Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/33—Heterocyclic compounds
  • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
  • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
  • A61K31/47—Quinolines; Isoquinolines
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/33—Heterocyclic compounds
  • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
  • A61K31/40—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/33—Heterocyclic compounds
  • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
  • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
  • A61K31/47—Quinolines; Isoquinolines
  • A61K31/475—Quinolines; Isoquinolines having an indole ring, e.g. yohimbine, reserpine, strychnine, vinblastine
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/33—Heterocyclic compounds
  • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
  • A61K31/535—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with at least one nitrogen and one oxygen as the ring hetero atoms, e.g. 1,2-oxazines
• • 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
  • A61P25/20—Hypnotics; Sedatives
• • 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
  • A61P25/24—Antidepressants
• • 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
  • A61P25/26—Psychostimulants, e.g. nicotine, cocaine
• • 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
  • 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
  • A61P25/00—Drugs for disorders of the nervous system
  • A61P25/30—Drugs for disorders of the nervous system for treating abuse or dependence
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P9/00—Drugs for disorders of the cardiovascular system
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P9/00—Drugs for disorders of the cardiovascular system
  • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis

Definitions

• This invention relates to a novel method of treatment and to novel compounds for use in such a method.
• European Published Patent Application No. 0126311 discloses substituted benzopyran compounds having blood pressure lowering activity, including 6-acetyl- trans-4-(4-fluorobenzoylamino)-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-3-ol.
• EP-A-0 376524, EP-A-0205 292, EP-A-0250077, EP-A-0093 535, EP-A-0 150 202, EP-A-0076075 and WO/89/05808 (Beecham Group pic) describe certain benzopyran derivatives which possess anti-hypertensive activity.
• WO ⁇ 89 ⁇ 11477 and WO ⁇ 89 ⁇ 07103 also describe certain compounds which are believed to possess anti-hypertensive activity.
• EP-A-0430 621 and EP-A-0385 584 (Beecham Group pic) describe the resolution of certain intermediates useful in the preparation of the compounds described in the above mentioned patent applications.
• EP-A-0 194 885 (E. Lilly) describes certain amino substituted benzopyran derivatives possessing anti-convulsant activity.
• EP-A-0509-762 (E.R. Squibb) describes certain indole and dihydroquinoline substituted derivatives which as disclosed as possessing inter alia anti-hypertensive activity.
• PCT/GB92/01045 (SmithKline Beecham pic; unpublished at the priority date), which describes certain fluorobenzoylamido benzopyrans, pyranopyridines and tetrahydronaphthylenes in which the 3 and 4 position substituents are trans to each other. These compounds are described as possessing inter alia anxiolytic and anti- convulsant activity.
• the present invention provides a method of treatment and/or prophylaxis of anxiety, mania, depression, the effects associated with withdrawal from substances of abuse such as cocaine, nicotine, alcohol and benzodiazepines; disorders treatable and/or preventable with anti-convulsive agents, such as epilepsy; and in the treatment or prevention of cerebral ischaemia, disorders resulting from sub- arrachnoid haemorrhage, Parkinson's Disease, migraine and/or psychosis, comprising administering to the sufferer in need thereof an effective or prophylactic amount of a compound of formula (I) or pharmaceutically acceptable salt thereof:
