title: 2-Carboxyquinoxalines kill mycobacterium tuberculosis through noncovalent inhibition of DprE1.
abstract: Phenotypic screening of a quinoxaline library against replicating Mycobacterium tuberculosis led to the identification of lead compound Ty38c (3-((4-methoxybenzyl)amino)-6-(trifluoromethyl)quinoxaline-2-carboxylic acid). With an MIC99 and MBC of 3.1 μM, Ty38c is bactericidal and active against intracellular bacteria. To investigate its mechanism of action, we isolated mutants resistant to Ty38c and sequenced their genomes. Mutations were found in rv3405c, coding for the transcriptional repressor of the divergently expressed rv3406 gene. Biochemical studies clearly showed that Rv3406 decarboxylates Ty38c into its inactive keto metabolite. The actual target was then identified by isolating Ty38c-resistant mutants of an M. tuberculosis strain lacking rv3406. Here, mutations were found in dprE1, encoding the decaprenylphosphoryl-d-ribose oxidase DprE1, essential for biogenesis of the mycobacterial cell wall. Genetics, biochemical validation, and X-ray crystallography revealed Ty38c to be a noncovalent, noncompetitive DprE1 inhibitor. Structure-activity relationship studies generated a family of DprE1 inhibitors with a range of IC50's and bactericidal activity. Co-crystal structures of DprE1 in complex with eight different quinoxaline analogs provided a high-resolution interaction map of the active site of this extremely vulnerable target in M. tuberculosis.
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|‡Global Health Institute, Ecole Polytechnique Fédérale de Lausanne, 1015 Lausanne, Switzerland.
authors.forename: João
authors.initials: J
authors.lastname: Neres
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|‡Global Health Institute, Ecole Polytechnique Fédérale de Lausanne, 1015 Lausanne, Switzerland.
authors.forename: Ruben C
authors.initials: RC
authors.lastname: Hartkoorn
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|§Department of Biology and Biotecnology "Lazzaro Spallanzani", University of Pavia, 27100 Pavia, Italy.
authors.forename: Laurent R
authors.initials: LR
authors.lastname: Chiarelli
authors.affiliation: ∥Tydock Pharma, srl Via Campi 183, 41125 Modena, Italy.|⊥Department of Life Sciences, University of Modena and Reggio Emilia, Via Campi 183, 41126 Modena, Italy.
authors.forename: Ramakrishna
authors.initials: R
authors.lastname: Gadupudi
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|§Department of Biology and Biotecnology "Lazzaro Spallanzani", University of Pavia, 27100 Pavia, Italy.
authors.forename: Maria Rosalia
authors.initials: MR
authors.lastname: Pasca
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|§Department of Biology and Biotecnology "Lazzaro Spallanzani", University of Pavia, 27100 Pavia, Italy.
authors.forename: Giorgia
authors.initials: G
authors.lastname: Mori
authors.affiliation: ∥Tydock Pharma, srl Via Campi 183, 41125 Modena, Italy.
authors.forename: Alberto
authors.initials: A
authors.lastname: Venturelli
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|#A. N. Bakh Institute of Biochemistry, Russian Academy of Science, 119071 Moscow, Russia.
authors.forename: Svetlana
authors.initials: S
authors.lastname: Savina
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|#A. N. Bakh Institute of Biochemistry, Russian Academy of Science, 119071 Moscow, Russia.
authors.forename: Vadim
authors.initials: V
authors.lastname: Makarov
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|‡Global Health Institute, Ecole Polytechnique Fédérale de Lausanne, 1015 Lausanne, Switzerland.
authors.forename: Gaelle S
authors.initials: GS
authors.lastname: Kolly
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|§Department of Biology and Biotecnology "Lazzaro Spallanzani", University of Pavia, 27100 Pavia, Italy.
authors.forename: Elisabetta
authors.initials: E
authors.lastname: Molteni
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|§Department of Biology and Biotecnology "Lazzaro Spallanzani", University of Pavia, 27100 Pavia, Italy.
authors.forename: Claudia
authors.initials: C
authors.lastname: Binda
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|‡Global Health Institute, Ecole Polytechnique Fédérale de Lausanne, 1015 Lausanne, Switzerland.
authors.forename: Neeraj
authors.initials: N
authors.lastname: Dhar
authors.affiliation: ∥Tydock Pharma, srl Via Campi 183, 41125 Modena, Italy.|⊥Department of Life Sciences, University of Modena and Reggio Emilia, Via Campi 183, 41126 Modena, Italy.
