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  • C07D207/04—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members
  • C07D207/08—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members with hydrocarbon radicals, substituted by hetero atoms, attached to ring carbon atoms
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  • C07D295/04—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms with substituted hydrocarbon radicals attached to ring nitrogen atoms
  • C07D295/10—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms with substituted hydrocarbon radicals attached to ring nitrogen atoms substituted by doubly bound oxygen or sulphur atoms
  • C07D295/104—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms with substituted hydrocarbon radicals attached to ring nitrogen atoms substituted by doubly bound oxygen or sulphur atoms with the ring nitrogen atoms and the doubly bound oxygen or sulfur atoms attached to the same carbon chain, which is not interrupted by carbocyclic rings
  • C07D295/108—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms with substituted hydrocarbon radicals attached to ring nitrogen atoms substituted by doubly bound oxygen or sulphur atoms with the ring nitrogen atoms and the doubly bound oxygen or sulfur atoms attached to the same carbon chain, which is not interrupted by carbocyclic rings to an acyclic saturated chain
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  • A61P25/00—Drugs for disorders of the nervous system
  • A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P27/00—Drugs for disorders of the senses
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  • A61P27/06—Antiglaucoma agents or miotics
• • A—HUMAN NECESSITIES
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  • C07D211/00—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings
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  • C07D211/06—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members
  • C07D211/08—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members with hydrocarbon or substituted hydrocarbon radicals directly attached to ring carbon atoms
  • C07D211/18—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members with hydrocarbon or substituted hydrocarbon radicals directly attached to ring carbon atoms with substituted hydrocarbon radicals attached to ring carbon atoms
  • C07D211/20—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members with hydrocarbon or substituted hydrocarbon radicals directly attached to ring carbon atoms with substituted hydrocarbon radicals attached to ring carbon atoms with hydrocarbon radicals, substituted by singly bound oxygen or sulphur atoms
  • C07D211/22—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members with hydrocarbon or substituted hydrocarbon radicals directly attached to ring carbon atoms with substituted hydrocarbon radicals attached to ring carbon atoms with hydrocarbon radicals, substituted by singly bound oxygen or sulphur atoms by oxygen atoms
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  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D213/00—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members
  • C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
  • C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
  • C07D213/24—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with substituted hydrocarbon radicals attached to ring carbon atoms
  • C07D213/28—Radicals substituted by singly-bound oxygen or sulphur atoms
  • C07D213/30—Oxygen atoms
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  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D213/00—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members
  • C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
  • C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
  • C07D213/60—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
  • C07D213/61—Halogen atoms or nitro radicals
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  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D213/00—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members
  • C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
  • C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
  • C07D213/60—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
  • C07D213/62—Oxygen or sulfur atoms
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  • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
  • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
  • C07D403/06—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings linked by a carbon chain containing only aliphatic carbon atoms

Definitions

• the present invention relates to novel compounds which are selective for muscarinic acetylcholine receptor subtypes, as well as to methods for activating muscarinic receptors and for treating or alleviating diseases in which modification of muscarinic receptor activity is beneficial.
• Muscarinic acetylcholine receptors play a central role in the central nervous system for higher cognitive functions, as well as in the peripheral parasympathetic nervous system. Cloning has established the presence of five distinct muscarinic receptor subtypes (termed ml-m5) (cf. T.I. Bonner et al, Science 237, 1987, pp. 527- 532; T.I. Bonner et al., Neuron 1, 1988, pp. 403-410).
• ml is the predominant subtype in the cerebral cortex and is believed to be involved in the control of cognitive functions
• m2 is predominant in heart and is believed to be involved in the control of heart rate
• m3 is believed to be involved in gastrointestinal and urinary tract stimulation as well as sweating and salivation
• m4 is present in brain
• m5 is present in brain and may be involved certain functions of the central nervous system associated with the dopaminergic system.
• muscarinic ligands Animal studies of various muscarinic ligands (S. Iversen, Life Sciences 60 (Nos. 13/14), 1997, pp. 1 145-1152) have shown that muscarinic compounds have a profound effect on cognitive functions, e.g. learning and memory. This would suggest a potential utility of muscarinic agonists in the improvement of cognitive functions in diseases characterized by cognitive impairment, both age-related (such as Alzheimer ' s disease or other dementias) and not age-related (such as attention deficit hyperactivity disorder).
• age-related such as Alzheimer ' s disease or other dementias
• attention deficit hyperactivity disorder such as attention deficit hyperactivity disorder
• muscarinic receptor subtype is the more abundant one in the cerebral cortex, basal ganglia and hippocampus where it accounts for 35-60% of all muscarinic receptor binding sites (cf. A. Levey, Proc. Natl. Acad. Sci.. USA 93, 1996, pp. 13541- 13546).
• the ml (and possibly m4) subtype plays a major role as a postsynaptic muscarinic receptor (located on cholinoceptive neurons in the neocortex and hippocampus) in various cognitive and motor functions and is likely to be a major contributor to the ml responses measured in these regions of the brain.
• conditions associated with cognitive impairment such as Alzheimer's disease, are accompanied by selective loss of acetylcholine in the brain. This is believed to be the result of degeneration of cholinergic neurons in the basal forebrain which innervate areas of the association cortex and hippocampus involved in higher processes (cf. S. Iversen, supra). This finding would suggest that such conditions may be treated or at least ameliorated with drugs that augment the cholinergic function in the affected areas of the brain.
