Potent catechol-O-methyltransferase Inhibitor Research Articles

Inhibition of human erythrocyte and gastroduodenal catechol-O-methyltransferase activity by nitecapone

The effect of increasing single oral doses of the novel catechol-O-methyltransferase (COMT) inhibitor, nitecapone, on enzyme activity in red cells (RBC) and gastroduodenal COMT activity has been studied in healthy male volunteers. A dose-dependent decrease in RBC COMT activity was seen in all cases after 1 to 150 mg of the drug. The highest dose of 300 mg did not produce much more inhibition of COMT than 150 mg. The inhibition was not complete; at the highest doses the COMT activity was reduced by 50-60%. The effect and the duration of the inhibition in RBC COMT was strongly correlated with plasma nitecapone concentrations in the dose range up to 150 mg. RBC COMT activity recovered fully in 4 h after medication. Gastric mucosal COMT activity was several-fold higher than that in RBCs. It was also dose-dependently inhibited at the two doses (25 and 100 mg) studied. The inhibition of gastric and duodenal COMT was greater than that in RBCs. This also indicates that nitecapone is locally active in the gastroduodenal tract. The results confirm nitecapone as a potent COMT inhibitor in human tissues. New COMT inhibitors may provide a valuable approach to the treatment of Parkinson's disease in combination with L-dopa and dopa decarboxylase inhibitor therapy.

Antiestrogen action of 2-hydroxyestrone on MCF-7 human breast cancer cells.

The estrogen responsive human breast cancer MCF-7 cell culture was examined for its response to 2-hydroxyestrone a principal metabolite of estradiol. Addition of 2-hydroxyestrone to the cell cultures in concentration of 10(-9) - 10(-6) M had no effect on cell growth and proliferation because of rapid O-methylation of the catechol estrogen by catechol O-methyltransferase which is highly active in these cells. In the presence of quinalizarin, a potent catechol O-methyltransferase inhibitor which reduces the O-methylation of the steroid, 10(-7) M and 10(-8) M 2-hydroxyestrone markedly suppresses the growth and proliferation of the cells. The tumor cell growth-inhibitory action of the catechol estrogen was neutralized by the presence of 10(-9) M estradiol. The catechol estrogen inhibition of cell growth is not observed in the estrogen receptor-negative human breast cancer cell lines MDA-MB-231 and MDA-MB-330 providing evidence that the inhibition is specific and is estrogen receptor-mediated. In contrast, the 16 alpha-hydroxylated metabolites of estradiol, estriol and 16 alpha-hydroxyestrone, are effective stimulators of MCF-7 cell proliferation with the latter exhibiting potency in excess of that expected from its estrogen receptor affinity. The present results represent the first observation of a specific receptor-mediated antiestrogenic action of 2-hydroxyestrone and suggest that the physiological regulation of the agonist activity of the primary estrogen may involve in situ generation of catechol estrogen.

Effects of short-term exposure to catecholestrogens on catecholamine turnover in the preoptic-hypothalamic brain of ovariectomized rats

The effects of a series of catecholestrogens (2-hydroxyestrogens and 4-hydroxyestrogens) were compared to those of a primary estrogen, ethynylestradiol (EE 2), on the catecholaminergic system in the preoptic-hypothalamic rat brain. Adult ovariectomized rats received single injections (100 μg s.c.) of EE 2, 2-hydroxyestradiol (2-OHE 2), 2-hydroxyethynylestradiol (2-OHEE 2), 4-hydroxyestradiol (4-OHE 2) or 4-hydroxyethynylestradiol (4-OHEE 2). Eight hours after estrogen administration the animals were killed, and the concentrations of dopamine (DA), noradrenaline (NA) and adrenaline (A) were determined radioenzymatically in the preoptic area (POA) and the mediobasal hypothalamus (MBH). EE 2 and all the catecholestrogens tested uniformly suppressed (P < 0.01) serum LH levels. This correlated well with the decreased turnover rate of A in the POA. In both the POA and the MBH of EE 2-treated animals the turnover rate of NA was markedly decreased whereas the concentrations of catecholamines remained unaffected. The catecholethynylestrogens, potent inhibitors of catechol O-methyltransferase, caused two-fold increases of NA concentrations in the POA (2-OHEE 2 and 4-OHEE 2) and MBH (2-OHEE 2) without affecting the turnover rate. Thus, although EE 2 and all the catecholestrogens tested uniformly suppressed LH release they induced highly different effects on the noradrenergic system in the preoptic-hypothalamic brain.

