Harold G. Selnick

First Demonstration of Cerebrospinal Fluid and Plasma Aβ Lowering with Oral Administration of a β-Site Amyloid Precursor Protein-Cleaving Enzyme 1 Inhibitor in Nonhuman Primates

β-Site amyloid precursor protein (APP)-cleaving enzyme (BACE) 1 cleavage of amyloid precursor protein is an essential step in the generation of the potentially neurotoxic and amyloidogenic Aβ42 peptides in Alzheimers disease. Although previous mouse studies have shown brain Aβ lowering after BACE1 inhibition, extension of such studies to nonhuman primates or man was precluded by poor potency, brain penetration, and pharmacokinetics of available inhibitors. In this study, a novel tertiary carbinamine BACE1 inhibitor, tertiary carbinamine (TC)-1, was assessed in a unique cisterna magna ported rhesus monkey model, where the temporal dynamics of Aβ in cerebrospinal fluid (CSF) and plasma could be evaluated. TC-1, a potent inhibitor (IC50 ∼ 0.4 nM), has excellent passive membrane permeability, low susceptibility to P-glycoprotein transport, and lowered brain Aβ levels in a mouse model. Intravenous infusion of TC-1 led to a significant but transient lowering of CSF and plasma Aβ levels in conscious rhesus monkeys because it underwent CYP3A4-mediated metabolism. Oral codosing of TC-1 with ritonavir, a potent CYP3A4 inhibitor, twice daily over 3.5 days in rhesus monkeys led to sustained plasma TC-1 exposure and a significant and sustained reduction in CSF sAPPβ, Aβ40, Aβ42, and plasma Aβ40 levels. CSF Aβ42 lowering showed an EC50 of ∼20 nM with respect to the CSF [TC-1] levels, demonstrating excellent concordance with its potency in a cell-based assay. These results demonstrate the first in vivo proof of concept of CSF Aβ lowering after oral administration of a BACE1 inhibitor in a nonhuman primate.

In Vivo Quantification of Calcitonin Gene-Related Peptide Receptor Occupancy by Telcagepant in Rhesus Monkey and Human Brain Using the Positron Emission Tomography Tracer [11C]MK-4232

Calcitonin gene-related peptide (CGRP) is a potent neuropeptide whose agonist interaction with the CGRP receptor (CGRP-R) in the periphery promotes vasodilation, neurogenic inflammation and trigeminovascular sensory activation. This process is implicated in the cause of migraine headaches, and CGRP-R antagonists in clinical development have proven effective in treating migraine-related pain in humans. CGRP-R is expressed on blood vessel smooth muscle and sensory trigeminal neurons and fibers in the periphery as well as in the central nervous system. However, it is not clear what role the inhibition of central CGRP-R plays in migraine pain relief. To this end, the CGRP-R positron emission tomography (PET) tracer [11C]MK-4232 (2-[(8R)-8-(3,5-difluorophenyl)-6,8-[6-11C]dimethyl-10-oxo-6,9-diazaspiro[4.5]decan-9-yl]-N-[(2R)-2′-oxospiro[1,3-dihydroindene-2,3′-1H-pyrrolo[2,3-b]pyridine]-5-yl]acetamide) was discovered and developed for use in clinical PET studies. In rhesus monkeys and humans, [11C]MK-4232 displayed rapid brain uptake and a regional brain distribution consistent with the known distribution of CGRP-R. Monkey PET studies with [11C]MK-4232 after intravenous dosing with CGRP-R antagonists validated the ability of [11C]MK-4232 to detect changes in CGRP-R occupancy in proportion to drug plasma concentration. Application of [11C]MK-4232 in human PET studies revealed that telcagepant achieved only low receptor occupancy at an efficacious dose (140 mg PO). Therefore, it is unlikely that antagonism of central CGRP-R is required for migraine efficacy. However, it is not known whether high central CGRP-R antagonism may provide additional therapeutic benefit.

Pharmacological Properties of MK-3207, a Potent and Orally Active Calcitonin Gene-Related Peptide Receptor Antagonist

