Classifications

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  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D235/00—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, condensed with other rings
  • C07D235/02—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, condensed with other rings condensed with carbocyclic rings or ring systems
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
  • A61P1/04—Drugs for disorders of the alimentary tract or the digestive system for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
  • A61P1/08—Drugs for disorders of the alimentary tract or the digestive system for nausea, cinetosis or vertigo; Antiemetics
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P11/00—Drugs for disorders of the respiratory system
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P11/00—Drugs for disorders of the respiratory system
  • A61P11/02—Nasal agents, e.g. decongestants
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P25/00—Drugs for disorders of the nervous system
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P25/00—Drugs for disorders of the nervous system
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• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P25/00—Drugs for disorders of the nervous system
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• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P3/00—Drugs for disorders of the metabolism
  • A61P3/04—Anorexiants; Antiobesity agents
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P3/00—Drugs for disorders of the metabolism
  • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
  • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P37/00—Drugs for immunological or allergic disorders
  • A61P37/08—Antiallergic agents
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D235/00—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, condensed with other rings
  • C07D235/02—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, condensed with other rings condensed with carbocyclic rings or ring systems
  • C07D235/04—Benzimidazoles; Hydrogenated benzimidazoles
  • C07D235/06—Benzimidazoles; Hydrogenated benzimidazoles with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached in position 2
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D405/00—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom
  • C07D405/02—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings
  • C07D405/12—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings linked by a chain containing hetero atoms as chain links

Definitions

• the present invention relates to novel imidazo heterocyclic compounds, to the use of these compounds as pharmaceutical compositions, to pharmaceutical compositions comprising the compounds, and to a method of treatment employing these compounds and compositions.
• the present compounds show a high and selective binding affinity to the histamine H3 receptor indicating histamine H3 receptor antagonistic, inverse agonistic or agonistic activity. As a result, the compounds are useful for the treatment and/or prevention of diseases and disorders related to the histamine H3 receptor.
• histamine H3 receptor The existence of the histamine H3 receptor has been known for several years and the receptor is of current interest for the development of new medicaments (see eg Stark, H.; Schlicker, E.; Schunack, W., Drugs Fut. 1996, 21, 507-520; Leurs, R.; Timmerman, H.; Vollinga, R. C, Progress in Drug Research 1995, 45, 107-165). Recently, the human histamine H3 receptor has been cloned, cf Lovenberg, T.W. et al, Molecular Pharmacology, June 1999, 55, 1101-1107.
• the histamine H3 receptor is a presynaptic autoreceptor located both in the central and the peripheral nervous system, the skin and in organs such as the lung, the intestine, probably the spleen and the gastrointestinal tract. Recent evidence suggests that the H3 receptor show intrinsic, constitutive activity, in vitro as well as in vivo (i.e. it is active in the absence of an agonist; see eg Morisset et al., Nature 2000, 408, 860-864). This activity can be inhibited by compounds acting as inverse agonists.
• the histamine H3 receptor has been demonstrated to regulate the release of histamine and also of other neurotransmitters such as serotonin and acetylcholine.
• a histamine H3 receptor antagonist or inverse agonist would be expected to increase the release of these neurotransmitters in the brain.
• a histamine H3 receptor agonist leads to an inhibition of the biosynthesis of histamine and an inhibition of the release of histamine and also of other neurotransmitters such as serotonin and acetylcholine.
• Helv. Chim. Acta 1979, 62(2), 507-10 discloses imidazole derivatives. However, they are neither disclosed nor suggested to possess histamine H3 receptor agonistic, inverse agonistic or antagonistic activity.
• novel compounds which interact with the histamine H3 receptor would be a highly desirable contribution to the art.
• the present invention provides such a contribution to the art being based on the finding that a novel class of imidazo heterocyclic compounds has a high and specific affinity to the histamine H3 receptor.
• the present compounds are useful in the treatment and/or prevention of a wide range of conditions and disorders in which an interaction with the histamine H3 receptor is beneficial.
• the compounds may find use eg in the treatment of diseases of the central nervous system, the peripheral nervous system, the cardiovascular system, the pulmonary system, the gastrointestinal system and the endocrinological system.
• Halogen designates an atom selected from the group consisting of F, Cl, Br and I.
• C ⁇ -alky in the present context designates a saturated, branched or straight hydrocarbon group having from 1 to 6 carbon atoms.
• Representative examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, butyl, isobutyl, sec-butyl, tert- butyl, n-pentyl, isopentyl, neopentyl, te/ -pentyl, n-hexyl, isohexyl and the like.
• C 2 -6-alkenyl represents a branched or straight hydrocarbon group having from 2 to 6 carbon atoms and at least one double bond.
• groups include, but are not limited to, vinyl, 1-propenyl, 2-propenyl, isopropenyl, 1 ,3- butadienyl, 1-butenyl, 2-butenyl, 3-butenyl, 2-methyl-1-propenyl, 1 -pentenyl, 2-pentenyl, 3- pentenyl, 4-pentenyl, 3-methyl-2-butenyl, 1-hexenyl, 2-hexenyl, 3-hexenyl, 2,4-hexadienyl, 5- hexenyl and the like.
• C 2 - 6 -alkyny represents a branched or straight hydrocarbon group having from 2 to 6 carbon atoms and at least one triple bond.
• groups include, but are not limited to, ethynyl, 1-propynyl, 2-propynyl, 1-butynyl, 2-butynyl, 3- butynyl, 1-pentynyl, 2-pentynyl, 3-pentynyl, 4-pentynyl, 1-hexynyl, 2-hexynyl, 3-hexynyl, 5- hexynyl, 2,4-hexadiynyl and the like.
• C-i-e-alkoxy in the present context designates a group -O-d-e-alkyl wherein C ⁇ e-alkyl is as defined above.
• Representative examples include, but are not limited to, meth- oxy, ethoxy, n-propoxy, isopropoxy, butoxy, isobutoxy, sec-butoxy, te/ -butoxy, n-pentoxy, isopentoxy, neopentoxy, te/t-pentoxy, n-hexoxy, isohexoxy and the like.
• C-i-e-alkylthio in the present context designates a group -S-Ci- 6 -alkyl wherein C ⁇ -alkyl is as defined above.
• Representative examples include, but are not limited to, methylthio, ethylthio, n-propylthio, isopropylthio, butylthio, isobutylthio, sec-butylthio, tert- butylthio, n-pentylthio, isopentylthio, neopentylthio, terf-pentylthio, n-hexylthio, isohexylthio and the like.
• Representative examples include, but are not limited to, methylcarbonyl, ethylcarbonyl, n-propylcarbonyl, isopropylcarbonyl, butylcarbonyl, isobutylcarbonyl, sec-butylcarbonyl, te/f-butylcarbonyl, n-pentylcarbonyl, isopentylcarbonyl, neopentylcarbonyl, ferf-pentylcarbonyl, n-hexylcarbonyl, isohexylcarbonyl and the like.
• Representative examples include, but are not limited to, methylsulfonyl, ethylsulfonyl, n-propylsulfonyl, isopropylsulfonyl, butylsulfonyl, iso- butylsulfonyl, sec-butylsulfonyl, ferf-butylsulfonyl, n-pentylsulfonyl, isopentylsulfonyl, neopen- tylsulfonyl, terf-pentylsulfonyl, n-hexylsulfonyl, isohexylsulfonyl and the like.
• C 3 - 10 -cycloalkyl represents a saturated mono-, bi-, tri- or spirocarbocyclic group having from 3 to 10 carbon atoms.
• Representative examples are cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl, cyclode- cyl, bicyclo[3.2.1]octyl, spiro[4.5]decyl, norpinyl, norbonyl, norcaryl, adamantyl and the like.
• C 3 - 8 -heterocyclyl represents a saturated 3 to 8 membered ring containing one or more heteroatoms selected from nitrogen, oxygen and sulfur.
• Repre- sentative examples are pyrrolidyl, piperidyl, piperazinyl, morpholinyl, thiomorpholinyl, azirid- inyl, tetrahydrofuranyl and the like.
• aryl represents a carbocyclic aromatic ring system such as phenyl, biphenyl, naphthyl, anthracenyl, phenanthrenyl, fluorenyl, indenyl, pentalenyl, az- ulenyl, biphenylenyl and the like.
• Aryl is also intended to include the partially hydrogenated derivatives of the carbocyclic aromatic systems enumerated above. Non-limiting examples of such partially hydrogenated derivatives are 1 ,2,3,4-tetrahydronaphthyl, 1 ,4-dihydronaphthyl and the like.
• arylthio as used herein represents a group -S-aryl wherein aryl is as defined above.
• aryloxy as used herein represents a group -O-aryl wherein aryl is as defined above.
• arylamino as used herein represents a group -NH-aryl wherein aryl is as defined above.
• aryl annulated with C 3 - 8 -heterocyclyl represents a ring system which contains an aryl group as defined herein to which a C - 8 -heterocyclyl group as de- fined herein is attached and which does not fall under the below definiton of heteroaryl.
• the aryl group and the heterocyclyl group may form fused, bridged or spirocyclic ring systems.
• Representative examples are 2,3-methylenedioxyphenyl, 3,4-methylenedioxyphenyl, 2,4-methylene- dioxyphenyl, 2,5-methylenedioxyphenyl, 3,5-methylenedioxyphenyl, 3,6-methylenedioxyphenyl, chromanyl, isochromanyl, 2,3-ethylenedioxyphenyl, 3,4-ethylenedioxyphenyl, 2,4-ethylenedioxy- phenyl, 2,5-ethylenedioxyphenyl, 3,5-ethylenedioxyphenyl, 3,6-ethylenedioxyphenyl and the like.
