Controlling of N-alkylpolyamine analogue metabolism by selective deuteration.

Replacing protium with deuterium is an efficient method to modulate drug metabolism. N-alkylated polyamine analogues are polyamine antimetabolites with proven anticancer efficacy. We have characterized earlier the preferred metabolic routes of N1,N12-diethylspermine (DESpm), N1-benzyl-N12-ethylspermine (BnEtSpm) and N1,N12-dibenzylspermine (DBSpm) by human recombinant spermine oxidase (SMOX) and acetylpolyamine oxidase (APAO). Here, we studied the above analogues, their variably deuterated counterparts and their metabolites as substrates and inhibitors of APAO, SMOX, semicarbazide-sensitive amine oxidase (SSAO), diamine oxidase (DAO) and monoamine oxidases. We found that targeted deuteration efficiently redirected the preferable cleavage site and suppressed reaction rate by APAO and SMOX in vitro We found a three- to six-fold decline in Vmax with moderate variable effect on Km when deuterium was located at the preferred hydrogen abstraction site of the analogue. We also found some of the metabolites to be potent inhibitors of DAO and SSAO. Surprisingly, analogue deuteration did not markedly alter the anti-proliferative efficacy of the drugs in DU145 prostate cancer cells, while in mouse embryonic fibroblasts, which had higher basal APAO and SMOX activities, moderate effect was observed. Interestingly, the anti-proliferative efficacy of the analogues did not correlate with their ability to suppress polyamine biosynthetic enzymes, induce spermidine/spermine-N1-acetyltransferase or deplete intracellular polyamine levels, but correlated with their ability to induce SMOX. Our data show that selective deuteration of N-alkyl polyamine analogues enables metabolic switching, offering the means for selective generation of bioactive metabolites inhibiting, e.g. SSAO and DAO, thus setting a novel basis for in vivo studies of this class of analogues.

Triethylenetetramine modulates polyamine and energy metabolism and inhibits cancer cell proliferation.

Polyamine metabolism is an attractive anticancer drug target, since polyamines are absolutely required for cellular proliferation, and increased levels of polyamines and their biosynthetic enzyme ornithine decarboxylase (ODC) are associated with cancer. Triethylenetetramine (TETA) is a charge-deficient isosteric analogue of the polyamine spermidine (Spd) and a Cu(II)-chelating compound used for the treatment of Wilson's disease, and it has been implicated as a potential anticancer therapeutic drug. In the present study, we studied the effects of TETA in comparison with two other Cu(II)-chelators, D-penicillamine (PA) and tetrathiomolybdate (TTM), on polyamine metabolism in DU145 prostate carcinoma, MCF-7 breast carcinoma and JEG-3 choriocarcinoma cells. TETA induced antizyme, down-regulated ODC and inhibited [(14)C] Spd uptake. Moreover, it completely prevented α-difluoromethylornithine (DFMO)-induced increase in [(14)C] Spd uptake, and inhibited [(14)C] putrescine (Put) uptake and ODC activity in vivo Seven-day treatment of DU145 cells with TETA caused growth cessation by reducing intracellular polyamine levels and suppressing the formation of hypusinated eukaryotic translation initiation factor 5A (eIF5A). TETA or its N-acetylated metabolites also inhibited spermine (Spm), diamine and semicarbazide-sensitive amine oxidases and decreased the level of intracellular reactive oxygen species. Moreover, TETA inhibited the utilization of Put as energy source via the tricarboxylic acid (TCA) cycle, as indicated by decreased production of (14)CO2 from [(14)C] Put. These results indicate that TETA attacks multiple proven anticancer drug targets not attributed to copper chelation, which warrants further studies to reveal its potential in cancer chemoprevention and cure.