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A Compound Of Formula (I) And A Composition Used As Allosteric Potentiators Of The M4 Muscarinic Receptor

Abstract: The present invention relates to selective allosteric potentiators of the Formula (I): or pharmaceutically acceptable salts thereof, for the treatment of disorders associated with M4 muscarinic receptors.

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Patent Information

Application #
Filing Date
25 May 2007
Publication Number
32/2007
Publication Type
INA
Invention Field
PHARMACEUTICALS
Status
Email
Parent Application
Patent Number
Legal Status
Grant Date
2011-01-28
Renewal Date

Applicants

ELI LILLY AND COMPANY
LILLY CORPORATE CENTER, CITY OF INDIANAPOLIS, INDIANA

Inventors

1. ALMUDENA RUBIO ESTEBAN
14469 TWIN OAKS DRIVE, CARMEL, INDIANA 46032
2. DARRYL WAYNE HILLIARD
11117 HARRISTON DRIVE, FISHERS, INDIANA 46038

Specification

FIELD OF THE INVENTION The present invention provides compounds of Formula (I), and a composition thereof, used as allosteric potentiation of the M4 subtype of muscarinic receptor. BACKGROUND OF THE INVENTION The present invention provides compounds that are selective allosteric modulators ot the M subtype of muscarinic receptor. The M4 muscarinic receptor is believed to play a role in modulating synaptic function in key areas of the brain involved in regulating mood, cravings, attention and cognition. As a result, it provides a novel therapculic target tor the treatment oi' psychosis; attention disorders, such as attention deficit hyperactivity disorder (ADHD): cognitive disorders, including memory loss; and drug addiction. The M2 and M4 subtypes of muscarinic receptor are also involved in muscarinic agonist- induced analgesic effects, but it is believed that side effects of such treatment are associated primarily with M2 receptor activation. Thus, compounds that selectively modulate M4 receptors would provide a novel treatment strategy for neuropathic pain, without unwanted side e ffeet s. Unlike compounds that act at the neurotransmitter binding site (orthosteric site), allosterie modulators act at a distinct site on the receptor. The use of allosteric modulators provides several advantages in the treatment of disease. Christoponlos Nature Reviews (2002) 1:19X-210. For instance, under saturating conditions (high concentrations of allosteric potentiator) one would not expect excessive stimulation of the M4 muscarinic receptor, since it is dependent on the endogenous neurotransmitter for activation. Second, allosteric agonists exert their physiological effects only in the presence of endogenous agonist. As a result, allosteric potentiators are less likely to produce the condition of receptor desensitzation or down-regulation that are associated with excessive cholinergic stimulation, finally, allosteric modulators are likely to show greater receptor selectivity, especially as the orthosteric binding site is well conserved between muscarinic receptor subtypes. At present, no selective allosteric modulators of the M4 subtype of muscarinic receptor have been reported. The development of selective M4 allosteric potentiators will therefore greatly enhance the ability to treat disorders such as psychosis and pain, without unwanted side effects. Thas, the present invention provides a class of allosteric. modulators of M4 muscarinic receptors, compositions comprising these compounds, and methods of using the compounds. BRIEF SUMMARY OF THE INVENTION The invention provides compounds of formula (I): n is 0. I, or 2: X is a bond. -O-. -SCp-C(O)-, -NR'X -C(0)-KR2-. or -NR2-C(0>: p is 0. 1 , or 2: R is hydrogen, hydroxyl. CVO alkyl. phenyl, pyridyl, pyrrolidinyi. pipcra/dnyl. morphol no. thiazolyF imida/'oivl, or I ..i-dinxalaiiyl: ^vhieh phenyl, pipera/.inyl, or tnla/ulyl group may be optionally substituted with one substiluenl selected from the group consisting of halo or d-C' alkyl; wherein n cannot be 0 when p is 0. or when X is -<)-, -KR\ or -r\R2-ClO)-: R* is hydrogen or C|-C> alkyl: which C'i-C: alkyl may be optionally substituted with one hydroxyl; or a pharmaceutically acceptable salt thereof. The compounds of Formula I are muscarinic receptor potentiators. Specifically, the compounds of formula 1 are allosleric potentiators of the M4 subtype of muscarinic receptor. Because these compounds potentiate the physiological effects associated with M.( receptor activation, the compounds arc useful in the treatment of disorders related 10 inadequate Mj receptor activation, fhese disorders include: pyschosis (particularly, schizophrenia): cognitive disorders (for example, memory loss); attention disorders (such as attention deficit hyperactivity disorder); and pain (in particular, neuropathic pain). In one embodiment, this invention provides a pharmaceutical composition comprising, as an active ingrethent, a compound of Formula 1. or a pharmaceutically acceptable salt thereof, in combination with one or more pharmaceutically acceptable carriers, diluents, or excipients. hi 60g, 7.0mmol) in THF (3ml) at room temperature is added dropwiss a solution of lithium bis(trimethylsilyl)amide (1.0M in hexanes)(3.50ml, 3.50mmol). The reaction mixture is stirred for 15 minutes and then is treated with 3-amino-5-chloro-6-rnethanesulfmyl-4-methyI-thieno[2,3-b]pyridine-2- carboxylic acid cyclopropylamide (0.345g, l.OOmmol). The reaction mixture is heated at 75°C in a sealed tube for 1.5 hours, cooled to room temperature, and then quenched by the addition of water (10ml). The reaction mixture is transferred to an erlenmeyer flask and diluted to 75ml with water. The mixture is cooled to 0°C and stirred. During this time, a yellow precipitate is formed. This solid is collected by filtration and slurried in a 2:1 EtOAc:hexane mixture. This slurry is stirred for 10 minutes, filtered, and the collected solid is placed in the vacuum oven to dry (60°C for 16 hours). This affords the title