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The recent discovery of several nonpeptide CGRP antagonists have led to significant advances in our understanding of CGRP receptor pharmacology. Specifically, these antagonists have demonstrated a clear species selectivity with >100-fold greater affinity for human CGRP receptor compared to receptors from other species, such as rat, rabbit and guinea pig. Therefore, nonhuman primate models are required to accurately assess the in vivo activity of these antagonists. The commonly used model in marmosets involves electrical stimulation of the trigeminal ganglia and is a technically difficult and terminal procedure. In this report, we describe a noninvasive pharmacodynamic model in which topical application of capsaicin is utilized to induce the release of endogenous CGRP and a vasodilatory response which can be measured using laser Doppler imaging. Using the potent and selective CGRP antagonist Compound 3, which is an analog of the well-characterized compound BIBN4096BS, we demonstrated 62% inhibition with 300 microg/kg, i.v., in the rat. When tested in the rhesus monkey, only 30 microg/kg of Compound 3 was needed to produce complete inhibition, suggesting that the rhesus CGRP receptor shares a pharmacological profile similar to marmoset and human receptors. Two separate measurements were obtained in this model to provide an indication of both the acute inhibitory effect as well as the prophylactic effect of the CGRP antagonist. At the doses studied, Compound 3 was equally effective on both the acute and prophylactic inhibition of CGRP-mediated vasodilation in rat and rhesus. In conclusion, this is the first report to describe and validate a noninvasive model in nonhuman primates that allows rapid evaluation of CGRP antagonist activity against endogenous CGRP.

This study was designed to test the ability of adenovirus-delivered vascular endothelial growth factor (Ad-VEGF) to stimulate angiogenesis and arteriogenesis in the rabbit hindlimb following the induction of ischemia and to evaluate the functional changes in the collateral circulation. Ten days after the surgical induction of hindlimb ischemia, either a control virus (1 x 10(9) pfu) or an adenovirus containing the gene for VEGF(165) (1 x 10(6), 1 x 10(7), 1 x 10(8), or 1 x 10(9) pfu) was administered intramuscularly into the ischemic limb. Thirty days after administration of the adenoviral vectors, skeletal muscle capillary density was assessed and angiography was performed as markers of angiogenesis and arteriogenesis, respectively. Hindlimb blood flow was directly measured and hyperemic tests were performed to evaluate the functional improvements in collateral blood flow. Animals treated with Ad-VEGF at 1 x 10(8) and 1 x 10(9) pfu showed elevated levels of circulating VEGF and dose-dependent hindlimb edema. These doses also led to a robust angiogenic response (i.e., increase in capillary density), but failed to improve collateral blood flow. Consistent with the lack of a functional response, there was no angiographic evidence of enhanced arteriogenesis with any dose of Ad-VEGF. Following the induction of hindlimb ischemia, administration of Ad-VEGF stimulated capillary sprouting (i.e., angiogenesis), but did not increase the growth and development of larger conduit vessels (i.e., arteriogenesis) or improve collateral blood flow. These results support the concept that VEGF may not be expected to have therapeutic utility for the treatment of peripheral or myocardial ischemia.

As part of an ongoing effort to prepare therapeutically useful orally active thrombin inhibitors, we have synthesized a series of compounds that utilize nonbasic groups in the P1 position. The work is based on our previously reported lead structure, compound 1, which was discovered via a resin-based approach to varying P1. By minimizing the size and lipophilicity of the P3 group and by incorporating hydrogen-bonding groups on the N-terminus or on the 2-position of the P1 aromatic ring, we have prepared a number of derivatives in this series that exhibit subnanomolar enzyme potency combined with good in vivo antithrombotic and bioavailability profiles. The oxyacetic amide compound 14b exhibited the best overall profile of in vitro and in vivo activity, and crystallographic studies indicate a unique mode of binding in the thrombin active site.

Methanesulfonanilide Class III antiarrhythmic agents have been shown to block a specific outward delayed rectifier K+ current, Ikr in cardiac cells. K+ conductance also is recognized to be an important regular of contractile tone in vascular smooth muscle. The purpose of the present investigation was to assess the effects of the new and potent methanesulfoanilide Class III agents E-4031, UK-68,798, UK-66,914 and the Class III standard d-sotalol in vitro in phasically active and electrically quiescent vascular smooth muscle preparations. All four Class III agents augmented phasic contractile tension in spontaneously active rat portal veins at concentrations similar to those effecting significant Class III electrophysiologic activity in cardiac muscle, but failed to contract electrically quiescent rabbit aortic rings. At concentrations exceeding effective cardiac Class III electrophysiologic concentrations, E-4031 relaxed methoxamine- and histamine-contracted rabbit aortic rings, and d-sotalol relaxed methoxamine-contracted aortic rings. UK-68,798 and UK-66,914 failed to relax spasmogen-contracted aortic rings. The similarity in effective concentrations required for the four Class III agents to augment phasic contractile tension in the rat portal vein and increase myocardial refractoriness in cardiac muscle is consistent with the presence of similar K+ channel subtypes in the two tissues. Alternatively, the observed activities in the two tissues may be due to actions of these four Class III agents on another, non-Ikr ion channel present in rat portal vein, with an order of potency for blockade similar to block of Ikr in cardiac tissue. © 1992 wiley-Liss, Inc.

Summary 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.

