Semicarbazide-sensitive amine oxidase inhibitors for use as analgesics in traumatic neuropathy and neurogenic inflammation

Abstract

The invention relates to a compound having SSAO/VAP-1 inhibitor activity for use in the treatment of hyperalgesia and allodynia implicated in traumatic neuropathy or neurogenic inflammation. Accordingly, the invention relates to a compound having SSAO/VAP-1 inhibitor activity for use in the inhibition of pathological activation and dysfunctions of peptidergic sensory nerves caused by mechanical damage or chemical activation of peptidergic sensory nerves in neurogenic inflammation.

Claims

1. A method for the treatment of hyperalgesia and allodynia implicated in traumatic neuropathy or neurogenic inflammation, said method comprising administering a compound having SSAO/VAP-1 inhibitor activity to a subject in need of said treatment, wherein the compound has the general formula of Ar(CH2)n-CR1=NOR2 (I) or salt, hydrate or solvate thereofwherein Ar is a group of the formula: ##STR00002## R1 is H or lower alkyl; R2 is H, lower alkyl, benzyl, (CH2)k-COOR13, (CH2)m-N(R14R15) or CONHR16, wherein R13 is lower alkyl, k is 1, 2 or 3; m is 1, 2 or 3; R14 and R15 are independently from each other lower alkyl, or R14 and R15 together with the nitrogen they are atteched form a 5 to 7 membered heteroring, optionally containing 1 to 3 further heteroatom(s) selected from nitrogen, oxygen and sulfur atoms, R16 is phenyl, optionally substituted with one or more group selected from halogen, lower alkyl and lower alkoxy; R3 and R4 together with the carbons they are attached to form a 5 to 7 membered heteroring containing one or two oxygen(s), preferably 1,3-dioxolane, optionally substituted with lower alkyl; or R3 is H, halogen, lower alkyl or OR17, wherein R17 is H, lower alkyl, lower alkenyl, optionally substituted with phenyl; R4 is H or OR19, wherein R19 is lower alkyl; R5 is H or halogen; R6 is H or halogen; R7 is H, halogen, OH, OR20 or a phenyl substituted with Z wherein R20 is lower alkyl or lower alkenyl, and Z is CHNOH or halogen; X is a 5 to 7 membered heteroring containing 1 or 3 heteroatom(s) selected from nitrogen, oxygen and sulfur atoms; Y is SR21, OR22, 5 to 7 membered heteroring containing 1 to 3 heteroatom(s) selected from nitrogen, oxygen and sulfur atoms, optionally substituted with phenyl or a (lower alkenyl)amino, optionally N-substituted with lower alkyl; wherein R21 is lower alkyl or phenyl and R22 is lower alkyl; R8 is lower alkyl or optionally substituted benzyl, wherein the substituent is 1 or 2 lower alkoxy; R9 is H or phenyl; R10 is di(lower alkyl)amino, preferably dimethylamino, 5 to 7 membered heteroring containing 1 or 3 heteroatom(s) selected from nitrogen, oxygen and sulfur atoms, optionally substituted with one or more group selected from lower alkyl, lower alkenyl and phenyl; or R9 and R10 together with the attached carbon atoms form an optionally substituted 5 to 8 membered heteroring containing 1 or 3 heteroatom(s) selected from nitrogen, oxygen and sulfur atoms, optionally substituted with one or more group selected from lower alkyl and benzyl, and optionally together with lower alkylene form a fused bicyclic group; W is a bond or a phenylene group; R11 is lower alkyl; R12 is phenyl, optionally substituted with halogen; n is integer of 0 to 4; Ar1 and Ar2 are the same or different and stand for phenyl, optionally substituted with one or more group selected from halogen, lower alkyl and lower alkoxy; and any stereoisomer, mixture of stereoisomers, E or Z forms, mixture of E and Z forms, crystalline form, non-crystalline form thereof; wherein lower alkyl refers to aliphatic and alicyclic groups including straight-chain (linear), branched-chain or cyclic groups having up to 6 carbon atoms; lower alkenyl refers to unsaturated aliphatic and alicyclic groups including straight-chain (linear), branched-chain, cyclic groups, and combinations thereof, having up to 6 carbon atoms, which contain at least one double bond (CC); lower alkoxyrefers to a (lower alkyl)-Ogroup, where the lower alkylhas the above-defined meaning.