• acetylcholine esterase (AChE) inhibitors such as 9-amino-l,2,3,4- tetrahydroacridine (tacrine) results in an increase of acetylcholine in the brain which indirectly causes stimulation of muscarinic receptors.
• AhE acetylcholine esterase
• tacrine 9-amino-l,2,3,4- tetrahydroacridine
• Tacrine treatment has resulted in a moderate and temporary cognitive improvement in Alzheimer's patients (cf. Kasa et al., supra).
• tacrine has been found to have cholinergic side effects due to the peripheral acetylcholine stimulation. These include abdominal cramps, nausea, vomiting, diarrhea, anorexia, weight loss, myopathy and depression. Gastrointestinal side effects have been observed in about a third of the patients treated.
• Tacrine has also been found to cause significant hepatotoxicity, elevated liver transaminases having been observed in about 30% of the patients (cf. P. Taylor, "Anticholinergic Agents". Chapter 8 in Goodman and Gilman: The Pharmacological Basis of Therapeutics, 9 th Ed., 1996, pp. 161-176).
• the adverse effects of tacrine have severely limited its clinical utility.
• Another AChE inhibitor, (R,S)-l-benzyl-4-[5,6- dimethoxy-l-indanon-2yl]methylpiperidine.HCl (donepezil) has recently been approved for the treatment of symptoms of mild to moderate Alzheimer ' s disease (cf. P. Kasa et al, supra). No hepatic damage has been observed for this compound but it has gastrointestinal effects similar to those of tacrine, probably due to stimulation of the m3 receptor caused by elevated parasympathetic tone.
• muscarinic agonists (believed to be ml selective) hitherto suggested for the treatment of Alzheimer's disease, such as arecoline, have not shown greater efficacy in clinical trials than AChE inhibitors (cf. S.V.P. Jones et al., supra).
• arecoline was found to have not so much cognitive enhancing effects as effects on behavioral changes often observed in Alzheimer's disease patients, such as a significant increase in motor activity, significant uplifting of mood, and significant decrease in anergia.
• ml agonists have later been found to be weak partial agonists selective for the m2 and/or m3 receptor subtypes (H. Brauner-Osborne et al., J. Med. Chem. 38, 1995, pp. 2188-2195).
• m2 subtype selectivity is presumed to be responsible for the cardiovascular effects observed for these agonists, e.g. tachycardia and bradycardia, and m3 activity is believed to account for the adverse gastrointestinal effects of the agonists.
• m2 and/or m3 activity is therefore a significant drawback for the muscarinic agonists proposed until now for the treatment of Alzheimer's disease, severely limiting the doses of the drugs which it has been possible to administer to patients who may therefore have received sub-optimal doses.
• the lack of subtype selectivity and low potency of the currently tested cholinergic compounds appear to favor the negative peripheral side effects and have limited cognitive effects because of weak and/or opposing actions in the brain. It would therefore be of great advantage to develop compounds which have an improved selectivity for the ml subtype, but which have little or no activity on the m2 and m3 subtypes.
• the present invention provides compounds with muscarinic agonist activity of the general formula (I):
• Xi, X 2 , X 3 , X4 and X5 are selected from C, N and O; k is 0 or 1; t is 0, 1 or 2;
• Ri is straight or branched-chain d-8 alkyl, C 2-8 alkenyl, C 2-8 alkynyl, C ⁇ -8 alkylidene, C ⁇ - alkoxy, C ⁇ -8 heteroalkyl, C ⁇ -8 aminoalkyl, C ⁇ -8 haloalkyl, C ⁇ -8 alkoxycarbonyl, C ⁇ -8 hydroxyalkoxy, C ⁇ -8 hydroxyalkyl, -SH, C ⁇ -8 alkylthio, -O-CH 2 -
• -C(O)-C 5-6 aryl substituted with C ⁇ -3 alkyl or halo; C 5-6 aryl or C 5-6 cycloalkyl optionally comprising 1 or more heteroatoms selected from N, S and O; -C(O)NR 3 R 4 ,
• R 4 and R are the same or different, each independently being selected from H, C ⁇ -6 alkyl; C 5 - 6 aryl optionally comprising 1 or more heteroatoms selected from N, O and
• A is C 5- ⁇ 2 aryl or C 5- cycloalkyl, each optionally comprising 1 or more heteroatoms selected from N, S and O;
• R 2 is H, amino, hydroxyl, halo, or straight or branched-chain C ⁇ -6 alkyl, C -6 alkenyl, C 2-6 alkynyl, C ⁇ - alkoxy, C ⁇ - heteroalkyl, C ⁇ - 6 aminoalkyl, C ⁇ - haloalkyl, C1.6 alkylthio, C ⁇ -6 alkoxycarbonyl, -CN, -CF 3 , -OR 3 , -COR 3 , NO 2 , -NHR 3 , - NHC(O)R 3 , -C(O)NR 3 R 4 , -NR3R1, -NR3C(O)NR4R 5 , -OC(O)R 3 , -C(O)R 3 R4, -O(CH 2 ) q NR 3 , -CNR 3 t or -(CH 2 ) q NR 3 R4; where q is an integer from 1 to 6; n is 0, 1, 2, 3
• Z is CR 8 R 9 wherein R 8 and R are independently selected from H, and straight or branched chain C ⁇ . 8 alkyl; or a pharmaceutically acceptable salt, ester or prodrug thereof.
