Chemical, pharmacodynamic and pharmacokinetic characterization of the GluN2B receptor antagonist 3-(4-phenylbutyl)-2,3,4,5-tetrahydro-1H-3-benzazepine-1,7-diol - starting point for PET tracer development.

GluN2B-NMDA receptors play a key role in several neurological and neurodegenerative disorders. In order to develop novel negative allosteric GluN2B-NMDA receptor modulators, the concept of conformational restriction was pursued, i.e. the flexible aminoethanol substructure of ifenprodil was embedded into a more rigid tetrahydro-3-benzazepine system. The resulting tetrahydro-3-benzazepine-1,7-diol (±)-2 (WMS-1410) showed promising receptor affinity in receptor binding studies (K i = 84 nM) as well as pharmacological activity in two-electrode-voltage-clamp experiments (IC 50 = 116 nM) and in cytoprotective assays (IC 50 = 18.5 nM). The interactions of (R)-2 with the ifenprodil binding site of GluN2B-NMDA receptors were analyzed on the molecular level and the "foot-in-the-door" mechanism was developed. Due to promising pharmacokinetic parameters (logD7.4 = 1.68, plasma protein binding of 76-77%, sufficient metabolic stability) F-substituted analogs were prepared and evaluated as tracers for positron emission tomography (PET). Both fluorine-18-labeled PET tracers [18F]11 and [18F]15 showed high brain uptake, specific accumulation in regions known for high GluN2B-NMDA receptor expression, but no interactions with σ 1 receptors. Radiometabolites were not observed in the brain. Both PET tracers might be suitable for application in humans.

In vitro ADME characterization of a very potent 3-acylamino-2-aminopropionic acid-derived GluN2C-NMDA receptor agonist and its ester prodrugs.

The GluN2C subunit exists predominantly, but not exclusively in NMDA receptors within the cerebellum. Antagonists such as UBP1700 and positive allosteric modulators including PYD-106 and 3-acylamino-2-aminopropionic acid derivatives such as UA3-10 ((R)-2-amino-3-{[5-(2-bromophenyl)thiophen-2-yl]carboxamido}propionic acid) represent promising tool compounds to investigate the role of GluN2C-containing NMDA receptors in the signal transduction in the brain. However, due to its high polarity the bioavailability and CNS penetration of the amino acid UA3-10 are expected to be rather low. Herein, three ester prodrugs 12a-c of the NMDA receptor glycine site agonist UA3-10 were prepared and pharmacokinetically characterized. The esters 12a-c showed higher lipophilicity (higher logD 7.4 values) than the acid UA3-10 but almost the same binding at human serum albumin. The acid UA3-10 was rather stable upon incubation with mouse liver microsomes and NADPH, but the esters 12a-c were fast hydrolyzed to afford the acid UA3-10. Incubation with pig liver esterase and mouse serum led to rapid hydrolysis of the esters 12a-c. The isopropyl ester 12c showed a promising logD 7.4 value of 3.57 and the highest stability in the presence of pig liver esterase and mouse serum. These results demonstrate that ester prodrugs of UA3-10 can potentially afford improved bioavailability and CNS penetration.

GluN2A-Selective NMDA Receptor Antagonists: Mimicking the U-Shaped Bioactive Conformation of TCN-201 by a [2.2]Paracyclophane System.

Under physiological conditions, N-Methyl-D-Aspartate (NMDA) receptors play a crucial role for synaptic plasticity, long-term potentiation and long-term depression. However, overactivation of NMDA receptors can result in excitotoxicity, which is associated with various neurological and neurodegenerative diseases. The physiological properties of NMDA receptors are strongly dependent on the GluN2 subunit incorporated into the heterotetrameric NMDA receptor. Therefore, subtype selective NMDA receptor modulators are of high interest. Since prototypical GluN2A-NMDA receptor antagonists TCN-201 and its MPX-analogs adopt a U-shaped conformation within the binding pocket, paracyclophanes were designed containing the phenyl rings in an already parallel orientation. Docking studies of the designed paracyclophanes show a similar binding pose as TCN-201. [2.2]Paracyclophanes with a benzoate or benzamide side chain were prepared in four-step synthesis, respectively, starting with a radical bromination in benzylic 1-position of [2.2]paracyclophane. In two-electrode voltage clamp experiments using Xenopus laevis oocytes transfected with cRNAs for the GluN1-4a and GluN2A subunits, the esters and amides (conc. 10 µM) did not show considerable inhibition of ion flux. It can be concluded that the GluN2A-NMDA receptor does not accept ligands with a paracyclophane scaffold functionalized in benzylic 1-position, although docking studies had revealed promising binding poses for benzoic acid esters and benzamides.

[2.2]Paracyclophane-Based TCN-201 Analogs as GluN2A-Selective NMDA Receptor Antagonists.

Recent studies have shown the involvement of GluN2A subunit-containing NMDA receptors in various neurological and pathological disorders. In the X-ray crystal structure, TCN-201 (1) and analogous pyrazine derivatives 2 and 3 adopt a U-shape (hairpin) conformation within the binding site formed by the ligand binding domains of the GluN1 and GluN2A subunits. In order to mimic the resulting π/π-interactions of two aromatic rings in the binding site, a [2.2]paracyclophane system was designed to lock these aromatic rings in a parallel orientation. Acylation of [2.2]paracyclophane (5) with oxalyl chloride and chloroacetyl chloride and subsequent transformations led to the oxalamide 7, triazole 10 and benzamides 12. The GluN2A inhibitory activities of the paracyclophane derivatives were tested with two-electrode voltage clamp electrophysiology using Xenopus laevis oocytes expressing selectively functional NMDA receptors with GluN2A subunit. The o-iodobenzamide 12 b with the highest similarity to TCN-201 showed the highest GuN2A inhibitory activity of this series of compounds. At a concentration of 10 µM, 12 b reached 36 % of the inhibitory activity of TCN-201 (1). This result indicates that the [2.2]paracyclophane system is well accepted by the TCN-201 binding site.