In Vitro and in Vivo Properties of 3-tert-Butyl-7-(5-methylisoxazol-3-yl)-2-(1-methyl-1H-1,2,4-triazol-5-ylmethoxy)-pyrazolo[1,5-d]-[1,2,4]triazine (MRK-016), a GABA_A Receptor α5 Subtype-Selective Inverse Agonist

Abstract

3-tert-Butyl-7-(5-methylisoxazol-3-yl)-2-(1-methyl-1H-1,2,4-triazol-5-ylmethoxy)-pyrazolo [1,5-d][1,2,4]triazine (MRK-016) is a pyrazolotriazine with an affinity of between 0.8 and 1.5 nM for the benzodiazepine binding site of native rat brain and recombinant human ␣1-, ␣2-, ␣3-, and ␣5-containing GABA A receptors. It has inverse agonist efficacy selective for the ␣5 subtype, and this ␣5 inverse agonism is greater than that of the prototypic ␣5-selective compound 3-(5-methylisoxazol-3-yl)-6-[(1-methyl-1,2,3-triazol-4-hdyl)methyloxy]-1,2,4-triazolo[3,4-a]phthalazine (␣5IA). Consistent with its greater ␣5 inverse agonism, MRK-016 increased long-term potentiation in mouse hippocampal slices to a greater extent than ␣5IA. MRK-016 gave good receptor occupancy after oral dosing in rats, with the dose required to produce 50% occupancy being 0.39 mg/kg and a corresponding rat plasma EC 50 value of 15 ng/ml that was similar to the rhesus monkey plasma EC 50 value of 21 ng/ml obtained using [11 C]flumazenil positron emission tomography. In normal rats, MRK-016 enhanced cognitive performance in the delayed matching-to-position version of the Morris water maze but was not anxiogenic, and in mice it was not proconvulsant and did not produce kindling. MRK-016 had a short half-life in rat, dog, and rhesus monkey (0.3-0.5 h) but had a much lower rate of turnover in human compared with rat, dog, or rhesus monkey hepatocytes. Accordingly, in human, MRK-016 had a longer half-life than in preclinical species (ϳ3.5 h). Although it was well tolerated in young males, with a maximal tolerated single dose of 5 mg corresponding to an estimated occupancy in the region of 75%, MRK-016 was poorly tolerated in elderly subjects, even at a dose of 0.5 mg, which, along with its variable human pharmacokinetics, precluded its further development.

Selective Modulators of α₅-Containing GABA_A Receptors and their Therapeutic Significance

“…One compound of this class, Compound 43 ( Table 1, 2), enhanced the cognitive performance of rats in the delayed matching-to-position (DMTP) version of the Morris water maze model without the anxiogenic or convulsive side effects typical of non-selective benzodiazepine receptor inverse agonists such as methyl-6, 7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM) [39][40][41][42]. A similar result was observed with an orally administered α 5 GABA A R-selective pyrazolotriazine compound, MRK-016 (Table 1, 2), although this drug was discontinued because it was poorly tolerated in elderly subjects and exhibited unpredictable pharmacokinetics [43,44]. Two structurally similar triazolophthalazines, α 5 IA and α 5 IA-II (Table 1, 2), were also developed to be orally bioavailable and selective for α 5 GABA A Rs.…”

Rapid Antidepressant Action and Restoration of Excitatory Synaptic Strength After Chronic Stress by Negative Modulators of Alpha5-Containing GABAA Receptors

“…Unlike nonselective negative allosteric modulators, these compounds are not anxiogenic, hallucinogenic, or epileptogenic in humans (Atack et al, 2009).…”

“…This dose should produce roughly 80% occupancy of α5-containing GABA A Rs (Atack et al, 2009). Within 24 h, a single injection of MRK-016 restored sucrose preference (F(3,33) = 20.63, po0.0001, n = 12 rats; Figure 4a and b) and social interaction (F = (2,14) = 11.84, p = 0.000978, n = 8 rats; Figure 4c) in rats subjected to CRS.…”

“…L-655,708 and MRK-016 are negative allosteric modulators of GABA A Rs with a 10-100 fold selectivity for GABA A Rs containing the α5-subunit (Atack et al, 2005;. The α5-selective negative allosteric modulators are not epileptogenic, hallucinogenic, or anxiogenic in humans (Atack et al, 2009).…”

Benzodiazepine receptor ligands: a patent review (2006 – 2012)

“…Data obtained from these studies disclosed the therapeutic utility of the a5 inverse agonists. In particular, L655708 [57] reverses memory deficit after exposure to the anesthetic isoflurane and MRK-016 [58] reverses the memory deficit associated with inflammation, both in young and aging brain.…”