2. The method of claim 1, wherein the compound is selected from the group consisting of: 3-methoxy-2-methylbenzaldehyde oxime; 8-Pyrrolidino-1-naphthaldehyde oxime; 5-Hydroxy-1,3-benzodioxole-4-carbaldehyde oxime; 5-Ethoxy-1,3-benzodioxole-4-carbaldehyde oxime; 5-(Allyloxy)-1,3-benzodioxole-4-carbaldehyde oxime; 5-Bromo-1,3-benzodioxole-4-carbaldehyde oxime; 5-{2-[(Hydroxyimino)methyl]phenyl}-1,3-benzodioxole-4-carbaldehyde oxime; 6-Ethoxy-1,3-dimethyl-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbaldehyde oxime; 6-Methoxy-1,3-dimethyl-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbaldehyde oxime; 1,3-Dimethyl-2,4-dioxo-6-propoxy-1,2,3,4-tetrahydropyrimidine-5-carbaldehyde oxime; 6-Ethoxy-1,3-dimethyl-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbaldehyde-O-methyloxime; 1,3-Dimethyl-2,4-dioxo-6-(propylthio)-1,2,3,4-tetrahydropyrimidine-5-carbaldehyde oxime; 1,3-Dimethyl-6-(methylthio)-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbaldehyde oxime; 6-(Ethylthio)-1,3-dimethyl-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbaldehyde oxime; 1,3-Dimethyl-2,4-dioxo-6-(phenylthio)-1,2,3,4-tetrahydropyrimidine-5-carbaldehyde oxime; 6-(Ethylthio)-1,3-dimethyl-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbaldehyde-O-methyloxime; 6-[Allyl(methyl)amino]-1,3-dimethyl-2,4-dioxo-1,2,3,4-tetrahydropyrimidine-5-carbaldehyde oxime; 3-(4,5-Diphenyl-1,3-oxazol-2-yl)propanal oxime and 1-(4-Chlorobenzoyl)-2-methyl-1H-indole-3-carbaldehyde oxime, or any stereoisomer, mixture of stereoisomers, E or Z forms, mixture of E and Z forms, crystalline form, non-crystalline form, hydrate, solvate or salt thereof.

3. The method of claim 2, wherein the compound is the 3-(4,5-diphenyl-1,3-oxazol-2-yl)propanal oxime.

4. A method for the treatment of hyperalgesia and allodynia implicated in traumatic neuropathy or neurogenic inflammation, said method comprising administering a compound having SSAO/VAP-1 inhibitor activity to a subject in need of said treatment, wherein the compound is; (2-phenyl-2-propen-1-yl)hydrazine or pharmaceutically acceptable salt thereof.

5. The method of claim 1, wherein the subject is suffering from traumatic neuropathy or neurogenic inflammation.

6. The method of claim 1, wherein the subject is suffering from pathological activation and a dysfunction of peptidergic sensory nerves caused by mechanical damage or chemical activation.

7. The method of claim 1, wherein the subject is suffering from severe hyperalgesia or allodynia.

Description

SUMMARY OF DRAWINGS

(1) FIGS. 1 A and B: The test protocol is shown schematically for measurement of the mechanical touch sensitivity and cold tolerance in the traumatic mononeuropathy model in rats (A) and mice (B).

(2) FIGS. 2 A and B: Effect of SZV-1911 and SZV-1287 on sciatic nerve ligation-induced mechanical allodynia on the injured (A) and the controlateral (B) paw of the rats 7 days after the operation in comparison with the vehicle.

