title: Kinase inhibitor profile for human nek1, nek6, and nek7 and analysis of the structural basis for inhibitor specificity.
abstract: Human Neks are a conserved protein kinase family related to cell cycle progression and cell division and are considered potential drug targets for the treatment of cancer and other pathologies. We screened the activation loop mutant kinases hNek1 and hNek2, wild-type hNek7, and five hNek6 variants in different activation/phosphorylation statesand compared them against 85 compounds using thermal shift denaturation. We identified three compounds with significant Tm shifts: JNK Inhibitor II for hNek1(Δ262-1258)-(T162A), Isogranulatimide for hNek6(S206A), andGSK-3 Inhibitor XIII for hNek7wt. Each one of these compounds was also validated by reducing the kinases activity by at least 25%. The binding sites for these compounds were identified by in silico docking at the ATP-binding site of the respective hNeks. Potential inhibitors were first screened by thermal shift assays, had their efficiency tested by a kinase assay, and were finally analyzed by molecular docking. Our findings corroborate the idea of ATP-competitive inhibition for hNek1 and hNek6 and suggest a novel non-competitive inhibition for hNek7 in regard to GSK-3 Inhibitor XIII. Our results demonstrate that our approach is useful for finding promising general and specific hNekscandidate inhibitors, which may also function as scaffolds to design more potent and selective inhibitors.
authors.affiliation: LaboratórioNacional de Biociências, Centro Nacional de PesquisaemEnergia e Materiais, Campinas, 13083-970 SP, Brazil. eduardo.moraes@bioetanol.org.br.
authors.forename: Eduardo Cruz
authors.initials: EC
authors.lastname: Moraes
authors.affiliation: LaboratórioNacional de Biociências, Centro Nacional de PesquisaemEnergia e Materiais, Campinas, 13083-970 SP, Brazil. gabriela.meirelles@lnbio.cnpem.br.
authors.forename: Gabriela Vaz
authors.initials: GV
authors.lastname: Meirelles
authors.affiliation: LaboratórioNacional de Biociências, Centro Nacional de PesquisaemEnergia e Materiais, Campinas, 13083-970 SP, Brazil. rodrigo.honorato@lnbio.cnpem.br.
authors.forename: Rodrigo Vargas
authors.initials: RV
authors.lastname: Honorato
authors.affiliation: Instituto Carlos Chagas, FIOCRUZ, Curitiba, 81350-010 PR, Brazil. tatiana.brasil@gmail.com.
authors.forename: Tatiana de Arruda Campos Brasil
authors.initials: Tde A
authors.lastname: de Souza
authors.affiliation: LaboratórioNacional de Biociências, Centro Nacional de PesquisaemEnergia e Materiais, Campinas, 13083-970 SP, Brazil. edmarcia.souza@lnbio.cnpem.br.
authors.forename: Edmarcia Elisa
authors.initials: EE
authors.lastname: de Souza
authors.affiliation: LaboratórioNacional de Biociências, Centro Nacional de PesquisaemEnergia e Materiais, Campinas, 13083-970 SP, Brazil. mario.murakami@lnbio.cnpem.br.
authors.forename: Mario Tyago
authors.initials: MT
authors.lastname: Murakami
authors.affiliation: LaboratórioNacional de Biociências, Centro Nacional de PesquisaemEnergia e Materiais, Campinas, 13083-970 SP, Brazil. paulo.oliveira@lnbio.cnpem.br.
authors.forename: Paulo Sergio Lopes
authors.initials: PS
authors.lastname: de Oliveira
authors.affiliation: Programa de Pós-graduação em Biologia Funcional e Molecular, Departamento de Bioquímica e BiologiaTecidual, Instituto de Biologia, UniversidadeEstadual de Campinas, Campinas, 13083-862 SP, Brazil. jorgkoba@unicamp.br.
authors.forename: Jörg
authors.initials: J
authors.lastname: Kobarg
mesh_terms: D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006801:Humans; D062105:Molecular Docking Simulation; D000072380:NIMA-Related Kinase 1; D000072379:NIMA-Related Kinases; D047428:Protein Kinase Inhibitors / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011994:Recombinant Proteins / Q000187:drug effects / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D018797:Cell Cycle Proteins; D047428:Protein Kinase Inhibitors; D011994:Recombinant Proteins; C000606914:NEK1 protein, human; C459456:NEK6 protein, human; C512685:NEK7 protein, human; D000072380:NIMA-Related Kinase 1; D000072379:NIMA-Related Kinases; D017346:Protein Serine-Threonine Kinases