title: Identification and evaluation of novel acetolactate synthase inhibitors as antifungal agents.
abstract: High-throughput phenotypic screening against the yeast Saccharomyces cerevisiae revealed a series of triazolopyrimidine-sulfonamide compounds with broad-spectrum antifungal activity, no significant cytotoxicity, and low protein binding. To elucidate the target of this series, we have applied a chemogenomic profiling approach using the S. cerevisiae deletion collection. All compounds of the series yielded highly similar profiles that suggested acetolactate synthase (Ilv2p, which catalyzes the first common step in branched-chain amino acid biosynthesis) as a possible target. The high correlation with profiles of known Ilv2p inhibitors like chlorimuron-ethyl provided further evidence for a similar mechanism of action. Genome-wide mutagenesis in S. cerevisiae identified 13 resistant clones with 3 different mutations in the catalytic subunit of acetolactate synthase that also conferred cross-resistance to established Ilv2p inhibitors. Mapping of the mutations into the published Ilv2p crystal structure outlined the chlorimuron-ethyl binding cavity, and it was possible to dock the triazolopyrimidine-sulfonamide compound into this pocket in silico. However, fungal growth inhibition could be bypassed through supplementation with exogenous branched-chain amino acids or by the addition of serum to the medium in all of the fungal organisms tested except for Aspergillus fumigatus. Thus, these data support the identification of the triazolopyrimidine-sulfonamide compounds as inhibitors of acetolactate synthase but suggest that targeting may be compromised due to the possibility of nutrient bypass in vivo.
authors.affiliation: Novartis Institutes for BioMedical Research, Cambridge, Massachusetts, USA.
authors.forename: Daryl L
authors.initials: DL
authors.lastname: Richie
authors.forename: Katherine V
authors.initials: KV
authors.lastname: Thompson
authors.forename: Christian
authors.initials: C
authors.lastname: Studer
authors.forename: Vivian C
authors.initials: VC
authors.lastname: Prindle
authors.forename: Thomas
authors.initials: T
authors.lastname: Aust
authors.forename: Ralph
authors.initials: R
authors.lastname: Riedl
authors.forename: David
authors.initials: D
authors.lastname: Estoppey
authors.forename: Jianshi
authors.initials: J
authors.lastname: Tao
authors.forename: Jessica A
authors.initials: JA
authors.lastname: Sexton
authors.forename: Thomas
authors.initials: T
authors.lastname: Zabawa
authors.forename: Joseph
authors.initials: J
authors.lastname: Drumm
authors.forename: Simona
authors.initials: S
authors.lastname: Cotesta
authors.forename: Jürg
authors.initials: J
authors.lastname: Eichenberger
authors.forename: Sven
authors.initials: S
authors.lastname: Schuierer
authors.forename: Nicole
authors.initials: N
authors.lastname: Hartmann
authors.forename: N Rao
authors.initials: NR
authors.lastname: Movva
authors.forename: John A
authors.initials: JA
authors.lastname: Tallarico
authors.forename: Neil S
authors.initials: NS
authors.lastname: Ryder
authors.forename: Dominic
authors.initials: D
authors.lastname: Hoepfner
mesh_terms: D000095:Acetolactate Synthase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000597:Amino Acids, Branched-Chain / Q000378:metabolism / Q000494:pharmacology; D000935:Antifungal Agents / Q000737:chemistry / Q000494:pharmacology*; D020134:Catalytic Domain / Q000187:drug effects; D057166:High-Throughput Screening Assays; D006801:Humans; D008826:Microbial Sensitivity Tests; D062105:Molecular Docking Simulation; D009154:Mutation; D011485:Protein Binding; D011743:Pyrimidines / Q000737:chemistry / Q000494:pharmacology*; D012441:Saccharomyces cerevisiae / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D044967:Serum / Q000737:chemistry / Q000378:metabolism; D013449:Sulfonamides / Q000737:chemistry / Q000494:pharmacology*; D013453:Sulfonylurea Compounds / Q000737:chemistry / Q000494:pharmacology*
chemical_list: D000597:Amino Acids, Branched-Chain; D000935:Antifungal Agents; D011743:Pyrimidines; D029701:Saccharomyces cerevisiae Proteins; D013449:Sulfonamides; D013453:Sulfonylurea Compounds; C112343:chlorimuron ethyl; D000095:Acetolactate Synthase