Spermine is often the most abundant
polyamine in human
tumors such as
breast carcinomas. However, its specific role in
tumor biology is still uncertain, since inhibitors of
ornithine decarboxylase such as
alpha-difluoromethylornithine depress cell growth while leaving
spermine content mostly unaffected. We have assessed the specific role of
spermine in
breast cancer cell growth using N-cyclohexyl-1,3-diaminopropane (C-DAP), a potent
spermine synthase inhibitor. In ZR-75-1 cells, C-DAP decreased net cell growth after 14 days by 65% at 50 microm, with an IC50 of about 5 microM, and was about 10 times more potent than N-(n-butyl)-1,3-diaminopropane, another
spermine synthase inhibitor. C-DAP acted as a specific inhibitor of
spermine biosynthesis, since (a) it depleted
spermine content while causing an equal or greater accumulation of
spermidine on a molar basis, (b) it rapidly induced
S-adenosylmethionine decarboxylase activity and the accumulation of its products due to relief of
spermine-dependent inhibition of
enzyme expression, and (c) exogenous
spermine (1 microM) completely reversed C-DAP-induced growth inhibition. C-DAP and related compounds were accumulated, at least in part, through a mechanism distinct from the
polyamine transport system, while also blocking
putrescine and
spermidine uptake with various potencies. Reversibility of C-DAP-induced growth inhibition by exogenous
spermine was progressively lost on prolonged treatment, in association with marked morphological changes. In 4 different human
breast cancer cell lines (ZR-75-1, T47-D, MCF-7, and MDA-MB-231), relative growth sensitivity to C-DAP was inversely related to the extent of
spermidine accumulation caused by
spermine synthase inhibition, suggesting that
spermidine overaccumulation can functionally replace
spermine. Interestingly, C-DAP strongly potentiated growth inhibition caused by
alpha-difluoromethylornithine in all cell lines tested by preventing conversion of residual
spermidine to
spermine, indicating that
spermine synthesis limits
alpha-difluoromethylornithine action and that under some critical threshold,
spermidine cannot fulfill cellular needs for
spermine. Thus,
spermine plays specific and important functions in
breast tumor growth, and
spermine synthase inhibitors could markedly improve the therapeutic effectiveness of existing
polyamine depletion strategies, especially in
spermine-rich
tumors.