• the present invention further provides pharmaceutical compositions comprising an effective amount of a compound of formula (I).
• compositions comprising a compound of formula (I).
• the present invention provides a method of treating the symptoms of a disease or condition associated with increased intraocular pressure. such as, for example, glaucoma, where the method comprises administering a therapeutically effective amount of a composition comprising a compound of formula
• Figure 1 is a graph showing raw data from one 96-well microtiter plate of screening of 35,000 small organic molecules in the assay described in Example XVI.
• Figure 2 is a graph showing data comparing the profile of the reference antagonist atropine with ml muscarinic receptor transfected cells stimulated with either carbachol (open triangles) or compound A (Example I) (closed triangles).
• the present invention provides compounds preferably showing a relatively high selectivity towards the ml receptor subtype relative to other muscarinic subtypes which may have a beneficial effect in the treatment of cognitive impairment such as Alzheimer's disease or other conditions associated with age-related cognitive decline while avoiding the adverse affects of the drugs hitherto suggested for this purpose.
• Compounds exhibiting this property have surprisingly been isolated by screening against ml-m5 receptor subtypes.
• the present invention provides compounds of formula (I), wherein
• - 5 - Xi, X 2 , X 3 , X4 and X 5 are C; or one of Xi, X 2 , X 3 , X 4 or X 5 is O or N and the others are C; k is 0 or 1 ; t is 1; Ri is straight or branched-chain C ⁇ -8 alkyl, C 2-8 alkenyl, C -8 alkynyl, C ⁇ -8 alkylidene, C ⁇ -8 alkoxy, C ⁇ -8 aminoalkyl, C ⁇ -8 haloalkyl, C ⁇ -8 alkoxycarbonyl, -C(O)NR 3 R 4 , -NR3R1, -NR 3 C(O)NR4R5, -OC(O)R 3 , or -(CH 2 ) S NR 3 R4; where R 3 , P and R 5 are the same or different, each independently being selected from H and C ⁇ -6 alkyl; and s is an integer from 1 to
• A is phenyl or naphthyl; where R 2 is straight or branched-chain C ⁇ -6 alkyl, C 2 . 6 alkenyl, C 2- alkynyl, C ⁇ -6 alkoxy, C ⁇ -6 aminoalkyl, C ⁇ -6 haloalkyl, C ⁇ -6 alkoxycarbonyl, -CN, -CF 3 , -OH, -COR 3 , -NHR 3 , -NHC(O)R 3 , -C(O)NR 3 R 4 , -NR 3 R 4 , -NR 3 C(O)NR4R 5 , -OC(O)R 3 , or -CH 2 ) q NR 3 R 4 ; where q is an integer from 1 to 6; or
• A is aryl comprising 1 or more heteroatoms selected from N, S and O;
• R 2 is H, halo, straight or branched-chain C ⁇ . 6 alkyl, C 2-6 alkenyl, C 2 . alkynyl, C ⁇ -6 alkoxy, C ⁇ -6 heteroalkyl, C ⁇ -6 aminoalkyl, C ⁇ -6 haloalkyl, C ⁇ - alkoxycarbonyl, -CN, - CF 3 , OH, -COR3, -NHR 3 , -NHC(O)R 3 , -C(O)NR 3 Rt, -NR3R,, -NR 3 C(O)NR4R 5 , - OC(O)R 3 or -(CH 2 ) q NR 3 R4; or a pharmaceutically acceptable salt, ester or prodrug thereof.
• the compound is of the formula (II):
• Preferred subgeneric embodiments of compounds of formula (II) include compounds of formulas (Ila) and (lib):
• t is 1 and Y is -C(O)-, -NHC(O)-, S, O or -OC(O)-.
• X 3 is C.
• Ri is alkyl, where preferably R 2 is alkyl, aminoalkyl, alkoxy or hydroxyl. In one embodiment, p is 3. In another. Ri is C 2 . 8 alkyl and R 2 is methyl, hydroxyl or alkoxy.
• n is 1 or 2; Y is -C(O)- or O and t is 1.
• R 2 is halo.
• X 3 may also be N, where according to one embodiment, Ri is alkyl or alkoxy; or Ri is benzyl or phenyl; where R 2 is alkyl or alkoxy.
• X 3 is O, where t may be, for example, 0.
• R 2 is alkyl or alkoxy; or R 2 is halo.
• Particular embodiments of the invention include: 4-Methoxy- 1 -[4-(2-methylphenyl)-4-oxo- 1 -butyljpiperidine: 4-Ethoxy- 1 -[4-(2-methylphenyl)-4-oxo- 1 -butyljpiperidine; 4-Propoxy- 1 -[4-(2-methylphenyl)-4-oxo- 1 -butyl]piperidine; 4-Butoxy- 1 -[4-(2-methylphenyl)-4-oxo- 1 -butyljpiperidine: 4-Methoxymethyl- 1 -[4-(2-methylphenyl)-4-oxo- 1 -butyljpiperidine; 4-Ethoxymethyl- 1 -[4-(2-methylphenyl)-4-oxo- 1 -butyljpiperidine; 4-Propoxymethyl- 1 -[4-(2-methylphenyl)-4-oxo- 1 -butyljpiperidine
• (CH 2 ) P -Y- is -(CH 2 ) 3 -C(O)- or -(CH 2 ) 3 -S-; and Xi through X 5 are C; such that -A-
• R 2 n and Ri are not together: o-methyl-phenyl and n-butyl, respectively; phenyl and n-butyl, respectively; or p-fluoro-phenyl and -O-(CH 2 ) 2 CH 3 , respectively).