• heteroaryl represents a heterocyclic aromatic ring system containing one or more heteroatoms selected from nitrogen, oxygen and sulfur such as fu- ranyl, thiophenyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, isoxazolyl, isothiazolyl, 1 ,2,3-tria- zolyl, 1 ,2,4-triazolyl, pyranyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, 1 ,2,3-triazinyl, 1 ,2,4-triazinyl, 1 ,3,5- triazinyl, 1 ,2,3-oxadiazolyl, 1 ,2,4-oxadiazolyl, 1 ,2,5-oxadiazolyl, 1 ,3,4-oxadiazolyl, 1 ,2,3-thiadiazolyl, 1 ,
• Heteroaryl is also intended to include the partially hydrogenated derivatives of the heterocyclic systems enumerated above.
• Non- limiting examples of such partially hydrogenated derivatives are 2,3-dihydrobenzofuranyl, pyr- rolinyl, pyrazolinyl, indolinyl, oxazolidinyl, oxazolinyl, oxazepinyl and the like.
• heteroarylthio represents a group -S-heteroaryl wherein heteroaryl is as defined above.
• heteroaryloxy represents a group -O-heteroaryl wherein heteroaryl is as defined above.
• heteroarylamino as used herein represents a group -NH-heteroaryl wherein heteroaryl is as defined above.
• Aryl-C ⁇ - 6 -alkyl means C ⁇ -alkyl as defined above, substituted by an aryl or heteroaryl as defined above, for example:
• a functional group which can be converted to hydrogen in vivo is intended to include any group which upon administering the present compounds to the subjects in need thereof can be converted to hydrogen eg enzymatically or by the acidic environment in the stomach.
• Non-limiting examples of such groups are acyl, carbamoyl, monoalkylated carbamoyl, dialkylated carbamoyl, alkoxycarbonyl, alkoxyalkyl groups and the like such as d-e-alkylcarbonyl, aroyl, d-e-alkylcarbamoyl, di-C ⁇ e-alkylcarbamoyl, C- ⁇ - 6 -alkoxy- carbonyl and d- 6 -alkoxy-d-e-alkyl.
• the phrase "diseases and disorders related to the histamine H3 receptor" is intended to include any disease or disorder in which an effect, either antagonistic or agonistic, on the histamine H3 receptor is beneficial.
• the present invention relates to a compound of the general formula (I):
• R 1 is hydrogen or a functional group, which can be converted to hydrogen in vivo
• R 2 is hydrogen, C ⁇ - 6 -alkyl, d- 6 -alkoxy, d-e-alkylthio, halogen, cyano, trifluoromethyl, hydroxy, thiol or -NR 5 R 6 , wherein R 5 and R 6 independently are hydrogen or d-e-alkyl,
• n 0, 1 or 2
• n 1 , 2, 3 or 4
• p 0, 1 , 2 or 3,
• q 0, 1 , 2 or 3
• aryl-C ⁇ - 6 -alkyl aryl or aryloxy, which are optionally substituted with one or more substituents selected from
• R 1 , R 2 , R 3 and R 4 are hydrogen, X is a valence bond, m is 0, n is 3, p is 0 and Y is -S-, -(CH 2 ) q -V must not be -(CH 2 ) 2 -NH 2 ,
• the present invention relates to a compound of the general formula (I"):
• R 1 is hydrogen or a functional group, which can be converted to hydrogen in vivo
• R 2 is hydrogen, d- 6 -alkyl, d- 6 -alkoxy, C ⁇ - 6 -alkylthio, halogen, cyano, trifluoromethyl, hydroxy, thiol or -NR 5 R 6 , wherein R 5 and R 6 independently are hydrogen or d- 6 -alkyl,
• n 0, 1 or 2
• n 1 , 2, 3 or 4,
• p 0, 1 , 2 or 3,
• q 0, 1 , 2 or 3
• Ci-e-alkyl C 2 . 6 -alkenyl or C 2 - 6 -alkynyl
• R 1 , R 2 , R 3 and R 4 are hydrogen, X is a valence bond, m is 0, n is 3, p is 0 and Y is -S-, -(CH 2 ) q -V must not be -(CH 2 ) 2 -NH 2 ,
• R 1 is hydrogen
• R 2 is also preferably hydrogen.
• R 3 and R 4 are both hydrogen.
• n is 2.
• n is 3 or 4.
• n is 1.
• p is preferably 1 or 2.
• X is preferably a valence bond or -O-.
• Y is preferably -O-.
• V is selected from heteroaryl, aryl and aryl annulated with C 3 - 8 - heterocyclyl, which are optionally substituted as defined for formula (I).
• V is selected from aryl and aryl annulated with C 3 . 8 -heterocyclyl, which are op- tionally substituted as defined as for formula (I).
• V is selected from phenyl, naphthyl, tetrahydronaphthyl and 3,4-methylene- . dioxyphenyl, which are optionally substituted as defined for formula (I).
• V is selected from phenyl and naphthyl, which are optionally substituted as defined for formula (I).
• V is unsubstituted or substituted with one or two substituents independently selected from
• aryl-C ⁇ - 6 -alkyl aryl and aryloxy, which are optionally substituted as defined for formula (I) above.
• V is unsubstituted or substituted with one or two substituents independently selected from
• V is unsubstituted or substituted with one or two substituents independently selected from phenyl, phenoxy and trifluoromethyl.
• V is C - 6 -alkyl or C 2 . 6 -alkenyl, which is optionally substituted as defined for formula (I) above.
• V is d-e-alkyl or C 2 -e-alkenyl, which is substituted with aryl, which is optionally substituted as defined for formula (I) above.
• V is C ⁇ - 6 -alkyl or C 2 -e-alkenyl, which is substituted with phenyl, which is optionally substituted as defined for formula (I) above.
• the invention relates to a compound of the general formula (le):
• R 1 , R 2 , R 3 , R 4 and V are as defined for formula (I) or in any of the above preferred embodiments.
• the invention relates to a compound of the general formula (If):
• R 1 , R 2 , R 3 , R 4 and V are as defined are as defined for formula (I) or in any of the above preferred embodiments.
• the invention relates to a compound of the general for- mula (Ih):
• R 1 , R 2 , R 3 , R 4 and V are as defined are as defined for formula (I) or in any of the above preferred embodiments.
• the invention relates to a compound of the general formula (li):
• R 1 , R 2 , R 3 , R 4 and V are as defined are as defined for formula (I) or in any of the above preferred embodiments.
• the compounds of the present invention may have one or more asymmetric centres and it is intended that any optical isomers, as separated, pure or partially purified optical isomers or racemic mixtures thereof are included within the scope of the invention. Furthermore, when a double bond or a fully or partially saturated ring system is present in the molecule geometric isomers may be formed. It is intended that any geometric isomers, as separated, pure or partially purified geometric isomers or mixtures thereof are included within the scope of the invention. Likewise, molecules having a bond with restricted rotation may form geometric isomers. These are also intended to be included within the scope of the present invention.
• the present invention also encompasses pharmaceutically acceptable salts of the present compounds.
• Such salts include pharmaceutically acceptable acid addition salts, pharmaceutically acceptable metal salts, ammonium and alkylated ammonium salts.
• Acid addition salts include salts of inorganic acids as well as organic acids. Representative examples of suitable inorganic acids include hydrochloric, hydrobromic, hydroiodic, phosphoric, sulfuric, nitric acids and the like.
• suitable organic acids include formic, acetic, trichloroacetic, trifluoroacetic, propionic, benzoic, cinnamic, citric, fumaric, gly- colic, lactic, maleic, malic, malonic, mandelic, oxalic, picric, pyruvic, salicylic, succinic, methanesulfonic, ethanesulfonic, tartaric, ascorbic, pamoic, bismethylene salicylic, ethanedi- sulfonic, gluconic, citraconic, aspartic, stearic, palmitic, EDTA, glycolic, p-aminobenzoic, glu- tamic, benzenesulfonic, p-toluenesulfonic acids and the like.
• pharma- ceutically acceptable inorganic or organic acid addition salts include the pharmaceutically acceptable salts listed in J. Pharm. Sci. 1977, 66, 2, which is incorporated herein by reference.
• metal salts include lithium, sodium, potassium, magnesium salts and the like.
• ammonium and alkylated ammonium salts include ammonium, methylam- monium, dimethylammonium, trimethylammonium, ethylammonium, hydroxyethylammonium, diethylammonium, butylammonium, tetramethylammonium salts and the like.
• Also intended as pharmaceutically acceptable acid addition salts are the hydrates, which the present compounds are able to form.
• the acid addition salts may be obtained as the direct products of compound synthesis.
• the free base may be dissolved in a suitable solvent containing the ap-litiste acid, and the salt isolated by evaporating the solvent or otherwise separating the salt and solvent.
• the compounds of the present invention may form solvates with standard low molecular weight solvents using methods well known to the person skilled in the art. Such solvates are also contemplated as being within the scope of the present invention.
