3-[4-(DIBENZO[b,f][1,4]OXAZEPIN-11-YL)-PIPERAZIN-1-YL]-2,2-DIMETHYL PROPANOIC ACID FOR USE IN THE TREATMENT OF SLEEP DISORDERS

? Acid 3– [4– (dibenzo [b, f] [1,4] oxazepin – 11-yl) –piperazin – 1-yl] –2,2-dimethyl-propanoic acid for use in the treatment of sleep disorders

Description

The invention relates to methods for treating sleep disorders and compositions useful in such procedures.

Difficulty falling asleep or staying asleep is a medical problem of relevance for a variety of reasons. Sometimes, these problems are due to endogenous conditions, such as sleep apnea or insomnia. Other times, these problems arise from exogenous stress conditions, such as the disturbing effect produced by changes in work shifts and jet lag. Whether it has an endogenous or exogenous origin, the difficulty of falling asleep or staying asleep can cause daytime sleepiness that affects the health, quality of life and safety of those who suffer from it.

Existing pharmaceutical treatments to induce sleep include sedatives or hypnotics, such as benzodiazepine and barbiturate derivatives. These treatments have numerous drawbacks, including rebound insomnia, delayed onset of desired sedative effects, persistence of sedative effects after the desired period of sleep and side effects due to nonspecific activity, such as psychomotor and memory deficits, Miorrelaxation and alteration of sleep patterns, including MOR sleep inhibition. In addition, sedatives and hypnotics can create dependence, may lose their effectiveness after prolonged use and some people may metabolize them more slowly. Therefore, doctors usually recommend or prescribe antihistamines as milder treatments for sleep disorders, in cases where hypnotics are less appropriate. However, many antihistamines have a number of side effects. These side effects include prolongation of the QT interval on the electrocardiogram of a subject, as well as side effects on the central nervous system (CNS), such as decreased muscle tone and droopy eyelids. Finally, such compounds can bind to muscarinic receptors, which leads to anticholinergic side effects, such as blurred vision, dry mouth, constipation, urinary problems, dizziness and anxiety.

As a result, there is a need for treatments that promote sleep with reduced side effects. In addition, although known sleep-inducing compounds are effective in treating initial insomnia, ie, a subject's difficulty in falling asleep at bedtime, there are currently no drugs indicated to treat sleep maintenance insomnia, ie, maintaining sleep of a subject during a normal period of sleep after having reconciled. Therefore, there is also a need for better pharmaceutical treatments to maintain sleep in subjects in need of such treatment.

The present invention relates to loxapine analogs and their use to modulate sleep. Loxapine (LOXAPAC ™, LOXITANE ™) ​​is a tricyclic dibenzoxazepine antipsychotic agent that is used in the treatment of schizophrenia manifestations. Loxapine (2-chloro-11– (4-methyl-1-piperazinyl) dibenz [b, f] [1,4] oxazepine) has the following structure:

Compound 1:

or a salt, a solvate or a hydrate thereof. For example, the invention relates to a solvate of Compound 1. In one embodiment, the invention relates to a hydrate of Compound 1.

In another embodiment, the invention relates to a pharmaceutically acceptable salt of Compound 1. For example, the salt may be an acid addition salt, such as a hydrochloride salt.

In one aspect, the invention relates to the compound:

In another aspect, the invention relates to the compound:

In another aspect, the invention relates to a composition comprising a compound of the formula:

or a salt, a solvate or a hydrate thereof, and at least one pharmaceutically acceptable excipient. In one embodiment, the composition includes a solvate of Compound 1. In another embodiment, the composition includes a hydrate of Compound 1. In another embodiment, the composition includes a pharmaceutically acceptable salt of Compound 1. For example, the salt may be a salt. of acid addition. An embodiment of an acid addition salt is a salt

fifteen hydrochloride

In one embodiment, the invention relates to a composition of and at least one pharmaceutically acceptable excipient. In another embodiment, the invention relates to a composition of

and at least one pharmaceutically acceptable excipient. In another aspect, the invention relates to Compound 1:

or a pharmaceutically acceptable salt, solvate or hydrate thereof for use in sleep modulation. For example, the subject is a human being. In one embodiment, sleep modulation is selected from, for example, decrease in time until sleep is reconciled, increase in the average duration of the sleep session and increase

10 of the maximum duration of the sleep session. In one embodiment, sleep modulation treats a sleep disorder. Examples of sleep disorders include disorders of the circadian rhythm, insomnia, parasomnia, sleep apnea syndrome, narcolepsy and hypersomnia.

In one embodiment, Compound 1, or a pharmaceutically acceptable salt, solvate or hydrate thereof is administered in combination with one or more additional therapies to modulate sleep in a subject. For example, him

fifteen Subject is a human being.

The foregoing description sets out in a fairly general manner the most important features of the present invention to allow understanding of the detailed description of the present invention presented below and so that the present contributions made to the technique can be better appreciated. Other objectives and features of the present invention will be apparent from the following detailed description considered in combination.

twenty With the examples.

The details of one or more embodiments of the invention are set forth in the description that follows. Although it is possible to use any procedure and material similar or equivalent to those described herein in practice or in the test of the present invention, the procedures and materials are now described. There are other features, objectives and advantages of the invention that will be apparent from the

25 description. Throughout memory, singular forms also include the plural, unless the context clearly states otherwise. Unless otherwise defined, all technical and scientific terms used herein have the same meanings that are commonly known to any person skilled in the art to which the present invention pertains. In case of conflict, we must abide by this report.

30 Definitions

For convenience, certain terms and expressions used in the report, the examples and the appended claims are collected here.

"Treat" includes any effect, eg, that diminishes, reduces, modulates or eliminates, resulting in the improvement of the condition, disease, disorder, etc. "Treating" includes any effect, eg, that diminishes, reduces, modulates or eliminates, that results in the improvement of the condition, disease, disorder, etc. "Treat" or "treatment" of a

Pathological status includes: (1) preventing the pathological state, ie, making the clinical symptoms of the pathological state not develop in a subject that may be exposed or predisposed to the pathological state, but does not yet show symptoms of the pathological state; (2) inhibit the pathological state, ie, stopping the development of the pathological state or its clinical symptoms; or (3) relieve the pathological state, ie, cause a temporary or permanent withdrawal of the pathological state or its clinical symptoms. "Pathological state" means any disease, condition, symptom or indication.

The terms "crystalline polymorphs" or "polymorphs" or "crystalline forms" mean crystalline structures in which a compound (or a salt or a solvate thereof) can crystallize in different crystal packaging distributions, all of which have The same elementary composition. Different crystalline forms usually have different X-ray diffraction patterns, infrared spectra, melting point, density, hardness, crystalline form, and optical and electrical properties, stability and solubility. The recrystallization solvent, crystallization rate, storage temperature and other factors can cause one crystalline form to dominate over another. The crystalline polymorphs of the compounds can be prepared by crystallization under different conditions. For example, by using different solvents or different solvent mixtures for recrystallization; crystallization at different temperatures; various modes of cooling, ranging from very fast to very slow cooling during crystallization, and the like. Polymorphs can also be obtained by heating or melting the disclosed compounds followed by gradual or rapid cooling. The presence of polymorphs is determined by solid magnetic nuclear resonance spectroscopy, infrared spectroscopy, differential scanning calorimetry, powder X-ray diffraction and other techniques known to those skilled in the art.

In addition, the compounds of the present invention, for example, the salts of the compounds, can exist either hydrated or dehydrated (the anhydrous form) or as solvates with other solvent molecules. Non-restrictive examples of hydrates include monohydrates, dihydrates, etc. Non-restrictive examples of solvates include ethanol solvates, acetone solvates, etc.

"Solvates" means solvent addition forms that contain either stoichiometric or non-stoichiometric amounts of solvent. Some compounds have a tendency to trap a fixed molar ratio of solvent molecules in the crystalline solid state, thus forming a solvate. If the solvent is water, the solvate formed is a hydrate, when the solvent is alcohol, the solvate formed is an alcoholate. Hydrates are formed by combining one or more water molecules with one of the substances in which water retains its molecular state as H2O, such combination being capable of forming one or more hydrates.

As used herein, the term "analog" refers to a chemical compound that is structurally similar to another, but differs slightly in its composition (as in the substitution of an atom for an atom of a different element or in presence of a specific functional group, or the replacement of a functional group by another functional group). Thus, an analog is a compound that is similar or comparable in function and appearance, but not in structure or origin to the reference compound.

As defined herein, the term "derivative" refers to compounds that have a common core structure and are substituted with various groups as described herein. For example, all compounds represented by Formula I are derivatives of loxapine, and have Formula I as a common nucleus.

The term "loxapine type compounds" or "loxapine analog compounds" or "loxapine derivative compounds" is intended to include loxapine analogs or antihistamines that include two aryl groups attached to the same atom that are linked through a tricyclic ring system, eg, a seven-membered oxygen and nitrogen ring (ie, similar to loxapine) attached to place a piperazine ring.

The term "antihistamine" refers to a compound that binds to an H1 receptor and blocks histamine activity and / or reduces the constitutive activity of the receptor.

As used herein, the term "sleep disorder" includes conditions recognized by those skilled in the art as sleep disorders, for example, conditions known in the art or conditions that are proposed as sleep disorders or that have been discovered that they are sleep disorders. A sleep disorder also appears in a subject that has other medical disorders, diseases or injuries, or in a subject that is being treated with other medications or medical treatments, so that the subject, as a result, has difficulty reconciling the sleep and / or remain asleep or do not have a restorative or restorative sleep, eg, the subject suffers from sleep deprivation.

The term "treatment of a sleep disorder" also includes treating a sleep disorder component of other disorders, such as CNS disorders (eg, mental or neurological disorders, such as anxiety). In addition, the expression "treat a sleep disorder" includes the beneficial effect of improving other symptoms associated with the disorder.

The term "maximum non-MOR sleep time" is defined as an absolute maximum amount of non-MOR sleep per hour after treatment, the administration of the drug occurring in the circadian hour (HC) 18, which corresponds to 6 hours after the light goes out in a night lab rat subjected to a light-dark cycle of

12:12 LO (12 hours of light and 12 hours of darkness). The nominal criteria of 55% non-MOR sleep per hour equals 33 minutes of non-MOR sleep per hour.

As used herein, the term "cumulative non-MOR sleep" is defined as the cumulative total net increase in the number of minutes of non-MOR sleep, measured over the entire duration of a drug's soporific effect, which commonly, but not always, takes place in the first 6 hours after treatment, adjusted for the cumulative total net number of minutes of non-MOR sleep produced during the corresponding reference hours without treatment of the day recorded 24 hours before compared to the control vehicle treatment.

As defined herein, the term "sleep session" refers to a differentiated episode of continuous or near-continuous sleep consisting of non-MOR sleep, MOR sleep or both MOR and non-MOR sleep phases, which is defined before and then for more than two contiguous periods of 10 seconds of waking state.

As used herein, the term "longest duration of the sleep session" is defined as the total number of minutes an animal remains asleep (non-MOR and / or MOR sleep phases) during the single episode or " Longest sleep session ”that took place starting at a given time after treatment. The measurement criteria of the “duration of the sleep session” assume that sleep is measured continuously in times of 10 seconds and the assignment of the value is made based on the predominant state, accounted for or otherwise determined as a phase of differentiated sleep (with the sleep phases defined as non-MOR sleep, MOR sleep or wakefulness) during the 10-second interval that defines the time.

The expression "average duration of the sleep session" is defined as the average duration (in minutes) of each sleep session that begins at a given time, regardless of the individual duration of each episode or session.

"Bounce insomnia" is defined as a period of rebound, paradoxical or compensatory wakefulness that takes place after the sleep-promoting effects of a hypnotic or soporific agent.

"MOR sleep inhibition" is defined as the reduction of MOR sleep time after treatment at HC 18 (6 hours after the light goes out, LO 12:12) or at HC 5 (5 hours after it is turned on the light; LO 12:12). Compounds that reduce MOR sleep time by more than 15 minutes (with respect to the reference and adjusted for vehicle treatment) when administered well to HC 18 or HC 5 are considered unacceptable.

Compared to non-MOR sleep or wakefulness, MOR sleep causes ventilatory depression and episodic cardiovascular changes. During rebound insomnia, the physiological effects of MOR sleep are magnified and normal sleep cycles are disrupted.

As defined herein, "disproportionate inhibition of locomotor activity" is a reduction in locomotive activity that exceeds the normal and expected reduction in behavior attributable to sleep.

"Combination therapy" (or "therapy") includes the administration of a compound of the invention and at least one second agent as part of a specific treatment regimen intended to provide the beneficial effect from the joint action of these therapeutic agents The beneficial effect of the combination includes, but is not limited to, the joint pharmacokinetic or pharmacodynamic action that results from the combination of therapeutic agents. The combinations of the compounds of the present invention and the other active agents can be co-administered in a single combination or separately. When a separate administration is used, the administration of an element may be prior, simultaneous or subsequent to the administration of other agents. The administration of these therapeutic agents in combination is usually carried out in a defined period of time (usually, in minutes, hours, days or weeks depending on the selected combination). In one embodiment, "combination therapy" encompasses the administration of two or more of these therapeutic agents as part of separate monotherapeutic guidelines that intentionally or arbitrarily result in the combinations of the present invention. In another embodiment, "combination therapy" is intended to encompass the administration of these therapeutic agents in a sequential manner. That is, in which each therapeutic agent is administered at a different time, as well as the administration of these therapeutic agents, or at least two of the therapeutic agents, in a substantially simultaneous manner. Substantially simultaneous administration can be performed, for example, by administration to the subject of a single capsule having a fixed proportion of each therapeutic agent or in multiple individual capsules for each of the therapeutic agents. Sequential or substantially simultaneous administration of each therapeutic agent can be accomplished by any appropriate route that includes, but is not limited to, oral routes, intravenous routes, intramuscular routes and direct absorption through mucous membrane tissues.

The therapeutic agents can be administered by the same route or by different routes. For example, a first therapeutic agent of the selected combination can be administered by intravenous injection, while the other therapeutic agents of the combination can be administered orally. Alternatively, for example, all therapeutic agents can be administered orally or all therapeutic agents can be administered by intravenous injection. The order in which the therapeutic agents are administered is irrelevant. "Combination therapy" also encompasses the administration of therapeutic agents as described above in another combination with other biologically active ingredients and non-pharmacological therapies (eg, surgery, radiation treatment or a medical device). When the combination therapy further comprises a non-pharmacological treatment, the non-pharmacological treatment can be applied at an appropriate time, as long as a beneficial effect of the joint action of the combination of the therapeutic agents and the non-pharmacological treatment is achieved. For example, in appropriate cases, a beneficial effect can still be achieved when non-drug treatment is temporarily withdrawn from the administration of therapeutic agents, perhaps for days or even weeks.

The terms "parenteral administration" and "parenterally administered" as used herein, refer to modes of administration other than enteral or topical administration, usually by injection, and include, without limitation, intramuscular, intramuscular injection and infusion. , intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal and intrasternal.

