RAPID INDUCTION OF APOPTOSIS BY DEREGULATED UPTAKE OF POLYAMINE ANALOGS

Treatment of Chinese hamster ovary cells with alpha-difluoromethylorni thine for 3 days, followed by exposure to cycloheximide, led to an unr egulated, rapid and massive accumulation of polyamine analogues. This accumulation led to cell death by apoptosis within a few hours. Clear evidence of DNA fragmentation was seen in response to both N-terminall y ethylated polyamines and to polyamines containing methyl groups on t he terminal carbon atoms. Programmed cell death was induced within 2-4 h of exposure to 1 mu M or higher concentrations of N-1,N-11-bis(ethy l)norspermine. The presence of cycloheximide increased the uptake of t he polyamine analogues and therefore led to cell death at lower analog ue concentrations, but it was not essential for the induction of apopt osis, since similar effects were seen when the protein synthesis inhib itor was omitted and the concentration of N-1,N-11-bis(ethyl)norspermi ne was increased to 5 mu M or more The induction of apoptosis was bloc ked both by the addition of the caspase inhibitor -benzyloxycarbonyl-V al-Ala-Asp-fluoromethylketone, or by the addition of the polyamine oxi dase inhibitor N-1-methyl-N-2-(2,3-butadienyl)butane-1,4-diamine (MDL 72,527). These experiments provide evidence to support the concepts th at: (1) polyamines or their oxidation products may be initiators of pr ogrammed cell death; (2) regulation of polyamine biosynthesis and upta ke prevents the accumulation of toxic levels of polyamines; and (3) th e antineoplastic effects of bis(ethyl) polyamine analogues may be due to the induction of apoptosis in sensitive tumour cells.