• the invention also encompasses prodrugs of the present compounds, which on administration undergo chemical conversion by metabolic processes before becoming active pharmacological substances. In general, such prodrugs will be functional derivatives of the present compounds, which are readily convertible in vivo into the required compound of the formula (I). Conventional procedures for the selection and preparation of suitable prodrug derivatives are described, for example, in "Design of Prodrugs", ed. H. Bundgaard, Elsevier, 1985.
• the invention also encompasses active metabolites of the present compounds.
• the compounds of the present invention interact with the histamine H3 receptor and are accordingly useful for the treatment and/or prevention of a wide variety of conditions and disorders in which histamine H3 receptor interactions are beneficial.
• the present invention relates to compounds of the general formula (I'):
• R 1 is hydrogen or a functional group, which can be converted to hydrogen in vivo
• n 0, 1 or 2
• n 1 , 2, 3 or 4,
• p 0, 1 , 2 or 3,
• q 0, 1 , 2 or 3
• R 10 and R 11 independently are hydrogen or d-e-alkyl, ⁇ aryl-C - 6 -alkyl, aryl or aryloxy, which are optionally substituted with one or more substituents selected from
• the invention also relates to pharmaceutical compositions comprising, as an active ingredient, at least one compound of the formula (I') or any optical or geometric isomer or tautomeric form thereof including mixtures of these or a pharmaceutically acceptable salt thereof together with one or more pharmaceutically acceptable carriers or diluents.
• the invention relates to the use of a compound of the general formula (I') or any optical or geometric isomer or tautomeric form thereof including mixtures of these or a pharmaceutically acceptable salt thereof for the preparation of a pharmaceutical composition for the treatment and/or prevention of diseases and disorders related to the histamine H3 receptor.
• the invention relates to a method for the treatment and/or prevention of diseases and disorders related to the histamine H3 receptor the method com- prising administering to a subject in need thereof an effective amount of a compound of the formula (I') or any optical or geometric isomer or tautomeric form thereof including mixtures of these or a pharmaceutically acceptable salt thereof or a pharmaceutical composition comprising the same.
• the invention relates to compounds with histamine H3 receptor antagonistic activity or inverse agonistic activity which may accordingly be useful in the treatment of a wide range of conditions and disorders in which histamine H3 receptor blockade is beneficial.
• the invention relates to compounds with histamine H3 receptor agonistic activity and which may accordingly be useful in the treatment of a wide range of conditions and disorders in which histamine H3 receptor activation is beneficial.
• the present compounds are used for the preparation of a pharmaceutical composition for the reduction of weight.
• the present compounds are used for the preparation of a pharmaceutical composition for the treatment and/or prevention of overweight or obesity.
• the present compounds are used for the preparation of a pharmaceutical composition for the suppression of appetite or satiety induction.
• the present compounds are used for the preparation of a pharmaceutical composition for the prevention and/or treatment of disorders and diseases related to overweight or obesity such as atherosclerosis, hypertension, IGT (impaired glucose tolerance), diabetes, especially Type 2 diabetes (NIDDM (non- insulin dependent diabetes mellitus)), dyslipidaemia, coronary heart disease, gallbladder dis- ease, osteoarthritis and various types of cancer such as endometrial, breast, prostate and colon cancers.
• disorders and diseases related to overweight or obesity such as atherosclerosis, hypertension, IGT (impaired glucose tolerance), diabetes, especially Type 2 diabetes (NIDDM (non- insulin dependent diabetes mellitus)), dyslipidaemia, coronary heart disease, gallbladder dis- ease, osteoarthritis and various types of cancer such as endometrial, breast, prostate and colon cancers.
• the present compounds are used for the preparation of a pharmaceutical composition for the prevention and/or treatment of eating disorders such as bulimia and binge eating. In a further preferred embodiment of the invention the present compounds are used for the preparation of a pharmaceutical composition for the treatment and/or prevention of IGT.
• the present compounds are used for the preparation of a pharmaceutical composition for the treatment and/or prevention of Type 2 diabetes.
• the present compounds are used for the preparation of a pharmaceutical composition for the delaying or prevention of the progression from IGT to Type 2 diabetes.
• the present compounds are used for the preparation of a pharmaceutical composition for the delaying or prevention of the progression from non-insulin requiring Type 2 diabetes to insulin requiring Type 2 diabetes.
• the compounds of the present invention may also be used for the treatment of airway disorders such as asthma, as anti-diarrhoeals and for the modulation of gastric acid secretion. Furthermore, the compounds of the present invention may be used for the treatment of diseases associated with the regulation of sleep and wakefulness and for the treatment of narcolepsy and attention deficit disorders.
• the compounds of the invention may be used as CNS stimulants or as sedatives.
• the present compounds may also be used for the treatment of conditions associated with epilepsy.
• the present compounds may be used for the treatment of motion sickness and vertigo.
• they may be useful as regulators of hypothalamo- hypophyseal secretion, antidepressants, modulators of cerebral circulation, and in the treatment of irritable bowel syndrome.
• the compounds of the present invention may be used for the treatment of dementia and Alzheimer's disease.
• the compounds of the present invention may also be useful for the treatment of allergic rhinitis, ulcer or anorexia.
• the compounds of the present invention may furthermore be useful for the treat- ment of migraine, see R.L. McLeod et al., The Journal of Pharmacology and Experimental Therapeutics 287 (1998), 43-50, and for the treatment of myocardial infarction, see C.J. Mackins and R. Levi, Expert Opinion on Investigational Drugs 9 (2000), 2537-2542.
• the present novel compounds may also interact with the vanilloid receptors, the serotonin receptors, and the adrenergic receptors and may be useful for the treatment of dis- eases associated with these receptors.
• the compounds of the present invention may be vanilloid receptor agonists, and thus be useful for the treatment of obesity by enhancement of the metabolic rate and energy expenditure.
• the compounds of the present invention may be useful for the treatment of pain or neurogenic inflammation or inflammatory painful conditions.
• the present compounds are used for the preparation of a pharmaceutical composition for the treatment and/or prevention of diseases and disorders related to the vanilloid receptor, such as for the treatment and/or prevention of pain, neurogenic inflammation or obesity.
• the present compounds may interact with the 5-HT3 receptor (sero- tonin-3-receptor), the compounds of the present invention may be useful as antiemetics, in particular the chemotherapy-induced emesis.
• 5-HT3 antagonists include treatment of central nervous system disorders such as anxiety, schizophrenia, drug abuse and withdrawal symptoms, and pathological and age-associated amnesia.
• the present compounds are used for the preparation of a pharmaceutical composition for the treatment and/or prevention of diseases and disorders related to the serotonin-3 receptor (5-HT3), such as for the treatment of emesis.
• the present compounds may interact with the adrenergic alpha-2- receptor and thus be useful for the treatment of hypertension and of conditions associated with overexpression or hypersensitization of the adrenergic alpha-2 receptor, especially obesity, withdrawal symptoms to an adrenergic alpha-2 agonist, neurological disorders (especially orthostatic hypotension), multiple system atrophy, diabetes mellitus, benign prostatic hyperplasia or drug induced sensitization of the adrenergic alpha-2 receptor.
• the compounds of the present invention by virtue of their interaction with the adrenergic alpha-2 receptor, may be useful as sedatives and hypnotics (sleep inducing agents) or as stimulants.
• the present compounds are used for the preparation of a pharmaceutical composition for the treatment and/or prevention of diseases and disorders related to the alpha-2 adrenergic receptor, such as for use as a sleep inducing agent.
• the present compounds are combined with diet and/or exercise.
• the present compounds may be administered in combination with one or more further pharmacologically active substances in any suitable ra- tios.
• Such further active agents may be selected from antiobesity agents, antidiabetics, anti- hypertensive agents, agents for the treatment and/or prevention of complications resulting from or associated with diabetes and agents for the treatment and/or prevention of complications and disorders resulting from or associated with obesity.
• the present compounds may be adminis- tered in combination with one or more antiobesity agents or appetite regulating agents.
• agents may be selected from the group consisting of CART (cocaine amphetamine regulated transcript) agonists, NPY (neuropeptide Y) antagonists, MC4 (melano- cortin 4) agonists, MC3 (melanocortin 3) agonists, orexin antagonists, TNF (tumor necrosis factor) agonists, CRF (corticotropin releasing factor) agonists, CRF BP (corticotropin releas- ing factor binding protein) antagonists, urocortin agonists, ⁇ 3 agonists, MSH (melanocyte- stimulating hormone) agonists, MCH (melanocyte-concentrating hormone) antagonists, CCK (cholecystokinin) agonists, serotonin re-uptake inhibitors, serotonin and noradren
• CART cocaine amp
• the antiobesity agent is leptin.
• the antiobesity agent is dexamphetamine or amphetamine.
• the antiobesity agent is fenfluramine or dexfenfluramine.
• the antiobesity agent is sibutramine. In a further embodiment the antiobesity agent is orlistat.
• the antiobesity agent is mazindol or phentermine.
• the antiobesity agent is phendimetrazine, diethylpropion, fluoxetine, bupropion, topiramate or ecopipam.
• Suitable antidiabetics comprise insulin, GLP-1 (glucagon like peptide-1 ) derivatives such as those disclosed in WO 98/08871 to Novo Nordisk A/S, which is incorporated herein by reference as well as orally active hypoglycaemic agents.