compound as a yellow solid (0.242g, 58%). Mass (m/z): 417.2 (M++l), 415.2 (IvT-1). Example 20 3-Amino-5-chloro-4-methyl-6-(2-pyridin-4-yl-ethoxy)-thieno[2,3-b]pyridine-2- carboxylic acid cyclopropylamide To a solution of 4-pyridineethanol (0.862g, 7.0mmol) in THF (3ml) at room temperature is added dropwise a solution of lithium bis(trimethylsilyl)amide (1.0M in hexanes)(3.50ml, 3.50mmol). The reaction mixture is stirred for 15 minutes and then is treated with 3-amino-5-chloro-6-methanesulfinyl-4-methyl-thieno[2,3-b]pyridine-2- carboxylic acid cyclopropylamide (0.345g, l.OOmmol). The reaction mixture is heated at 75°C in a sealed tube for 1,5 hours, cooled to room temperature, and then quenched by the addition of water (10ml). The reaction mixture is transferred to an erlenmeyer flask and diluted to 75ml with water. The mixture is cooled to 0°C and stirred. During this time, a yellow precipitate is formed. This solid is collected by filtration and slurried in a 2:1 EtOAc:hexane mixture. This slurry is stirred for 10 minutes, filtered, and the collected solid is placed in the vacuum oven to dry (60°C for 16 hours). This affords the title compound as a yellow solid (0.23 lg, 57%). Mass (m/'z): 403.1 (M++l), 401.1 (M+-1). Example 21 3-Amino-6-{2-[bis-(2-hydroxyethyl)-amino]-ethoxy}-5-chloro-4-methyl-thieno[2,3- b]pyridine-2-carboxylic acid cyclopropylamide To a solution of triethanolamine (1.044g, 7.0mmol) in THF (3ml) at room temperature is added dropwisea solution of lithium bis(trimethylsilyl)amide (1.0M in hexanes)(3.50ml, 3.50mmol). The reaction mixture is stirred for 15 minutes and then is treated with 3-amino-5-chloro-6-methanesulfinyl-4-mcthyl-thieno[2,3-b]pyridinc-2- carboxylic acid cyclopropylamide (0.345g, l.OOmmol). The reaction mixture is heated at 75CC in a sealed tube for 1.5 hours, cooled to room temperature, and then quenched by the addition of water (10ml). The reaction mixture is transferred to an erlenmeyer flask and diluted to 75ml with water. The mixture is cooled to 0°C and stirred. During this time, a yellow precipitate is formed. This solid is collected by filtration and slurried in a 2:1 EtOAc: hexane mixture. This slurry is stirred for 10 minutes, filtered, and the collected solid is placed in the vacuum oven to dry (60°C for 16 hours). This affords the title compound as a yellow solid (0.179g, 41%). Mass (rn/z): 429.2 (M++l), 427.2 (M+-l). Example 22 3-Amino-5-chloro-6-([l,3]clioxolari-4-ylmethoxy)-4rmethyl-thieno[2,3-b]pyridine-2- carboxylic acid cyclopropylamide To a solution of glycerol formal (0.729g, 7.0mmol) in THF (3ml) at room temperature is added dropwise a solution of lithium bis(trimethylsilyl)amide (1.0M in hexanes)(3.50ml, 3.50mmol). The reaction mixture is stirred for 15 minutes and then is treated with 3-amino-5-chloro-6-methanesulfinyl-4-methyl-thieno[2,3-b]pyridine-2- carboxylic acid cyclopropylamide (0.345g, l.OOmmol). The reaction mixture is heated at 75°C in a sealed tube for 1.5 hours, cooled to room temperature, and then quenched by the addition of water (10ml). The reaction mixture is transferred to an erlenmeyer flask and diluted to 75ml with water. The mixture is cooled to 0°C and stirred. During this time, a white precipitate is formed. This solid is collected by filtration and slurried in a 2:1 EtOAc: hexane mixture. This slurry is stirred for 10 minutes, filtered, and the collected solid is placed in the vacuum oven to dry (60°C for 16 hours). This affords the title compound as a white solid (0.195g, 51%). Mass (m/z): 384.1 (M++l), 382.1 (M+-l). Example 23 3-Amino-5-chloro-4-methyl-6-[(2-pyrrolidin-l-yl-ethylcarbamoyl)-methoxy]-thieno[2,3- b]pyridine-2-carboxylic acid cyclopropylamide To a solution of (3-amino-5-chtoro-2-cyclopropylcarbamoyl-4-methyl-thieno[2,3- b]pyridin-6-yloxy)-acetic acid (0.300g, O.843mmol) in a 1:1 mixture of THF:DMF (2.0ml each) are adejed 1-hydroxybenzotriazole hydrate (0.148g, 1.096mmol), N,N- diisopropylethylamine (0.163g, 1.265mmol), l-[3-(dimethylamino)propyi]-3- ethylcarbodiimide hydrochloride (0.242g, 1.265mmol), and l-(2-aminoethyl)pyrrolidine (0.289g, 2.529mmol). The resulting solution is stirred at room temperature for 16 hours. The reaction is quenched by the addition of water (25ml). A yellow precipitate formed. The mixture is cooled to 0°C and stirred for 10 minutes, then filtered. The collected solid is dried in the vacuum oven for 3 hours at 60CC, This affords the title compound as a yellow solid (0.069g, 18%). Mass (m/z): 452.2 (Mf+1), 450.2 (Ivf-1). Example 24 3-Amino-5-chloro-6-[(4-fluoro-benzylcarbamoyl)-methoxy]-4-methyl-r.hieno[2,3- b]pyridine-2-earboxylic acid cyclopropylamide To a solution of (3-amino-5-ehloro-2-cyclopropylcarbamoyl-4-methyl-thieno[2,3- b]pyridin-6-yloxy)-acetic acid (0,300g, 0.843mmol) in a 1:1 mixture of THF:DMF (2.0ml each) are added 1 -hydroxybenzotriazole hydrate (0.148g, 1.096mmol), N,N- diisopropylethylamine (0.163g, 1.265mmol), l-[3-(dimethylamino)propyl]-3- ethylcarbodiimide hydrochloride (0.242g, 1.265mmol), and 4-fluorobenzylamine (0.317g, 2.529mmol). The resulting solution is stirred at room temperature for 16 hours. The reaction is quenched by the addition of water (25ml). A yellow precipitate is formed. The mixture is cooled to 0°C and stirred for 10 minutes, then filtered. The collected solid is slurried in 10: 1 hexane:EtOAc and stirred for 10 minutes. The solid is collected by filtration and dried in the vacuum oven at 50°C for 1 hour. This affords the title compound as a pale yellow solid (0.l43g, 37%). Mass (m/z): 463.1 (M++l), 461.1 (M+- 1). Example 25 3-Amino-5-chloro-4-methyl-6-[(2-morpholin-4-yl-ethylcarbamoyl)-methoxy]-thieno[2,3- b]pyridine-2-carboxylic acid cyclopropylamide To a solution of (3-amino-5-chloro-2-cyclopropylcarbamoyl-4-methy!