Enhancement of 1,3-bis(2-chloroethyl)-1-nitrosourea-induced cytotoxicity and DNA damage by alpha-difluoromethylornithine in L1210 leukemia cells.
Polyamine depletion by pretreatment with alpha-difluoromethylornithine (DFMO), a specific and irreversible inhibitor of ornithine decarboxylase, potentiates the cytotoxicity of 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) in L1210 leukemia cells grown in a modified soft agar system. The dose enhancement ratio was 1.97 at a control colony formation level of 5%. The basis for this enhancement was investigated at the level of DNA damage using a modified fluorometric assay to quantitate the production of alkaline-labile strand breaks per relative DNA molecular mass. Pretreatment of cultured L1210 cells for 48 hr with 5 mM DFMO depleted intracellular putrescine and spermidine (but not spermine) pools and resulted in a 2.3-fold increase in BCNU-induced (10 micrograms/ml, 2 hr) DNA strand breaks per relative DNA molecular mass. The inclusion of 10 microM spermidine during the DFMO pretreatment fully prevented growth inhibition and enhancement of BCNU-induced DNA damage while maintaining cellular spermidine pools at control levels. The inclusion of 2 microM putrescine or spermidine also prevented growth inhibition and enhancement of DNA damage while maintaining spermidine pools at only 25 to 35% of control. Thus, the portion of spermidine essential for cell growth appears to be associated with DNA. BCNU itself was found to reduce cellular polyamine levels by causing their leakage from cells. In addition, BCNU was found to react directly with spermidine in a cell-free system, resulting in a major reaction product detectable by high-performance liquid chromatography. While decreased interaction of BCNU with polyamines could account, in part, for enhancement effects of DFMO, it is more probable that alterations in DNA structure secondary to polyamine depletion are responsible for these effects.
D000818:Animals; D002330:Carmustine / Q000633:toxicity*; D002470:Cell Survival / Q000187:drug effects; D004273:DNA, Neoplasm / Q000378:metabolism; D004357:Drug Synergism; D000518:Eflornithine; D007700:Kinetics; D007939:Leukemia L1210 / Q000503:physiopathology*; D051379:Mice; D009952:Ornithine / Q000031:analogs & derivatives* / Q000633:toxicity; D065108:Ornithine Decarboxylase Inhibitors; D011073:Polyamines / Q000378:metabolism; D011700:Putrescine / Q000494:pharmacology; D013095:Spermidine / Q000494:pharmacology; D013096:Spermine / Q000494:pharmacology
D004273:DNA, Neoplasm; D065108:Ornithine Decarboxylase Inhibitors; D011073:Polyamines; D013096:Spermine; D009952:Ornithine; D002330:Carmustine; D013095:Spermidine; D011700:Putrescine; D000518:Eflornithine