Calcitonin gene-related peptide (CGRP) has long been hypothesized to play a key role in migraine pathophysiology, and the advent of small-molecule antagonists has clearly demonstrated a clinical link between blocking the CGRP receptor and migraine efficacy. 2-[(8R)-8-(3,5-Difluorophenyl)-10-oxo-6,9-diazaspiro[4.5]dec-9-yl]-N-[(2R)-2′-oxo-1,1′,2′,3-tetrahydrospiro[indene-2,3′-pyrrolo[2,3-b]pyridin]-5-yl]acetamide (MK-3207) represents the third CGRP receptor antagonist to display clinical efficacy in migraine trials. Here, we report the pharmacological characterization of MK-3207, a potent and orally bioavailable CGRP receptor antagonist. In vitro, MK-3207 is a potent antagonist of the human and rhesus monkey CGRP receptors (Ki = 0.024 nM). In common with other CGRP receptor antagonists, MK-3207 displays lower affinity for CGRP receptors from other species, including canine and rodent. As a consequence of species selectivity, the in vivo potency was assessed in a rhesus monkey pharmacodynamic assay measuring capsaicin-induced changes in forearm dermal blood flow via laser Doppler imaging. MK-3207 produced a concentration-dependent inhibition of dermal vasodilation, with plasma concentrations of 0.8 and 7 nM required to block 50 and 90% of the blood flow increase, respectively. The tritiated analog [3H]MK-3207 was used to study the binding characteristics on the human CGRP receptor. [3H]MK-3207 displayed reversible and saturable binding (KD = 0.06 nM), and the off-rate was determined to be 0.012 min−1, with a t1/2 value of 59 min. In vitro autoradiography studies on rhesus monkey brain slices identified the highest level of binding in the cerebellum, brainstem, and meninges. Finally, as an index of central nervous system penetrability, the in vivo cerebrospinal fluid/plasma ratio was determined to be 2 to 3% in cisterna magna-ported rhesus monkeys.

Discovery and X-ray Crystallographic Analysis of a Spiropiperidine Iminohydantoin Inhibitor of β-Secretase‡

A high-throughput screen at 100 microM inhibitor concentration for the BACE-1 enzyme revealed a novel spiropiperidine iminohydantoin aspartyl protease inhibitor template. An X-ray cocrystal structure with BACE-1 revealed a novel mode of binding whereby the inhibitor interacts with the catalytic aspartates via bridging water molecules. Using the crystal structure as a guide, potent compounds with good brain penetration were designed.

Cardiac electrophysiologic and inotropic actions of new and potent methanesulfonanilide class III antiarrhythmic agents in anesthetized dogs.

The effects of cumulative intravenous (i.v.) administration of potent and selective methanesulfonanil-ide class III antiarrhythmic agents on cardiac electrophysiologic and hemodynamic parameters were compared with those of D-sotalol in chloralose-anesthetized dogs. The new class III agents tested were E-4031 (l-(2-(6-methyl-2-pyridyl)ethyl)-(4-methanesulfonamidobenzo-yDpiperidine]; UK-66,914 [N -(4-< l-hydroxy-2-(4-(4-pyri-dinyl)-l-piperazinyl)ethyl)phenyl)methanesulfonamide], and UK-68,798 [l-(4-methanesulfonamidophenoxy)-2-(N -(4-methanesulfonamidophenethy I )-yV-methylamino (ethane]. The class III agents produced significant and dose-dependent increases in ventricular refractoriness, with effective doses required to increase ventricular relative refractory period 20 ms above baseline (ED20. (μg/kg i.v., with 959? confidence limits) of 5.2 (4.2–6.6) for UK-68,798. 17 (13–23) for E-4031. 75 (58–99) for UK-66,914, and 3.700 (2.600–5,800) for D-sotalol. Significant increases in the electrocardiographic QT and QTc intervals paralleled the increases in ventricular refractoriness for the four class III agents. Significant increases in left ventricular (LV) + dP/dt also paralleled increases in ventricular refractoriness and QT intervals for E-4031 (10–1,000 μg/kg i.v.), UK-66,914 (100–1,000 μ,g/kg i.v.). and UK-68.798 (30–1.000 μg/kg i.v.). but not for D-sotalol. No concomitant alterations in LV – dPIdt were observed for the new and potent methanesulfonanilide class III agents. resulting in significant increases in the ratio of LV + dPI dtl-dPldt for E-4031, UK-66.914, and UK-68,798. Potent and selective methanesulfonanilide class III agents therefore may augment cardiac contractility in addition to prolonging ventricular refractoriness.

A facile three-step synthesis of 1,2-amino alcohols using the Ellman homochiral tert-butylsulfinamide

Abstract Addition of organometallic reagents to tert -butylsulfinimines derived from tert -butyldimethylsiloxyacetaldehyde stereoselectively generates protected 1,2-amino alcohols. Removal of the acid labile protecting groups affords amino alcohols in high yield. The predominant diastereomer is opposite to that predicted by the traditional Ellman model; therefore, a chelation model invoking rapid E / Z isomerization of the imine is proposed to rationalize the observed selectivity.