-thieno[2,3- b]pyridin-6-yloxy)-acetic acid (OJOOg, 0.843mmol) in a 1:1 mixture of THF:DMF (2.0ml each) are added 1-hydroxybenzotriazole hydrate (0.148g, 1.096mmol), N,N- diisopropylethylamine (0.163g, 1.265mmol), l-[3-(dimethylamino)propyl]-3- ethylcarbodiimide hydrochloride (0.242g, 1.265mmol), and 4-(2-aminoethyl)morpholine (0.329g, 2.529mmol). The resulting solution is stirred at room temperature for 16 hours. The reaction is quenched by the addition of water (25ml), If no precipitate forms, the mixture may be acidified to pH2 with IN HC1 and extracted with ElOAc (25ml). The organic layer is discarded and the aqueous layer is made basic (pH12) with 5N NaOH and extracted with EtOAc(40ml). The organic layer is dried (anhydrous magnesium sulfate), filtered, and concentrated to give a yellow solid. The solid is slurried in 3:1 hexane:EtOAc (10ml), then filtered. The collected solid is dried in the vacuum oven at 50°C for 72 hours. This affords the title compound as a yellow solid (0.053g, 13%). Mass (m/z): 468.2 (M++l), 466.2 (MM), Example 26 3-amino-5-chloro-4-methyl-6-{2-[(pyridine-4-earbonyl)-amino]-ethoxy}-thieno[2,3- b]pyridine-2-carboxylic acid cyclopropylamide To a solution of N-(2-hydroxyethyl)-isonicotinamide (0,83 lg, 5.00mmol) in THF (2.5ml) at room temperature is added dropwise a solution of lithium bis(trimethylsilyl) amide (1.0M in hexanes)(2.50ml, 2.50mmol). The reaction mixture is stirred for 15 minutes and then is treated with 3-amino-5-chloro-6-inethanesulfinyl-4-methyl- thieno[2,3-b]pyridine-2-carboxylic acid cyclopropylamide (0.345g, l.OOmmol). The reaction mixture is heated at 80°C in a sealed tube for 2 hours, cooled to room temperature, and then quenched by the addition of water (15ml). The mixture is diluted to 100ml with water and then cooled to 0°C and stirred. A yellow precipitate is formed. This solid is collected by filtration, slurried in EtOAc, and filtered again to give a yellow solid which is placed in the vacuum oven to dry (50CC for 3 hours). This affords the title compound as a yellow solid (0.046g, 10%). Mass (m/z): 446.3 (M++l), 444.2 (M+-l). Example 27 3-Amino-6-benzyloxy-5-chloro-4-methyl-thieno[2,3-b]pyridine-2-carboxylic acid cycloprcpylamide To a solution of benzyl alcohol (0.541g, 5.00mraol) in THF (2.5ml) at room temperature is added dropwise a solution of lithium bis(trimethylsilyl')amide (1.0M ;n hexanes)(2.50ml, 2.50mmol), The reaction mixture is stirred for 15 minutes and then is treated with 3-amino-5-chloro-6-methanesulfinyl-4-methyl-thieno[2,3-b]pyridine-2- carboxylic acid cyclopropylamide (0.345g, l.OOmmol). The reaction mixture is heated at 80°C in a sealed tube for 2 hours, cooled to room temperature, and then quenched by the addition of water (15ml). The mixture is diluted to 100ml with water and then cooled to 0CC and stirred. A yellow precipitate is formed. This solid is collected by filtration and purified by flash chromatography (1.5: 1 hexane: EtOAc) to give the title compound as a pale yellow solid (0.130g, 33%). Mass (m/z): 388.2 (M+-l), 386.2 (M+-l). Example 28 3-Amino-5-chloro-4-methyl-6-(2-morpholin-4-yl-ethoxy)-thieno[2,3-bjpyridine-2- carboxylic acid cyclopropylamide hydrochloride A mixture of 3-amino-5-chloro-4-methyl-6-(2-morpholin-4-yl-ethoxy)-thieno[2,3- b]pyridine-2-carboxylic acid cyclopropylamide (0.800g, 1,95mmol) and MeOH (50ml) is heated to 50°C. The mixture is treated with THF (5ml) and DMF (5ml). While still at 50°C, the mixture is acidified to pHl with concentrated HC1. A homogeneous solution is formed. The solution is allowed to cool slowly to room temperature. During this time, a white precipitate is formed. This solid is collected by Filtration and dried in the vacuum oven for 72 hours at 50°C. This affords the title compound as a white solid (0.562g, 65%). Mass (m/z): 411.1 (M++1)-HC1, 445.1 (M+-1). Example 29 3-Amino-5-chloro-6-ethoxy-4-methyl-thieno[2,3-b]pyridine-2-carboxylic acid cyclopropylamide Sodium hydride (0.037g, 0.925mmol) is suspended in anhydrous 1,2-dimethoxy- ethane (5.0ml) under nitrogen. The suspension is cooled to 0°C. To this cold suspension is added via cannula a solution of 3-amino-5-chloro-6-hydroxy-4-methyl-thieno[2,3- b]pyridine-2-carboxylic acid cyclopropylamide (0.300g, l.Olmmol) in anhydrous DMF (5.0ml). The ice bath is removed, and the reaction is warmed to room temperature. After the reaction is stirred 25 minutes at room temperature, lithium bromide (0.175g, 2.02mmol) is added as a solid, and the reaction is stirred for an additional two hours. Finally, ethyl iodide is added (0.24ml, 0.47g, 3.00mmol), and the reaction is stirred for 36 hours at room temperature. At the completion of the reaction time, the mixture is quenched with water. The solid is collected by filtration, then purified by flash chromatography (hexanes:EtOAc grathent) to give the title compound as a white solid (24.3mg, 8% yield). Mass (m/z): 326.0 (M1"), 324.0 (M-). Alternatively, Example 33 may be prepared following the procedures outlined for the preparation of Example 29, substituting EtOH for benzyl alcohol. Example 30 (3-Amino-5-chloro-2-cyclopropylcarbamoyl-4-methyl-thieno[2,3-b]pyridin-6-yloxy)- acetic acid (imidazol-l-yl)-amide To a solution of (3-amino-5-chloro-2-cyclopropylcarbamoyl-4-methyl-thieno[2,3- b]pyridin-6-yloxy)-acetic acid (0.350g, 0.98mmol) in DMF (4ml) at room temperature is added 1,1 '-carbonyldiimidazole (0.239g, 1.48mmol). The resulting solution is heated and stirred at 40°C for 20 minutes. During this time, a cream colored precipitate is formed. The mixture is cooled to 10°C, and neat 2-aminopyridine (0.231g, 2.46mmol) is added. The mixture is stirred at room temperature for 30 minutes, then quenched by adding saturated sodium bicarbonate solution (10ml). A thick, white precipitate is formed. The mixture is diluted with water (25ml), stirred for 10 minutes, then filtered. The collected solid is dried in the vacuum oven for 16 hours. This affords the title compound as a white solid (0.228g, 58%). Mass (m/z): 404.0 (MM). The compounds of the present invention can be administered alone or in the form of a pharmaceutical composition, that