MXPA04011206A - Treatment of neuropathic pain. - Google Patents

Abstract

A method for treating a patient suffering from neuropathic pain, comprising administering to a patient in need of such treatment an effective amount of an agonist drug capable of binding to the neuronal nicotinic receptor (NNR) but which does not readily cross the blood-brain barrier.

Description

TREATMENT OF NEUROPATHIC PAIN TECHNICAL FIELD The present invention refers to a new treatment of neuropathic pain. Specifically, the invention relates to a method for treating a patient suffering from neuropathic pain, including allodynia and the reduction of side effects associated with the activation of a combatant of central neuronal nicotinic receptors.

BACKGROUND OF THE INVENTION Pain is a sensation and a perception that is comprised of a series of mechanisms complexes. In its simplest construction, it is a signal of the ignition of nociception, receptors of touch and pressure in the periphery that is transmitted to the spinal cord and finally to the centers, inferior and superior, of the brain. However, this signal can be modified in a multitude of ways at each level of the pain trajectory. (See, for example, Millan, M.J. (1999) The Induction of Pain: An Integrative Review, Progress in Neurobiology, 57, 1-164 (Pergamon Press) for a thorough review). There are mainly three types of pain. Acute pain, called nociception, is the instantaneous onset of a painful sensation in response to a noxious stimulus. It is considered to be adaptive because it can prevent an organism from harming itself. For example, removing a hand from a hot stove as quickly as the pain is felt will prevent serious burns. The second type of pain is persistent pain. Unlike acute pain, it usually has a delayed onset but can last from hours to days. It is considered predominantly adaptive because the occurrence of persistent pain following the injury can prevent further damage to the tissue. For example, the pain associated with a sprained ankle will prevent the patient from using the foot in such a way as to prevent further trauma and help with healing. The final category of pain is chronic pain. It has a delayed onset and can last from months to years. In contrast to persistent and acute pain, chronic pain is considered poorly adaptive and is associated with conditions such as arthritis, nerve injury, AIDS and diabetes. Neuropathic or chronic pain occurs in a variety of forms including spontaneous pain (painful sensation without an external stimulus), allodynia (painful sensation in response to a normally innocuous stimulus) and hyperalgesia (strong painful sensation to a mildly painful stimulus). It may be this diversity of symptoms that has made this condition difficult to treat clinically. In fact, the current treatments are predominantly the use of inactive label of antidepressants and anticonvulsants. However, both antidepressants and anticonvulsants present problems for the patient. Tricyclic antidepressants have the longest history of use in the treatment of neuropathic pain. Such drugs typically target the serotonergic and noradrenergic systems and increase the extracellular levels available of both serotonin and norepinephrine. It has been proposed that the postsynaptic activation of a2-adrenoreceptors by norepinephrine may be the mechanism through which these compounds reduce neuropathic pain. However, since these antidepressants easily cross the blood-brain barrier, their ability to increase levels of serotonin and norepinephrine can cause unwanted activation of other receptors that lead to a high risk of centrally mediated side effects. Side effects of antidepressants vary from medium but symptoms such as dry mouth and sedation are irritated to aggravate life threatening side effects such as postural hypotension and cardiac arrhythmia. The elderly, who represent a large number of neuropathic patients, are particularly vulnerable to the more serious side effects of antidepressants. The effectiveness of anticonvulsants in the treatment of several pain states, including neuropathic pain, has been recently evaluated (McQuay et al. (1995) Anticonvulsant Drugs for The Management of Pain: A Systematic Review, British Medical Journal 311, 1047- 52). Similar to antidepressants, side effects often occur with these medications.

Due to the common occurrence of side effects with antidepressants and anticonvulsants and the limitations of these located on the use of these compounds, there is a need for treatment for neuropathic pain that does not fall on antidepressants or anticonvulsants. In addition, there is a need for a treatment for neuropathic pain that avoids centrally mediated side effects.

