title: Inhibition of the mitochondrial pyrimidine biosynthesis enzyme dihydroorotate dehydrogenase by doxorubicin and brequinar sensitizes cancer cells to TRAIL-induced apoptosis.
abstract: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising agent in selectively killing tumor cells. However, TRAIL monotherapy has not been successful as many cancer cells are resistant to TRAIL. Chemotherapeutic agents, such as doxorubicin have been shown to act synergistically with TRAIL, but the exact mechanisms of actions are poorly understood. In this study, we performed high-throughput small interfering RNA screening and genome-wide gene expression profiling on doxorubicin-treated U1690 cells to explore novel mechanisms underlying doxorubicin-TRAIL synergy. The screening and expression profiling results were integrated and dihydroorotate dehydrogenase (DHODH) was identified as a potential candidate. DHODH is the rate-limiting enzyme in the pyrimidine synthesis pathway, and its expression was downregulated by doxorubicin. We demonstrated that silencing of DHODH or inhibition of DHODH activity by brequinar dramatically increased the sensitivity of U1690 cells to TRAIL-induced apoptosis both in 2D and 3D cultures, and was accompanied by downregulation of c-FLIPL as well as by mitochondrial depolarization. In addition, uridine, an end product of the pyrimidine synthesis pathway was able to rescue the sensitization effects initiated by both brequinar and doxorubicin. Furthermore, several other cancer cell lines, LNCaP, MCF-7 and HT-29 were also shown to be sensitized to TRAIL by brequinar. Taken together, our findings have identified a novel protein target and its inhibitor, brequinar, as a potential agent in TRAIL-based combinatorial cancer therapy and highlighted for the first time the importance of mitochondrial DHODH enzyme and pyrimidine pathway in mediating TRAIL sensitization in cancer cells.
authors.affiliation: Medical Biotechnology, VTT Technical Research Centre of Finland, Turku, Finland.
authors.forename: T
authors.initials: T
authors.lastname: He
authors.affiliation: Medical Biotechnology, VTT Technical Research Centre of Finland, Turku, Finland.
authors.forename: S
authors.initials: S
authors.lastname: Haapa-Paananen
authors.affiliation: Division of Toxicology, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden.
authors.forename: V O
authors.initials: VO
authors.lastname: Kaminskyy
authors.affiliation: 1] Medical Biotechnology, VTT Technical Research Centre of Finland, Turku, Finland [2] Division of Molecular Toxicology, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden.
authors.forename: P
authors.initials: P
authors.lastname: Kohonen
authors.affiliation: Medical Biotechnology, VTT Technical Research Centre of Finland, Turku, Finland.
authors.forename: V
authors.initials: V
authors.lastname: Fey
authors.affiliation: Division of Toxicology, Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden.
authors.forename: B
authors.initials: B
authors.lastname: Zhivotovsky
authors.affiliation: 1] Medical Biotechnology, VTT Technical Research Centre of Finland, Turku, Finland [2] FIMM-Institute for Molecular Medicine Finland, University of Helsinki, Helsinki, Finland.
authors.forename: O
authors.initials: O
authors.lastname: Kallioniemi
authors.affiliation: Medical Biotechnology, VTT Technical Research Centre of Finland, Turku, Finland.
authors.forename: M
authors.initials: M
authors.lastname: Perälä
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D001713:Biphenyl Compounds / Q000494:pharmacology*; D045744:Cell Line, Tumor; D019295:Computational Biology; D000090722:Dihydroorotate Dehydrogenase; D004305:Dose-Response Relationship, Drug; D004317:Doxorubicin / Q000494:pharmacology*; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D020869:Gene Expression Profiling; D020868:Gene Silencing; D006801:Humans; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism*; D044925:Oxidoreductases Acting on CH-CH Group Donors / Q000037:antagonists & inhibitors* / Q000172:deficiency / Q000235:genetics; D011743:Pyrimidines / Q000096:biosynthesis*; D034741:RNA, Small Interfering / Q000235:genetics; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000378:metabolism*; D014529:Uridine / Q000494:pharmacology; D011798:Quinaldines
chemical_list: D000970:Antineoplastic Agents; D001713:Biphenyl Compounds; D000090722:Dihydroorotate Dehydrogenase; D004317:Doxorubicin; D044925:Oxidoreductases Acting on CH-CH Group Donors; D011743:Pyrimidines; D034741:RNA, Small Interfering; D053221:TNF-Related Apoptosis-Inducing Ligand; D014529:Uridine; C046943:brequinar; D011798:Quinaldines; C030986:pyrimidine