• the present invention further provides a method of agonizing a muscarinic receptor comprising contacting the receptor with an effective amount of a compound of formula (I), inclusive of all compounds within the scope of formula (I)(i.e., including 4-rc-Butyl- 1 -[4-phenyl-4-oxo- 1 -butyljpiperidine; 4-n-Butyl- 1 -[4-(2- methylphenyl)-4-oxo-l -butyl] piperazine; 2-[3-(3- «-Butylpiperidine)propanesulfanyl] toluene; and 4-Propyloxy- 1 -[4-(4-fluorophenyl)-4-oxo- 1 -butyljpiperidine).
• a compound of formula (I) inclusive of all compounds within the scope of formula (I)(i.e., including 4-rc-Butyl- 1 -[4-phenyl-4-oxo- 1
• compositions comprising an effective amount of a compound of formula (I), inclusive of all compounds within the scope of formula (I)(i.e., including - «-Butyl-l-[4-phenyl-4-oxo-l -butyljpiperidine; 4- ⁇ -Butyl- l-[4-(2-methylphenyl)-4-oxo-l-butyl]piperazine; 2-[3-(3- «-
• Butylpiperidine propanesulfanyl] toluene; and 4-Propyloxy-l-[4-(4-fluorophenyl)-4- oxo-1 -butyljpiperidine).
• the present invention further also provides methods of treating the symptoms of a disease or condition associated with reduced levels of acetylcholine, the method comprising administering a therapeutically effective amount of a composition described herein.
• diseases or conditions include neurogenerative disease, cognitive impairment, age-related cognitive decline or dementia.
• Glaucoma is a disease in which an abnormality is observed in the circulation-control mechanism of the aqueous humor filling up the anterior chamber, i.e., the space formed between the cornea and the lens. This leads to an increase in the volume of the aqueous humor and an increase in intraocular pressure, consequently leading to the visual field defect and even to loss of eyesight due to the compulsion and contraction of the papillae of the optic nerve.
• the compounds of the present invention preferably show selective agonist activity towards the ml receptor.
• Such an agonist is defined as a compound that increases the activity of the ml muscarinic receptor when it contacts the receptor.
• Selectivity is defined as a property of a muscarinic ml agonist whereby an amount of agonist effective to increase the activity of the ml receptor causes little or no increase in the activity of the m3 and m5 subtypes, and preferably the m2 and m4 subtypes.
• alkyl means a straight or branched-chain alkane group with 1-6 carbon atoms in the chain, for instance methyl, ethyl, propyl, isopropyl, n-
• heteroalkyl is intended to indicate an alkane group containing 1 or 2 heteroatoms selected from O, S or N.
• alkenyl means a straight or branched-chain alkene group with 2-6 carbon atoms in the chain; the term “alkynyl” is intended to indicate a straight or branched-chain alkyne group with 2-6 carbon atoms in the chain.
• aryl and “cycloalkyl” preferably refer to mono- and bicyclic ring structures comprising 5 to 12 carbon atoms, more preferably monocyclic rings comprising 5 to 6 carbon atoms. Where such rings comprise one or more heteroatoms, selected from N, S and O, (i.e., heterocyclic rings) such rings comprise a total of 5 to 12 atoms, more preferably 5 to 6 atoms.
• Heterocyclic rings include, but are not limited to, furyl, pyrrolyl, pyrazolyl, thienyl, imidazolyl, isoxazolyl, oxazolyl, thiazolyl, isothiazolyl, pyridyl, piperidinyl, piperazinyl, pyridazinyl, pyrimidinyl, pyrazinyl, morpholinyl, oxadiazolyl, thiadiazolyl, imidazolinyl, imidazolidinyl and the like.
• the ring may be substituted by one or more of the groups included in the definition of R 2 above.
• substituents C ⁇ -6 alkyl, C ⁇ -6 alkenyl, C ⁇ -6 alkynyl, C ⁇ -6 alkoxy, C ⁇ - heteroalkyl, C ⁇ -6 aminoalkyl, C ⁇ - haloalkyl or C ⁇ - 6 alkoxycarbonyl may, if present, be substituted by one or more of hydroxyl, C alkoxy, halogen, cyano, amino or nitro.
• halogen or halo includes chlorine, fluorine, iodine and bromine.
• the starting compound having formula (X) may be prepared by general methods of organic synthesis.
• general methods of preparing compounds of formula (X) reference is made to Fuller, R. W. et al., J. Med. Chem. 14:322-325 (1971); Foye, W. O., et al.. J. Pharm.Sci. 68:591-595 (1979); Bossier, J. R. et al.. Chem. Abstr. 66:46195h and 67:21527a (1967); Aldous, F. A. B., J. Med. Chem. 17:1100-1111 (1974); Fuller, R. W. et al., J. Pharm. Pharmacol.
• radiolabelled derivatives having formula (XX) may be prepared by, for example, using a tritiated reducing agent to form the reductive amination or by utilizing a l4 C-labelled starting material.
• the compound having the formula (XXII) may be reduced with, for example, AIH 3 , diborane:methyl sulfide or other standard carbonyl reducing reagents to produce the ligand having the formula (XXX).