BRIEF DESCRIPTION OF THE INVENTION In one aspect of the invention, a method for treating a patient suffering from neuropathic pain is provided by administering to a patient in need of such treatment an effective amount of a combatant drug of neuronal nicotinic receptor (NN) capable of of joining peripheral NNRs but that does not easily cross the blood-brain barrier. In another aspect of the invention, there is provided a method for treating a patient suffering from allodynia, said method comprising administering to a patient in need of such treatment an effective amount of a combatant drug capable of binding to at least peripheral NNR that does not cross easily the blood-brain barrier in doses for which an analgesic effect is observed. In yet a further aspect of the invention, there is provided a method for treating a patient suffering from neuropathic pain, said method comprising administering to a patient in need of such treatment an effective amount of a quaternary nicotinic compound. In yet another aspect of the invention, there is provided a method for treating a patient suffering from neuropathic pain by performing peripheral NNRs at the level of the dorsal root ganglion (DRG) by administering to a patient in need of such treatment an effective amount of a nicotinic combatant compound that does not easily cross the blood-brain barrier. In still another aspect of the invention, there is provided a method for reducing the side effects associated with combatant activation of central NNRs, said method comprising administering to a patient in need of such treatment an effective amount of a non-crossing nicotinic combatant compound. easily the blood-brain barrier. DESCRIPTION OF THE DRAWINGS The following is a description of the figures presented for the purposes of illustration of the invention and not for purposes of limitation thereof. FIG. 1 shows the effect of systemically administered chlorisondamine pretreatment (0.4 μg / kg, ip 30 minutes before) on antinociception induced by systemically administered A-85380 (0.75 μg / kg, ip) as measured for the increase in claw extraction latency (average + sem) in the rat claw extraction model of acute thermal pain using normal rats. The averages represent the average of the left and right claw records collapsed through measurements of 1, 30 and 45 minutes in order to show the main dose effect. Five rats were used in each group. * p < 0.05 vs. saline / saline FIG. 2A shows the effect of pretreatment of chlorisondamine (10 μg 24 hours before) i.c.v. (intracerebroventricular) on analgesia induced by systemically administered A-85380 (0.75 μg / kg, i .p.9 as measured by the reduction in quarters (average + sem) in the persistent pain formalin model. Five rats were used in each group, Figure 2B shows the effect of pretreatment of chlorisondamine (0.4 μg / kg ip 45-50 minutes before) systemically administered on analgesia induced by systemically administered A-85380 (0.75 μGt). ??? / kg, i.p.) as measured by the reduction in quarters (average + sem) in the persistent pain formalin model.The quarters were measured during 20 minutes during phase 2. Six to seven rats were used in each group. * p < 0.05 vs. saline / saline Fig. 3 shows the effects of multiple doses of fighter NNR A-85380 on the start of claw extraction (average + sem) in a mechanical allodynia test in the model of nerve ligation esp of neuropathic pain. The start of claw extraction (gram force) was determined using the von Frey filament stimulation and the Dixon ascending-descending method. The measurements were taken before the combatant's injection (baseline) and at 15, 30, 60 and 120 minutes after the combatant injection. Six rats were used in each dose. FIG. 4 shows the effect of mecamylamine pretreatment (1 μ? T? / Kg, ip 30 minutes before) systemically administered on anti-allodynia induced by systemically administered A-85380 (0.75 μ ???? / kg, ip) according to it is measured by the increase in the beginning of claw extraction (average + sem) for the mechanical stimulation in the rat spinal nerve ligation model of neuropathic pain. The averages represent the beginnings of extraction collapsed through measurements of 15, 30 and 60 minutes in order to show the main treatment effect. Six rats were used in each group. * p <; 0.05 vs. saline / saline FIG. 5A shows the effect of pre-treatment of chlorisondamine (10 μg 24 hours before) intra-ventricular-ventricular (icv) on anti-allodynia induced by systemically administered A-85380 (0.75 μg / kg, ip) as measured by the increase in the start of claw extraction (average + sem) for mechanical stimulation in the rat spinal nerve ligation model of neuropathic pain. FIG. 5B shows the effect of pretreatment of chlorisondamine (0.4 μg / kg, ip 30 minutes before) systemically administered on anti-allodynia induced by systemically administered A-85380 (0.75 μg / kg, ip) as measured by the increase in the beginning of claw extraction (average + sem) for the mechanical stimulation in the rat nerve spinal nerve ligation model of neuropathic pain. The averages represent the beginnings of extraction collapsed through measurements of 15, 30 and 60 minutes in order to show the main treatment effect. Four to six rats were used in each group. * p < 0.05 vs. saline / saline The FI G. 6 shows the effect of pretreatment of hexamethonium (19 μg / kg, ip 30 minutes before) systemically administered on anti-allodynia induced by systemically administered A-85380 (0.75 mol / kg, i.p.) as measured by the increase in the beginning of claw extraction (average + sem) for the mechanical stimulation in the rat nerve spinal nerve ligation model of neuropathic pain. The averages represent the beginnings of extraction collapsed through measurements of 15, 30 and 60 minutes in order to show the main treatment effect. Five rats were used in each group. * p < 0.05 vs. saline / saline FI G. 7 shows the effect of chlorisondamine pretreatment (0.4 μg / kg, i.p., 30 minutes before) systemically administered on antinociception induced by systemically administered A-85380 (0.75 μ? T ??? kg, ip) as measured by the increase in claw extraction latency (average + sem) in the rat claw extraction model of acute thermal pain using neutropathic rats. The averages represent the average of the left and right claw records collapsed through measurements of 1, 5, 30 and 45 minutes in order to show the main treatment effect. Six rats were used in each group. * p < 0.05 vs. saline / saline FIG. 8A shows the effect of multiple doses of A-85380 injected towards the plantar surface of the rear claw on the affected side (ipsalateral) on the beginning of claw extraction (average + sem) for the mechanical stimulation of the neuropathic claw in the model of the nerve spinal nerve ligation of neuropathic pain. The F IG. 8B shows the effect of multiple doses of A-85830 injected into the plantar surface of the rear claw on the unaffected side (contralateral) on the affected side (ipsalateral) on the start of claw extraction (average +. Sem) for the Mechanical stimulation of the neuropathic claw in the rat spinal nerve ligation model of neuropathic pain. The measurements were taken before the combatant injection (baseline) and at 1 5, 30, 60 and 1 20 minutes after the combatant injection. Six rats were used in each dose / claw combination. * p < 0.05 vs. saline at the same time point. The F IG. 9A shows the effect of different doses of infused A-85830 on the L5 DRG on the start of claw extraction (averaged io + sem) for mechanical stimulation in the spinal nerve ligation model of neuropathic pain rat. The measurements were taken before the combatant injection (baseline) and at 1 5, 30, and 60 minutes after the combatant injection. Six rats were used in each dose / claw combination. * p < 0.05 vs. saline at the same time point.