(3) FIGS. 3 A and B: Effect of SZV-1911 and SZV-1287 on the mechanonociceptive thresholds of the ipsilateral (A) and of the contralateral (B) paw of sham-operated rats in comparison with the vehicle.

(4) FIGS. 4 A-D: Effect of SZV-1911 (A-B) and SZV-1287 (C-D) on the mechanonociceptive thresholds of the injured paw of sciatic nerve operated mice in comparison with the vehicle on Day 3 and on Day 7, respectively.

(5) FIGS. 5 A-D: Effect of SZV-1911 (A-B) and SZV-1287 (C-D) on the paw withdrawal latency of the injured paw of sciatic nerve operated mice in comparison with the vehicle on Day 3 and on Day 7, respectively.

(6) FIGS. 6 A and B: The test protocol is shown schematically for measurement of the noxious heat thresholds and mechanonociceptive thresholds after i. pl. resiniferatoxin (A) and formalin (B) administration.

(7) FIGS. 7 A and B: Effects of SZV-1911 and SZV-1287 on RTX-induced thermal (A.) and mechanical (B.) hyperalgesia in mice in comparison with the vehicle.

(8) FIGS. 8 A-C: Effects of SZV-1287 on formalin-induced acute nocifensive reactions (A) and the drop of thermo-(B) and mechanonociceptive (C) thresholds of mice.

METHODOLOGY

Experimental Models, Protocols and Measuring Techniques

(9) Solution Preparation

(10) The solutions were prepared freshly on the day of the experiment with sterile saline. SZV-1287 (prepared according to WO 2010/029379 A1) dissolved in saline and the solution was clear. The SZV-1911 (prepared according to WO2006/094201 and WO2005/014530) solution was prepared with Tween 80 and ethanol, stirred properly with a vortex, heated moderately to 42 C. and sonicated, but it was still opalescent. The i.p. administered volume was 0.2 ml/100 g body weight from 10 mg/ml in rats solution and 0.1 m/10 g body weight from 2 mg/ml solution to achieve the 20 mg/kg doses. The solvent of SZV-1911 was given to the vehicle-treated animals in the same volume.

(11) Experimental Animals and Ethical Issues

(12) Sciatic nerve ligation-induced traumatic mononeuropathy experiments were performed on male Wistar rats (250-300 g), and on male C57Bl/6 mice as well as resiniferatoxin- or formalin-induced hyperalgesia. The original breeding pairs of the animals were purchased from Charles River Ltd. All the animals were kept in the Animal Facility of the Department of Pharmacology and Pharmacotherapy at the University of Pcs at 24-25 C. provided with standard chow and water ad libitum.

(13) All experimental procedures were carried out according to the European Communities Council Directive of 86/609/EEC. The studies were approved by the Ethics Committee on Animal Research, University of Pecs. All efforts were made to minimize animal-suffering, to reduce number of used animals and to utilize in vivo techniques if available.

(14) I. Neuropathy Models

(15) Experimental Protocol, Model and Investigational Techniques

(16) Before all experiments one conditioning and three reliable control threshold measurements were performed on three consecutive days. All operations were performed by the same operator, who was blinded to the animal group assignments and the treatment allocation (FIG. 1A for the rat, B for the mouse experimental paradigm).

(17) Investigation of Mechanical Hyperalgesia and Cold Allodynia in Sciatic Nerve Ligation-Induced Traumatic Mononeuropathy

(18) On the 3.sup.rd and on the 7.sup.th day following the nerve ligation, the mechano-nociceptive thresholds were determined directly before and 15 min after i.p. drug administration to observe the difference between pre-injection and post-injection thresholds in mice. We determined the developing cold allodynia following the sciatic nerve ligation at Days 3 and 7. In rats we examined the developing mechanical allodynia just 7 days after the sciatic nerve ligation on the same way like in mice. Only animals with a minimum of 25% hyperalgesia/allodynia were included in the study (Seltzer et al., Pain., 1990, 43(2):205-18; Pintr et al., Naunyn Schmiedebergs Arch Pharmacol., 2002, 366(2):142-50.).

