ES2353702T3 - DERIVATIVES OF 1- [2H-1-BENZOPIRAN-2-ONA-8-IL] -PIPERAZINE FOR THE TREATMENT OF PAIN. - Google Patents

Abstract

Use of a compound of formula (1): (R 2) n NNR 3 OO (R 4) p (R 1) m R1 is (C1-4) alkyl, (C1-4) alkoxy, hydroxyl, (C1-) alkoxy 4) (C1-4) alkyl, pyrrolidinyl, piperidinyl, morpholinyl, halogen, cyano, trifluoromethyl, amino or amino mono- or disubstituted amino acids where the substituents are (C1-4) alkyl or (C1-4) alkyl carbonyl, m has the value 0, 1 or 2, R2 is (C1-4) alkyl, (C1-4) alkoxy, halogen or trifluoromethyl, n is 0 or 1, with the proviso that (m + n) is at least 1, R3 is hydrogen, (C1-3) alkyl or (C2-3) alkenyl, R4 is (C1-4) alkyl and p has the value 0, 1 or 2, and all stereoisomers and pharmacologically acceptable salts thereof, for the preparation of a Pharmaceutical composition for pain treatment.

Description

The invention relates to a new use of known derivatives of 1- [2H-1benzopyran-2-on-8-yl] -piperazine, broad-spectrum compounds that bind the 5-HT receptor, which have among other functional activities of the receptor of serotonin, potent agonist activity of the 5-HT1A receptor as well as 5-HT1D receptor antagonist activity. The invention also relates to the use of a compound disclosed therein for the manufacture of a medicament, as specified in the claims. A beneficial effect is revealed in this or is apparent to a person understood in the subject from the descriptive memory and the general knowledge included in the technique. In embodiments of the invention specific compounds disclosed therein are used for the manufacture of pain medications.

Acute pain is a normal sensation triggered in the nervous system to alert the individual about possible damage. Chronic pain results from persistent signs of pain in the nervous system that continue after the initial injury or damage has disappeared. Chronic pain can occur in the absence of any injury or evidence of past body damage, the so-called psychogenic pain.

There are many different types of analgesics. None of them is perfect, nor is it possible with the currently available drugs to adequately treat any type of pain, quickly and without any side effects.

The invention was intended to provide a medicament for the treatment of pain that contains at least one compound with a molecular mechanism of action different from that of all currently commercialized analgesics and therefore has therapeutic value under pain conditions not satisfactorily treatable with known analgesics.

Surprisingly, it was found that 3-amino-8- (1-piperazinyl) -2H-1-benzopyran-2-one monohydrate monohydrate (hereinafter referred to as "compound 1"), a compound that binds the 5-HT receptor of broad spectrum, which has, among other functional activities of the serotonin receptor, potent 5-HT1A agonist activity, 5-HT1D antagonist activity, as well as 5-HT7 agonist activity (see receptor ligation profile, below), it has potent activity in experimental animal models of pain. The compound has no sedative effects when administered in dosages of up to 100 mg / kg p.o., and it was also shown to be highly active as a growth factor inducer. This last activity indicates neuroprotective effects and improvement of brain plasticity required for neuroregeneration

and also indicates potential therapeutic effects in neuropathic pain (see, P. Anand., "Neurotrophic factors and their receptors in human sensory neuropathies", Prog. Brain Res., 146, 477-492, 2004 and R. Wang et al., "Glial cell line-derived neurotrophic factor normalizes neurochemical changes in injured dorsal root ganglion neurons and 5 prevents the expression of experimental neuropathic pain", Neuroscience, 121, 815824, 2003) and diabetes-induced pain (JA Christianson et al., "Beneficial effects of neurotrophin treatment on diabetes-induced hypoalgesia in mice ”, J. Pain, 4, 493-504, 2003). When administered orally, the compounds of the invention show good bioavailability, resulting in an action of high potency and duration.

10 prolonged. The pharmacological activities of the compounds of the invention and their salts represent a new class of analgesic compounds for the treatment of chronic pain disorders or to treat other conditions in which there is a hypersensitization to painful signals, hyperalgesia, allodynia, increased perception.

15 pain and increased pain memory. The invention relates to compounds of the general formula (1)

image 1

where:

R1 is (C1-4) alkyl, (C1-4) alkoxy, hydroxy, (C1-4) alkoxy (C1-4) alkyl, pyrrolidinyl, piperidinyl, morpholinyl, halogen, cyano, trifluoromethyl, amino or amino mono-o disubstituted where the substituents are (C1-4) alkyl or (C14) alkylcarbonyl, m has the value 0, 1 or 2,

R2 is C1-4 alkyl, C1-4 alkoxy, halogen or trifluoromethyl, n is 0 or 1, with the proviso that (m + n) is at least 1, R3 is hydrogen, C1- alkyl 3) or (C2-3) alkenyl, R4 is (C1-4) alkyl and

p has the value 0, 1 or 2, as well as pharmacologically acceptable salts for use in the treatment of pain thereof.

Derivatives of 1- [2H-1-benzopyran-2-on-8-yl] -piperazine, compounds that bind the broad-spectrum 5-HT receptor, which have among other functional activities of the serotonin receptor, agonist activity 5- Powerful HT1A as well as 5-HT1D antagonist activity, originally developed as antidepressants (European patent EP 0 650 964). The presence of 5-HT1D antagonism is considered to have therapeutic value. The 5-HT1D receptors are presynaptically located on the nerve terminal and have a negative modulatory influence on the release of 5-HT. Therefore, blocking these receptors increases the release of 5-HT from their terminals. The additional presence of presynaptic 5-HT1D antagonism will result in a similar effect as observed after the administration of 5-HT reabsorption inhibitors. When the 5-HT1D antagonism is combined with 5-HT1A agonism, the latter activity is intensified.

That the preceding reasoning is probably valid for the analgesic activity of the compounds of the invention was shown by an interaction study with sumatriptan, the prototype 5-HT1D agonist developed as an anti-migraine drug. Although migraine often manifests itself as an acute headache, "pain" and "migraine" are certainly not synonymous. Until the development of "triptans" as selective anti-migraine drugs, the disorders were completely refractory to classical analgesics. The reverse is also applicable: "triptans" are not analgesics. Sumatriptan was found to be completely inactive in the "hot plate" test (see below). Surprisingly, however, it was found that sumatriptan almost completely blocked the analgesic effect of compound 1 (see below), clearly demonstrating that the 5-HT1D antagonism of this compound plays an important role in its analgesic activity.

International patents WO 99/39128 and WO 99/52907 disclose substituted azabicyclic compounds as agonists and antagonists of 5HT1 receptors specifically one or both 5-HT1A and 5-HT1D receptors. It was claimed that these compounds were useful in the treatment or prevention of migraine, depression and other disorders for which 5-HT1 is indicated agonist or antagonist.

To the invention belong all the compounds having the formula (1), racemates, mixtures of diastereomers and individual stereoisomers. Therefore, the compounds in which the substituents on potentially asymmetric carbon atoms are in the R configuration or in the S configuration belong to the invention.