authors.forename: Stefania
authors.initials: S
authors.lastname: Ferrari
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|∇Inserm U1019 - CNRS UMR 8204, Institut Pasteur de Lille, Université de Lille, 1 rue du Professeur Calmette, 59019, Lille, France.
authors.forename: Priscille
authors.initials: P
authors.lastname: Brodin
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|∇Inserm U1019 - CNRS UMR 8204, Institut Pasteur de Lille, Université de Lille, 1 rue du Professeur Calmette, 59019, Lille, France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Delorme
authors.affiliation: ∇Inserm U1019 - CNRS UMR 8204, Institut Pasteur de Lille, Université de Lille, 1 rue du Professeur Calmette, 59019, Lille, France.
authors.forename: Valérie
authors.initials: V
authors.lastname: Landry
authors.affiliation: §Department of Biology and Biotecnology "Lazzaro Spallanzani", University of Pavia, 27100 Pavia, Italy.
authors.forename: Ana Luisa
authors.initials: AL
authors.lastname: de Jesus Lopes Ribeiro
authors.affiliation: ⊥Department of Life Sciences, University of Modena and Reggio Emilia, Via Campi 183, 41126 Modena, Italy.
authors.forename: Davide
authors.initials: D
authors.lastname: Farina
authors.affiliation: ⊥Department of Life Sciences, University of Modena and Reggio Emilia, Via Campi 183, 41126 Modena, Italy.
authors.forename: Puneet
authors.initials: P
authors.lastname: Saxena
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|‡Global Health Institute, Ecole Polytechnique Fédérale de Lausanne, 1015 Lausanne, Switzerland.
authors.forename: Florence
authors.initials: F
authors.lastname: Pojer
authors.affiliation: ○Department of Chemistry and Pharmacy, University of Sassari, 07100 Sassari, Italy.
authors.forename: Antonio
authors.initials: A
authors.lastname: Carta
authors.affiliation: ⊥Department of Life Sciences, University of Modena and Reggio Emilia, Via Campi 183, 41126 Modena, Italy.
authors.forename: Rosaria
authors.initials: R
authors.lastname: Luciani
authors.affiliation: ◆Department of Chemistry, University of Pavia, 27100 Pavia, Italy.
authors.forename: Alessio
authors.initials: A
authors.lastname: Porta
authors.affiliation: ◆Department of Chemistry, University of Pavia, 27100 Pavia, Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Zanoni
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|§Department of Biology and Biotecnology "Lazzaro Spallanzani", University of Pavia, 27100 Pavia, Italy.
authors.forename: Edda
authors.initials: E
authors.lastname: De Rossi
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|∥Tydock Pharma, srl Via Campi 183, 41125 Modena, Italy.|⊥Department of Life Sciences, University of Modena and Reggio Emilia, Via Campi 183, 41126 Modena, Italy.
authors.forename: Maria Paola
authors.initials: MP
authors.lastname: Costi
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|§Department of Biology and Biotecnology "Lazzaro Spallanzani", University of Pavia, 27100 Pavia, Italy.
authors.forename: Giovanna
authors.initials: G
authors.lastname: Riccardi
authors.affiliation: †More Medicines for Tuberculosis (MM4TB) Consortium (www.mm4tb.org).|‡Global Health Institute, Ecole Polytechnique Fédérale de Lausanne, 1015 Lausanne, Switzerland.
authors.forename: Stewart T
authors.initials: ST
authors.lastname: Cole
mesh_terms: D000429:Alcohol Oxidoreductases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000995:Antitubercular Agents / Q000138:chemical synthesis / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D002473:Cell Wall / Q000187:drug effects / Q000201:enzymology; D018360:Crystallography, X-Ray; D055808:Drug Discovery; D004791:Enzyme Inhibitors / Q000138:chemical synthesis / Q000494:pharmacology*; D015870:Gene Expression; D006860:Hydrogen Bonding; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000201:enzymology / Q000254:growth & development; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011810:Quinoxalines / Q000138:chemical synthesis / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D054852:Small Molecule Libraries / Q000138:chemical synthesis / Q000494:pharmacology*; D013329:Structure-Activity Relationship
chemical_list: D000995:Antitubercular Agents; D001426:Bacterial Proteins; D004791:Enzyme Inhibitors; D011810:Quinoxalines; D011994:Recombinant Proteins; D054852:Small Molecule Libraries; D000429:Alcohol Oxidoreductases; C583625:DprE1 protein, Mycobacterium tuberculosis