FIG. 9B shows the effect of pretreatment of chlorisondamine (0.4 μ? T / kg, ip 30 minutes before) systemically administered on the anti-allodynia induced by A-85830 infused on the DRG (20 μg) as measured by the increase in the beginning of claw extraction (average + sem) for the mechanical stimulation in the spinal nerve ligation model of neuropathic pain rat. The averages represent the beginnings of extraction collapsed through measurements of 15, 30 and 60 minutes in order to show the main treatment effect. Six rats were used in each group. * p < 0.05 vs. saline / saline FIG. 10 shows the comparison of the effects of A-85830 administered either i.p., i.d. in the affected claw or on the L5 DRG on the beginning of claw extraction (average + sem) for the mechanical stimulation in the rat spinal nerve ligation model of neuropathic pain. The doses of A-85380 administered i.d. or DRG were converted into systemic doses based on the body weight of each rat and were represented by graphs as the average equivalent to systemic dose. The averages of the beginning of claw extraction represent the time point of only fifteen minutes. FIG. 11 shows the comparison of the effects of A-85830 administered alone or with chlorisondamine administered i.p. or i.c.v. on the start of claw removal of neuropathic rats.

DETAILED DESCRIPTION The term used to describe pain reduction is analgesia (antinociception, anti-allodynia), which can be described as the reduction of pain and, from the perspective of this invention, the reduction of pain associated with neuropathic pain, as well as also allodynia is often associated with neuropathic pain. The term "combatant" is defined as a compound that shows 30% or more of efficacy in a functional assay compared to the endogenous binder acetylcholine or the exogenous binder nicotine. The terms "substantially unable to cross the brain blood barrier" and "not readily able to cross the brain blood barrier" refer to the inability of the compounds to cross the blood-brain barrier and activate the central NNRs in doses that they are fully capable of activating NNRs in the periphery. This invention provides a method for treating neuropathic pain in a patient that requires such treatment while minimizing the risk of centrally mediated side effects in the patient. The inventors have made the surprising discovery that neuropathic pain can be lessened, at least in part, by the combatant's union to the peripheral NNR, where the combatant drug is substantially unable to penetrate the blood-brain barrier thereby preventing the induction of side effects of the centrally mediated central nervous system (CNS) . The inventors are the first to demonstrate a substantial role for peripheral NNRs in neuropathic pain frequently associated with allodynia. Any suitable NNR fighter unable to easily cross the blood-barrier barrier can be used in the method of the present invention. The inventors have made the surprising discovery that agonistic binding to peripheral NNRs provides relief from absent combatant binding of allodynia to centrally located NNRs. This new discovery provides a new method to treat a patient in need of allodynia relief by administering to the patient an NNR combatant who is substantially unable to cross the blood-brain barrier. This new method to treat allodynia dramatically reduces the potential for CNS centrally mediated side effects. In this way, the new method of treatment is of particular value to a patient who requires such treatment for allodynia without incurring centrally mediated side effects. All references contained herein are incorporated by reference in their entirety.

METHODS Animals Sprague Dawley Male rats (80-100 g for nerve ligation alone or 200-225 g for nerve ligation plus cannulation Lev. Or DRG catheterization) were purchased from Charles River (Portage, MI). Before the surgery, the animals were housed per group and kept in a regulating environment by temperature (lights lit between 7:00 a.m. and 8:00 p.m.). Following the nerve ligation surgery alone, the animals were housed per group. Two weeks after the surgery, experimentation began. The animals were between 250-350 g during the experiments. Next to the nerve ligation surgery plus either cannulation i.c.v. or DRG catheterization, the animals were housed individually. Experimentation began one week after surgery when the animals were between 250-350 g. The rats had access to food and water ad libitum.

Surgical Procedures Under halothane anesthesia, the spinal nerves L5 and L6 were tightly ligated in the manner previously described by Chung et al. (Kim S.H., Chung J.M., An experimental model for neuropathy produced by segmental spinal nerve ligation in the mouse, Pain 1992; 50: 355-363). In summary, an incision was made on the dorsal part of the hip, and the muscle was dissected blunt to reveal the spinal processes. The transverse process L6 was removed, and the left spinal nerves L5 and L6 were tightly ligated with 5.0 braided silk suture. The wound was cleaned, the membrane was sewed with 4.0 dissolvable Vicryl suture and the skin was closed with wound clips. The implantation of the DRG catheters was performed immediately following the nerve ligation, with animals maintained under halothane for the entire procedure (35-40 minutes). The catheters were constructed of PE 20 with small pieces of tygon tube cemented along the length of the catheter to allow for suture tie-downs during surgery. The left DRG L5 was exposed by removing the posterior articular process of the L5 vertebra. Gel foam was inserted into the cavity to prevent the catheter from damaging the ganglion. The catheter was sutured to the muscle and aponeurosis, then inserted subcutaneously to externalize between the shoulder blades. The saline was infused into the catheter, and sealed by heat. The implantation of Lev cannulas. were conducted under anesthesia of sodium pentobarbital (50 mg / kg; i.p. Nembutal) with nerve ligation that originated just after implantation; the anals were kept under anesthesia for the entire procedure (40-45 minutes). The animals were implanted with a probe guide cannula 22 (Plastics One, Roanoke VA), cut to 6 mm. The stereotaxic coordinates of bregma were 1.0 mm posterior, 1.6 mm lateral, and 4.5 ventral (Paxinos G. and Watson C, The mouse brain stereotactic coordinates, New York: Academic Press, 1 997). Three skull screws were attached for stability, and the implant was covered with cranioplastic acrylic.