Pharmaceutically acceptable salts can be obtained by standard procedures well known in the art, for example by mixing a compound of the present invention with a suitable acid, for example an inorganic acid such as hydrochloric acid or with an organic acid. The active compounds and their salts can be processed to form compositions by methods

10 standard, for example pills, tablets, coated tablets, capsules, powders, injectable liquids and the like, using auxiliary substances such as liquid and solid excipient materials. Especially preferred are the compounds having the formula (1) where (R2) n and (R4) p are hydrogen, R3 has the meanings indicated above and (R1) m is a

Substituent at position 3 selected from the group consisting of pyrrolidinyl, piperidinyl, morpholinyl, amino and mono- or disubstituted amino where the substituents are (C1-4) alkyl or (C1-4) alkylcarbonyl.

The invention particularly relates to the compound 3-amino-8- (1-piperazinyl) 2H-1-benzopyran-2-one and the salts thereof, that is to say the compound of formula (1) where (R1) m is 3 -NH2, (R2) n, R3 and (R4) p are hydrogen, which therefore has the

formula (2):

NH2

image2

(2)

Especially preferred is 3-amino-825 (1-piperazinyl) -2H-1-benzopyran-2-one monohydrate monohydrate, which is referred to in the following as "Compound 1".

The compounds of the invention are active with doses in the range of 0.1-100 mg / kg after oral administration and their unique pharmacological profile makes them particularly useful in the treatment of pain.

30 According to what is used in it, the term pain refers to all types of pain. Preferably, the term should refer to all types of chronic pain, including nociceptive, neuropathic, psychogenic and mixed category pain (nociceptive and neuropathic components). This particularly includes, but is not limited to, diabetic neuropathy, neurogenic pain, central pain, somatic pain, visceral and cancer pain, inflammatory pain, post-operative pain, chronic waist pain, sciatica, cervical and lumbar pain, aches of tension headache, migraine neuralgia, chronic daily headaches, herpes neuralgia and post-herpes neuralgia, facial and oral neuralgia and myofacial pain syndromes, phantom limb pain, stump pain and paraplegic pain, dental pain, resistant pain to opioids, post-surgical pain including cardiac surgery and mastectomy, labor and delivery pain, postpartum pain, post-infarction pain, angina pain, genito-urinary tract pain including pelvic pain and cystitis and vulvar vestibulitis and orchyalgia, irritable bowel syndrome, pain of premenstrual syndrome, pain resulting from burns or chemical damage or sunburn and pain of damage Bone.

Subtypes of nociceptive pain are somatic pain and visceral pain.

Somatic pain includes inflammatory pain, post-operative pain, chronic waist pain, cervical and lumbar pain, migraine neuralgia, dental pain, labor and delivery pain, postpartum pain, pain resulting from burns or chemical damage or sunburn and bone damage pain.

Visceral pain includes cancer pain, post-surgical pain including cardiac surgery, angina pain, genito-urinary tract pain including pelvic pain and cystitis and vulvar vestibulitis and orchyalgia and premenstrual pain syndrome.

Subtypes of neuropathic pain are diabetic neuropathy, cancer pain, neurogenic pain, central pain, sciatica, herpes neuralgia, post-herpes neuralgia, facial and oral neuralgia, phantom limb pain, stump pain and paraplegic pain, opioid-resistant pain, post-surgical pain including mastectomy and post-infarction pain.

Subtypes of psychogenic pain are chronic daily headaches and tension headaches.

Subtypes of mixed category pain are cancer pain, myofacial syndromes and tension headaches (for example McCaffery M, Pasero C. Pain: Clinical Manual p. 19 St. Louis: Mosby 1999; Merskek H and Bogduk (eds) Classification of chronic pain, 2nd edition, IASP Task Force on Taxonomy, pp. 209214, IASP Press, Seattle 1994; The Merck Manual, Section 14, Chapter 167, Pain, 17 Merck & Co Edition 1999).

PHARMACEUTICAL PREPARATIONS

The compounds of the invention can be brought into forms suitable for administration by usual processes, using auxiliary substances such as liquid or solid excipient material. The pharmaceutical compositions of the invention can be administered enterically, orally, parenterally (intramuscularly or intravenously), rectally or locally (topically). They can be administered in the form of solutions, powders, tablets, capsules (including microcapsules), ointments (creams or gels) or suppositories. Suitable excipients for such formulations are liquid or solid fillers and diluents, solvents, emulsifiers, lubricants, flavorings, colorants and / or pharmaceutically usual buffer substances. Frequently used auxiliary substances that may be mentioned are magnesium carbonate, titanium dioxide, lactose, mannitol and other sugars or sugar alcohols, talc, lactoprotein, gelatin, starch, cellulose and their derivatives, animal and vegetable oils such as liver oil fish, sunflower oil, peanuts or sesame, polyethylene glycol and solvents such as, for example, sterile water and mono- or polyhydric alcohols such as glycerol.

Types of pharmaceutical compositions that may be used include, but are not limited to, tablets, chewable tablets, capsules, solutions, parenteral solutions, suppositories, suspensions and other types disclosed therein or apparent to a person skilled in the art herein and General knowledge in the subject. In embodiments of the invention, a pharmaceutical package or equipment is provided comprising one or more containers filled with one or more of the ingredients of a pharmaceutical composition of the invention. Various written materials may be associated with such a container (such containers), such as instructions for use or a note in the form required by a government agency that regulates the manufacture, use or sale of pharmaceutical products, a note that reflects approval by the agency of manufacture, use or sale for human or veterinary administration. PROTOCOLS FOR PHARMACOLOGICAL TESTS CHRONIC CONSTRICTIVE DAMAGE OF THE NERVIO: A NEUROPATHIC PAIN MODEL

The purpose of these studies is to evaluate the potential analgesic properties of a test substance in the Bennett and Xie model of peripheral mononeuropathy (see:

G.J. Bennett and Y-K. Xie, "A peripheral mononeuropathy in rat that produces disorders of pain sensation like those seen in man", Pain, 33, 87-107, 1988). Study design

Male Sprague-Dawley rats (approximately 250 g; Harlan, R.U., or other accredited supplier). The observer is blind to the treatment groups.

Five treatment groups (n = 10 rats / group); control vehicle, reference substance and 3 dosage groups for the test substance. 2 days of baseline trial, surgery to induce peripheral mononeuropathy and monitoring the development of neuropathy using a behavioral test on days 10 and 11 post-operative (PO). Chronic administration of the drug, subcutaneously (either by injection twice a day or by continuous infusion via osmotic mini-pumps), starting on day 12 PO for 14 days and behavioral testing in four instants of post-dose time by mechanical allodynia. Process:

The rats are prepared in batches (with members of each treatment group in each batch) and behavioral and dosing tests are performed for each batch at fixed intervals after surgery. Behavioral tests (see below) are performed with all rats for a period of 2 days prior to surgery, to establish baseline values. A peripheral mononeuropathy is then induced by placing four loosely constrictive ligatures around the right common sciatic nerve under aseptic conditions. The animals are allowed to recover from the surgery for a minimum of 4 days before restarting the behavioral tests (B. Lynn and SE Carpenter, “Primary afferent units from the hairy skin of the rat hind limb”, Brain Research, 238, 29 -43, 1982).