The term "pulmonary", as used herein, refers to any part, tissue or organ whose main function is the exchange of gas with the outside environment, eg, exchange of O2 / CO2, in a patient . "Pulmonary" usually refers to the tissues of the respiratory tract. Thus, the phrase "pulmonary administration" refers to the administration of the formulations described herein in any part, tissue or organ whose main function is the gas exchange with the external environment (eg, mouth, nose, pharynx, oropharynx, laryngopharynx, larynx, trachea, carina, bronchi, bronchioles, alveoli). For the purposes of the present invention, "pulmonary" also includes a tissue or a cavity that is contiguous with the respiratory tract, in particular, the sinuses.

A "pharmaceutical composition" is a formulation that contains the compounds disclosed in a form suitable for administration to a subject. In one embodiment, the pharmaceutical composition is in a pharmaceutical or unit dosage form. The unit dosage form is any of a variety of forms that includes, for example, a capsule, an IV bag, a tablet, a simple pump on an aerosol inhaler or a vial. The amount of active ingredient (eg, a formula of the disclosed compound or salts thereof) in a unit dose of composition is an effective amount and is varied according to the treatment in question. The person skilled in the art will appreciate that it is sometimes necessary to make routine variations of the dose depending on the age and condition of the patient. The dose will also depend on the route of administration. A variety of routes are contemplated, including oral, pulmonary, rectal, parenteral, transdermal, subcutaneous, intravenous, intramuscular, intraperitoneal, intranasal and the like. Pharmaceutical forms for topical or transdermal administration of a compound of the present invention include powders, powders, ointments, pastes, creams, lotions, gels, solutions, patches and inhalants. In one embodiment, the active compound is mixed under sterile conditions with a pharmaceutically acceptable carrier and with any necessary preservative, buffer or propellant.

The term "instant dose" refers to formulations of compounds that are rapidly dispersed pharmaceutical forms.

The term "immediate release" is defined as a release of compound in a pharmaceutical form in a relatively short period of time, generally, up to about 60 minutes. The term "modified release" is defined to include delayed release, prolonged release and pulsed release. The term "pulsed release" is defined as a series of drug releases from a pharmaceutical form. The term "sustained release" or "prolonged release" is defined as the continuous release of a compound from a pharmaceutical form for a prolonged period of time.

A "subject" includes mammals, eg, humans, companion animals (eg, dogs, cats, birds and the like), farm animals (eg, cows, sheep, pigs, horses, poultry and the like) and laboratory animals (eg, rats, mice, guinea pigs and the like). Most preferably, the subject is a human being.

As used herein, the term "pharmaceutically acceptable" refers to those compounds, materials, compositions, vehicles and / or pharmaceutical forms that are, based on a solid medical opinion, suitable for use in contact with tissues. of human beings and animals without causing excessive toxicity, irritation, allergic response or other problem or complication, according to a reasonable benefit / risk ratio.

"Pharmaceutically acceptable excipient" means an excipient that is useful in the preparation of a pharmaceutical composition that is, in general, safe, non-toxic and neither biologically nor otherwise undesirable, and includes excipients that are acceptable for veterinary use, as well as A pharmaceutical use in humans. A "pharmaceutically acceptable excipient," as used herein and in the claims, includes both one and more than one such excipient.

The compounds of the invention can also form salts. All these forms are also contemplated within the scope of the claimed invention.

The "pharmaceutically acceptable salt" of a compound means a salt that is pharmaceutically acceptable and that possesses the desired pharmacological activity of the precursor compound.

As used herein, "pharmaceutically acceptable salts" refers to derivatives of the disclosed compounds in which the precursor compound is modified by the formation of acid or base salts thereof. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues, such as amines; alkaline or organic salts of acidic residues, such as carboxylic acids and the like. Pharmaceutically acceptable salts include conventional non-toxic salts or quaternary ammonium salts of the precursor compound formed, for example, from non-toxic organic or inorganic acids. For example, such conventional non-toxic salts include, but are not limited to, those derived from organic and inorganic acids selected from 2-acetoxybenzoic, 2-hydroxyethanesulfonic, acetic, ascorbic, benzene-sulfonic, benzoic, bicarbonic, carbonic, citric , edetic, ethanedisulfonic, 1,2-ethane-sulphonic, fumaric, glucoheptonic, gluconic, glutamic, glycolic, glycolsanilic, hexylresorcinic, hydrabamic, hydrobromic, hydrochloric, iohydric, hydroxyloleic, hydroxynaphthoic, isethionic, lactic, lactobionic, lauryl-sulfonic, maleic, malic, mandelic, methane-sulphonic, napsyl, nitric, oxalic, pamoic, pantothenic, phenylacetic, phosphoric, polygalacturonic, propionic, salicylic, stearic, subacetic, sulphonic sulfanyl, sulfuric, tannic, tartaric, toluene-sulfonic, and common amino acids, eg, glycine, alanine, phenylalanine, arginine, etc.

Other examples include hexanoic acid, cyclopentane-propionic acid, pyruvic acid, malonic acid, 3- (4-hydroxy-benzoyl) benzoic acid, cinnamic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4-toluenesulfonic acid, camphor sulfulfic acid, 4-methylbicyclo acid [2.2.2] -oct-2-ene-1-carboxylic acid, 3-phenylpropionic acid, trimethylacetic acid, tertiary butylacetic acid, muconic acid and the like. The invention also encompasses salts formed when an acid proton present in the precursor compound is well replaced by a metal ion, eg, an alkali metal ion, an alkaline earth metal ion or an aluminum ion; or is coordinated with an organic base such as ethanolamine, diethanolamine, triethanolamine, tromethamine, N-methylglucamine and the like.

It is to be understood that all references made to pharmaceutically acceptable salts include solvent addition forms (solvates) or crystalline forms (polymorphs) as defined herein of the same salt.

The pharmaceutically acceptable salts of the present invention can be formed from a precursor compound containing a basic or acidic moiety by conventional chemical procedures. Generally, it is possible to prepare such salts by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent or in a mixture of both; generally, non-aqueous media such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile are used. For example, the scheme below shows the formation of a pharmaceutically acceptable hydrochloric salt of the precursor compound, Compound 1, after treatment with hydrochloric acid.

The number of protonated atoms and counterions associated with the salt can be controlled and depends on the number of acidic / basic atoms of the parent compound and the amount of acid used to treat the parent compound. In one embodiment of the invention, the monohydrochloride salt of the precursor compound is formed after treatment with hydrochloric acid. In another embodiment, the hydrochloride salt of the precursor compound is formed after treatment with hydrochloric acid.

In "Remington's Pharmaceutical Sciences", XVIII ed. (Mack Publishing Company, 1990), lists of suitable salts are found. For example, salts may include, but are not limited to, hydrochloride and acetate salts of the compounds of the present invention that contain aliphatic amine, that contain hydroxylamine and that contain imine.

The compounds of the present invention can also be prepared as esters, for example, pharmaceutically acceptable esters. For example, it is possible to convert a carboxylic acid function group of a compound into its corresponding ester, eg, a methyl ester, ethyl ester or other ester. In addition, an alcohol group of a compound can be converted into its corresponding ester, eg, an acetate, propionate or other ester.

In memory, singular forms also include the plural, unless the context clearly indicates otherwise. Unless otherwise defined, all technical and scientific terms used herein have the same meanings that are commonly understood by any person skilled in the art to which the present invention belongs. In case of conflict, this report will be taken as a reference.

All percentages and proportions used herein, unless otherwise indicated, are given by weight.

An "effective amount" of a compound of the disclosed invention is the amount that, when administered to a subject having a disease or disorder, causes regression of the disease or disorder in the subject. The amount of the disclosed compound that will be administered to a subject will depend on the particular disorder, the mode of administration, the co-administered compounds, if any, and the characteristics of the subject, such as general health, other diseases, age, sex, genotype, body weight and drug tolerance. The person skilled in the art will be able to determine the appropriate doses based on these and other factors.

As used herein, the term "effective amount" refers to an amount of a compound, or a combination of compounds, of the present invention effective when administered alone or in combination in the form of a sleep inducing agent. For example, an effective amount refers to an amount of the compound present in a formula or in a medical device administered to a patient or recipient subject sufficient to cause biological activity. The combination of compounds is optionally a synergistic combination. Synergy, as described, for example, by Chou and Talalay, Adv. Enzyme Regul. vol. 22, pp. 27–55 (1984), occurs when the effect of the compounds administered in combination is greater than the additive effect of the compounds when administered alone as a single agent. In general, synergistic effects are most clearly demonstrated at sub-optimal concentrations of the compounds. Synergy can occur in terms of a lower cytotoxicity or a greater sleep-promoting effect, a lower hangover effect or some other beneficial effect of the combination compared to the individual components.

"A therapeutically effective amount" means the amount of a compound that, when administered to a mammal to treat a disease, is sufficient to effect such a treatment of the disease. The "therapeutically effective amount" will vary depending on the compound, the disease and its severity, and the age, weight, etc. of the mammal to be treated.

"Pharmacological effect", as used herein, encompasses effects produced in the subject that achieve the intended purpose of a therapy. In one embodiment, a pharmacological effect means that the main indications of the subject being treated are prevented, relieved or reduced. For example, a pharmacological effect would be one that produced the prevention, relief or reduction of the main indications in a treated subject. In another embodiment, a pharmacological effect means that the disorders or symptoms of the main indications of the subject being treated are prevented, relieved or reduced. For example, a pharmacological effect would be one that produced the prevention or reduction of the main indications in a treated subject.

The invention provides a method of modulating sleep by administering an effective amount of a loxapine analog of the invention that is a moiety that antagonizes a histamine receptor or a group of histamine receptors. The invention also relates to new analogs of loxapine.

Effective sleep modulators have certain characteristics that correspond to greater efficacy and lower side effects. These characteristics include a desired half-life in a subject, control of the onset of the desired sedative effects and a minimal and even undetectable effect on the side effects produced in the psychomotor system or the central nervous system (CNS) (eg, deficit memory, decreased muscle tone, droopy eyelids or daytime sleepiness). For example, effective sleep modulators have a half-life in humans of less than 7 hours, less than 6 hours, less than 5 hours, less than 4 hours, approximately, 3 hours,

or in the interval of 3 to 7 hours.

One approach to the development of an effective sleep modulator is the strategic derivation of a known compound or known family of compounds with sleep modulating activity. The derivation of a known compound may increase one or more biological properties to allow the resulting compound to function in a better way. Examples of favorable biological properties include, but are not limited to, the induction of a hypnotic or differentiated sleep state, the activity of the therapeutic compound for a differentiated period of time, penetration through the blood brain barrier in the CNS, p .ej., as a consequence of the lipophilicity of the substituents or the configurational lipophilicity (ie, lipophilicity as a result of a certain configuration, such as the formation of internal salts between a carboxylate anion and a protonated amine), the modulation of the half-life of the therapeutic compound, a modification of the load, a modification of the pharmacokinetics, a modification of the log P in a value of one or more one, an increase in receptor selectivity, a decrease in peripheral half-life, the ability to increase the dose, the increase in peripheral elimination, the decrease in anti-muscarinic activity, Anticholinergic decline and any combination thereof.

The derivation of a compound produces a variety of effects and can modify the mechanism of action. For example, in some cases, a compound containing a certain functional group, such as, for example, an ester, carboxylic acid or alcohol group, has a better selectivity for a desired receptor against unwanted receptors compared to a Composed without this group. In other cases, the compound containing the functional group in question is more active as a therapeutic agent for treating sleep disorders than the corresponding compound without this group. The effect of the derived compound depends on the identity of the addition.

By deriving a compound in order to increase favorable biological properties and decrease unwanted side effects, it is possible to implement a strategy based on possible mechanistic effects or interactions. For example, in some compounds, the presence of a carboxylic acid results in the ability to form an intramolecular ionic bond that includes the corresponding carboxylate ion, e.g., formation of zwitterionic species with a nitrogen atom in the compound or formation. of a link with salt. These interactions produce favorable biological effects such as configurational lipophilicity, ie, increased lipophilicity as a consequence of a certain configuration, such as formation of internal salts between an anion.

10 carboxylate and a protonated amine. Such configurational lipophilicity allows penetration through the blood brain barrier in the CNS, although it is believed that the presence of two polar ions inhibits the crossing of the non-polar blood brain barrier. Another benefit of the presence of the carboxylic acid is a greater ability of the compound to selectively bind to the desired receptor.

There is a group of compounds useful in sleep modulation that is related to loxapine, which is an agent

fifteen psychotherapeutic that belongs to the family of compounds commonly known as tricyclic antidepressants (TCA). Loxapine is an antipsychotic agent of dibenzoxazepine that produces pharmacological responses in various animal species that are characteristic of those observed in most antipsychotic drugs. Although the exact mechanism of action is unknown, administration of loxapine succinate produces a potent inhibition of spontaneous motor activity. Loxapine is recommended to treat schizophrenia.

twenty In one embodiment, the loxapine analogs of the invention are used in the treatment of a circadian rhythm disturbance, such as, for example, jet lag, shift shifts, delayed sleep phase syndrome, the advanced soil phase and sleep-cycle disorder different from 24 hours.

In another embodiment, loxapine analogs are used in the treatment of insomnia, including, for example, extrinsic insomnia, psychophysiological insomnia, height insomnia, restless legs syndrome, disorder of

25 periodic limb movement, medication-dependent insomnia, drug dependence insomnia, alcoholism insomnia and insomnia associated with mental disorders.

In one embodiment, the loxapine analogs of the invention are used to treat a parasomnia disorder, such as,

p. eg, sleepwalking, night dread, behavior disorder associated with MOR sleep, sleep bruxism and sleep enuresis.

30 In another embodiment, loxapine analogs are used to treat a sleep apnea disorder, such as, for example, central sleep apnea, obstructive sleep apnea and mixed sleep apnea.

Some examples include:

Preferably, the compounds of the present invention modulate sleep with fewer side effects: e.g.,

35 Compounds do not inhibit MOR sleep (therefore, the sleep induced by these compounds may resemble the natural sleep cycles of the person), the use of the compounds does not cause rebound insomnia and / or the compounds do not inhibit activity Locomotive or negatively affect body temperature.

The in vitro selection criteria for the loxapine analogs of the invention are shown in Table 2.

Table 2

<dl><dt>Union to H1 (main Diana) </dt><dd>Ki &lt;500nMolar </dd></dl>

<dl><dt>Union without target </dt><dd /></dl>

<dl><dt>• Cholinergic M1, M2, M3 </dt><dd>• Ki&gt; 10 times the measured Ki of the H1 receiver </dd></dl>

<dl><dt>• Dopamine D1, D2 </dt><dd>• Ki&gt; 10 times the measured Ki of the H1 receiver </dd></dl>

<dl><dt>• Adrenergic a1, a2 </dt><dd>• Ki&gt; 10 times the measured Ki of the H1 receiver </dd></dl>

In one embodiment, the binding Ki without a target is 50 times the measured Ki of the H1 receptor. In some embodiments, the binding Ki without a target is 100 times the measured Ki of the H1 receptor.

In vitro binding assays are used to determine H1 binding (ie, binding to the main target) and binding to M1, M2 and M3 (ie, binding without target). These binding assays measure the ability of loxapine analogs to displace known patterns from the H1, M1, M2 and M3 receptors, in which H1 is a histamine receptor, and M1, M2 and M3 are cholinergic (muscarinic) receptors. Similar tests are performed with the H1 and dopamine receptors (D1 and D2) and with the H1 and adrenergic receptors (a1 and a2).

