Stephen J. Medhurst

Novel histamine H3 receptor antagonists GSK189254 and GSK334429 are efficacious in surgically-induced and virally-induced rat models of neuropathic pain.

&NA; Several studies have implicated a potential role for histamine H3 receptors in pain processing, although the data are somewhat conflicting. In the present study we investigated the effects of the novel potent and highly selective H3 receptor antagonists GSK189254 (6‐[(3‐cyclobutyl‐2,3,4,5‐tetrahydro‐1H‐3‐benzazepin‐7‐yl)oxy]‐N‐methyl‐3‐pyridinecarboxamide hydrochloride) and GSK334429 (1‐(1‐methylethyl)‐4‐({1‐[6‐(trifluoromethyl)‐3‐pyridinyl]‐4‐piperidinyl}carbonyl)hexahydro‐1H‐1,4‐diazepine) in two rat models of neuropathic pain, namely the chronic constriction injury (CCI) model and the varicella‐zoster virus (VZV) model. Both GSK189254 (0.3, 3 and/or 10 mg/kg p.o.) and GSK334429 (1, 3 and 10 mg/kg p.o.) significantly reversed the CCI‐induced decrease in paw withdrawal threshold (PWT) measured using an analgesymeter and/or von Frey hairs. In addition, GSK189254 (3 mg/kg p.o.) and GSK334429 (10 mg/kg p.o.) both reversed the VZV‐induced decrease in PWT using von Frey hairs. We also investigated the potential site of action of this analgesic effect of H3 antagonists using autoradiography. Specific binding to H3 receptors was demonstrated with [3H]‐GSK189254 in the dorsal horn of the human and rat spinal cord, and in human dorsal root ganglion (DRG), consistent with the potential involvement of H3 receptors in pain processing. In conclusion, we have shown for the first time that chronic oral administration of selective H3 antagonists is effective in reversing neuropathic hypersensitivity in disease‐related models, and that specific H3 receptor binding sites are present in the human DRG and dorsal horn of the spinal cord. These data suggest that H3 antagonists such as GSK189254 and GSK334429 may be useful for the treatment of neuropathic pain.

Activation of CB1 and CB2 receptors attenuates the induction and maintenance of inflammatory pain in the rat.

&NA; The aim of the present study was to investigate the effects of cannabinoid agonists on established inflammatory hyperalgesia. We have compared the effects of pre‐administration versus post‐administration of a potent non‐selective cannabinoid agonist HU210 and a selective CB2 receptor agonist JWH‐133 on hindpaw weight bearing and paw oedema in the carrageenan model of inflammatory hyperalgesia. For comparative purposes we also determined the effects of the μ‐opioid receptor agonist morphine and the COX2 inhibitor rofecoxib in this model. At 3 h following intraplantar injection of carrageenan (2%, 100 μl) there was a significant (P<0.001) reduction in weight bearing on the ipsilateral hindpaw, compared to vehicle treated rats and a concomitant increase in ipsilateral hindpaw volume (P<0.001), compared to vehicle treated rats. Systemic administration of HU210 (10 μg/kg) and JWH‐133 (10 mg/kg) at 3 h following injection of carrageenan, significantly attenuated decreases in ipsilateral hindpaw weight bearing (P<0.05 for both) and paw volume (P<0.001 for both). Pre‐administration of HU210 and JWH‐133 had similar effects on weight bearing in this model. Pre‐administered HU210 also significantly decreased carrageenan‐induced changes in paw volume (P<0.001), this was not the case for JWH‐133. Effects of post‐administered HU210 and JWH‐133 on ipsilateral hindpaw weight bearing and paw volume were comparable to the effect of systemic post‐administration of morphine and rofecoxib (3 mg/kg for both). In summary, both HU210 and JWH‐133 attenuated established inflammatory hypersensitivity and swelling, suggesting that cannabinoid‐based drugs have clinical potential for the treatment of established inflammatory pain responses.

