title: The DNA repair protein ALKBH2 mediates temozolomide resistance in human glioblastoma cells.
abstract: BACKGROUND
  Glioblastoma multiforme (GBM; World Health Organization astrocytoma grade IV) is the most frequent and most malignant primary brain tumor in adults. Despite multimodal therapy, all such tumors practically recur during the course of therapy, causing a median survival of only 14.6 months in patients with newly diagnosed GBM. The present study was aimed at examining the expression of the DNA repair protein AlkB homolog 2 (ALKBH2) in human GBM and determining whether it could promote resistance to temozolomide chemotherapy.
  
  
  METHODS
  ALKBH2 expression in GBM cell lines and in human GBM was determined by quantitative real-time PCR (qRT-PCR) and gene expression analysis, respectively. Drug sensitivity was assessed in GBM cells overexpressing ALKBH2 and in cells in which ALKBH2 expression was silenced by small-interfering (si)RNA. ALKBH2 expression following activation of the p53 pathway was examined by western blotting and qRT-PCR.
  
  
  RESULTS
  ALKBH2 was abundantly expressed in established GBM cell lines and human GBM, and temozolomide exposure increased cellular ALKBH2 expression levels. Overexpression of ALKBH2 in the U87 and U251 GBM cell lines enhanced resistance to the methylating agents temozolomide and methyl methanesulfonate but not to the nonmethylating agent doxorubicin. Conversely, siRNA-mediated knockdown of ALKBH2 increased sensitivity of GBM cells to temozolomide and methyl methanesulfonate but not to doxorubicin or cisplatin. Nongenotoxic activation of the p53 pathway by the selective murine double minute 2 antagonist nutlin-3 caused a significant decrease in cellular ALKBH2 transcription levels.
  
  
  CONCLUSIONS
  Our findings identify ALKBH2 as a novel mediator of temozolomide resistance in human GBM cells. Furthermore, we place ALKBH2 into a new cellular context by showing its regulation by the p53 pathway.
authors.affiliation: Department of Biomedicine, University of Bergen, Jonas Lies vei 91, N-5009, Bergen, Norway. tor-christian.johannessen@biomed.uib.no
authors.forename: Tor-Christian Aase
authors.initials: TC
authors.lastname: Johannessen
authors.forename: Lars
authors.initials: L
authors.lastname: Prestegarden
authors.forename: Amra
authors.initials: A
authors.lastname: Grudic
authors.forename: Monika E
authors.initials: ME
authors.lastname: Hegi
authors.forename: Berit Bølge
authors.initials: BB
authors.lastname: Tysnes
authors.forename: Rolf
authors.initials: R
authors.lastname: Bjerkvig
mesh_terms: D000071499:AlkB Homolog 2, Alpha-Ketoglutarate-Dependent Dioxygenase; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology*; D015153:Blotting, Western; D001932:Brain Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D002448:Cell Adhesion / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D045643:DNA Repair Enzymes / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D003606:Dacarbazine / Q000031:analogs & derivatives* / Q000494:pharmacology; D049308:Dioxygenases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D019008:Drug Resistance, Neoplasm; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D005909:Glioblastoma / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D006801:Humans; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000077204:Temozolomide; D014407:Tumor Cells, Cultured; D014410:Tumor Stem Cell Assay
chemical_list: D018906:Antineoplastic Agents, Alkylating; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D003606:Dacarbazine; D049308:Dioxygenases; C486440:ALKBH2 protein, human; D000071499:AlkB Homolog 2, Alpha-Ketoglutarate-Dependent Dioxygenase; D045643:DNA Repair Enzymes; D000077204:Temozolomide