Binding studies for the histamine receptor, H1, indicate a binding affinity and, therefore, the results of binding assays indicate the activity of the loxapine analog compound. Binding studies for muscarinic receptors indicate the extent to which the compounds bind with muscarinic receptors responsible for the anticholinergic activity of the compound. Binding to muscarinic receptors causes several unwanted side effects of many known antihistamines, e.g. eg dry mouth. A decrease in the binding of the compounds with the M1-M3 receptors compared to the binding of the compound with the H1 receptor indicates a greater specificity of the compound for the histamine receptor versus the muscarinic receptor. In addition, a drug with greater specificity for the histamine receptor has fewer anticholinergic side effects.

The H1 binding of the loxapine analogs of the invention (also referred to herein as "test compounds" or "compounds of the invention") is determined by measuring the specific binding of a given test compound or a series of compounds of test with the H1 receptor and comparing it with the specific binding of a known pattern (ie, reference compound). Reference compounds used in this H1 binding assay include, for example, triprolidine (3.3nM Ki), chlorpheniramine (Ki 103.0nM), pyrilamine (Ki 1.9nM), cyproheptadine (Ki 8.5nM), cimetidine (Ki&gt; 10,000) and dimaprit (Ki&gt; 10,000). (See, eg, Chang et al., J. Neurochem., 32: 1653–63 (1979) (with modifications); Martinez – Mir, et al., Brain Res., 526: 322–27 (1990 ); and Haaksme, et al., Pharmac. Ther., 47: 73-104 (1990).

For example, in one embodiment of the H1 binding assay, the H1 receptor is derived from bovine cell membranes, and a radioligand [3H] Pyrilamine (15-25 Ci / mmol) is used at a final ligand concentration of 2.0nM for detect specific binding for the H1 receptor. Assay characteristics include a KD (binding affinity) of 1.3nM and a Bmax (number of receptors) of 6.2 fmol / mg of tissue (wet weight). Tripolidine (10IM) was used as a nonspecific determinant, reference compound and positive control. Binding reactions are carried out in 50Mm NA-KPO4 (pH 7.5) at 25 ° C for 60 minutes. The reaction was terminated with a rapid vacuum filtration on fiberglass filters. The level of radioactivity trapped on the filters is measured and compared with the control values ​​to determine any interaction between a given test compound and the H1 binding site.

The M1 binding assay determines the binding to M1 of a test compound by measuring the specific binding of a test compound given to M1 and its comparison with the specific binding of a reference compound. (See, eg, Buckley, et al., Mol. Pharmacol. 35: 469-76 (1989) (with modifications)). Reference compounds used in the M1 binding assay include, for example, scopolamine, MethylBr (0.09nM Ki); 4-DAMP iodomethylate (0.27nM Ki); pirenzepine (Ki 2.60nM); HHSID (Ki 5.00nM); and methoctramine (Ki 29,70nM).

For example, in one embodiment of the M1 binding assay, the M1 muscarinic receptor is a human recombinant M1 expressed in CHO cells, and a radioligand, [3 H] -espopolamine, N-methyl chloride (80-100 Ci /) is used. mmol) at a final ligand concentration of 0.5nM to detect specific binding to M1. Assay characteristics include a KD (binding affinity) of 0.05nM and a Bmax (number of receptors) of 4.2 pmol / mg protein. (-) - scopolamine, methyl–, bromide (methylcopolamine bromide) (1.0IM) is used as a nonspecific determinant, reference compound and positive control. Binding reactions are carried out in PBS for 60 minutes at 25 ° C. The reaction was terminated with a rapid vacuum filtration on fiberglass filters. The level of radioactivity trapped on the filters is measured and compared with the control values ​​to determine any interaction between a given test compound and the cloned muscarinic M1 binding site.

The M2 binding assay determines the binding to M2 of a test compound by measuring the specific binding of a test compound given to M2 and its comparison with the specific binding of a reference compound. (See, eg, Buckley, et al., Mol. Pharmacol. 35: 469-76 (1989) (with modifications)). Reference compounds used in the M2 binding assay include, for example, scopolamine, MethylBr (0.3nM Ki); 4-DAMP iodomethylate (Ki 20.7nM); methoctramine (Ki 20,460nM); HHSID (Ki 212.7nM); and pirenzepine (Ki 832.9nM).

For example, in one embodiment of the M2 binding assay, the M2 muscarinic receptor is a human recombinant M2 expressed in CHO cells, and a radioligand, [3H] -espopolamine, N-methyl chloride (80-100 Ci /) is used. mmol) at a final ligand concentration of 0.5nM to detect specific binding to M1. Assay characteristics include a KD (binding affinity) of 0.29 and a Bmax (number of receptors) of 2.1 pmol / mg protein. It's used

5 (-) - scopolamine, methyl - bromide (methylcopolamine bromide) (1.0μM) as a non-specific determinant, reference compound and positive control. Binding reactions are carried out in PBS for 60 minutes at 25 ° C. The reaction was terminated with a rapid vacuum filtration on fiberglass filters. The level of radioactivity trapped on the filters is measured and compared with the control values ​​to determine any interaction between a given test compound and the cloned muscarinic M2 binding site.

10 The M3 binding assay determines the binding to M3 of a test compound by measuring the specific binding of a test compound given to M3 and its comparison with the specific binding of a reference compound. (See, eg, Buckley, et al., Mol. Pharmacol. 35: 469-76 (1989) (with modifications)). Reference compounds used in the M3 binding assay include, for example, scopolamine, MethylBr (0.3nM Ki); 4-DAMP iodomethylate (0.8nM Ki); HHSID (Ki 14.5nM); pyrenzepine (Ki 153.3nM) and metoctramine (Ki 700.0nM).

fifteen For example, in one embodiment of the M3 binding assay, the M3 muscarinic receptor is a human recombinant M3 expressed in CHO cells, and a radioligand, [3H] -espolamine, N-methyl chloride (80-100 Ci /) is used. mmol) at a final concentration of 0.2nM ligand to detect specific binding to M1. Assay characteristics include a KD (binding affinity) of 0.14nM and a Bmax (number of receptors) of 4.0 pmol / mg protein. (-) - scopolamine, methyl–, bromide (methylcopolamine bromide) (1.0μM) is used as a nonspecific determinant,

twenty reference compound and positive control. Binding reactions are carried out in 50mM Tris-HCl (pH 7.4) containing 10mM MgCl2, 1mM EDTA for 60 minutes at 25 ° C. The reaction was terminated with a rapid vacuum filtration on fiberglass filters. The level of radioactivity trapped on the filters is measured and compared with the control values ​​to determine any interaction between a given test compound and the cloned muscarinic M3 binding site.

25 The in vitro selection criteria for the loxapine analogs of the invention are shown in Table 3.

Table 3

<dl><dt>Union to H1 (main Diana) </dt><dd>Ki &lt;300nMolar </dd></dl>

<dl><dt>Union without target • M1 cholinergic • M2 cholinergic • M3 cholinergic </dt><dd>• Ki&gt; 1uM • Ki&gt; 1uM • Ki&gt; 1uM </dd></dl>

Other in vitro selection criteria for the loxapine analogs of the invention are shown in Table 4.

Table 4

<dl><dt>Union to H1 (main Diana) </dt><dd>Ki &lt;150nMolar </dd></dl>

<dl><dt>Union without target • M1 cholinergic • M2 cholinergic • M3 cholinergic </dt><dd>• Ki&gt; 10uM • Ki&gt; 10uM • Ki&gt; 10uM </dd></dl>

30 Binding to H1 (binding to the main target) and binding to M1, M2 and M3 (binding without target) are determined using the H1, M1, M2 and M3 binding assays described above.

Other in vitro selection criteria for loxapine analogs include hERG binding. The binding to the main target and the binding without target are determined as described above. If the test compound shows binding to the

35 main target (H1) and the ratio between the main target binding / desired targetless binding, hERG binding (targetless binding) is determined using a comparative hERG blocking study to assess the effect of a given test compound in cloned hERG channels expressed in mammalian cells. (See, eg, Brown and Rampe, Pharmaceutical News 7: 15–20 (2000); Rampe et al., FEBS Lett., 417: 28–32 (1997); Weirich and Antoni, Basic Res. Cardiol. 93 Suppl. 1: 125–32 (1998); and Yap and Camm, Clin. Exp. Allergy, 29 Suppl 3, 174–81 (1999)).

40 The non-target junction of hERG, the cardiac potassium channel responsible for the rapid delayed rectifying current (IKr) of human ventricles, is assessed because inhibition of IKr is the most common cause of possible prolongation of cardiac action by non-cardiac drugs . (See Brown and Rampe (2000), Weirich and Antoni (1998); and Yap and Camm (1999)). The possible duration of the increased action causes the prolongation of the QT interval that has been associated with a dangerous ventricular arrhythmia, the twisting of the tips. (Brown and Rampe (2000)).

Four. Five In the hERG assay, hERG channels are expressed in a human embryonic kidney cell line (HEK293) that lacks endogenous Ikr. Expression in a mammalian cell line is preferred to transient expression in Xenopus oocytes, as this demonstrates a constant sensitivity of 10–100 times lower towards hERG channel blockers. (See, Rampe 1997).

In one embodiment of the hERG test, the positive control (ie, the reference compound) is terfenadine (Sigma, St. Louis, MO) which has been observed that, at a concentration of 60nM, blocks the hERG current at approximately 75% The test compounds are administered in physiological saline buffered with HEPES (HB-PS ) + 0.1% dimethylsulfoxide (DMSO). Each test compound is applied at a concentration of 10IM to HEK293 cells that express hERG (n 3, where n = the number of cells). The cells are exposed to the test compound for the time necessary to reach a stable blockage, but not exceeding 10 minutes. The positive control (60mM terfenadine) is applied to two cells (n 2).

The cells exposed to hERG are then transferred to a recording chamber and superfused with HB-PS solution. The pipette solution for whole cell registries includes potassium aspartate (130mM), MgCl2 (5mM), EGTA (5mM), ATP (4mM) and HEPES (10mM) at a pH adjusted to 7.2 with KOH. The initial and stable blockage of the hERG current due to the test compound is measured using a pulse pattern with fixed amplitudes (depolarization: +20 mV for 2 seconds; repolarization: –50 mV for 2 seconds), repeated at intervals of 10 seconds from a retention potential of –80 mV. The maximum tail current is measured during the 2-second stage up to –50 mV. A stable state is maintained for at least 30 seconds before applying the test compound or the positive control compound. Maximum tail currents are measured until a new stable state is reached.

In addition to the in vitro selection criteria described above, the loxapine analogs of the invention are selected using the following sleep-wake and physiological evaluations.

Non-MOR sleep: loxapine analogues are selected if in adult male Wistar rats: (i) the maximum non-MOR amount exceeds 55% of non-MOR sleep per hour no later than the third hour after treatment; and (ii) the nature of this increase in non-MOR sleep is such that the total net cumulative increase in non-MOR sleep in the first 6 hours after treatment (adjusted with respect to the reference to the corresponding 24 hours prior to the circadian cycle or in relation to the control vehicle treatment) is not less than 20 minutes in total for the dose of compound that produces the maximum sleep consolidation measured by the duration of the sleep session when the drug is administered orally.

The term "maximum non-MOR sleep time" is defined as an absolute maximum amount of non-MOR sleep per hour after treatment, with the administration of the drug being performed in the circadian hour (HC) 18, which corresponds to 6 hours after turning off the light in a night lab rat that undergoes a light – dark cycle LO

12:12 (12 hours of light and 12 hours of darkness). The nominal criteria of 55% non-MOR sleep per hour equals 33 minutes of non-MOR sleep per hour.

As used herein, the term "cumulative non-MOR sleep" is defined as the total net cumulative increase in the number of minutes of non-MOR sleep measured during the entire soporific effect of the drug that, commonly, but not always, occurs in the first 6 hours after treatment, adjusted with respect to the total net accumulated number of minutes of non-MOR sleep that occurs during the corresponding reference times without treatment of the day recorded 24 hours before compared to treatment with similar control vehicle.

As defined herein, the term "sleep session" refers to a differentiated episode of continuous or near-continuous sleep consisting of non-MOR sleep, MOR sleep or non-MOR sleep phases and MOR sleep, the episode being defined earlier. and then for more than two contiguous 10-second stages of waking state. The following non-restrictive description illustrates this concept: WWWWSSSSWSSSSSSSWWSSSSSSSWWWW, in which each letter represents the predominant state of awakening (S = sleep, W = wakefulness) observed every 10 seconds. The "sleep" session lasts 21 times of 10 seconds or 3.5 minutes.

Sleep consolidation: Loxapine analogues are selected if, in adult male Wistar rats, (i) the absolute duration of the longest continuous sleep episodes (ie, "sleep session") after treatment is greater than 13 minutes; (ii) the longest net sleep session after treatment is greater than or equal to 3 minutes adjusted with respect to a reference 24 hours before and calculated with respect to vehicle treatment; and (iii) the average absolute duration of each sleep session when the average per hour is calculated, in an hour calculated every hour, is greater than or equal to 5 minutes. The selection criteria mentioned above assume that the sleep and wake phases are determined continuously every 10 seconds (eg, "times" of 10 seconds of sleep score), that sleep and wakefulness are measured polygraphically using EEG and EMG criteria, and that sleep episodes (composed of non-MOR and / or MOR sleep) are defined as continuous "sessions" until the episode is interrupted in more than two contiguous periods of 10 seconds of wakefulness.

As used herein, the term "longest duration of the sleep session" is defined as the total number of minutes an animal remains asleep (non-MOR and / or MOR sleep phases) during the single most sleep session. long that takes place at the beginning of a given time after treatment. The measurement criteria of the “duration of the sleep session” assume that sleep is measured continuously in times of 10 seconds and the value is assigned based on the predominant state, calculated or otherwise determined as a sleep phase differentiated (in which the phases of the dream are defined as non-MOR sleep, MOR sleep or wakefulness) during the 10-second interval that defines the time.

The term "average duration of the sleep session" is defined as the average duration (in minutes) of each and every episode or sleep session that begins at a given time regardless of the individual duration of each episode or session.

Side effects measured simultaneously: loxapine analogs are selected if, in adult male Wistar rats, these compounds (i) do not produce appreciable amounts of rebound insomnia; (ii) they do not significantly inhibit MOR sleep; and (iii) they do not disproportionately inhibit locomotive activity and / or motor tone compared to the normal effects of sleep itself. The threshold of the definitions of these three side effect variables is as follows:

"Rebound insomnia" is defined as the period of rebound, paradoxical or compensatory rebound that occurs after the sleep-promoting effects of a hypnotic or soporific agent. Bounce insomnia is commonly observed during the usual resting phase of the circadian cycle 6–18 hours after treatment at HC 18 (6 hours after the light goes out, given LO 12:12), but it can occur to anyone of the first 30 hours after treatment. Bounce is considered unacceptable when, in an adult male Wistar rat, the excess accumulated wakefulness associated with rebound insomnia is greater than 10% of the reduction in the average non-MOR sleep times per hour during the resting phase of the circadian cycle after treatment (lights on).

