title: Gemcitabine reactivates epigenetically silenced genes and functions             as a DNA methyltransferase inhibitor.
abstract: Gemcitabine is indicated in combination with cisplatin as first-line therapy             for solid tumours including non-small cell lung cancer (NSCLC), bladder cancer             and mesothelioma. Gemcitabine is an analogue of pyrimidine cytosine and functions             as an anti-metabolite. Structurally, however, gemcitabine has similarities to             5-aza-2-deoxycytidine (decitabine/Dacogen®), a DNA methyltransferase inhibitor             (DNMTi). NSCLC, mesothelioma and prostate cancer cell lines were treated with             decitabine and gemcitabine. Reactivation of epigenetically silenced genes was             examined by RT-PCR/qPCR. DNA methyltransferase activity in nuclear extracts and             recombinant proteins was measured using a DNA methyl-transferase assay, and alterations             in DNA methylation status were examined using methylation-specific PCR (MS-PCR)             and pyrosequencing. We observe a reactivation of several epigenetically silenced             genes including GSTP1, IGFBP3 and RASSF1A. Gemcitabine functionally inhibited             DNA methyltransferase activity in both nuclear extracts and recombinant proteins.             Gemcitabine dramatically destabilised DNMT1 protein. However, DNA CpG methylation             was for the most part unaffected by gemcitabine. In conclusion, gemcitabine both             inhibits and destabilises DNA methyltransferases and reactivates epigenetically             silenced genes having activity equivalent to decitabine at concentrations significantly             lower than those achieved in the treatment of patients with solid tumours. This             property may contribute to the anticancer activity of gemcitabine.
authors.affiliation: Thoracic Oncology Research Group, Institute of Molecular Medicine, Trinity College Dublin, Dublin 2, Republic of Ireland. sgray@stjames.ie
authors.forename: Steven G
authors.initials: SG
authors.lastname: Gray
authors.forename: Anne-Marie
authors.initials: AM
authors.lastname: Baird
authors.forename: Fardod
authors.initials: F
authors.lastname: O'Kelly
authors.forename: Georgios
authors.initials: G
authors.lastname: Nikolaidis
authors.forename: Malin
authors.initials: M
authors.lastname: Almgren
authors.forename: Armelle
authors.initials: A
authors.lastname: Meunier
authors.forename: Eilis
authors.initials: E
authors.lastname: Dockry
authors.forename: Donal
authors.initials: D
authors.lastname: Hollywood
authors.forename: Tomas J
authors.initials: TJ
authors.lastname: Ekström
authors.forename: Antoinette S
authors.initials: AS
authors.lastname: Perry
authors.forename: Kenneth J
authors.initials: KJ
authors.lastname: O'Byrne
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D045744:Cell Line, Tumor; D018899:CpG Islands; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D019175:DNA Methylation; D000077209:Decitabine; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D004795:Enzyme Stability; D044127:Epigenesis, Genetic; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D020868:Gene Silencing; D056284:Histone Deacetylase 1 / Q000378:metabolism; D006801:Humans; D011401:Promoter Regions, Genetic; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism; D040281:Vascular Endothelial Growth Factor Receptor-1 / Q000235:genetics / Q000378:metabolism; D000093542:Gemcitabine
chemical_list: D000964:Antimetabolites, Antineoplastic; C411762:RASSF1 protein, human; D025521:Tumor Suppressor Proteins; D003841:Deoxycytidine; D000077209:Decitabine; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; C000619380:DNMT1 protein, human; D040281:Vascular Endothelial Growth Factor Receptor-1; C110550:HDAC1 protein, human; D056284:Histone Deacetylase 1; D001374:Azacitidine; D000093542:Gemcitabine