• the orally active hypoglycaemic agents preferably comprise imidazolines, sulphony- lureas, biguanides, meglitinides, oxadiazolidinediones, thiazolidinediones, insulin sensitizers, ⁇ -glucosidase inhibitors, agents acting on the ATP-dependent potassium channel of the ⁇ - cells eg potassium channel openers such as those disclosed in WO 97/26265, WO 99/03861 and WO 00/37474 (Novo Nordisk A/S) which are incorporated herein by reference, nateglinide, glucagon antagonists such as those disclosed in WO 99/01423 and WO 00/39088 (Novo Nordisk A/S and Agouron Pharmaceuticals, Inc.), which are incorporated herein by reference, GLP-1 agonists such as those disclosed in WO 00/42026 (Novo Nordisk A/S and Agouron Pharmaceuticals, Inc.), which are incorporated herein by reference, D
• the present compounds are administered in combination with insulin.
• the present compounds are administered in combination with a sulphonylurea eg tolbutamide, glibenclamide, glipizide, glimepiride, gli- malariaide or glyburide.
• a sulphonylurea eg tolbutamide, glibenclamide, glipizide, glimepiride, gli- malariaide or glyburide.
• the present compounds are administered in combination with a biguanide eg metformin.
• the present compounds are adminis- tered in combination with a meglitinide eg repaglinide or senaglinide.
• the present compounds are administered in combination with a thiazolidinedione eg troglitazone, ciglitazone, pioglitazone, rosigli- tazone or the compounds disclosed in WO 97/41097, WO 97/41119, WO 97/41120, WO 00/41121 and WO 98/45292 (Dr. Reddy's Research Foundation), which are incorporated herein by reference.
• a thiazolidinedione eg troglitazone, ciglitazone, pioglitazone, rosigli- tazone or the compounds disclosed in WO 97/41097, WO 97/41119, WO 97/41120, WO 00/41121 and WO 98/45292 (Dr. Reddy's Research Foundation), which are incorporated herein by reference.
• the present compounds may be administered in combination with an insulin sensitizer eg such as those disclosed in WO 99/19313, WO 00/50414, WO 00/63191 , WO 00/63192, WO 00/63193 (Dr. Reddy's Research Foundation) and WO 00/23425, WO 00/23415, WO 00/23451 , WO 00/23445, WO 00/23417, WO 00/23416, WO 00/63153, WO 00/63196, WO 00/63209, WO 00/63190 and WO 00/63189 (Novo Nordisk A/S), which are incorporated herein by reference.
• an insulin sensitizer eg such as those disclosed in WO 99/19313, WO 00/50414, WO 00/63191 , WO 00/63192, WO 00/63193 (Dr. Reddy's Research Foundation) and WO 00/23425, WO 00/2
• the present compounds are administered in combination with an ⁇ -glucosidase inhibitor eg miglitol or acarbose.
• an ⁇ -glucosidase inhibitor eg miglitol or acarbose.
• the present compounds are administered in combination with an agent acting on the ATP-dependent potassium channel of the ⁇ -cells eg tolbutamide, glibenclamide, glipizide, glicazide or repaglinide.
• an agent acting on the ATP-dependent potassium channel of the ⁇ -cells eg tolbutamide, glibenclamide, glipizide, glicazide or repaglinide.
• the present compounds may be administered in combination with nateglinide.
• the present compounds are administered in combination with an antilipidemic agent eg cholestyramine, colestipol, clofibrate, gemfibrozil, Iovastatin, pravastatin, simvastatin, probucol or dextrothyroxine.
• an antilipidemic agent eg cholestyramine, colestipol, clofibrate, gemfibrozil, Iovastatin, pravastatin, simvastatin, probucol or dextrothyroxine.
• the present compounds are administered in com- bination with more than one of the above-mentioned compounds eg in combination with met- formin and a sulphonylurea such as glyburide; a sulphonylurea and acarbose; nateglinide and metformin; acarbose and meformin; a sulfonylurea, metformin and troglitazone; insulin and a sulfonylurea; insulin and metformin; insulin, metformin and a sulfonylurea; insulin and troglitazone; insulin and Iovastatin; etc.
• a sulphonylurea such as glyburide
• a sulphonylurea and acarbose such as glyburide
• a sulphonylurea and acarbose such as glyburide
• the present compounds may be administered in combination with one or more antihypertensive agents.
• antihypertensive agents are ⁇ -blockers such as alprenolol, atenolol, timolol, pindolol, propranolol and metoprolol, ACE (angiotensin converting enzyme) inhibitors such as benazepril, captopril, enalapril, fosinopril, lisinopril, quinapril and ramipril, calcium channel blockers such as nifedipine, felodipine, nicardipine, isradipine, nimodipine, diltiazem and verapamil, and ⁇ -blockers such as doxazosin, urapidil, prazosin and terazosin. Further reference can be made to Remington: The Science and Practice of Pharmacy, 19 th Edition, Gennaro, Ed., Mack Publishing
• the compounds of the invention may be administered alone or in combination with pharmaceutically acceptable carriers or excipients, in either single or multiple doses.
• the pharmaceutical compositions according to the invention may be formulated with pharmaceutically acceptable carriers or diluents as well as any other known adjuvants and excipients in accordance with conventional techniques such as those disclosed in Remington: The Science and Practice of Pharmacy, 19 th Edition, Gennaro, Ed., Mack Publishing Co., Easton, PA, 1995.
• compositions may be specifically formulated for administration by any suitable route such as the oral, rectal, nasal, pulmonary, topical (including buccal and sublingual), transdermal, intracistemal, intraperitoneal, vaginal and parenteral (including subcutaneous, intramuscular, intrathecal, intravenous and intradermal) route, the oral route being preferred. It will be appreciated that the preferred route will depend on the general condi- tion and age of the subject to be treated, the nature of the condition to be treated and the active ingredient chosen.
• compositions for oral administration include solid dosage forms such as capsules, tablets, dragees, pills, lozenges, powders and granules. Where appropri- ate, they can be prepared with coatings such as enteric coatings or they can be formulated so as to provide controlled release of the active ingredient such as sustained or prolonged release according to methods well known in the art.
• Liquid dosage forms for oral administration include solutions, emulsions, suspensions, syrups and elixirs.
• Pharmaceutical compositions for parenteral administration include sterile aqueous and non-aqueous injectable solutions, dispersions, suspensions or emulsions as well as sterile powders to be reconstituted in sterile injectable solutions or dispersions prior to use. Depot injectable formulations are also contemplated as being within the scope of the present invention.
• Other suitable administration forms include suppositories, sprays, ointments, cremes, gels, inhalants, dermal patches, implants etc.
• a typical oral dosage is in the range of from about 0.001 to about 100 mg/kg body weight per day, preferably from about 0.01 to about 50 mg/kg body weight per day, and more preferred from about 0.05 to about 10 mg/kg body weight per day administered in one or more dosages such as 1 to 3 dosages.
• the exact dosage will depend upon the frequency and mode of administration, the sex, age, weight and general condition of the subject treated, the nature and severity of the condition treated and any concomitant diseases to be treated and other factors evident to those skilled in the art.
• a typical unit dosage form for oral administration one or more times per day such as 1 to 3 times per day may contain of from 0.05 to about 1000 mg, preferably from about 0.1 to about 500 mg, and more preferred from about 0.5 mg to about 200 mg.
• parenteral routes such as intravenous, intrathecal, intramuscular and similar administration
• typically doses are in the order of about half the dose employed for oral administration.
• the compounds of this invention are generally utilized as the free substance or as a pharmaceutically acceptable salt thereof.
• One example is an acid addition salt of a compound having the utility of a free base.
• a compound of the formula (I) contains a free base such salts are prepared in a conventional manner by treating a solution or suspension of a free base of the formula (I) with a chemical equivalent of a pharmaceutically acceptable acid, for example, inorganic and organic acids. Representative examples are mentioned above.
• Physiologically acceptable salts of a compound with a hydroxy group include the an- ion of said compound in combination with a suitable cation such as sodium or ammonium ion.
• solutions of the novel compounds of the formula (I) in sterile aqueous solution, aqueous propylene glycol or sesame or peanut oil may be employed.
• aqueous solutions should be suitable buffered if necessary and the liquid diluent first rendered isotonic with sufficient saline or glucose.
• the aqueous solutions are par- ticulariy suitable for intravenous, intramuscular, subcutaneous and intraperitoneal administration.
• the sterile aqueous media employed are all readily available by standard techniques known to those skilled in the art.
• Suitable pharmaceutical carriers include inert solid diluents or fillers, sterile aqueous solution and various organic solvents.
• solid carriers are lactose, terra alba, su- erase, cyclodextrin, talc, gelatine, agar, pectin, acacia, magnesium stearate, stearic acid or lower alkyl ethers of cellulose.
• liquid carriers are syrup, peanut oil, olive oil, phospholipids, fatty acids, fatty acid amines, polyoxyethylene or water.
• the carrier or diluent may include any sustained release material known in the art, such as glyceryl monostearate or glyceryl distearate, alone or mixed with a wax.
• the pharmaceutical compo- sitions formed by combining the novel compounds of the formula (I) and the pharmaceutically acceptable carriers are then readily administered in a variety of dosage forms suitable for the disclosed routes of administration.
• the formulations may conveniently be presented in unit dosage form by methods known in the art of pharmacy.
• Formulations of the present invention suitable for oral administration may be pre- sented as discrete units such as capsules or tablets, each containing a predetermined amount of the active ingredient, and which may include a suitable excipient. These formulations may be in the form of powder or granules, as a solution or suspension in an aqueous or non-aqueous liquid, or as an oil-in-water or water-in-oil liquid emulsion.