In adult male Wistar rats, rebound insomnia manifests as an increase in wakefulness with respect to the corresponding reference times (24 hours before) after the drug's inducing sleep effect, and rebound insomnia is measured cumulatively .

“MOR sleep inhibition” is defined as the reduction of MOR sleep time after treatment to HC 18 (6 hours after the light goes out, LO 12: 112) or to HC 5 (5 hours after the light is turned on ; LO 12:12). Compounds that reduce MOR sleep time by more than 15 minutes (in relation to the reference and adjustment made for vehicle treatment) are considered unacceptable when administered well to HC 18 or HC 5.

As defined herein, "disproportionate inhibition of locomotive activity" is a reduction in locomotive activity greater than normal and the expected reduction in behavioral activity attributable to sleep. Logic dictates that if an animal is asleep, normally , there will be a corresponding reduction in locomotive activity. If the hypnotic or soporiferous compound reduces locomotor activity levels by an excess of 20% more than is solely attributable to sleep, the compound is considered unacceptable. It is possible to quantify the locomotive activity (ALM) or the motor tone objectively using any form of monitoring of the behavioral locomotive activity (nonspecific movements, telemetric activity monitoring, three-dimensional movement detection devices, movement activity on wheels, scanning measures , electromyographic record, etc.), as long as the measurement is performed simultaneously to the desired sleep-wake measurements in the same animal.

In one embodiment, the locomotive activity in the animal's cage is measured using a biotelemetric device surgically implanted in the peritoneal cavity of the animal; The implanted device and the associated telemetric receiver detect if and how much the animal moves in the cage. Sleep and wakefulness are measured at times of 10 seconds simultaneously. Locomotive activity counts per unit of time are divided by the concurrent amount of vigil for the same unit, obtaining a measure of the "intensity of locomotive activity" (IALM) for that unit of time. Hypnotic or soporiferous compounds administered to HC 18 (6 hours after the light goes out; LO 12:12) that would decrease locomotive activity per unit of wake time by more than 20% with respect to the vehicle would be considered unacceptable.

In another embodiment, the loxapine analogs of the invention are selected using sleep-wake and physiological in vivo measurement criteria shown in Table 5:

Table 5

<dl><dt>SCORE – 2000 </dt><dd>Absolute value Change of reference value with respect to vehicle only </dd></dl>

<dl><dt>Maximum no MOR time </dt><dd>&gt; 55% of maximum sleep / hour Not applicable </dd></dl>

<dl><dt>No cumulative MOR </dt><dd>Not applicable &gt; 20 minutes at ED 100 for MSBL at T1–6 </dd></dl>

<dl><dt>Longest sleep session </dt><dd>Absolute maximum of&gt; 17 minutes &gt; 5 minutes </dd></dl>

<dl><dt>Average sleep session </dt><dd>Absolute maximum of&gt; 6 minutes Not used in SAR cuts </dd></dl>

<dl><dt>Bounce insomnia </dt><dd>&lt;10% reduction in non-MOR sleep times per hour during the resting phase of the circadian cycle after treatment (lights on) Not applicable </dd></dl>

<dl><dt>MOR sleep inhibition </dt><dd>Not applicable Does not exceed 15 minutes, Rx to HC 5 </dd></dl>

<dl><dt>IALM </dt><dd>Not applicable Does not exceed 20% of the IALM reduction </dd></dl>

The procedures to evaluate these sleep-wake and physiological criteria are described above. The "absolute value" shown in the second column of Table 5 refers to the value determined for each compound of

5 test, while the “change” value shown in the third column of Table 5 reflects an adjusted value in which the absolute value is the difference with respect to the vehicle, when the vehicle values ​​are adjusted with respect to the reference value .

In some embodiments, the longest sleep session lasts longer than 13 minutes. In others, it lasts more than 17 minutes. In some embodiments, the longest net sleep session after treatment

10 It lasts greater than or equal to 3 minutes. In others, it lasts greater than or equal to 6 minutes.

Other in vivo sleep-wake and physiological evaluation criteria used to select the loxapine analogs of the invention include the measurement of acute body temperature and latent body temperature as a change in the reference value with respect to the vehicle. The change in acute body temperature should not exceed –0.50ºC and the change in latent body temperature should not exceed + 0.50ºC in the time of 1–6 hours. The

fifteen Acute body temperature (T1–6) is adjusted with respect to the corresponding reference value measured 24 hours before with respect to the vehicle (the decrease with respect to the vehicle). The latent body temperature, measured at 7–18 hours after treatment with the drug (T7–18), is adjusted with respect to the corresponding reference value measured 24 hours before with respect to the vehicle (the decrease with respect to the vehicle).

The compounds modulate sleep in several ways, including the decrease in time until sleep is reconciled, the increase in the average duration of the sleep session and the increase in the maximum duration of the sleep session.

The compounds, or their pharmaceutically acceptable salts, are administered oral, nasal, transdermal, pulmonary, by inhalation, buccal, sublingual, intraperitoneal, intravenous, rectal, intrapleural, intrathecal and parenterally. In one embodiment, the compound is administered orally. Any person skilled in the art will recognize the advantages of certain routes of administration.

25 The compounds of the present invention serve for the treatment of a variety of sleep disorders, including alteration of the circadian rhythm, insomnia, parasomnia, sleep apnea syndrome, narcolepsy and / or hypersomnia. In one embodiment, the compounds treat circadian rhythm disturbances, such as, for example, jet lag, shift shifts, delayed sleep phase syndrome, advanced sleep phase syndrome and sleep cycle disorder– 24 hour different vigil. In another embodiment, the compounds treat the

30 insomnia, including, for example, extrinsic insomnia, psychophysiological insomnia, height insomnia, restless legs syndrome, periodic limb movement disorder, medication-dependent insomnia, drug dependence insomnia, alcoholism insomnia and insomnia associated with mental disorders.

In another embodiment, the compounds treat parasomnias, including sleepwalking, night dread, behavioral disorder associated with MOR sleep, sleep bruxism and sleep enuresis. In yet another embodiment, the

35 Procedure treats sleep apnea disorder, including central sleep apnea, obstructive sleep apnea and mixed sleep apnea. In addition, the compounds treat other sleep disorders such as narcolepsy or hypersomnia.

In some embodiments, a compound is administered in the form of a pharmaceutically acceptable salt. Any person skilled in the art will recognize the various methods for creating pharmaceutically acceptable salts and

40 Identify the appropriate salt. In one embodiment, the compound or a pharmaceutically acceptable salt thereof is included in a pharmaceutical composition.

As used herein, the term "sleep disorder" includes conditions recognized by one skilled in the art as sleep disorders, for example, conditions known in the art or conditions proposed as sleep disorders or discovered as sleep disorders See, for example, Thorpy, MJ, "International Classification of Sleep Disorders"; Revised: Diagnostic and Coding Manual. American Sleep Disorders Association; Rochester, Minnesota 1997; and “ICD – 9 – CM, International Classification of Diseases,” ninth review, clinically modified, National Center for Health Statistics, Hyattsville, MD.

For example, sleep disorders can be classified in general into dysomnias, eg, intrinsic, extrinsic or circadian rhythm disorders; parasomnias, eg, disorders associated with awakening, sleep-wake transition and rapid eye movement (MOR), and other parasomnias; disorders associated with mental, neurological and other medical disorders; and other sleep disorders.

Intrinsic sleep disorders include, for example, psychophysiological insomnia, misperception of sleep status, idiopathic insomnia, narcolepsy, recurrent hypersomnia, idiopathic hypersomnia, post-traumatic hypersomnia, obstructive sleep apnea syndrome, central apnea syndrome sleep, central alveolar hypoventilation syndrome, periodic limb movement disorder and restless legs syndrome.

Extrinsic sleep disorders include, for example, inadequate sleep hygiene, environmental sleep disorder, height insomnia, sleep adjustment disorder, sleep insufficiency syndrome, disorder of sleep behavior patterns, association disorder with the onset of sleep, food allergy insomnia, nighttime food or drink syndrome, hypnotic dependence sleep disorder, stimulant dependence sleep disorder, sleep disorder due to alcoholism or toxin-induced sleep disorder.

Sleep disorders related to the circadian rhythm include, for example, time zone change syndrome (jet lag), sleep disorder due to changes in work shifts, irregular sleep-wake pattern, delayed sleep phase syndrome, Advanced sleep phase syndrome and sleep-cycle disorder different from 24 hours.

Sleep disorders upon waking include, for example, confusing awakenings, sleepwalking and night terrors.

Sleep-wake transition disorders include, for example, rhythmic movement disorder, early sleep, speaking in dreams and nocturnal cramps in the lower extremities.

Sleep disorders associated with MOR include, for example, nightmares, sleep paralysis, sleep-related erection problems, painful sleep-related erections, sleep-related sinus stop MOR, and MOR behavioral sleep disorder.

Other parasomnias include, for example, sleep bruxism, sleep enuresis, difficulty swallowing syndrome associated with sleep, paroxysmal nocturnal dystonia, sudden and unexplained nocturnal death syndrome, primary snoring, infant sleep apnea, congenital central hypoventilation syndrome , sudden infant death syndrome and benign neonatal sleep myoclonus.

"Sleep disorders" also appear in subjects suffering from other medical disorders, diseases or injuries, or in subjects who are being treated with other medications or medical treatments and, as a consequence, have difficulty falling asleep and / or remain asleep, or do not enjoy a restful or restorative sleep, eg, a subject with sleep deprivation. For example, some subjects have difficulty sleeping after medical treatment for other conditions, e.g. eg, chemotherapy or surgery, or for suffering pain or other effects of physical injury.

It is widely known in the art that certain medical disorders, for example, central nervous system (CNS) disorders, e.g., mental or neurological disorders, e.g., anxiety, may have a sleep disorder component, e.g. .ej., sleep deprivation. Thus, "treatment of a sleep disorder" also includes treating a sleep disorder component of other disorders, e.g. eg CNS disorders. In addition, treatment of the sleep disorder component of CNS disorders may also have a beneficial effect of improving other symptoms associated with said disorder. For example, in some subjects suffering from anxiety linked to sleep deprivation, treating the sleep deprivation component also treats the anxiety component. Thus, the present invention also includes a method of treating such medical disorders.

For example, sleep disorders linked to mental disorders include psychosis, mood disorders, anxiety disorders, panic disorder, addictions and the like. Specific mental disorders include, for example, depression, obsessive compulsive disorder, affective disorder / neurosis, neurosis / depressive disorder, anxiety neurosis, dysthymic disorder, behavioral disorder, mood disorders, schizophrenia, manic depression, delirium and alcoholism .

Sleep disorders associated with neurological disorders include, for example, brain degeneration disorders, dementia, Parkinson's disease, Huntington's disease, Alzheimer's disease, fatal familial insomnia, sleep-related epilepsy, electrical epileptic state of sleep and pains of Head related to sleep. Sleep disorders associated with other medical disorders include, for example, African trypanosomiasis, nocturnal cardiac ischemia, chronic obstructive pulmonary disease, sleep-related asthma, sleep-related gastrointestinal reflux, peptic ulcer disease and fibrositis syndrome.

In some circumstances, sleep disorders are also associated with pain, eg, neuropathic pain linked to restless legs syndrome; migraine; hyperalgesia; fibromyalgia, pain; normal or exaggerated increase in pain sensation, such as hyperalgesia, causalgia and allodynia; acute pain; burn pain; atypical facial pain; neuropathic pain; Back pain; complex regional pain syndromes type I and II; arthritic pain; sports injury pain; pain related to an infection, eg, HIV, postpolio syndrome and post-herpetic neuralgia; phantom limb pain; labor pain; cancer pain; pain after chemotherapy; pain after a stroke; pain after an operation; neuralgia; conditions associated with visceral pain, including irritable bowel syndrome, migraine and angina.

Other sleep disorders include, for example, a person who sleeps little, a person who sleeps a lot, subvigilia syndrome, fragmentary myoclonus, sleep hyperhidrosis, sleep disorder associated with the menstrual cycle, sleep disorder associated with pregnancy, terrifying hypnagogic hallucinations , neurogenic sleep-related tachypnea, sleep-related laryngospasms and sleep asphyxiation syndrome.

Insomnia is commonly classified as initial insomnia, in which the subject needs more than 30 minutes to fall asleep; and sleep maintenance insomnia, in which the subject spends more than 30 minutes awake during an expected period of sleep or, for example, wakes up before the time he wishes to wake up with difficulties or inability to fall asleep again . The disclosed compounds may be effective in the treatment of the initial and maintenance of sleeplessness, of insomnia as a result of disorders of circadian rhythm adjustment or insomnia caused by CNS disorders. An embodiment consists in treating a subject of a circadian rhythm adjustment disorder. Another embodiment is to treat a subject of insomnia caused by a mood disorder. In other embodiments, a subject of sleep apnea, sleepwalking, night terrors, restless legs syndrome, initial insomnia and / or sleep maintenance insomnia is treated; or more preferably, initial insomnia or sleep maintenance insomnia. The disclosed compounds may be effective in the treatment of insomnia of sleep onset. The disclosed compounds may also be effective in the treatment of sleep maintenance insomnia.

The dosage schedule of the compounds is selected according to a variety of factors including type, species, age, weight, sex and health status of the patient; severity of the condition to be treated; route of administration; renal and hepatic function of the patient; and the compound or salt of the same employee in particular. Any doctor or veterinarian skilled in the art can easily determine and prescribe the effective amount of the drug necessary to prevent, counteract or stop the progress of the condition.

The oral doses of the present invention, when used for the indicated effects, will vary between about 0.05 to 5,000 mg / day orally. The effective amounts of the disclosed compounds commonly range between about 0.01 mg / kg per day and about 100 mg / kg per day and, preferably, between 0.1 mg / kg per day and about 10 mg / kg / day. In "Remington: the Science and Practice of Pharmacy", XIX edition, Mack Publishing Co., Easton, PA (1995), techniques for the administration of the disclosed compounds of the invention can be found.

For example, in some embodiments, an acid salt of a compound containing an amine or other basic group is obtained by reacting the compound with a suitable organic or inorganic acid, such as hydrogen chloride, hydrogen bromide, acetic acid, acid perchloric and similar. Compounds with a quaternary ammonium group also contain a counterion, such as chloride, bromide, iodide, acetate, perchlorate and the like. Other examples of such salts include hydrochlorides, hydrobromides, sulfates, methanesulfonates, nitrates, maleates, acetates, citrates, smokers, tartrates (e.g., (+) - tartrates, (-) - tartrates or mixtures thereof, including mixtures thereof racemic), succinates, benzoates and salts with amino acids such as glutamic acid.

Salts of compounds containing a carboxylic acid or other acid functional groups are prepared by reaction with a suitable base. Such a pharmaceutically acceptable salt is formed with a base that provides a pharmaceutically acceptable cation, which includes alkali metal salts (especially sodium and potassium), alkaline earth metal salts (especially calcium and magnesium), aluminum and ammonium salts, as well. as salts formed from physiologically acceptable organic bases, such as trimethylamine, triethylamine, morpholine, pyridine, piperidine, picoline, dicyclohexylamine, N, N'-dibenzylethylenediamine, 2-hydroxyethylamine, bis- (2-hydroxyethyl) amine, tri– (2-hydroxyethyl) amine, procaine, dibenzyl-piperidine, N-benzyl- -phenethylamine, dehydroabyethylamine, N, N'-bisdehydroiethylamine, glucamine, N-methylglucamine , collidine, quinine, quinoline and a basic amino acid such as lysine and arginine.