(19) The Sciatic Nerve Operation Procedure

(20) Mice were anaesthesized with a combination of ketamine and xylazine (100 mg/kg and 5 mg/kg i.p. respectively), but in rats Na-pentobarbital (40 mg/kg i.p) were used. The common sciatic nerve was exposed unilaterally on the right side high in the thigh and - of the nerve trunk was carefully separated and tightly ligated using a siliconised silk suture (Ethicone 5-0 and 8.0, respectively). Then the wound was closed and the animals were allowed to survive for 8 days (Seltzer, 1990; Malmberg and Basbaum 1998; Bolcskei et al. 2005; Sandor et al. 2010). During this period, signs of spontaneous pain (holding the legs in elevated position transiently for 2-3 days) and mechanical hyperalgesia and cold allodynia developed. In order to investigated the efficacy of these drugs we examined the drop of the mechanical touch sensitivity on Day 3, however the decrease of the mechanical threshold develops fully 7 days after the surgery.

(21) Cold Stimulation

(22) A noxious cold stimulus was applied by immersing one of the murine hindpaws in icy water, remaining the temperature steady at 0 C. Cold perception was assessed as the time taken by the animal to pulling out its paw from the cold water, the cut-off time was 180 seconds. The paw withdrawal behaviour represents a consistent and reproducible method for the determination of the hypersensitivity of cold nociception after sciatic nerve ligation.

(23) Measurement of the Mechanical Touch Sensitivity

(24) The mechanical touch sensitivity thresholds of the plantar surface of the paws was determined by aesthesiometry (Ugo Basile Dynamic Plantar Aesthesiometer 37400; Comerio, Italy). This is an electronic von Frey device, in which the animals moved about freely in one of the compartments of the enclosure positioned on the metal mesh surface. Following acclimation after cessation of exploratory behaviour, the touch stimulator unit was placed under the animal's paw, using the adjustable angled-mirror to position the filament below the target area of the plantar surface. Then an electrodynamic actuator of proprietary design lifted a straight metal filament, which touched the plantar surface and began to exert an increasing upward force at a preset rate of application until a stop signal (removal of the paw) was attained. The paw withdrawal threshold was obtained in grams. Hyperalgesia or allodynia (decrease of the withdrawal thresholds) was expressed as percentage by comparing the data of each individual animal to the averaged three initial control thresholds.

(25) Statistical Analysis

(26) Results are expressed as the means.e.m. The pre-drug and the post-drug data pairs within each group were compared with 2-way ANOVA following Bonferroni's post-test. *p<0.05, **p<0.01; ***p<0.001 were considered to be significant.

(27) The test protocol is shown schematically on FIGS. 1 A and B.

(28) Analgesic Action of SSAO Inhibitors in Traumatic Mononeuropathy

(29) Effects on Sciatic Nerve Ligation-Induced Hyperalgesia in Rats

(30) Seven days after the nerve ligation an approximately 40-50% mechanical allodynia developed on the operated limb of the rats. The vehicle (0.2 ml/100 g body weight) did not alter the mechanical allodynia 15 minutes later (41.5+7.2% vs 46.7+5.7% pre-injection value), but pretreatment with the 20 mg/kg i.p. doses of both SZV-1287 and SZV-1911 significantly reduced the 48.86.2% and 47.87.9% pre-injection allodynia to 34.67.6% and 32.93.4%, respectively. The approximately 30% inhibitory effects of the two compounds did not differ significantly from each other. No change of the mechanonociceptive thresholds was detected either on the contralateral side of the operated (FIG. 2.B) or in the sham-operated group. The results are shown in FIGS. 2 A and B.

