Identification and Mode of Action of a Plant Natural Product Targeting Human Fungal Pathogens

Abstract

Candida albicans is a major cause of fungal diseases in humans, and its resistance to available drugs is of concern. In an attempt to identify novel antifungal agents, we initiated a small-scale screening of a library of 199 natural plant compounds (i.e., natural products [NPs]). In vitro susceptibility profiling experiments identified 33 NPs with activity against C. albicans (MIC50s ≤ 32 μg/ml). Among the selected NPs, the sterol alkaloid tomatidine was further investigated. Tomatidine originates from the tomato (Solanum lycopersicum) and exhibited high levels of fungistatic activity against Candida species (MIC50s ≤ 1 μg/ml) but no cytotoxicity against mammalian cells. Genome-wide transcriptional analysis of tomatidine-treated C. albicans cells revealed a major alteration (upregulation) in the expression of ergosterol genes, suggesting that the ergosterol pathway is targeted by this NP. Consistent with this transcriptional response, analysis of the sterol content of tomatidine-treated cells showed not only inhibition of Erg6 (C-24 sterol methyltransferase) activity but also of Erg4 (C-24 sterol reductase) activity. A forward genetic approach in Saccharomyces cerevisiae coupled with whole-genome sequencing identified 2 nonsynonymous mutations in ERG6 (amino acids D249G and G132D) responsible for tomatidine resistance. Our results therefore unambiguously identified Erg6, a C-24 sterol methyltransferase absent in mammals, to be the main direct target of tomatidine. We tested the in vivo efficacy of tomatidine in a mouse model of C. albicans systemic infection. Treatment with a nanocrystal pharmacological formulation successfully decreased the fungal burden in infected kidneys compared to the fungal burden achieved by the use of placebo and thus confirmed the potential of tomatidine as a therapeutic agent.

Tomatidine, a natural steroidal alkaloid shows antiviral activity towards chikungunya virus in vitro

“…Unfortunately, to date literature on those aspects is scarce. Tomatidine has been used in several in vivo mouse studies and no toxicity was observed up to a concentration of 50 mg/kg 19,[36][37][38][39][40] . Only one study measured the steady-state tomatidine plasma levels and revealed a plasma concentration of 287 ng tomatidine per mL after 2 month of oral treatment with 0.05% (w/w) tomatidine added to standard chow 36 .…”

Tomatidine Is a Lead Antibiotic Molecule That Targets Staphylococcus aureus ATP Synthase Subunit C

“…On the other hand, TO, the aglycone version of ␣-tomatine, loses inhibitory activity against most phytopathogens (42) and does not possess sterol binding activity (43). In Saccharomyces cerevisiae and several protozoans, however, it was reported that TO perturbs ergosterol biosynthesis by targeting Erg6 (C-24 sterol methyltransferase) and also of Erg4 (C-24 sterol reductase), two enzymes absent in mammals and most bacteria (44)(45)(46)(47). Such findings in fungi and protozoans suggest a different mode of action for the potent and specific antibacterial activity of TO against SCVs of the Bacillales (13).…”

Systematic Analysis of Functionally Related Gene Clusters in the Opportunistic Pathogen, Candida albicans

“…Conditions that induced a broad range of cellular transcriptional responses for analysis were selected for analysis. The microarray and RNA-sequencing gene expression datasets utilized were: the transcriptional changes in response to carbon shift from glucose to xylose (GSE50476), response to the anti-fungal thiosemicarbazone, NSC319726, (GSE106486), response to the sterol alkaloid tomatidine (GSE96965), and the response to mucin (GSE149196) [ 46 , 47 , 48 ].…”