In some embodiments, certain compounds and their salts also come in the form of solvates, for example, hydrates, and the present invention includes each solvate and mixtures thereof.

In one embodiment, the compounds described herein, and their pharmaceutically acceptable salts, are used in pharmaceutical preparations in combination with a pharmaceutically acceptable carrier or diluent. Suitable pharmaceutically acceptable carriers include inert solid fillers or diluents and sterile organic or aqueous solutions. The compounds will be present in such pharmaceutical compositions in amounts sufficient to provide the desired dose amount in the range described herein. In "Remington: the Science and Practice of Pharmacy" above, techniques for the formulation and administration of the disclosed compounds of the invention are found.

Commonly, the compound is prepared for oral administration in which the disclosed compounds or their salts are combined with a liquid or solid carrier or diluent suitable to form capsules, tablets, pills, powders, syrups, solutions, suspensions and the like.

Tablets, pills, capsules and the like contain from about 1 to about 99 percent by weight of the active ingredient and a binder, such as gum tragacanth, acacia, corn starch or gelatin; excipients such as dicalcium phosphate; a disintegrating agent, such as corn starch, potato starch or alginic acid; a lubricant such as magnesium stearate; and / or a sweetening agent such as sucrose, lactose, saccharin, xylitol and the like. When a unit dosage form is a capsule, it usually contains, in addition to materials of the above type, a liquid carrier such as a fatty oil.

In some embodiments, various other materials are present as coatings or to modify the physical form of the dosage unit. For example, in some embodiments, the tablets are coated with lacquer, sugar or both. In some embodiments, a syrup or elixir contains, in addition to the active ingredient, sucrose as a sweetening agent, methyl and propylparabenos as preservatives, a colorant and a flavoring agent, such as cherry or orange aroma and the like.

For some embodiments related to parenteral administration, the disclosed compounds, or their salts, solvates or polymorphs, can be combined with sterile organic or aqueous media to form injectable solutions or suspensions. Injectable compositions are preferably aqueous isotonic solutions or suspensions. The compositions may be sterilized and / or contain adjuvants such as preservatives, stabilizers, humectants or emulsifiers, dissolution promoters, salts for regulating osmotic pressure and / or buffers. In addition, they may also contain other therapeutically valuable substances. The compositions are prepared according to conventional mixing, granulation or coating procedures, respectively, and contain about 0.1 to 75%, preferably, about 1 to 50%, of the active ingredient.

For example, injectable solutions are produced using solvents such as sesame oil or peanut oil or aqueous propylene glycol, as well as aqueous solutions of pharmaceutically acceptable water-soluble salts of the compounds. In some embodiments, the dispersions are prepared in glycerol, liquid polyethylene glycols and mixtures thereof in oils. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent the growth of microorganisms. The terms "parenteral administration" and "parenterally administered", as used herein, mean modes of administration other than enteral and topical administration, usually by injection, and include, without limitation, intravenous, intramuscular injection and infusion, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal and intrasternal.

For rectal administration, suitable pharmaceutical compositions are, for example, topical preparations, suppositories or enemas. Suppositories are advantageously prepared from fat emulsions or suspensions. The compositions may be sterilized and / or contain adjuvants such as preservatives, stabilizers, humectants or emulsifiers, dissolution promoters, salts for regulating osmotic pressure and / or buffers. In addition, they may also contain other therapeutically valuable substances. The compositions are prepared according to conventional mixing, granulation or coating procedures, respectively, and contain about 0.1 to 75%, preferably, about 1 to 50%, of the active ingredient.

In some embodiments, the compounds are formulated to administer the active agent by pulmonary administration, eg, the administration of an aerosol formulation containing the active agent from, for example, a manual spray pump, a nebulizer or an inhaler. pressurized dispenser In some embodiments, suitable formulations of this type also include other agents, such as antistatic agents, to maintain the disclosed compounds as effective aerosols.

A drug delivery device for administering aerosols comprises a suitable aerosol can with a metering valve containing the pharmaceutical aerosol formulation as described and an actuating housing adapted to hold the can and allow the administration of the drug. The canister of the drug delivery device has an air chamber that represents more than about 15% of the total volume of the canister. Usually, the polymer intended for pulmonary administration is dissolved, suspended or emulsified in a mixture of a solvent, surfactant and propellant. The mixture is kept under pressure in a canister that has been sealed with a metering valve.

For a nasal administration, a solid or liquid vehicle can be used either. The solid carrier includes a coarse-grained powder having a particle size in the range of, for example, about 20 to about 500 micrometers, and such formulation is administered by rapid inhalation through the nostrils. In some embodiments in which the liquid carrier is used, the formulation is administered as a spray or nasal drops and includes oil or aqueous solutions of the active ingredients.

Also contemplated are formulations that are rapid dispersion pharmaceutical forms, also known as "instant dose" forms. In particular, some embodiments of the present invention are formulated as compositions that release their active ingredients in a short period of time, e.g., commonly, in less than about five minutes, preferably, in less than about ninety seconds, more preferably , in less than about thirty seconds, and most preferably, in less than about ten or fifteen seconds. Such formulations are suitable for administration to a subject by a variety of routes, for example, by introduction into a body cavity or application on a wet body surface or an open wound.

Commonly, an "instant dose" is a solid pharmaceutical form that is administered orally, which quickly disperses in the mouth and hence does not require a great deal of swallowing and allows the compound to be swallowed or absorbed quickly through the oral mucous membranes. In some embodiments, rapidly dispersed pharmaceutical forms are also used in other applications, including the treatment of wounds, and other bodily lesions and pathological conditions in which it is not possible to release the medication by an externally supplied lotion.

The "instant dose" forms are known in the art; see, for example, effervescent pharmaceutical forms and insoluble microparticle quick release coatings in U.S. Patent Nos. 5,578,322 and 5,607,697; freeze-dried foams and liquids in U.S. Patent Nos. 4,642,903 and 5,631,023; Fusion spinning of pharmaceutical forms, in US Pat. Nos. 4,855,326; 5,380,473 and 5,518,730; manufacture of solid free forms, in US Patent No. 6,471,992; vehicle matrix based on saccharin and a liquid binder, in US Pat. Nos. 5,587,172; 5,616,344; 6,277,406 and 5,622,719; and other forms known in the art.

The loxapine analogs of the invention are also formulated as "pulsed release" formulations in which the analog is released from the pharmaceutical compositions in a series of releases (ie, pulses). Loxapine analogs are also formulated as "sustained release" formulations, in which the analog is continuously released from the pharmaceutical composition for a prolonged period.

Also contemplated are formulations, e.g., liquid formulations, which include cyclic or acyclic encapsulation or solvation agents, e.g., cyclodextrins, polyethers or polysaccharides (e.g., methylcellulose) or, more preferably, derivatives of - Polyanionic cyclodextrin with a group of sodium sulphonate salt separated from the lipophilic cavity by an alkyl ether or polysaccharide spacer group. In one embodiment, the agent is methyl cellulose. In another embodiment, the agent is a polyanionic cyclodextrin derivative with a sodium sulphonate salt separated from the lipophilic cavity by a butyl ether spacer group, eg, CAPTISOL® (CyDex, Overland, KS). The person skilled in the art can evaluate the proportions of the formulation between the suitable agent and the disclosed compound by preparing a solution of the agent in water, eg, a solution of 40% by weight; preparing serial dilutions, eg, to make solutions of 20%, 10%, 5%, 2.5%, 0% (control) and the like; adding an excess (compared to the amount that can be dissolved by the agent) of the disclosed compound; mixing under appropriate conditions, eg, by heating, stirring, ultrasonic treatment and the like; centrifuging or filtering the resulting mixtures to obtain transparent solutions; and analyzing the concentration of the revealed compound of the solutions.

In addition to the therapeutic formulations described above, a therapy that includes the compounds of the present invention optionally includes administration together with one or more additional therapies, e.g., drugs or physical or behavioral treatments (e.g., light therapy , electrostimulation, modification of habits, cognitive therapy, modification of the circadian rhythm and the like). Such a practice is called "combination therapy." The other or other combination therapy therapies include therapies recognized by the person skilled in the art as desirable in combination with the compound of the invention, for example, therapies known in the art or therapies that have been proposed or discovered in the art for the treatment of sleep disorders or the treatment of diseases associated with sleep disorders, for example, therapies for any of the sleep disorders or other conditions disclosed herein. In some embodiments, the compound is administered as a combination therapy while, in other embodiments, it is administered as a single therapy.

Commonly, the compound is administered as a single therapy.

The person skilled in the art will appreciate that a therapy administered in combination with the compounds of the present invention is directed to the same or a different target disorder to which the compounds of the present invention are being directed. First the administration of the compound of the invention is carried out, followed by the other therapy or, alternatively, it may be that the administration of the other therapy is performed first. The other therapy is any known in the art to treat, prevent or reduce the symptoms of the target disorder, eg, a sleep disorder or other disorders, eg, other CNS disorders. In addition, some embodiments of the present invention have compounds administered in combination with other therapies known for the target disorder. Moreover, the other therapy includes any agent beneficial to the patient when administered in combination with the disclosed compound.

For example, in some embodiments in which the other therapy is a drug, it is administered as a separate formulation or in the same formulation as the compound of the invention. A compound of the invention is administered in a combination therapy with any one or more of the medications that require or do not require a commercially available prescription, including, but not limited to, antihistamines, antimicrobials, fungicides, germicides, hormones, antipyretics, antidiabetics , bronchodilators, antidiarrheals, antiarrhythmic agents, coronary dilation agents, glycosides, spasmolytics, antihypertensive agents, antidepressants, anxiolytics, other psychotherapeutic agents, steroids, corticosteroids, analgesics, cold medications, vitamins, sedatives, hypnotics, contraceptives, non-steroidal anti-inflammatory drugs, hypoglycemic, hypocholesterolemic, anticonvulsant, other antiepileptic, immunomodulatory, anticholinergic, sympatholytic, sympatholytic, sympatholytic, sympatholytic prostaglandins that have various pharmacological, diuretic, sleep inducing activities, antihistamines, antineoplastic, oncolytic, antiandrogen, antimalarial, antileprosy and other various types of drugs. See Goodman and Gilman, "The Basis of Therapeutics" (VIII edition, Pergamon Press, Inc., USA, 1990) and "The Merck Index" (XI edition, Merck &amp; Co., Inc., USA). , 1989).

Examples of drugs used in combination with the compounds of the invention include, but are not limited to, AMBIEN® STILNOX® (zolpidem tartrate), indiploon, ESTORRA ™ (eszopiclone), NEURONTIN® (gabapentin), LYRICA® (pregabalin) , eplivanserin, SONATA® (zaleplon), ESTORRA ™ (eszopiclone), ZOPICLONE ™ (imovane), DESYREL ™ (trazodone hydrochloride), SEROQUEL® (quetiapine fumarate), CLOZARIL® (clozapine), ZYPREXA ™ (RYPREXA ™) ® (risperidone), M100907 and LUNESTA ™.

In one embodiment, the compounds of the invention are useful in combination with a mechanical therapy such as PPC. The "PPC of the respiratory tract" or the "continuous positive pressure (PPC) of the respiratory tract" is a mechanical device for the treatment of sleep apnea and other sleep-related breathing disorders (including snoring). Treatment with a PPC airway device is commonly administered by the patient's nose or mouth.

A subject undergoing treatment with PPC of the airways should wear a plastic mask fitted over the nose while sleeping. The mask is attached to a compressor that forces air through the nose creating positive pressure on the subject's airways. The principle of the procedure is that the pressure generated on the airways produces a "splinting" action that prevents or decreases the collapse of the airways and, therefore, obstructive sleep apnea. Although an effective therapeutic response is observed in the majority of subjects receiving PPC treatment of the respiratory tract, many subjects cannot withstand the apparatus or pressure and refuse treatment. In addition, recent covert monitoring studies have shown that long-term maintenance of airway PPC treatment is very unsatisfactory. It is known that subjects remove their masks while they sleep.

In one aspect, the compound of the invention is administered in combination with a PPC airway device to enhance sleep. In another aspect, the compound of the invention is administered in combination with a PPC airway device to improve sleep. In another aspect, the compound of the invention is administered in combination with a respiratory tract PPC device to improve the maintenance of the respiratory tract PPC treatment. Without wishing to be bound by theory, it is believed that by administering an effective amount of a sleep-promoting compound of the invention to a subject in combination with a PPC treatment of the respiratory tract, the subject will sleep better and more. deeply and, therefore, will not tend to remove the mask.

In one embodiment, the compound of the present invention is administered prior to the treatment of respiratory tract PPC. In another embodiment, the compound of the present invention is administered substantially at the same time as the treatment of airway PPC. In one embodiment, parallel administration of an effective amount of the compound is performed by adding an additional aerosol channel to the air pressure treatment portion of the airway PPC device, thereby administering the compound of the present invention in a form. nebulized by a nasal or oral mask of the PPC airway device. Alternatively, an effective amount of the compound may be added to the water or liquid reservoir that is commonly part of the respiratory tract PPC treatment device.

With the use of the PPC mask treatment of the respiratory tract, the compound of the invention is administered at a low concentration overnight or at higher concentrations, in the form of bowling, at different times at the beginning and during the night.

All publications and patent documents cited herein are incorporated herein by reference as if it were indicated that each such document or publication was specifically and individually incorporated herein by reference. It is not intended that citations from publications and patent documents be recognized as the relevant prior art, nor that they constitute any recognition as to their content or dates. From the written description of the invention, those skilled in the art will appreciate that it is possible to implement it in a variety of embodiments, and that the above description and the examples presented below are intended to be illustrative and not restrictive of the claims listed below.

Example 1: Synthesis of loxapine analogues

The compound of the invention, and its salts, solvates or related hydrates, can be synthesized by methods known to those skilled in the art. Example 10 reveals the synthesis of Compound 1.

Example 2: Sleep inducing properties of the compounds of the invention

Mammalian sleep can be divided into sleep that occurs during periods of rapid eye movement (MOR), accompanied by substantial brain activity, and periods of non-MOR sleep (NMOR), accompanied by a decrease in activity. cerebral. Commonly, a period of normal nighttime sleep is primarily occupied by NMOR sleep and, therefore, the accumulation of NMOR can serve to measure the total accumulation of sleep, e.g., a significant decrease in NMOR may be associated with insomnia. and an accumulation of "lack of sleep", eg, an accumulated physiological need for sleep that tends to last until a sufficient amount of additional sleep accumulates. Thus, an increase in NMOR linked to a treatment may indicate the efficacy of the treatment in the treatment of insomnia.

