title: Consequences of combining siRNA-mediated DNA methyltransferase 1 depletion with 5-aza-2'-deoxycytidine in human leukemic KG1 cells.
abstract: 5-azacytidine and 5-aza-2'-deoxycytidine are clinically used to treat patients with blood neoplasia. Their antileukemic property is mediated by the trapping and the subsequent degradation of a family of proteins, the DNA methyltransferases (DNMT1, DNMT3A, and DNMT3B) leading to DNA demethylation, tumor suppressor gene re-expression and DNA damage. Here we studied the respective role of each DNMT in the human leukemia KG1 cell line using a RNA interference approach. In addition we addressed the role of DNA damage formation in DNA demethylation by 5-aza-2'-deoxycytidine. Our data show that DNMT1 is the main DNMT involved in DNA methylation maintenance in KG1 cells and in mediating DNA damage formation upon exposure to 5-aza-2'-deoxycytidine. Moreover, KG1 cells express the DNMT1 protein at a level above the one required to ensure DNA methylation maintenance, and we identified a threshold for DNMT1 depletion that needs to be exceeded to achieve DNA demethylation. Most interestingly, by combining DNMT1 siRNA and treatment with low dose of 5-aza-2'-deoxycytidine, it is possible to uncouple DNA damage formation from DNA demethylation. This work strongly suggests that a direct pharmacological inhibition of DNMT1, unlike the use of 5-aza-2'-deoxycytidine, should lead to tumor suppressor gene hypomethylation and re-expression without inducing major DNA damage in leukemia.
authors.affiliation: Unité de Service et de Recherche n°3388 CNRS-Pierre Fabre, ETaC Epigenetic Targeting of Cancer, CRDPF, Toulouse, France.
authors.forename: Stéphane
authors.initials: S
authors.lastname: Vispé
authors.affiliation: Unité de Service et de Recherche n°3388 CNRS-Pierre Fabre, ETaC Epigenetic Targeting of Cancer, CRDPF, Toulouse, France.
authors.forename: Arthur
authors.initials: A
authors.lastname: Deroide
authors.affiliation: Unité de Service et de Recherche n°3388 CNRS-Pierre Fabre, ETaC Epigenetic Targeting of Cancer, CRDPF, Toulouse, France.
authors.forename: Emeline
authors.initials: E
authors.lastname: Davoine
authors.affiliation: Unité de Service et de Recherche n°3388 CNRS-Pierre Fabre, ETaC Epigenetic Targeting of Cancer, CRDPF, Toulouse, France.
authors.forename: Cécile
authors.initials: C
authors.lastname: Desjobert
authors.affiliation: Molecular and Cellular Biology Department, Centre de Recherche Pierre Fabre, Castres, France.
authors.forename: Fabrice
authors.initials: F
authors.lastname: Lestienne
authors.affiliation: Unité de Service et de Recherche n°3388 CNRS-Pierre Fabre, ETaC Epigenetic Targeting of Cancer, CRDPF, Toulouse, France.
authors.forename: Lucie
authors.initials: L
authors.lastname: Fournier
authors.affiliation: Unité de Service et de Recherche n°3388 CNRS-Pierre Fabre, ETaC Epigenetic Targeting of Cancer, CRDPF, Toulouse, France.
authors.forename: Natacha
authors.initials: N
authors.lastname: Novosad
authors.affiliation: Informatique de Recherche (Bioinformatics and Statistics), Centre de Recherche Pierre Fabre, Castres, France.
authors.forename: Sophie
authors.initials: S
authors.lastname: Bréand
authors.affiliation: Informatique de Recherche (Bioinformatics and Statistics), Centre de Recherche Pierre Fabre, Castres, France.
authors.forename: Jérôme
authors.initials: J
authors.lastname: Besse
authors.affiliation: Laboratory for Epigenetics and Environment, Centre National de Génotypage, CEA-Institut de Génomique, Evry, France.
authors.forename: Florence
authors.initials: F
authors.lastname: Busato
authors.affiliation: Laboratory for Epigenetics and Environment, Centre National de Génotypage, CEA-Institut de Génomique, Evry, France.
authors.forename: Jörg
authors.initials: J
authors.lastname: Tost
authors.affiliation: Molecular and Cellular Biology Department, Centre de Recherche Pierre Fabre, Castres, France.
authors.forename: Luc
authors.initials: L
authors.lastname: De Vries
authors.affiliation: Molecular and Cellular Biology Department, Centre de Recherche Pierre Fabre, Castres, France.
authors.forename: Didier
authors.initials: D
authors.lastname: Cussac
authors.affiliation: Unité de Service et de Recherche n°3388 CNRS-Pierre Fabre, ETaC Epigenetic Targeting of Cancer, CRDPF, Toulouse, France.
authors.forename: Joëlle
authors.initials: J
authors.lastname: Riond
authors.affiliation: Unité de Service et de Recherche n°3388 CNRS-Pierre Fabre, ETaC Epigenetic Targeting of Cancer, CRDPF, Toulouse, France.
authors.forename: Paola B
authors.initials: PB
authors.lastname: Arimondo
mesh_terms: D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000235:genetics; D018899:CpG Islands / Q000235:genetics; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000037:antagonists & inhibitors* / Q000235:genetics*; D004249:DNA Damage / Q000235:genetics; D019175:DNA Methylation / Q000187:drug effects / Q000235:genetics*; D000090205:DNA Methyltransferase 3A; D004268:DNA-Binding Proteins / Q000235:genetics; D000077209:Decitabine; D059565:G2 Phase Cell Cycle Checkpoints / Q000187:drug effects / Q000235:genetics; D006657:Histones / Q000378:metabolism; D006801:Humans; D007938:Leukemia / Q000188:drug therapy*; D009687:Nuclear Proteins / Q000235:genetics; D010766:Phosphorylation; D011401:Promoter Regions, Genetic / Q000235:genetics; D034622:RNA Interference; D034741:RNA, Small Interfering; D000072160:Tumor Protein p73; D025521:Tumor Suppressor Proteins / Q000235:genetics; D000097584:DNA Methyltransferase 3B
chemical_list: D001374:Azacitidine; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; D000090205:DNA Methyltransferase 3A; D004268:DNA-Binding Proteins; D000077209:Decitabine; D006657:Histones; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D000072160:Tumor Protein p73; D025521:Tumor Suppressor Proteins; D000097584:DNA Methyltransferase 3B; C000717967:DNMT3A protein, human; C468783:H2AX protein, human; C000619380:DNMT1 protein, human
keywords: 5-aza-2’-deoxycytidine; DNA damage; DNA methylation; DNMT; leukemia