Inhibition of mammalian spermine synthase by N‐alkylated‐1,3‐diaminopropane derivatives in vitro and in cultured rat hepatoma cells

Abstract

A number of N-alkylated-l,3-diaminopropane derivatives [H2N-(CH2)3-NH-(CH2),,H, where n = 1 -91 have been tested as potential inhibitors of partially purified rat hepatoma (HTC) cell or pure bovine spleen spermine synthase.Among the compounds described in this paper, the most potent competitive inhibitor of spermine synthase, with respect to spermidine, is N-butyl-l,3-didminopropane with Ki values of 11.9 nM and 10.4 nM for the HTC cell and bovine spleen enzymes respectively. Inhibition of spermine synthase by this alkylated amine is selective since spermidine synthase activity is not affected up to 100 pM N-butyl-1,3-diaminopropane at a range of 5 -200 pM putrescine.Added to the culture medium of growing HTC cells, N-butyl-l,3-diaminopropane causes the expected changes in the polyamine levels with a marked decrease of spermine and an increase of spermidine. Under these conditions cell growth continues unabated.Such N-alkylated-I ,3-diaminopropane derivatives may have considerable potential as tools for studying the role of polyamines and in particular the functions of spermine in cell multiplication and differentiation.The requirement of polyamine synthesis for eukaryotic cell multiplication has been demonstrated mainly by the use of specific irreversible inhibitors of L-ornithine decarboxylase, one of the key enzymes of the polyamine biosynthetic pathway (reviewed in [l]). Inhibition of L-ornithine decarboxylase has been shown to exert beneficial effects in animal models of proliferative diseases [2 -51 and in some protozoan infections in humans [6]. Altogether these studies emphasize the importance of the polyamine metabolic pathway as promising target for the design of new anti-proliferative agents [l, 21.Use of L-ornithine-decarboxylase inhibitors for creating polyamine deficiency in mammalian tissues has however certain limitations. In vivo administration of these compounds causes putrescine and spermidine deficiency but scarcely affects tissue spermine concentrations [7, 81. As hypothesized [9, lo], maintenance of residual cell growth and lack of cytotoxicity of these L-ornithine decarboxylase inhibitors may possibly result from their ineffectiveness in altering initial spermine levels. There is some experimental support for this hypothesis, at least in cultured cells [lo, I I]. However, availability of specific and very effective inhibitors of spermine synthase should provide more direct information on particular biological functions of spermine both in vitro and in vivo.Correspondence to P. S. Marnont, Merrell Dow Research Institute,

Polyamine metabolism and cancer

“…The reactions catalyzed by PAPT and SAPT are similar, except for the utilization of putrescine by PAPT and spermidine by SAPT. Attempts to synthesize inhibitors of PAPT and SAPT have resulted in cyclohexylamine for PAPT and N-(n-butyl)-1,3-diaminopropane (BDAP) and N-(3-aminopropyl) cyclohexylamine (APCHA) for SAPT [57,58]. Sadenosyl-1,8-diamino-3-thiooctane (AdoDATO) is a more potent and specific inhibitor of PAPT, with no inhibitory effect on SAPT [59].…”

Effect of spermine synthase deficiency on polyamine biosynthesis and content in mice and embryonic fibroblasts, and the sensitivity of fibroblasts to 1,3-bis-(2-chloroethyl)-N-nitrosourea

“…Yeast mutants that lack spermine synthase are viable and do not require exogenous spermine for growth [8]. Treatment of rodents with inhibitors of spermine synthase did not produce any striking effects [10], and studies in which mammalian cells in culture have been exposed to inhibitors of spermine synthase have given variable results [11][12][13][14][15]. Serious problems in the interpretation of these experiments are caused by : (a) the possible lack of specificity of the inhibitors ; (b) the possibility that the inhibitors might substitute for spermine ; and (c) the difficulty in obtaining a major reduction in spermine levels in both cultured cells and in tissues of rodents treated with the maximal tolerated dose of the inhibitors.…”

Circular Dichroism and NMR Studies of Metabolically Stableα-Methylpolyamines: Spectral Comparison with Naturally Occurring Polyamines

“…Of these structural modifications, an ␣-methyl derivative showed the most potent biological activity in a graft-vs-host disease mouse model. While the biological utility of other ␣-substituted polyamines (Me 2 , CF 3 ) has been reported (26,27), both the ␣-Me 2 -and ␣-CF 3 -DSG analogs were inactive. In addition, the ␣-methyl derivative was the most active drug in a heart allotransplantation rat model, surpassing even DSG and showing improved pharmacokinetic properties.…”