Inactivated MGMT by O6-benzylguanine is associated with prolonged G2/M arrest in cancer cells treated with BCNU.
HCT116 and HCT15 cells that highly express O(6)-methylguanine-DNA-methyltransferase (MGMT) displayed a transient cell cycle G2/M arrest in response to exposure to 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) alone; however, 70-80% of cells were arrested in G2/M after treatment with O(6)-benzylguanine (BG) and BCNU. Cells accumulated in G2/M showed elevated levels of an inactive form of cyclin B1/p-Cdc2 (Tyr15) complex that was not associated with activation of Chk1/p-Cdc25C and was independent of p53/p21 status. The most prominent feature of cell death was the appearance of enlarged and multinucleated cells that was related to the inhibition of mitotic entry. In contrast, BG-resistant cell lines, HCT116 BBR and HCT15 BBR cells that contain mutations K165E and K165N of MGMT, respectively, displayed a normal cell cycle progression with a slight and transient increase in G2/M arrest at 24 h after treatments with either BCNU alone or BG combined with BCNU. The differences in the ability to progress toward G2/M after treatment with BG and BCNU between cells expressing wild-type MGMT and mutated MGMT were confirmed in CHO cells transfected with human wild type and K165E mutant MGMT cDNA, respectively. Thus, our findings suggest that BG-inactivated MGMT may be linked to cell signaling events, forcing cells into a permanent G2/M arrest in response to the DNA damages induced by BCNU.
Yan
Ling
L
Division of Hematology and Oncology, Department of Medicine, Case Western Reserve University, Cleveland, OH 44106, USA.
Donze
Jon R
JR
Liu
Lili
L
D019943:Amino Acid Substitution; D002330:Carmustine / Q000633:toxicity*; D002453:Cell Cycle / Q000187:drug effects*; D016923:Cell Death / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D004249:DNA Damage; D019008:Drug Resistance, Neoplasm; D016195:G2 Phase / Q000187:drug effects; D006147:Guanine / Q000031:analogs & derivatives* / Q000633:toxicity; D006801:Humans; D016297:Mutagenesis, Site-Directed; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000037:antagonists & inhibitors* / Q000235:genetics
C064976:O(6)-benzylguanine; D006147:Guanine; D019853:O(6)-Methylguanine-DNA Methyltransferase; D002330:Carmustine