HK1182015B - Use of active pharmaceutical compounds for the treatment of central nervous system conditions - Google Patents

Abstract

The present invention relates to the pharmaceutical use of selective GABA A α5 negative allosteric modulators for the treatment, prevention and/or delay of progression of central nervous system (CNS) conditions related to excessive GABAergic inhibition in the brain.

Description

Use of active pharmaceutical compounds for the treatment of central nervous system disorders

The present invention relates to the pharmaceutical use of selective GABAA α 5 negative allosteric modulators for the treatment, prevention and/or delay of progression of Central Nervous System (CNS) disorders associated with excessive gabaergic inhibition in the cortex and hippocampus. More particularly, the present invention relates to the pharmaceutical use of selective GABAA α 5 negative allosteric modulators for the treatment, prevention and/or delay of progression of CNS disorders caused by neurodevelopmental defects leading to excessive gabaergic inhibition in the cortex and hippocampus, or for post-stroke recovery, wherein said CNS disorders are selected from the group consisting of cognitive deficit in down syndrome, cognitive deficit in autism, cognitive deficit in neurofibromatosis type I.

In particular, the present invention relates to the use of a selective GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a CNS disorder, wherein said selective GABAA α 5 negative allosteric modulator is selected from a compound of formula (I) and/or a compound of formula (II) or a pharmaceutically acceptable salt thereof

Wherein R is¹、R²、R³、R⁴、R⁵And R⁶As defined herein.

Down Syndrome (DS) caused by trisomy of chromosome 21 is the most common genetic cause of intellectual disability, with about one live births in every 650 to 1000 live births worldwide [ bittersah et al, eur jpublic health (2007)17 (2): 221-225]. Although the cause of DS cognitive deficits remains uncertain, prenatal and perinatal forebrain and cerebellar cellular and anatomical abnormalities indicate altered early brain development in subjects with DS. Similar Central Nervous System (CNS) abnormalities have been described in the DS mouse model. In particular, Ts65Dn mouse (the most widely used DS model) has abnormal forebrain and cerebellar development, synaptogenesis and neurophysiological and behavioral deficits.

Recent studies have shown that a major functional defect in the post-partum Ts65Dn brain can be an imbalance between excitation and inhibition, such as a decrease in the number of excitatory synapses and a relative increase in inhibitory synaptic markers in the cortex and hippocampus. Further studies have shown that an increase in inhibitory synaptic driving force may be a general physiological phenotype in the anterior brain of Ts65 Dn.

There are currently no treatment options available for treating cognitive deficits in DS patients. It has now been found that inhibition of GABAA receptor function is an attractive mechanism for the treatment of DS cognitive impairment.

GABAA receptors that regulate chloride channels (chloridephannels) are the major inhibitory neurotransmitter receptors in the mammalian central nervous system and have been widely used as targets for neuromodulatory drugs. Many compounds used clinically (e.g. anxiolytics, sedatives, hypnotics or antiepileptics) via allosteric benzodiazepines(BZD) binding sites to increase GABAA receptor activation. Such compounds have been referred to as "BZD site receptor agonists". BZD binding site ligands that produce the opposite effect (i.e., decrease receptor activation) are referred to as "BZD site receptor inverse agonists". "BZD site receptor antagonists" are ligands that bind to the receptor without modulating its function, but block the activity of both agonists and inverse agonists [ HaefelyWE, eurarchpsychiatyrneurol sci (1989) 238: 294-301]. To date, BZD receptor inverse agonists have only been tested in animal behavior experiments and in a very few exploratory human studies. The results show beneficial activity, however, further development of compounds into the clinic is prevented by anxiogenic effects that may result from the lack of selectivity shown by these agents for particular BZD receptor subtypes.

Non-selective antagonists of the GABAA receptor (also known as channel blockers), such as printed defense toxin or PTZ, most likely increase the risk of convulsions through their effects on receptors containing GABAA α 1, α 2 and α 3 subunits, and therefore cannot be safely used in DS patients. It is therefore a prerequisite that suitable GABAA receptor inhibitors are selective for receptor subtypes primarily involved in memory formation.

The importance of different α subunit subtypes has been elucidated by the production of transgenic mice lacking normal diazepam sensitivity (α 4 and α 6 are insensitive to diazepam) for the α 1, α 02, α 3, or α 5 subunits, results indicate that α 1 is responsible for the sedative effect of BZD receptor ligand agonists, and that α 2 and perhaps α 3 are responsible for the anxiolytic effect of BZD receptor ligand agonists [ the ] GABAA receptor is a pentamer composed mostly of 2 α, 2 p, and 1 γ subunitsK et al, Science (2000)290 (5489): 131-134;H，CellTissueRes(2006)326(2)：505-516]the results of improved pharmacology of the α 5 subunit are less pronounced, but reduced or no expression of this subunit can be associated with cognitive promotion in hippocampal-dependent tasks and, importantly, have no effect on the anxiety or convulsive paradigm, consistent with preferential localization of the α 5 subunit in the hippocampus.

Thus, it is hypothesized that a BZD site ligand that has inverse agonist selectivity for GABAA receptors containing the α 5 subunit should enhance cognitive function without anxiogenic and proconvulsive side effects.

Selectivity of the BZD site ligands can be achieved by different affinities for GABAA receptor subtypes ("binding selectivity"). Alternatively, in the case of similar subtype affinities, different degrees of receptor modulation ("functional selectivity") may be attempted, i.e., inverse agonism for the GABAA α 5 receptor subtype and no activity for other subtypes. Compounds may also have a combination of both binding and functional selectivity, but this has so far been rare. Recently, a number of compounds have been synthesized which have been described as inverse agonists with activity at α 5 subunit-containing GABAA receptors [ WO2006/045429, WO2006/045430, WO2007/042421, WO2009/071476 ]. It has now been found that certain of these compounds have beneficial pharmacological properties as well as excellent binding and functional selectivity for GABAA receptors containing the α 5 subunit. The results confirm the following assumptions: compounds with such pharmacological properties may improve cognitive function without CNS-mediated side effects, including anxiety and/or convulsions [ ballard et al, Psychopharmacology, (2009) 202: 207-223].

The pharmaceutically active compounds used in the present invention are molecules with a combination of both binding and functional selectivity for α 5 subunit containing GABAA receptors that improve cognition. Importantly, the pharmaceutically active compounds used in the present invention have no anxiogenic or convulsant effect under the exposure tested in toxicological studies.

In the present invention it has been found that selective negative allosteric modulators of GABAA α 5 have a pro-cognitive effect on a variety of animal models, but do not cause anxiety or convulsions. Ts65Dn and control (euploid) mice were chronically administered the active pharmaceutical compound used in the present invention and subjected to a series of behavioral tests including assessment of sensorimotor performance, anxiety and cognition. The active pharmaceutical compounds used in the present invention improved the performance of Ts65Dn mice (but not the control mice) in the Morris water maze (morriswaterfastermaze) and did not affect the sensorimotor performance, general activity, motor coordination or anxiety of Ts65Dn or the control mice. Plasma concentrations of the active pharmaceutical compound in blood samples obtained from treated Ts65Dn and control mice correlated with 25-75% GABAA α 5 receptor occupancy levels in vivo binding simulation studies. Importantly, these experiments demonstrate selective occupancy of brain GABAA α 5 receptors and reinforce the following view: the dual binding and functional selectivity confers the desired cognitive enhancing effector properties without the undesirable side effects associated with activity at other GABAA receptor subtypes.

Interestingly, it has been shown that long term administration of the active pharmaceutical compounds used in the present invention:

1. does not alter either Ts65Dn or the tested sensorimotor performance of the control mice;

2. does not affect the coordination of movement in the rotameter test (rotameter);

3. does not alter spontaneous locomotor activity in the cage during the light or dark phase of the cycle;

4. anxiety or motor activity was not altered in the open field test (OpenFieldtest) for Ts65Dn and control mice;

5. reduction of hyperactivity found in vehicle treated Ts65Dn mice in the well plate test (HoleBoardtest);

6. performance of Ts65Dn mice during the learning (acquisition) and cued (cued) phase was improved in the Morris water maze.

It was further found that the active pharmaceutical compounds used in the present invention:

a) reversing the spatial learning deficit of Nfl +/-mutant mice under conditions in which the compound does not enhance learning in control mice;

b) does not affect performance under conditions that mask Nfl +/-mice's behavioral deficits;

c) does not affect Nfl +/-and does not affect the motor learning in the rotarod test of the control mice;

d) can be used as a potential treatment for cognitive deficits associated with NFl.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, suitable methods and materials are described below.

Unless otherwise indicated, the nomenclature used in this application is based on the IUPAC systematic nomenclature.

Any open valency appearing on a carbon, oxygen, sulfur or nitrogen atom in the structures herein indicates the presence of hydrogen, unless otherwise indicated.

The definitions set forth herein apply regardless of whether the terms in question appear alone or in combination. It is contemplated that the definitions described herein may be supplemented to form chemically related combinations, such as "heterocycloalkyl-aryl," haloalkyl-heteroaryl, "" aryl-alkyl-heterocycloalkyl, "or" alkoxy-alkyl. The last member of the combination is a group that is substituted in reverse order by the other members of the combination.

The term "one or more" when referring to the number of substituents refers to the range from one substituent to the highest possible number of substitutions, i.e., replacement of one hydrogen by a substituent to replacement of at most all hydrogens.

The terms "optional" or "optionally" mean that the subsequently described event or circumstance may, but need not, occur, and that the description includes instances where the event or circumstance occurs and instances where it does not.

The term "substituent" means an atom or group of atoms that replaces a hydrogen atom on a parent molecule.

The term "substituted" means that the specified group bears one or more substituents. Where any group may carry multiple substituents and a variety of possible substituents are provided, the substituents are independently selected and need not be the same. The term "unsubstituted" means that the specified group bears no substituents. The term "optionally substituted" means that the specified group is unsubstituted or substituted with one or more substituents independently selected from the group of possible substituents. The term "one or more" when referring to the number of substituents refers to from one substituent to the highest possible number of substitutions, i.e., replacement of one hydrogen by a substituent to replacement of at most all hydrogens.

The terms "compound used in the present invention" and "compound used in the present invention" mean a compound of formula (I) or (II) and stereoisomers, tautomers, solvates, and salts (e.g., pharmaceutically acceptable salts) thereof.

The term "pharmaceutically acceptable salt" refers to salts that are not biologically or otherwise undesirable. Pharmaceutically acceptable salts include acid addition salts and base addition salts.

The term "pharmaceutically acceptable acid addition salts" denotes those pharmaceutically acceptable salts formed using the following acids: inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, carbonic acid, phosphoric acid; and organic acids selected from the group consisting of aliphatic, alicyclic, aromatic, araliphatic, heterocyclic, carboxylic and sulfonic organic acids, such as formic acid, acetic acid, propionic acid, glycolic acid, gluconic acid, lactic acid, pyruvic acid, oxalic acid, malic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, aspartic acid, ascorbic acid, glutamic acid, anthranilic acid, benzoic acid, cinnamic acid, mandelic acid, embonic acid (embonic acid), phenylacetic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, and the like.

The term "pharmaceutically acceptable base addition salts" denotes those pharmaceutically acceptable salts formed using organic or inorganic bases. Examples of acceptable inorganic bases include sodium, potassium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, and aluminum salts. Salts derived from pharmaceutically acceptable organic non-toxic bases include: primary, secondary and tertiary amines, substituted amines (including naturally occurring substituted amines), cyclic amines, and salts of basic ion exchange resins, such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, 2-diethylaminoethanol, tromethamine, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine (procaine), hydrabamine (hydrabamine), choline, betaine, ethylenediamine, glucosamine, methyl reduced glucamine, theobromine, purine, piperazine, piperidine, N-ethylpiperidine, and salts of polyamine resins.