• P is a ring system selected from the following:
• R-P is hydrogen or C 1 -5 alkyl
• M a is R 5 ;
• J is carbon and M is nitrogen and J a and M a are hydrogen; or d)
• R 1 and R 2 is hydrogen and the other is selected from the class of hydrogen, C 3-8 cycloalkyl, C 1 -6 alkyl optionally interrupted by oxygen or substituted by hydroxy, C 1 -6 alkoxy or substituted aminocarbonyl, C 1 -6 alkylcarbonyl, C 1 -6 alkoxycarbonyl, C 1 -6 alkylcarbonyloxy, C 1 -6 alkoxy, nitro, cyano, halo,
• alkylsulphonyl C 1-6 alkylsulphonyl, C 1 -6 alkoxysulphinyl, C 1 -6 alkoxysulphonyl, aryl, heteroaryl, arylcarbonyl, heteroaiylcarbonyl, phosphono, arylcarbonyloxy, heteroarylcarbonyloxy, arylsulphinyl, heteroarylsulphinyl, arylsulphonyl,
• heteroarylsulphonyl in which any aromatic moiety is optionally substituted, C 1 -6 alkylcarbonylamino, C 1 -6 alkoxycarbonylamino, C 1 -6 alkyl-thiocarbonyl, C 1 -6 alkoxy-thiocarbonyl, C 1 -6 alkyl-thiocarbonyloxy, 1-mercapto C 2-7 alkyl, formyl, or aminosulphinyl, aminosulphonyl or aminocarbonyl, any amino moiety being optionally substituted by one or two C 1 -6 alkyl groups, or C 1 -6 alkylsulphinylamino, C 1 -6 alkylsulphonylamino,C 1-6 alkoxysulphinylamino or C 1 -6
• R 3 and R 4 is hydrogen or C 1 -4 alkyl and the other is C 1 -4 alkyl, CF3 or CH2 X a where X a is fluoro, chloro, bromo, iodo, C 1 -4 alkoxy, hydroxy, C 1 -4
• R 5 is C 1-6 alkylcarbonyloxy, benzoyloxy, ONO2, benzyloxy, phenyloxy or C 1 -6 alkoxy and R 6 and R 9 are hydrogen or R 5 is hydroxy and R 6 is hydrogen or C 1-2 alkyl and R 9 is hydrogen; in which R 1 1 and R 12 are independently selected from hydrogen, alkyl, haloalkyl, cycloalkyl, aralkyl, cyano, nitro, COR 13 , CONHR 13 , CONR 13 R 14 or halo where R 13 and R 14 are independently selected from hydrogen, alkyl, haloalkyl, aryl, aralkyl, cycloalkyl or (cycloalkyl)-alkyl;
• R 10 is hydrogen, alkyl, haloalkyl, cycloalkyl, O-R 15 , cyano, nitro, CF3, halo, S-alkyl, COR 15 , COOR 15 , NR 15 CO alkyl or OCO alkyl where R 15 is hydrogen, alkyl, haloalkyl, aryl, aralkyl, cycloalkyl or (cycloalkyl)-alkyl; m is O or 1;
• the R 7 group is cis or trans to the R 5 group.
• All C 1-6 alkyl or C 1 -4 alkyl or alkyl containing groups in formula (I) are preferably selected from methyl, ethyl, n - and iso -propyl, n -, iso -, sec - and tert-butyl.
• Suitable C 3-8 cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl and cyclooctyl.
• Suitable halo substituents include fluoro, chloro and bromo.
• Aryl whenever mentioned herein includes but is not limited to phenyl and naphthyl.
• Heteroaryl whenever mentioned herein includes a 5- or 6- membered monocyclic or 9- or 10- membered bicyclic of which 5- or 6- membered monocyclic heteroaryl is preferred. In addition, 5- or 6-membered monocyclic or 9- or
• 10-membered bicyclic heteroaryl preferably contains one, two or three heteroatoms which are selected from the class of oxygen, nitrogen and sulphur and which, in the case of there being more than one heteroatom, are the same or different.
• 5- or 6-membered monocyclic heteroaryl containing one, two or three heteroatoms which are selected from the class of oxygen, nitrogen and sulphur include furyl, thienyl, pyrryl, oxazolyl, thiazolyl, imidazolyl and thiadiazolyl, and pyridyl, pyridazyl, pyrimidyl, pyrazolyl and triazolyl.
• Preferred examples of such groups include furanyl, thienyl, pyrryl and pyridyl, in particular 2- and 3-furyl, 2- and
• 3-pyrryl, 2- and 3-thienyl, and 2-, 3- and 4-pyridyl examples of 9- or 10-membered bicyclic heteroaryl containing one, two or three heteroatoms which are selected from the class of oxygen, nitrogen and sulphur include benzofuranyl, benzothienyl, indolyl and indazolyl, quinolyl and isoquinolyl, and quinazolyl.
• Preferred examples of such groups include 2- and 3-benzofuryl, 2- and 3-benzothienyl, and 2- and 3-indolyl, and 2- and 3-quinolyl.