The behavior test is restarted on day 10 post-operative (PO) and repeated on day 11 to monitor the development of allodynia. Drug treatment is carried out from day 12 PO (the instant of time corresponding to the maximum behavioral changes) or as defined in the study protocol and behavioral tests are carried out for a certain time. Animals that show any signs of autonomy from the affected parts are eliminated. Any animal that does not develop a peripheral mononeuropathy (as determined by the results of the behavioral tests used in a particular protocol) is not used in the study. Behavioral tests:

Mechanical allodynia test: the animal is placed in a glass wire cage and Von Frey filaments are applied to the surface of the hind leg. The filaments are applied in ascending order (starting with the weakest force) and the withdrawal threshold is evaluated for both hind legs, the ipsilateral and the contralateral. The withdrawal threshold is defined as the lowest force of two or more consecutive Von Frey filaments to elicit a withdrawal reflex response (ie a short tap of the leg).

Analysis of the results:

Standard statistical methods are used to evaluate effects related to the test substance. The data is analyzed for homogeneity and appropriate parametric or non-parametric methods are applied. LIGADURA OF SPINAL NERVES: A NEUROPATHIC PAIN MODEL

Neuropathic pain test (Chung test) in the rat according to Kim and Chung (Pain 1992, 50: 355-363): slight ligation of spinal nerves in rats is associated with hyperalgesia, allodynia and spontaneous pain and constitutes both a model for peripheral neuropathic pain in humans. Antihyperalgesics reduce these chronic signs of pain hypersensitivity. Rats (180220 g) (sodium pentabarbital 40 mg / kg i.p.) are anesthetized and an incision is made at the L4-S2 level to expose the left L5 and L6 spinal nerves. A ligature is tightly bound around each nerve. Then the wound is sutured. Rats receive an intramuscular injection (i.m.) of 50,000 IU Penicillin and are allowed to recover. At least 2 weeks after surgery, when the chronic state was completely installed, the rats are consecutively subjected to thermal and tactile stimulation of both hind legs, the uninjured and the injured. For thermal stimulation, the device consists of 6 boxes of individual Plexiglas (17 x 11 x 13 cm) placed on a raised glass floor. A rat is placed in the box and left freely to get used for 10 minutes. Then, the lower part of the undamaged and injured hind legs is focused with a mobile source of infrared radiation (setting 20) and the latency of leg removal latency is automatically recorded. The removal of the leg interrupts the reflected radiation and disconnects the meter and the light source. To prevent tissue damage, if no reaction is observed, the test is terminated after 45 seconds. For tactile stimulation, the animal is placed under an inverted Plexiglas box (17 x 11 x 13 cm) on a grid floor. The tip of a Von Frey electronic probe with increasing pressure is then applied to the uninjured and injured hind legs and the force required to induce the removal of the leg is automatically recorded. This procedure is carried out 3 times and the average force per leg is calculated to provide a base classification per animal. Prior to receiving drug treatment, all animals are subjected to tactile stimulation and assigned to treatment groups based on their response to pain.

IN VIVO ELECTROPHYSIOLOGY: A MODEL FOR INFLAMMATORY PAIN

In vivo electrophysiology is a powerful means to directly observe the sensory responses of spinal neurons to supra-threshold stimuli (as opposed to behavioral tests using thresholds) and thus is more related to the clinical pain states.

Male Sprague-Dawley rats (200-250 g) are anesthetized with 2-3% halothane (in 66% N2O and 33% O2) and subsequently maintained with 1.8% halothane. The temperature of the animal's body nucleus is monitored using a rectal thermometer probe coupled to a heating blanket. At the end of the experiment, the animals are eliminated with an anesthetic overdose. A laminectomy is performed to expose the L4-L5 segments of the spine and a tungsten electrode coated with parylene is inserted into the dorsal horn using a Stepper Epson device. The depth of the registration site is observed from the microlector readings. Extracellular records are made of individual neurons of the dorsal horn that receive inputs of C and A fibers from the skin of the hind leg, identified by their ability to respond to both harmful and harmless stimuli (prick and touch). Neural responses are caused by transcutaneous electrical stimulation applied in the center of the receptive field of the neuron in the ipsilateral rear, with a current of 3 times the threshold current required for the activity evoked by fiber C. At 10 minute intervals tests are carried out consisting of a series of 16 stimuli (pulses of 2 ms amplitude at 0.5 Hz) and post-stimulus histograms are constructed. These evoked responses are separated, according to latency, in activity evoked by fiber A� (0-20 ms post-stimulus); activity evoked by A� fiber (20-90 ms); activity evoked by fiber C (90-300 ms) and post-discharge of the neuron (300-800 ms). The neuronal response evoked by the first stimulus of the series is referred to as "input" and consists of the number of action potentials (90-800 ms) evoked by this stimulus. Wind-up, a measure of the increased neuronal response evoked by repetitive stimulation, is quantified as the difference between the total number of action potentials produced by the 16 stimuli (90-800 ms) and the x16 input. , the wind-up measure includes both the increased responses evoked by fiber C and the post-discharge generated as the wind-up develops. Von Frey hair (1-75 g) and heat (30-48 ° C) are also used to quantify responses to natural mechanical and thermal stimuli.

The tests are carried out at intervals of 10 minutes, until the neuronal responses evoked in each trial differ by less than 10%. Then, the results of the last three tests are averaged to give control values for each parameter.

Then cumulative doses of the compound (in a volume of 50 µl) are applied to the exposed spine, in a "cavity" formed by laminectomy containing the volume of 50 µl of drug. Neural responses are followed for 60 minutes after each dose, after which the solution can be taken out to apply the next dose. Similarly, systemic effects can be measured using subcutaneous injection in 0.2 ml.

The data is shown as a percentage of the pre-drug control values, with a time course of 60 minutes for each dose of the drug. To generate a dose response curve, the maximum effects of each dose on each neuron are averaged. Data are presented as mean ± mean square error. For each route, 10 neurons are needed. A comparison of the effects of the compound (by the most effective route determined by the preceding studies) is made in normal animals with those observed 3 hours after inflammation with carrageenan. In this way a total of 30 experiments are performed - one neuron per animal and each study lasts one day. FORMALINE INJECTION TEST IN LA PATA: A MODEL FOR INFLAMMATORY PAIN

The formalin injection test in the mouse or rat leg according to Wheeler-Aceto et al., (Psychopharmacology 1991, 104: 35-44): the animals are given an injection in the plant of the left hind leg of 5% formalin (25 for the mouse, 50 for the rat). This treatment induces a recognizable recoil response in control animals. The number of setbacks is counted for 10 minutes, starting immediately after the formalin injection (early phase) and again for 5 minutes in mice or 15 minutes in rats, starting 20 minutes after the injection. EDRA ESSAY TESTED BY CARRAGENINE: A MODEL FOR INFLAMMATORY PAIN