• the preparation may be tabletted, placed in a hard gelatine capsule in powder or pellet form or it can be in the form of a troche or lozenge.
• the amount of solid carrier will vary widely but will usually be from about 25 mg to about 1 g.
• the preparation may be in the form of a syrup, emulsion, soft gelatine capsule or sterile injectable liquid such as an aqueous or non-aqueous liquid suspension or solution.
• a typical tablet which may be prepared by conventional tabletting techniques, may contain:
• Active compound (as free compound or salt thereof) 5.0 mg
• the pharmaceutical composition of the invention may comprise the compound of the formula (I) in combination with further pharmacologically active substances such as those described in the foregoing.
• the preparation of the compounds of this invention can be realised in many different ways.
• the starting imidazole derivatives of the formula (II) may be prepared according to procedures described in literature (see eg Croat. Chem. Acta. 1973, 45, 297. J. Am. Chem. Soc. 1976, 98, 984).
• the other reactants are either known compounds or compounds, which may be prepared in analogy with the preparation of similar known compounds.
• the alcohol of a compound of the formula (II) wherein m, n and p are as defined above and Pg represents a protecting group like eg triphenylmethyl (trityl) may be reacted with a compound of the formula (III) wherein V is as defined above.
• This reaction is carried out in the presence of a catalyst like eg triphenylphosphine and a base like eg diethyl azodicarboxylate in a suitable solvent like eg THF at a temperature of up to reflux for the solvent used for eg 1-200 hours. Removal of the trityl group of a compound of the formula (IV) is accomplished with dilute acid to give a compound of the formula la).
• a compound of the formula (V) wherein m, n and p are as defined above and Pg represents a protecting group like eg triphenylmethyl (trityl) and L represents a leaving group such as eg halogen or mesylate may be reacted with a compound of the formula (VI) wherein V and q are as defined above.
• This reaction is carried out in the presence of a strong base like eg sodium hydride in a suitable solvent like eg THF or DMF at a temperature of up to reflux for the solvent used for eg 1-200 hours. Removal of the trityl group from a compound of the formula (VII) is accomplished with dilute acid to give a compound of the formula (lb).
• a compound of the formula (Ic), wherein m, n, p, q, and V are defined as in formula (I), may be prepared as outlined below:
• a suitable base such as eg sodium hydride, potassium hydride, lithium hydride in a suitable solvent such as tetrahydrofuran, ether or a hydrocarbon such as toluene or hexane.
• halogenide such as a bromide as shown in formula (VIII) or a chloride as shown in formula (IX) or a synthetic equivalent thereof
• Catalysts such as tetrabutylammo- nium iodide may be added optionally.
• the reaction temperature might be between - 78 °C and the boiling point of the solvent applied for a reaction time of 0.5 - 200 hours. Removal of the protection group of a compound of the formula (X) may be accomplished as described in the literature (eg T. W. Greene, P. G. M.
• triphenylmethyl group may be removed with acid, such as acetic acid which may be diluted with another solvent such as water at a temperature between room temperature and 100 °C.
• a compound of the formula (Id), wherein n, m, p, q, X, Y, and V are defined as in formula (I), may be prepared as outlined below:
• a cyclic ketone, of formula (XI) in which n, m, p, q and V are defined as in formula (I), and L is a leaving group such as bromo, chloro, or methansulfonyl, may be warmed with DMF to a suitable temperature such as a temperature between 100 °C and 200 °C for a time between 0.5 and 200 hours.
• CDI carbonyldiimidazole
• DBU 1 ,8-diazabicyclo[5.4.0]undec-7-ene
• DCM dichloromethane, methylenechloride
• DIPEA diisopropylethylamine
• TFA trifluoroacetic acid
• THF tetrahydrofuran
• HibarTM RT 125-4 5.0 ⁇ m, 4.0 x 125 mm; gradient elution, 5% to 80% solvent B (0.1% TFA in acetonitrile) in solvent A (0.1% TFA in water) within 10 min, 2.0 ml/min, detection at 214 nm, temperature 35 °C.
• Step 1 4,5,6,7-Tetrahvdro-1H-benzimidazole-5-carboxylic acid, hydrochloride salt
• Step 2 1 (3)-Triphenylmethyl-4.5.6.7-tetrahydro-1H-benzimidazole-5-carboxylic acid methyl ester
• Triethylamine (9.5 ml, 69 mmol) was added to the mixture followed by portion wise addition of triphenylmethyl chloride (6.4 g, 23 mmol). The mixture was stirred overnight at room temperature and then filtered. The solvent was evaporated from the filtrate to give a residue, which was stirred with diethyl ether (200 ml). The mixture was filtered and the solvent was evaporated from the filtrate.
• Example 11 4-(4,5,6,7-Tetrahydro-1H-benzimidazol-5-ylmethoxy)benzoic acid ethyl ester, oxalic acid salt
• Step 1 1-Triphenylmethyl-4.5,6,7-tetrahydro-1H-benzimidazole-4-carboxylic acid ethyl ester
• Step 2 4-Hydroxymethyl-1 -triphenylmethyl-4,5,6,7-tetrahydro-1 H-benzimidazole
• Step 3 To a solution of 4-hydroxymethyl-1 -triphenylmethyl-4,5,6,7-tetrahydro-1 H-benzimidazole (0.21 g, 0.54 mmol) in THF (3 ml) were added triphenylphosphine (0.20 g, 0.75 mmol), 2-ethylphenol (0.073 ml, 0.60 mmol), and DEAD (0.118 ml, 0.75 mmol). The mixture was shaken at 20 °C overnight, concentrated under reduced pressure, and the residue was purified by column chromatography (silica gel, gradient elution with heptane/ethyl acetate 9:1 to 1 :1).
• the resulting product (100 mg) was dissolved in 5 ml of a mixture of glacial acetic acid and H 2 O (9:1), and kept at 70 °C for 20 hours. The mixture was concentrated and the residue was re-dissolved in ethyl acetate. To this solution was added a solution of oxalic acid (45 mg) in ethyl acetate (5 ml), whereby 23 mg (12%) of the title compound precipitated as a colourless solid.
• Step 1 l-Triphenylmethyl-I . ⁇ .e . ⁇ -hexahvdrocvcloheptaimidazole ⁇ -carboxylic acid methyl ester
• Example 28 4-(2,4-Difluorophenoxymethyl)-1 ,4,5,6,7,8-hexahydrocycloheptaimidazole, hydrochloride salt
• Step 1 3-(4,5,6,7-Tetrahvdro-1H-benzimidazol-5-vDpropionitrile hydrochloride
• Step 3 3-(1 (3)-Triphenylmethyl-4,5,6 -tetrahydro-1 H-benzimidazol-5-yl)-1 -propanol
• the aqueous phase was extracted with ethyl acetate (3 x 100 ml).
• the combined organic layers were washed with a saturated aqueous solution of sodium hydrogencarbonate (200 ml) and dried over magnesium sulphate.
• the solvent was removed in vacuo.
• the crude product was purified by flash chromatography on silica (200 g), using ethyl acetate/heptane 1 :1 as eluent, to give. .0 g of ((4-(methoxy)cyclohex-3-enyloxy)methyl)benzene, which was slightly contaminated with 4-benzyloxycyclohexanone.
• the title compound was transferred into its hydrochloride salt, by dissolving in ethyl acetate (100 ml) and addition of hydrogen chloride in ethyl acetate (3.0
• Step 1 5-((Biphenyl-2-yl)methoxymethyl)-1-(triphenylmethyl)-4,5.6.7-tetrahvdro-1 H-benzimidazole and 5-((Biphenyl-2-yl)methoxymethyl)-3-(triphenylmethv ⁇ -4.5,6.7-tetrahvdro-3H- benzimidazole.
• the crude product was purified by flash chromatography on silica, using first ethyl acetate/heptane (1 :1) and then DCM/methanol/25% aqueous ammonia (first: 200:10:1 , then: 100:10:1 ) as eluent, to give 0.24 g of a mixture of 5-((biphenyl-2-yl)methoxymethyl)-1-(triphenylmethyl)-4,5,6,7-tetra- hydro-1 H-benzimidazole and 5-((biphenyl-2-yl)methoxymethyl)-3-(triphenylmethyl)-4,5,6,7- tetrahydro-3H-benzimidazole.
• Step 1 5-((1-Naphthyl)methoxymethv ⁇ -1-(triphenylmethyl)-4,5,6,7-tetrahydrobenzimidazole and 5-((1-naphthyl)methoxymethv ⁇ -3-(triphenylmethyl)-4,5.6,7-tetrahvdrobenzimidazole
• the crude product was purified by flash chromatography on silica (90 g), using a gradient of ethyl acetate/heptane (from 50:50 to 100:0) as eluent, to give 400 mg of a mixture of 5-((1-naphthyl)methoxymethyl)-1- (triphenylmethyl)-4,5,6,7-tetrahydrobenzimidazole and 5-((1 -naphthyl)methoxymethyl)-3- (triphenylmethyl)-4,5,6,7-tetrahydrobenzimidazole.