is, combined with pharmaceutically acceptable carriers, or excipients, the proportion and nature of which are determined by the solubility and chemical properties of the compound selected, the chosen route of administration, and standard pharmaceutical practice. The compounds of the present invention, while effective themselves, may be formulated and administered in the form of their pharmaceutically acceptable salts, for purposes of stability, convenience of crystallization, increased solubility, and the like. Thus, the present invention provides pharmaceutical compositions comprising a compound of the Formula 1 and a pharmaceutically acceptable diluent. The compounds of Formula I can be administered by a variety of routes. In effecting treatment of a patient afflicted with disorders described herein, a compound of Formula I can be administered in any form or mode that makes the compound bioavailable in an effective amount, including oral and parenteral routes. For example, compounds of Formula I can be administered orally, by inhalation, or by the subcutaneous, intramuscular, intravenous, transdermal, intranasal, rectal, occular, topical, sublingual, buccal, or other routes. Oral administration is generally preferred for treatment of the neurological and psychiatric disorders described herein. One skilled in the art of preparing formulations can readily select the proper form and mode of administration depending upon the particular characteristics of the compound selected, the disorder or condition to be treated, the stage of the disorder or condition, and other relevant circumstances. (Remington's Pharmaceutical Sciences, 18th Edition, Mack Publishing Co. (1990)). The pharmaceutical compositions are prepared in a manner well known in the pharmaceutical art. The carrier or excipient may be a solid, semi-solid, or liquid material that can serve as a vehicle or medium for the active ingrethent. Suitable carriers or excipients are well known in the art. The pharmaceutical composition may be adapted for oral, inhalation, parenteral, or topical use and may be administered to the patient in the form of tablets, capsules, aerosols, inhalants, suppositories, solutions, suspensions, or the like. The compounds of the present invention may be administered orally, for example, with an inert diluent or capsules or compressed into tablets. For the purpose of oral therapeutic administration, the compounds may be incorporated with excipients and used in the form of tablets, troches, capsules, elixirs, suspensions, syrups, wafers, chewing gums and the like. These preparations should contain at least 4% of the compound of the present invention, the active ingrethent, but may be varied depending upon the particular form and may conveniently be between 4% to about 70% of the weight of the unit. The amount of the compound present in compositions is such that a suitable dosage will be obtained. Preferred compositions and preparations according to the present invention may be determined by a person skilled in the art. The tablets, pills, capsules, troches, and the like may also contain one or more of the following adjuvants: binders such as povidone, hydroxypropyl cellulose, microcrystalline cellulose, gum tragacanth or gelatin; excipients such as dicalcium phosphate, starch, or lactose; disintegrating agents such as alginic acid, Primogel, corn starch and the like; lubricants such as talc, magnesium stearate or Sterotex; glidants such as colloidal silicon dioxide; and sweetening agents, such as sucrose, aspartame, or saccharin, or a flavoring agent, such as peppermint, methyl salicylate or orange flavoring, may be added. When the dosage unit form is a capsule, it may contain, in addition to materials of the above type, a liquid carrier such as polyethylene glycol or a fatty oil. Other dosage unit forms may contain other various materials that modify the physical form of the dosage unit, for example, coalings. Thus, tablets or pills may be coated with sugar, shellac, or other coating agents. A syrup may contain, in addition to the present compounds, sucrose as a sweetening agent and certain preservatives, dyes and colorings and flavors, Materials used in preparing these various compositions should be pharmaceutically pure and non-toxic in the amounts used. For the purpose of parenteral therapeutic administration, the compounds of the present invention may be incorporated into a solution or suspension. These preparations typically contain at least 0.001% of a compound of the invention, but may be varied to be between 0.001 and about 90% of the weight thereof. The amount of the compound of Formula I present in such compositions is such that a suitable dosage will be obtained. The solutions or suspensions may also include one or more of the following adjuvants: sterile diluents, such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents; antibacterial agents, such as benzyl alcohol or methyl paraben; antioxidants, such as ascorbic acid or sodium bisulfite; chelating agents, such as ethylene diaminetetraacetic acid, buffers, such as acetates, citrates or phosphates; and agents for the adjustment of tonicity, such as sodium chloride or dextrose. The parenteral preparation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic. Preferred compositions and preparations are able to be determined by one skilled in the art. The compounds of the present invention may also be administered topicaily, and when done so, the carrier may suitably comprise a solution, ointment, or gel base. The base, for example, may comprise one or more of the following: petrolatum, lanolin, polyethylene glycols, bees wax, mineral oil, diluents such as water and alcohol, and emulsifiers, and stabilizers. Topical formulations may contain a concentration of a compound of Formula I or its pharmaceutical salt from about 0.1 to about 10% w/v (weight per unit volume). The compounds of Formula 1 are allosteric potentiators