title: Constitutive activation of the ATM/BRCA1 pathway prevents DNA damage-induced apoptosis in 5-azacytidine-resistant cell lines.
abstract: 5-Azacytidine (AZA) exerts its anti-tumor effects by exerting cytotoxicity via its incorporation into RNA and DNA, which causes the reactivation of aberrantly silenced growth-regulatory genes by promoter demethylation, as well as DNA damage. AZA is used for patients with myelodysplastic syndrome and acute myeloid leukemia. However, some patients demonstrate resistance to AZA, the mechanisms of which are not fully elucidated. We therefore sought to better characterize the molecular mechanism of AZA resistance using an in vitro model of AZA resistance. We established AZA-resistant cell lines by exposing the human leukemia cell lines U937 and HL-60 to clinical concentrations of AZA, and characterized these cells. AZA-resistant cells showed a down-regulation of the DNMT3A protein, in correlation with their marked genome-wide DNA hypomethylation. Furthermore, genes involved in pyrimidine metabolism were down-regulated in both AZA-resistant cell lines; AZA sensitivity was restored by inhibition of CTP synthase. Of note is that the DNA damage response pathway is constitutively activated in the AZA-resistant cell lines, but not in the parental cell lines. Inhibition of the DNA damage response pathway canceled the AZA resistance, in association with an increase in apoptotic cells. We found that the molecular mechanism underlying AZA resistance involves pyrimidine metabolism and the DNA damage response through ATM kinase. This study therefore sheds light on the mechanisms underlying AZA resistance, and will enable better understanding of AZA resistance in patients undergoing AZA treatment.
authors.affiliation: Institute for Medical Science, Tokyo Medical University, 6-7-1 Nishi-shinjuku, Shinjuku, Tokyo 160-0023, Japan. Electronic address: s-ima@tokyo-med.ac.jp.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Imanishi
authors.affiliation: Institute for Medical Science, Tokyo Medical University, 6-7-1 Nishi-shinjuku, Shinjuku, Tokyo 160-0023, Japan; Department of Molecular Science, Tokyo Medical University, 6-7-1 Nishi-shinjuku, Shinjuku, Tokyo 160-0023, Japan.
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Umezu
authors.affiliation: Institute for Medical Science, Tokyo Medical University, 6-7-1 Nishi-shinjuku, Shinjuku, Tokyo 160-0023, Japan; First Department of Internal Medicine, Tokyo Medical University, 6-7-1 Nishi-shinjuku, Shinjuku, Tokyo 160-0023, Japan.
authors.forename: Kazushige
authors.initials: K
authors.lastname: Ohtsuki
authors.affiliation: First Department of Internal Medicine, Tokyo Medical University, 6-7-1 Nishi-shinjuku, Shinjuku, Tokyo 160-0023, Japan.
authors.forename: Chiaki
authors.initials: C
authors.lastname: Kobayashi
authors.affiliation: First Department of Internal Medicine, Tokyo Medical University, 6-7-1 Nishi-shinjuku, Shinjuku, Tokyo 160-0023, Japan.
authors.forename: Kazuma
authors.initials: K
authors.lastname: Ohyashiki
authors.affiliation: Institute for Medical Science, Tokyo Medical University, 6-7-1 Nishi-shinjuku, Shinjuku, Tokyo 160-0023, Japan.
authors.forename: Junko H
authors.initials: JH
authors.lastname: Ohyashiki
mesh_terms: D017209:Apoptosis / Q000502:physiology*; D064007:Ataxia Telangiectasia Mutated Proteins / Q000235:genetics / Q000378:metabolism*; D001374:Azacitidine / Q000494:pharmacology*; D019313:BRCA1 Protein / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics / Q000378:metabolism; D004249:DNA Damage / Q000502:physiology*; D015536:Down-Regulation; D019008:Drug Resistance, Neoplasm / Q000502:physiology*; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology; D006801:Humans; D011743:Pyrimidines / Q000378:metabolism
chemical_list: D019313:BRCA1 Protein; C492913:BRCA1 protein, human; D011743:Pyrimidines; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D001374:Azacitidine
keywords: 2-desoxy-5-azacytidine (PubChem CID: 451668); 3-deazauridine (PubChem CID: 3000824); 5-Azacytidine; 5-Azacytidine (PubChem CID: 9444); DNA damage; DNA demethylation; KU55933 (PubChem CID: 5278396); RG108 (PubChem CID: 702558); caffeine (PubChem CID: 2519); cisplatin (PubChem CID: 441203); etoposide (PubChem CID: 36462); mitomycin C (PubChem CID: 5746)