30 , 31 ].…”