of the M4 subtype of muscarinic receptors. Furthermore, the compounds of Formula 1 selectively potentiate M4 receptors relative to other muscarinic receptors. The activity of the compounds of the present invention may be determined by the methods below. Calcium mobilization in whole cells stably expressing human muscarinic receptors A. Stable Cell Lines Standard molecular cloning techniques may be used to generate stable cell lines expressing the human muscarinic M1-M5 receptors. M,, M3 an M5 receptors arc expressed in Chinese hamster ovary (CHO) cell lines, whereas, M2 an M4 are expressed in AVI2 Gai5 cell lines. The cDNA encoding these muscarinic receptors correspond to the published sequence in the NCBI nucleotide database of accession numbers: AF498915, AF498916, AF498917, AF498918 and AF498919, forM,-M5 respectively. B. Methods Using a calcium-sensitive fluorescent dye, agonist or potentiation activity of a given compound can be detected in a single assay using a Fluorescence Light Imaging Plate Reader (FLIPR) instrument. Cells are plated in Poly-D-Lysine coated black plates/clear bottom (Becton Dickinsons) at 40,000 cells per ml (lOOul/well) in growth media 24 hours prior to running the assay. The medium is removed before the addition of 50u,l fluorescence dye solution (HBSS containing 20mM Hepes, 10u.M Fluo-3-AM, 0.05% pluronic acid F127; supplemented with 2.5mM probenecid for CHO cell assays). The cells are incubated with the dye for 75 minutes before replacing with assay buffer (20mM Hepes in Hanks balanced salt solution (Gibco); supplemented with 2.5mM probenecid for CHO cells). The plate is transferred to the FLIPR machine (Molecular Devices) for fluorescence recordings. The cells are periodically excited by 488nm light, and the emitted fluorescent light passed through a 510-570nm filter and then detected by a cooled CCD camera. Automated multiple compound additions are timed by computer program. Cells are pre-incubated with increasing concentration of compounds. After 2 minutes, a range of acetylcholine concentrations is added to each concentration of compound. If the compound is an allosteric enhancer, a compound concentration- dependent potentiation of acetylcholine response would be detectable. The effectiveness of potentiator compounds can be ranked by their affinity and cooperativity. An alternative method may be used to provide an estimate of the affinity of compounds of the present invention, and to rank order the compounds based on this estimated affinity. In this method, a single concentration of carbachol that is approximately 10% of a saturating concentration is added to all wells, and increasing concentrations of the test compounds of the present invention are added. An estimated affinity value is derived by calculation of the EC50 for potentiation of the 10% carbachol response. This method may be used to rank order the compounds described herein as Examples. As confirmed with the compound of Example 1, acetylcholine and carbachol (both non-selective full muscarinic receptor agonists) are potentiated in an equivalent fashion in the presence of an allosteric modulator, C. Data Analysis and Results Allosteric parameters may be estimated using the equations by Lazareno et al., Mo!. Pharmacol. (1995) 48: 362-378. The effect of increasing concentrations of the compound of Example 1 on cellular ACh concentration-response curves in recombinant cell lines (AV12 Ga15 hM2 or hM4 and CHO hMi, I1M3 or hMs) may be tested using FLIPR. Data are collected in duplicates from at least three independent experiments. No significant allosteric effect is observed in CHO cells stably expressing hMi, hM3 or hM5 receptors. The cooperativity factor and affinity of the compound of Example 1 for hM4 receptors is estimated to be 34.5 ± 3.5 and 200 ± 42 nM, respectively. A modest allosteric effect on hM2 receptors is also observed but is too modest to be accurately estimated. Using the FLIPR assay to potentiate 10% carbachol, as described above for rank ordering of the compounds of the present invention, each of the compounds described herein as EXAMPLES has an estimated affinity for hM^ receptors of < 500 11M. Neurotransmitter release assay A. Methods Two male Lister Hooded rats are killed by carbon dioxide asphyxiation and cervical dislocation. The brains are rapidly removed, and the striatum dissected out and cross-chopped three times at 150pm. The slices are suspended in 12ml of HEPES buffer (128mM NaCl, 2.4mM KCI, 3.2mM CaCl2, 1.2mM KH2PO4, 1.2 mM MgSO47H20, 25mM HEPES, lOmM Glucose, pH 7.5). The slices are washed twice, with resuspending in fresh buffer each time, then incubated at 37°C for 30 minutes with [3H]-choline chloride (250nM). After 30 minutes, a further four washes are carried out, and 100|il of slices are placed in each well of a 96 well filter plate (Millipore MABCN 96-well multiscreen plate). The bathing solution is removed by vacuum filtration (Millipore Univac manifold system), then a further 70p.l of HEPES buffer (+/- compound) is added to each well, and the plate is returned to the incubator for 5 minutes. After a 5-minute incubation, the buffer is removed by vacuum filtration into a collecting plate (Wallac 96 well flexible sample plates). Stimulating solution (70nl/well: 20mM potassium 1-/- compound) is then added, and the plate returned to the incubator for a further 5-minutes. The stimulating buffer is then removed by vacuum filtration into a second collecting plate. At the end of the experiment, the amount of tissue in each well is estimated by placing the plates in a freezer for one hour, punching out the filter discs, and adding Soluene® (to digest the slices) and leaving for a further hour. The radioactivity in the digested tissue is measured using liquid scintillation counting. Neurotransmitter release is calculated as a fraction of total radioactivity present in the well. B. Results The potentiation effect of compounds in native