• the receptor ligands having formula (XXXII) may be prepared by nucleophilic displacement of an electrophile (E) by the amino derivative (XXXI).
• electrophiles which may be used for this purpose include halides such as I, Cl, Br, tosylate or mesylate.
• this compound may be prepared from oxidation of an sec. alcohol with for example pyridinium chlorocromate or N- chlorosuccimmide or CrO 3 -H 2 SO or nickel peroxide or metal (Al, K) or DCC- DMSO.
• this compound may be prepared by alkylation of an alcohol with arylhalides under for example Cu catalysis.
• this compound may be prepared by alkylation of a thiol with arylhalides under for example Cu catalysis.
• Suitable pharmaceutically acceptable salts of the compounds of this invention include acid addition salts which may, for example, be formed by mixing a solution of the compound according to the invention with a solution of a pharmaceutically acceptable acid such as hydrochloric acid, sulphuric acid, fumaric acid, maleic acid, succinic acid, acetic acid, benzoic acid, oxalic acid, citric acid, tartaric acid, carbonic acid or phosphoric acid.
• suitable pharmaceutically acceptable salts thereof may include alkali metal salts, e.g. sodium or potassium salts; alkaline earth metal salts, e.g. calcium or magnesium salts; and salts formed with suitable organic ligands. e.g.
• quarternary ammonium salts examples include the acetate, benzenesulfonate, benzoate, bicarbonate, bisulfate, bitartrate, borate, bromide, calcium, carbonate, chloride, clavulanate, citrate, dihydrochloride, fumarate, gluconate, glutamate, hydrobromide, hydrochloride, hydroxynaphthoate, iodide, isothionate.
• lactate lactobionate, laurate, maleate, mandelate, mesylate, methylbromide, methylnitrate, methylsulfate, nitrate, N-methylglucamine ammonium salt, oleate, oxalate, phosphate/diphosphate, salicylate. stearate, sulfate, succinate. tannate, tartrate, tosylate, triethiodide and valerate salt.
• the present invention includes within its scope prodrugs of the compounds of this invention.
• prodrugs are inactive derivatives of the compounds of this invention which are readily convertible in vivo into the required compound.
• Conventional procedures for the selection and preparation of suitable prodrug derivatives are described, for example, in "Design of Prodrugs,” ed. H. Bundgaard, Elsevier, 1985. Metabolites of these compounds include active species produced upon introduction of compounds of this invention into the biological milieu.
• the compounds according to the invention may exist as a racemate or as enantiomers. It should be noted that all such isomers and mixtures thereof are included in the scope of the present invention. Furthermore, some of the crystalline forms for compounds of the present invention may exist as polymorphs and as such are intended to be included in the present invention. In addition, some of the compounds of the present invention may form
• solvates with water (i.e., hydrates) or common organic solvents. Such solvates are also included in the scope of this invention.
• Such isomers may be separated by conventional techniques such as preparative chiral chromatography.
• the compounds may be prepared in racemic form, or individual enantiomers may be prepared either by stereoselective synthesis or by resolution.
• the compounds may, for example, be resolved into their component enantiomers by standard techniques, such as the formation of diastereomeric pairs by salt formation with an optically active acid, such as (-)-di-p-toluoyl-d-tartaric acid and/or (+)-di-p-toluoyl-l-tartaric acid followed by fractional crystallization and regeneration of the free base.
• the compounds may also be resolved by formation of diastereomeric esters or amides, followed by chromatographic separation and removal of the chiral auxiliary.
• any of the processes for preparation of the compounds of the present invention it may be necessary and/or desirable to protect sensitive or reactive groups on any of the molecules concerned. This may be achieved by means of conventional protecting groups, such as those described in Protective Groups in Organic Chemistry, ed. J.F.W. McOmie, Plenum Press, 1973; and T.W. Greene & P.G.M. Wuts, Protective Groups in Organic Synthesis, John Wiley & Sons, 1991.
• the protecting groups may be removed at a convenient subsequent stage using methods known from the art.
• Compounds of the present invention may be administered in any of the foregoing compositions and according to dosage regimens established in the art whenever specific pharmacological modification of the activity of muscarinic receptors is required.
• compositions comprising one or more compounds of the invention together with a pharmaceutically acceptable diluent or excipient.
• compositions are in unit dosage forms such as tablets, pills, capsules (including sustained-release or delayed-release formulations), powders, granules, elixirs, tinctures, syrups and emulsions, sterile parenteral solutions or suspensions, aerosol or liquid sprays, drops, ampoules, auto-injector devices or suppositories; for oral, parenteral (e.g. intravenous, intramuscular or subcutaneous), intranasal, sublingual or rectal administration, or for administration by inhalation or insufflation, and may be formulated in an appropriate manner and in accordance with
• compositions may be in sustained-release form suitable for once-weekly or once-monthly administration; for example, an insoluble salt of the active compound, such as the decanoate salt, may be adapted to provide a depot preparation for intramuscular injection.
• suitable topical formulations for administration e.g. eye or skin or mucosa.
• the active drug component can be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water and the like.
• suitable binders, lubricants, disintegrating agents, flavoring agents and coloring agents can also be incorporated into the mixture.