The rat carrageenan-induced edema test follows that described by Winter et al. (Proc. Soc. Exp. Biol. Med. 1,962, 111: 544-547): solution on the lower surface of the right hind leg (0.75 mg per leg in 0.05 ml of physiological saline solution). 2 hours later, the rats are subjected consecutively to thermal and tactile stimulation, both of the non-inflamed and inflamed hind legs. For thermal stimulation, the device (Ugo Basile, Reference: 7371) consists of 6 boxes of individual Plexiglas (17 x 11 x 13 cm) placed on a raised glass floor. A rat is placed in the box and is released to get used for 10 minutes. Then, the inflamed and non-inflamed hind legs are focused with a mobile infrared radiation source (setting 20) and the leg withdrawal latencies are automatically registered. The removal of the leg interrupts the reflected radiation and disconnects the meter and the light source. To prevent tissue damage, if no reaction is observed, the test is terminated after 45 seconds. For tactile stimulation, the animal is placed under an inverted Plexiglas box (17 x 11 x 13 cm) on a grid floor. Then the tip of a Von Frey electronic probe with increasing pressure is applied to the non-inflamed and inflamed hind legs and the force required to induce the removal of the leg is automatically recorded. This procedure is carried out 3 times and the average force per leg is calculated to provide a base classification per animal. 3.5 hours later, the animals are sacrificed by a blow to the cervical vertebra and the hind legs are sectioned and weighed. An increase in leg weight (edema) indicates inflammation. This last procedure can also be applied to mice.

TEST HYPERTHERMIA TEST: A MODEL FOR INFLAMMATORY PAIN

Yeast hyperthermia test in the mouse or rat according to Teotino et al (J. Med. Chem. 1.963, 6: 248): First, the rectal temperature of the animals is measured using a rectal probe. Then a yeast suspension (512 mg / kg s.c.) is injected. 8 hours later, the test substance is administered. The rectal temperature of the mice is measured immediately before administration of the test substance and again 60 and 120 minutes later. COLORECTAL DISTENSION IN RATS: A MODEL FOR VISCERAL PAIN Experimental Procedure:

Animals (female Sprague Dawley rats, body weight: 208-257 g, five to seven per group) are fasted for 24 hours prior to experiments with free access to water. Acetic acid (0.6%, 1.5 ml) is injected into the colon (within 10 cm of the anus). After 50 minutes, a 5 cm long rubber balloon (volume 6-7 ml) is inserted rectally into the descending colon and is secured by connecting the attached tube to the rat's tail. The pressure of the balloon is adjusted to 100 mbar for 10 minutes. During this time the number of abdominal constrictions is recorded by visual inspection. Experiments are continued only with animals that respond to colorectal distention with more than 10 abdominal constrictions. These animals receive a single subcutaneous dose of an active compound or vehicle (10% Arlatone G, 10% Ethanol (96%) in Water) and the protocol for colorectal distension is repeated at 30, 60, 90 and 120 min after the administration.

Analysis of data:

The results are given as mean ± standard deviation. Both the number of abdominal constrictions at 30, 60, 90 and 120 minutes after the administration of substance or vehicle as well as the average values (30-120 min) are compared with previous values (time 0) by t-paired tests for bilateral limits. Values of p <0.5 are taken as statistically significant.

In addition, for each animal the relative number of constrictions (% of previous values) is calculated and the average values of the groups are provided. NEUROPROTECTING ACTIVITY: INDUCTION OF GROWTH FACTORS

The compounds of the invention (3 mg / kg, p.o.) or vehicle are administered once per day for a period of 3 weeks (n = 8 animals per treatment group). 24 hours after the last dose, the animals were sacrificed (using CO2 / O2 anesthesia), the brains were separated and dissected. From the individual brain samples the RNA is extracted and the induction of the growth factors BDNF and GDNF is determined by quantitative PCR. Total RNA is isolated from the brain pieces with the Trizol (Invitrogen) method. CDNA is prepared starting with 2 µg of total RNA (pretreated for 30 min with DNase (Ambion) in first strand buffer) using the reverse transcriptase Superscript II (Invitrogen). The quantification of mRNA by real-time PCR makes use of the observation that early PCR cycles are characterized by an exponential increase in target amplification. The accumulation of the PCR product is measured using Sybergreen II. Primers are designated using the Primer Express software package (Applied Biosystems). Expression levels of the maintenance genes ornithine decarboxylase (ODC_ex8) and alpha tubulin (TUBA) were used for normalization and control of a good cDNA synthesis.

HOT PLATE TEST: A MODEL FOR ACUTE PAIN

Mouse or rat hot plate assay according to Eddy and Leimbach (J. Pharmacol. Exp. Ther. 1953, 107: 385-393): Mice or rats are placed on a hot metal plate maintained at 54 ° C for mice or 52ºC for rats, surrounded by a Plexiglas cylinder (Height: 13 cm; Diameter: 19 cm). The latency time is measured until the first leg withdrawal (maximum: 30 seconds). TAIL WITHDRAWAL TEST: AN ACUTE PAIN MODEL

Mouse or rat tail removal test according to D'Amour and Smith (J. Pharmacol. Exp. Ther. 1.941, 72: 74-79): The animal's tail is heated by a thermal light source. The latency time is measured before the animal removes the tail (maximum: 15 seconds for mice, 30 seconds for rats). TRIALS OF RETORCIMIENTO WITH FENILBUTAZONA / ACETIC ACID: MODELS FOR ACUTE PAIN

Twisting assays with phenylbenzoquinone and acetic acid in mice follow the methods described by Hendershot et al (J. Pharmacol. Exp. Ther. 1959,

125: 237-240): Mice are injected with phenylbenzoquinone (PBQ) (1.25 mg / kg i.p.) or acetic acid (0.5% i.p.). This treatment induces a recognizable twisting response in control animals. The number of contortions is counted for 10 minutes, starting 5 minutes after injection of PBQ or acetic acid.

FREUND ASSISTANT TEST: A CHRONIC INFLAMMATORY PAIN MODEL

Chronic inflammatory pain test (trial with Freund's adjuvant) in the rat according to Whiteley (Current Protocols in Pharmacology, Wiley, NY, 5.5, 1999): An injection of Freund's adjuvant in rats induces chronic clinical symptoms of polyarthritis with pain . On Day 1, the rats are weighed and a suspension of Mycobacterium butyricum (Freund's adjuvant) is injected intradermally into the tail quarter (1 mg in 0.1 ml of paraffin oil). Control rats received a similar injection of paraffin oil. On Day 18, when the chronic condition was completely installed, the rats were weighed again and clinical symptoms of inflammation were evaluated. Then, the rats are consecutively subjected to thermal and tactile stimulation of both hind legs. To assess clinical symptoms, each leg is classified by inflammation according to a 5-point scale (0-4) and the tail is classified according to a 4-point scale (0-3), that is, a maximum classification of 19 per animal. For thermal stimulation, the device (Ugo Basile, Reference: 7.371) consists of 6 boxes of individual Plexiglas (17 x 11 x 13 cm) placed on a raised glass floor. A rat is placed in the box and left freely to get used for 10 minutes. Then, each posterior leg is focused with a mobile source of infrared radiation (setting 20) and the latency times of leg withdrawal are automatically recorded. The removal of the leg interrupts the reflected radiation and disconnects the meter and the light source. To prevent tissue damage, if no reaction is observed, the test is terminated after 45 seconds. For tactile stimulation, the animal is placed under an inverted Plexiglas box (17 x 11 x 13 cm) on a grid floor. Then the tip of a Von Frey electronic probe (Bioseb, Model 1610) with increasing pressure is applied to each rear leg and the force required to induce the removal of the leg is automatically recorded. This procedure is carried out 3 times and the average force per leg is calculated to provide a base classification per animal. Prior to receiving drug treatment, all animals are subjected to tactile stimulation and assigned to treatment groups based on their pain response.