Injection and Infusion Procedures For infusion i .c.v. (intracerebroventricular), the chlorisondamine (10 μg) was dissolved in 5 μ? of saline regulated by phosphate and infused by syringe pump at a speed of 5 μ? / min. The infusion was performed 24 hours before the test in order to reduce the strain to the animals immediately before the behavior test. For all other routes of administration including intraperitoneal (i.p.) > intradermal (i.d.) or on the DRG, the chlorisondamine was dissolved in saline. The final volumes for i.p. injections were 1 ml / kg, for i.d. were 50 μ? and for DRG injections were 10 μ ?. All injections were made by syringe attached to the hand.

Behavioral Assessment For the assessment of acute pain, a claw thermal stimulator was used to evaluate the nociceptive responses to an acute thermal stimulus (Anesthesiology Research Laboratory, Department of Anesthesiology, University of California at San Diego, La Jolla, CA). The device and behavioral procedure have been previously described (Hargreaves et al., A new and sensitive method for measuring thermal nociception in cutaneuos hyperalgesia, Pain 1988, 32: 77-88).; Dirig et al. Characterization of variables defining hindrance paw withdrawal latency evoked by radiant thermal stímuli, J. Neurosci. Meth. 1997; 76: 183-191). In summary, the rats were placed in Plexiglas boxes on a glass surface maintained at 30 ° C and allowed to acclimate for 30 minutes. The measured nociceptive response was the lagging to extract the rear claw when heated by light from a focused projector lamp (current setting at 4.8 A). The final record was the average of the scores for the left and right claw. To avoid tissue damage, the maximum response latency before the automatic lamp shutdown was 20.5 sec. The formalin test was used as a persistent pain assessment. The method, described above (Bannon et al., 1998) is in summary as follows. Following acclimation to the test environment, 50 μ? 5% formalin solution was injected subcutaneously into the dorsal surface of one of the posterior claws. Only phase two, defined as the 30-minute period of 30 to 50 minutes post-formalin injection, was recorded for the nocifensive behaviors that include contraction, adherence, and biting. For the assessment of neuropathic pain, mechanical allodynia in the affected claw of animals that had overcome the spinal nerve ligation was evaluated using the von Frey filaments. As described previously (Chaplan et al., Quantitative assessment of tactile allodynia in the rat paw J Neurosci Meth, 1994; 53: 55-62), two weeks after surgery, the rats were acclimated to the test box that It was constructed of plexiglass with a wire mesh floor to allow access to the plantar surface of the rear claws. Using the Ascending-Descending Dixons method, a baseline level of allodynia was taken with allodynia defined as start of extraction # 4 g. The test compounds were thus administered and consequently the beginnings of extraction were determined.

Experimental Provenance For the acute thermal pain experiments, a start of baseline extraction was determined, and then the chlorisondamine or its vehicle was administered: Next at 30 minutes, A-85380 or its vehicle was determined and the measurements were taken at 15, 30 and 45 minutes post fighter injection. For formalin experiments, chlorisondamine or its vehicle was either fused 24 hours or 40-50 minutes before further handling. A-85380 was injected like this and, 5 minutes later, the formalin was injected into the claw. The measurements were taken between 30 and 50 minutes after the formalin injection. For spinal nerve ligation experiments, the timing of antagonist administration was dependent on the route of administration. With i.c.v. administration, the chlorisondamine or its vehicle was given 24 hours before further handling. On the day of the experiment, a record of baseline allodynia was determined and then A-85380 or its vehicle control was given and measurements were taken in 15, 30 and 60 minutes post-combatant injection. The dose response curves for administration i.p., i.d. and DRG were also determined using this timeline. In contrast, with the i.p. of an antagonist, the The baseline allodynia was determined first, the antagonist or its vehicle was injected and then, 30 minutes later, A-85380 or its vehicle was injected and the measurements were taken at 15, 30 and 60 minutes post-combatant injection.

Compounds The following compounds were dissolved in their vehicles suitable for injections i.c.v., i.p., i.d. or DRG: A-85380 ie, 3- (2 (S) -azetidinylmethoxy) pyridine dihydrochloride (available from RBI, Natick, MA, Sullivan et al., 1996); chlorisondamine diiodide (available from Tocris, Ballwin, MO; Clarke et al., 1994); Mecamylamine hydrochloride (Sigma, St. Louis, MO); and hexamethonium (Sigma).

