title: Design, synthesis, biological evaluation and X-ray structural studies of potent human dihydroorotate dehydrogenase inhibitors based on hydroxylated azole scaffolds.
abstract: A new generation of potent hDHODH inhibitors designed by a scaffold-hopping replacement of the quinolinecarboxylate moiety of brequinar, one of the most potent known hDHODH inhibitors, is presented here. Their general structure is characterized by a biphenyl moiety joined through an amide bridge with an acidic hydroxyazole scaffold (hydroxylated thiadiazole, pyrazole and triazole). Molecular modelling suggested that these structures should adopt a brequinar-like binding mode involving interactions with subsites 1, 2 and 4 of the hDHODH binding site. Initially, the inhibitory activity of the compounds was studied on recombinant hDHODH. The most potent compound of the series in the enzymatic assays was the thiadiazole analogue 4 (IC5016 nM). The activity was found to be dependent on the fluoro substitution pattern at the biphenyl moiety as well as on the choice/substitution of the heterocyclic ring. Structure determination of hDHODH co-crystallized with one representative compound from each series (4, 5 and 6) confirmed the brequinar-like binding mode as suggested by modelling. The specificity of the observed effects of the compound series was tested in cell-based assays for antiproliferation activity using Jurkat cells and PHA-stimulated PBMC. These tests were also verified by addition of exogenous uridine to the culture medium. In particular, the triazole analogue 6 (IC50against hDHODH: 45 nM) exerted potent in vitro antiproliferative and immunosuppressive activity without affecting cell survival.
authors.affiliation: Department of Science and Drug Technology, University of Torino, via Pietro Giuria 9, 10125 Torino, Italy.
authors.forename: Stefano
authors.initials: S
authors.lastname: Sainas
authors.affiliation: Department of Science and Drug Technology, University of Torino, via Pietro Giuria 9, 10125 Torino, Italy.
authors.forename: Agnese C
authors.initials: AC
authors.lastname: Pippione
authors.affiliation: Department of Science and Drug Technology, University of Torino, via Pietro Giuria 9, 10125 Torino, Italy.
authors.forename: Marta
authors.initials: M
authors.lastname: Giorgis
authors.affiliation: Department of Oncology, University of Torino, via Michelangelo 27/B, 10126 Torino, Italy.
authors.forename: Elisa
authors.initials: E
authors.lastname: Lupino
authors.affiliation: Department of Chemistry and Molecular Biology, University of Gothenburg, Box 462, S-40530 Gothenburg Sweden.
authors.forename: Parveen
authors.initials: P
authors.lastname: Goyal
authors.affiliation: Department of Oncology, University of Torino, via Michelangelo 27/B, 10126 Torino, Italy.
authors.forename: Cristina
authors.initials: C
authors.lastname: Ramondetti
authors.affiliation: Department of Oncology, University of Torino, via Michelangelo 27/B, 10126 Torino, Italy.
authors.forename: Barbara
authors.initials: B
authors.lastname: Buccinnà
authors.affiliation: Department of Oncology, University of Torino, via Michelangelo 27/B, 10126 Torino, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Piccinini
authors.affiliation: LabMol, Faculty of Pharmacy, Federal University of Goias, 74605-170 Goiania, Brazil.
authors.forename: Rodolpho C
authors.initials: RC
authors.lastname: Braga
authors.affiliation: LabMol, Faculty of Pharmacy, Federal University of Goias, 74605-170 Goiania, Brazil.
authors.forename: Carolina H
authors.initials: CH
authors.lastname: Andrade
authors.affiliation: Department of Chemistry and Molecular Biology, University of Gothenburg, Box 462, S-40530 Gothenburg Sweden.
authors.forename: Mikael
authors.initials: M
authors.lastname: Andersson
authors.affiliation: Department of Chemistry and Molecular Biology, University of Gothenburg, Box 462, S-40530 Gothenburg Sweden; Department of Chemistry, Biochemistry, University of Bielefeld, Universitätsstraße 25, 33615 Bielefeld, Germany.
authors.forename: Ann-Christin
authors.initials: AC
authors.lastname: Moritzer
authors.affiliation: Department of Chemistry and Molecular Biology, University of Gothenburg, Box 462, S-40530 Gothenburg Sweden; Department of Structural Biology, Stanford University School of Medicine, 299 Campus Drive West Stanford, CA 94305-5126, USA.
authors.forename: Rosmarie
authors.initials: R
authors.lastname: Friemann
authors.affiliation: Stephenson Institute for Renewable Energy, Department of Chemistry, University of Liverpool, L69 7ZF Liverpool, UK.
authors.forename: Stefano
authors.initials: S
authors.lastname: Mensa
authors.affiliation: Department of Biochemistry and Structural Biology, Lund University, Sweden.
authors.forename: Salam
authors.initials: S
authors.lastname: Al-Kadaraghi
authors.affiliation: Department of Science and Drug Technology, University of Torino, via Pietro Giuria 9, 10125 Torino, Italy.
authors.forename: Donatella
authors.initials: D
authors.lastname: Boschi
authors.affiliation: Department of Science and Drug Technology, University of Torino, via Pietro Giuria 9, 10125 Torino, Italy. Electronic address: marco.lolli@unito.it.
authors.forename: Marco L
authors.initials: ML
authors.lastname: Lolli
mesh_terms: D001393:Azoles / Q000737:chemistry*; D001665:Binding Sites; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D000090722:Dihydroorotate Dehydrogenase; D015195:Drug Design; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000494:pharmacology; D006801:Humans; D006900:Hydroxylation; D007165:Immunosuppression Therapy; D019169:Jurkat Cells; D008958:Models, Molecular; D015394:Molecular Structure; D044925:Oxidoreductases Acting on CH-CH Group Donors / Q000037:antagonists & inhibitors*; D013329:Structure-Activity Relationship; D014961:X-Ray Diffraction
chemical_list: D001393:Azoles; D000090722:Dihydroorotate Dehydrogenase; D004791:Enzyme Inhibitors; D044925:Oxidoreductases Acting on CH-CH Group Donors
keywords: Autoimmune diseases; Bioisosterism; Brequinar; Dihydroorotate dehydrogenase (DHODH) inhibitors; Leflunomide; X-ray-crystallography