• Preferred variables are as those mentioned in the above mentioned patents which are incorporated herein by reference. Especially preferred variables are those mentioned in EP-0509762 which are incorporated herein by reference.
• the compounds of formula (I) may have chiral carbon atoms at positions around the saturated portion of ring system P especially at the atoms attached to R 5 and the moiety R 7 and therefore may exist as enantiomers.
• the present invention extends to each enantiomer and to mixtures thereof including racemates.
• R 1 substituents also have chiral centres and therefore may exist as enantiomers.
• the present invention extends to each enantiomer and to mixtures thereof including racemates
• pharmaceutically acceptable salt thereof also includes solvates of such compounds, such as for example the hydrate.
• the administration to the mammal may be by way of oral or parenteral administration.
• a unit dose will normally contain 1 to 1000 mg, suitably 1 to 500 mg, for example an amount in the range of from 2 to 400 mg such as 2, 5, 10, 20, 30, 40, 50, 100, 200, 300 and 400 mg of the active compound.
• Unit doses will normally be administered once or more than once per day, for example 1, 2, 3, 4, 5 or 6 times a day, more usually 1 to 4 times a day, such that the total daily dose is normally in the range, for a 70 kg adult of 1 to 1000 mg, for example 1 to 500 mg, that is in the range of approximately 0.01 to 15 mg/kg/day, more usually 0.1 to 6 mg/kg/day, for example 1 to 6 mg/kg/day.
• the compound of formula (I) is administered in the form of a unit-dose composition, such as a unit dose oral, rectal, topical or parenteral (especially intravenous) composition.
• compositions are prepared by admixture and are suitably adapted for oral or parenteral administration, and as such may be in the form of tablets, capsules, oral liquid preparations, powders, granules, lozenges, reconstitutable powders, injectable and infusable solutions or suspensions or suppositories.
• Orally administrable compositions are preferred, in particular shaped oral compositions, since they are more convenient for general use.
• Tablets and capsules for oral administration are usually presented in a unit dose, and contain conventional excipients such as binding agents, fillers, diluents, tabletting agents, lubricants, disintegrants, colourants, flavourings, and wetting agents.
• the tablets may be coated according to well known methods in the art.
• Suitable fillers for use include cellulose, mannitol, lactose and other similar agents.
• Suitable disintegrants include starch, polyvinylpyrrolidone and starch derivatives such as sodium starch glycollate.
• Suitable lubricants include, for example, magnesium stearate.
• Suitable pharmaceutically acceptable wetting agents include sodium lauryl sulphate.
• solid oral compositions may be prepared by conventional methods of blending, filling, tabletting or the like. Repeated blending operations may be used to distribute the active agent throughout those compositions employing large quantities of fillers. Such operations are, of course, conventional in the art.
• Oral liquid preparations may be in the form of, for example, aqueous or oily suspensions, solutions, emulsions, syrups, or elixirs, or may be presented as a dry product for reconstitution with water or other suitable vehicle before use.
• Such liquid preparations may contain conventional additives such as suspending agents, for example sorbitol, syrup, methyl cellulose, gelatin, hydroxyethylcellulose,
• non-aqueous vehicles which may include edible oils
• preservatives for example methyl or propyl p-hydroxybenzoate or sorbic acid, and if desired conventional flavouring or colouring agents.
• Oral formulations also include conventional sustained release formulations, such as tablets or granules having an enteric coating.
• fluid unit dose forms are prepared containing the compound and a sterile vehicle.
• the compound depending on the vehicle and the concentration, can be either suspended or dissolved.
• Parenteral solutions are normally prepared by dissolving the compound in a vehicle and filter sterilising before filling into a suitable vial or ampoule and sealing.
• adjuvants such as a local anaesthetic, preservatives and buffering agents are also dissolved in the vehicle.
• the composition can be frozen after filling into the vial and the water removed under vacuum.
• Parenteral suspensions are prepared in substantially the same manner except that the compound is suspended in the vehicle instead of being dissolved and sterilised by exposure to ethylene oxide before suspending in the sterile vehicle.
• a surfactant or wetting agent is included in the composition to facilitate uniform distribution of the compound of the invention.
• the compositions will usually be accompanied by written or printed directions for use in the medical treatment concerned.