• suitable binders include, without limitation, starch, gelatin, natural sugars such as glucose or beta-lactose, natural and synthetic gums such as acacia, tragacanth or sodium alginate, carboxymethylcellulose, polyethylene glycol, waxes and the like.
• Lubricants used in these dosage forms include, without limitation, sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride and the like.
• Disintegrators include, without limitation, starch, methyl cellulose, agar, bentonite, xanthan gum and the like.
• a suitable pharmaceutical excipient e.g. such as the ones described above
• other pharmaceutical diluents e.g. water
• the active ingredient is dispersed evenly throughout the composition so that the composition may be readily subdivided into equally effective unit dosage forms such as tablets, pills and capsules.
• the solid preformulation composition may then be subdivided into unit dosage forms of the type described above containing from 0.1 to about 50 mg of the active ingredient of the present invention.
• the tablets or pills of the present composition may be coated or otherwise compounded to provide a dosage form affording the advantage of prolonged action.
• the tablet or pill can comprise an inner core containing the active compound and an outer layer as a coating surrounding the core.
• the outer coating may be an enteric layer which serves to resist disintegration in the stomach and permits the inner core to pass intact into the
• compositions can be incorporated for administration orally or by injection include aqueous solutions, suitably flavored syrups, aqueous or oil suspensions, and flavored emulsions with edible oils such as cottonseed oil, sesame oil, coconut oil or peanut oil, as well as elixirs and similar pharmaceutical carriers.
• Suitable dispersing or suspending agents for aqueous suspensions include synthetic and natural gums such as tragacanth, acacia, alginate. dextran, sodium carboxymethylcellulose, gelatin. methylcellulose or polyvinyl-pyrrolidone. Other dispersing agents which may be employed include glycerin and the like.
• sterile suspensions and solutions are desired.
• Isotonic preparations which generally contain suitable preservatives are employed when intravenous administration is desired.
• the compositions can also be formulated as an ophthalmic solution or suspension formation, i.e., eye drops, for ocular administration
• the present invention also relates to a method of alleviating or treating a disease or condition in which modification of muscarinic receptor activity, in particular ml receptor activity, has a beneficial effect by administering a therapeutically effective amount of a compound of the present invention to a subject in need of such treatment.
• diseases or conditions may, for instance arise from inadequate stimulation or activation of muscarinic receptors.
• compounds which are selective for a particular muscarinic receptor subtype, in particular ml the problems with adverse side effects observed with the known muscarinic drugs, such as tachycardia or bradycardia or gastrointestinal effects, may substantially be avoided.
• subject refers to an animal, preferably a mammal, most preferably a human, who has been the object of treatment, observation or experiment.
• therapeutically effective amount means that amount of active compound or pharmaceutical agent that elicits the biological or medicinal response in a tissue, system, animal or human that is being sought by a researcher, veterinarian, medical doctor or other clinician, which includes alleviation of the symptoms of the disease being treated.
• compounds of general formula I exhibit subtype selectivity for the muscarinic ml receptor subtype.
• the compounds exhibit selectivity for the muscarinic ml receptor subtype compared to other human G-protein coupled receptors tested including serotonin, histamine, dopamine or adrenergic receptors.
• selectivity is that these compounds may be effective in the treatment or amelioration of a number of diseases and disorders of the central nervous system without the undesirable side effects previously observed with non- selective compounds.
• muscarinic ml receptor subtype selectivity makes them potentially very useful in treating a number of diseases and disorders characterized by cognitive impairment such as attention deficit disorder, or neurodegenerative diseases, e.g. Alzheimer's disease, other forms of age-related cognitive decline, e.g. senile dementia, or dementia-related symptoms such as decreased motor activity, mood changes, anergia, apathy, restlessness and aggressive behavior. It is currently believed that the muscarinic ml receptor may also be involved in control of intraocular pressure, and that muscarinic ml agonists may therefore be used to treat or alleviate ocular diseases such as glaucoma.
• cognitive impairment such as attention deficit disorder, or neurodegenerative diseases, e.g. Alzheimer's disease, other forms of age-related cognitive decline, e.g. senile dementia, or dementia-related symptoms such as decreased motor activity, mood changes, anergia, apathy, restlessness and aggressive behavior.
• muscarinic ml receptor may also
• compounds of the present invention may be administered in a single daily dose, or the total daily dosage may be administered in divided doses two, three or four times daily.
• compounds for the present invention may be administered in intranasal form via topical use of suitable intranasal vehicles, or via transdermal routes, using those forms of transdermal skin patches well known to persons skilled in the art.
• the dosage administration will, of course, be continuous rather than intermittent throughout the dosage regimen.
• the dosage regimen utilizing the compounds of the present invention is selected in accordance with a variety of factors including type, species, age, weight, sex and medical condition of the patient; the severity of the condition to be treated; the route of administration; the renal and hepatic function of the patient; and the particular compound employed.
• a physician or veterinarian of ordinary skill can readily determine and prescribe the effective amount of the drug required to prevent, counter or arrest the progress of the disease or disorder which is being treated.
• the daily dosage of the products may be varied over a wide range from 0.01 to 100 mg per adult human per day.
• the compositions are preferably provided in the form of tablets containing 0.01, 0.05, 0.1, 0.5, 1.0, 2.5, 5.0, 10.0, 15.0, 25.0 or 50.0 mg of the active ingredient for the symptomatic adjustment of the dosage to the patient to be treated.