RECEIVER LIGHTING EXPERIMENTS

Receiver ligation data were obtained by CEREP (128, rue Danton, 92500 Rueil-Malmaison, France) or Solvay Pharmaceuticals B.V., using well-documented standard procedures. For example, affinity for 5-HT1A receptors was determined by testing the ability of the compounds of the invention to displace [3H] -2- (di-n-propylamino) -8-hydroxytetralin ([3H] -8-OH-DPAT ) of their specific ligation sites in rat frontal cortex homogenates. This test is based on the method described by Gozlan et al. (Nature, 305, (1983), pages 140-142). DOSE

The affinity of the compounds of the invention for serotonin receptors was determined as described above. From the measured binding affinity for a given compound of formula (1), a lower effective theoretical dose can be estimated. At a concentration of the compound equal to twice the measured Ki value, 100% of the receptors will probably be occupied by the compound. Converting that concentration to mg of compound per kg of patient provides a lower theoretical effective dose, assuming an ideal bioavailability. Pharmacokinetic, pharmacodynamic and other considerations may modify the dose currently administered to a greater or lesser value. The conveniently administered dosage is 0.001-1000 mg / kg, preferably 0.1-100 mg / kg of the patient's body weight.

EXAMPLE I. ANALYTICAL METHODS USED DURING SYNTHESIS NUCLEAR MAGNETIC RESONANCE SPECTROCOPY (NMR)

NMR spectra were recorded with a Bruker AM400 spectrometer or a Varian VXR400S spectrometer. Chemical shifts (�) were indicated in

5 ppm below TMS as internal standard. A sample of 10-50 mg was dissolved in a deuterated solvent, usually CDCl3 or a mixture of DMSO-d6 / CDCl3 (4: 1 v / v). The solvent was selected to ensure complete dissolution of the sample. Free induction decays were generally obtained at room temperature under the following conditions: Digital resolution 0.2 Hz Scanning amplitude 18 ppm Pulse amplitude 20 degrees Pulse repetition time 4.5 s or greater if necessary for relaxation

complete Carrier wave frequency 6.0 ppm Number of acquisitions 128 or more, if necessary. The signs

C-13 satellites at 0.5% signal strength should be clearly visible. 10 To determine relative contents it was used as an NMR method.

TITULOMETRY (CHLORINE AND WATER DETERMINATIONS)

For potentiometric titrations a Metrohm E636 model (Switzerland) was used. In this synthesis potentiometric chloride determinations were used to determine chloride. The titration was performed with a combined silver electrode and

15 using silver nitrate as titration reagent. The method is specific for chloride because it can distinguish iodide chloride and bromide based on potentials other than electrodes.

Voltammetric titrations were performed for the determination according to Karl Fisher using a Metrohm 633 KF apparatus (Metrohm, Switzerland) according to the USP method.

20 EXAMPLE II. SYNTHESIS OF 3-AMINO-8- (1-PIPERAZINIL) -2H-1-BENZOPIRAN-2ONA AND ITS MONOCLORHYDRATE MONOHIDRATE (Compound 1) STAGE 1: NITRATION

The first stage was the nitration of 5-bromo-2-hydroxybenzaldehyde (1 *) that provided 5-bromo-2-hydroxy-3-nitrobenzaldehyde (2 *):

25

image3 OR image3 OR

image4 Br HBr

H AcOH OH

OH stage 1 NO2 recrist. (1 *) (stage 1a) (2 *)

)

A solution of 1.0 mol of 5-bromo-2-hydroxybenzaldehyde (1 *) in 3.75 liters of acetic acid (98%) was formed by heating the mixture to approximately 60 ° C. 1.5 moles of nitric acid were slowly added for approximately 1 hour

5 concentrate (137 g = 97 ml). After completing the addition, stirring was continued at 65 ° C for an additional 10 minutes. Then the solution was cooled to 45 ° C and the product was precipitated by adding 4 liters of water. After stirring for at least 3 hours, the product was collected on a filter and washed with water until the pH of the mother liquor was approximately 6. The material was

Dry as much as possible by centrifugation. The crude product was dissolved in 800 ml of acetone under reflux and with stirring. 400 ml of acetone were removed by distillation. After cooling to 20 ° C, the mixture was stirred for 3 hours. The precipitate was collected on a filter and washed with petroleum ether 40-65 ° C. The solid was dried overnight in a stream of air at 40 ° C. Finally the

The crude (2 *) product was recrystallized from acetone to provide a final product with a purity of 98% as indicated by NMR analysis. 5-Bromo-2-hydroxybenzaldehyde (1 *) was identified by its characteristic chemical shift � 9.84 ppm; 5-Bromo-2-hydroxy-3-nitrobenzaldehyde (2 *) had a characteristic chemical shift � of 10.4 ppm.

20 The total yield of this stage was approximately 60% (crude / crude).

STAGE 2: CONDENSATION OF ERLENMEYER

The second stage was Erlenmeyer's condensation of 5-bromo-2-hydroxy-3-nitrobenzaldehyde (2 *) with N-acetyl-glycine to provide N- (6-bromo-8-nitro-2-oxo2H-1-benzopyran-3 -il) acetamide (3 *).

25

image3 OR image3 OR

image3 OH H

N

Br image3 NCH3

Br

H

OR

H O

OH Ac2O

OO NO2 NaOAc NO2

(2 *) DMF (3 *) stage 2

To a mixture of 1.0 mol of 5-bromo-2-hydroxy-3-nitrobenzaldehyde (2 *), 1.0 mol of N-acetylglycine and 1.0 mol of anhydrous sodium acetate, 800 ml of N5 are added methyl-2-pyrrolidone. The mixture was stirred and heated to 50 ° C. Then 2.2 moles of acetic anhydride were introduced into the reaction vessel for approximately 30 minutes. The reaction mixture was heated to 100 ° C. During heating, the reaction mixture became homogeneous for a while; briefly after a solid formed that hindered the agitation. After heating at 100 ° C for 4-10 hours, the mixture was cooled to 80 ° C and 1,100 ml of acetic acid (98%) was added. Then stirring the mixture was easy. As a next step, the mixture was cooled to room temperature and stirred for 60 minutes. The precipitate was collected on a filter and washed twice with 625 ml of acetic acid (80%), five times with 900 ml of water and once with 300 ml of acetone. The product was dried in a stream of

15 air at 40 ° C for 24 hours, and had a purity of 98% as indicated by the NMR analysis. 5-Bromo-2-hydroxy-3-nitrobenzaldehyde (2 *) had a characteristic displacement of � 10.4 ppm; the characteristic chemical shift of N- (6-bromo8-nitro-2-oxo-2H-1-benzopyran-3-yl) acetamide (3 *) was � 8.72 ppm.