tissues is tested by its ability to potentiate auto-inhibition of acetylcholine release in striatal slices, as induced by 20mM potassium stimulation. This is considered to be an M4-mediated process via pre-synaptic auto-regulation in the striatum. Zhang,W. et al, J.Neurosci. (2002)22: 1709-1717. Using the methods above, a representative compound of the present invention (Example 1) exhibits concentration-dependent potentiation of auto-inhibition with an 1C5[) of 1.5 uM. Several preclinical laboratory animal models have been described for a number of the disorders associated with muscarinic receptors. For instance, inhibition of the conditioned avoidance response (CAR) by neuroleptic and atypical antipsychotics is one of the most stuthed pharmacological models of psychosis. To date, all clinically effective antipsychotics have been demonstrated to selectively suppress CAR (cf. Wadenberg & Hicks, 1999. Neuroscience & Biobehavioral Reviews, 23: 851-8). Conditioned Avoidance Response A. Methods Male Fisher-344 rats (N = 5-8) are trained in an avoidance paradigm in which the rat must make a shuttle response to avoid or escape a footshock. The apparatus is a Coulbourn Instruments shuttle operant chamber. Each session equals a total of 50 trials, presented on a 30 second inter-trial interval, and each trial begins with simultaneous illumination of a houselight and rising of a guillotine door. The rat is given 10 seconds to cross over to the other side (an avoidance response) before a 1 mA footshock is initiated. The shock remains on until the rat crosses over to the other side (an escape response) or 10 seconds has elapsed (an escape failure). Rats are well trained on this task, with baseline avoidance performance > 95 %. The number of avoidance, escape and escape failure responses during each session is recorded and used for analysis. Groups of rats are dosed with (1) vehicle (10% acacia plus sterile water), (2) a sub-efficacious dose of the muscarinic agonist oxotremorine sesquifumarate (Oxo alone), or (3) Oxo in the presence of increasing doses of the test compound (10 mg/kg to 60 mg/kg), followed by (4) a retest of Oxo alone. Each test compound is administered orally 2 hours prior to testing. Oxo (0.03 mg/kg) is administered subcutaneously 30 minutes prior to testing. Data are analyzed via a one-way (within-group design) Analysis of Variance (ANOVA). In cases of a significant ANOVA (p < 0.05), post-hoc comparisons may be made in which compound doses are compared back to the Oxo alone group (paired t-test). The conditioned avoidance assay is highly predictive of antipsychotic efficacy in the clinic. Representative muscarinic M4 receptor potentiators exhibit an antipsychotic- like profile in the conditioned avoidance responding paradigm. Although these M4 potentiators are inactive when tested alone (data not shown), these compounds potentiate the efficacy of an inactive dose of the muscarinic agonist oxotremorine. The results of calcium mobilization and neurotransmitter release stuthes demonstrate the ability of compounds of the present invention to act as potentiators of M4 muscarinic receptors. It is recognized that the compounds of the present invention would be expected to potentiate the effects of M4 receptor activation. Thus, the compounds of the present invention are expected tc be useful in the treatment of various disorders associated with muscarinic receptors, as described to be treated herein, and other disorders that can be treated by such allosteric potentiators, as are appreciated by those skilled in the art. The disorders associated with M4 muscarinic receptors are treated by administering an effective amount of a compound or pharmaceutical composition of Formula I. An effective amount can be readily determined by the attending diagnostician, as one skilled in the art, by the use of conventional techniques and by observing results obtained under analogous circumstances. In determining an effective amount, the dose of a compound of Formula I, a number of factors are considered by the attending diagnostician, including, but not limited to: the compound of Formula I to be administered; the species of mammal - its size, age, and general health; the specific disorder involved; the degree of involvement or the severity of the disorder; the response of the individual patient; the mode of administration; the bioavailability characteristics of the preparation administered; the dose regimen selected; the use of other concomitant medication; and other relevant circumstances. An effective amount of a compound of Formula 1 is expected to vary from about 0.001 milligram per kilogram of body weight per day (mg/kg/day) to about 100 mg/kg/day. Preferred amounts may be readily determined by one skilled in the art. We Claim: 1. Novel thienopyridine modulators of the M4 Muscarinic Receptor of Formula I: wherein: m is 1 or 2 n is 0 or 1 X is a bond R1 is hydrogen or morpholino or a pharmaceutically acceptable salt thereof. 2. The compound as claimed in claim 1, wherein the compound is selected from i) 3-Amino 5-chloro 6 methoxy-4-methyl-thieno[2,3-b]pyridine-2-carboxylic acid cyclopropylamide or pharmaceutically acceptable salt thereof; ii) 3-Amino-5-chloro-4-methyl-6-(2-morpholin-4-yl-ethoxy)-thieno[2,3-b]pyridine-2- carboxylic acid cyclopropylamide or pharmaceutically acceptable salt thereof; 3. A pharmaceutical composition comprising the compound of claim 1 or a pharmaceutically salt thereof in combination with pharmaceutically acceptable carrier, excipients or diluents, 4. A compound and a pharmaceutical composition to treat the conditions associated with M4 muscarinic receptor activity, substantially as herein described and as illustrated in the foregoing examples. The present invention relates to selective allosteric potentiators of the Formula (I): or pharmaceutically acceptable salts thereof, for the treatment of disorders associated with M4 muscarinic receptors.