• the present invention further provides a pharmaceutical composition for use in the treatment and/or prophylaxis of anxiety, mania, depression, disorders associated with a subarachnoid haemorrhage, neural shock, the effects associated with withdrawal from substances of abuse such as cocaine, nicotine, alcohol and benzodiazepines; disorders treatable or preventable with anti-convulsive agents, such as epilepsy; Parkinson's disease, psychosis, migraine and/or cerebral ischaemia, which comprises a compound of formula (I) or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.
• the invention provides the use of a compound of formula (I) or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the treatment and/or prophylaxis of anxiety, mania, depression and/or disorders associated with a subarachnoid haemorrhage, neural shock the effects associated with withdrawal from substances of abuse such as cocaine, nicotine, alcohol and benzodiazepines; disorders treatable or preventable with anti-convulsive agents, such as epilepsy; Parkinson's disease, psychosis, migraine and/or cerebral ischaemia.
• compositions may be prepared in the manner as hereinbefore described.
• the invention also extends to novel compounds of formula (I) and
• compounds of formula (I) which have trans isomerisation between the groups attached to the atom attached to the group R 7 and the group attached to the atom adjacent, on the right-hand side of the diagram as shown herein may be prepared by procedures generally described or analogous to those described in EP- 0126311, EP-0376524, EP-205292, EP-0250077, EP-0093535,
• compounds of formula (I) which have cis isomeration between the groups attached to the atom attached to the group R 7 and the group attached to the atom adjacent on the right-hand side of the diagram as shown herein may be prepared according to the procedures generally described on or analogous to those described in EP-A-0139992.
• Cis compound of formula (I) may also be prepared according to the procedures described by G. Burrell et al, Tet. Letters, 31, 3649-3652 (1996) or by the procedures described by U. Quast and E. Villhauer, Eur. J. Pharmacol, Molecular Pharmacology Section 245, 165-171 (1993). It should be appreciated that racemates for formula (I) may be resolved or enantiomerically purified compounds of formula (I) may be prepared using procedures conventional in the art and in particular using the procedures outlined in EP-0430631 and EP-0355584.
• the compounds of formula (I) may be prepared in the required enantiomeric form by forming a chirally pure epoxide using catalysts and conditions generally outlined in WO91 ⁇ 14694 or WO 93 ⁇ 17026 and thereafter converted to the required compound of formula (I) using procedures outlined herein.
• the invention also provides a pharmaceutical composition
• a pharmaceutical composition comprising novel compounds of formula (I), or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.
• the invention also provides the use of novel compounds of formula (I), or a pharmaceutically acceptable salt thereof as a therapeutic agent, in particular in the treatment and/or prophylaxis of anxiety, mania, depression, the effects associated with withdrawal from substances of abuse such as cocaine, nicotine, alcohol and benzodiazepines; disorders treatable and/or preventable with anti-convulsive agents, such as epilepsy; cerebral ischaemia disorders resulting from subarachnoid haemorrhage, Parkinson's disease, migraine and/or psychosis.
• Suitable examples of compounds of formula (I) include examples 1 to 4 in EP-A-0509762 which are incorporated herein by reference.
• mice Male Sprague - Dawley rats (Charles River, U.K., 250 - 300g) are singly housed for 3 days prior to testing. On the test day, the animals are then randomly assigned to groups of 8 - 16 and dosed orally at a dose volume of 1 ml/kg with various doses of compound (1 - 300 mg/kg) or vehicle. At ⁇ min post dose the rats are placed with a weight - and treatment - matched pair male (encountered for the first time) in the social interaction box under high - light, unfamiliar conditions. The box is made of white perspex 54 x 37 x 26 cm with a transparent perspex front side. The floor is divided into 24 equal squares and is brightly lit (115 lux). Time spent (sees) in active social interaction (sniffing, grooming, following, mounting, climbing over or under, boxing, biting) is scored "blind" by remote monitoring as is the number of squares crossed (as an index of locomotion).
• Drugs are suspended in 1% methyl cellulose.
• the maximal electroshock seizure (MES) threshold test in rodents is particularly sensitive for detecting potential anticonvulsant properties 1 .
• anticonvulsant agents elevate the threshold to electrically - induced seizures whilst proconvulsants lower the seizure threshold.