Catechol-O-methyltransferase inhibition by U-0521 increases striatal utilization of levodopa.

The effects of U-0521, a catechol-O-methyltransferase (COMT) inhibitor, were studied on this enzyme activity and on Dopa metabolism in rat striatum. In vivo maximal inhibition (95%) of COMT activity was obtained at 5 min with enzyme recovery to 64% of basal activity at 120 min. When injected in increasing doses U-0521 (200 mg . kg-1) inhibited, at 10 min, COMT activity by 85% with an IC50 = 80 mg . kg-1. In rats pretreated with U-0521 and then with DOPA the accumulation of 3-O-methyldopa-(OMD) in the plasma was essentially blocked while Dopa, dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) accumulation in the striatum was significantly higher than in DOPA treated controls. U-0521, a potent COMT inhibitor, enhances the availability and utilization of levodopa in the brain and may thus be helpful in future treatment of parkinsonian patients.

Catechol-o-methyltransferase biochemical genetics: human lymphocyte enzyme.

Human erythrocyte (RBC) catechol-O-methyltransferase (COMT) is under genetic control. Experiments were performed to determine whether COMT in the human lymphocyte is regulated in parallel with RBC COMT. Supernatants of lymphocyte homogenates contained COMT activity. However, they also contained a potent COMT inhibitor, the effect of which could be negated by dilution. Lymphocyte COMT activity was maximal at a reaction pH of 7.7 an at a MgCl2 concentration of 0.67 mM. The apparent Km value for 3,4-dihydroxybenzoic acid, the catechol substrate for the reaction, was 1.2 x 10(-5) M and that for S-adenosyl-L-methionine, the methyl donor, was 2.3 x 10(-6) M. An average of 48.3 /+- 3.3% (mean /+- SEM) of the enzyme activity in crude lymphocyte homogenates from 3 subjects was removed by centrifugation at 100,000 g for 1 hr and was presumed to be membrane associated. The average COMT activity in lymphocytes isolated from blood of 23 randomly selected adult subjects was 14.0 /+- 1.2 units/10(6) cells (mean /+- SEM) or 913 /+- 69 units/mg protein. There was a significant correlation of relative RBC with relative lymphocyte COMT activity in these 23 subjects. The correlation coefficient was 0.733 (P less than 0.001) when lymphocyte enzyme activity was expressed per milligram of protein and 0.649 (P less than 0.001) when lymphocyte activity was expressed per 10(6) cells. These results are compatible with the conclusion that the genetic polymorphism which regulates RBC COMT activity may also regulate the level of human lymphocyte COMT activity.

Inhibition of rat liver catechol- O-methyltranferase by lanthanum, neodymium and europium

Partially purified rat liver catechol- O-methyltransferase (COMT) is inhibited by lanthanum, and by the lanthanides, neodymium and europium. The 50 per cent inhibitory concentrations are 3 × 10 −6, 1.2 × 10 −6, and 8.5 × 10 −7 M for La 3+, Nd 3+ and Eu 3+ respectively. The inhibition of COMT by these ions is reversible. Lineweaver-Burk plots of the results of experiments in which the enzyme activity was measured in the presence of varying concentrations of magnesium, an activator of COMT, and different concentrations of La 3+, Nd 3+ and Eu 3+ were compatible with non-competitive or “mixed” inhibition. Double reciprocal plots of the results of experiments in which COMT activity was determined in the presence of different concentrations of La 3+ and varying quantities of the two co-substrates for the reaction, S-adenosyl-1-methionine and 3,4-dihydroxybenzoic acid, were also compatible with non-competitive or “mixed inhibition”. The characteristics of the inhibition of rat liver COMT by La 3+, Nd 3+ and Eu 3+ are similar to those of the inhibition of COMT by Ca 2+. However, lanthanum, neodymium and europium are two to three orders of magnitude more potent inhibitors of COMT than is calcium.