Documents

Application Documents

# Name Date
1 abstract-01871-kolnp-2007.jpg 2011-10-07
1 LP-1871-KOLNP-2007-01-02-2023-ALL DOCUMENTS.pdf 2023-02-01
2 1871-kolnp-2007-translated copy of priority document 1.1.pdf 2011-10-07
2 1871-KOLNP-2007_EXAMREPORT.pdf 2016-06-30
3 1871-kolnp-2007-reply to examination report.pdf 2011-10-07
3 1871-KOLNP-2007-(29-01-2014)-FORM-27.pdf 2014-01-29
4 1871-kolnp-2007-others.pdf 2011-10-07
4 1871-KOLNP-2007-(11-03-2013)-CORRESPONDENCE.pdf 2013-03-11
5 1871-KOLNP-2007-OTHERS DOCUMENTS 1.1.pdf 2011-10-07
5 1871-KOLNP-2007-(11-03-2013)-FORM-27.pdf 2013-03-11
6 1871-kolnp-2007-granted-specification.pdf 2011-10-07
6 1871-KOLNP-2007-(27-01-2012)-CORRESPONDENCE.pdf 2012-01-27
7 1871-kolnp-2007-granted-letter patent.pdf 2011-10-07
7 1871-KOLNP-2007-(27-01-2012)-FORM 27.pdf 2012-01-27
8 1871-kolnp-2007-granted-form 2.pdf 2011-10-07
8 1871-KOLNP-2007-(10-10-2011)-OTHER PATENT DOCUMENTS.pdf 2011-10-10
9 01871-kolnp-2007-abstract.pdf 2011-10-07
9 1871-kolnp-2007-granted-form 1.pdf 2011-10-07
10 01871-kolnp-2007-claims.pdf 2011-10-07
10 1871-kolnp-2007-granted-description (complete).pdf 2011-10-07
11 01871-kolnp-2007-correspondence others.pdf 2011-10-07
11 1871-kolnp-2007-granted-claims.pdf 2011-10-07
12 01871-kolnp-2007-description complete.pdf 2011-10-07
12 1871-kolnp-2007-granted-abstract.pdf 2011-10-07
13 01871-kolnp-2007-form 1.pdf 2011-10-07
13 1871-kolnp-2007-form 5 1.1.pdf 2011-10-07
14 01871-kolnp-2007-form 2.pdf 2011-10-07
14 1871-KOLNP-2007-FORM 3.1.pdf 2011-10-07
15 01871-kolnp-2007-form 3.pdf 2011-10-07
15 1871-KOLNP-2007-FORM 3-1.2.pdf 2011-10-07
16 01871-kolnp-2007-form 5.pdf 2011-10-07
16 1871-KOLNP-2007-FORM 3-1.1.pdf 2011-10-07
17 1871-kolnp-2007-form 3 1.1.pdf 2011-10-07
17 01871-kolnp-2007-gfa.pdf 2011-10-07
18 01871-kolnp-2007-international publication.pdf 2011-10-07
18 1871-kolnp-2007-form 26 1.1.pdf 2011-10-07
19 01871-kolnp-2007-priority document.pdf 2011-10-07
19 1871-KOLNP-2007-FORM 2.1.pdf 2011-10-07
20 1871-KOLNP-2007-ABSTRACT 1.1.pdf 2011-10-07
20 1871-kolnp-2007-form 18 1.1.pdf 2011-10-07
21 1871-kolnp-2007-assignment 1.1.pdf 2011-10-07
21 1871-KOLNP-2007-FORM 1.1.pdf 2011-10-07
22 1871-KOLNP-2007-ASSIGNMENT.pdf 2011-10-07
22 1871-KOLNP-2007-EXAMINATION REPORT REPLY RECIEVED.pdf 2011-10-07
23 1871-KOLNP-2007-CLAIMS 1.1.pdf 2011-10-07
23 1871-kolnp-2007-examination report 1.1.pdf 2011-10-07
24 1871-KOLNP-2007-DESCRIPTION (COMPLETE) 1.1.pdf 2011-10-07
24 1871-kolnp-2007-correspondence 1.1.pdf 2011-10-07
25 1871-KOLNP-2007-CORRESPONDENCE OTHERS 1.1.pdf 2011-10-07
26 1871-kolnp-2007-correspondence 1.1.pdf 2011-10-07
26 1871-KOLNP-2007-DESCRIPTION (COMPLETE) 1.1.pdf 2011-10-07
27 1871-KOLNP-2007-CLAIMS 1.1.pdf 2011-10-07