• Step 1 5-(4-Trifluoromethylbenzyloxymethyl)-1-(triphenylmethyl)-4,5.6,7-tetrahydro- benzimidazole and 5-(4-Trifluoromethylbenzyloxymethyl)-3-(triphenylmethvD-4, 5,6,7- tetrahvdrobenzimidazole
• reaction mixture was stirred for 16 hours at room temp. Water (100 ml) was added carefully. The mixture was extracted with ethyl acetate (3 x 80 ml). The combined organic layers were washed with a saturated aqueous solution of sodium hydrogencarbonate (100 ml) and dried over magnesium sulphate. The solvent was removed in vacuo.
• the crude product was purified by flash chromatography on silica (90 g), using a gradient of ethyl acetate/heptane (from 50:50 to 100:0) as eluent, to give 1.65 g of a mixture of 5-(4-trifluoromethylbenzyloxymethyl)- 1 -(triphenylmethyl)-4,5,6,7-tetrahydrobenzimidazole and 5-(4-trifluoromethylbenzyl- oxymethyl)-3-(triphenylmethyl)-4,5,6,7-tetrahydrobenzimidazole.
• Step 1 5-((3-(TrifluoromethvDbenzyloxy)methyl)-1 -triphenylmethyl-4,5,6,7-tetrahvdro-1 H- benzimidazole and 5-((3-(trifluoromethyl)benzyloxy)methv0-3-triphenylmethyl-4,5,6,7- tetrahvdro-1 H-benzimidazole
• the crude product was purified by flash chromatography on silica (90 g), using a gradient of ethyl acetate/heptane (from 50:50 to 100:0) as eluent, to give 1.50 g of a mixture of 5-((3-(trifluoromethyl)benzyloxy)methyl)-1-triphenylmethyl-4,5,6,7-tetrahydro-1H- benzimidazole and 5-((3-(trifluoromethyl)benzyloxy)methyl)-3-triphenylmethyl-4,5,6,7- tetrahydro-3H-benzimidazole.
• Phosphorus tribromide (5.5 ml, 57.1 mmol) was added dropwise over a period of 10 min to a solution of 2-phenoxybenzylalcohol (4.08 g, 20.4 mmol) in dioxane (165 ml). The reaction mixture was stirred for 40 min. Another portion of phosphorus tribromide (5.5 ml, 57.1 mmol) was added. The reaction mixture was stirred for 56 hours at room temperature. It was cooled to 0 °C. Water (45 ml) was added carefully. The reaction mixture was stirred for 30 min at room temperature. The mixture was extracted with ethyl acetate (100 ml).
• the ethyl acetate phase was washed with 1 N hydrochloric acid.
• the combined aqueous layers were extracted with ethyl acetate.
• the combined organic layers were washed with a satu- rated aqueous soludtion of sodium hydrogen carbonate (100 ml) and dried over magnesium sulphate.
• the solvent was removed in vacuo.
• the crude product was purified by flash chromatography on silica (90 g), using ethyl acetate/heptane (1 :3) as eluent, to give 4.80 g of 2- phenoxybenzyl bromide.
• Step 4 5-((2-Phenoxybenzyloxy)methyl)-1 -triphenylmethyl-4.5,6,7-tetrahvdro-1 H- benzimidazole and 5-((2-phenoxybenzyloxy)methyl)-3-triphenylmethyl-4,5,6,7-tetrahvdro-3H- benzimidazole
• the crude prod- uct was purified by flash chromatography on silica (90 g), using ethyl acetate/heptane (gradient from 50:50 to 100:0) as eluent, to give 1.21 g of 5-((2-phenoxybenzyloxy)methyl)-1- triphenylmethyl-4,5,6,7-tetrahydro-1 H-benzimidazole and 5-((2-phenoxybenzyloxy)methyl)-3- triphenylmethyl-4,5,6,7-tetrahydro-3H-benzimidazole.
• the solvent was removed in vacuo.
• the crude product was purified by flash chromatography on silica (40 g), using DCM/methanol/25% aqueous ammonia (first: 100:10:1 , then 100:20:2) as eluent to give 74 mg of the title compound.
• the title compound was transferred into its hydrochloride salt by lyophiliza- tion of its solution in 0.1 N hydrochloric acid (40 ml).
• the aqueous phase was extracted with ethyl acetate (100 ml).
• the combined organic layers were washed with a saturated aqueous solution of sodium hydrogencarbonate (100 ml) and dried over magnesium sulphate.
• the solvent was removed in vacuo.
• the crude product was purified by flash chromatography on silica (200 g), using ethyl acetate/heptane (1 :1 ) as eluent, to give 2.36 g of 2-phenoxymethyl-2,3-dihydropyran-4-one.
• Step 1 Allyloxy-1 -triphenylmethyl-4,5,6 -tetrahvdro-1 H-benzimidazole and allyloxy-3- triphenylmethyl-4.5.6.7-tetrahvdro-3H-benzimidazole
• the reaction mixture was stirred for 16 hours, while it was warming up to room temperature. It was cooled to 0 °C. Water (1 ml) was added carefully dropwise. A 1 N aqueous solution of sodium hydroxide (30 ml) was added. The mixture was extracted with tert- butyl methyl ether (3 x 30 ml). The combined organic layers were dried over magnesium sulphate. The solvent was removed.
• the crude product was purified by flash chromatography on silica (80 g), using ethyl acetate as eluent, to give 290 mg of a mixture of allyloxy-1- triphenylmethyl-4,5,6,7-tetrahydro-1 H-benzimidazole and allyloxy-3-triphenylmethyl-4,5,6,7- tetrahydro-1 H-benzimidazole.
• the title compound was transferred into its hydrochloride salt: The title compound (58 mg) was dissolved in ethyl acetate (2 ml). A 3.5 M solution of hydro- gen chloride in ethyl acetate (1 ml) was added. The solvent was removed in vacuo. The residue was dissolved in ethyl acetate (1 ml) and heptane (3 ml). The solvent was removed. The residue was dried in vacuo.
• Step 1 5-(((2-Naphthvnmethoxy)methvn-1-triphenylmethyl-4.5.6.7-tetrahvdro-1H- benzimidazole and 5-(((2-naphthyl)methoxy)methv ⁇ -3-triphenylmethyl-4,5,6.7-tetrahydro-3H- benzimidazole
• the crude product was purified by flash chromatography on silica (90 g), using a gradient of ethyl acetate/heptane (from 50:50 to 100:0), to give 590 mg of a mixture of 5-(((2-naphthyl)methoxy)methyl)-1 -triphenylmethyl-4,5,6,7-tetrahydro-1 H-benzimidazole and 5-(((2-naphthyl)methoxy)methyl)-3-triphenylmethyl-4,5,6,7-tetrahydro-3H-benzimidazole.
• the title compound was transferred into its hydrochloride salt, by dissolving it in ethyl acetate (30 ml) and addition of a 3.5 M solution of hydrogen chloride in ethyl acetate (3 ml). The precipitation was collected and dried in vacuo.
• Step 1 5-(2-Chloro-5-trifluoromethylbenzyloxymethyl)-1-triphenylmethyl-4.5,6,7-tetrahydro- 1 H-benzimidazole and 5-(2-chloro-5-trifluoromethylbenzyloxymethyl)-3-triphenylmethyl-
• the crude product was purified by flash chromatography on silica (40 g), using a gradient of ethyl acetate/heptane (from 50:50 to 100:0) as eluent, to give 560 mg of a mixture of 5-(2-chloro-5-trifluoromethylbenzyloxymethyl)-1 ⁇ triphenylmethyl-4,5,6,7-tetrahydro-1 H- benzimidazole and 5-(2-chloro-5-trifluoromethylbenzyloxymethyl)-3-triphenylmethyl-4,5,6,7- tetrahydro-3H-benzimidazole.
• Step 1 (£)-5-(3-Phenylallyloxymethvn-1-triphenylmethyl-4.5.6.7-tetrahvdro-1H- benzimidazole and (£)-5-(3-phenylallyloxymethv ⁇ -3-triphenylmethyl-4,5.6,7-tetrahvdro-3H- benzimidazole
• the crude product was purified by flash chromatography on silica (90 g), using a gradient of ethyl acetate/heptane (1 :2 to pure ethyl acetate) as eluent, to give 501 mg of a mixture of (E)-5-(3-phenylallyloxymethyl)-1-triphenylmethyl-4,5,6,7-tetrahydro-1 H-benzimidazole and (E)-5-(3-phenylallyloxymethyl)-3-triphenylmethyl-4,5,6,7-tetrahydro-3H-benz- imidazole.
• Step 1 5-((Biphenyl-4-yl)methoxymethyl)-1-triphenylmethyl-4,5.6,7-tetrahydro-1H- benzimidazole and 5-((biphenyl-4-yl)methoxymethv ⁇ -3-triphenylmethyl-4,5,6,7-tetrahydro- 3H-benzimidazole
• the crude product was purified by flash chromatography on silica (90 g), using a gradient of ethyl acetate/heptane (from 1 :2 to 1.0) as eluent, to give 440 mg of a mixture of 5-((biphenyl-4-yl)- methoxymethyl)-1 -triphenylmethyl-4,5,6,7-tetrahydro-1 H-benzimidazole and 5-((biphenyl-4- yl)methoxymethyl)-3-triphenylmethyl-4,5,6,7-tetrahydro-3H-benzimidazole.