Statistical Analysis For the experiments with repeated measurements, ie, mechanical allodynia and acute thermal pain, the data were first analyzed using a two-way variability repeated measures analysis (ANOVA) with two independent factors. For the experiments without repeated measurements, ie, formalin, the data were first analyzed using a two-way ANOVA with two independent factors. If there was a significant interaction of both factors or both factors such as time, the subsequent post hoc hoc ANOVAs were performed using each treatment combination as an independent group. The difference in claw extraction latents between the affected and unaffected claw of the neuropathic animals tested in acute thermal pain was assessed using a duplicate t-test. All post hoc importance was determined using Fishers LSD. The use of preclinical models to study pain has allowed a deeper investigation of the different pain states including the determination of multiple sites and mechanisms of action for different types of pain. For example, studies indicate that acute pain and persistent pain are influenced by tonic-descendant inhibitory components that limit the transmission of pain at the level of the spinal cord (Proudfit HK, and Hammond DL, Alterations in nociceptive threshold and morphine-induced analgesia produced by intrathecally administered amine antagonists Brain Res 1981, 218: 393-399; Ornote et al., 1998; Kaneko M, Hammond ID, Role of spinal? -aminobutyric acidA receptors in formalin-induced nociception in the rat. Exp Ther 1997, 282: 928-938). In contrast, in neuropathic pain, while similar descending inhibitory influences may limit the expression of mechanical allodynia (Xu et al., Endogenous noradrenergic tone controls symtoms of allodynia in the spinal nerve ligation model of neuropathic pain, Eur J Pharmacol 1999, 366: 41-45), researchers have identified a downward tonic facilitation of tactile allodynia as well as evidence for additional upward facilitation (Ossipov et al., Mediation of spinal nerve injury induced tactile allodynia by descending facilitatory pathways in the dorsolateral funiculus in rats, Neurosci Lett 2000, 290: 129-132; Sung et al., Supraspinal in the production of mechanical allodynia by spinal injury in rats, Neurosci Lett 1998, 246: 117-119; Sheen K and Chung JM. Signs of neuropathic pain depend on signal from injured nerve fibers in a rat model, Brain Res 1993, 610: 62-68). In the demonstration of additional differences in pain states, the downward facilitation required for the expression of mechanical allodynia does not seem to play a role in thermal hyperalgesia caused by spinal nerve ligation (Bian et al., Tactile allodynia, but not Thermal hyperalgesia, of the hind limbs is blocked by transection spinal in rats with nerve injury, Neurosci Lett 1998, 241: 79-82; Kauppila et al., Influence of spinalization on spinal withdrawal reflex responses varíes depending on the submodality of the stimulus test and the experimental pathophysiological condition in the mouse, Brain Res 1998, 797: 234-242). Neuronal mechanisms that emphasize antinociception induced by NNR combatants in acute pain models have also been shown to differ based on the type of acute pain produced or on the specific NNR combatant used (Curzon et al., Differences between the antinociceptive effects of the cholinergic channel activators A-85380 and (+) - epibatidine in rats, J Pharmacol Exp Ther 1998; 287: 847-853). Since NNR fighters have also been shown to be effective in neuropathic and persistent pain models (Bannon et al., Broad-spectrum, non-opioid analgesic activity by selective modulation of neuronal nicotinic acetylcholine receptors, Science 1998, 279: 77-81; Kesingland et al., Analgesia profile of the nicotinic acetylcholine receptor agonist, (+) - epibatidine and ABT-594 in models of persistent inflammatory and neuropathic pain, Pain 2000, 86: 113-118) an objective of the present study was to delineate more the different possible neuronal trajectories that underline the antinociception induced by combatant NNR, analgesia and anti-allodynia. The combatant NNR A-85380 was used completely in order to facilitate comparison between the various pain models.

EXAMPLE 1 The effect of chlorisondamine systemically administered in an acute thermal pain model was examined. Chlorisondamine, an almost irreversible quaternary NNR antagonist that does not readily cross the blood-brain barrier was used to identify the peripheral and central actions of A-85380. The effect of systemically administered chlorisondamine (0.4 [mu] g / kg, i.p.) in A-85380 (0.75 [mu] t / kg, i.p.) was assessed. As shown in FIG. 1, chlorisondamine had no effect on the antinociceptive action of A-85380 in acute thermal pain (combatant-antagonist interaction, p = 0.42, antagonist effect, p = 0.35).

EXAMPLE 2 The effect of chlorisondamine centrally and systemically administered in a persistent pain model was examined. Using phase two of the persistent pain formalin model, chlorisondamine (10 μg) was administered Lev. 24 hours before the systemic administration of A-85380. The chlorisondamine administered i.c.v. completely blocked the analgesic effects of systemically administered A-85380 (0.75 μg / kg, i.p. antagonist-combatant interaction, p = 0.048, antagonist / combatant treatment combination effect p = 0.0002, FIG.2A). In contrast, when chlorisondamine was administered systemically 40-50 minutes before the formalin injection, there was no significant attenuation in analgesia induced by A-85380 in phase two of the formalin test (combatant-antagonist interaction, p = 0.12). , antagonist effect p = 0.19; FIG 2B).

