Inhibition of human ornithine decarboxylase activity by enantiomers of difluoromethylornithine

Q. U. Ning, Natalia Ignatenko, Phillip Yamauchi, David E. Stringer, Corey Levenson, Patrick Shannon, Scott Perrin, Eugene W. Gerner

Research output: Contribution to journalArticle

31 Citations (Scopus)

Abstract

Racemic difluoromethylornithine (D/L-DFMO) is an inhibitor of ODC (ornithine decarboxylase), the first enzyme in eukaryotic polyamine biosynthesis, D/L-DFMO is an effective anti-parasitic agent and inhibitor of mammalian cell growth and development. Purified human ODC-catalysed ornithine decarboxylation is highly stereospecific. However, both DFMO enantiomers suppressed ODC activity in a time- and concentration-dependent manner. ODC activity failed to recover after treatment with either L- or D-DFMO and dialysis to remove free inhibitor. The inhibitor dissociation constant (K D) values for the formation of enzyme-inhibitor complexes were 28.3 ± 3.4, 1.3 ± 0.3 and 2.2 ± 0.4 μM respectively for D-, L- and D/L-DFMO. The differences in these KD values were statistically significant (P < 0.05). The inhibitor inactivation constants (Kinact) for the irreversible step were 0.25 ± 0.03, 0.15 ± 0.03 and 0.15 ± 0.03 min-1 respectively for D-, L- and D/L-DFMO. These latter values were not statistically significantly different (P > 0.1). D-DFMO was a more potent inhibitor (IC50 ∼ 7.5 μM) when compared with D-ornithine (IC50 ∼ 1.5 mM) of ODC-catalysed L-ornithine decarboxylation. Treatment of human colon tumour-derived HCT116 cells with either L- or D-DFMO decreased the cellular polyamine contents in a concentration-dependent manner. These results show that both enantiomers of DFMO irreversibly inactivate ODC and suggest that this inactivation occurs by a common mechanism. Both enantiomers form enzyme-inhibitor complexes with ODC, but the probability of formation of these complexes is 20 times greater for L-DFMO when compared with D-DFMO. The rate of the irreversible reaction in ODC inactivation is similar for the L- and D-enantiomer. This unexpected similarity between DFMO enantiomers, in contrast with the high degree of stereospecificity of the substrate ornithine, appears to be due to the α-substituent of the inhibitor. The D-enantiomer may have advantages, such as decreased normal tissue toxicity, over L- or D/L-DFMO in some clinical applications.

Original languageEnglish (US)
Pages (from-to)465-470
Number of pages6
JournalBiochemical Journal
Volume375
Issue number2
DOIs
StatePublished - Oct 15 2003

Fingerprint

Eflornithine
Ornithine Decarboxylase
Enantiomers
Ornithine
Decarboxylation
Polyamines
Enzyme Inhibitors
Inhibitory Concentration 50
HCT116 Cells
Dialysis
Biosynthesis
Cell growth
Growth and Development
Toxicity
Tumors
Colon
Tissue
Substrates
Enzymes

Keywords

  • Difluoromethylornithine enantiomer
  • Enzyme inactivation parameter
  • Ornithine decarboxylase
  • Polyamine

ASJC Scopus subject areas

  • Biochemistry

Cite this

Ning, Q. U., Ignatenko, N., Yamauchi, P., Stringer, D. E., Levenson, C., Shannon, P., ... Gerner, E. W. (2003). Inhibition of human ornithine decarboxylase activity by enantiomers of difluoromethylornithine. Biochemical Journal, 375(2), 465-470. https://doi.org/10.1042/BJ20030382

Inhibition of human ornithine decarboxylase activity by enantiomers of difluoromethylornithine. / Ning, Q. U.; Ignatenko, Natalia; Yamauchi, Phillip; Stringer, David E.; Levenson, Corey; Shannon, Patrick; Perrin, Scott; Gerner, Eugene W.

In: Biochemical Journal, Vol. 375, No. 2, 15.10.2003, p. 465-470.

Research output: Contribution to journalArticle

Ning, QU, Ignatenko, N, Yamauchi, P, Stringer, DE, Levenson, C, Shannon, P, Perrin, S & Gerner, EW 2003, 'Inhibition of human ornithine decarboxylase activity by enantiomers of difluoromethylornithine', Biochemical Journal, vol. 375, no. 2, pp. 465-470. https://doi.org/10.1042/BJ20030382
Ning, Q. U. ; Ignatenko, Natalia ; Yamauchi, Phillip ; Stringer, David E. ; Levenson, Corey ; Shannon, Patrick ; Perrin, Scott ; Gerner, Eugene W. / Inhibition of human ornithine decarboxylase activity by enantiomers of difluoromethylornithine. In: Biochemical Journal. 2003 ; Vol. 375, No. 2. pp. 465-470.
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N2 - Racemic difluoromethylornithine (D/L-DFMO) is an inhibitor of ODC (ornithine decarboxylase), the first enzyme in eukaryotic polyamine biosynthesis, D/L-DFMO is an effective anti-parasitic agent and inhibitor of mammalian cell growth and development. Purified human ODC-catalysed ornithine decarboxylation is highly stereospecific. However, both DFMO enantiomers suppressed ODC activity in a time- and concentration-dependent manner. ODC activity failed to recover after treatment with either L- or D-DFMO and dialysis to remove free inhibitor. The inhibitor dissociation constant (K D) values for the formation of enzyme-inhibitor complexes were 28.3 ± 3.4, 1.3 ± 0.3 and 2.2 ± 0.4 μM respectively for D-, L- and D/L-DFMO. The differences in these KD values were statistically significant (P < 0.05). The inhibitor inactivation constants (Kinact) for the irreversible step were 0.25 ± 0.03, 0.15 ± 0.03 and 0.15 ± 0.03 min-1 respectively for D-, L- and D/L-DFMO. These latter values were not statistically significantly different (P > 0.1). D-DFMO was a more potent inhibitor (IC50 ∼ 7.5 μM) when compared with D-ornithine (IC50 ∼ 1.5 mM) of ODC-catalysed L-ornithine decarboxylation. Treatment of human colon tumour-derived HCT116 cells with either L- or D-DFMO decreased the cellular polyamine contents in a concentration-dependent manner. These results show that both enantiomers of DFMO irreversibly inactivate ODC and suggest that this inactivation occurs by a common mechanism. Both enantiomers form enzyme-inhibitor complexes with ODC, but the probability of formation of these complexes is 20 times greater for L-DFMO when compared with D-DFMO. The rate of the irreversible reaction in ODC inactivation is similar for the L- and D-enantiomer. This unexpected similarity between DFMO enantiomers, in contrast with the high degree of stereospecificity of the substrate ornithine, appears to be due to the α-substituent of the inhibitor. The D-enantiomer may have advantages, such as decreased normal tissue toxicity, over L- or D/L-DFMO in some clinical applications.

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