• Step 1 (2-Chloro-3-(trifluoromethv0phenv0methanol
• Phosphorus tribromide (11.75 ml, 125 mmol) was added to a solution of crude (2- chloro-3-(trifluoromethyl)phenyl)methanol (9.37 g, 44.5 mmol) in dioxane (200 ml). The reac- tion mixture was stirred for 1 hour. Another portion of phosphorus tribromide (11.75 ml, 125 mmol) was added. The reaction mixture was stirred for 16 hours at room temperature. It was diluted with ethyl acetate (200 ml). A 10% aqueous solution of sodium hydrogensulphate (200 ml) was added, while cooling with a water bath. The phases were separated.
• Step 3 5-(2-Chloro-3-(trifluoromethv0benzyloxymethyl)-1 -triphenylmethyl-4,5,6,7-tetrahvdro- 1 H-benzimidazole and 5-(2-chloro-3-(trifluoromethv0benzyloxymethyl)-3-triphenylmethyl- 4,5,6,7-tetrahvdro-3H-benzimidazole
• Tetrabutylammonium iodide (12 mg, 0.03 mmol) was added. The reaction mixture was stirred for 56 hours, while the reaction mixture was warming up to room tern- perature. It was diluted with THF (10 ml). Water (10 ml) was added carefully dropwise, while the reaction mixture was cooled with a water bath. It was diluted with ethyl acetate (20 ml) and washed with a 10% aqueous solution of sodium hydrogensulphate (20 ml). The aqueous solution was extracted with ethyl acetate (20 ml). The combined organic layers were washed with a saturated aqueous solution of sodium hydrogencarbonate (100 ml) and dried over magnesium sulphate.
• Step 1 3-(((1-Triphenylmethyl-4,5,6,7-tetrahvdro-1H-benzimidazol-5- yl)methoxy)methyl)benzonitrile and 3-(((3-triphenylmethyl-4.5.6.7-tetrahydro-3H- benzimidazol-5-vDmethoxy)methyl)benzonitrile
• the crude product was purified by flash chromatography on silica (90 g), using a gradient of ethyl acetate/heptane (from 1 :1 to 1:0) as eluent, to give 0.18 g of a mixture of 3-(((1 -triphenylmethyl-4,5,6,7-tetrahydro-1 H-benzimidazol-5-yl)methoxy)methyl)- benzonitrile and 3-(((3-triphenylmethyl-4,5,6,7-tetrahydro-3H-benzimidazol-5-yl)methoxy)- methyl)benzonitrile.
• the title compound was transformed into its hydrochloride salt, by dissolving it in ethyl acetate (10 ml) and addition of a 3.5 M solution of hydrogen chloride in ethyl acetate (1.5 ml, 5.25 mmol). The solvent was removed in vacuo. The residue was crystallized form acetone.
• Phosphorus tribromide (17.6 ml, 186 mmol) was added dropwise to a solution of 4- phenyl-1-butanol (10.0 g, 66.6 mmol) in dioxane (200 ml). The reaction mixture was stirred for 1 hour at room temperature. Another portion of phosphorus tribromide (17.6 ml, 186 mmol) was added dropwise. The reaction mixture was stirred for 16 hours at room tempera- ture. It was cooled to 0 °C. Water (130 ml) was added dropwise. The reaction mixture was stirred for 30 min at room temperature.
• Step 2 5-(4-Phenylbutoxymethyl)-1 -triphenylmethyl-4,5,6,7-tetrahvdro-1 H-benzimidazole and 5-(4-phenylbutoxymethv ⁇ -3-triphenylmethyl-4,5.6.7-tetrahvdro-3H-benzimidazole
• Tetrabutylammonium iodide (20 mg, 0.06 mmol) and (4-bromobutyl)benzene (0.80 g, 3.80 mmol) were added successively.
• the reaction mixture was stirred for 2 days at 60 °C. It was cooled to room temperature. Water (50 ml) was added dropwise.
• the reaction mixture was extracted with ethyl acetate (3 x 70 ml). The combined organic layers were washed with a saturated aqueous solution of sodium hydrogencarbonate (100 ml) and dried over magnesium sulphate. The solvent was removed in vacuo.
• the crude product was purified by flash chromatography on silica (40 g), using a gradient of ethyl acetate/heptane (from 1 :1 to 4:1) as eluent, to give 0.52 g of a mixture of 5-(4-phenylbutoxymethyl)-1-triphenyl- methyl-4,5,6,7-tetrahydro-1 H-benzimidazole and 5-(4-phenylbutoxymethyl)-3-triphenyl- methyl-4,5,6,7-tetrahydro-3H-benzimidazole.
• Phosphorus(lll) tribromide (0.41 ml, 4.37 mmol) was added to a solution of 1-[2-(2,4- dichlorophenoxy)phenyl]methanol (420 mg, 1.56 mmol) in dioxane (10 ml). The reaction mix- ture was stirred for 1 hour at room temperature. Another portion of phosphorus(lll) tribromide (0.41 ml, 4.37 mmol) was added. The reaction mixture was stirred for 16 hours at room temperature. It was cooled to 0 °C. Water (5 ml) was added dropwise. The reaction mixture was warmed to room temperature and diluted with ethyl acetate (400 ml).
• the aqueous phase was extracted with tert-butyl methyl ether (50 ml). The combined organic layers were dried over magnesium sulphate. The solvent was removed in vacuo.
• the crude product was purified by flash chromatography on silica, using ethyl acetate/heptane 1:1 followed by pure ethyl acetate as eluent, to give 319 mg of a mixture of 5-[2-(2,4-dichlorophenoxy)benzyloxymethyl]-1 -triphenylmethyl-4,5,6,7-tetrahydro-1 H- benzimidazole and 5-[2-(2,4-dichlorophenoxy)benzyloxymethyl]-3-triphenylmethyl-4,5,6,7- tetrahydro-3H-benzimidazole.
• Step 5 A soution of a mixture of 5-[2-(2,4-dichlorophenoxy)benzyloxymethyl]-1 -triphenyl- methyl-4,5,6,7-tetrahydro-1 H-benzimidazole and 5-[2 ⁇ (2,4-dichlorophenoxy)benzyloxy- methyl]-3-triphenylmethyl-4,5,6,7-tetrahydro-3H-benzimidazole (356 mg, 0.55 mmol) in a mixture of glacial acetic acid (9 ml) and water (1 ml) was heated to 70 °C for 2 hours. The reaction mixture was cooled to room temperature. The solvent was removed in vacuo. The crude product was purified by flash chromatography on silica (80 g), using DCM/methanol/-
• the title compound was transferred into its hydrochloride salt by dissolving it in ethyl acetate (20 ml) and addition of a 3.8 M solution of hydrogen chloride in ethyl acetate (5 ml). The solvent was removed in vacuo. The crystals were dried in vacuo.
• the reaction mixture was stirred for 16 hours at room temperature. It was added to ice water (2000 ml). The phases were separated. The aqueous phase was extracted with DCM (400 ml). The combined organic layers were extracted with a saturated aqueous solution of sodium hydrogencarbonate (3 x 400 ml). The aqueous sodium hydrogencarbonate phase was acidified with 1 N hydrochloric acid to pH 2. The precipitation was isolated by filtration and dissolved in ethyl acetate (300 ml). The ethyl acetate solution was dried over magnesium sulphate.
• the crude product was purified by flash chromatography on silica (90 g), using ethyl acetate/heptane (first 1:1, then 1:0) as eluent, to give 0.62 g of a mixture of 5-[2-(4-fluorobenzyl)benzyloxymethyl]1-triphenylmethyl-4,5,6,7- tetrahydro-1 H-benzimidazole 5-[2-(4-fluorobenzyl)benzyloxymethyl]-3-triphenylmethyl- 4,5,6,7-tetrahydro-3H-benzimidazole.
• the aqueous phase was extracted with ethyl acetate (2 x 50 ml). The combined organic layers were washed with water (100 ml) and dried over magnesium sulphate. The solvent was removed in vacuo.
• the crude product was purified by flash chromatography on silica (200 g), using ethyl acetate/heptane (1 :3) as eluent, to give 9.80 g of 2-(3-(difluoromethoxy)phenoxy)benzonitrile.
• Phosphorus tribromide (5 ml, 53 mmol) was added to a solution of [2-(3-(trifluoro- methoxy)phenoxy)phenyl]methanol (5.33 g, 18.9 mmol) in dioxane (100 ml). The reaction mixture was stirred for 1.5 hours. Another portion of phosphorus tribromide (5 ml, 53 mmol) was added. The reaction mixture was stirred for 16 hours at room temperature. It was diluted with ethyl acetate (500 ml) and washed with a 10% aqueous solution of sodium hydrogen sulphate (300 ml). The aqueous phase was extracted with ethyl acetate (2 x 100 ml).
• Step 5 5-r2-(3-(Trifluoromethoxy)phenoxy)benzyloxymethvH-1 -triphenylmethyl-4,5,6,7- tetrahydro-1 H-benzimidazole and 5-f2-(3-(trifluoromethoxy)phenoxy)benzyloxymethyl1-3- triphenylmethyl-4.5,6 -tetrahydro-3H-benzimidazole
• Tetrabutylammonium iodide (18 mg, 0.05 mmol) was added. The reaction mixture was stirred for 16 hours, while it was warming to room temperature. The reaction mixture was cooled to 0 °C. Water (5 ml) was added carefully dropwise, until the hydrogen evolution had ceased. The mixture was diluted with ethyl acetate (100 ml) and a 10% aqueous solution of sodium hydrogensulphate (100 ml) was added. The phases were separated. The aqueous phase was extracted with ethyl acetate (2 x 50 ml). The combined organic layers were washed with a saturated aqueous solution of sodium hydrogencarbonate (100 ml) and dried over magnesium sulphate.
