title: Potentiation of nitrosourea cytotoxicity in human leukemic cells by inactivation of O6-alkylguanine-DNA alkyltransferase.
abstract: The HL-60 promyelocytic leukemia cell line is resistant to nitrosoureas and contains high levels of the DNA repair protein O6-alkylguanine-DNA alkyltransferase (alkyltransferase). We examined the protective role of the alkyltransferase in the nitrosourea resistance observed in this myeloid leukemia cell line to determine whether inactivation of the alkyltransferase with the modified base, O6-methylguanine (O6mGua), could sensitize these cells to nitrosoureas. The HL-60 cells were sensitized approximately 3.0-fold to five different nitrosoureas when the alkyltransferase was inactivated by 88% following a 24-h preincubation in 0.5 mM O6mGua. No effect of O6mGua preincubation was observed in the K562 chronic myelogenous leukemia cell line which is sensitive to nitrosoureas and has low levels of alkyltransferase activity. When regeneration of HL-60 alkyltransferase activity after exposure to nitrosoureas was prevented by maintaining cells in O6mGua, HL-60 became even more sensitive (3.7- to 8.5-fold) to nitrosoureas but remained slightly more resistant than K562. Next, we compared the dose of methyl- and chloroethylnitrosoureas which were cytotoxic in HL-60 with the dose which caused repair-induced inactivation of the alkyltransferase. Both methyl- and chloroethyl-nitrosoureas caused the dose-dependent inactivation of the alkyltransferase and with both, cytotoxicity was increased with O6mGua exposure. However, chloroethylnitrosoureas, which form a variety of O6 alkylation adducts, some of which are poorly repaired, exhibited 7-12 times more cytotoxicity relative to repair-induced inactivation of the alkyltransferase whereas methylnitrosoureas became cytotoxic only when the alkyltransferase had been inactivated. These data suggest that leukemic cells are sensitized to both methyl- and chloroethylnitrosoureas when O6mGua is used to persistently inactivate the alkyltransferase. However, the alkyltransferase provides more efficient protection from methylnitrosoureas than chloroethylnitrosoureas most likely because the latter form adducts which are poorly repaired by the protein and which if unrepaired may become cytotoxic cross-links.
authors.affiliation: Department of Medicine, University Hospitals of Cleveland, Case Western Reserve University School of Medicine, Ohio 44106.
authors.forename: S L
authors.initials: SL
authors.lastname: Gerson
authors.forename: J E
authors.initials: JE
authors.lastname: Trey
authors.forename: K
authors.initials: K
authors.lastname: Miller
mesh_terms: D002470:Cell Survival / Q000187:drug effects; D004260:DNA Repair; D004305:Dose-Response Relationship, Drug; D004351:Drug Resistance; D004357:Drug Synergism; D006147:Guanine / Q000031:analogs & derivatives / Q000494:pharmacology; D006801:Humans; D015470:Leukemia, Myeloid, Acute / Q000201:enzymology* / Q000473:pathology; D008780:Methyltransferases / Q000032:analysis / Q000037:antagonists & inhibitors / Q000502:physiology*; D009607:Nitrosourea Compounds / Q000494:pharmacology*; D019853:O(6)-Methylguanine-DNA Methyltransferase; D014407:Tumor Cells, Cultured
chemical_list: D009607:Nitrosourea Compounds; D006147:Guanine; C008449:O-(6)-methylguanine; D008780:Methyltransferases; D019853:O(6)-Methylguanine-DNA Methyltransferase