(31) Allodynia values were calculated as % by comparing the mechano-nociceptive thresholds measured on the 7.sup.th day to the mean initial threshold values. Column pairs represent allodynia before drug injection and 15 min after compound/vehicle administration. Results are expressed as meanss.e.m. of the mechanonociceptive threshold changes of n=7 rats in the vehicle-treated and 8-8 rats in both compound-treated groups. Data were analysed with One-way ANOVA following Bonferroni's post-test in comparison to determine differences between the pre- and post-injection values of the respective groups, *p<0.05.

(32) The effects of SZV-1911 and SZV-1287 on the mechanonociceptive thresholds of the ipsilateral (A) and of the contralateral (B) paw of sham-operated rats in comparison with the vehicle are shown on FIGS. 3A and 3B.

(33) Effects on Sciatic Nerve Ligation-Induced Hyperalgesia in Mice

(34) Investigating the effects of SSAO inhibitors we performed the sciatic nerve ligation in TRPA1 wildtype mice. 3 days after the operation the developing mechanical hyperalgesia was approximately 40% in all groups which was even increased on Day 7. The hyperalgesia was significantly diminished to 15 minutes after the i.p. administration of a single dose of 20 mg/kg SZV-1911 compared to their postoperative control values.

(35) Investigating the cold allodynia which is a symptom of the sciatic nerve ligation caused traumatic mononeuropathy, we did not observed any significant changes.

(36) The effects of SZV-1911 (A-B) and SZV-1287 (C-D) on the mechanonociceptive thresholds of the injured paw of sciatic nerve operated mice in comparison with the vehicle on Day 3 and on Day 7, respectively, are shown on FIGS. 4 A, B, C and D.

(37) The mechanical hyperalgesia values were determined as % by comparing the nociceptive thresholds measured on day 3 and 7 after the operation to the mean initial mechanonociceptive threshold values. Column pairs represent hyperalgesia before drug injection and 15 min after compound/solvent administration. Results are expressed as meanss.e.m. of the thermo- and mechanonociceptive threshold changes of n=6-6 in the vehicle/compound-treated TRPA1.sup.+/+ as well as TRPA1.sup./ mice groups. Data were analysed with One-way ANOVA following Bonferroni's post-test in comparison to determine differences between the pre- and post-injection values of the respective groups. *p<005

(38) The effects of SZV-1911 (A-B) and SZV-1287 (C-D) on the paw withdrawal latency of the injured paw of sciatic nerve operated mice in comparison with the vehicle on Day 3 and on Day 7, respectively, are shown on FIGS. 5 A, B, C and D.

(39) The paw withdrawal latency values were determined as % by comparing the latencies measured on day 3 and 7 after the operation to the mean initial latency values. Column pairs represent the change of paw withdrawal latency before drug injection and 15 min after compound/solvent administration. Results are expressed as meanss.e.m of the withdrawal behaviour latency changes of n=6-6 mice per groups. Data were analysed with One-way ANOVA following Bonferroni's post-test in comparison to determine differences between the pre- and post-injection values of the respective groups.*p<0.05.

(40) II. Neurogenic Inflammation Models

(41) Resiniferatoxin-Evoked Thermal and Mechanical Hyperalgesia

(42) Resiniferatoxin (RTX) is a potent agonist of Transient Receptor Potential Vanilloid 1 (TRPV1) receptors, which are mainly expressed on capsicin-sensitive sensory nerve ending. Intraplantarly (i.pl.) injection of RTX (0.03 g/ml; 20 l) into one of the hindpaws induced an acute inflammation and a robust drop of heat and mechanical thresholds. After applying single dose of pretreatment 10 minutes before testing-, thermonociceptive measuring were repeated at 5, 10, 15, 20 minutes, while mechanonociceptive testing was observed at 2, 4, 6, 24 hours.

