title: Mechanisms of resistance to azacitidine in human leukemia cell lines.
abstract: The DNA methylation inhibitor azacitidine (5-azacytidine) is used against myelodysplastic syndrome and acute myeloid leukemia, but drug resistance is an ongoing, intractable problem. To investigate resistance mechanisms, we generated two azacitidine-resistant cell lines, THP-1/AR and HL60/AR, and studied genetic disparities between them and their corresponding parental lines. In cells treated with azacitidine, significant mitotic variations were noted in parental cells which were absent in resistant cells, suggesting that resistance arises from negating azacitidine-mediated activation of apoptosis signaling and reestablishing G2/M checkpoint. Importantly, both resistant cell lines have common point mutations in the uridine-cytidine kinase 2 (UCK2) gene, which encodes the rate-limiting enzyme of the azacitidine activation pathway. Forced expression of mutated UCK2 in parental THP-1 cells abrogated azacitidine-induced apoptosis, whereas overexpression of wild type UCK2 in resistant THP-1/AR cells restored sensitivity to azacitidine, implying that UCK2 gene mutations perturb azacitidine activation and advance azacitidine resistance. Our study provides new insights into azacitidine resistance and establishes models useful in developing effective strategies to overcome it.
authors.affiliation: Division of Hematology, Jichi Medical University, Tochigi, Japan.
authors.forename: Piyanuch
authors.initials: P
authors.lastname: Sripayap
authors.affiliation: Division of Hematology, Jichi Medical University, Tochigi, Japan. Electronic address: t-nagai@jichi.ac.jp.
authors.forename: Tadashi
authors.initials: T
authors.lastname: Nagai
authors.affiliation: Division of Hematology, Jichi Medical University, Tochigi, Japan.
authors.forename: Mitsuyo
authors.initials: M
authors.lastname: Uesawa
authors.affiliation: Division of Hematology, Jichi Medical University, Tochigi, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Kobayashi
authors.affiliation: Division of Genetic Therapeutics, Center for Molecular Medicine, Jichi Medical University, Tochigi, Japan.
authors.forename: Tomonori
authors.initials: T
authors.lastname: Tsukahara
authors.affiliation: Division of Hematology, Jichi Medical University, Tochigi, Japan.
authors.forename: Ken
authors.initials: K
authors.lastname: Ohmine
authors.affiliation: Division of Cell Transplantation and Transfusion, Jichi Medical University, Tochigi, Japan.
authors.forename: Kazuo
authors.initials: K
authors.lastname: Muroi
authors.affiliation: Division of Hematology, Jichi Medical University, Tochigi, Japan.
authors.forename: Keiya
authors.initials: K
authors.lastname: Ozawa
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D001374:Azacitidine / Q000494:pharmacology*; D002455:Cell Division / Q000187:drug effects* / Q000235:genetics; D019008:Drug Resistance, Neoplasm / Q000187:drug effects* / Q000235:genetics; D016195:G2 Phase / Q000187:drug effects* / Q000235:genetics; D018922:HL-60 Cells; D006801:Humans; D007951:Leukemia, Myeloid; D017354:Point Mutation; D014541:Uridine Kinase / Q000235:genetics / Q000378:metabolism*
chemical_list: D000964:Antimetabolites, Antineoplastic; C483834:UCK2 protein, human; D014541:Uridine Kinase; D001374:Azacitidine