1-s2.0-S107455211500201X-main.pdf (2.75 MB)
Thiophenecarboxamide derivatives activated by EthA kill Mycobacterium tuberculosis by inhibiting the CTP synthetase PyrG
journal contributionposted on 2020-09-02, 14:07 authored by Giorgia Mori, Laurent R Chiarelli, Marta Esposito, Vadim Makarov, Marco Bellinzoni, Ruben C Hartkoorn, Giulia Degiacomi, Francesca Boldrin, Sean Ekins, Ana Luisa de Jesus Lopes Ribeiro, Leonardo B Marino, Ivana Centárová, Zuzana Svetlíková, Jaroslav Blaško, Elena Kazakova, Alexander Lepioshkin, Nathalie Barilone, Giuseppe Zanoni, Alessio Porta, Marco Fondi, Renato Fani, Alain R Baulard, Katarína Mikušová, Pedro M Alzari, Riccardo Manganelli, Luiz Pedro S de Carvalho, Giovanna Riccardi, Stewart T Cole, Maria Rosalia Pasca
To combat the emergence of drug-resistant strains of Mycobacterium tuberculosis, new antitubercular agents and novel drug targets are needed. Phenotypic screening of a library of 594 hit compounds uncovered two leads that were active against M. tuberculosis in its replicating, non-replicating, and intracellular states: compounds 7947882 (5-methyl-N-(4-nitrophenyl)thiophene-2-carboxamide) and 7904688 (3-phenyl-N-[(4-piperidin-1-ylphenyl)carbamothioyl]propanamide). Mutants resistant to both compounds harbored mutations in ethA (rv3854c), the gene encoding the monooxygenase EthA, and/or in pyrG (rv1699) coding for the CTP synthetase, PyrG. Biochemical investigations demonstrated that EthA is responsible for the activation of the compounds, and by mass spectrometry we identified the active metabolite of 7947882, which directly inhibits PyrG activity. Metabolomic studies revealed that pharmacological inhibition of PyrG strongly perturbs DNA and RNA biosynthesis, and other metabolic processes requiring nucleotides. Finally, the crystal structure of PyrG was solved, paving the way for rational drug design with this newly validated drug target.
Activation, MetabolicAnimalsAntitubercular AgentsBacterial ProteinsCarbon-Nitrogen LigasesDrug DesignDrug Evaluation, PreclinicalHep G2 CellsHigh-Throughput Screening AssaysHumansMiceMicrobial Sensitivity TestsModels, MolecularMycobacterium tuberculosisOxidoreductasesProtein ConformationThiophenesde Carvalho MC_UP_A253_1111Organic Chemistry0304 Medicinal and Biomolecular Chemistry0305 Organic Chemistry0601 Biochemistry and Cell Biology