title: Fluorescence assay of dihydroorotate dehydrogenase that may become a cancer biomarker.
abstract: We developed an assay method for measuring dihydroorotate dehydrogenase (DHODH) activity in cultured HeLa cells and fibroblasts, and in stage III stomach cancer and adjacent normal tissues from the same patient. The assay comprised enzymatic reaction of DHODH with a large amount of dihydroorotic acid substrate, followed by fluorescence (FL) detection specific for orotic acid using the 4-trifluoromethyl-benzamidoxime fluorogenic reagent. The DHODH activities in the biologically complex samples were readily measured by the assay method. Our data indicate significantly higher DHODH activity in HeLa cells (340 ± 25.9 pmol/105cells/h) than in normal fibroblasts (54.1 ± 7.40 pmol/105cells/h), and in malignant tumour tissue (1.10 ± 0.19 nmol/mg total proteins/h) than in adjacent normal tissue (0.24 ± 0.11 nmol/mg total proteins/h). This is the first report that DHODH activity may be a diagnostic biomarker for cancer.
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14 Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Sheng
authors.initials: S
authors.lastname: Yin
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14 Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Kabashima
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14 Bunkyo-Machi, Nagasaki 852-8521, Japan.|Department of Pharmacy, School of Medicine, Shenzhen University, 3688 Nanhai Boulevard, Shenzhen, Guangdong 518060, China.
authors.forename: Qinchang
authors.initials: Q
authors.lastname: Zhu
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14 Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Takayuki
authors.initials: T
authors.lastname: Shibata
authors.affiliation: Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14 Bunkyo-Machi, Nagasaki 852-8521, Japan.
authors.forename: Masaaki
authors.initials: M
authors.lastname: Kai
mesh_terms: D001681:Biological Assay; D014408:Biomarkers, Tumor; D002384:Catalysis; D000090722:Dihydroorotate Dehydrogenase; D004789:Enzyme Activation; D057075:Enzyme Assays; D005347:Fibroblasts; D005453:Fluorescence; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D044925:Oxidoreductases Acting on CH-CH Group Donors / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D014408:Biomarkers, Tumor; D000090722:Dihydroorotate Dehydrogenase; D044925:Oxidoreductases Acting on CH-CH Group Donors