The terms "halo", "halogen" and "halide" are used interchangeably herein and denote fluorine, chlorine, bromine or iodine. In particular, halogen represents F, C1 or Br, most particularly F.

The term "alkyl" denotes a monovalent straight or branched chain saturated hydrocarbon group having 1 to 12 carbon atoms, especially 1 to 7 carbon atoms, more especially 1 to 4 carbon atoms, for example, methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, sec-butyl or tert-butyl. In particular, alkyl represents methyl or isopropyl, most particularly methyl.

The term "alkoxy" denotes a group of formula-O-R ', wherein R' is alkyl. Examples of alkoxy moieties include methoxy, ethoxy, isopropoxy, and tert-butoxy.

The term "haloalkyl" denotes an alkyl group wherein at least one hydrogen atom of the alkyl group has been replaced by the same or a different halogen atom, especially a fluorine atom. Examples of haloalkyl include monofluoro-, difluoro-or trifluoromethyl, -ethyl or-propyl, such as 3, 3, 3-trifluoropropyl, 2-fluoroethyl, 2, 2, 2-trifluoroethyl, fluoromethyl or trifluoromethyl. The term "perhaloalkyl" denotes an alkyl group wherein all of the hydrogen atoms of the alkyl group have been replaced by the same or different halogen atoms. In particular, haloalkyl denotes monofluoromethyl and difluoromethyl.

The term "hydroxyalkyl" denotes an alkyl group wherein at least one hydrogen atom of the alkyl group has been replaced by a hydroxyl group. Examples of hydroxyalkyl groups include: hydroxymethyl, 2-hydroxyethyl, 2-hydroxypropyl, 3-hydroxypropyl, 1- (hydroxymethyl) -2-methylpropyl, 2-hydroxybutyl, 3-hydroxybutyl, 4-hydroxybutyl, 2, 3-dihydroxypropyl, 2-hydroxy-1-hydroxymethylethyl, 2, 3-dihydroxybutyl, 3, 4-dihydroxybutyl or 2- (hydroxymethyl) -3-hydroxypropyl.

The term "heterocycloalkyl" refers to a monovalent saturated or partially unsaturated monocyclic or bicyclic ring system of 4 to 9 ring atoms, comprising 1, 2 or 3 ring heteroatoms selected from N, O and S, the remaining ring atoms being carbon. Bicyclic means consisting of two rings having two ring atoms in common, i.e. the bridge separating the two rings is a single bond or a chain of one or two ring atoms. Examples of monocyclic saturated heterocycloalkyl groups are: azetidinyl, pyrrolidinyl, tetrahydrofuranyl, tetrahydro-thienyl, pyrazolidinyl, imidazolidinyl, oxazolidinyl, isoxazolidinyl, thiazolidinyl, piperidinyl, tetrahydropyranyl, tetrahydrothiopyranyl, piperazinyl, morpholinyl, thiomorpholinyl, 1-dioxo-thiomorpholin-4-yl, azepinyl, diazepanyl, homopiperazinyl, or oxazepinyl. Examples of bicyclic saturated heterocycloalkyl are: 8-aza-bicyclo [3.2.1] octyl, quinuclidinyl, 8-oxa-3-aza-bicyclo [3.2.1] octyl, 9-aza-bicyclo [3.3.1] nonyl, 3-oxa-9-aza-bicyclo [3.3.1] nonyl or 3-thia-9-aza-bicyclo [3.3.1] nonyl. Examples of partially unsaturated heterocycloalkyl groups are dihydrofuranyl, imidazolinyl, dihydro-oxazolyl, tetrahydro-pyridyl or dihydropyranyl. Heterocycloalkyl groups may be optionally substituted as described herein. In particular, heterocycloalkyl represents morpholinyl, thiomorpholinyl, dioxothiomorpholinyl, 2-oxa-6-aza-spiro [3.3] hept-6-yl, pyrrolidinyl and oxopyrrolidinyl. Most particularly, heterocycloalkyl represents morpholinyl, thiomorpholinyl or dioxothiomorpholinyl.

The term "heterocycloalkylalkyl" denotes an alkyl group having at least one hydrogen atom of the alkyl group replaced by a heterocycloalkyl group. Examples of heterocycloalkyl alkyl groups include pyrrolidinyl-methyl and pyrrolidinyl-methyl.

The term "aryl" denotes a monovalent aromatic carbocyclic mono-or bicyclic ring system comprising 6 to 10 carbon ring atoms. Examples of aryl moieties include phenyl and naphthyl. Aryl groups may be optionally substituted as described herein. Particular aryl groups are phenyl and monofluoro-phenyl.

The term "heteroaryl" denotes a monovalent aromatic heterocyclic mono-or bicyclic ring system having 5 to 12 ring atoms, comprising 1, 2, 3 or 4 heteroatoms selected from N, O and S, the remaining ring atoms being all carbon. Examples of heteroaryl moieties include: pyrrolyl, furyl, thienyl, imidazolyl, oxazolyl, thiazolyl, triazolyl, oxadiazolyl, thiadiazolyl, tetrazolyl, pyridyl, pyrazinyl, pyrazolyl, pyridazinyl, pyrimidinyl, triazinyl, azepinyl, diazepitriptenyl, isoxazolyl, benzofuranyl, isothiazolyl, benzothienyl, indolyl, isoindolyl, isobenzofuranyl, benzimidazolyl, benzoxazolyl, benzisoxazolyl, benzothiazolyl, benzisothiazolyl, benzooxadiazolyl, benzothiadiazolyl, benzotriazolyl, purinyl, quinolyl, isoquinolyl, quinazolinyl, quinoxalinyl, carbazolyl, or acridinyl. Heteroaryl groups may be optionally substituted as described herein. In particular, heteroaryl represents pyridyl, monofluoropyridyl and 5, 6-dihydro-8H- [1, 2, 4] triazolo [4, 3-a ] pyrazin-7-yl, most particularly pyridyl and monofluoropyridyl.

The term "oxo" denotes a divalent oxygen atom ═ O.

The term "active pharmaceutical ingredient" (or "API") denotes a compound in a pharmaceutical composition having a specific biological activity.

The term "pharmaceutically acceptable" refers to the property of a material that is useful in the preparation of pharmaceutical compositions that are generally safe, non-toxic, and not biologically or otherwise undesirable, and that is acceptable for veterinary as well as human pharmaceutical use.

The term "pharmaceutically acceptable excipient" means any ingredient that is not therapeutically active and is non-toxic, such as disintegrants, binders, fillers, solvents, buffers, tonicity agents, stabilizers, antioxidants, surfactants or lubricants used in formulating pharmaceutical products.

The term "pharmaceutical composition" (or "composition") means a mixture or solution comprising a therapeutically effective amount of an active pharmaceutical ingredient, and a pharmaceutically acceptable excipient, to be administered to a mammal, such as a human, in need thereof.

The term "modulator" refers to a molecule that interacts with a target receptor. Interactions include, for example, agonistic, antagonistic, or inverse agonistic activity.

The term "inhibitor" refers to a compound that competes with a particular ligand for binding to a particular receptor, reduces or prevents binding of a particular ligand to a particular receptor, or which reduces or prevents inhibition of a particular protein function.

The term "agonist" refers to a compound that has affinity for a receptor binding site and activity to enhance a receptor-mediated response, as defined, for example, in Goodman and Gilman, "the pharmacological basis therapeutics, 7 th edition," page 35, macmillan publication. A "full agonist" achieves a complete response, whereas a "partial agonist" achieves less than complete activation, even if it occupies the entire receptor population. An "inverse agonist" produces an opposite effect to an agonist by binding to the same agonist binding site, or attenuates the effect of an agonist by binding to a different allosteric binding site.

The term "antagonist" means a compound that attenuates or prevents the action of other compounds or receptor sites, as defined, for example, in Goodman and Gilman, "the pharmacological basis of therapeutics, 7 th edition," page 35 macmillan publication. In particular, antagonists refer to compounds that attenuate the effects of agonists. A "competitive antagonist" binds to the same site as an agonist, but does not activate that site, thereby blocking the action of the agonist. A "noncompetitive antagonist" binds to an allosteric (non-agonist) site on the receptor to prevent receptor activation. The "reversible antagonist" binds non-covalently to the receptor and can therefore be "washed out". An "irreversible antagonist" covalently binds to a receptor and is not replaced by competing ligands or washes.

The term "allosteric modulator" refers to a compound that binds to a site of the receptor other than the binding site for an agonist ("allosteric site"). It induces a conformational change in the receptor which in turn alters the affinity of the receptor for the endogenous ligand or agonist. A "positive allosteric modulator" increases affinity, while a "negative allosteric modulator" (NAM) decreases affinity and thus indirectly decreases the activity of the receptor. In the present invention, negative allosteric modulators specifically bind to benzodiazepinesA binding site and inverse agonist selectivity for the GABAA receptor comprising the α 5 subunit.

The term "inhibition constant" (Ki) denotes the absolute binding affinity of a particular inhibitor for a receptor. It is measured using a competitive binding assay and is equal to the concentration at which the particular inhibitor occupies 50% of the receptor in the absence of a competing ligand (e.g., a radioligand). The Ki value pairs can be converted numerically to pKi values (-logKi), where an increase in the number indicates an exponential increase in potency.

The term "next to the maximum effective concentration" (EC10) means the concentration of a particular compound required to achieve 10% of the maximum value of the particular effect.

The term "binding selectivity" refers to the ratio between the binding affinities of a particular compound for two or more different receptor subtypes. A particular compound is said to be "binding selective" if its binding selectivity is 10 or greater, more particularly if its binding selectivity is 20 or greater, and most particularly if its binding selectivity is 50 or greater.

The term "functionally selective" refers to different degrees of modulation of different receptor subtypes by a particular compound, e.g., as an inverse agonist for one particular receptor subtype, and as an antagonist for another receptor subtype. In the present invention, a compound is particularly functionally selective if it acts as an inverse agonist of the GABAA α 5 β 3 γ 2 receptor subtype and reduces the effect of GABA by more than 30% but has an effect on other GABAA receptor subtypes of less than 15%, especially less than 10%.

The terms "disorder," "defect," "disability," "disorder," "disease," or "disease condition" are used interchangeably to refer to any disease, disorder, symptom, disorder, or indication.

The term "neurodevelopmental defect" denotes a neurodevelopmental disorder in which the growth and development of the brain or central nervous system has been impaired (ReynoldCR et al, thought of, handbook of neuro-developmental and networkic disorder of the same, N.Y.).

The term "GABAergic inhibition" means GABA-mediated neurotransmission with inhibition to mature neurons in vertebrates (BernardC et al, Epilepsia (2000)41 (S6): S90-S95).

The term "excessive gabaergic inhibition" denotes an increase in GABA-mediated neurotransmission that results in a disruption of the excitatory/inhibitory (E/I) circuit balance tending to inhibit (xleschevinikova. m. et al, j. neurosci. (2004) 24: 8153-8160).

The term "cognitive deficit" or "cognitive impairment" describes any feature that is a disorder of cognitive performance. The term denotes a deficiency in the overall intellectual performance, such as mental retardation, which denotes a specific deficiency in cognitive ability (learning disability, dyslexia), or which denotes drug-induced memory impairment. Cognitive deficits may be congenital or caused by environmental factors such as brain injury, neurological disorders, or psychiatric disorders. The term "cognitive deficit in down syndrome" or "cognitive impairment in down syndrome" denotes a cognitive deficit in a subject exhibiting trisomy of chromosome 21, in particular an abnormality in learning, memory and language that results in a mild to extreme impairment of the mental function of said subject.

