Discovery of {4-[4,9-bis(ethyloxy)-1-oxo-1,3-dihydro-2H-benzo[f]isoindol-2-yl]-2-fluorophenyl}acetic acid (GSK726701A), a novel EP4 receptor partial agonist for the treatment of pain.

A novel series of EP4 agonists and antagonists have been identified, and then used to validate their potential in the treatment of inflammatory pain. This paper describes these novel ligands and their activity within a number of pre-clinical models of pain, ultimately leading to the identification of the EP4 partial agonist GSK726701A.

Impact of chronic inflammation on the pharmacokinetic-pharmacodynamic relationship of naproxen.

OBJECTIVES: The use of biomarkers for predicting the clinical doses of analgesic drugs relies on the understanding of the relationship between drug exposure and response under disease conditions. In this study, we demonstrate the relevance of such a relationship for COX-inhibitors by modelling the effect of naproxen on prostaglandin E2 (PGE(2)) and thromboxane B2 (TXB(2)) in a chronic inflammation model in rats. METHODS: Rats were treated with Freund's complete adjuvant (FCA) by intraplantar injection. On post-inoculation days (PID) 7-21, animals received single or chronic (qd until day 21) doses of naproxen (10mg/kg). Blood samples were collected at various intervals after dosing to characterise naproxen pharmacokinetics and its effects on PGE(2) and TXB(2) production. PK-PD modelling was performed using nonlinear mixed effects in NONMEM. RESULTS: The inhibition of PGE(2) and TXB(2) could be described by a sigmoid E(max) model. A decrease in the potency estimates of both biomarkers was observed under chronic inflammation, as compared to healthy animals. IC(50) values for PGE(2) inhibition showed a shift from 2840+/-510 to 4000+/-677ng/ml(mean+/-SD), whilst IC(50) values for TXB(2) inhibition increased from 1180+/-323 to 3360+/-453ng/ml in healthy and FCA-inoculated animals, respectively. CONCLUSIONS: Our results show that chronic inflammation causes a significant change in the potency estimates for COX-inhibition. These findings illustrate the implications of pathophysiological processes on pharmacodynamics and consequently on the required exposure levels for achieving response during chronic treatment.

Differences in the sensitivity of behavioural measures of pain to the selectivity of cyclo-oxygenase inhibitors.

OBJECTIVES: Freund's complete adjuvant (FCA) is an animal model of inflammatory pain commonly used in the screening of COX-inhibitors. However, there is little understanding of how behavioural measures of the anti-inflammatory effect in the FCA model correlate to differences in mechanism of action and whether such endpoints equally reflect drug activity in humans. In the current investigation we evaluate the time course of the analgesic effect for different endpoints after treatment with drugs with varying degrees of selectivity for COX-1 and COX-2. We also assess prostaglandin (PGE(2)) and thromboxane (TXB(2)) inhibition to establish the correlation between behavioural measures and the degree of selectivity for COX-1 and COX-2. METHODS: Sprague-Dawley rats were treated with FCA by intra-plantar injection. On post-inoculation day (PID) 7, rats received a single oral dose of naproxen, diclofenac, ketorolac or rofecoxib. Drug treatment continued until PID 21. A control group received placebo only. Behavioural endpoints for inflammatory pain and blood samples for biomarkers were obtained at various time points before and after dosing to characterise the time course of drug effect and disease progression. RESULTS: COX-inhibitors showed no effect on the dynamic plantar test. In contrast, full analgesia was observed after drug administration for weight bearing capacity (WBC) and paw pressure (PP), with varying duration of the effect for each of the endpoints. No tolerance to drug effect was observed up to 14 days of chronic treatment. Rofecoxib showed an increase in baseline pain threshold values after chronic treatment, which may be related to its pharmacokinetic characteristics. CONCLUSIONS: Changes in paw pressure threshold seem to best reflect the anti-hyperalgesic properties of COX-inhibitors with enough sensitivity to enable estimation of the dose-exposure-response curve.

1,5-Biaryl pyrrole derivatives as EP1 receptor antagonists: Structure-activity relationships of 4- and 5-substituted benzoic acid derivatives.

This paper details the SAR of 1,5-biaryl pyrrole derivatives with substituents in the 2-, 4-, and 5-positions of the benzoic acid group as EP1 receptor antagonists. Substitution at the 2-position was poorly tolerated, whereas only fluorine was tolerated at the 4-position. In contrast, a range of substituents at the 5-position were discovered which enhanced the in vitro affinity and led to compounds with promising oral exposure. Three derivatives showed efficacy in a preclinical model of inflammatory pain when dosed orally to rats.

1,5-Biaryl pyrrole derivatives as EP1 receptor antagonists. Structure-activity relationships of 6-substituted and 5,6-disubstituted benzoic acid derivatives.