20 The total yield of this stage was approximately 80% (crude / crude).

STAGE 3: REDUCTION

The third stage was the catalytic hydrogenation of N- (6-bromo-8-nitro-2-oxo2H-1-benzopyran-3-yl) acetamide (3 *) to N- (8-amino-2-oxo-2H- 1-benzopyran-3-yl) acetamide (4 *).

H

H

image5 image3 N CH3 Br image3 NCH3 OOOPd / C

OO OR EtOH NH2

NO2 K2CO3

(4*)

stage 3

(3*)

5

10

fifteen

twenty

25

A mixture of 1.0 mol of N- (6-bromo-8-nitro-2-oxo-2H-1-benzopyran-3-yl) acetamide (3 *), 50 g of palladium on 10% carbon paste (which contains 61% water), 1.0 mol of potassium carbonate and 15 liters of ethanol was heated to 60 ° C. At this temperature the starting material was reduced with hydrogen at an overpressure of 4x105 Pa (4 bar) at 147 rad / s (1,400 rpm). After completing the reaction (1 hour), the catalyst was separated by filtration, using an auxiliary medium for filtration and washing with 4.5 liters of methyl ethyl ketone (MEK). The filtrate was concentrated to 2 liters and 2.3 liters of MEK were added. To change the ethanol solvent to MEK, 2 liters of the solvent mixture was removed by distillation at normal pressure and 2 liters of MEK were added. This was repeated 4 times. Then 5 liters of MEK and 2.6 liters of water were added and the mixture was stirred. The layers separated. The top layer was concentrated under normal pressure to approximately 3.5 liters. The residue was cooled to 25 ° C. During this cooling the product crystallized. The mixture was then cooled to -10 ° C and stirred for two hours. The solid was filtered and washed three times with 800 ml of hexane. The product was dried (50 ° C, 20 cm Hg, N2) to constant weight.

N- (6-Bromo-8-nitro-2-oxo-2H-1-benzopyran-3-yl) acetamide (3 *) had a characteristic chemical shift of � 8.72 ppm; the displacement of N- (8 amino-2-oxo-2H-1-benzo-piran-3-il) acetamide (4) was � 8.55 ppm.

The total yield of this stage was approximately 70% (crude / crude).

STAGE 4: CONSTRUCTION OF THE PIPERAZINE ANNULAR SYSTEM

Step 4 was the alkylation of N- (8-amino-2-oxo-2H-1-benzopyran-3-yl) acetamide (4 *) with bis-chloroethylamine providing N- (8- (1-piperazinyl) -2-oxo -2H1-benzopyran-3-yl) acetamide (5 *).

HCl image3

image3 Cl

image3 N CH3H image3 N CH3 NO

H

OO

OR image3 N image3 OO

MCB

NH2

stage 4

N

recrist

(4 *) .H (5 *)

(stage 4a)

to)

A mixture of 2.5 liters of monochlorobenzene, 1.0 mol of N- (8-amino-2-oxo2H-1-benzo-pyran-3-yl) acetamide (4 *) and 1.2 moles of bischloroethylamine hydrochloride It was heated to reflux under nitrogen. Part of the monochlorobenzene (0.5 liters) was distilled off. This mixture was heated at reflux for 10 days. The reaction was followed by HPLC. After the reaction, the mixture was cooled to 20 ° C and stirred overnight. The solid product was collected on a filter and washed once with 360 ml of monochlorobenzene and 3 times with 360 ml of ethanol. The product was dried under vacuum at 50 ° C.

5 Half of the crude product was dissolved in 3 liters of water. After the addition of 18 g of Celite and 50 g of charcoal, the mixture was stirred for 1 hour at room temperature. After filtration the solution was concentrated by distillation of water. Meanwhile, the second half of the crude product was treated as previously described. When the total volume of aqueous solutions

10 combined was approximately 1.5 liters, distillation was stopped and the mixture was cooled to room temperature. Then 125 g of sodium bicarbonate were added portionwise. After stirring for 1.5 hours at 15 ° C, the precipitate formed was collected on a filter. After washing with 360 ml of water and 2 times with 270 ml of ethanol, the product was dried under vacuum at 50 ° C.

The N- (8-amino-2-oxo-2H-1-benzo-pyran-3-yl) acetamide (4 *) had a characteristic chemical shift of � 8.55 ppm; the displacement of N- (8- (1piperazinyl) -2-oxo-2H-1-benzopyran-3-yl) acetamide (5 *) was � 8.57 ppm. The total yield of this stage was approximately 50% (crude / crude). STAGE 5: AMIDA HYDROLYSIS

Step 5 was the hydrolysis of the amide function of N- (8- (1-piperazinyl) -2-oxo2H-1-benzopyran-3-yl) acetamide (5 *) using hydrochloric acid. This resulted in the trichlorohydrate salt of 3-amino-8- (1-piperazinyl) -2H-1-benzopyran-2-one (6 *).

image3 NH2

H image6 N CH3 OO O

image3 N image3

OO image3 N image3

.3HCl EtOH

N H2O H N

H (5 *) stage 5 (6 *)

At room temperature, 2.9 liters of concentrated hydrochloric acid were added for about 10 minutes to a 1.0 mol suspension of N- (8- (1piperazinyl) -2-oxo-2H-1-benzopyran-3-yl) acetamide (5 *) and 1.4 liters of absolute ethanol. During this aggregate the temperature increased to 40 ° C. After the addition the mixture was stirred at a temperature of 50 ° C for 1.5 hours. The mixture was cooled to

20-20 ° C and, after crystallization started, 1.4 liters of absolute ethanol was added. The mixture was then stirred for 1 hour at 20 ° C and for 2 hours at 0 ° C. The crystals were isolated by filtration and washed twice with 0.6 liters of acetone. The isolated product was dried under vacuum (40 ° C, 200 mm Hg, N2, 24 hours).

N- (8- (1-piperazinyl) -2-oxo-2H-1-benzopyran-3-yl) acetamide (5 *) had a

5 characteristic chemical shift of � 8.57 ppm; the trichlorohydrate salt of 3-amino-8- (1-piperazinyl) -2H-1-benzopyran-2-one (6 *) had a characteristic chemical shift of � 6.77 ppm.

The total yield of this stage was approximately 85% (crude / crude).

STAGE 6: PARTIAL NEUTRALIZATION

10 The final stage, the sixth, was the partial neutralization of the trichlorohydrate (6 *) with sodium bicarbonate to produce the desired product: COMPOUND 1, the monohydrate of the 3-amino-8- (1-piperazinyl) -2H monohydrochloride -1-benzopiran-2ona (7 *).

image3 NH2 image3 NH2

O O NaHCO3

OO image3 N image3

image3 N image3 EtOH

.3HCl .HCl.H2O

N stage 6 NH

recrist H (stage 6a)

(6 *) (7 *)

fifteen

To a suspension of 1.0 mol of the trichlorohydrate salt (6 *) in 3.5 liters of ethanol was added for approximately 30 minutes a solution of 2.2 moles of sodium bicarbonate in 2.8 liters of water. The temperature was between 20 ° C and 25 ° C. Then the suspension was stirred for 3 hours. The reaction mixture is

20 filtered and subsequently washed with 1.1 liters of water, 1.1 liters of ethanol and 1.1 liters of hexane. The isolated crude product was dried in vacuo (40 ° C, 200 mm Hg, N2, 24 hours).