Sleep quality can be associated with continuity or maintenance of sleep. For example, a subject with sleep apnea wakes up numerous times during a period of sleep, eg, the subject has difficulty staying asleep continuously. Although such a subject may accumulate a duration of common nighttime sleep, eg, 8 hours, sleep is not restorative or restorative due to the awakening caused by sleep apnea. Thus, an increase in the longer uninterrupted sleep session (SSIL, also known as a longer sleep session) associated with a treatment may indicate the effectiveness of the treatment in increasing the continuity of sleep and, therefore, in the Sleep maintenance insomnia treatment.

Sleep-wakefulness, locomotor activity and body temperature are monitored in male Wistar rats treated with a test compound (ie, loxapine analog) initially at a concentration of 10 mg / kg. Higher and lower doses of the selected compounds are analyzed (eg, as high as 45 mg / kg and as low as necessary to establish a dose without effect). The treatments are administered to HC 18, the maximum of the period dominating by activity (6 hours after the light goes out) and causes soporific effects (sleep inducers) characterized by an increase in non-MOR sleep time, an increase in continuity of sleep, but without evidence of MOR sleep inhibition or rebound insomnia.

Sleep-wakefulness, locomotor activity and body temperature were monitored in vivo with compound. Adult male Wistar rats (250 g at the time of surgery, Charles River Laboratories, Wilmington MA) (2% isoflouran in medical grade oxygen) were anesthetized and surgically prepared with a cranial implant to allow electroencephalogram (EEG) registration ) chronic and electromyogram (EMG). Body temperature and locomotive activity were monitored by a miniature transmitter (Mini-Mitter, Bend, OR) placed by surgery on the abdomen. The cranial implant consisted of stainless steel screws (two frontal (+3.2 AP of bregma, ± 2.0 ml) and two occipital (–6.9 AP, ± 5.5 ml)) for registration by EEG. Two stainless steel cables coated with Teflon® were placed under the trapezius muscles in the neck for an EMG registration. All ends were welded to a miniature connector before surgery and sterilized with ethylene oxide gas. The implant assembly was fixed to the skull with a dental acrylic. A minimum of three weeks was left for surgical recovery.

Each rat remained in its individual log cage located in separate ventilated compartments of stainless steel cabinets with the usual design. An upper filter lifter and a low torque switch are incorporated into each cage. They were given food and water at will. During the study, a 24-hour light-dark cycle was maintained (12 hours of light, 12 hours of darkness). The animals were not disturbed for at least 48 hours before and after the treatments.

Sleep and wakefulness were determined using “SCORE – 2000 ™” (Hypnion, Worcester, MA), an Internet-based physiological and sleep-wake monitoring system. The system monitored amplified EEG (1–30 Hz bandpass filter), integrated EMG (10–100 Hz bandpass filter), body temperature and nonspecific locomotive activity (ALM) via telemetry, and fluid intake activity continuously and simultaneously . Wake-up states were classified as non-MOR sleep (NMOR), MOR sleep, wake or wake dominated by theta range every 10 seconds. The total fluid intake, locomotor activity and body temperature were quantified and recorded every minute, using algorithms for pattern matching and EEG feature extraction. From this data, the longest uninterrupted sleep session (SSIL) was obtained. The classification algorithm used EEG wake-up templates individually shown plus EMG criteria to differentiate the MOR sleep from the waking state dominated by theta range, plus the behavioral-dependent contextual rules (eg, if the animal was drinking, I was awake). Fluid intake and locomotor activity intensity (ALM) were recorded every 10 seconds, while body temperature was recorded every minute. The locomotive activity was detected by a telemetric receiver (Mini – Mitter) located under the

cage. Telemetry measurements (ALM and body temperature) were not part of the scoring algorithm; Thus, sleep score and telemetric data were independent measures.

Compound 1 was administered to HC 18, the maximum point of the period dominated by the activity, leaving sufficient time to observe how the effect of the treatment elapsed in time before switching on the light (6 hours after treatment). Compound 1 was suspended in 0.25% or 0.5% sterile methylcellulose (1-2 ml / kg). The treatments were administered orally in the form of a bolus.

A parallel group study design was used. The control vehicles were extracted from a large mixture (N&gt; 200): a subset of the mixed control vehicles was selected based on the computerized coincidence with the reference value of 24 h before the treatment of the active treatment group.

The results of the NMOR and SSIL parameters for Compound 1 were measured. Table 6 shows the representative results.

Table 6: Sleep-inducing properties of compounds

<dl><dt>No. </dt><dd>Dose NMOR SSIL </dd></dl>

<dl><dt>1 </dt><dd /><dt>1 </dt><dd> 6,0 ± 2,3 </dd></dl>

<dl><dt>3 </dt><dd> 39 ± 6 18,8 ± 3,0 </dd></dl>

<dl><dt>10 </dt><dd> 19,7 ± 6,7 </dd></dl>

<dl><dt>* the dose is in mg / kg; NMOR and SSIL are in minutes.</dt><dd /></dl>

Example 3: Irwin trace side effects

Irwin screening can provide useful information about the possible side effects of the compounds on general physiological and behavioral functions. Tracing is performed by administering test compounds orally in 0.25% aqueous methylcellulose using male Wistar rats, a species frequently used in such studies and for which reference data are available.

Irwin tracing analyzes numerous parameters in animals that have received the test compound. For example, tracking may include: effects in the cage, eg, dispersion, respiratory velocity, locomotor activity, restlessness, aggressiveness, alertness, apathy and exophthalmia; effects on the ring, eg, transfer awakening, spatial locomotion, ptosis, startle, tail lift, piloerection, touch escape, positional passivity, catalepsy, strong tightening reflex, visual positioning, clamping force, canopy auditory, cornea, pain response and cable manipulation; the parameters observed during treatment, eg, cyanosis, cutaneous blood supply, hypothermia, body tone, pupil size, pupil response to light, tearing, cleaning habits, redness, salivation and bite provocation; general scores, eg, fear, irritability, abnormal gait, abnormal body posture, tremors, tics, seizures, strange behavior, torsion, vocalization, diarrhea, number of bowel movements, number of urination, dying state, lethality and abnormalities detected. In Irwin, S; “Comprehensive observational assessment: I a. A systematic, quantitative procedure for assessing the behavioral and physiological state of the mouse ”. Psychopharmacology (Berl.) 13: 222-257, 1968, whose teachings are incorporated herein by reference in their entirety, more information can be found in this regard.

Irwin's screening of the revealed sleep inducing agents is performed by Covance (Princeton, NJ) according to Irwin's previous; Covance Standard Operating Procedure (current revision of SOP PHARM 8.10); Guide of the ICH (International Harmonization Committee) of guidelines of the competent regulatory bodies (Topic S7A; CPMP / ICH / 539/00) on pharmacological safety studies for pharmaceutical compounds in humans (November 2000); and all procedures carried out on live animals comply with the provisions of British legislation, in particular, the Law of (scientific procedures with) animals of 1986, which obliges all UK laboratories to maintain an ethical review procedure at the local level that guarantees a respectful and justified use of all animals in the facilities; that all possibilities of reduction, improvement or replacement be taken into account, and that the high standards of accommodation and care are met.

All chemical compounds were purchased from Colorcon, Ltd, Dartford Kent, UK, unless otherwise indicated and are of a purity of ACS reagent quality or higher. All formulations of test compounds are prepared on the day of dosing by the Covance Harrogate pharmacy. Test compounds are formulated in 0.25% aqueous methylcellulose at the highest concentration required. The lower doses are obtained by serial dilution of the highest concentration, using 0.25% aqueous methylcellulose. The dose levels are expressed in terms of the amount of the test compound administered regardless of purity or active content. All formulations are stored at room temperature (typically 10 to 30 ° C) in tightly closed containers and protected from light.

An adequate number of Wistar rats (Crl: WI (Glx / BRL / Han) BR: WH) are obtained from Charles River Ltd. (Margate, Kent, United Kingdom). Rats are approximately 5 weeks old and weigh between 150 and 170 g upon arrival. Animals are introduced in groups of no more than six individuals in polypropylene cages (33 x 15 x 13 cm)

or (45 x 28 x 20 cm) with solid floors and lamellae of quality wood 10 (Datesand Ltd., Cheshire, United Kingdom) as beds. The cages are cleaned and dried before use. Aspen chewable blocks are placed in the cages as a form of environmental enrichment. Typically, confinement rooms are maintained between acceptable temperature and relative humidity limits (nominally 19 to 25 ° C and 40% to 70%, respectively). These rooms are illuminated with fluorescent light for 12 of the 24 hours of the cycle and are designed to receive at least 15 changes of pure air per hour. Diet (RM1 (E) .SQC. (Special Diets Services Ltd. Witham, United Kingdom) and main tap water are provided at will (except during treatment). Their specific constituents are periodically analyzed and not found to contain any entity biological or chemical that may interfere with the analysis system. Upon arrival, the health status of all animals is examined. The acclimatization period of the animals is at least 5 days. During this time, animals identify their cage tags. The veterinarian performs an exam before starting any experimental procedure to ensure its suitability for the study. Before starting the study, the animals are randomly distributed in treatment groups and individually marked in the tail as they come to hand. At the end of the study, the animals are subjected to euthanasia.

Each animal receives a single oral administration of vehicle or test article, using a constant dose of 1 mg / kg. Individual doses are based on individual body weights obtained on the day of dosing.

The previous Irwin tracking parameters are systematically analyzed according to the relevant controls. In general, changes induced by the drug, absent in normal animals, are recorded using increasing integers, in which 0 equals normal (being able to also use +/– = present / absent). The parameters present in normal animals are scored using an integer that allows the recording of increases and decreases. Detailed observations are made at 30, 60, 90, 180 and 300 minutes after the dose. The animals are kept for a period of 7 days after the dose, during which time they are observed daily for gross signs of toxicity and mortality.

Example 4: Side effects on hERG of disclosed agents

The cardiac potassium channel, hERG, is responsible for the rapid delayed rectifier current (Ikr) produced in the ventricles of humans. This channel has been chosen for evaluation, because Ikr inhibition is the most common cause of possible prolongation of undesirable cardiac action by non-cardiac drugs. The increase in the possible duration of the action causes the prolongation of the QT interval, which has been associated with dangerous ventricular arrhythmia, torsion of the tips (Brown, AM; Rampe, D. (2000). “Drug – induced long QT syndrome: is hERG the root of all evil? ”; and Pharmaceutical News 7, 15–20; Rampe, D; Roy, ML; Dennis, A; Brown, AM. (1997), all of whose teachings are incorporated into the present memory by reference). The hERG channels were expressed in a human embryonic kidney cell line (HEK293) that lacked endogenous Ikr. Expression in a mammalian cell line is preferred to transient expression in Xenopus oocytes, as this demonstrates a constant sensitivity of 10–100 times lower towards hERG channel blockers. See, for example: "A mechanism for the pro-arrhythmic effects of cisapride (Propulsid): high affinity blockade of the human cardiac potassium channel hERG". FEBS Lett. 417, 28–32; Weirich, J; Antoni, H. (1998); “Rate – dependence of anti – arrhythmic and pro – arrhythmic properties of class I and class III anti – arrhythmic drugs”. Basic Res Cardiol 93 Suppl 1, 125-132; and Yap, YG; Camm, AJ. (1999); and "Arrhythmogenic mechanisms of non-sedating antihistamines." Clin. Exp. Allergy 29 Suppl 3, 174-181. All the teachings of the preceding articles are incorporated herein by reference.

The in vitro effects of the sleep inducing agents revealed on the hERG channel current (gene related to human ether – a – go – go) were determined (Ikr, the rapidly activating delayed cardiac potassium rectifier current) by the Chan test (Cheveland, OH) according to the standard operating procedures of the Chan test.

All chemical compounds used were purchased from Sigma (St. Louis, MO) unless otherwise indicated and were of a grade ACS reagent purity or higher. The stock solutions of the test articles and terfernadine (positive control) were prepared using dimethylsulfoxide (DMSO) and stored in a freezer. The concentrations of the test article and the positive control were prepared by diluting the stock solutions in a physiological saline buffered with HEPES (N- [2-hydroxyethyl] piperazin-N '- [2-ethanesulfonic acid]) (composition in mM): NaCl, 137; KCl, 4.0; CaCl2, 1.8; MgCl2, 1; HEPES, 10; Glucose, 10; pH adjusted to 7.4 with NaOH (prepared weekly and refrigerated before use). As previous results have shown that 0.3% DMSO does not affect channel current, all test and control solutions will contain 0.1% DMSO. If the final DMSO concentration must be greater than 0.3%, to achieve a concentration of the specified test item, a separate test with a control vehicle with n&gt; 2 was performed at the highest final DMSO concentration. Test and control solutions were prepared daily from stock solutions.

The cells used were human embryonic epithelial kidney cells (HEK293; strain of origin, American Type Culture Collection, Manassas, VA; sub-strain, ChanTest, Cleveland, OH) transformed with adenovirus 5 DNA and transfected with hERG cDNA. Stable transfectants were selected by coexpression with the G418 resistance gene incorporated into the expression plasmid. The selection pressure was maintained including G418 in the culture medium. The cells were cultured in Dulbecco-modified Eagle's medium / nutrient mixture F-12 (D-MEM / F-12) supplemented with 10% fetal bovine serum, 100 U / ml of penicillin G sodium, 100Ig / ml of Streptomycin sulfate and 500 Ig / ml of G418.

Data acquisition and analysis were performed using a pCLAMP software package (Axon Instruments,

5 AC). The stable state was a constant increase restrictive over time (linear dependence on time) before and after the application of the test article. The decrease in the amplitude of the current was used after reaching the stable state to calculate the percentage of blocking with respect to the control.

All experiments were performed at room temperature (18ºC – 24ºC). Each cell acted as its own control. A concentration (10μM) of each test article was applied to the cells expressing hERG (n&gt; 3, where n = the number of cells). The duration of exposure to each concentration was limited to the time needed to reach the stable state block, but not more than 10 minutes. A concentration of the positive control article (60nM terfenadine) was applied to two cells (n&gt; 2). The cells were transferred to the recording chamber and superfused with HB-PS solution. The pipette solution for whole cell records were (composition in mM): potassium aspartate, 130; MgCl2.5; EGTA (ethylene glycol tetraacetate), 5; ATP (adenosine triphosphate), 4; HEPES, 10; pH

fifteen adjusted to 7.2 with KOH. The pipette solution was prepared in batches, aliquots were taken, stored in a freezer and a fresh aliquot was thawed every day. Patch pipettes were made with glass capillary tubes using a P-97 micropipette extractor (Sutter Instruments, CA). A commercial patch clamp amplifier was used for whole cell records. Before scanning, the current registers were passed through low-pass filters to a fifth of the sampling frequency.

The initial and steady state blockage of the hERG current due to the test article was measured using a pulse pattern with fixed amplitudes (depolarization: +20 mV for 2 s; repolarization: –50 mV for 2 s) repeated at intervals of 10 s, from a clamping potential of –80 mV. The maximum tail current was measured during the 2 second stage up to –50 mV. A stable state was maintained for at least 30 seconds before applying the test article or positive control. Maximum tail currents were measured until a

25 New stable state.

Table 7 shows the% blockage of the hERG channel at the concentrations indicated for various sleep inducing agents revealed. Commonly, values ​​of about 10% or less are considered desirable, values ​​of about 12% to about 30% may be acceptable if the compound has a good sleep-inducing behavior and does not produce other side effects; and values ​​greater than approximately 30% are considered undesirable.