• mice Male, Charles River, U.K. CD - 1 strain, 25 - 30g are randomly assigned to groups of 10 - 20 and dosed orally or intraperitoneally at a dose volume of 10 ml/kg with various doses of compound (0.3 - 300 mg/kg) or vehicle. Mice are then subjected at 30 or 60 min post dose to a single electroshock (0.1 sec, 50Hz, sine wave form) administered via corneal electrodes.
• the mean current and standard error required to induce a tonic seizure in 50% (CC 50 ) of the mice in a particular treatment group is determined by the 'up and down' method of Dixon and Mood (1948) 2 .
• the CC 50 is usually 14 - 18 mA. Hence the first animal in the control group is subjected to a current of 16 mA. If a tonic seizure does not ensue, the current is increased for a subsequent mouse. If a tonic convulsion does occur, then the current is decreased, and so on until all the animals in the group have been tested.
• the percentage increase or decrease in CC 50 for each group compared to the control is calculated.
• the X-maze test of anxiety examines the exploratory response of naive rats in an environment which offers both anxiogenic (open arms) and relatively non-anxiogenic (closed arms) areas. A selective increase in exploration of the open arms following drug pretreatment is therefore postulated to indicate anxiolytic effects.
• the X-maze was raised 70cm above the floor and consisted of two enclosed arms 45cm (long) x 15cm (wide) x 10cm (high) and two open arms 45 x 10 x 1cm arranged such that the two arms of each type were opposite each other. Both arm types were marked into two equal sections. Rats were placed onto the centre of the X-maze and observed for a period of 10 minutes during which time the following parameters were recorded: 1) the number of entries onto, and the time spent on, (a) open arms, (b) closed arms, (c) end of open arms and (d) end of closed arms. 2) the number of sections crossed. The fear-drive evoked in the open arms exceeds that in the enclosed arms and rats typically show a clear preference for the enclosed arms.
• Anxiolytic drugs increase the number of entries made onto, and the time spent on, the outer half of the open arms, and also the percentage of entries made onto, and the time spent on, the whole of the open arms. These four measures of anxiety, and also the total number of sections traversed, were calculated for each animal. Drugs are administered intraperitoneally or orally to groups of 6 to 12 rats 30 to 60 mins before testing. Statistical comparisons between vehicle- and drug-treated groups were made using a Mann- Whitney 'U' test (two tailed).
• End-tidal CO 2 (et CO 2 ) is monitored continuously and arterial blood gas analysis was performed periodically to assure stable and adequate ventilation throughout each experiment.
• Polyethylene cannulae are placed in the left external jugular vein and the right femoral artery and vein for drug administration, monitoring arterial blood pressure, and blood sampling, respectively.
• Transfemoral catheterization of the left vertebral artery is then performed via the left femoral artery using a 5 french Lehman dacron catheter ( Bard, Tewksbury MA). Anaesthesia is supplemented as needed with pentobarbital (5 mg/kg, i.v.) prior to the experimental period.
• the effects of the compounds of this invention are also examined in the chronic canine model of delayed cerebral vasospasm (two haemorrhage model of cerebral vasospasm).
• a control vertebral angiogram is obtained and autologous blood is administered intracisternally on day 1 (as above).
• day 3 the intracisternal administration of blood is repeated and the severe delayed vasospasm is quantitated angiographically on day 7 in all animals.
• the effect of an infusion of test compounds on the reversal of significantly delayed cerebral vasospasm is observed. 5.
• the compounds of formula (I) or pharmaceutically acceptable salts thereof are tested for therapeutic utility using the procedures outlined as follows: 1) Anti-Parkinsonian Activity
• Transient forebrain ischemia is produced by bilateral carotid artery ligation under 2.5% isoflourane in 100% O 2 anesthesia, the animals being placed onto a heating pad to maintain body temperature at 37° C.
• the common carotid arteries are exposed and aneurism clips are placed on both arteries for a certain period of time indicated in the figure legends.
• PBN dissolved in saline was administered intraperitoneally as a bolus 30 min before occlusion (pretreatments) or immediately after and again at 6 h of reperfusion, followed by the same dose b.i.d. for 2 days (post- treatment).