An animal model of chronic inflammatory pain: Pharmacological and temporal differentiation from acute models

Clinically, inflammatory pain is far more persistent than that typically modelled pre‐clinically, with the majority of animal models focussing on short‐term effects of the inflammatory pain response. The large attrition rate of compounds in the clinic which show pre‐clinical efficacy suggests the need for novel models of, or approaches to, chronic inflammatory pain if novel mechanisms are to make it to the market. A model in which a more chronic inflammatory hypersensitivity phenotype is profiled may allow for a more clinically predictive tool. The aims of these studies were to characterise and validate a chronic model of inflammatory pain. We have shown that injection of a large volume of adjuvant to the intra‐articular space of the rat knee results in a prolonged inflammatory pain response, compared to the response in an acute adjuvant model. Additionally, this model also results in a hypersensitive state in the presence and absence of inflammation. A range of clinically effective analgesics demonstrate activity in this chronic model, including morphine (3 mg/kg, t.i.d.), dexamethasone (1 mg/kg, b.i.d.), ibuprofen (30 mg/kg, t.i.d.), etoricoxib (5 mg/kg, b.i.d.) and rofecoxib (0.3–10 mg/kg, b.i.d.). A further aim was to exemplify the utility of this chronic model over the more acute intra‐plantar adjuvant model using two novel therapeutic approaches; NR2B selective NMDA receptor antagonism and iNOS inhibition. Our data shows that different effects were observed with these therapies when comparing the acute model with the model of chronic inflammatory joint pain. These data suggest that the chronic model may be more relevant to identifying mechanisms for the treatment of chronic inflammatory pain states in the clinic.

Activation of the α7-Nicotinic Acetylcholine Receptor Reverses Complete Freund Adjuvant–Induced Mechanical Hyperalgesia in the Rat Via a Central Site of Action

UNLABELLED The role of specific nicotinic receptor (nAChR) subtypes in antinociception has not been fully elucidated because of the lack, until recently, of selective tool compounds. (R)-N-(1-azabicyclo[2.2.2]oct-3-yl)(5-(2-pyridyl)thiopene-2-carboxamide) (compound B) is reported to be an agonist selective for the alpha(7)nAChR and in the present study was found to be efficacious in inflammatory pain models in 2 species. Compound B reversed complete Freund adjuvant-induced reductions in paw withdrawal thresholds in rat and mouse in a dose-related manner, producing maximum reversals of 65% +/- 4% at 10 mg/kg and 87% +/- 15% at 20 mg/kg. When rats and mice were predosed with the centrally penetrant, broad-spectrum nicotinic receptor antagonist mecamylamine, the efficacy of the agonist was significantly inhibited, producing reversals of only 11% +/- 5% at 10 mg/kg and 5% +/- 13% at 20 mg/kg, confirming activity via nicotinic receptors. Rats were also predosed systemically with the selective low-brain penetrant alpha(7)-antagonist methyllycaconitine, which had no effect on agonist activity (90% +/- 18% at 10 mg/kg), suggesting a central involvement. This hypothesis was further established with methyllycaconitine completely inhibited the agonist effect when dosed intrathecally (1% +/- 7%). PERSPECTIVE These studies provide good rationale for the utility of selective, central nervous system penetrant agonists at the alpha(7)-nicotinic receptor for the treatment of inflammatory pain.

Design and synthesis of 6-phenylnicotinamide derivatives as antagonists of TRPV1

6-Phenylnicotinamide (2) was previously identified as a potent TRPV1 antagonist with activity in an in vivo model of inflammatory pain. Optimization of this lead through modification of both the biaryl and heteroaryl components has resulted in the discovery of 6-(4-fluorophenyl)-2-methyl-N-(2-methylbenzothiazol-5-yl)nicotinamide (32; SB-782443) which possesses an excellent overall profile and has been progressed into pre-clinical development.

Discovery and structure-activity relationships of a series of pyroglutamic acid amide antagonists of the P2X7 receptor.

