title: siRNA knockdown of mitochondrial thymidine kinase 2 (TK2) sensitizes human tumor cells to gemcitabine.
abstract: Nucleoside metabolism enzymes are determinants of chemotherapeutic drug activity. The nucleoside salvage enzyme deoxycytidine kinase (dCK) activates gemcitabine (2', 2'-difluoro-2'-deoxycytidine) and is negatively regulated by deoxycytidine triphosphate (dCTP). Reduction of dCTP in tumor cells could, therefore, enhance gemcitabine activity. Mitochondrial thymidine kinase 2 (TK2) phosphorylates deoxycytidine to generate dCTP. We hypothesized that: (1) TK2 modulates human tumor cell sensitivity to gemcitabine, and (2) antisense knockdown of TK2 would decrease dCTP and increase dCK activity and gemcitabine activation. siRNA downregulation of TK2 sensitized MCF7 and HeLa cells (high and moderate TK2) but not A549 cells (low TK2) to gemcitabine. Combined treatment with TK2 siRNA and gemcitabine increased dCK. We also hypothesized that TK2 siRNA-induced drug sensitization results in mitochondrial damage that enhances gemcitabine effectiveness. TK2 siRNA and gemcitabine decreased mitochondrial redox status, DNA content, and activity. This is the first demonstration of a direct role for TK2 in gemcitabine resistance, or any independent role in cancer drug resistance, and further distinguishes TK2 function from that of other dTMP-producing enzymes [cytosolic TK1 and thymidylate synthase (TS)]. siRNA knockdown of TK1 and/or TS did not sensitize cancer cells to gemcitabine indicating that, among the 3 enzymes, only TK2 is a candidate therapeutic target for combination with gemcitabine.
authors.affiliation: Department of Microbiology and Immunology, The University of Western Ontario, London, Ontario, Canada.|Cancer Research Laboratory Program, Lawson Health Research Institute and London Regional Cancer Program, London, Ontario, Canada.
authors.forename: Christine
authors.initials: C
authors.lastname: Di Cresce
authors.affiliation: Department of Oncology, The University of Western Ontario, London, Ontario, Canada.|Cancer Research Laboratory Program, Lawson Health Research Institute and London Regional Cancer Program, London, Ontario, Canada.
authors.forename: Rene
authors.initials: R
authors.lastname: Figueredo
authors.affiliation: Department of Microbiology and Immunology, The University of Western Ontario, London, Ontario, Canada.|Cancer Research Laboratory Program, Lawson Health Research Institute and London Regional Cancer Program, London, Ontario, Canada.
authors.forename: Mateusz
authors.initials: M
authors.lastname: Rytelewski
authors.affiliation: Department of Microbiology and Immunology, The University of Western Ontario, London, Ontario, Canada.|Cancer Research Laboratory Program, Lawson Health Research Institute and London Regional Cancer Program, London, Ontario, Canada.
authors.forename: Saman
authors.initials: S
authors.lastname: Maleki Vareki
authors.affiliation: Department of Microbiology and Immunology, The University of Western Ontario, London, Ontario, Canada.|Cancer Research Laboratory Program, Lawson Health Research Institute and London Regional Cancer Program, London, Ontario, Canada.
authors.forename: Colin
authors.initials: C
authors.lastname: Way
authors.affiliation: Department of Oncology, The University of Western Ontario, London, Ontario, Canada.|Cancer Research Laboratory Program, Lawson Health Research Institute and London Regional Cancer Program, London, Ontario, Canada.
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Ferguson
authors.affiliation: Department of Oncology, The University of Western Ontario, London, Ontario, Canada.|Cancer Research Laboratory Program, Lawson Health Research Institute and London Regional Cancer Program, London, Ontario, Canada.
authors.forename: Mark D
authors.initials: MD
authors.lastname: Vincent
authors.affiliation: Department of Microbiology and Immunology, The University of Western Ontario, London, Ontario, Canada.|Department of Oncology, The University of Western Ontario, London, Ontario, Canada.|Department of Pathology, The University of Western Ontario, London, Ontario, Canada.|Department of Physiology and Pharmacology, The University of Western Ontario, London, Ontario, Canada.|Cancer Research Laboratory Program, Lawson Health Research Institute and London Regional Cancer Program, London, Ontario, Canada.
authors.forename: James
authors.initials: J
authors.lastname: Koropatnick
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D045744:Cell Line, Tumor; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D003842:Deoxycytidine Kinase / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D061986:MCF-7 Cells; D008928:Mitochondria / Q000201:enzymology; D009369:Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000628:therapy*; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000235:genetics; D013937:Thymidine Kinase / Q000235:genetics* / Q000378:metabolism*; D014162:Transfection; D000093542:Gemcitabine
chemical_list: D000964:Antimetabolites, Antineoplastic; D034741:RNA, Small Interfering; D003841:Deoxycytidine; C105339:thymidine kinase 2; D013937:Thymidine Kinase; D003842:Deoxycytidine Kinase; D000093542:Gemcitabine
keywords: cancer; deoxycytidine kinase; gemcitabine; mitochondria; thymidine kinase