Herein we describe the SAR of 1,5-biaryl pyrrole derivatives, with substituents in the 6-position of the benzoic acid moiety, as EP(1) receptor antagonists. Substitution at this position was well tolerated and led to the identification of several analogues with high affinity for the EP(1) receptor that displayed good efficacy in the established FCA model of inflammatory pain. Furthermore, several analogues were prepared which combined substitution at the 5- and 6-positions as well as derivatives with an aromatic ring fused to the 5- and 6-positions.

Pyridone derivatives as potent, orally bioavailable VLA-4 integrin antagonists.

A series of pyridone-N-benzyl-propanoic acids have been optimised to afford potent orally bioavailable VLA-4 antagonists.

Structure-activity relationships of 1,5-biaryl pyrroles as EP1 receptor antagonists.

The preliminary SAR of a series of novel 1,5-biaryl pyrrole EP1 receptor antagonists derived from compound 1 is described. Replacement of the benzyl group of 1 with isosteric groups was investigated. The most effective replacement was found to be the isobutyl group. The cyclopentylmethyl and cyclohexylmethyl groups were also effective benzyl replacements. The cyclohexylmethyl derivative 19 demonstrated the lowest metabolic clearance within this series. In addition, several high affinity substituted benzyl analogues were also identified. Compound 39 was found to have good bioavailability in rats and demonstrated efficacy in the established FCA preclinical model of inflammatory pain with a calculated ED50 of 9.2mg/kg.

Correlation between in vitro and in vivo concentration-effect relationships of naproxen in rats and healthy volunteers.

Understanding the mechanisms underlying the analgesic effect of new cyclooxygenase inhibitors is essential to identify dosing requirements in early stages of drug development. Accurate extrapolation to humans of in vitro and in vivo findings in preclinical species is needed to optimise dosing regimen in inflammatory conditions. The current investigation characterises the inhibition of prostaglandin E2 (PGE(2)) and thromboxane B2 (TXB(2)) by naproxen in vitro and in vivo in rat and human blood. The inhibition of PGE(2) in the absence or presence of increasing concentrations of naproxen (10(-8)-10(-1) M) was measured by ex vivo whole blood stimulation with LPS, whereas inhibition of TXB(2) was measured in serum following blood clotting. In further experiments, inhibition of PGE(2) and TXB(2) levels was also assessed ex vivo in animals treated with naproxen (2.5, 10, 25 mg kg(-1)). Subsequently, pharmacokinetic (PK)/pharmacodynamics (PD) modelling of in vitro and in vivo data was performed using nonlinear mixed effects in NONMEM (V). Inhibition of PGE(2) and TXB(2) was characterised by a sigmoid E(max) model. The exposure-response relationships in vitro and in vivo were of the same order of magnitude in both species. IC(80) estimates obtained in vitro were similar for PGE(2) inhibition (130.8 +/- 11 and 131.9 +/- 19 10(-6) M, mean +/- s.d. for humans and rats, respectively), but slightly different for TXB(2) inhibition (103.9+/-15 and 151.4 +/- 40 10(-6) M, mean +/- s.d. for humans and rats, respectively, P < 0.05). These differences, however, may not be biologically relevant. The results confirm the value of exposure-effect relationships determined in vitro as a means to predict the pharmacological activity in vivo. This analysis also highlights the need to parameterise concentration-effect relationships in early drug development, as indicated by the estimates of IC(80) for PGE(2) and TXB(2) inhibition.

Discovery of novel biaryl heterocyclic EP1 receptor antagonists.

We describe the generation of novel EP(1) receptor antagonists by investigation of thiophene isosteres. In addition, we disclose preliminary in vitro and in vivo DMPK for selected compounds.

Improving the in vitro prediction of in vivo central nervous system penetration: integrating permeability, P-glycoprotein efflux, and free fractions in blood and brain.

This work examines the inter-relationship between the unbound drug fractions in blood and brain homogenate, passive membrane permeability, P-glycoprotein (Pgp) efflux ratio, and log octanol/water partition coefficients (cLogP) in determining the extent of central nervous system (CNS) penetration observed in vivo. The present results demonstrate that compounds often considered to be Pgp substrates in rodents (efflux ratio greater than 5 in multidrug resistant Madin-Darby canine kidney cells) with poor passive permeability may still exhibit reasonable CNS penetration in vivo; i.e., where the unbound fractions and nonspecific tissue binding act as a compensating force. In these instances, the efflux ratio and in vitro blood-brain partition ratio may be used to predict the in vivo blood-brain ratio. This relationship may be extended to account for the differences in CNS penetration observed in vivo between mdr1a/b wild type and knockout mice. In some instances, cross-species differences that might initially seem to be related to differing transporter expression can be rationalized from knowledge of unbound fractions alone. The results presented in this article suggest that the information exists to provide a coherent picture of the nature of CNS penetration in the drug discovery setting, allowing the focus to be shifted away from understanding CNS penetration toward the more important aspect of understanding CNS efficacy.