The Use of Novel C-Methylated Spermidine Derivatives To Investigate the Regulation of Polyamine Metabolism

Abstract

The polyamines are organic polycations present at millimolar concentrations in eukaryotic cells where they participate in the regulation of vital cellular functions including proliferation and differentiation. Biological evaluation of rationally designed polyamine analogs is one of the cornerstones of polyamine research. Here we have synthesized and characterized novel C-methylated spermidine analogs, that is, 2-methylspermidine, 3-methylspermidine, and 8-methylspermidine. 3-Methylspermidine was found to be metabolically stable in DU145 cells, while 8-methylspermidine was a substrate for spermidine/spermine N(1)-acetyltransferase (SSAT) and 2-methylspermidine was a substrate for both SSAT and acetylpolyamine oxidase. All the analogs induced the splicing of the productive mRNA splice variant of SSAT, overcame growth arrest induced by 72-h treatment with ornithine decarboxylase (ODC) inhibitor α-difluoromethylornithine, and were transported via the polyamine transporter. Surprisingly, 2-methylspermidine was a weak downregulator of ODC activity in DU145 cells. Our data demonstrates that it is possible to radically alter the biochemical properties of a polyamine analog by changing the position of the methyl group.

Spermidine promotes adipogenesis of 3T3-L1 cells by preventing interaction of ANP32 with HuR and PP2A

“…The use of metabolically stable polyamine analogues combined with other compounds targeting polyamine metabolism such as DFMO is of great value in studies aiming to elucidate the functions of the individual polyamines [8,25]. However, the functions of the individual polyamines are difficult to assess because they are readily metabolized and complete depletion of one polyamine but not the other is difficult to achieve.…”

“…The optical isomers of (R)/(S)-MeSpd (α-MeSpd) and RR-Me 2 Spm were synthesized as described in [9] and racemic γ -MeSpd as in [8]. (Manassas, VA, U.S.A).…”

“…This stimulus triggers a rapid induction of immediate-early genes (c-myc, cjun and c-fos) and the early adipogenic transcription factors C/EBP (CCAAT/enhancer-binding protein) β and C/EBPδ, which then induce SREBP-1c (sterol-regulatory-element-binding protein 1c), PPARγ (peroxisome-proliferator-activated receptor γ ) and C/EBPα, the master regulators of adipogenesis, which promote terminal differentiation. Since spermidine and spermine are readily interconvertible, and even catabolically stable α-MeSpd is a substrate of spermine synthase, in the present study we used their novel metabolically stable functional mimetics, γ -MeSpd and RR-Me 2 Spm [(R,R)-α,ωbismethylspermine] [8] to investigate the molecular mechanisms responsible for the spermidine depletion-mediated inhibition of adipogenesis. We have shown previously that the expression levels of PPARγ and C/EBPα are dramatically reduced in spermidine-depleted 3T3-L1 cells [6].…”

Enantiomers of 3-Methylspermidine Selectively Modulate Deoxyhypusine Synthesis and Reveal Important Determinants for Spermidine Transport

“…We used SC instead of aminoguanidine (AG), another widely used serum amine oxidase inhibitor, because we found that AG treatment alone inhibits DHS and retards cell growth 15 . Our previous studies indicated that racemic 1-, 2-, and 3-MeSpd were not cytotoxic in the absence of serum amine oxidase inhibitor, while Spd and rac-8-MeSpd were 16 . Thus, racemic 1-, 2-and 3-MeSpd were not considered to be degraded by the serum amine oxidases to cytotoxic aldehydes 16 .…”

Chemical activation of SAT1 corrects diet-induced metabolic syndrome

“…[25]. A standard assay mixture contained 100 mM Tris-HCl, pH 7.8, 1 mM semicarbazide, 5 mM EDTA, 3 mM 8-MeSpd [26], and 50 nCi (50-60 mCi/mmol) of 14 C-acetyl-CoA (Moravek Inc.) in a total volume of 100 μl. 8-MeSpd has higher affinity for SAT1 in comparison to its natural substrate spermidine (K m 78 ± 3 µM and V max of 7.35 ± 0.10 µmol/min/mg vs. K m 151 ± 15 µM and V max of 4.28 ± 0.13 µmol/min/mg).…”