The term "intellectual impairment" (ID) or "mental retardation" means early onset cognitive impairment manifested by a significantly reduced ability to understand new information or complex information, learn new skills, and independently cope with the reduced ability, which begins before adulthood and has a lasting effect on development.

The term "pro-cognitive" describes any feature that alleviates or reverses a disorder such as confusion, disorientation, delirium, or cognitive deficits, or improves cognition.

The term "neurofibromatosis type 1" (NF1) denotes a disorder caused by mutation of a gene on chromosome 17 encoding a protein called neurofibromin involved in intracellular signaling (CuiY et al, Cell (2008) 1735: 549-60).

The term "autism" refers to a neurodevelopmental disorder characterized by impaired social interaction and communication and by limited and repetitive behaviors (american psychiatric associations inc., diagnostic manual of mental disorders (DSM-IV-TR) (2000), 4 th edition).

The term "stroke" is a rapidly progressing loss of brain function caused by a disturbance in the supply of blood to the brain. This can be caused by ischemia (lack of blood flow) or hemorrhage due to occlusion (thrombosis, arterial embolism) (SimsNR et al, Biochimica tBiophysica acta (2009)1802 (1): 80-91).

The term "post-stroke recovery" refers to the ability to recover impaired brain function after stroke (dimyanm.a. et al, NatRevNeurol (2011)7 (2): 76-85).

The term "treating" of a disease condition includes (1) preventing the disease condition, even if clinical symptoms of the disease condition do not develop in a subject that may be exposed to the disease condition or predisposed to the disease condition, but does not yet experience or exhibit symptoms of the disease condition, (2) inhibiting the disease condition, i.e., arresting the development of the disease condition or its clinical symptoms, or (3) alleviating the disease condition, i.e., causing temporary or permanent regression of the disease condition or its clinical symptoms.

The term "therapeutically effective amount" means the amount of a compound of the invention that, when administered to a subject, achieves the following effect: (i) treating or preventing a particular disease, condition, or disorder, (ii) attenuating, ameliorating, or eliminating one or more symptoms of a particular disease, condition, or disorder, or (iii) preventing or delaying the onset of one or more symptoms of a particular disease, condition, or disorder described herein. The therapeutically effective amount will vary depending on the following factors: the compound, the condition being treated, the severity of the condition being treated, the age and relative health of the subject, the route and form of administration, the judgment of the attending physician or veterinary practitioner, and other factors.

The term "subject" or "patient" means an animal, more particularly a vertebrate. In certain embodiments, the vertebrate is a mammal. Mammals including humans; non-human primates, such as chimpanzees and other apes and monkey species; farm animals such as cattle, horses, sheep, goats, and pigs; domestic animals such as rabbits, dogs, and cats; laboratory animals, including rodents, such as rats, mice and guinea pigs. In certain embodiments, the mammal is a human. The term subject does not denote a particular age or gender.

Detailed Description

In particular, the present invention relates to the use of GABAA α 5 negative allosteric modulators for the treatment, prevention and/or delay of progression of Central Nervous System (CNS) disorders caused by neurodevelopmental defects leading to excessive gabaergic inhibition in the cortex and hippocampus.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect resulting in excessive gabaergic inhibition in the cortex and hippocampus, wherein said CNS disorder is selected from the group consisting of cognitive deficit in down syndrome, cognitive deficit in autism, cognitive deficit in neurofibromatosis type I or cognitive deficit after stroke.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect resulting in excessive gabaergic inhibition in the cortex and hippocampus, wherein said CNS disorder is selected from the group consisting of cognitive deficits in down syndrome.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect resulting in excessive gabaergic inhibition in the cortex and hippocampus, wherein said CNS disorder is selected from the group consisting of autism cognitive deficit.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect resulting in excessive gabaergic inhibition in the cortex and hippocampus, wherein said CNS disorder is selected from the group consisting of type I neurofibromatosis cognitive deficits.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect resulting in excessive gabaergic inhibition in the cortex and hippocampus, wherein said CNS disorder is selected from post-stroke cognitive deficits.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect resulting in excessive gabaergic inhibition in the cortex and hippocampus, wherein said CNS disorder is selected from the group consisting of intellectual impairment.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator, wherein said GABAA α 5 negative allosteric modulator is a ligand of the BZD binding site and acts as a inverse agonist of the GABAA receptor containing the α 5 subunit.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is a ligand of the BZD binding site of the GABAA receptor and is an inverse agonist of the GABAA α 5 β 3 γ 2 receptor subtype.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator has binding selectivity for a GABAA receptor comprising an α 5 subunit.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is functionally selective for a GABAA receptor comprising an α 5 subunit.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator binds to the human GABAA α 5 β 3 γ 2 receptor subtype with a binding selectivity greater than 10 times the binding affinity for the human GABAA α 1 β 2/3 γ 2, α 2 β 3 γ 2 and α 3 β 3 γ 2 receptor subtypes.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator has binding selectivity for a GABAA receptor comprising an α 5 subunit.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator exhibits functional selectivity as follows: as inverse agonists of the human GABAA α 5 β 3 γ 2 receptor subtype, the effect of GABA is reduced by more than 30% and additionally the GABA effect is less than 15% on the human GABAA α 1 β 2/3 γ 2, α 2 β 3 γ 2 and α 3 β 3 γ 2 receptor subtypes.

In a particular embodiment, the present invention relates to the use of a GABAA alpha 5 negative allosteric modulator, wherein said GABAA alpha 5 negative allosteric modulator is selected from a compound of formula (I) or a compound of formula (II) or a pharmaceutically acceptable salt thereof,

wherein

R¹Is hydrogen, halogen, alkyl, haloalkyl or cyano;

R²is hydrogen, halogen, alkyl, haloalkyl or cyano;

R³is hydrogen, alkyl or heterocycloalkylalkyl, wherein the heterocycloalkylalkyl is optionally substituted with one or more of the following substituents: hydroxy, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, halogen or cyano;

R⁴is aryl or heteroaryl, each optionally substituted with 1, 2 or 3 halogens;

R⁵is hydrogen, alkyl, haloalkyl or hydroxyalkyl;

R⁶is-C (O) -NR⁷R⁸

R⁷Is hydrogen;

R⁸is an alkyl group;

or R⁷And R⁸Together with the nitrogen to which they are attached form a heterocycloalkyl or heteroaryl, each of which is optionally substituted with one or more of the following substituents: hydroxy, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, halogen or cyano.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of a compound of formula (I) or a pharmaceutically acceptable salt thereof, wherein

R¹Is hydrogen, halogen, alkyl, haloalkyl or cyano;

R²is hydrogen, halogen, alkyl, haloalkyl or cyano;

R³is hydrogen, alkyl or heterocycloalkylalkyl, wherein the heterocycloalkylalkyl is optionally substituted with one or more of the following substituents: hydroxy, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, halogen or cyano.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of a compound of formula (I) or a pharmaceutically acceptable salt thereof, wherein

R¹Is hydrogen, halogen, haloalkyl or cyano;

R²is halogen or haloalkyl;

R³is hydrogen, alkyl or heterocycloalkylalkyl substituted with 1 oxo.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of:

3-fluoro-10-fluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-bromo-10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-cyano-10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-chloro-10-fluoromethyl-6-methyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-chloro-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3, 10-dichloro-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-chloro-3-cyano-6-methyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-chloro-3-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-bromo-10-chloro-6-methyl-9H-imidazo [1, 5-a][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-bromo-3-fluoro-9H-imidazo [1, 5-a][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-bromo-10-methyl-6- (2-oxo-pyrrolidin-1-ylmethyl) -9H-imidazo [1, 5-a][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepineOr a pharmaceutically acceptable salt thereof.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is 3-bromo-10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepineOr a pharmaceutically acceptable salt thereof.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is not 3-bromo-10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepineOr a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of a compound of formula (II) or a pharmaceutically acceptable salt thereof, wherein

R⁴Is aryl orHeteroaryl, each of said aryl or heteroaryl being optionally substituted with 1, 2 or 3 halogens;

R⁵is hydrogen, alkyl, haloalkyl or hydroxyalkyl;

R⁶is-C (O) -NR⁷R⁸；

R⁷Is hydrogen;

R⁸is an alkyl group;

or R⁷And R⁸Together with the nitrogen to which they are attached form a heterocycloalkyl or heteroaryl, each of which is optionally substituted with one or more of the following substituents: hydroxy, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, halogen or cyano.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of a compound of formula (II) or a pharmaceutically acceptable salt thereof, wherein

R⁴Is aryl or heteroaryl, each optionally substituted with 1 halogen;

R⁵is an alkyl group;

R⁶is C (O) -NR⁷R⁸；

R⁷Is hydrogen, and R⁸Is an alkyl group;

or R⁷And R⁸Together with the nitrogen to which they are attached, form a heterocycloalkyl group optionally substituted by 1 or 2 oxo groups, or form a heteroaryl group.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of:

n-isopropyl-6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -nicotinamide;

(5, 6-dihydro-8H- [1, 2, 4] triazolo [4, 3-a ] pyrazin-7-yl) - [6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -methanone;

[6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] - (2-oxa-6-aza-spiro [3.3] hept-6-yl) -methanone;

(1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (4-fluoro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone;

{6- [3- (4-chloro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -morpholin-4-yl-methanone;

[6- (5-methyl-3-pyridin-2-yl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -morpholin-4-yl-methanone;

6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -isopropyl-nicotinamide;

(1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone;

{6- [3- (5-chloro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -thiomorpholin-4-yl-methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator, wherein said GABAA α 5 negative allosteric modulator is N-isopropyl-6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -nicotinamide; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator, wherein said GABAA α 5 negative allosteric modulator is (5, 6-dihydro-8H- [1, 2, 4] triazolo [4, 3-a ] pyrazin-7-yl) - [6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator, wherein said GABAA α 5 negative allosteric modulator is [6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] - (2-oxa-6-aza-spiro [3.3] hept-6-yl) -methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is (1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (4-fluoro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator, wherein said GABAA α 5 negative allosteric modulator is {6- [3- (4-chloro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -morpholin-4-yl-methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator, wherein said GABAA α 5 negative allosteric modulator is [6- (5-methyl-3-pyridin-2-yl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -morpholin-4-yl-methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator, wherein said GABAA α 5 negative allosteric modulator is 6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -isopropyl-nicotinamide; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is (1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator, wherein said GABAA α 5 negative allosteric modulator is {6- [3- (5-chloro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -thiomorpholin-4-yl-methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator wherein said GABAA α 5 negative allosteric modulator is used alone or in combination with a second active pharmaceutical compound, sequentially or simultaneously.

In a particular embodiment, the present invention relates to a method for treating, preventing and/or delaying the progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect resulting in excessive gabaergic inhibition in the cortex and hippocampus in a subject in need of such treatment, comprising: administering to the subject a therapeutically effective amount of a pharmaceutically acceptable form of a GABAA α 5 negative allosteric modulator as described herein.

In a particular embodiment, the present invention relates to a method for treating, preventing and/or delaying the progression of a Central Nervous System (CNS) disorder in a subject in need of such treatment, wherein said CNS disorder is selected from the group consisting of down syndrome cognitive deficits, said method comprising: administering to the subject a therapeutically effective amount of a pharmaceutically acceptable form of a GABAA α 5 negative allosteric modulator as described herein.

In a specific embodiment, the present invention relates to a method for treating, preventing and/or delaying the progression of a Central Nervous System (CNS) disorder in a subject in need of such treatment, wherein the CNS disorder is selected from the group consisting of autism cognitive deficit, the method comprising: administering to the subject a therapeutically effective amount of a pharmaceutically acceptable form of a GABAA α 5 negative allosteric modulator as described herein.