Table 7: hERG Lock

Example 5: Specificity for histamine H1 receptors

Binding assays were performed using compound 1 sleep inducing agent revealed in binding assays.

35 Competitive with known patterns for the histamine H1 receptor and M1, M2 and M3 muscarinic receptors, alpha 1 and alpha 2 receptors, and D1 and D2 receptors.

Histamine H1 analyzes are described in Chang, et al., "Heterogeneity of Histamine H1-Receptors: Species Variation in [3H] Mepyramine Binding of Brain Membranes". Journal of Neurochemistry. 32: 1653-1663 (1979); Martinez– Mir, MI, Pollard, H., Moreau, J., et al. "Three Histamine Receptors (H1, H2, and H3) Visualized in the Brain of Human and Non-Human Primates." Brain Res. 526: 322–327 (1990); Haaksma, EEJ, Leurs, R. and Timmerman, H. "Histamine Receptors: Subclasses and Specific Ligands." Pharmac. Ther. 47: 73-104 (1990). Muscarinic analyzes are described in Buckley, NJ, Bonner, TI, Buckley, CM, and Brann, MR “Antagonist Binding Properties of Five Cloned Muscarinic Receptors Expressed in CHO – K1 Cells”. Mol. Pharmacol 35: 469-476 (1989). The analyzes were performed according to the previous articles with the following modifications. Chemical reagents that

Four. Five used in what is described below were obtained in Sigma, St. Louis, MO.

For histamine H1 analyzes, the receptors were obtained from bovine cerebellar membrane tissue with a Bmax (number of receptors) of 6.2 femtomol / mg of tissue (wet weight) and a KD (binding affinity) of 1 , 3nM. A radioligand ([3 H] pyrilamine (15-25) Ci / mmol), 1.9nM Ki, final concentration of 2.0nM), and 10 IM of triprolidine (3.3nM Ki) was used as a nonspecific determinant, reference compound and positive control. The receptor and radioactive ligand were combined with the test compound at a range of concentrations of test compound from about 10–10 to about 10-M, and the sample was incubated in 50 mM Na-KPO4 (pH 7.5) at 25 ° C for 60 minutes. The reaction was terminated with a rapid vacuum filtration on fiberglass filters. The radioactivity of the displaced radioactive ligand trapped on the filters was determined and compared with the control values ​​to measure any interaction of the test compound with the histamine H1 binding site.

55 For muscarinic assays, the receptors were obtained from human recombinant receptors expressed in CHO cells (PerkinElmer, Inc., Wellesley, MA). The radioactive ligand used was [3 H] -espopolamine, N-methyl chloride (80–100 Ci / mmol). (-) - Methylcopolamine bromide (1.0μM) was used as a nonspecific determinant, reference compound and positive control. After incubation, the reactions were terminated with rapid vacuum filtration on fiberglass filters. The radioactivity of the displaced radioactive ligand trapped on the filters was determined and compared with the control values ​​to measure any interaction of the test compound with the respective one

5 receiver.

For the test with the M1 receptor, the Bmax (number of receptors) was 4.2 picomol / mg protein and the KD (binding affinity) of the receptor was 0.05nM. The radioactive ligand was used at a final concentration of 0.5nM, while the (-) - methylcopolamide bromide had a Ki of 0.09nM. The receptor and radioactive ligand were combined with the test compound at a concentration range of the test compound of approximately 10–12 to

10 approximately 10-M, they were incubated in Dulbecco phosphate buffered saline (PBS) for 60 minutes at 25 ° C and processed as described above.

For the test with the M2 receptor, the Cmax (number of receptors) was 2.1 picomol / mg protein and the KD (binding affinity) of the receptor was 0.29nM. The radioactive ligand was used at a final concentration of 0.5nM, while (-) - methylcopolamide bromide had a Ki of 0.3nM. The receptor and the radioactive ligand were combined with the

fifteen Test compound at a concentration range of the test compound from about 10-12 to about 10-5M, were incubated in Dulbecco phosphate buffered saline (PBS) for 60 minutes at 25 ° C and processed as described above.

For the test with the M3 receptor, the Bmax (number of receptors) was 4.0 picomol / mg protein and the KD (binding affinity) of the receptor was 0.14nM. The radioactive ligand was used at a final concentration of 0.2nM, while

twenty (-) - Methylcopolamide bromide had a Ki of 0.3nM. The receptor and radioactive ligand were combined with the test compound at a range of concentrations of test compound from about 10-12 to about 10-M, incubated in 50mM Tris-HCl (pH 7.4) containing 10mM MgCl2 , 1mM EDTA for 60 minutes at 25 ° C, and were processed as described above.

The purinergic A1 binding assay, adenosine, was performed according to published procedures. See, e.g., Bruns,

25 et al., Naunyn Schmiedebergs Arch. Pharmacol., 335 (1): 59–63 (1987), with minimal modifications; and Ferlany, et al. Drug Dev. Res. 9: 85–93 (1986).

The purinergic A2 binding assay, adenosine, was performed according to published procedures. See, eg, Jarvis, et al., J. Phannacol. Exper. Ther. 251 (3): 888–93 (1989) with modifications; and Bruns, et al., Mol. Pharmacol 29 (4): 331–46 (1986) with modifications.

30 The D1 (recombinant human) binding assay, dopamine, was performed according to published procedures. See, eg: Jarvie, et al. J. Recept Res., 13 (1–4): 573–90 (1993); and Billard, et al. Life Sciences, 35 (18): 1885–93 (1984), with modifications.

The D1 (recombinant human) binding assay, dopamine, was performed according to published procedures. See, eg: Jarvie, et al. J. Recept Res., 13 (1–4): 573–90 (1993); and Gundlach, et al. Life Sciences, 35 (19): 1981–8 (1984), with

35 modifications.

Binding to H1 may indicate the desired sleep inducing activity of the compound. Binding to muscarinic receptors shows nonspecific binding and may indicate anticholinergic activity that may result in unwanted side effects, e.g. eg, the side effects of many known antihistamines, eg, blurred vision, dry mouth, constipation, urinary problems, dizziness, anxiety and the like. A decrease in the union of

40 Compounds with the M1-M3 receptors compared to the binding of the compound with the H1 receptor indicate a greater specificity of the compound for the histamine receptor versus the muscarinic receptor. In addition, a drug with greater specificity for the histamine receptor would have fewer anticholinergic side effects.

Table 8 shows the Ki inhibition constant in nM for H1 and muscarinic receptors. It can be seen that the revealed compound is very specific to H1 against muscarinic receptors. So, hopefully the

Four. Five Revealed compound exhibits good sleep-inducing activity with limited side effects associated with muscarinic receptor inhibition.

Table 8: Specificity for histamine H1 receptors

<dl><dt>Comp No. </dt><dd>H1 (bovine) M1 M2 M3 Alpha 1 Alpha 2 D1 D2 </dd></dl>

<dl><dt>1 </dt><dd>40.31 &gt; 10,000 &gt; 10,000 &gt; 10,000 &gt; 10,000 &gt; 10,000 &gt; 10,000 rat and human being &gt; 10,000 rat and human being </dd></dl>

<dl><dt>1 11.5 in rat, 23.7 in human </dt><dd /></dl>

Example 6: Evaluation of loxapine analogues

The following pharmacokinetic parameters are calculated from the individual plasma concentrations of the modified antihistamine compound using a compartment-free approach and an appropriate validated pharmacokinetic computer program (eg, WinNonlin Professional). The concentration values ​​published as BLQ are set to zero. If concentration data is available, if possible, intermediate calculations (non QC.d data) are made between periods. The dose increase does not depend on pharmacokinetic stones.

For each pharmacokinetic parameter per dose group, descriptive statistical data are calculated, including the mean, standard deviation, coefficient of variation, geometric mean, median, minimum and maximum. For each dose level, descriptive statistical data for ABC (0 – t), ABC (0 – inf) and Cmax transformed by natural logarithms are provided. In addition, the average and median concentration versus the time graphs are provided.

The proportionality of the dose after the study medication is examined by analyzing the pharmacokinetic variables ABC (0 – t), ABC (0 – inf) and Cmax transformed by natural logarithms with a linear model that includes the dose transformed by natural logarithms as it would cova It is concluded that there is proportionality of the dose if the 95% confidence interval for the covariant slope includes the value 1. The linearity of the dose for ABC (O – t), ABC (O – inf) and Cmax is also examined using a linear model. See, eg: Gibaldi and Perrier, Pharmacokinetics, II Ed., Marcel Dekker: New York, New York (1982). In the calculations, nominal sample collection times were used, except when the actual sampling times were not included in the acceptable time intervals specified by protocol. The following parameters were estimated:

<dl><dt>Cma x</dt><dd>Plasma concentration </dd></dl>

<dl><dt>Tmax </dt><dd>Time to reach the maximum concentration </dd></dl>

<dl><dt>Cmax and Tmax </dt><dd>they were presented directly from the concentration – time data. </dd></dl>

<dl><dt>ABC0 – t </dt><dd>Surface under the plasma – time concentration curve from hour 9 to the last time point with measurable concentrations, estimated by the linear trapezoid law. </dd></dl>

<dl><dt>ABC0–00 </dt><dd>Surface under the plasma concentration curve - time extrapolated to infinity, calculated using the following formula </dd></dl>

<dl><dt>ABC0–00 = ABC0–1 + C0 / A0 </dt><dd /></dl>

<dl><dt>Where Ct is the last measurable concentration in plasma and Az is the estimated terminal phase elimination rate using a log – linear regression during the terminal phase of elimination. The number of points used in the calculation of Az was determined by ocular inspection of the data describing the terminal phase. At least the last three time points with measurable values ​​were used in the calculation of Az. The number of points used in the calculation of Az is based on the best correlation (adjusted with respect to r2) obtained for the time points that describe the phase of terminal elimination. It is considered that an adjusted value with respect to r2 for the regression line accurately defines the phase of terminal elimination if the value is&gt; 0.7.</dt><dd /></dl>

<dl><dt>T1 / 2 </dt><dd>Elimination half-life determined by In (2) Az. </dd></dl>

<dl><dt>CL </dt><dd>Systemic clearance; for intravenous infusion or infusion, calculated using the formula:</dd></dl>

<dl><dt>CL = Dose / ABC0–00 </dt><dd /></dl>

<dl><dt>It is given as CL / F, in which F = absolute bioavailability for all other routes of administration. </dt><dd /></dl>

<dl><dt>V2 </dt><dd>Volume of distribution for all routes of administration calculated using the following formula: </dd></dl>

<dl><dt>Vz = CL A2 </dt><dd /></dl>

<dl><dt>CL / F is used to calculate V2 / F for extravascular administration routes. </dt><dd /></dl>

Pharmacokinetic analysis is performed using the professional edition of the WinNonlin program (Pharsight Corporation, Version 3.3 or 4.1). Descriptive statistical data such as the mean and standard deviation are calculated in Microsoft Excel (version 8.0e).

The metabolism of the test articles in cryopreserved hepatocytes of monkey and human was analyzed as follows:

MATERIALS

<dl><dt>Materials: </dt><dd>Manufacturer, lot number and expiration date </dd></dl>

<dl><dt>Cellzdirect Hepatocytes </dt><dd>Monkey </dd></dl>

<dl><dt>Human being </dt><dd /></dl>

<dl><dt>Williams Medium E </dt><dd>Sigma W1878, expiration: 11–2004 </dd></dl>

<dl><dt>Fetal Bovine Serum </dt><dd>Fisher BW 14–501F, lot 01104637, expiration February 17, 2010. </dd></dl>

<dl><dt>Trypan Blue 0.45 </dt><dd>Biowhittaker 17–942E, lot 01104637, expiration January 14. </dd></dl>

<dl><dt>Stock solution of test material </dt><dd>CB – 1 / III / 6 </dd></dl>

<dl><dt>DMSO </dt><dd>Fisher BP231–100, lot 041215, expiration July 12, 2009 </dd></dl>

<dl><dt>10mM ethoxycoumarin in methanol </dt><dd>PSLB 22 – A – 15, expiration 25–09–04 </dd></dl>

<dl><dt>ACN </dt><dd>Fisher A998–4, lot 041181, expiration 07/6 </dd></dl>

<dl><dt>formic acid </dt><dd>Fisher 032879, expiration 14–03–06 </dd></dl>

5 Pre-incubation preparation:

The sample is diluted with DMSO to prepare 100IM and 10IM stock solutions. 0.1% formic acid in acetonitrile is prepared by adding 1 ml of formic acid per liter of acetonitrile (stored at RT for 3 months). 96-well plates of sudden cooling are prepared at 10 minutes, 60 and 120 minutes with 150 L of acetonitrile + 0.1% formic acid in each well. They are stored on ice or refrigerated.

10 The hepatocytes are then thawed and 100 L of cell suspension is placed in a microcentrifuge tube with 100 L of 0.4% trypan blue solution and gently shaken by inversion. A small amount of stained cell suspension (approximately 15 Il) is placed in a clean hemacytometer with a lid. The hemacytometer is placed on the microscope stage, and the focus and power are adjusted until a single counting square fills the field. The number of cells in the four squares subdivided into

fifteen outer corners of the hemacytometer. Viable cells are opalescent, round and pale with a darker contour. Non-viable cells are dark and dull blue.

The% viability is calculated as the number of viable cells divided by the total cells x 100.

The density of viable cells and the total number of viable cells are calculated:

Density of viable cells (D) = Average of 3 viable cell counts (C) x 104xf2;

twenty Total number of viable cells (E) = D x 26 (resuspension volume).

The additional medium necessary to reach a concentration of 1 x 106 cells / ml was calculated:

Volume of additional medium = total viable cells (E) –26 ml 1 x 106

The cells are diluted in that way and stored at room temperature.

25 Incubations

198 Il of hepatocytes are transferred to the relevant wells of a dosage plate. The remaining hepatocyte suspension is combined, and placed in a suitable boiling water container and left for 5 minutes to deactivate the cells (for deactivated controls and preparation of the standard curve).

198 Il of deactivated hepatocytes are transferred to control wells and 198 Il of empty media are transferred to the

30 control wells with buffer. Plates are pre-incubated for at least 15 min. The reactions are initiated with 2 L of appropriate dilution of test compound from the dosage plate. The plates are incubated in an incubator at 37 ° C for approximately 10 minutes, then 50 Il of incubation are removed to a 10 minute cooling plate containing 150 Il of acetonitrile + 0.1% formic acid and stored with refrigeration or over ice. After 60 minutes, 50 μl of incubation is removed to a sudden cooling plate in 60 minutes containing 150 μl of

35 Acetonitrile + 0.1% formic acid and stored with refrigeration or on ice. After 120 minutes, 50 µl of incubation is removed to a 120 minute sudden cooling plate containing 150 µl of acetonitrile + 0.1% formic acid and stored with refrigeration or on ice. The remaining 50 Il are frozen in incubation plates. The tubes are then centrifuged at –4 ° C at –1,400 xg for –10 minutes. 100 Il of supernatant is diluted with 100 Il of water in analysis plates, the plates are stored with refrigeration at –20 ° C before analysis.