EXAMPLE 3 The effect of Fighter NNR A-85380 in a model of neuropathic pain was examined. A-85380 systemically administered induced a dose-dependent anti-allodynia in rats with neuropathy secondary to the narrow ligation of spinal nerves L5 and L6 (dose-time interaction, p <0.0001, effects of A-85380 to 15, 30 and 60 minutes, p <0.0001), see FIG. 3. A-85380 at 0.5-1.0 μ ???? / kg, i.p. induced behaviors such as prostration and ataxia immediately following the injection. However, these effects were abated by the 15 minute time point and did not interfere with the behavioral test.

EXAMPLE 4 The effect of chlornisamine NNR antagonists systemically administered on anti-allodynia induced by A-85380, was examined. To determine whether the anti-allodynic action of A: 85380 was mediated by NNRs, the ability of the NNR channel blocker mecamylamine to block the anti-allodynia induced by A-85380 was assessed. Mecamylamine, 1 μ? T ??? / kg, i.p. given 30 minutes before A-85380, it completely blocked the anti-allodynia of A-85380 (antagonist-combatant interaction, p <0.0001, antagonist / combatant treatment combination effect p <0.0001, FIG 4). Prostration and ataxia that immediately followed the combatant's injection were also blocked.

EXAMPLE 5 The site of anti-allodynia action induced by A-85380 was examined. To determine the site of action of the anti-allodynia induced by A-85380, the quaternary NNR antagonist chlorisondamine is given either i.c.v. or i.p. before the administration of A-85380. When i.c.v. 24 hours before the behavioral test, chlorisondamine (10 completely blocked the anti-allodynia induced by systemically administered A-85380 (0.75 μG ???? / kg, ip, interaction of combatant and antagonist, p = 0.01; of antagonist / combatant treatment combination p = 0.0004; FIG 5A) Surprisingly, in contrast to the lack of effects administered to block the anti-allodynia induced by A-85380 was in fact due to the penetration of chlorisondamine into the CNS through a permanent weakness in the BBB incidentally originated during the surgery.To point out this interest, the neuropathic rats were evaluated in the acute thermal model of claw extraction.As demonstrated above, the chloridodamine peripherally administered is completely ineffective in antagonizing the antinociception induced by A-85380 indicating a completely central site of action of the combatant in this model of pain. was crossing the BBB in neuropathic animals, the chlorisondamine peripherally administered in these animals should partially or completely block the antinociceptive effects of A-85380. This was not the case as shown in FIG. 7. Systemically administered chlorisondamine (0.4 μG ??? / kg / ip) did not alter the antinociception induced by systemically administered A-85380 (0.75 μG ??? / kg, ip) when neuropathic rats were used in the extraction model. claw of acute thermal pain (interaction of antagonist and combatant, p = 0.46, effect of antagonist p = 0.41). Interestingly, in the baseline measurement alone, there was a small but significantly lower start of claw removal for the affected claw (7.2 + 0.3 s) vs. the affected claw (8.5 + 0.3 s; p = 0.003; data not revealed).

EXAMPLE 8 The site of action for the peripheral effects of A-85380 in a neuropathic pain model was examined. The ability of A-85380 to induce an anti-allodynic action when applied directly to the primary receptive field was assessed by injecting A 853.80 into the plantar surface of either the affected claw (ipsalateral to the ligation) or unaffected (contralateral to the ligation) and then the degree of allodynia in the affected claw is measured. A-85380, in doses of 10, 20 and 50 μg / rat, i.d. induced changes in the start of claw extraction that depended on time, claw and dose (interaction, p = 0.01). Therefore, analyzes of each claw were made individually. For ipsalateral claw injection, only 50 μg of dose of A-85380 had an anti-allodynic effect, predominantly at the 15 minute time point even though the 60 minute time point was also significantly different from saline ( time and dose interaction, p <0.0001, dose effect at 15 min, p <0.0001, at 30 min, p = 0.47, at 60 min, p = 0.005, FIG 8A). For the contralateral claw injection, all three doses of A-85380 induced a significant anti-allodynia, again the predominant effect being in the time of 15 minutes with a very small but significant effect of 10 μg in only 60 minutes (interaction of time and dose, p <0.0001, dose effect at 15 min p <0.0001, at 30 min, p = 0.36, at 60 min, p = 0.03, FIG 8B).

EXAMPLE 9 The ability of A-85380 to induce an anti-allodynic action when applied directly to the DRG was examined. A-85380 was fused on the L5 DRG on the side of the ligation. Both 10 and 20 μ9 of A-85380 infused on it. DRG induced a significant anti-allodynia that lasted 15 and 30 minutes, respectively (time and dose interaction, p <0.0001, dose effect at 15 min, p <0.0001, at 30 min, p = 0.007, at 60 min , p = 0.33, FIG 9A). The pretreatment with chlorisondamine, 0.4 μ ???? /? ^, I.p. 30 minutes before, it completely blocked the anti-allodynia induced by 20 μg of infused A-85380 on the DRG (p <0.0001, FIG 9B).

