Abrogation of the Chk1-mediated G(2) checkpoint pathway potentiates temozolomide-induced toxicity in a p53-independent manner in human glioblastoma cells.
Temozolomide (TMZ) produces O(6)-methylguanine in DNA, which in turn mispairs with thymine, triggering futile DNA mismatch repair (MMR) and ultimately cell death. We found previously that in p53-proficient human glioma cells, TMZ-induced futile DNA MMR resulted not in apoptosis but rather in prolonged, p53- and p21-associated G(2)-M arrest and senescence. Additionally, p53-deficient cells were relatively more TMZ resistant than p53-deficient glioma cells, which underwent only transient G(2)-M arrest before death by mitotic catastrophe. These results suggested that prolonged G(2)-M arrest might protect cells from TMZ-induced cytotoxicity. In the present study, we therefore focused on the mechanism by which TMZ induces G(2)-M arrest and on whether inhibition of such G(2)-M arrest might sensitize glioma cells to TMZ-induced toxicity. U87MG glioma cells treated with TMZ underwent G(2)-M arrest associated with Chk1 activation and phosphorylation of both cdc25C and cdc2. These TMZ-induced effects were inhibited by the Chk1 kinase inhibitor UCN-01. Although not in itself toxic, UCN-01 increased the cytotoxicity of TMZ 5-fold, primarily by inhibiting cellular senescence and increasing the percentage of cells bypassing G(2)-M arrest and undergoing mitotic catastrophe. In addition to enhancing TMZ-induced cytotoxicity in p53-proficient cells, UCN-01 also blocked TMZ-induced Chk1 activation and transient G(2)-M arrest in p53-deficient U87MG-E6 cells and similarly enhanced TMZ-induced mitotic catastrophe and cell death. Taken together, these results indicate that Chk1 links TMZ-induced MMR to G(2)-M arrest. Furthermore, inhibition of the cytoprotective G(2) arrest pathway sensitizes cells to TMZ-induced cytotoxicity and may represent a novel, mechanism-based means of increasing TMZ efficacy in both p53 wild-type and p53 mutant glioma cells.
Hirose
Y
Y
Brain Tumor Research Center, Department of Neurological Surgery, University of California-San Francisco, 2340 Sutter Street, San Francisco, CA 94143-0875, USA.
Berger
M S
MS
Pieper
R O
RO
D000470:Alkaloids / Q000494:pharmacology; D018906:Antineoplastic Agents, Alkylating / Q000633:toxicity*; D016203:CDC2 Protein Kinase / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000378:metabolism; D000071877:Checkpoint Kinase 1; D003606:Dacarbazine / Q000031:analogs & derivatives / Q000633:toxicity*; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016195:G2 Phase / Q000187:drug effects / Q000502:physiology*; D005909:Glioblastoma / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D006801:Humans; D008938:Mitosis / Q000187:drug effects / Q000502:physiology; D010766:Phosphorylation / Q000187:drug effects; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases; D019311:Staurosporine / Q000031:analogs & derivatives; D000077204:Temozolomide; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism / Q000502:physiology*; D020687:cdc25 Phosphatases / Q000378:metabolism
D000470:Alkaloids; D018906:Antineoplastic Agents, Alkylating; D018797:Cell Cycle Proteins; D004791:Enzyme Inhibitors; D047428:Protein Kinase Inhibitors; D016159:Tumor Suppressor Protein p53; C054852:7-hydroxystaurosporine; D003606:Dacarbazine; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D016203:CDC2 Protein Kinase; C517009:CDC25C protein, human; D020687:cdc25 Phosphatases; D019311:Staurosporine; D000077204:Temozolomide