The dried product (1 mol) was dissolved in 9 liters of methanol by heating at reflux temperature. The solution did not become completely transparent. After cooling to 20 ° C the mixture was filtered. 300 ml of water and 150 ml of methanol were added to the filtrate, after which approximately 3 liters of the solvent mixture was distilled at normal pressure. The entire procedure was repeated with another mole of the dried product. The combined fractions were then concentrated to a volume of approximately 12 liters by distillation. After the addition of

6 liters of ethanol, 6 liters of the solvent mixture were removed by distillation under normal pressure. The mixture was then cooled to 0 ° C and stirred for 2 hours. The precipitate was collected on a filter and washed twice with 750 ml of acetone. The product was dried under vacuum (40 ° C, 200 mm Hg, N2, 24 hours) and then homogenized by crushing and, if necessary, by micronization.

The trichlorohydrate salt of 3-amino-8- (1-piperazinyl) -2H-1-benzopyran-2-one (6 *) had a characteristic chemical shift of � 6.77 ppm; The displacement of the final product, COMPOUND 1, was 6.7 ppm.

The total yield of this stage was approximately 85% (crude / crude).

COMPOUND 1, the 3-amino-8- (1piperazinyl) -2H-1-benzopyran-2-one monohydrate monohydrate, had a molecular formula C13H18ClN3O3 and a molecular mass of 299.5. The pure product (99.8%, NMR) was a white to yellowish powder. Its chloride content was 11.7% (m / m), as determined by titulometry. Its water content, determined by the water titration test according to Karl Fisher, was 6.5% (m / m). EXAMPLE III: FORMULATION OF COMPOUND 1

For oral administration (p.o.): to the desired amount (0.5-15 mg) of Compound 1 in a glass tube some glass beads were added and the substance was crushed by swirling for 2 minutes. After the addition of 1 ml of a 1% aqueous solution of methylcellulose, the compound was suspended by stirring for 10 minutes with vortexing. For concentrations up to and greater than 1 mg / ml, the particles that remained in the suspension were still suspended by an ultrasonic bath.

For intraperitoneal administration (i.p.): to the desired amount (0.5-15 mg) of solid compound 1 in a glass tube, some glass beads were added and the solid was triturated by swirling for 2 minutes. After the addition of 1 ml of a 1% aqueous solution of methylcellulose and 5% mannitol, the compound was suspended by stirring for 10 minutes with vortexing. Finally, the pH was adjusted to 7. EXAMPLE IV: RESULTS OF PHARMACOLOGICAL TESTS COLORECTAL DISTENSION IN RATS: A VISCERAL PAIN MODEL

Colorectal distention after sensitization with acetic acid (according to the protocol given above) resulted in comparable numbers of abdominal constrictions in all groups. The lowest dose of Compound 1 (0.1 µmol / kg) was dissolved in 10% Arlatone G, 10% Ethanol (96%) in Water and reduced visceral hypersensitivity to 90 minutes after administration. A comparable vehicle (20% Arlatone G, 20% Ethanol (96%) in Water) was tested in a control group. No significant reduction in the number of abdominal constrictions was observed during the observation of the distention protocol.

5 repetitive colorectal. Compound 1 exhibited a significant inhibition of the number of abdominal contractions due to colorectal distension after sensitization with acetic acid in the dose range between 3 and 10 µmoles / kg, 90 minutes after subcutaneous administration. The vehicle used for this dose did not interfere with the experimental protocol.

10 INDUCTION OF GROWTH FACTORS Treatment with Compound 1 increased the levels of GDNF and BDNF RNA in the thalamus, striatum, prefrontal cortex, nucleus accumbens and hippocampus (see table).

RNA regulation of growth factors by treatment with Compound 1 as determined by quantitative PCR. The data are expressed as a multiple of levels obtained by vehicle treatment.

Brain area

GDNF BDNF

thalamus

2.6 (p <0.001) 2.3 (p <0.0001)

striated body

2.2 (p <0.00005) 1.8 (p = 0.12)

prefrontal cortex

1.3 (p = 0.24) 2.1 (p <0.0005)

nucleus accumbens

1.5 (p <0.05) 5.1 (p <0.05)

hippocampus

1.7 (p <0.001) 2.4 (p <0.0001)

HOT PLATE TEST: A MODEL FOR ACUTE PAIN

Compound 1 and other compounds were tested on the hot plate,

minutes after intraperitoneal administration. A description of the method is

He gave before.

Drug treatment

Latency of the animal to withdraw from the hot plate, seconds + Standard deviation

physiological saline solution

4.5 + 0.3

morphine (7.5 mg / kg i.p.)

7.8 + 0.3

compound 1 (0.1 mg / kg i.p.)

8.4 + 0.5

sumatriptan (10 mg / kg i.p.)

3.9 + 0.3

comp 1 (0.1 i.p.) + sumatriptan (10 i.p.)

4.7 + 0.3

From the data indicated in the previous table it is clear that morphine almost doubles the latency of animals to remove from the hot plate: a relevant analgesic effect. Compound 1 is as effective as morphine, but at a much lower dose. The sumatriptan 5-HT1D selective agonist is inactive at the very high dose

10 of 10 mg / kg i.p., but at that dose it almost completely antagonizes the analgesic effect of compound 1 (P value = 0.0001, Wilcoxon test). Compound 1 is also orally active in this test in rats: its ED50 is 3.2 mg / kg (p.o.)

TEST OF REINFORCEMENT WITH ACETIC ACID: ACUTE PAIN MODEL 15

Compound 1 is active in this assay: its ED50 is 1.82 mg / kg p.o.

BINDING PROFILE TO THE COMPOUND RECEIVER 1.

Ligation data collected in the following table were obtained by CEREP (128, rue Danton, 92500 Rueil-Malmaison, France) or Solvay Pharmaceuticals 20 B.V., using well-documented standard procedures.