EXAMPLE 10 A comparison was made between the administration i.p., i.d., and DRG of A-85380. In order to allow a more direct comparison, the doses i.d. and DRG were first converted into their equivalent systemic dose, μ ???? / kg, by determining the dose received by each rat according to their body weight and then calculating the given average dose. The fifteen minute time point of the i.p., ipsalateral, i.d., and DRG routes of administration are shown in FIG. 10. While the power to increase the start of claw extraction is similar between the i.p. and i.d., A-85380 is decided more potent when administered directly on the DRG.

DISCUSSION The action site (s) highlighting the reduction in pain response capacity induced by combatant NNR A-85380 differ according to the type of pain model used. Chlorisondamine, an almost irreversible quaternary NNR antagonist that does not readily cross the blood-brain barrier was used to separate the peripheral and central actions of A-85380 (Clarke et al., The pharmacology of the nicotinic antagonist, chlorisondamine, investigated in rat brain and autonomic ganglion, Br J Pharmacol 1994, 111: 397-405). For acute thermal pain, studies with systemically administered chlorisondamine vs. Centrally administered indicated that the antinociceptive site is mediated centrally (see Curzon et al., Differences between the antinociceptive effects of the cholinergic channel activators A-85380 and (+) - epibatidine in rats, J Pharmacol Exp Ther 1998, 287: 847- 853, see also FIG 1). Similar to acute thermal pain, the analgesic action of A-85380 in a persistent pain model was also shown to be mediated predominantly in the CNS, as indicated by the ability of centrally administered chlorisondamin to block analgesia induced by A-85380 while the peripheral administration of the antagonist had no significant effect (FIG 2). In contrast to the acute thermal and persistent pain model, the present study demonstrated a substantial role for peripheral NNRs in the anti-allodynia induced by the combatant NNR A-85380. A-85380 induced a dose-dependent anti-allodynia in the spinal nerve ligation model of neuropathic pain, an effect mediated by NNRs as demonstrated by total blocking of anti-allodynia with the NNR antagonist, mecamylamine (FIGS.3 and 4). In addition, the ability of chlorisqndamine, both centrally and peripherally administered to antagonize the anti-allodynia induced by A-85380, demonstrated that there is both a peripheral and central site of anti-allodynic action (FIG 5). Because the peripheral site of the anti-allodynic action for a NNR combatant is a new discovery, it was considered necessary to verify this result. Thus, the peripheral antagonism of A-85380 was also shown to utilize the NNR antagonist, hexamethonium, (FIG 6). In addition, a defect in the blood-brain barrier caused by the ligation surgery was regulated by the discovery that the peripherally administered chlorisondamine does not alter the antinociception in the claw extraction test of acute thermal pain in neuropathic rats (FIG. 7). With verification of the peripheral action of A-85380 established, the exact location of the peripheral anti-allodynia was investigated. Injection of A-85380 directly into the respective primary field of sensory neurons on the plantar surface of the ipsalateral claw induced a small but important anti-allodynia. However, equivalent or greater anti-allodynia was achieved when A-85380 was injected towards the contralateral claw, indicating that both effects were more likely due to a systemic effect of A-85380, not a local effect in the primary receptive field ( FIG 8). In contrast to the lack of a selective effect of A-85380 in the primary receptive field. A-85380 induced a dose-dependent anti-allodynic effect when infused directly onto the L5 DRG (FIG 9). This effect of A-85380 was blocked by the pre-treatment with the systemically administered chlorisondamine indicating that it is mediated by NNR. A comparison of the effects of A-85380 when administered systemically, in the primary receptive field or on the DRG show that A-85380 is more potent when infused onto the DRG, suggesting in this way that this may be a peripheral site of an important action for anti-allodynia induced by NNR combatant in a neuropathic pain rat model (FIG 10). While this seems to be a good interest of similarity between antinociception and analgesia, in terms of the mechanisms of action delineated in preclinical studies that include NNR binders as well as other receptor binders and in terms of the therapeutic pharmacology found in the Clinically, studies indicate that anti-allodynia in neuropathic pain is very different (Curzon, et al., J. Pharmacol Exp Ther 1998; 287: 847-853; Rueter et al., Brain Res. 2000; 872: 93-101; Rogers and Iwamoto, J. Pharmacol Exp Ther 1993; 267: 341-349; Ornotete et al., Brain Res. 1998; 814: 194-198; Sugimoto er al., Neuropharmacology 1986; 25: 481-485; Kaneko and Hammond, J. Pharmacol Exp Ther 1997; Millan, Prog. Neurobiol 1999; 57: 1-164). Specifically, neuropathic pain clearly alters the functioning of both peripheral and central processes such as the induction of ectopic discharge in sensory neurons and the induction of central sensitization, respectively (Go \ d, Pain 2000; 84: 117-120; Attal and Bouhassira, Acta Neurol Scand 1999, Suppl 173: 12-24, Sheen and Chung, Brain Res. 1993, 610: 62-68, Sung et al., Neurosci Lett 1998; 246: 117-119; Bian ei al ., Neurosci Lett 1998; 241: 79-82; Ossipov et al., Neurosci Lett 2000; 290: 129-132; Read al., Neurosci Lett 2000; 290: 129-132). Therefore, theoretically, the inhibition of pain transmission and the subsequent relief of mechanical allodynia could occur either through the actions of a therapeutic agent in the periphery or in the CNS. The differential effects of the site injections found in the present study suggest that there is a peripheral site of anti-allodynic action mediated by NNR in the DRG but not in the primary receptive field (FIGS 8, 9, 10). The present study is the first to report the ability of an NNR combatant to reduce neuropathic pain at the DRG level and, in turn, the level of pain relief following the infusion of NNR combatant into the DRG is much larger that of any compound of any other kind previously reported (FIG 9, Liu et al., 2000, Pain 2000; 85: 503-521; Lyu et al., Brain Res. 2000; 871: 98-103). Most of the NNR subunits have been reported to exist in the DRG including, but not limited to, 7, ct3, and a4 (Ninkovic and Hunt, Brain Res. 1983; 272: 57-60; Boyd ef a /., 1991 , J. Neurobiol 1991; 22: 1-14). In this way, the NNRs are in a position to alter the transmission of pain at the DRG level, and the present study suggests that they may play an important role in reducing the transmission of neuropathic pain at this site.