In a specific embodiment, the present invention relates to a method for treating, preventing and/or delaying the progression of a Central Nervous System (CNS) disorder in a subject in need of such treatment, wherein said CNS disorder is selected from the group consisting of type I neurofibromatosis cognitive deficits, said method comprising: administering to the subject a therapeutically effective amount of a pharmaceutically acceptable form of a GABAA α 5 negative allosteric modulator as described herein.

In a specific embodiment, the present invention relates to a method for treating, preventing and/or delaying the progression of a Central Nervous System (CNS) disorder in a subject in need of such treatment, wherein said CNS disorder is selected from the group consisting of post-stroke cognitive deficits, said method comprising: administering to the subject a therapeutically effective amount of a pharmaceutically acceptable form of a GABAA α 5 negative allosteric modulator as described herein.

In a particular embodiment, the present invention relates to a method for treating, preventing and/or slowing the progression of a Central Nervous System (CNS) disorder in a subject in need of such treatment, wherein the CNS disorder is selected from the group consisting of intellectual impairment (disability), the method comprising: administering to the subject a therapeutically effective amount of a pharmaceutically acceptable form of a GABAA α 5 negative allosteric modulator as described herein.

In a particular embodiment, the present invention relates to a pharmaceutical composition comprising a GABA a α 5 negative allosteric modulator as described herein in a pharmaceutically acceptable form for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect resulting in excessive gabaergic inhibition in the cortex and hippocampus.

In a specific embodiment, the present invention relates to a pharmaceutical composition comprising a GABAA α 5 negative allosteric modulator as described herein in a pharmaceutically acceptable form for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of cognitive deficits in down syndrome.

In a specific embodiment, the present invention relates to a pharmaceutical composition comprising a gaba a α 5 negative allosteric modulator as described herein in a pharmaceutically acceptable form for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of autism cognitive deficit.

In a specific embodiment, the present invention relates to a pharmaceutical composition comprising a gaba a α 5 negative allosteric modulator as described herein in a pharmaceutically acceptable form for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of type I neurofibromatosis cognitive deficits.

In a specific embodiment, the present invention relates to a pharmaceutical composition comprising a gaba a α 5 negative allosteric modulator as described herein in a pharmaceutically acceptable form for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of post-stroke cognitive deficits.

In a specific embodiment, the present invention relates to a pharmaceutical composition comprising a gaba a α 5 negative allosteric modulator as described herein in a pharmaceutically acceptable form for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of a intellectual impairment.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described herein for the preparation of a medicament for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect resulting in excessive gabaergic inhibition in the cortex and hippocampus.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described herein for the preparation of a medicament for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of down's syndrome cognitive deficits.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described herein for the preparation of a medicament for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of autism cognitive deficit.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described herein for the preparation of a medicament for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of type I neurofibromatosis cognitive deficits.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described herein for the preparation of a medicament for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from post-stroke cognitive deficits.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described herein for the preparation of a medicament for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of a intellectual disorder.

In a particular embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of Central Nervous System (CNS) disorders caused by neurodevelopmental defects leading to excessive gabaergic inhibition in the cortex and hippocampus.

In a particular embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of down's syndrome cognitive deficits.

In a particular embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of autism cognitive deficit.

In a particular embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of type I neurofibromatosis cognitive deficits.

In a particular embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of post-stroke cognitive deficits.

In one embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator as described herein for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, wherein said CNS disorder is selected from the group consisting of intellectual impairment, in one embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder, said Central Nervous System (CNS) disorder being caused by a neurodevelopmental defect resulting in excessive GABAergic inhibition in the cortex and hippocampus, wherein said CNS disorder is selected from the group consisting of cognitive deficits in Down syndrome, and wherein said GABAA α 5 negative allosteric modulator is 3-bromo-10-difluoromethyl-9H-imidazo [1, 5-a ] 1][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepineOr a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect leading to excessive gabaergic inhibition in the cortex and hippocampus, wherein said CNS disorder is selected from the group consisting of cognitive deficits in down syndrome, and wherein said GABAA α 5 negative allosteric modulator is (1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (4-fluoro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect resulting in excessive gabaergic inhibition in the cortex and hippocampus, wherein said CNS disorder is selected from the group consisting of cognitive deficits in down syndrome, and wherein said GABAA α 5 negative allosteric modulator is 3-bromo-10-difluoromethyl-9H-imidazo [1, 5-a ] and][1，2，4]triazolo [1, 5-d][1，4]BenzodiazepineOr a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect leading to excessive gabaergic inhibition in the cortex and hippocampus, wherein said CNS disorder is selected from the group consisting of cognitive deficits in down syndrome, and wherein said GABAA α 5 negative allosteric modulator is (1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (4-fluoro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone; or a pharmaceutically acceptable salt thereof.

In another embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder associated with excessive gabaergic inhibition in the cortex and hippocampus, said GABAA α 5 negative allosteric modulator being selected from a compound of formula (I) or a compound of formula (II) or a pharmaceutically acceptable salt thereof

Wherein

R¹Is hydrogen, halogen, alkyl, haloalkyl or cyano；

R²Is hydrogen, halogen, alkyl, haloalkyl or cyano;

R³is hydrogen, alkyl or heterocycloalkylalkyl, wherein the heterocycloalkylalkyl is optionally substituted with one or more of the following substituents: hydroxy, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, halogen or cyano;

R⁴is aryl or heteroaryl, each optionally substituted with 1, 2 or 3 halogens;

R⁵is hydrogen, alkyl, haloalkyl or hydroxyalkyl;

R⁶is-C (O) -NR⁷R⁸

R⁷Is hydrogen;

R⁸is an alkyl group;

or R⁷And R⁸Together with the nitrogen to which they are attached form a heterocycloalkyl or heteroaryl, each of which is optionally substituted with one or more of the following substituents: hydroxy, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, halogen or cyano.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described above for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder associated with excessive gabaergic inhibition in the cortex and hippocampus, wherein said excessive gabaergic inhibition in the cortex and hippocampus is caused by a neurodevelopmental defect.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described above for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder caused by a neurodevelopmental defect resulting in excessive gabaergic inhibition in the cortex and hippocampus.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described above for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder associated with excessive gabaergic inhibition in the cortex and hippocampus, in particular caused by a neurodevelopmental defect, wherein said CNS disorder is selected from the group consisting of cognitive deficit in down syndrome, cognitive deficit in autism or cognitive deficit in neurofibromatosis type I.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described above for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder associated with excessive gabaergic inhibition in the cortex and hippocampus, in particular caused by a neurodevelopmental defect, wherein said CNS disorder is a cognitive deficit in down syndrome.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described above for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder associated with excessive gabaergic inhibition in the cortex and hippocampus, in particular caused by a neurodevelopmental defect, wherein said CNS disorder is an autism cognitive deficit.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described above for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder associated with excessive gabaergic inhibition in the cortex and hippocampus, in particular caused by a neurodevelopmental defect, wherein said CNS disorder is a cognitive deficit of neurofibromatosis type I.

In a specific embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator as described above for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder associated with excessive gabaergic inhibition in the cortex and hippocampus, wherein said CNS disorder is characterized by post-stroke disability.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator binds to the human GABAA α 5 β 3 γ 2 receptor subtype with a binding selectivity which is more than 10 times the binding affinity for the human GABAA α 1 β 2/3 γ 2, α 2 β 3 γ 2 and α 3 β 3 γ 2 receptor subtypes.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator exhibits functional selectivity as follows: as inverse agonists of the human GABAA α 5 β 3 γ 2 receptor subtype, the effect of GABA is reduced by more than 30% and additionally the GABA effect is less than 15% on the human GABAA α 1 β 2/3 γ 2, α 2 β 3 γ 2 and α 3 β 3 γ 2 receptor subtypes.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is selected from a compound of formula (I) or a pharmaceutically acceptable salt thereof, wherein R is¹、R²And R³As defined herein.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is selected from a compound of formula (I) or a pharmaceutically acceptable salt thereof, wherein R is¹Is hydrogen, halogen, haloalkyl or cyano; r²Is halogen or haloalkyl; r³Is hydrogen, alkyl or heterocycloalkylalkyl substituted with 1 oxo.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of:

3-fluoro-10-fluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-bromo-10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-cyano-10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-chloro-10-fluoromethyl-6-methyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-chloro-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3, 10-dichloro-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-chloro-3-cyano-6-methyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-chloro-3-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-bromo-10-chloro-6-methyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-bromo-3-fluoro-9H-imidazo [1, 5-a][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-bromo-10-methyl-6- (2-oxo-pyrrolidin-1-ylmethyl) -9H-imidazo [1, 5-a][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepineOr a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is selected from a compound of formula (II) or a pharmaceutically acceptable salt thereof, wherein R is⁴、R⁵And R⁶As defined herein.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described hereinWherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of compounds of formula (II) or a pharmaceutically acceptable salt thereof, wherein R⁴Is aryl or heteroaryl, each optionally substituted with 1 halogen; r⁵Is an alkyl group; r⁶Is C (O) -NR⁷R⁸；R⁷Is hydrogen, and R⁸Is an alkyl group; or R⁷And R⁸Together with the nitrogen to which they are attached, form a heterocycloalkyl group optionally substituted by 1 or 2 oxo groups, or form a heteroaryl group.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of:

n-isopropyl-6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -nicotinamide;

(5, 6-dihydro-8H- [1, 2, 4] triazolo [4, 3-a ] pyrazin-7-yl) - [6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -methanone;

[6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] - (2-oxa-6-aza-spiro [3.3] hept-6-yl) -methanone;

(1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (4-fluoro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone;

{6- [3- (4-chloro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -morpholin-4-yl-methanone;

[6- (5-methyl-3-pyridin-2-yl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -morpholin-4-yl-methanone:

6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -isopropyl-nicotinamide;

(1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone;

{6- [3- (5-chloro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -thiomorpholin-4-yl-methanone;

or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is N-isopropyl-6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -nicotinamide; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is (5, 6-dihydro-8H- [1, 2, 4] triazolo [4, 3-a ] pyrazin-7-yl) - [6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is [6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] - (2-oxa-6-aza-spiro [3.3] hept-6-yl) -methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is (1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (4-fluoro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is {6- [3- (4-chloro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -morpholin-4-yl-methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is [6- (5-methyl-3-pyridin-2-yl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -morpholin-4-yl-methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is 6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -isopropyl-nicotinamide; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is (1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is {6- [3- (5-chloro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -thiomorpholin-4-yl-methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to the use of a GABAA α 5 negative allosteric modulator for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is used alone, or in combination, sequentially or simultaneously, with a second active pharmaceutical compound.

In a particular embodiment, the present invention relates to a method for treating, preventing and/or delaying the progression of cognitive deficits in down syndrome, cognitive deficits in autism, cognitive deficits in neurofibromatosis type I, or for post-stroke recovery in a subject in need of such treatment, comprising: administering to the subject a therapeutically effective amount of a pharmaceutically acceptable form of a GABAA α 5 negative allosteric modulator as described herein.

In a particular embodiment, the present invention relates to a pharmaceutical composition comprising a GABAA α 5 negative allosteric modulator as described herein in a pharmaceutically acceptable form for use in the treatment, prevention and/or delay of progression of cognitive deficits in down syndrome, cognitive deficits in autism, cognitive deficits in neurofibromatosis type I, or for post-stroke recovery.

In a particular embodiment, the invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of cognitive deficits in down syndrome, cognitive deficits in autism, cognitive deficits in neurofibromatosis type I, or for post-stroke recovery.

