title: Mechanisms operative in the antitumor activity of temozolomide in glioblastoma multiforme.
abstract: OBJECTIVE
  Glioblastoma multiforme (GBM) is the most frequent and incurable brain tumor in adults. Although temozolomide (TMZ) does not cure GBM, it has demonstrated anti-GBM activity and has improved survival (8-14 months) and quality of life. We studied the mechanisms by which TMZ affects 2 human GBM cell lines; U251-MG and U87-MG, aiming to unravel the drug-activated cascades to enable the development of combination therapies that will improve the efficacy of TMZ.
  
  
  METHODS
  The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium assay was used to assess cell viability. Modulation of gene expression by TMZ therapy was assayed by gene profiling and verified by quantitative real-time polymerase chain reaction. Protein levels influenced by the treatment were studied by Western blots and immunocytochemistry.
  
  
  RESULTS
  Increasing concentrations of TMZ decreased cell viability in a concentration-dependent manner. The expression of 1,886 genes was altered >2-fold after TMZ treatment. We focused on the 81 genes similarly altered by TMZ treatment in both cell lines to neutralize tissue-specific characteristics. Fourteen target genes of hypoxia-inducible factor (HIF-1), were found to be up-regulated after TMZ treatment including vascular endothelial growth factor (VEGF). HIF-1alpha expression was constant at the mRNA level; however, its post-treatment protein levels increased compared with those of untreated control cells.
  
  
  CONCLUSIONS
  The genetic analyses suggest that treatment with TMZ activates stress mechanisms in GBM cells that include the angiogenesis-inducing proteins HIF-1alpha and VEGF. We propose that treatment with TMZ be supplemented with either an antibody to VEGF or down-regulators of HIF-1alpha to improve clinical results of TMZ in the treatment of GBM.
authors.affiliation: Department of Pediatric Hemato-Oncology, Safra Children's Hospital and the Institute of Hematology, Tel-Hashomer, Israel. tamar.fisher@sheba.health.gov.il
authors.forename: Tamar
authors.initials: T
authors.lastname: Fisher
authors.forename: Gil
authors.initials: G
authors.lastname: Galanti
authors.forename: Gad
authors.initials: G
authors.lastname: Lavie
authors.forename: Jasmine
authors.initials: J
authors.lastname: Jacob-Hirsch
authors.forename: Iris
authors.initials: I
authors.lastname: Kventsel
authors.forename: Sharon
authors.initials: S
authors.lastname: Zeligson
authors.forename: Robert
authors.initials: R
authors.lastname: Winkler
authors.forename: Amos J
authors.initials: AJ
authors.lastname: Simon
authors.forename: Ninette
authors.initials: N
authors.lastname: Amariglio
authors.forename: Gideon
authors.initials: G
authors.lastname: Rechavi
authors.forename: Amos
authors.initials: A
authors.lastname: Toren
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D001932:Brain Neoplasms / Q000378:metabolism / Q000473:pathology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D003606:Dacarbazine / Q000031:analogs & derivatives* / Q000494:pharmacology; D020869:Gene Expression Profiling; D005909:Glioblastoma / Q000378:metabolism / Q000473:pathology*; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000096:biosynthesis; D007150:Immunohistochemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000077204:Temozolomide
chemical_list: D000970:Antineoplastic Agents; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D003606:Dacarbazine; D000077204:Temozolomide