EXAMPLE 11 A-420503 (IR, 5S) 5- (3,6-diazabicyclo [3.2.0] hept-6-yl) -N-hydroxy-nicotinamide) infusion of significant anti-allodynia at 500 mg which is equivalent to 4.77 μ ???? / kg, ip The effects on the DRG were blocked by pre-treatment with an NMR antagonist. When i.p. was injected, this induces significant anti-allodynia at 62 μg / kg, with no noticeable side effects.

EXAMPLE 12 When A-85380 is given systemically (i.p.), it easily crosses the blood-brain barrier and stimulates both peripheral and central NNRs. It also induces an important dose-dependent reduction in the symptoms of neuropathic pain (anti-allodynia) in the rat spinal nerve ligation model of neuropathic pain. This can be seen in FIG. 1 (black boxes). Along with the reduction in allodynia, there are a variety of side effects such as prostration, ataxia, shingles and dyspnea that limit the range of therapeutic doses. Chlorisondamine is a quaternary NNR antagonist that does not easily cross the blood-brain barrier. When i.c.v. in small quantities, it only blocks the centrally located NNRs. When central NNRs are blocked with chlorisondamine (10 μg), subsequent systemic injections of A-85380 should activate only the peripheral NNRs. As also shown in FIG. 11, when this is done in neuropathic rats, A-85380 is still able to induce a significant reduction in allodynia. A higher dose is required to see this effect (3 μG ??? / kg when given without an antagonist) as can be seen in the previous graph (black circles). However, in addition to the increase in dose, there are almost no side effects suggesting that side effects such as prostration, ataxia, and shoring are mediated by centrally located NNRs. When a small amount of chlorisondamine is given i.p., it only blocks the peripherally located NNRs. When peripheral NNRs are blocked with chlorisondamine (0.4 μ? P ?? / kg, i.p.), subsequent systemic injections of A-85380 should only activate the central NNRs. Also as shown in FIG. 11, when this is done in neuropathic rats, A-85380 is still able to induce a significant reduction in allodynia. A higher dose is required to observe this effect (1.5 μG ??? / kg vs. 0.75 μ? T ??? / kg when it occurs without an antagonist) as can be observed in the previous graph (blue triangles). However, with the increase in the dose there is a significant increase in side effects including shingles, dizziness, prostration and ataxia, suggesting further that the side effects that limit the dose of A-85380 are mediated centrally. Together these data also support a role for peripherally located NNRs in the anti-allodynia induced by the NNR combatant in a rat neuropathic pain model. In addition, they demonstrate for the first time that this anti-allodynia can be achieved following a systemic injection of an NNR combatant and in the absence of stimulation of the centrally located NNRs. In addition, they demonstrate that this anti-allodynia can be achieved with lower side effects than when centrally located NNRs are stimulated. While the invention is described above in connection with the illustrative or preferred embodiments, these embodiments are not intended to be exhaustive or limiting of the invention. In turn, the invention is intended to cover all alternatives, modifications and equivalents included within its spirit and scope of the invention, as defined by the appended claims.

Claims (6)

1. CLAIMS 1. A method for treating a patient suffering from neuropathic pain, comprising administering to a patient in need of such treatment an effective amount of a combatant drug capable of binding peripheral neuronal nicotinic receptors but not easily crossing the blood barrier -brain.
2. 2. The method for treating a patient suffering from neuropathic pain according to claim 1, characterized in that the drug provides the patient with relief of allodynia.
3. 3. A method for treating a patient suffering from allodynia, comprising administering to a patient in need of such treatment an effective amount of a combatant drug capable of binding to at least one peripheral neuronal nicotinic receptor but not easily crossing the blood barrier -brain.
4. 4. A method for treating a patient suffering from neuropathic pain, comprising administering to a patient in need of such treatment an effective amount of a quaternary nicotinic compound.
5. 5. A method for treating a patient suffering from neuropathic pain by performing peripheral neuronal nicotinic receptors at the level of the dorsal root ganglion, comprising administering to a patient in need of such treatment an effective amount of a nicotinic combatant compound that does not cross easily the blood-brain barrier.
6. 6. A method for reducing side effects associated with combatant activation of central neuronal nicotinic receptors, comprising administering to a patient in need of such treatment an effective amount of a nicotinic combatant compound that does not readily cross-the blood barrier. brain.