In a particular embodiment, the invention relates to a GABAA α 5 negative allosteric modulator as described herein for the preparation of a medicament for the treatment, prevention and/or delay of progression of cognitive deficits in down syndrome, cognitive deficits in autism, cognitive deficits in neurofibromatosis type I, or for post-stroke recovery.

In a particular embodiment, the invention relates to the use of a GABAA α 5 negative allosteric modulator as described herein for the preparation of a medicament for the treatment, prevention and/or delay of progression of cognitive deficits in down syndrome, cognitive deficits in autism, cognitive deficits in neurofibromatosis type I, or for post-stroke recovery.

In another embodiment, the invention relates to a GABAA alpha 5 negative allosteric modulator selected from the group consisting of a compound of formula (I) or a compound of formula (II) or a pharmaceutically acceptable salt thereof,

wherein

R¹Is hydrogen, halogen, alkyl, haloalkyl or cyano;

R²is hydrogen, halogen, alkyl, haloalkyl or cyano;

R³is hydrogen, alkyl or heterocycloalkylalkyl, wherein the heterocycloalkylalkyl is optionally substituted with one or more of the following substituents: hydroxy, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, halogen or cyano;

R⁴is aryl or heteroaryl, each optionally substituted with 1, 2 or 3 halogens;

R⁵is hydrogen, alkyl, haloalkyl or hydroxyalkyl;

R⁶is-C (O) -NR⁷R⁸

R⁷Is hydrogen;

R⁸is an alkyl group;

or R⁷And R⁸Together with the nitrogen to which they are attached form a heterocycloalkyl or heteroaryl, each of which is optionally substituted with one or more of the following substituents: hydroxy, oxo, alkyl, alkoxy, haloalkyl, hydroxyalkyl, halogen or cyano;

the GABAA α 5 negative allosteric modulators are useful for the treatment, prevention and/or delay of progression of Central Nervous System (CNS) disorders associated with excessive gabaergic inhibition in the cortex and hippocampus.

In a particular embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of Central Nervous System (CNS) disorders associated with excessive gabaergic inhibition in the cortex and hippocampus caused by a deficit in neurodevelopment.

In a particular embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of Central Nervous System (CNS) disorders caused by neurodevelopmental defects leading to excessive gabaergic inhibition in the cortex and hippocampus.

In a particular embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder associated with excessive gabaergic inhibition in the cortex and hippocampus, in particular caused by a neurodevelopmental defect, wherein said CNS disorder is selected from the group consisting of cognitive deficit in down syndrome, cognitive deficit in autism or cognitive deficit in neurofibromatosis type I.

In a particular embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder associated with excessive gabaergic inhibition in the cortex and hippocampus, in particular caused by a neurodevelopmental defect, wherein said CNS disorder is a cognitive deficit in down syndrome.

In a particular embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder associated with excessive gabaergic inhibition in the cortex and hippocampus, in particular caused by a neurodevelopmental defect, wherein said CNS disorder is an autism cognitive deficit.

In a particular embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder associated with excessive gabaergic inhibition in the cortex and hippocampus, in particular caused by a neurodevelopmental defect, wherein said CNS disorder is a cognitive deficit of neurofibromatosis type I.

In a specific embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder associated with excessive gabaergic inhibition in the cortex and hippocampus, wherein said CNS disorder is characterized by post-stroke disability.

In a particular embodiment, the invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator binds to the human GABAA α 5 β 3 γ 2 receptor subtype with a binding selectivity which is more than 10 times the binding affinity for the human GABAA α 1 β 2/3 γ 2, α 2 β 3 γ 2 and α 3 β 3 γ 2 receptor subtypes.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator exhibits functional selectivity as follows: as inverse agonists of the human GABAA α 5 β 3 γ 2 receptor subtype, the effect of GABA is reduced by more than 30% and additionally the GABA effect is less than 15% on the human GABAA α 1 β 2/3 γ 2, α 2 β 3 γ 2 and α 3 β 3 γ 2 receptor subtypes.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein,wherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of compounds of formula (I) or a pharmaceutically acceptable salt thereof, wherein R is¹、R²And R³As defined herein.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is selected from a compound of formula (I) or a pharmaceutically acceptable salt thereof, wherein R is¹Is hydrogen, halogen, haloalkyl or cyano; r²Is halogen or haloalkyl; r³Is hydrogen, alkyl or heterocycloalkylalkyl substituted with 1 oxo.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of:

3-fluoro-10-fluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-bromo-10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-cyano-10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-chloro-10-fluoromethyl-6-methyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-chloro-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3, 10-dichloro-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-chloro-3-cyano-6-methyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-chloro-3-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-bromo-10-chloro-6-methyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

10-bromo-3-fluoro-9H-imidazo [1, 5-a][1，2，4]Triazolo [1, 5-d][1，4]Benzodiazepine

3-bromo-10-methyl-6- (2-oxo-pyrrolidin-1-ylmethyl) -9H-imidazo [1, 5-a][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepineOr a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is 3-bromo-10-difluoromethyl-9H-imidazo [1, 5-a ] 1][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepineOr a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is not 3-bromo-10-difluoromethyl-9H-imidazo [1, 5-a ] or a pharmaceutically acceptable salt thereof][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepineOr a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is selected from a compound of formula (II) or a pharmaceutically acceptable salt thereof, wherein R is⁴、R⁵And R⁶As defined herein.

In a particular embodiment, the present invention relates to GABAA α 5 negative allosteric modulators as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described hereinWherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of compounds of formula (II) or a pharmaceutically acceptable salt thereof, wherein R⁴Is aryl or heteroaryl, each optionally substituted with 1 halogen; r⁵Is an alkyl group; r⁶Is C (O) -NR⁷R⁸；R⁷Is hydrogen, and R⁸Is an alkyl group; or R⁷And R⁸Together with the nitrogen to which they are attached, form a heterocycloalkyl group optionally substituted by 1 or 2 oxo groups, or form a heteroaryl group.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is selected from the group consisting of:

n-isopropyl-6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -nicotinamide;

(5, 6-dihydro-8H- [1, 2, 4] triazolo [4, 3-a ] pyrazin-7-yl) - [6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -methanone;

[6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] - (2-oxa-6-aza-spiro [3.3] hept-6-yl) -methanone;

(1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (4-fluoro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone;

{6- [3- (4-chloro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -morpholin-4-yl-methanone;

[6- (5-methyl-3-pyridin-2-yl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -morpholin-4-yl-methanone;

6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -isopropyl-nicotinamide;

(1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone;

{6- [3- (5-chloro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -thiomorpholin-4-yl-methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is N-isopropyl-6- (5-monomethyl-3-phenyl-isoxazol-4-ylmethoxy) -nicotinamide; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is (5, 6-dihydro-8H- [1, 2, 4] triazolo [4, 3-a ] pyrazin-7-yl) - [6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is [6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] - (2-oxa-6-aza-spiro [3.3] hept-6-yl) -methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is (1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (4-fluoro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is {6- [3- (4-chloro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -morpholin-4-yl-methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is [6- (5-methyl-3-pyridin-2-yl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -morpholin-4-yl-methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is 6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -isopropyl-nicotinamide; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is (1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is {6- [3- (5-chloro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -thiomorpholin-4-yl-methanone; or a pharmaceutically acceptable salt thereof.

In a particular embodiment, the present invention relates to a GABAA α 5 negative allosteric modulator as described herein for use in the treatment, prevention and/or delay of progression of a Central Nervous System (CNS) disorder as described herein, wherein said GABAA α 5 negative allosteric modulator is used alone or in combination, sequentially or simultaneously, with a second active pharmaceutical compound.

Examples

Materials and methods

a animals

Table 1 shows the number of male animals used in this study. Ten control and ten Ts65Dn6 month old mice received 8581 at the start of treatment; 16 control and 15 Ts65Dn5-6 month old mice received R1 at the start of treatment, and two additional groups of control (n ═ 13) and Ts65Dn (n ═ 13) mice received vehicle.

b. Active pharmaceutical compounds

The active pharmaceutical compounds used in the present invention are prepared as previously described in WO2006/045429, WO2006/045430, WO2007/042421 and WO 2009/071476:

compound 8580

3-fluoro-10-fluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepinePrepared as described in WO2006/045430, page 17, example 3.

Compound 8581

3-bromo-10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepinePrepared as described in WO2006/045430, page 21, example 7.

Compound 8582

3-cyano-10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepinePrepared as described in WO2006/045430, page 23, example 13.

Compound 8583

10-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepinePrepared as described in WO2006/045430, page 26, example 16.

Compound 8584

3-chloro-10-fluoromethyl-6-methyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepinePrepared as described in WO2006/045430, page 28, example 20.

Compound 8585

10-chloro-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepinePrepared as described in example 1, page 15 of WO 2006/045429.

Compound 8586

3, 10-dichloro-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepineAs described in example 20 on page 23 of WO2006/045429And (4) preparation.

Compound 8587

10-chloro-3-cyano-6-methyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepinePrepared as described in example 47, page 37 of WO 2006/045429.

Compound 8588

10-chloro-3-difluoromethyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepinePrepared as described in example 32, page 29 of WO 2006/045429.

Compound 8589

3-bromo-10-chloro-6-methyl-9H-imidazo [1, 5-a ]][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepinePrepared as described in WO2006/045429, page 33, example 38.

Compound 8590

10-bromo-3-fluoro-9H-imidazo [1, 5-a][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepinePrepared as described in example 47, page 37 of WO 2006/045429.

Compound 8591

3-bromo-10-methyl-6- (2-oxo-pyrrolidin-1-ylmethyl) -9H-imidazo [1, 5-a][1，2，4]Triazolo [1, 5-d][1，4]BenzodiazepinePrepared as described in example 101 on page 67 of WO 2007/042421.

Compound O1

N-isopropyl-6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -nicotinamide was prepared as described in WO2009/071476 page 50, example 26.

Compound P1

(5, 6-dihydro-8H- [1, 2, 4] triazolo [4, 3-a ] pyrazin-7-yl) - [6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -methanone was prepared as described in WO2009/071476 page 62, example 75.

Compound Q1

[6- (5-methyl-3-phenyl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] - (2-oxa-6-aza-spiro [3.3] hept-6-yl) -methanone was prepared as described in WO2009/071476 page 64, example 81.

Compound R1

(1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (4-fluoro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone was prepared as described in WO2009/071476 page 75, example 112.

Compound S1

{6- [3- (4-chloro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -morpholin-4-yl-methanone was prepared as described in WO2009/071476, page 78, example 123.

Compound T1

[6- (5-methyl-3-pyridin-2-yl-isoxazol-4-ylmethoxy) -pyridin-3-yl ] -morpholin-4-yl-methanone was prepared as described in WO2009/071476 page 123 example 274.

Compound U1

6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -isopropyl-nicotinamide was prepared as described in WO2009/071476 page 127 example 289.

Compound V1

(1, 1-dioxo-1 λ 6-thiomorpholin-4-yl) - {6- [3- (5-fluoro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone was prepared as described in WO2009/071476, page 127, example 293.

Compound W1

{6- [3- (5-chloro-pyridin-2-yl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -thiomorpholin-4-yl-methanone was prepared as described in WO2009/071476 page 132, example 310.

Has passed through³H]Flumazenil (85 Ci/mmol; Roche) competes for binding to HEK293 cells expressing the rat (stable transfection) or human (transient transfection) α 1 β 2/3 γ 2, β 02 β 13 γ 2, β 23 β 33 γ 2 and β 45 β 53 γ 2 constitutive receptors, measuring the binding affinity of the above active pharmaceutical compounds to GABAA receptor subtypes it can be seen from table 2a that the active pharmaceutical compounds used in the present invention exhibit high affinity for the α 5 β 3 γ 2 receptor subtype and good selectivity for the α 1 β 2/3 γ 2, α 2 β 3 γ 2 and α 3 β 3 γ 2 receptor subtypes.