• pretreatments pretreatments
• reperfusion followed by the same dose b.i.d. for 2 days (post- treatment).
• post- treatment For quantification of CA1 neurons, animals are sacrificed at 7 days postischemia and perfused with buffered formalin.
• Brains were removed, stored in formalin for 3 days, embedded in paraffin, cut at 7- ⁇ m-thick coronal sections (1.5-1.9 mm posterior to bregma ⁇ ) and stained with thionin. The number of intact neurons over a 750- ⁇ m length of the CA1 layer on both hippocampal sides of 3 sections is counted for each animal. b) MCAO Method
• SHR Three strains of mature male rats (SHR) are obtained from commercial vendors (Taconic Farms, Germantown, NY; Charles River, Danvers,
• the tubing is lead subdermally from the artery and exteriorized between the scapula just below the back of the neck and cleared/filled with sterile isotonic saline. Incisions are closed using 2- 0 silk suture and treated with 5% lidocaine ointment (Astra).
• MCAO() or sham surgery is carried out in the SHR, SD rats under sodium pentobarbital (65 mg/kg, i.p. and supplemented as needed) anesthesia. All animals are allowed free access to food and water prior to and after surgery. Body temperature is maintained at 37°C using a heating pad throughout the surgical procedure. Surgery is conducted similar to that described previously (2.4). The right dorsal surface to the head and shaved and prepped with providone-iodine, and the rat placed in a stereotaxic device (David Kopf Instruments, Tujunga, CA) with the surgery (right) side of the head superior. A 1-2 cm incision was made between the orbit and the external auditory canal.
• the temporal muscle is dissected from the skull and retracted without damaging the zygomatic bond or mandibular nerve. Under an operating microscope and with saline irrigation, a 2-3 mm craniotomy is made just rostral to the zygomatic-squamosal skull suture. The dura is opened over the artery using the modified tip of a 30-gauge needle. For permanent right MCAO, using electrocoagulation (Force 2
• Electrosurgical Generator Valley Lab Inc., Boulder, CO
• the artery was stimultaneously occluded and cut dorsal to the lateral olfactory tract at the level of the inferior cerebral vain.
• a small piece of sterile saline-soaked Gelfoam (Upjohn, Kalamazoo, M1) is then positioned over the craniotomy and the temporails muscle and skin are closed in two layers. Animals are allowed to recover from anesthesia under a heating lamp and then are returned to their cages. The animals are sacrificed 24 hours following MCAO and the brains are prepared from reactive histologic examination.
• rats are euthanized with an overdose of sodium pentobarbital.
• brains are removed and six coronal forebrain slices (2 mm thick) are made from the level of the olfactory bulbs to the cortical-cerebellar junction using a rat brain slicer [(59); Zivic-Miller Laboratories Inc.,
• the 11 planar images were obtained from each side of the six 2 mm thick sections and correspond approximately to 1 mm section surfaces from + 5mm to -5 mm from bregma (97) and include the complete forebrain. These planar image surfaces (from the photographs) are digitized and used in the Image Analysis System for planimetry determination of infarct size and swelling. Two parameters of ischemic damage due to MCAO are determined for each slice as described previously (2,4,98,122).
• “Hemispheric swelling” is expressed as the percent increase in size of the ipsilateral (i.e., surgery side) hemisphere over the contralateral (normal) hemisphere and is calculated as:
• Infarct size which was expressed as the percent infarcted tissue in reference to the contralateral (normal) hemisphere and is calculated as:
• the swelling and infarct size are expressed in reference to the contralateral hemisphere (i.e., ipsilateral ischemic damage is normalized to the normal contralateral hemisphere). These parameters are determined for each slice to evaluate the profile of damage throughout the forebrain (i.e., "fore-brain profile") and for “total” forebrain changes by using the sum of all individual slice data in these formulas.

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• 1992
  • 1992-12-11 GB GB929225859A patent/GB9225859D0/en active Pending
• 1993
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  • 1993-12-08 EP EP94902047A patent/EP0674519A1/en not_active Withdrawn
  • 1993-12-08 WO PCT/GB1993/002515 patent/WO1994013297A1/en not_active Application Discontinuation

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