title: DCK is frequently inactivated in acquired gemcitabine-resistant human cancer cells.
abstract: Although gemcitabine is the most effective chemotherapeutic agent against pancreatic cancer, a growing concern is that a substantial number of patients acquire gemcitabine chemoresistance. To elucidate the mechanisms of acquisition of gemcitabine resistance, we developed gemcitabine-resistant cell lines from six human cancer cell lines; three pancreatic, one gastric, one colon, and one bile duct cancer. We first analyzed gemcitabine uptake using three paired parental and gemcitabine resistant pancreatic cancer cell lines (PK-1 and RPK-1, PK-9 and RPK-9, PK-59 and RPK-59) and found that uptake of gemcitabine was rapid. However, no DNA damage was induced in resistant cells. We further examined the microarray-based expression profiles of the cells to identify genes associated with gemcitabine resistance and found a remarkable reduction in the expression of deoxycytidine kinase (DCK). DCK is a key enzyme that activates gemcitabine by phosphorylation. Genetic alterations and expression of DCK were studied in these paired parental and derived gemcitabine-resistant cell lines, and inactivating mutations were found only in gemcitabine-resistant cell lines. Furthermore, siRNA-mediated knockdown of DCK in the parental cell lines yielded gemcitabine resistance, and introduction of DCK into gemcitabine-resistant cell lines invariably restored gemcitabine sensitivities. Mutation analyses were expanded to three other different paired cell lines, DLD-1 and RDLD-1 (colon cancer cell line), MKN-28 and RMKN-28 (gastric cancer cell line), and TFK-1 and RTFK -1 (cholangiocarcinoma cell line). We found inactivating mutations in RDLD-1 and RTFK-1 and decreased expression of DCK in RMKN-28. These results indicate that the inactivation of DCK is one of the crucial mechanisms in acquisition of gemcitabine resistance.
authors.affiliation: Department of Molecular Pathology, Tohoku University Graduate School of Medicine, Sendai, Japan.
authors.forename: Yuriko
authors.initials: Y
authors.lastname: Saiki
authors.forename: Yuki
authors.initials: Y
authors.lastname: Yoshino
authors.forename: Hiroko
authors.initials: H
authors.lastname: Fujimura
authors.forename: Tatsuya
authors.initials: T
authors.lastname: Manabe
authors.forename: Yuki
authors.initials: Y
authors.lastname: Kudo
authors.forename: Miki
authors.initials: M
authors.lastname: Shimada
authors.forename: Nariyasu
authors.initials: N
authors.lastname: Mano
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Nakano
authors.forename: Yoonha
authors.initials: Y
authors.lastname: Lee
authors.forename: Shinjiro
authors.initials: S
authors.lastname: Shimizu
authors.forename: Shinya
authors.initials: S
authors.lastname: Oba
authors.forename: Sho
authors.initials: S
authors.lastname: Fujiwara
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Shimizu
authors.forename: Na
authors.initials: N
authors.lastname: Chen
authors.forename: Zhaleh Kashkouli
authors.initials: ZK
authors.lastname: Nezhad
authors.forename: Guo
authors.initials: G
authors.lastname: Jin
authors.forename: Shinichi
authors.initials: S
authors.lastname: Fukushige
authors.forename: Makoto
authors.initials: M
authors.lastname: Sunamura
authors.forename: Masaharu
authors.initials: M
authors.lastname: Ishida
authors.forename: Fuyuhiko
authors.initials: F
authors.lastname: Motoi
authors.forename: Shinichi
authors.initials: S
authors.lastname: Egawa
authors.forename: Michiaki
authors.initials: M
authors.lastname: Unno
authors.forename: Akira
authors.initials: A
authors.lastname: Horii
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000493:pharmacokinetics / Q000494:pharmacology*; D045744:Cell Line, Tumor; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000493:pharmacokinetics / Q000494:pharmacology; D003842:Deoxycytidine Kinase / Q000235:genetics / Q000378:metabolism*; D019008:Drug Resistance, Neoplasm; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D020868:Gene Silencing; D006657:Histones / Q000378:metabolism; D006801:Humans; D010190:Pancreatic Neoplasms / Q000201:enzymology*; D010766:Phosphorylation; D034741:RNA, Small Interfering / Q000235:genetics; D000093542:Gemcitabine
chemical_list: D000964:Antimetabolites, Antineoplastic; C468783:H2AX protein, human; D006657:Histones; D034741:RNA, Small Interfering; D003841:Deoxycytidine; D003842:Deoxycytidine Kinase; D000093542:Gemcitabine