• a unit dose typically contains from about 0.001 mg to about 50 mg of the active ingredient, preferably from about 1 mg to about 10 mg of active ingredient.
• An effective amount of the drug is ordinarily supplied at a dosage level of from about 0.0001 mg/kg to about 25 mg/kg of body weight per day.
• the range is from about 0.001 to 10 mg/kg of body weight per day, and especially from about 0.001 mg/kg to 1 mg/kg of body weight per day.
• the compounds may be administered on a regimen of 1 to 4 times per day.
• Compounds according to the present invention may be used alone at appropriate dosages defined by routine testing in order to obtain optimal pharmacological effect on a muscarinic receptor, in particular the muscarinic ml receptor subtype, while minimizing any potential toxic or otherwise unwanted effects.
• co-administration or sequential administration of other agents which improve the effect of the compound may, in some cases, be desirable.
• the pharmacological properties and the selectivity of the compounds of this invention for specific muscarinic receptor subtypes may be demonstrated by a number of different assay methods using recombinant receptor subtypes, preferably of the human receptors if these are available, e.g. conventional second messenger or binding assays.
• a particularly convenient functional assay system is the receptor selection and amplification assay disclosed in US 5,707,798 describing a method of screening for bioactive compounds by utilizing the ability of cells transfected with receptor DNA, e.g. coding for the different muscarinic subtypes, to amplify in the presence of a ligand of the receptor. Cell amplification is detected as increased levels of a marker also expressed by the cells.
• Example I 4-n-Butyl-l-[4-(2-methylphenyl)-4-oxo-l-butyl]piperidine (5) l-Benzyl-4-n-butylidenepiperidine (2).
• the resulting suspension was heated to 90°C for 30 min, until the evolution of hydrogen ceased.
• the suspension was cooled on an ice-bath for 20 min followed by addition of a slurry of butyltriphenylphosphonium bromide (26.6 g, 67 mmol) in DMSO (70 mL).
• Example II 3-Hydroxymethyl-[4-(2-methylphenyl)-4-oxo-l-butyl]piperidine (7) 4-(3-Hydroxymethyl-piperidin-l-yl)-butyronitrile (6).
• piperidine-3-yl-methanol (1.12 g, 10 mmol) in acetonitrile (10 mL)
• potassium carbonate (1.38 g, 10 mmol
• 4- bromobutyronitrile (0.90 mL, 9 mmol).
• the reaction mixture was stirred at rt. for 12 h.
• the mixture was filtered and evaporated to dryness.
• reaction mixture was quenched by addition of H 2 SO (10 mL, 2 M) and stirred at rt for 12 hours followed by addition of NaOH (10 mL, 2 M).
• Addition of THF (15 mL) was followed by extraction with ethyl acetate (3 x 50 mL) and the combined organic phases are washed with brine (10 mL) and NaOH (10 mL, 2 M), dried (MgS0 4 ) and evaporated to dryness to produce 0.43 g of crude 9.
• Example VII 4-Methyl-l-[4-(2-methylphenyl)4-oxo-l-butyl]piperazine (15) 4-(4-Methyl-piperazin-l-yl)-butyronitrile (14).
• 1- methyl-piperazine 0.52 g, 5.1 mmol
• 4-bromobutyronitrile 0.78 g, 5.3 mmol
• potassium carbonate 0.71 g, 5.3 mmol
• Example IX 4-n-Butyl-l-[4-(2-ethoxyphenyl)-4-oxo-l-butylJpiperidine (18)
• Mg turnings 94 mg, 3.8 mmol
• Et 2 0 3 mL
• reaction mixture was quenched by addition of H 2 S0 (7 mL, 2 M) and stirred at rt for 3 hours, followed by addition of NaOH (20 mL, 2 M) until basic conditions.
• the reaction mixture was extracted with ethyl acetate (3 x 50 mL) and the combined organic phases are washed with brine (10 mL) and NaOH (10 mL, 2 M), and the organic phases were dried (MgS0 4 ) and evaporated to dryness to produce 0.69 g of crude 19.
• the crude product was subjected to CC [eluent: CH 2 C1 2 : MeOH (99:1)] to give pure 19 (0.40 g, 64 %); LC-MS [M + H] + 315 (cald. 315.5).
• reaction mixture was quenched by addition of H 2 S0 (8 mL, 2 M) and stirred at rt for 4 hours, then the reaction mixture was basified by addition of NaOH (20 mL, 2 M).
• Addition of THF (20 mL) was followed by extraction with ethyl acetate (3 x 50 mL) and the combined organic phases are washed with brine (10 mL) and NaOH (10 mL, 2 M), and the organic phases were dried (MgS0 ) and evaporated to dryness to produce 0.61 g of crude 20.
• the crude product was subjected to CC [eluent: CH 2 C1 2 : MeOH (99:1)] to give pure 20 (0.21 g, 35 %); LC-MS [M+ Hf 315 (cald. 315.5).
• the reaction mixture was basified by addition of NaOH (2 mL, 2.0 M) and filtered through celite.
• the aqueous phase was extracted with ethyl acetate (3 x 50 mL) and the combined organic phases were washed with brine (10 mL) and NaOH (10 mL, 2 M), dried (MgS0 4 ) and evaporated to dryness to produce 0.42 g of crude 27.
• the crude product was subjected to CC [eluent: : CH 2 C1 2 : MeOH (99:1)] to give pure 27 (0.21 g, 58 %); LC-MS [M+ H] + 303 (cald. 303.2).