As can be seen from table 2b, the active pharmaceutical compounds used in the present invention also exhibit substantial functional selectivity. Subtype selective effects were determined against cloned receptors expressed in Xenopus oocytes. Human recombinant GABAA receptors were expressed in xenopus laevis (xenopus laevis) oocytes. The amperometric response was induced by applying GABA of EC10 before and during the co-application of the test compound under a two microelectrode voltage clamp condition. The magnitude of the response in the presence of the test compound is expressed as a percentage of the magnitude before drug addition.

c. Pharmaceutical composition

For mouse studies, the active pharmaceutical compounds of the invention were formulated in chocolate-milk (Puleva, Barcelona, Spain). The active pharmaceutical compound or vehicle of the present invention was administered orally at a dose of 20mg/kg for 6 weeks. Their administration was delayed during the 30 day behavioral assessment period.

For human use, pharmaceutical compositions or medicaments comprising an active pharmaceutical compound as hereinbefore described together with a therapeutically inert carrier, diluent or excipient may be prepared, as well as methods of using the compounds of the invention to prepare such compositions and medicaments.

The compositions may be formulated, metered, and administered in a manner consistent with good medical practice. Factors to be considered in this regard include the particular condition being treated, the particular mammal being treated, the clinical condition of the individual patient, the etiology of the condition, the site of drug delivery, the method of administration, the timing of administration, and other factors known to the practitioner.

The active pharmaceutical compounds for use in the present invention may be administered by any suitable means, including oral, topical (including buccal and sublingual), rectal, vaginal, transdermal, parenteral, subcutaneous, intraperitoneal, intrapulmonary, intradermal, intrathecal and epidural and intranasal, and, if desired for topical treatment, intralesional administration. Parenteral infusion includes intramuscular, intravenous, intraarterial, intraperitoneal or subcutaneous administration.

The active pharmaceutical compounds used in the present invention may be administered in any convenient administration form, such as tablets, powders, capsules, solutions, dispersions, suspensions, syrups, sprays, suppositories, gels, emulsions, patches and the like. Such compositions may contain ingredients conventional in pharmaceutical formulations, such as diluents, carriers, pH adjusting agents, preservatives, solubilizers, stabilizers, wetting agents, emulsifiers, sweeteners, colorants, flavorants, salts for varying the osmotic pressure, buffers, masking agents, antioxidants, and additional active agents. They may also contain other therapeutically valuable substances.

Typical pharmaceutical compositions are prepared by mixing the active pharmaceutical compound used in the present invention with a carrier or excipient. Suitable carriers and excipients are well known to those skilled in the art and are described in detail, for example, in Ansel h.c. et al, Ansel's pharmaceutical dosage forms and drug delivery systems (2004) Lippincott, Williams & Wilkins, philiadelphia; gennaroa.r. et al, Remington: the science and practice of pharmacy (2000) Lippincott, Williams & Wilkins, philiadelphia; and rower.c, handbook of pharmaceutical excipients (2005) pharmaceutical press, Chicago. The pharmaceutical compositions may also include one or more buffering agents, stabilizing agents, surfactants, wetting agents, lubricating agents, emulsifying agents, suspending agents, preservatives, antioxidants, opacifiers, glidants, processing aids, colorants, sweeteners, fragrances, flavors, diluents and other known additives to provide an excellent appearance to the drug (i.e., a compound of the present invention or a pharmaceutical composition thereof) or to aid in the preparation of the drug product (i.e., a medicament).

The dosage of the active pharmaceutical compounds used in the present invention to be administered may vary within wide limits and will of course be adapted to the individual requirements of each particular case. Generally, in the case of oral administration, a daily dosage of about 0.1 to 1500mg, more specifically 1 to 1000mg, most specifically 5 to 500mg of the active pharmaceutical compound used according to the invention per person will be appropriate, although the above upper limit may also be exceeded if necessary.

An example of a suitable oral dosage form is a tablet comprising about 100mg to 500mg of the active pharmaceutical compound used in the present invention complexed with about 90-30mg of anhydrous lactose, about 5-40mg of croscarmellose sodium, about 5-30mg polyvinylpyrrolidone (PVP) K30 and about 1-10mg magnesium stearate. These powdered ingredients are first mixed together and then mixed with a solution of PVP. The resulting composition may be dried, granulated, mixed with magnesium stearate and compressed into tablet form using conventional equipment.

An example of an aerosol composition can be prepared as follows: for example 10-100mg of the active pharmaceutical compound used according to the invention is dissolved in a suitable buffer solution, for example phosphate buffer, and, if desired, a tonicity agent (tonifier), for example a salt such as sodium chloride, is added. The solution may be filtered, for example, using a 0.2gm filter, to remove impurities and contaminants.

d. Statistical analysis

Data were analyzed using a two-factor ("genotype" x "treatment") ANOVA. The Morris water maze data was analyzed using a two-way ANOVA with repeated measures ("phase" x "genotype" x "treatment"). Thereafter the mean values for each experimental group were compared by Student's t-test if two groups were compared, or by Bonferroni test if more than two groups were compared. All analyses were performed using SPSS for Windows version 17.0 (SPSSAG, zurich, switzerland).

Brief description of the tables

Table 1 experimental animal groups used in examples 1 to 6 in the present invention.

Table 2a. binding affinity and binding selectivity of active pharmaceutical compounds used in the present invention.

Table 2b modulation of GABAA receptor subtypes expressed in xenopus oocytes by active pharmaceutical compounds. Effects on human GABAA α 5 receptor: % change from the next to maximal (EC10) response to GABA determined by flumazenil binding assay at a value of 30 xKi. Effects on human GABAA α 1, α 2 and α 3 receptors: % change from the next to maximal (EC10) response to GABA determined by flumazenil binding assay at 3 μ M or at 30xKi values (if Ki is less than 0.1 μ M).

Table 3.8581, R1 and vehicle treated Ts65Dn and a sensorimotor test series of control mice (mean score ± s.e.m.).

Table 4 well plate test results (mean score ± s.e.m.) for R1, 8581 and vehicle treated Ts65Dn and control mice. **: p < 0.01Ts65Dn relative to control

TABLE 5 concentration of 8581 (ng/ml) in serum of Nfl +/-and control mice at 0.5, 3, 7 and 24 hours after intraperitoneal injection.

Drawings

Figure 1.8581, R1, and mean ± s.e.m. of the lag time of vehicle-treated Ts65Dn and control mice falling from the rotarod at different constant speeds.

Figure 2.8581, R1, and mean ± s.e.m. of the lag time of vehicle-treated Ts65Dn and control mice from the stalk fall during the acceleration cycle.

Figure 3 mean ± s.e.m. of spontaneous activity in their cages of Ts65Dn and control mice in a 24 hour full dark-light cycle under vehicle, R1 or 8581 treatment conditions.

Figure 4 mean ± s.e.m. of mean activity made by Ts65Dn and control mice during the light and dark phase of the cycle under vehicle, R1 or 8581 treatment conditions.

Figure 5 mean ± s.e.m. of the number of crossovers made in the center and periphery of the open field by R1, 8581 and vehicle treated Ts65Dn and control mice. *: p is less than 0.05; **: p < 0.01, Bonferroni test after significant ANOVA.

Figure 6 mean ± s.e.m. of hind leg stance times in open field of R1, 8581 and vehicle treated Ts65Dn and control mice.

Figure 7 mean ± s.e.m. of the delay times to reach the plateau in MWM during eight learning periods.

Figure 8 mean ± s.e.m. of the delay times to plateau during eight learning periods for vehicle (a), 8581(B) and R1 treated Ts65Dn and control mice and vehicle and 8581 treated Ts65Dn (C) and control (D) mice. *: p is less than 0.05; **: p is less than 0.01; ***: p < 0.001, T-test after ANOVA.

Figure 9 mean ± s.e.m. of the delay times to plateau during eight learning periods for vehicle and 8581 treated Ts65Dn (a) and control (B) mice and vehicle and R1 treated Ts65Dn (C) and control (D) mice. *: p is less than 0.05; **: p is less than 0.01; ***: p < 0.001, significant post ANOVA t-test.

Figure 10 mean ± s.e.m. of the delay times to reach the plateau during the cue period. *: p < 0.05, Ts65Dn relative to control; #: p is less than 0.05; # #: p < 0.01, 8581 and R1 relative to the excipient.

Figure 11. 8581 reverses long-term potentiation defects in hippocampal slices of Ts65Dn mice after long-term treatment. Data are presented as mean ± s.e.m. of evoked EPSPs recorded from hippocampal slices of vehicle, 8581 treated Ts65Dn (Ts) and Control (CO) mice. After a stable baseline phase of 20min, tonic stimulation was applied to hippocampal slices to induce LTP. The field ESPS slopes were normalized and presented as mean ± SEM (n-5-7/cohort). P < 0.05, relative to vehicle (V).

Figure 12R 1 reversed long-term potentiation in hippocampal slices of Ts65Dn mice after long-term treatment. Data are presented as mean ± s.e.m. of evoked EPSPs recorded from hippocampal slices of vehicle, R1 treated TS and CO mice. After a stable baseline phase of 20min, tonic stimulation was applied to hippocampal slices to induce LTP. The field ESPS slopes were normalized and presented as mean ± SEM (n-5-7/cohort). P < 0.05, relative to vehicle (V).

FIG. 13.8581 rescues neuronal proliferation in the hippocampus of TS and CO mice. Data are expressed as mean ± s.e.m. of Ki67+ cell density in vehicle and 8581 treated TS and CO mice. ANOVA "genotype": f (1, 20) ═ 7.39, p ═ 0.024; "treat": f (1, 20) ═ 6.30, p ═ 0.033; "genotype x treatment": f (1, 20) ═ 1.81, p ═ 0.21 ·: p < 0.01TS vs CO; #: p < 0.05, #': p < 0.01 vehicle vs 8581 treated mice; bonferroni test after significant ANOVA.

Figure 14.8581 rescued granular cell density in TS mice hippocampus. Data are expressed as mean ± s.e.m. (a) of DAPI + cell density in the granular cell layer of vehicle and 8581 treated TS and CO mice. ANOVA "genotype": f (1, 20) ═ 0.51, p ═ 0.49; "treat": f (1, 20) ═ 7.09, p ═ 0.026; "genotype x treatment": f (1, 20) ═ 4.00, p ═ 0.076. *: p < 0.05TS vs CO; # #: p < 0.01 vehicle vs 8581 treated mice; bonferroni test after significant ANOVA. (B) Vehicle and representative images of DAPI immunostaining of 8581 treated TS and CO mice.

FIG. 15.8581 normalizes the percentage area occupied by GAD + synapse nodules in the hippocampus of TS mice. Data are expressed as mean ± s.e.m. (a) percentage of GAD + synaptosomes in hippocampus of TS and CO mice treated with vehicle and 8581. ANOVA "genotype": f (1, 20) ═ 0.085, p ═ 0.77; "treat": f (1, 20) ═ 1.14, p ═ 0.30; "genotype x treatment": f (1, 20) ═ 7.15, and p ═ 0.017. *: p < 0.05TS vs CO; #: p < 0.05 vehicle vs 8581 treated mice; bonferroni test after significant ANOVA. (B) Representative images of GAD immunostaining of vehicle and RO 4938581-treated TS and CO mice.

FIG. 16.8581(1mg/kg) rescued the spatial learning deficit of Nfl +/-mice. The average percentage of time spent in each quadrant during the probe trial (QL: left side of target quadrant; QT: target quadrant; QR: right side of target quadrant; QO: opposite side of target quadrant).