• the title compound was transformed into its hydrochloride salt, by dissolving it in ethyl acetate (10 ml) and addition of a 3.5 M solution of hydrogen chloride in ethyl acetate (1.5 ml, 5.25 mmol). The solvent was removed in vacuo.
• Phosphorus tribromide (4.7 ml, 49.8 mmol) was added dropwise to a solution of 5- phenylpentan-1-ol (3 ml, 17.8 mmol) in dioxane (70 ml). The reaction mixture was stirred for 2 hours at room temperature. Another portion of phosphorus tribromide (4.7 m. 49.8 mmol) was added. The reaction mixture was stirred for 16 h at room temprature. It was cooled to 0 °C. Water (60 ml) was added dropwise. The reaction mixture was diluted with ethyl acetate (100 ml). The phases were separated.
• the aqueous phase was extracted with ethyl acetate (2 x 100 ml).
• the combined organic layers were washed with a saturated aqueous solution of sodium hydrogencarbonate (200 ml) and dried over magnesium sulfate.
• the solvent was removed in vacuo.
• the crude product was purified by flash chromatography on silica (50 g), using ethyl acetate/heptane (1 :10) as eluent, to give 1.35 g of 1-bromo-5-phenylpentane.
• Step 2 5-((5-Phenylpentlvoxy)methyl)-1-trityl-4.5.6.7-tetrahvdro-1 H-benzimidazole and 5-((5- phenylpentlvoxy)methv ⁇ -3-trityl-4,5,6.7-tetrahvdro-3H-benzimidazole
• PHARMACOLOGICAL METHODS The ability of the compounds to interact with the histamine H3 receptor can be determined by the following in vitro binding assays.
• Rat cerebral cortex is homogenized in ice cold K-Hepes, 5 mM MgCl 2 pH 7.1 buffer. After two differential centrifugations the last pellet is resuspended in fresh Hepes buffer containing 1 mg/ml bacitracin. Aliquots of the membrane suspension (400 ⁇ g/ml) are incubated for 60 min at 25°C with 30 pM [ 125 l]-iodoproxifan, a known histamine H3 receptor antagonist, and the test compound at various concentrations. The incubation is stopped by dilution with ice-cold medium, followed by rapid filtration through Whatman GF/B filters pretreated for 1 hour with 0.5% polyethyleneimine. The radioactivity retained on the filters is counted using a Cobra II auto gamma counter. The radioactivity of the filters is indirectly proportional to the binding affinity of the tested compound. The results are analyzed by nonlinear regression analysis.
• the H3-receptor agonist ligand R- ⁇ -methyl[ 3 H]histamine is incubated with isolated rat cortex cell-membranes at 25 °C for 1 hour, followed by a filtration of the incubate through Whatman GF/B filters. Radioactivity retained on the filters is measured using a beta counter.
• Rat cerebral cortex is homogenized in 10 volumes (w/w) ice-cold Hepes buffer (20 mM Hepes, 5 mM MgCI 2 pH 7.1 (KOH) + 1 mg/ml bacitracin) using an Ultra-Turrax homogenizer for 30 seconds. The homogenate is centrifuged at 140 g in 10 min. The supernatant is transferred to a new test tube and centrifuged for 30 min at 23 000 g.
• Pellet is resuspended in 5-10 ml Hepes buffer, homogenized and centrifuged for 10 min at 23 000 g. This short centrifugation step is repeated twice. After the last centrifugation the pellet is resuspended in 2-4 ml Hepes buffer and the protein concentration is determined. The membranes are diluted to a protein concentration of 5 mg/ml using Hepes buffer, aliquoted and stored at -80 °C until use
• test-compound 100 ⁇ l membrane (200 ⁇ g/ml), 300 ⁇ l Hepes buffer and 50 ⁇ l R- ⁇ -methyl[ 3 H]histamine (1 nM) are mixed in a test tube.
• the compounds to be tested are dissolved in DMSO and further diluted in H 2 O to the desired concentrations.
• Radioligand and membranes are diluted in Hepes buffer + 1 mg/ml bacitracin. The mixture is incubated for 60 min at 25 °C. Incubation is terminated by adding 5 ml ice-cold 0.9% NaCl, followed by rapid filtration through Whatman GF/B filters pre-treated for 1h with 0.5% polyethyleneimine.
• the human H3 receptor is cloned by PCR and subcloned into the pcDNA3 expression vector.
• Cells stably expressing the H3 receptor are generated by transfecting the H3- expression vectors into HEK 293 cells and using G418 to select for H3 clones.
• the human H3-HEK 293 clones are cultured in DMEM (GIBCO-BRL) with glutamax, 10% foetal calf serum, 1 % penicillin/streptavidin and 1 mg/ml G 418 at 37 °C and 5% CO 2 . Before harvesting, the confluent cells are rinsed with PBS and incubated with Versene (proteinase, GIBCO- BRL) for approximately 5 min.
• Versene proteinase, GIBCO- BRL
• the cells are flushed with PBS and DMEM and the cellsus- pension collected in a tube and centrifuged for 5-10 min at 1500 rpm in a Heraeus Sepatech Megafuge 1.0.
• the pellet is resuspended in 10-20 vol. Hepes buffer (20 mM Hepes, 5 mM MgCI 2 , pH 7.1 (KOH)) and homogenized for 10-20 seconds using an Ultra-Turrax homoge- nizer.
• the homogenate is centrifuged for 30 min at 23 000 g.
• the pellet is resuspended in 5- 10 ml Hepes buffer, homogenized 5-10 seconds with the Ultra-Turrax and centrifuged for 10 min at 23 000 g.
• the membrane pellet is resuspended in 2- 4 ml Hepes buffer, homogenized with a syringe or teflonhomogenizer, and the protein concentration determined.
• the membranes are diluted to a protein concentration of 1-5 mg/ml in Hepes buffer, aliquoted and kept at -80 °C until use.
• the compounds according to the invention have an IC 50 value as determined by one or more of the assays of less than 10 ⁇ M, more preferred of less than 1 ⁇ M, and even more preferred of less than 500 nM, such as of less than 100 nM.
• the ability of the compounds to interact with the histamine H3 receptor as agonists, inverse agonists and/or antagonists, is determined by an in vitro functional assay utilizing membranes from HEK 293 cell expressing the human H3 receptors.
• the H3 receptor is cloned by PCR and subcloned into the pcDNA3 expression vector.
• Cells stably expressing the H3 receptor are generated by transfecting the H3-expression vectors into HEK 293 cells and using G418 to select for H3 clones.
• the human H3-HEK 293 clones are cultured in DMEM with glutamax, 10% foetal calf serum, 1% penicillin/streptavidin and 1 mg/ml G 418 at 37 °C and 5% CO 2 .
• the H3 receptor expressing cells are washed once with phosphate buffered saline (PBS) and harvested using versene (GIBCO-BRL). PBS is added and the cells are centrifuged for 5 min at 188 g. The cell pellet is resuspended in stimulation buffer to a concentration of 1 x 10 6 cells/ml. cAMP accumulation is measured using the Flash Plate® cAMP assay (NENTM Life Science Products). The assay is generally performed as described by the manufacturer.
• test compound either agonists or inverse agonists alone, or agonist and antagonist in combination.
• the assay can be run in "agonist-mode" which means that the test compound is added, in in- creasing concentration, on its own, to the cells, and cAMP is measured. If cAMP goes up, it is an inverse agonist; if cAMP does not change, it is a neutral antagonist, and if cAMP goes down, it is an agonist.
• the assay can also be run in the "antagonist-mode" which means that a test compound is added, in increasing concentrations, together with increasing concentrations of a known H3 agonist (eg RAMHA). If the compound is an antagonist, increasing con- centrations of it cause a right-ward shift in the H3-agonist's dose-response curves.
• the final volume in each well is 100 ⁇ l. Test compounds are dissolved in DMSO and diluted in H 2 O. The mixture is shaken for 5 min, and allowed to stand for 25 min at room temperature. The reaction is stopped with 100 ⁇ l "Detection Mix" per well.
• EC 50 values are calculated by non-linear regression analysis of dose response curves (6 points minimum) using GraphPad Prism.
• Kb values are calculated by Schild plot analysis.
• the ability of the present compounds to reduce weight is determined using the in vivo open cage Schedule-fed rat model.
• Sprague-Dawley (SD) male rats of an age of about VA to 2 months and a weight of about 250 g are habituated to the presence of food (Altromin pelleted rat chow) in their home cage only during three hours in the morning from 9 to12 a.m. all days a week. Water is pre- sent ad libitum. As the consumption of food stabilised after 7 to 9 days, the animals are ready for use.
• food Altromin pelleted rat chow
• the animals are tested twice a week.
• the test compound is administered intraperitoneally 30 min before the start of the sessions.
• One group of 0 ani- mals is administered the test compound at a dose of 15 mg/kg and another group of 11 animals is administered the test compound at a dose of 30 mg/kg.
• a control group of 11 animals is administered the vehicle consisting of NaCl 0.0% and Chremophor 5%. Food and water intake are monitored at 1 , 2 and 3 hours post administration.
• the animals are weighed weekly and if necessary extra food is given in order to ensure that the weight gain is 3 to 5 g per week corresponding to the normal weight gain for SD male rats at this age.

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