William K. Hagmann

[18F]MK-9470, a positron emission tomography (PET) tracer for in vivo human PET brain imaging of the cannabinoid-1 receptor

[18F]MK-9470 is a selective, high-affinity, inverse agonist (human IC50, 0.7 nM) for the cannabinoid CB1 receptor (CB1R) that has been developed for use in human brain imaging. Autoradiographic studies in rhesus monkey brain showed that [18F]MK-9470 binding is aligned with the reported distribution of CB1 receptors with high specific binding in the cerebral cortex, cerebellum, caudate/putamen, globus pallidus, substantia nigra, and hippocampus. Positron emission tomography (PET) imaging studies in rhesus monkeys showed high brain uptake and a distribution pattern generally consistent with that seen in the autoradiographic studies. Uptake was blocked by pretreatment with a potent CB1 inverse agonist, MK-0364. The ratio of total to nonspecific binding in putamen was 4–5:1, indicative of a strong specific signal that was confirmed to be reversible via displacement studies with MK-0364. Baseline PET imaging studies in human research subject demonstrated behavior of [18F]MK-9470 very similar to that seen in monkeys, with very good test–retest variability (7%). Proof of concept studies in healthy young male human subjects showed that MK-0364, given orally, produced a dose-related reduction in [18F]MK-9470 binding reflecting CB1R receptor occupancy by the drug. Thus, [18F]MK-9470 has the potential to be a valuable, noninvasive research tool for the in vivo study of CB1R biology and pharmacology in a variety of neuropsychiatric disorders in humans. In addition, it allows demonstration of target engagement and noninvasive dose-occupancy studies to aid in dose selection for clinical trials of CB1R inverse agonists.

Potent, orally absorbed glucagon receptor antagonists.

The SAR of 2-pyridyl-3,5-diaryl pyrroles, ligands of the human glucagon receptor and inhibitors of p38 kinase, were investigated. This effort resulted in the identification of 2-(4-pyridyl)-5-(4-chlorophenyl)-3-(5-bromo-2-propyloxyphenyl)pyrr ole 49 (L-168,049), a potent (Kb = 25 nM), selective antagonist of glucagon.

Substituted 2-aminopyridines as inhibitors of nitric oxide synthase

A series of substituted 2-aminopyridines was prepared and evaluated as inhibitors of human nitric oxide synthases (NOS). 4,6-Disubstitution enhanced both potency and specificity for the inducible NOS with the most potent compound having an IC50 of 28 nM.

Discovery of N-[(1S,2S)-3-(4-Chlorophenyl)-2- (3-cyanophenyl)-1-methylpropyl]-2-methyl-2- {[5-(trifluoromethyl)pyridin-2-yl]oxy}propanamide (MK-0364), a novel, acyclic cannabinoid-1 receptor inverse agonist for the treatment of obesity.

The discovery of novel acyclic amide cannabinoid-1 receptor inverse agonists is described. They are potent, selective, orally bioavailable, and active in rodent models of food intake and body weight reduction. A major focus of the optimization process was to increase in vivo efficacy and to reduce the potential for formation of reactive metabolites. These efforts led to the identification of compound 48 for development as a clinical candidate for the treatment of obesity.

Substituted imidazoles as glucagon receptor antagonists.

A modestly active, nonselective triarylimidazole lead was optimized for binding affinity with the human glucagon receptor. This led to the identification of a 2- and/or 4-alkyl or alkyloxy substituent on the imidazole C4-aryl group as a structural determinant for significant enhancement in binding with the glucagon receptor (e.g., 41, IC(50)=0.053 microM) and selectivity (>1000x) over p38MAP kinase in this class of compounds.

Characterization of a Novel, Non-peptidyl Antagonist of the Human Glucagon Receptor

We have identified a series of potent, orally bioavailable, non-peptidyl, triarylimidazole and triarylpyrrole glucagon receptor antagonists. 2-(4-Pyridyl)-5-(4-chlorophenyl)-3-(5-bromo-2-propyloxyphenyl)pyrrole (L-168,049), a prototypical member of this series, inhibits binding of labeled glucagon to the human glucagon receptor with an IC50 = 3.7 ± 3.4 nm(n = 7) but does not inhibit binding of labeled glucagon-like peptide to the highly homologous human glucagon-like peptide receptor at concentrations up to 10 μm. The binding affinity of L-168,049 for the human glucagon receptor is decreased 24-fold by the inclusion of divalent cations (5 mm). L-168,049 increases the apparent EC50 for glucagon stimulation of adenylyl cyclase in Chinese hamster ovary cells expressing the human glucagon receptor and decreases the maximal glucagon stimulation observed, with aK b (concentration of antagonist that shifts the agonist dose-response 2-fold) of 25 nm. These data suggest that L-168,049 is a noncompetitive antagonist of glucagon action. Inclusion of L-168,049 increases the rate of dissociation of labeled glucagon from the receptor 4-fold, confirming that the compound is a noncompetitive glucagon antagonist. In addition, we have identified two putative transmembrane domain residues, phenylalanine 184 in transmembrane domain 2 and tyrosine 239 in transmembrane domain 3, for which substitution by alanine reduces the affinity of L-168,049 46- and 4.5-fold, respectively. These mutations do not alter the binding of labeled glucagon, suggesting that the binding sites for glucagon and L-168,049 are distinct.