FIG. 17.8581(1mg/kg) rescued the spatial learning deficit of Nfl +/-mice. Average proximity to the target platform during probe trials.

FIG. 18. 8581(1mg/kg) did not affect the performance of Nfl +/-mice under conditions that masked their behavioral deficits. The average percentage of time spent in each quadrant during the probe trial (QL: left side of target quadrant; QT: target quadrant; QR: right side of target quadrant; QO: opposite side of target quadrant).

FIG. 19. 8581(1mg/kg) does not affect the performance of Nfl +/-mice under conditions that mask their behavioral deficits. Average proximity to the target platform during probe trials.

FIG. 20. contextual conditioned reflex: dose response curves (0.3, 1.0 and 3.0mg/kg) for 8581-treated control mice (P < 0.05).

FIG. 21. contextual conditioned reflex in control mice when administered 8581(1mg/kg) continuously for two days.

FIG. 22 Performance on the rotarod of vehicle or 8581(1mg/kg) treated control mice.

FIG. 23. Nfl +/-mice treated with vehicle or 8581(1mg/kg) performed on a rotating rod.

Abbreviations

ANOVA ═ analysis of variance

BZD ═ benzodiazepines

CNS (central nervous system)

Control of CO

Down syndrome with DS

F ═ F-test value

GABA (gamma-aminobutyric acid)

i.p. ═ intraperitoneally

LTP-long term enhancement

MANOVA ═ multivariate analysis of variance

MWM (Morris water maze)

probability of p ═ probability

p.o. ═ oral

Standard error of mean (s.e.m.)

TS＝Ts65Dn

veh excipient

Example 1 sensorimotor testing

Following RuedaN et al, [ neurosci lett (2008)433 (1): 22-27] performing a series of sensorimotor tests. Cerebellum and vestibular function were evaluated in the visual placement reflex test. In 3 consecutive trials, mice were gently dropped from a height of 15cm with the tail towards a flat surface. Paw extension responses were scored on a 0-4 scale [ 4: the animal extends the forepaw when positioned at the highest elevation; 3: the anterior paw is extended before the whisker contacts the surface; 2: the anterior paw extends behind the tentacle contact surface; 1: extension of the anterior paw after the nose contacts the surface; 0: not stretched ].

The startle response to sudden auditory stimuli was measured to evaluate auditory sensitivity. The mouse was placed facing the wall of an unfamiliar cage and auditory stimuli were generated by bumping two stainless steel tweezers (7cm long) together. A score (0-3 points) was assigned based on the magnitude of the response: jumping is larger than 1cm (3 minutes); jump less than 1cm (2 minutes); ear retraction (puril reflex, 1 point); or no reaction (0 min).

Whisker placement reflections were analyzed by noting the reflection response to touching the whisker with a cotton swab. Animals exposed to the stimulated whisker with ipsilateral paw in three consecutive trials were assigned a score of 1 and 0 if unresponsive.

The grip was evaluated by quantifying the resistance when pulling the tail to separate it from the aluminium bar mesh covers (2mm) (0: no resistance, complete loss of grip; 1: slight; 2: medium; 3: powerful; 4: very strong, normal grip).

To evaluate the equilibration capacity, four 20 second equilibration tests were performed on elevated (40 cm high) level (50 cm long) rods. Tests 1 and 2 were carried out on flat wooden bars (width 9 mm); tests 3 and 4 were carried out on cylindrical aluminum rods (1 cm diameter). In each experiment, animals were placed in a marked central zone (10cm) on an elevated rod. If the animal fell within 20 seconds, a score of 0 was given; score 1 if it stays in the central zone for more than 20 seconds; score 2 if it leaves the central zone; and if it reaches one end of the stick, it is scored as 3.

The grab reflex (three 5 second trials) was measured as the ability of the animal to remain suspended by grasping the elevated horizontal wire (2mm diameter) with the forepaws. The maximum possible score of 3(1 point per test) was obtained when the animals were kept suspended with the forepaws in all three tests. Traction capacity (traction capacity) was also scored by assessing the number of times the animal lifted the hind limb to reach the wire (0: none; 1: one limb; 2: two limbs).

Table 3 shows the scores of 8581, R1 and vehicle treated Ts65Dn and control mice in different sensorimotor tests. 8581 or R1 treatment did not alter either of the Ts65Dn or the tested sensorimotor performance (visual, auditory, strength, balance ability, grip reflex, traction ability or motor coordination in the coat hanger hanging test (coathangtest)) of the control mice.

Example 2. movement coordination: rotating rod

The coordination of the movements was evaluated using a rotating rod device (UgoBasile; Comerio, Italy) consisting of a plastic rod 37cm long and 3cm in diameter, which was rotated at different speeds. In one session, 4 trials were performed, each with a maximum duration of 60 seconds. In the first three tests, the rods were rotated at constant speeds of 5, 25 and 50rpm, respectively. The last trial consisted of an acceleration cycle in which the rod was rotated progressively faster and the animals had to adapt to progressively higher test requirements. The length of time each animal stayed on the rotating bar was recorded.

As shown in figures 1 and 2, after 8581 or R1 treatment, the motor coordination on the rotarod was unchanged for either genotype mouse. Ts65Dn and control mice showed no difference in the delay time (1atency) of the rod fall at different constant speeds (ANOVA "genotype": vel 2: F (1, 76) ═ 0.63, p ═ 0.42; vel 3: F (1, 76) ═ 1.54, p ═ 0.21) or during the acceleration period (F (1, 76) ═ 1.87, p ═ 0.17)).

In addition, no 8581 or R1 and vehicle treated Ts65Dn or control mice were found at different constant rates (ANOVA "treatment" vel 2: F (1, 76) ═ 0.08, p ═ 0.92); vel 3: there is a difference in fall delay time for F (1, 76) ═ 1.42, p ═ 0.24) or during the acceleration period (F (1, 76) ═ 1.40, p ═ 0.25).

MANOVA revealed that the factors "genotype" and "treatment" had no significant interplay under any of the rotarod test conditions (vel 2: F (1, 76) ═ 0.31, p ═ 0.72, vel 3: F (1, 76) ═ 0.48, p ═ 0.61, acceleration. F (1, 76) ═ 0.43, p ═ 0.64).

Example 3. spontaneous Activity: activity measurement (Actimetry)

In this test, the circadian variation of the animal's spontaneous motor activity during a complete 24-hour light-dark cycle is evaluated. The apparatus is a device (Acti-system II, Panlab, Barcelona) that detects changes in magnetic field caused by movement of a mouse. It records the movement of the animals during a continuous 24 hour period (12 hours light and 12 hours dark).

Figures 3 and 4 show that Ts65Dn and control mice (ANOVA "genotype" dark: F (1, 76) ═ 2.79, p ═ 0.10; bright: F (1, 76) ═ 2.24, p ═ 0.14) have no difference in the amount of spontaneous activity made during the dark or light phase of the cycle in their cages under vehicle, R1 or 8581 treatment conditions (ANOVA "treatment" dark: F (1, 76) ═ 2.20, p ═ 0.12, bright: F (1, 76) ═ 0.27, p ═ 0.76; ANOVA "genotype x treatment": dark: F (1, 76) ═ 0.79, p ═ 0.45; bright: F (1, 76) ═ 0.39, p ═ 0.67).

Example 4 open field

Exploratory behavior and anxiety were evaluated using a square shaped open field (55cmx55cm, surrounded by a 25cm tall fence and divided into 25 equal squares). Animals were placed in the center of the field and scored for the number of vertical (hind leg standing) movements and horizontal crossings (between squares, subdivided into center vs peripheral crossings) in one 5 minute trial.

In this open field test, no significant difference was found in the activity of the two genotyped mice in the center of the maze (ANOVA "genotype: F (1, 76) ═ 2.77, p ═ 0.10; fig. 5) or the number of hind legs (F (1, 76) ═ 0.01, p ═ 0.90; fig. 6). However, vehicle treated Ts65Dn mice were overactive under the same treatment when compared to control mice, as shown by increased peripheral activity (ANOVA "genotype": F (1, 76) ═ 15.86, p < 0.001; fig. 5) and total activity (F (1, 76) ═ 17.39, p < 0.001; fig. 6).

MANOVA revealed that R1 or 8581 treatment did not significantly affect the level of either genotype mice (ANOVA "treatment": peripheral: F (1, 76) ═ 1.08, p ═ 0.34; total number of crossings: F (1, 76) ═ 1.27, p ═ 0.28) or vertical (hind leg standing F (1, 76) ═ 1.75, p ═ 0.18) activity. The fact that long term administration of the two compounds did not affect the activity of the maze center (ANOVA "treatment": center: F (1, 76) ═ 2.42, p ═ 0.096) indicates that these compounds did not cause anxiety-causing effects in either genotype mice.

No significant interplay was found between "genotype" and "treatment" in terms of horizontal activity (center: F (1, 44) ═ 0.64, p ═ 0.71, periphery: F (1, 76) ═ 1.06, p ═ 0.35, total: F (1, 76) ═ 1.00, p ═ 0.37), but ANOVA revealed that these two factors had a significant effect on the number of hind leg standings (F (1, 76) ═ 3.36, p ═ 0.04).

Example 5 exploratory activity: orifice plate

The well plate is a wood box (32x32x30cm) with four wells. The bottom surface is divided into nine squares of 10 cm. In a 5 minute experiment, the number of seeks, the time taken to explore each well and the total activity in the device were measured. The repetition index was also calculated based on the number of ABA alternations (exploring previously explored wells).

Table 4 shows the scores of R1, 8581 and vehicle treated Ts65Dn and control mice in the well plate test. Under all treatment conditions, Ts65Dn mice crossed more times than control mice. 8581 and R1 treatment caused Ts65Dn mice to show a reduction in this hyperactivity. Ts65Dn mice also showed an increased number of explorations made under all treatment conditions. No difference was found between Ts65Dn and control mice vertical activity under different treatment conditions in this maze. No significant difference was found in the time spent exploring the wells between the two genotype mice and the treatment. Ts65Dn mice showed attention changes because they repeated more times to explore the most recently explored well (ABA index). After 8581 treatment (but not after R1 treatment), the ABA index of Ts65Dn mice was normalized.

Example 6. spatial learning: morris water maze

The Morris water maze was used to evaluate spatial learning. The device is a circular trough of 110cm diameter filled with water (22-24 ℃) and made opaque by the addition of powdered milk. Inside the tank, the platform is hidden 1cm below the water level.

At the end of the treatment period, animals were tested for 12 consecutive periods of one day: 8 learning periods (platform submerged in water) followed by 4 cued periods (platform visible). All tests were recorded with a camera located 2m above the horizontal. In each experiment, the mouse path was analyzed using a SMART computerized tracking system (panlabs.a., barcelona, spain) for each animal, measuring escape delay time, travel distance, and swimming speed.

Period of training

During the learning period (S1-S8), the platform is hidden 1cm below the water level. From one learning period to the next in one phase each day, the platform is placed in different positions (east, southwest, center and northwest); each site is used for four consecutive periods of time one period per day. The 8 learning periods and 4 cued periods (one period per day) included four pairs of trials at intervals of 30-45min, respectively. In each trial, mice were randomized from one of four positions (north, south, east, west) and the starting positions of both trials remained unchanged. The first pair of trials was terminated when the mouse found a platform or 60 seconds had elapsed; a second trial was started after a period of 20 seconds during which the animals were allowed to rest on the platform. Several fixation cues located outside the maze are always visible in the pool.

Cued period

During the cued period, the platform is visible: the horizontal plane is located 1cm below the platform and its position is marked with a mark. During each phase, eight trials were performed following the same experimental procedure as the learning phase.