The effects of E4031, a new class III antiarrhythmic agent similar to sotalol, were tested in isometrically contracting rabbit papillary muscles and in anesthetized, open-chest dogs. In papillary muscles, E4031 caused a modest dose-dependent increase of 26 +/- 8% in developed tension and 38 +/- 8% in its maximal rate of rise. Since there was no significant change in the maximal rate of relaxation, the ratio between both maximal velocities increased from 0.92 +/- 0.03 to 1.19 +/- 0.10. Time to peak tension did not change significantly, whereas time to half relaxation increased from 72 +/- 3 to 85 +/- 4 ms. The effective refractory period in the rabbit papillary muscles increased from 179 +/- 10 to 414 +/- 45 ms. In the open-chest dog, the i.v. administration of E4031 did not induce significant changes in heart rate, mean arterial pressure, or left ventricular end diastolic pressure. +dP/dt increased from 1,839 +/- 162 to 2,470 +/- 247 mm Hg/s with no significant change in -dP/dt after 100 micrograms/kg of E4031. Consequently, (+dP/dt)/(-dP/dt) increased from 0.97 +/- 0.07 to 1.18 +/- 0.08. To further evaluate the effects of E4031 on myocardial relaxation, the time constant of isovolumic left ventricular pressure decay was measured by two different methods (tau 1 and tau 2) before and after administering 10 micrograms/kg E4031. Tau 1 increased from 27 +/- 1.8 to 33 +/- 1.6 ms and tau 2 increased from 30 +/- 2.3 to 41 +/- 3.3 ms.(ABSTRACT TRUNCATED AT 250 WORDS)

1 (L-374,087) is a potent, selective, efficacious, and orally bioavailable thrombin inhibitor that contains a core 3-amino-2-pyridinone moiety. Replacement of the C6 pyridinone methyl group of 1 by a propyl group gave 5 (L-375,052), which retained all the excellent properties of 1, and also yielded higher plasma levels after oral dosing in dogs and rats.

We studied the effects of NE-10064 (azimilide), a new antiarrhythmic agent reported to be a selective blocker of the slowly activating component of the delayed rectifier, IKs. In ferret papillary muscles, NE-10064 increased effective refractory period (ERP) and decreased isometric twitch tension in a concentration-dependent manner (0.3-30 microM). Increases in ERP showed reverse use-dependence, and were greater at 1 than at 3 Hz. In contrast, changes in tension were use dependent, with larger decreases observed at 3 than at 1 Hz. In guinea pig ventricular myocytes, NE-10064 (0.3-3 microM) significantly prolonged action potential duration (APD) at 1 Hz. At 3 Hz, NE-10064 (0.3-1 microM) increased APD only slightly, and at 10 microM decreased APD and the plateau potential. NE-10064 potently blocked the rapidly activating component of the delayed rectifier, IKr (IC50 0.4 microM), and inhibited IKs (IC50 3 microM) with nearly 10-fold less potency. NE-10064 (10 microM) did not block the inward rectifier potassium current (IKl). NE-10064 (10 microM) blocked the L-type calcium current (ICa) in a use-dependent manner; block was greater at 3 than at 1 Hz. We conclude that (a) NE-10064's block of potassium currents is relatively selective for IKr over IKs, (b) NE-10064 inhibits ICa in a use-dependent fashion, and (c) NE-10064's effects on ERP and tension in papillary muscle as well as APD and action potential plateau level in myocytes may be explained by its potassium and calcium channel blocking properties.

Synthetic human tumor hypercalcemic factor (1–34, hHF) was compared with parathyroid hormone (human sequence, 1–34; hPTH) for vasorelaxant activity in isolated rabbit renal artery segments. The hHF exhibited a potent (IC 50 = 1.3×10 −9 M) and profound (98%) relaxation which was significantly greater in magnitude than that obtained for hPTH (IC 50 = 4.5×10 −9 M; maximal relaxation = 78%). The relaxations to both peptides were concentration-dependent and not associated with changes in cyclic AMP levels. These results demonstrate a parathyroid hormone-like response, independent of adenylate cyclase activation, in isolated renal arteries. Renal vasodilation may be important for the effects on renal function shared by these two peptides.

The effects of atriopeptins I and II on relaxation and cyclic GMP levels were studied on rat and rabbit aortas. Atriopeptin I was 2- and 100-fold less potent than atriopeptin II in causing relaxation of rat and rabbit aortas, respectively. The atriopeptin-elevated cyclic GMP levels generally correlated with the amount of relaxation. These results demonstrate that the vasodilator profile and, presumably, the receptor for atrial natriuretic factor, varies among different blood vessels and species.

Characterization of the vascular pharmacology and receptor binding of atrial natriuretic factor (ANF) has been achieved utilizing a synthetic peptide which contains the sequence and biological activity of ANF. The synthetic ANF (sANF) relaxes aortic segments contracted by agonists or by low (15 to 20 mM) but not high (greater than or equal to 40 mM) concentrations of K+. The relaxation to sANF is well maintained, reversible, independent of the vascular endothelium and correlated with increases in cyclic GMP (with no change in cyclic AMP). Plasma membranes prepared from rabbit aorta and kidney possess high affinity (Kd = 100 pM) specific binding sites for sANF. An excellent correlation exists between the receptor binding and pharmacology for several synthetic analogs of ANF. The presence of these receptors appears consistent with the activation of particulate (but not soluble) guanylate cyclase by sANF. sANF does exhibit a profound regional vasodilator selectivity which can be explained, in part, by changes in receptor density.