• NIH 3T3 cells available from the American Type Culture Collection as ATCC CRL 1658
• NIH 3T3 cells were grown at 37°C in a humidified atmosphere (5% C0 2 ) incubator in Dulbecco ' s modified Eagle's medium (DMEM) supplemented with 4.5 g/1 glucose, 4 mM glutamine, 50 units/ml penicillin, 50 units/ml streptomycin (available from Advanced Biotechnologies, Inc., Gaithersburg, MD) and 10% calf serum (available from Sigma, St. Louis, MI).
• DMEM Dulbecco ' s modified Eagle's medium
• the cells were treated with trypsin-EDTA, spun down and plated at 2xl0 6 per 15 cm dish in 20 ml of DMEM containing 10% calf serum.
• the ml-m5 muscarinic receptor subtypes were cloned substantially as described by Bonner et al., Science 237, 1987, p. 527, and Bonner et al., Neuron 1, 1988, p. 403.
• the cells were co-transfected with DNA encoding a chimera between the Gq protein and the five carboxy-terminal amino acids of the Gi protein (the Gq-i5 construct is described by Conklin et al., Nature 363, 1993, p.274).
• the cells were transfected using the Superfect transfection reagent (available from Qiagen, Valencia, CA) in accordance with the manufacturer's instructions.
• 60 ⁇ l Superfect was added to the DNA and mixed thoroughly by pipetting up and down several times. The mixture was incubated at room temperature for 10-15 minutes.
• the media were aspirated, and 12 ml fresh DMEM containing 10% calf serum and 50 units/ml penicillin/streptomycin was added to the plates.
• the DNA-Superfect solution was mixed once more with a pipette and added to the plate which was swirled to distribute the DNA mixture evenly over the surface.
• the cells were incubated overnight at 37°C and 5% C0 2 .
• the media were aspirated and the plates were rinsed once with a 15 ml volume of Hank's Buffered Saline. The plates were swirled to ensure thorough rinsing. 20 ml fresh DMEM supplemented with 10% calf serum and 50 units/ml penicillin/streptomycin was added to the plates. The cells were incubated for 24-48 hours until the plates were 100% confluent.
• DMEM containing 2% Cyto-SF3 was heated at 37°C in a water bath under sterile conditions.
• Sterile working stock solutions of test compounds to be assayed were prepared by diluting the compounds in DMEM to 8x the final concentration for testing.
• the compound (carbachol) to be included in the assay as a positive control was also diluted in DMEM to 8x the final concentration.
• 50 ⁇ l of the DMEM containing 2% Cyto-SF3 was added to each well of 96-well microtiter plates under sterile conditions.
• HBS Hank's Buffered Saline
• the plates were rinsed with this medium, and the cells were pipetted into a tube.
• the cells were centrifuged at 1000 rpm for 5-10 min. in an IEC Centra CL2 centrifuge (produced by Sorvall). Afterwards, the medium was carefully aspirated so as not to dislodge the cells.
• the cell pellet was suspended in 1600 ⁇ l DMEM containing 10% calf serum and 50 units/ml penicillin and 50 units/ml streptomycin, after which 20 ml DMEM supplemented with 2% Cyto-SF3 was added. 50 ⁇ l aliquots of this cell suspension was added to the wells of the 96-well microtiter plates prepared above (except for the plate control wells). The plates were then incubated for 4 days at 37°C and 5% C0 2 .
• NIH 3T3 cells were co- transfected with DNAs encoding the ml, m3 and m5 receptor subtypes.
• a compound library containing approximately 35,000 small organic compounds (1 per well) was screened against the receptors by the procedures described above.
• Fig. 1 illustrates data from one 96-well plate from the screen. On this plate, two compounds were active at one or more of the transfected receptors. In the total screen, four related compounds were identified that showed activity. To determine which of the receptors were activated in the screen, the compounds were tested as described above against each of the receptors transfected into separate cell cultures. Compound A only activated the ml receptor subtype, at which it was a potent partial agonist, inducing a lower maximal response than the reference compound carbachol.
• the four compounds were found to selectively activate the ml receptor with no significant activity at the m2, m3, m4 or m5 muscarinic receptors.
• the most active compound, compound A was not an antagonist of carbachol-induced responses of the five muscarinic receptor subtypes.
• Compound A was further tested for agonist activity against several other receptors at the ⁇ -adrenergic receptor subtypes ID, IB, 1A, 2A, 2B and 2C, the histamine HI and the serotonin 5-HT1A and 5-HT2A subtypes. The compound showed no significant activity in these assays. In antagonist experiments, compound A did not inhibit responses of the ⁇ -adrenergic receptor subtypes 2 A, 2B or 2C, or the serotonin receptor subtypes 5-HT1A or 5-HT2A. As illustrated in Figure 2, the responses induced by compound A were blocked by the muscarinic antagonist atropine with the same potency as were responses induced by the muscarinic agonist carbachol.
• R-SAT assays (see U.S. patent no. 5,707,798, incorporated herein by reference) were carried out where cells transfected with ml, m3 or m5 receptors were exposed to seven compounds at 1.5 ⁇ M concentration. Cellular response is expressed as a percentage of the maximum response of the cells (defined as response to 10 ⁇ M carbachol). The results are presented in the following table.
• the compounds are selective agonists of the ml receptor.

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