As shown in figure 7, all groups of mice know the platform position throughout the learning period because their delay time to reach the platform is shortened (ANOVA 'phase': F (7, 65) ═ 26.8, p < 0.001).

Ts65Dn mice showed significant learning deficit in MWM (ANOVA "genotype": F (1, 65) ═ 39.26, p < 0.001; fig. 8A), but the difference between Ts65Dn and the control learning curve decreased after 8581(ANOVA "genotype": F (1, 18) ═ 4.69, p < 0.05; fig. 8B) and R1 treatment (ANOVA "genotype": F (1, 26) ═ 13.57, p < 0.01).

As shown in fig. 9A, 8581 treatment significantly improved the performance of Ts65Dn mice (ANOVA "treatment": F (1, 24) ═ 32.43, p < 0.001). Chronic R1 treatment also improved cognition in Ts65Dn mice (F (1, 24) ═ 9.2, p < 0.01; fig. 9C). 8581 (FIG. 9B) or R1 (FIG. 9D) did not significantly affect the performance of control mice.

During the suggestive period (fig. 10), vehicle treated Ts65Dn mice showed prolonged delay time to plateau relative to control mice (ANOVA "genotype:f (1, 46) ═ 5.35, p ═ 0.024). R1 and 8581 treatment shortened the delay time to the plateau in Ts65Dn mice (ANOVA "treatment": F (1, 46) ═ 6.52, p ═ 0.003), but not in control mice (ANOVA "genotype x treatment": F (1, 46) ═ 3.44, p ═ 0.038)).

Example 7 Long Term Potentiation (LTP)

The effect of long-term administration of 8581 and R1 on LTP was evaluated in a Ts65Dn mouse model of down syndrome. 8581, R1(20mg/kg, oral) or vehicle was administered for six weeks. At 1 hour after the last administration, mice were decapitated and brains were removed rapidly. The hippocampus was dissected and cut into 400- μm sections with a microtome. The sections were allowed to recover in an interface bath (interface chamber) at room temperature for at least 1 hour with artificial cerebrospinal fluid (ACSF) containing (in mM): 120NaCl, 3.5KCl, 2.5CaCl₂、1.3MgSO₄、1.25NaH₂PO₄、26NaHCO₃And 10D-glucose, and is coated with 95% O₂And 5% CO₂And (4) saturation. From the CAl drainage layer with a glass micropipette (1-4M Ω) containing 2M NaCl, the field excitatory postsynaptic potential (fEPSP) was recorded and it was caused by stimulating the sandfly side branch with an insulated bipolar platinum/iridium electrode > 500 μ M from the recording electrode. The stimulation intensity was adjusted to cause fEPSP equal to 50% of the relative maximum amplitude without overlapping cluster peaks. After recording a stable baseline (pulse duration 100 μ s, 0.033Hz), long-term potentiation (LTP) was induced by TBS (1 burst of 5 pulses at 100Hz, 10 bursts total, and 200ms apart). During catalepsy (tetanus), the duration of the stimulation pulse is doubled. After a baseline recording of 20min, LTP was induced in each individual hippocampal slice and recorded for 80 min. The signal from the recording electrode was amplified, band-pass filtered (1Hz-1kHz), and stored in a computer using the Spike2 program (Spike2, Cambridge electronic design, Cambridge, UK). For the analysis, the fEPSP slope is expressed as a percentage of the baseline value recorded. Results from several sections are expressed as mean ± SEM. Statistical analysis was performed by Repeated Measures (RM) MANOVA ("time" x "treatment" x "genotype"). All analyses were performed using SPSS for Windows version 18.0.

As shown in figures 11 and 12, hippocampal slices of vehicle-treated Ts65Dn mice exhibited LTP deficiency. In contrast, LTP induced in hippocampal slices of 8581 or R1 treated animals was not different from that induced in hippocampal slices of control mice (fig. 11 and 12, respectively). This suggests that long-term treatment of Ts65Dn mice with 8581 or R1 may rescue LTP deficiency, possibly by reducing the excessive GABA-mediated inhibition observed in these animals.

Example 8 salvaged neurogenesis

Changes in hippocampal morphology (e.g., decrease in granulocytic density and hippocampal neurogenesis) were also associated with learning deficiencies shown in Ts65Dn mice. Spatial learning is known to be dependent on the functional integrity of the hippocampus, a structure in the CNS that plays an important role in information encoding and retrieval. We have studied the neogenetic cell population in Dentate Gyrus (DG) by labeling proliferating cells with anti-Ki 67 (a cell marker that undergoes late G1 and G2 and M phases). We demonstrated that hippocampus neurogenesis was reduced in these mice and showed that chronic administration of 8581 completely restored the density of proliferating cells in TS mice (p ═ 0.033; fig. 13). Moreover, neuronal survival of cells that have undergone maturation in TS mice was also normalized after long-term administration of 8581, as shown by the increase in DAPI + cells found in TS mice (p ═ 0.026; fig. 14).

Thus, the compound promotes cell proliferation and survival of neurons that have undergone maturation. Since both nascent and mature neurons appear to be involved in hippocampal-dependent learning and memory, restoration of proliferation and mature neuron density may be associated with a cognitive enhancing effect of 8581 in TS mice.

In addition, we found an increase in the number of gabaergic synapses in the hippocampus of TS mice compared to control animals. Importantly, long-term treatment with 8581 rescues the change because the number of synapses that were GAD positive after long-term treatment with the selective GABAA α 5NAM decreased significantly (p ═ 0.017; FIG. 15). This treatment produced an insignificant trend in CO mice to increase the number of these synapses.

Example 9, Nfl +/-spatial learning of mice in the Morris Water maze

Mice were trained for 8-9 days in two consecutive trials 1 week post treatment, 30 minutes after intraperitoneal injection of 8581 or vehicle per day. In each experiment, mice were given 60 seconds to look for a platform. After each experiment, mice were placed on the platform for 15 seconds. On the day of the probing test (day 3, day 5, day 7 and day 9), a 60-second probing test was performed after training. For probe trial 1 (day 3), none of the test groups know to search unambiguously in the target quadrant; probing trials 3 and 4 (days 7 and 9) showed that the Nfl/vehicle group was significantly impaired compared to CO/vehicle (two-factor ANOVA, quadrant x genotype interaction, F (3, 51) ═ 5.662, P < 0.01). Importantly, for all probing trials, the 8581 treated Nfl +/-mice showed comparable performance to control animals, indicating that 8581 rescued the spatial learning deficit of Nfl +/-mice. Two different measures of spatial learning (search in quadrant% (fig. 16) and mean proximity to target platform (fig. 17)) also showed that the active drug compounds used in the present invention rescued the spatial learning deficit of Nfl +/-mice (CO/vehicle (n-10), Nfl/vehicle (n-9), CO/8581 (n-10), and Nfl/8581 (n-11)).

The effect of the active pharmaceutical compounds used in the present invention on the performance of Nfl +/-mice was tested under conditions where the spatial learning of the mutant groups did not differ from the control group (less attenuation due to fewer number of probing experiments). The average percentage of time spent in each quadrant during the probing trial is shown in fig. 18. The average proximity to the target quadrant is shown in fig. 19. The results show that Nfl +/-mice treated with vehicle did not differ from control mice treated in a similar manner. Comparison between the% sought for the target quadrant and the proximity to the target quadrant does not show any differences between the groups. For probe trials 1 and 2 (days 5 and 7), all groups were selectively searched in the target quadrant (p < 0.01) and there was no difference between groups.

Example 10 fear conditioned reflex

Control mice (B16; 129F1) were trained using a one-day (FIG. 20) or two-day (FIG. 21) continuous trial using a contextual fear conditioning protocol. On the day of training, mice were placed in the training chamber 30 minutes after intraperitoneal injection of 8581 or vehicle. A foot shock (1 second, 0.4mA) was delivered 40 seconds after placement. Conditioned responses (percent rigor-time in mice) were recorded by using an automated procedure 24h after training. The average freezing level during the first 30 seconds of each training day and 24 hours after the last training trial was plotted.

The dose response curves shown in figure 20 (0.3, 1.0 and 3.0mg/kg) indicate a dose-dependent increase in contextual fear conditioning in control mice. 3mg/kg8581 significantly enhanced rigor 24 hours after training (p < 0.05). As can be seen from fig. 21, 1mg/kg8581 also caused a significant enhancement of the contextual conditioned reflex after two consecutive days of training of mice. Drug-treated mice had a marked increase in rigidity compared to the control/vehicle group (F1, 18) ═ 5.254, p ═ 0.034).

EXAMPLE 11 rotating Bar

Control mice (B16; 129F1) and Nfl +/-mice (n-10 for each of the 4 groups) were treated with vehicle or 8581. Mice were tested 30 minutes after intraperitoneal injection of 8581(1mg/kg) or vehicle using a rotarod protocol using an accelerated speed (4-40rpm, maximum duration 300 seconds) with four trials performed and a 30min time interval between trials. FIG. 22 shows the performance of control mice on the turnstiles, and FIG. 23 explains the performance of Nf1 +/-mice on the turnstiles. 8581 did not affect the performance of Nf1 +/-mutant or control mice.

Claims (10)

1. Use of a GABAA alpha 5 negative allosteric modulator for the preparation of a medicament for the treatment, prevention and/or delay of progression of CNS disorders which are associated with excessive GABAergic inhibition in the cortex and hippocampus,

   wherein the CNS disorder is selected from cognitive deficits in Down syndrome, cognitive deficits in autism, or cognitive deficits in neurofibromatosis type I,

   wherein the GABAA α 5 negative allosteric modulator is (1, 1-dioxo-1, 6-thiomorpholin-4-yl) - {6- [3- (4-fluoro-phenyl) -5-methyl-isoxazol-4-ylmethoxy ] -pyridin-3-yl } -methanone or a pharmaceutically acceptable salt thereof.
2. 2. The use of claim 1, wherein the excessive gabaergic inhibition in the cortex and hippocampus is caused by a neurodevelopmental defect.
3. 3. The use of claim 1, wherein the CNS disorder is caused by a neurodevelopmental defect resulting in excessive gabaergic inhibition in the cortex and hippocampus.
4. 4. The use according to any one of claims 1-3, wherein the CNS disorder is cognitive deficit in Down syndrome.
5. 5. The use according to any one of claims 1-3, wherein the CNS disorder is autism cognitive deficit.
6. 6. The use according to any one of claims 1-3, wherein the CNS disorder is cognitive deficits in neurofibromatosis type I.
7. 7. The use according to any one of claims 1 to 6, wherein the GABAA α 5 negative allosteric modulator binds to the human GABAA α 5 β 3 γ 2 receptor subtype with a binding selectivity of more than 10 times compared to the binding affinity to the human GABAA α 1 β 2/3 γ 2, α 2 β 3 γ 2 and α 3 β 3 γ 2 receptor subtypes.
8. 8. The use according to any one of claims 1 to 7, wherein the GABAA alpha 5 negative allosteric modulator exhibits functional selectivity as follows: as inverse agonists of the human GABAA α 5 β 3 γ 2 receptor subtype, the effect of GABA is reduced by more than 30% and additionally the GABA effect is less than 15% on the human GABAA α 1 β 2/3 γ 2, α 2 β 3 γ 2 and α 3 β 3 γ 2 receptor subtypes.
9. 9. The use according to any one of claims 1 to 8, wherein the GABAA α 5 negative allosteric modulator is used alone or in combination with a second active pharmaceutical compound, sequentially or simultaneously.
10. 10. Use of a GABAA α 5 negative allosteric modulator according to any one of claims 1-9 for the preparation of a medicament for the treatment, prevention and/or delay of progression of cognitive deficits in down syndrome, autism cognitive deficits, neurofibromatosis type I and/or for post-stroke recovery.