27 1871-kolnp-2007-examination report 1.1.pdf 2011-10-07
28 1871-KOLNP-2007-ASSIGNMENT.pdf 2011-10-07
28 1871-KOLNP-2007-EXAMINATION REPORT REPLY RECIEVED.pdf 2011-10-07
29 1871-kolnp-2007-assignment 1.1.pdf 2011-10-07
29 1871-KOLNP-2007-FORM 1.1.pdf 2011-10-07
30 1871-KOLNP-2007-ABSTRACT 1.1.pdf 2011-10-07
30 1871-kolnp-2007-form 18 1.1.pdf 2011-10-07
31 01871-kolnp-2007-priority document.pdf 2011-10-07
31 1871-KOLNP-2007-FORM 2.1.pdf 2011-10-07
32 01871-kolnp-2007-international publication.pdf 2011-10-07
32 1871-kolnp-2007-form 26 1.1.pdf 2011-10-07
33 01871-kolnp-2007-gfa.pdf 2011-10-07
33 1871-kolnp-2007-form 3 1.1.pdf 2011-10-07
34 01871-kolnp-2007-form 5.pdf 2011-10-07
34 1871-KOLNP-2007-FORM 3-1.1.pdf 2011-10-07
35 1871-KOLNP-2007-FORM 3-1.2.pdf 2011-10-07
35 01871-kolnp-2007-form 3.pdf 2011-10-07
36 1871-KOLNP-2007-FORM 3.1.pdf 2011-10-07
36 01871-kolnp-2007-form 2.pdf 2011-10-07
37 01871-kolnp-2007-form 1.pdf 2011-10-07
37 1871-kolnp-2007-form 5 1.1.pdf 2011-10-07
38 01871-kolnp-2007-description complete.pdf 2011-10-07
38 1871-kolnp-2007-granted-abstract.pdf 2011-10-07
39 01871-kolnp-2007-correspondence others.pdf 2011-10-07
39 1871-kolnp-2007-granted-claims.pdf 2011-10-07
40 01871-kolnp-2007-claims.pdf 2011-10-07
40 1871-kolnp-2007-granted-description (complete).pdf 2011-10-07
41 01871-kolnp-2007-abstract.pdf 2011-10-07
41 1871-kolnp-2007-granted-form 1.pdf 2011-10-07
42 1871-KOLNP-2007-(10-10-2011)-OTHER PATENT DOCUMENTS.pdf 2011-10-10
42 1871-kolnp-2007-granted-form 2.pdf 2011-10-07
43 1871-KOLNP-2007-(27-01-2012)-FORM 27.pdf 2012-01-27
43 1871-kolnp-2007-granted-letter patent.pdf 2011-10-07
44 1871-KOLNP-2007-(27-01-2012)-CORRESPONDENCE.pdf 2012-01-27
44 1871-kolnp-2007-granted-specification.pdf 2011-10-07
45 1871-KOLNP-2007-(11-03-2013)-FORM-27.pdf 2013-03-11
45 1871-KOLNP-2007-OTHERS DOCUMENTS 1.1.pdf 2011-10-07
46 1871-kolnp-2007-others.pdf 2011-10-07
46 1871-KOLNP-2007-(11-03-2013)-CORRESPONDENCE.pdf 2013-03-11
47 1871-kolnp-2007-reply to examination report.pdf 2011-10-07
47 1871-KOLNP-2007-(29-01-2014)-FORM-27.pdf 2014-01-29
48 1871-KOLNP-2007_EXAMREPORT.pdf 2016-06-30
48 1871-kolnp-2007-translated copy of priority document 1.1.pdf 2011-10-07
49 LP-1871-KOLNP-2007-01-02-2023-ALL DOCUMENTS.pdf 2023-02-01
49 abstract-01871-kolnp-2007.jpg 2011-10-07

ERegister / Renewals

3rd: 14 Mar 2011

From 18/10/2007 - To 18/10/2008

4th: 14 Mar 2011

From 18/10/2008 - To 18/10/2009

5th: 14 Mar 2011

From 18/10/2009 - To 18/10/2010

6th: 14 Mar 2011

From 18/10/2010 - To 18/10/2011

7th: 10 Oct 2011

From 18/10/2011 - To 18/10/2012

8th: 11 Oct 2012

From 18/10/2012 - To 18/10/2013

9th: 09 Oct 2013

From 18/10/2013 - To 18/10/2014