receiver

S1 radioligating Ki (nM) Compound 1

5-HT1A

h [3H] -8-OH-DPAT 0.25

5-HT1B

r [125I] -cianopindolol 2.0

5-HT1D

b [3H] -serotonin 13

5-HT2A

h [3H] -quetanserin 630

5-HT2B

h [3H] -LSD 320

5-HT2C

h [125I] -DOI > 1,000

5-HT3

h [3H] -BRL 43694 250

5-HT4

h [3H] -GR 113808 > 1,000

5-HT5

h [3H] -LSD 100

5-HT6

h [3H] -LSD > 1,000

5-HT7

h [3H] -LSD 3.2

5-HTreabsorption

h [3H] -paroxetine > 1,000

�1-adrenergic

r [3H] -prazosin > 1,000

�1A-adrenergic

r [3H] -prazosin 630

�1B-adrenergic

r [3H] -prazosin > 1,000

�2-adrenergic

r [3H] -RX 821002 > 1,000

�1-adrenergic

h [3H] -CGP 12177 fifty

�2-adrenergic

h [3H] -CGP 12177 40

�3-adrenergic

h [125I] -iodocianopindolol > 1,000

NAreabsorption

h [3H] -nisoxetine > 1,000

Dopamine-D1

h [3H] -SCH 23390 > 1,000

Dopamine-D2

h [3H] -spiperone > 1,000

Dopamine-D3

h [3H] -spiperone > 1,000

Dopamine-D4

h [3H] -spiperone > 1,000

Dopamine-D5

h [3H] -SCH 23390 > 1,000

Dopamine reabsorption

h [3H] -GBR 12935 > 1,000

Muscarina-M1

h [3H] -pirenzepine > 1,000

Muscarina-M2

h [3H] -AFDX-384 > 1,000

Muscarina-M3

h [3H] -4-DAMP > 1,000

Muscarina-M4

h [3H] -4-DAMP > 1,000

Muscarina-M5

h [3H] -4-DAMP > 1,000

Histamine-H1

h [3H] -pyrilamine > 1,000

Histamine-H2

h [125I] -APT > 1,000

Histamine-H3

r [3H] -�-methylhistamine > 10,000

tryptamine

r [3H] -triptamine > 10,000

melatonin

C [125I] -2-iodomelatonin > 10,000

nicotine

r [3H] -citisin > 10,000

�-opiate

r [3H] -DAMGO > 1,000

�-opiate

r [3H] -U 69593 > 1,000

�-opiate

r [3H] -DPDPE > 1,000

nociceptin

h [3H] -nociceptin > 1,000

receiver

S1 radioligating Ki (nM) Compound 1

(ORL1)

sigma

r [3H] -DTG > 1,000

sigma-SG1

g [3H] -pentazocone > 1,000

sigma-SG2

r [3H] -DTG > 1,000

cannabinoid CB1

h [3H] -WIN 55.212-2 > 10,000

Ca ++ channel

p [3H] -fluspirileno > 10,000

Ca ++ channel

r [3H] -nitrendipine > 10,000

Ca ++ channel

r [3H] -diltiazem > 10,000

Ca ++ channel

r [125I] -�-conotoxin > 1,000

Ca ++ channel

r [3H] -D-888 130

Ca ++ channel

r [3H] -devapamil > 10,000

Na + channel

r [3H] -batracotoxinin > 10,000

K + channel

r [125I] -�-dendrotoxin > 1,000

K + channel

r [125I] -apamine > 1,000

Adenosine-A1

h [3H] -DPCPX > 1,000

Adenosine-A2A

h [3H] -CGS 21680 > 1,000

Adenosine-A3

h [3H] -AB-MECA > 1,000

Purina-P2X

r [3H] -ab-MeATP > 1,000

GABAA

r [3H] -muscimol > 10,000

GABAB

r [3H] -PK 11195 > 1,000

Glycine

r [3H] -stricnin > 10,000

Strychnine insensitive glycine

r [3H] -MDL105519 > 10,000

NMDA

r [3H] -CGS 19755 > 10,000

angiotensin-AT1

h [125I] -angiotensin II > 1,000

angiotensin-AT2

h [125I] -CPG 42112A > 1,000

benzodiazepine

r [3H] -diazepam > 10,000

bombesina

r [125I] -bombesin > 1,000

bradykinin

h [3H] -bradiquinine > 1,000

CCKA

h [3H] -devazepide > 1,000

CCKB

h [3H] -CCK8 > 1,000

CCR1

h [125I] -MIP-1a > 1,000

CGRP

h [125I] -CGRPa > 1,000

CRF

h [125I] -oCRF > 10,000

Endothelin-ETA

h [125I] -endothelin-1 > 1,000

Endothelin-ET8

h [125I] -endothelin-1 > 1,000

Galanina-GAL1

h [125I] -galanine > 1,000

Galanina-GAL2

h [125I] -galanine > 1,000

Interleukin-6

h [125I] -interleucine-6 > 10,000

Interleukin-8

h [125I] -interleucine-8 > 1,000

LTB4

g [3H] -LTB4 > 10,000

LTD4

g [3H] -LTD4 > 10,000

melanocortin

h [125I] -NDP-a-MSH > 1,000

receiver

S1 radioligating Ki (nM) Compound 1

Neuroquinine NK1

h [3H] -substance P > 1,000

Neuroquinine NK2

h [125I] -neurokininA > 1,000

Neuroquinine NK3

h [3H] -SR 142801 > 1,000

Neuropeptide Y1

h [125I] -PYY > 1,000

Neuropeptide Y2

h [125I] -PYY > 1,000

Neurotensin-NT1

h [125I] -neurotensin > 1,000

PACAP

r [125I] -PACAP 1-27 > 1,000

Prostaglandin-I2

h [3H] -iloprost > 1,000

Prostaglandin H2

h [3H] -SQ 29548 > 1,000

somatostatin

m [125I] -somatostatin > 1,000

HRT

r [3H] -TRH > 10,000

Tumor necrosis factor

r [125I] -TNF� > 1,000

Vasopressin-V1A

h [3H] -vasopressin > 1,000

VIP1

h [125I] -VIP > 1,000

S1: b = bovine, c = chicken, g = guinea pig, h = human being, m = mouse, p = pig; r = rat.

Claims (2)

1. Use of a compound of formula (1): image 1 5 where: R1 is (C1-4) alkyl, (C1-4) alkoxy, hydroxyl, (C1-4) alkoxy (C1-4) alkyl, pyrrolidinyl, piperidinyl, morpholinyl, halogen, cyano, trifluoromethyl, amino or mono- or disubstituted amino where the substituents are (C1-4) alkyl or (C14) alkylcarbonyl, 10 m has the value 0, 1 or 2, R2 is (C1-4) alkyl, (C1-4) alkoxy, halogen or trifluoromethyl, n is 0 or 1, with the proviso that (m + n) is at least 1, R3 is hydrogen, (C1-3) alkyl or (C2-3) alkenyl, R4 is (C1-4) alkyl and 15 p has the value 0, 1 or 2, and all stereoisomers and pharmacologically acceptable salts thereof, for the preparation of a pharmaceutical composition for the treatment of pain. 2. Use according to claim 1, wherein said pain is chronic pain including nociceptive, neuropathic, psychogenic and mixed category 20 pain (nociceptive and neuropathic components), including, but not limited to, diabetic neuropathy, pain neurogenic, central pain, somatic pain, visceral and cancer pain, inflammatory pain, post-operative pain, chronic waist pain, sciatica, cervical and lumbar pain, tension headaches, outbreak headaches, chronic daily headaches , herpes neuralgia and post neuralgia 25 herpes, facial and oral neuralgia and myofacial pain syndromes, phantom limb pain, stump pain and paraplegic pain, dental pain, opioid-resistant pain, post-surgical pain including cardiac surgery and mastectomy, labor and delivery pain childbirth, postpartum pain, post-infarction pain, angina pain, genito-urinary tract pain including pelvic pain and cystitis and vulvar vestibulitis and orchialgia, irritable bowel syndrome, premenstrual syndrome pain, pain resulting from burns or chemical damage or sunburn and bone damage pain.