Effects of new and potent methanesulfonanilide class III antiarrhythmic agents on myocardial refractoriness and contractility in isolated cardiac muscle

The effects of the new and potent methanesulfonanilide class III antiarrhythmic agents (E-4031, UK-66,914, and UK-68,798) on myocardial refractoriness and contractility were compared to those of d-sotalol in ferret isometrically contracting right ventricular papillary muscle preparations. During 1 Hz pacing at 37°C, the four class III agents elicited concentration-dependent increases in ventricular effective refractory period (ERP), with a relative order of potency of UK-68,798 > E-4031 > UK-66,914 > d-sotalol. EC25 values (effective concentration required to increase ERP 25% above baseline) were (in μM) UK-68,798. 0.018; E-4031, 0.058; UK-66,914, 0.501; and d-sotalol, 43.76. Maximal increases in ERP relative to baseline (% of baseline value) for the class III agents at 37°C (range of 44.5 ± 4.5 to 63.0 ± 3.1%) were greater than the maximal increases observed at 27°C (range of 15.0 ± 3.3 to 31.2 ± 4.8%), whereas the maximal absolute (ms) increases in ERP above baseline were comparable for the class III agents at both temperatures. Increases in ERP produced by the four class III agents at 37°C were significantly greater at a pacing frequency of 1 Hz (range of 70.0 ± 7.6 to 102.0 ± 2.3 ms) than at 3 Hz (range of 18.3 ± 4.4 to 31.0 ± 4.8 ms). During a temporary period of hypoxic perfusion at 37°C, increases in ERP produced by the four class III agents were reversed, such that “hypoxic” ERP values approximated pretreatment, baseline values. During 1 Hz pacing at 37°C, modest increases in developed tension (range of + 18 ± 8 to +27 ± 8% above baseline), with balanced increases in the rates of tension development and decline, were observed with the administrations of E-4031, UK-66,914, and UK-68,798. In contrast, d-sotalol produced minimal effects on myocardial contractility.

Suppression of lethal ischemic ventricular arrhythmias by the class III agent E4031 in a canine model of previous myocardial infarction

The antiarrhythmic efficacy of a new and potent class III agent E4031 [1-[2-(6-methyl-2-pyridyl)-ethyl]-4-(4-methylsulfonylaminobenzoyl)piperidine] was evaluated in several canine models of recent myocardial infarction. In anesthetized dogs with baseline inducible ventricular arrhythmias studied 4–10 days after anterior myocardial infarction, 30–300 μg/kg i.v. E4031 sup-pressed induction of ventricular tachyarrhythmias by programmed ventricular stimulation in 7 of 10 animals tested, while significantly prolonging refractoriness in both noninfarcted and infarcted ventricular myocardium. The incidence of lethal ischemic ventricular arrhythmias developing in response to acute posterolateral myocardial ischemia in the presence of previous anterior infarction was reduced from 10 of 10 (100%) in a vehicle pretreatment group to 3 of 10 (30%, p < 0.01) in an E4031 (300 μg/kg intravenously, i.v.) pretreatment group. Neither the sizes of the underlying anterior myocardial infarctions (26.9 ± 3.7 vs. 33.2 ± 2.1% of left ventricle) nor the times to development of acute posterolateral myocardial ischemia (43 ± 11 vs. 40 ± 8 min) differed significantly between the vehicle and E4031 pretreatment groups, respectively, suggesting that the reduction in the incidence of lethal ischemic arrhythmias in the E4031 pretreatment group was not due to smaller underlying, electrically unstable myocardial substrates nor to a delay in onset of the acute ischemic insult. In conscious dogs with spontaneous ventricular ectopy at 48 h after myocardial infarction and in anesthetized dogs with no baseline inducible arrhythmias at 4–10 days after myocardial infarction, E4031 (30–3,000 μg/kg i.v.) produced no facilitation or aggravation of spontaneous or inducible ventricular arrhythmias. These findings suggest that pharmacologic agents such as E4031 that increase ventricular refractoriness (class III electrophysiologic activity) may provide significant protection against development of malignant ischemic ventricular arrhythmias in the setting of previous myocardial infarction.

Selective α1a adrenergic receptor antagonists based on 4-aryl-3,4-dihydropyridine-2-ones

A series of alpha1a receptor antagonists derived from a 4-aryl-3,4-dihydropyridine-2-one heterocycle is disclosed. Potency in the low nanomolar to picomolar range along with high selectivity was obtained. In vivo efficacy in a prostate contraction model in rats was observed with a few derivatives.

Pharmacokinetic optimization of 3-amino-6-chloropyrazinone acetamide thrombin inhibitors. Implementation of P3 pyridine N-oxides to deliver an orally bioavailable series containing P1 N-benzylamides.

In this manuscript we demonstrate that a modification principally directed toward the improvement of the aqueous solubility (i.e., introduction a P3 pyridine N-oxide) of the previous lead compound afforded a new series of potent orally bioavailable P1 N-benzylamide thrombin inhibitors. An expedited investigation of the P1 SAR with respect to oral bioavailability, plasma half-life, and human liver microsome stability revealed 5 as the best candidate for advanced evaluation.