Synthesis and Pharmacological Evaluation of Enantiomerically Pure GluN2B Selective NMDA Receptor Antagonists.

To determine the eutomers of potent GluN2B-selective N-methyl-d-aspartate (NMDA) receptor antagonists with a 3-benzazepine scaffold, 7-benzyloxy-3-(4-phenylbutyl)-2,3,4,5-tetrahydro-1H-3-benzazepin-1-ols (S)-2 and (R)-2 were separated by chiral HPLC. Hydrogenolysis and subsequent methylation of the enantiomerically pure benzyl ethers of (S)-2 and (R)-2 provided the enantiomeric phenols (S)-3 and (R)-3 [3-(4-phenylbutyl)-2,3,4,5-tetrahydro-1H-3-benzazepine-1,7-diol] and methyl ethers (S)-4 and (R)-4. All enantiomers were obtained with high enantiomeric purity (≥99.7 % ee). The absolute configurations were determined by CD spectroscopy. R-configured enantiomers turned out to be the eutomers in receptor binding studies and two-electrode voltage clamp experiments. The most promising ligand of this compound series is the R-configured phenol (R)-3, displaying high GluN2B affinity (Ki =30 nm), high inhibition of ion flux (IC50 =61 nm), and high cytoprotective activity (IC50 =93 nm). Whereas the eudismic ratio in the receptor binding assay is 25, the eudismic ratio in the electrophysiological experiment is 3.

Fluorinated GluN2B Receptor Antagonists with a 3-Benzazepine Scaffold Designed for PET Studies.

To analyze the N-methyl-d-aspartate (NMDA) receptor distribution in the central nervous system, fluorinated ligands that selectively address the ifenprodil binding site of GluN2B-subunit-containing NMDA receptors were developed. Various strategies to introduce a fluorine atom into the potent GluN2B ligand 2 (3-(4-phenylbutyl)-2,3,4,5-tetrahydro-1H-3-benzazepin-1,7-diol) were pursued, including replacement of the benzylic OH moiety with a fluorine atom (13) and introduction of fluoroethoxy moieties at various positions (14 (7-position), 17 (9-position), 18a-c (1-position)). With respect to GluN2B affinity and selectivity over related receptors, the fluoroethoxy derivatives 14 and 18a are the most promising ligands. Radiosynthesis of fluoroethoxy derivative [18 F]14 was performed by nucleophilic substitution of the phenol 2 with 2-[18 F]fluoroethyl tosylate. On rat brain slices the fluorinated PET tracer [18 F]14 accumulated in regions with high density of NMDA receptors containing GluN2B subunits. The bound radioactivity could not be replaced by (S)-glutamate. However, the GluN2B ligands eliprodil, Ro 25-6981, and the non-labeled 3-benzazepine 14 were able to abolish the specific binding of [18 F]14.

Evaluation of 11C-Me-NB1 as a Potential PET Radioligand for Measuring GluN2B-Containing NMDA Receptors, Drug Occupancy, and Receptor Cross Talk.

Clinical and preclinical research with modulators at the N-methyl-d-aspartate (NMDA) receptor GluN2B N-terminal domain (NTD) aims for the treatment of various neurologic diseases. The interpretation of the results is hampered by the lack of a suitable NMDA PET tracer for assessing the receptor occupancy of potential drugs. We have developed 11C-Me-NB1 as a PET tracer for imaging GluN1/GluN2B-containing NMDA receptors and used it to investigate in rats the dose-dependent receptor occupancy of eliprodil, a GluN2B NTD modulator. Methods:11C-Me-NB1 was synthesized and characterized by in vitro displacement binding experiments with rat brain membranes, in vitro autoradiography, and blocking and displacement experiments by PET and PET kinetic modeling. Receptor occupancy by eliprodil was studied by PET with 11C-Me-NB1. Results:11C-Me-NB1 was synthesized at 290 ± 90 GBq/µmol molar activity, 7.4 ± 1.9 GBq total activity at the end of synthesis (n = 17), and more than 99% radiochemical purity. 11C-Me-NB1 binding in rat brain was blocked in vitro and in vivo by the NTD modulators Ro-25-6981 and eliprodil. Half-maximal receptor occupancy by eliprodil occurred at 1.5 µg/kg. At 1 mg/kg of eliprodil, a dose with reported neuroprotective effects, more than 99.5% of binding sites were occupied. In vitro, 11C-Me-NB1 binding was independent of the σ-1 receptor (Sigma1R), and the Sigma1R agonist (+)-pentazocine did not compete for high-affinity binding. In vivo, a 2.5 mg/kg dose of (+)-pentazocine abolished 11C-Me-NB1-specific binding, indicating an indirect effect of Sigma1R on 11C-Me-NB1 binding. Conclusion:11C-Me-NB1 is suitable for the in vivo imaging of NMDA GluN1/GluN2B receptors and the assessment of receptor occupancy by NTD modulators. GluN1/GluN2B NMDA receptors are fully occupied at neuroprotective doses of eliprodil. Furthermore, 11C-Me-NB1 enables imaging of GluN1/GluN2B NMDA receptor cross talk.

Synthesis, biological evaluation and molecular docking studies of benzyloxyacetohydroxamic acids as LpxC inhibitors.

The inhibition of the UDP-3-O-[(R)-3-hydroxymyristoyl]-N-acetylglucosamine deacetylase (LpxC) represents a promising strategy to combat infections caused by multidrug-resistant Gram-negative bacteria. In order to elucidate the functional groups being important for the inhibition of LpxC, the structure of our previously reported hydroxamic acid 4 should be systematically varied. Therefore, a series of benzyloxyacetohydroxamic acids was prepared, of which the diphenylacetylene derivatives 28 (Ki=95nM) and 21 (Ki=66nM) were the most potent inhibitors of Escherichia coli LpxC. These compounds could be synthesized in a stereoselective manner employing a Sharpless asymmetric dihydroxylation and a Sonogashira coupling in the key steps. The obtained structure-activity relationships could be rationalized by molecular docking studies.