The discovery of sulfonylated dipeptides as Potent VLA-4 antagonists

Directed screening of a carboxylic acid-containing combinatorial library led to the discovery of potent inhibitors of the integrin VLA-4. Subsequent optimization by solid-phase synthesis afforded a series of sulfonylated dipeptide inhibitors with structural components that when combined in a single hybrid molecule gave a sub-nanomolar inhibitor as a lead for medicinal chemistry. Preliminary metabolic studies led to the discovery of substituted biphenyl derivatives with low picomolar activities. SAR and pharmacokinetic characterization of this series are presented.

CHAPTER 24. INHIBITION OF MATRIX METALLOPROTEINASES

Publisher Summary This chapter briefly describes the metalloproteinase (MMP) family of enzymes and the putative role it plays in both physiology and pathology, the structures of these enzymes and the current status of MMP inhibitor development for the treatment of this wide range of diseases. The MMP family currently includes fourteen members encoded by unique genes, 10 of which are secreted from cells in a soluble form and 4 new members that are bound to the cell membrane. The MMPs are zinc dependent, calcium requiring enzymes, which are expressed as inactive zymogens. These enzymes are also inhibited by one or more members of the tissue inhibitor of metalloproteinase (TIMP) family, of which three have been cloned and sequenced. The physiological activators of many of these enzymes remain unknown; however, some of the members of the MMP family have the capacity to activate other family members. Therefore, inhibition of the enzyme(s), ultimately responsible for this activation cascade, could result in blocking the activity of multiple enzymes without directly inhibiting their activity. Recently four membrane type MMPs (MT-MMPs) have been cloned and sequenced. Two of these enzymes have the capacity to activate MMP-2 suggesting that they may be a target to consider for the intervention of tumor metastasis. These enzymes are unique among the MMPs because they appear to be associated with the cell membrane and not released into the extracellular space. Using MMP inhibitors, shedding of a number of cell associated proteins, including TNFα, IL-6 and tumor necrosis factors (TNF) receptors, FAS ligand, ACE, TSH receptor ectodomain, and CD-23, may be mediated by membrane associated MMPs.

Conformational Analysis and Receptor Docking of N-[(1S,2S)-3-(4-Chlorophenyl)-2-(3-cyanophenyl)-1-methylpropyl]-2-methyl-2-{[5-(trifluoromethyl)pyridin-2-yl]oxy}propanamide (Taranabant, MK-0364), a Novel, Acyclic Cannabinoid-1 Receptor Inverse Agonist

X-ray crystallographic, NMR spectroscopic, and computational studies of taranabant afforded similar low-energy conformers with a significant degree of rigidity along the C11-N13-C14-C16-C17 backbone but with more flexibility around bonds C8-C11 and C8-O7. Mutagenesis and docking studies suggested that taranabant and rimonabant shared the same general binding area of CB1R but with significant differences in detailed interactions. Similar to rimonabant, taranabant interacted with a cluster of aromatic residues (F(3.36)200, W(5.43)279, W(6.48)356, and Y(5.39)275) through the two phenyl rings and with F(2.57)170 and L(7.42)387 through the CF 3-Pyr ring. The notable distinction between taranabant and rimonabant was that taranabant was hydrogen-bonded with S(7.39)383 but not with K(3.28)192, while rimonabant was hydrogen-bonded with K(3.28)192 but not with S(7.39)383. The strong hydrogen bonding between the amide NH of taranabant and hydroxyl of S(7.39)383 was key to the superior affinity of taranabant to CB1R.

N-Aryl 2,6-Dimethoxybiphenylalanine Analogues as VLA-4 Antagonists

A series of N-arylated phenylalanine derivatives has been synthesized and has been shown to be potent inhibitors of the integrin VLA-4. N-phenyl and N-heteroaryl derivatives with hydrogen bond acceptors in the meta position demonstrated low nanomolar activity against VLA-4.