(43) FormalinInduced Acute Somatic Nocifensive Behaviour

(44) Formalin (Formaldehydum solutum 37%; Ph.Hg. VII.; 20 l, 2.5%, i.pl.) injected into the right hindpaw known as a selective agonist of Transient Receptor Potential Ankyrin 1 (TRPA1) induces nocifensive reactions in two phases, the firea of which (0-5 min) is thought to be due to a direct chemonociceptive effect of formalin, while the second one (20-45 min) is mainly mediated by inflammatory reactions (Tjolsen et al., 1992). After applying single dose of pretreatment 10 minutes before testing-, thermonociceptive measuring was performed at 1 hour while mechanonociceptive testing was observed at 2, 4 hours.

(45) Determination of the Noxious Heat Thresholds

(46) The noxious heat threshold of the paw, defined as the lowest temperature evoking nocifensive behaviourwas measured with an increasing-temperature hot plate (IITC Life Science, Woodland Hills, Calif., USA). After habituation, mice were placed onto the plate, which was then heated up from room temperature at a rate of 12 C./min until the animals showing nocifensive behaviour (licking, lifting or shaking one of the hindpaws). The corresponding plate temperature was considered as the noxious heat threshold.

(47) The test protocol is shown schematically on FIGS. 6 A and B.

(48) Effects on RTX-Induced Hyperalgesia

(49) The average control heat threshold value was 43.730.5 C. in male C57Bl/6 animals. The intraplantarly administration of RTX caused firstly, after few minutes a robust drop of heat threshold, approx. 8 degree in solvent pretreated animal group. This decrease was significantly lower (1-2 C.) in mice injected intraperitoneally pretreatment with 20 mg/kg SZV-1287, and maintained during 15 minutes. Two hours after the application the mechanical hyperalgesia was developed. The change of the mechanonociceptive threshold in solvent treated group was 47.16% at 2 hours and 35.33% at 4 hours. These changes were significantly reduced in SZV-1911 injected group, 14.76% and 6.56% respectively at 2 and 4 hours. This drop of mechanonociceptive threshold was 22.17% in SZV-1287 treated group at 2 hours.

(50) The effects of SZV-1911 and SZV-1287 on RTX-induced thermal (A) and mechanical (B) hyperalgesia in mice in comparison with the vehicle are shown on FIGS. 7 A and B.

(51) Lines represent thermal and mechanical hyperalgesia before drug injection and 5, 10, 15, 20 min as well as 2, 4, 6, 24 hours, respectively after compound/solvent administration. Results are expressed as meanss.e.m. of the thermo- and mechanonociceptive threshold changes of n=7-8 per groups. Data were analysed with Two-way ANOVA following Bonferroni's post-test in comparison to determine differences between the pre- and post-injection values of the respective groups **p<0.01, *** p<0.001 SZV-1287 vs. saline and ### p<0.001 SZV-1911 vs. saline.

(52) Effects on Formalin-Induced Acute Nocifensive Behaviours and Hyperalgesia

(53) Nocifensive behaviour expressed as the total duration of paw lickings and liftings was significantly lower in mice treated with SZV-1287 both in the early phase (0-5 min) referring to acute chemonociception and late phase (20-45 min) evoked by the inflammatory reaction.

(54) The basal thermonociceptive thresholds were 44.81.7 and 44.90.7 C. which were significantly reduced by formalin application to 37.54 and 37.43 C. in SZV-1287 treated as well as in saline treated mice, respectively.

(55) The effects of SZV-1287 on formaline-induced acute nocifensive reactions (A) and the drop of thermo-(B) and mechanonociceptive (C) thresholds of mice are shown on FIGS. 8 A, B and C.

(56) The total duration of paw licking and lifting were expressed to representing the nociceptive behaviour. Column pairs represent thermal hyperalgesia before drug injection and 1 hour after compound/solvent administration. Mechanical hyperalgesia values were determined as % by comparing the nociceptive thresholds measured at 2 and 4 hours after the application to the initial threshold values. Results are expressed as meanss.e.m. of the thermo- and mechanonociceptive threshold changes of n=11-11 animals/groups. Data were analysed with One-way ANOVA following Bonferroni's post-test in comparison to determine differences between the pre- and post-injection values of the respective groups.