A computational lead-hopping exercise identified compound 4 as a structurally distinct P2X(7) receptor antagonist. Structure-activity relationships (SAR) of a series of pyroglutamic acid amide analogues of 4 were investigated and compound 31 was identified as a potent P2X(7) antagonist with excellent in vivo activity in animal models of pain, and a profile suitable for progression to clinical studies.

Identification of 2-oxo-N-(phenylmethyl)-4-imidazolidinecarboxamide antagonists of the P2X(7) receptor.

A backup molecule to compound 2 was sought by targeting the most likely metabolically vulnerable site in this molecule. Compound 18 was subsequently identified as a potent P2X(7) antagonist with very low in vivo clearance and high oral bioavailability in all species examined. Some evidence to support the role of P2X(7) in the etiology of pain is also presented.

Structure-activity relationships and in vivo activity of (1H-pyrazol-4-yl)acetamide antagonists of the P2X7 receptor

Structure-activity relationships (SAR) of analogues of lead compound 1 were investigated and compound 16 was selected for further study in animal models of pain. Compound 16 was shown to be a potent antihyperalgesic agent in both the rat acute complete Freunds adjuvant (CFA) model of inflammatory pain [Iadarola, M. J.; Douglass, J.; Civelli, O.; Naranjo, J. R. rain Res.1988, 455, 205] and the knee joint model of chronic inflammatory pain [Wilson, A. W.; Medhurst, S. J.; Dixon, C. I.; Bontoft, N. C.; Winyard, L. A.; Brackenborough, K. T.; De Alba, J.; Clarke, C. J.; Gunthorpe, M. J.; Hicks, G. A.; Bountra, C.; McQueen, D. S.; Chessell, I. P. Eur. J. Pain2006, 10, 537].

434 ANTINOCICEPTIVE EFFECTS OF A SELECTIVE ERBETA AGONIST THROUGH MODULATION OF THE ENDOGENOUS OPIOID SYSTEM

Background and Aims: The effects of estrogens on pain perception remain controversial. To investigate this GSK1522814A, a selective estrogen receptor beta ligand, was tested in rat models of inflammatory and acute pain. Mechanisms by which estrogens modulate pain remain unknown, however Smith et al. (2006; J. Neurosci 26: 5777) reported that estradiol influences opioid neurotransmission. To explore this further, GSK1522814A was dosed in the presence of the opioid antagonist naloxone, in both paradigms. Methods: Using rats previously dosed with CFA (100ml, intraplantar), weight bearing differences between hind limbs were measured before and after the administration of GSK1522814A (0.1–1mg/kg, p.o.) when administered alone, and in the presence of naloxone (1mg/kg, s.c.). In naïve rats, paw withdrawal thresholds to a mechanical stimulus were taken before and after the administration of GSK1522814A (1–10mg/kg, p.o.) when administered alone, and in the presence of naloxone (1mg/kg, s.c.). Results: GSK1522814A significantly reversed CFA-induced inflammatory pain at all doses (p < 0.001). In a separate study, a reversal of 87±13% at 1mg/kg was inhibited by naloxone to 4±10%. GSK1522814A (3+10mg/kg) also produced an anti-nociceptive response in naïve rats (p < 0.001). In a separate study, pre/postdose paw withdrawal threshold difference of ~125 g in rats dosed with GSK1522814A (3mg/kg) was reduced to ~15 g by naloxone. Conclusions: These results suggest that estrogens are involved in inflammatory pain and anti-nociception in rats via modulation of the endogenous opioid system. 435 ASSOCIATION OF HYDROMORPHONE AND PREGABALIN IN PHANTOM LIMB PAIN F. Cannata*, R. Bortone, G. Lunghi, M. Grio, P. Favaro, S. Del Monte, F. Debach, A. Canneti, M. Luzi, P. Di Marco, C. Reale. University “La Sapienza” Rome Dipartment of Anaesthesiology, Critical care and pain therapy, Rome, Italy