Preparation of standard curves

0.1IM standard is prepared by adding 2L of 10IM dosing solutions to 198L of hepatocytes

5 deactivated in a standard preparation plate. 150 Il of acetonitrile + 0.1% formic acid are added to the standard sudden cooling plate. 150 Il of the 0.1IM standard is transferred to a column of a standard plate. 75 Il of deactivated hepatocytes are added to the remaining wells. 75 Il of 0.1IM standard is transferred to the adjacent well of the plate column and the well is mixed by titration. Serial dilution is continued. 75 Il are removed from the final standard (when the wells contain 75 Il). The plates are incubated at approximately 37 ° C

10 for 10 minutes. 50 Il are transferred to the standard sudden cooling plate containing 150 Il of acetonitrile

+ 0.1% formic acid. The plates are centrifuged together with samples and the supernatant diluted 1: 1 with water as explained above. Frozen samples are stored at ~ 20 ° C.

Example 7: Clinical evaluation of loxapine analogues

The objective of a clinical trial with humans is to collect information on the effects of

fifteen Loxapine derivatives. Such information includes, for example, clinical signs and symptoms from a physical examination, adverse events, laboratory safety (eg, hematology, clinical serum chemistry, urine tests), vital signs (eg, pressure blood, heart rate, temperature, respiratory rate) and electrocardiogram (ECG) data.

Clinical trials are performed as follows:

twenty I. Choice of the subject

A minimum of 18 subjects are used (2 registered groups of 9 subjects each). Candidates who meet the following inclusion criteria will be part of the study:

• Healthy adult male subjects aged 18–45 years.

• Subjects weighing at least 60 kg and having a difference of 15% from their ideal weight (see 25 Table of desirable adult weights of Metropolitan Life Insurance Company, 1983).

• Healthy subjects with clinically irrelevant trace results (eg, laboratory profiles, medical records, ECG, physical examination).

Candidates who meet one of the following exclusion criteria will not be part of the study:

• History or presence of cardiovascular, pulmonary, hepatic, renal, hematologic, gastrointestinal, endocrine, immunological, dermatological, neurological or psychiatric disease.

<dl><dt>• </dt><dd>History or presence of sleep disorder. </dd></dl>

<dl><dt>• </dt><dd>History of chronic or seasonal allergies that require treatment with H1 receptor antagonists (ie, terfenadine, astemizole) in the 90 days prior to the study. </dd></dl>

<dl><dt>• </dt><dd>History or presence of alcoholism or drug addiction in the last 2 years. </dd></dl>

35 • Tobacco or nicotine consumption in the 90 days prior to the study.

• Hypersensitivity or idiosyncratic reaction known to the study drug, possible excipients of the study formulation (Captisol®; sodium saccharin, FCC; glycerin, USP; orange aroma, methylcellulose 400 cps, USP; purified water) or related compounds.

• Donation (standard or larger donation amount) of blood or blood products in the 90 days prior to the study.

<dl><dt>• </dt><dd>Participation in another clinical trial in the 90 days prior to the study. </dd></dl>

<dl><dt>• </dt><dd>History or presence of any disease, medical condition or surgery, which may have an effect on the absorption, metabolism, distribution or excretion of the drug. </dd></dl>

<dl><dt>• </dt><dd>Weight loss or gain ( 10%) in the 30 days prior to the study. </dd></dl>

Four. Five • Regular consumption of (eg, more days than you consume than you do not consume) excessive amounts of caffeine-containing drinks (eg, more than 5 cups of coffee or equivalent per day) in the previous 30 days to study

<dl><dt>• </dt><dd>Any condition that, in the opinion of the researcher or sponsor, makes the subject not suitable for the study. </dd></dl>

<dl><dt>• </dt><dd>Use of any previous or concomitant medication prohibited. </dd></dl>

Each of the subjects that completes the study selection evaluations, meets all the selection criteria and is accepted for the study is assigned a unique identification number and receives designated doses of modified antihistamine or placebo according to a randomized scheme. The random scheme is only accessible to the pharmaceutical personnel preparing the drug (which does not participate in the administration of the drug) and is not accessible to the subjects, analysts or staff members responsible for monitoring and evaluating adverse events.

The principal investigator may expel study subjects for the following reasons:

• secondary occurrence of a main exclusion criterion;

<dl><dt>• </dt><dd>to protect your health; </dd></dl>

<dl><dt>• </dt><dd> adverse events; </dd></dl>

<dl><dt>• </dt><dd>difficulties in collecting blood; </dd></dl>

<dl><dt>• </dt><dd>to protect the integrity of the study; </dd></dl>

<dl><dt>• </dt><dd>for violation of the protocol; and</dd></dl>

<dl><dt>• </dt><dd>for breach of study instructions. </dd></dl>

The clinical report includes the reasons for the expulsion of the subject, as well as the data related to said expulsion. Subjects withdrawn from the trial before the end of the study will pass all the procedures scheduled for the completion of the study. The investigator or a supervising physician will evaluate the retired subjects for any adverse event (either serious or not serious) or for clinically relevant abnormal laboratory test values, and will be treated and / or monitored until symptoms or values return to normal or acceptable levels, according to the opinion of the researcher.

II. Study Restrictions

Subjects do not take medication that requires or does not require a prescription (including herbal products) during the 7 days preceding the study until the last sample of the final pharmacokinetic sample collection period has been collected. In addition, the consumption of food and beverages containing the following substances is prohibited as indicated:

<dl><dt>• </dt><dd>Ethylxanthine: 72 hours before each dose and during the sample collection period, ie, caffeine drinks and equivalents are prohibited (eg, chocolate bars). </dd></dl>

<dl><dt>• </dt><dd>Alcohol: 72 hours before each dose and during the sample collection period. </dd></dl>

All medications taken during the 30 days before the start of the study are recorded. Any medication taken by chronic or seasonal allergies is recorded in the 90 days prior to the study.

Selection of the subject prior to the study: for the tracking, an informed consent form is administered. In the 14 days prior to dosing, medical history and demographic data are recorded, including name, sex, age, race, body weight (kg), height (cm), alcohol consumption and tobacco. Each subject undergoes a physical examination that includes vital signs, 12-lead ECG and laboratory analysis as specified. Laboratory tests include the following:

a) Hematology, which includes hemoglobin, MCV, red blood cell count, hematocrit, MCHC, white blood cell count with differential platelet count and MCH;

b) Serum chemistry, which includes albumin, ALT (SGOT), creatinine, alkaline phosphatase, glucose, total bilirubin, creatine phosphokinase (CPK), sodium, uric acid, AST (SGOT) and triglycerides;

c) Urinalysis, which includes appearance and color, glucose, nitrite, pH, ketones, urobilinogen, specific gravity, bilirubin, leukocytes, protein and blood;

d) Other tests, including HIV, urine drug screening, HbsAg, cannabinoids, HCV, benzodiazepines, HCV, amphetamines, hepatitis A (1gM), opiates, alcohol, cocaine and continin.

Treatment of the subjects: the subjects are locked at least 36 hours before dosing until the events of the 24 hours after the dose end. They will return for a follow-up visit one week after the last dose or after early withdrawal.

The subjects remain semi-lying in bed for the first 4 hours after drug administration. However, if adverse events occur at any time, the subjects are placed in an appropriate position or allowed to lie on their right side. The subjects do not perform any strenuous physical activity at any time during the confinement period.

Standard meals are provided on day 1 and day 2. On day 1, subjects must remain fasting for a minimum of 10 hours overnight before receiving the dose and for at least 4 hours later. However, if the option of a previous dose is used in the feeding state in period 3 of Group 2, a standard fat-rich meal is administered 30 minutes before the dose. In this case, the high-fat breakfast (ie, approximately 50% of the calories come from fat) consists of two eggs fried in butter, two strips of bacon, two toasts with butter, approx. 120 g of fries and approx. 235 ml of whole milk. During confinement, foods and beverages that contain caffeine or equivalent (e.g., chocolate bars) are prohibited.

It is not allowed to drink water from the previous 2 hours until after 2 hours of the dose. It is allowed to drink water the rest of the hours. Standard meals are provided at approximately 4 and 9 hours after dosing and at the appropriate subsequent hours.

III. Drug Administration

Subjects receive the dose of each period as assigned according to the random dose sequence schedule for each dose group (enrollment). The subjects receive the assigned dose in a glass measuring cup and, within each dose group, all active and placebo doses are administered at the same volume to maintain double blindness. Subjects are instructed to swallow the dose.

A total of 240 ml of water is administered with the dose. A designated portion of water (assigned by the pharmacist based on the dosing volume) is added to the drained dosing cup, moved to rinse and swallowed. This procedure is repeated twice and then the subject drinks the rest of the water.

The initial dose for the first dose level in humans is based on the toxicity and safety profiles of preclinical studies. The conversion of the equivalent body surface of human being into rat is 1/6 ("Toxicological Handbook", Michael J. Dereleko, CRC press, Boca Raton, FL). Based on NOAEL of 30 mg / kg / day for rat and the body surface equivalence criteria, the equivalent dose in a 60 kg individual is 300 mg / day (1/6 x 30 mg / kg / day [NOAEL in rat] x 60 kg). Based on the dose of NOAEL in rat (30 mg / kg / day), the dose of 3 mg is approximately 1/10 of the dose of NOAEL in rats. The proposed highest dose of 160 mg is also below NOAEL in rats.

If a dose-limiting toxicity is observed (grade 3 or 4 according to the modified grade scale according to the WHO common toxicity criteria, Annex I) in relation to the study medication in any 2 of the 6 subjects at any level of doses, dose increases are stopped and the previous dose is considered the maximum tolerated dose (MDT).

If a subject at any dose level has a dose-limiting toxicity, the principal investigator (in conference with the sponsor) decides, with good clinical judgment, whether to continue to the next planned dose level or adjust the next dose level down with respect to the planned dose. This consultation is done for all the groups that go after the previous dose group to decide whether to continue with the planned doses or reduce the doses. In addition, it is possible to replace the planned doses with intermediate doses if obvious safety or tolerability problems (ie, there does not have to be a fact of grade 3 or 4) from the previous dose that suggest the need for further increases. slowly.

Dosage increases are only allowed if, in the opinion of the principal investigator, adequate safety and tolerability has been demonstrated in the previous lower dose. In all cases, the principal investigator relies on good clinical judgment to decide whether to adjust the dose or stop the study based on the evaluation of all relevant safety factors of the subjects.

The principal investigator reviews the entry record data (e.g., those from physical examination, vital signs, questionnaire and clinical laboratory results (e.g., serum chemistry, hematology, analysis of urine and urine drug screening)) in search of clinically relevant changes since the screening or the previous period. The principal investigator determines whether the subject will receive the dose or be withdrawn from the study based on this review.

IV. Clinical observation

A hematology panel, a serum chemistry panel and a urine analysis are performed at the screening, at each entry register, 24 hours after each dose and one week after the final dose, or after an early withdrawal. Blood samples (approximately 7 ml) are collected with a permanent intravenous catheter that is emptied into glass tubes containing a predose of sodium heparin and at 0.25, 0.5, 0.75, 1.0, 1 , 5, 2, 3, 4, 6, 8, 10, 12, 18, and 24 hours after the dose. Urine samples are collected before the dose and during the 0–8 hour interval of each period. The samples collected during the interval are not mixed. Each urination is considered a sample. Urination is performed at discretion, not scheduled (except for pre-dose urination and urination at the end of the 8 hour interval).

Vital signs are measured during traces. When the moment of vital signs only coincides with an ECG, the vital signs are taken 10 minutes before the ECG. When the timing of the vital signs coincides with a blood draw or with a blood draw and an ECG, the vital signs are taken 10 minutes before the blood draw. Breathing and temperature are monitored at reception, 24 hours after each dose and one week after the final dose or after an early withdrawal. Individual blood pressure and heart rate measurements are taken after a minimum of 5 minutes in a semi-reclined position. The measures taken during the confinement of the study are monitored with an AVS machine at the reception; 0 (predose); at 0.25, 0.5, 0.75, 1, 1.5, 2, 3, 4, 6, 8, 10, 12, 18 and 24 hours after the dose; and one week after the final dose or after early withdrawal. For any heart rate measurement greater than 100 beats per minute, the heart rate will be rechecked two minutes later. On day 1, approximately 24 hours before dosing, 3 blood pressure and heart rate measurements are taken, 2 minutes apart, as described above.

An ECG of 12 standard leads is performed for each subject undergoing screening on day 1 at the hours that coincide with the hours of day 1 of 1 hour before the dose and at 1, 1,5, 2, 3, 4 and 6 hours after the dose; on day 1, 1 hour before the dose and at 1, 1.5, 2, 3, 4, 6 and 24 hours after the dose; and one week after the last dose or after early withdrawal. More ECGs may be performed at other times if deemed necessary. All standard 12 lead ECGs are recorded for 10 seconds. The timing of the completion and the ECG registration technique is normalized for all subjects. Subjects must be lying down for at least 1 minute before each 12-lead ECG evaluation. The principal investigator evaluates the PR, QRS, QT and QTc intervals. When the timing of the ECG coincides with a blood draw, the ECG will be taken after the extraction.

A doctor examines each subject of the screening, upon registration, 24 hours after each dose and one week after the final dose or after early withdrawal. More tests may be done at other times if deemed necessary.

Immediately before the measurement of the vital signs 1 hour before the dose and at 1, 2, 6 and 24 hours after the dose (the vital signs are taken 10 minutes before the blood draw at this time), it is presented The subjects have a similar visual scale and are asked to mark on a vertical line of 100 mm the point that best describes their level of alert at that time, between very sleepy and alert / very awake.

Subjects are informed to inform the doctor or study staff of any adverse events or illnesses experienced during the trial. In addition, a specific survey is conducted regarding adverse events before dosing, at 2, 4, 8 and 24 hours after the dose and one week after the last dose, or after an early withdrawal. The questions are posed in a non-specific way so as not to influence the answer.

The investigator or a supervising physician evaluates any subject who has suffered any adverse event (very serious

<dl><dt /><dd>or not serious) or from which clinically relevant abnormal laboratory test values ​​have been obtained, and are treated and / or monitored until the symptoms or values ​​return to normal or acceptable levels, according to the opinion of the researcher. Doctors, either on site or in the emergency department of a nearby hospital, administer the treatment in case of any serious adverse event. When appropriate, medical tests and examinations are performed to document the resolution of the event (s). The results are classified, for example, as resolved, improved, unaltered, worsened, fatal or unknown (lack of follow-up).</dd></dl>

<dl><dt>V.</dt><dd> Preparation of reports </dd></dl>

All adverse events that occur during the clinical trial are recorded. Adverse events are coded with MedDRA (version 4.1). An adverse event / experience (HA) is any unjustified medical occurrence in a patient or subject undergoing a clinical investigation to which a pharmaceutical product has been administered that does not necessarily have a causal relationship with this treatment (ICH / WHO). Therefore, an adverse event (HA) is any unfavorable or unintended sign (including, for example, an abnormal laboratory finding), symptom

or disease temporarily associated with the use of a medical product, regardless of whether it is considered or not related to the medical product (ICH / WHO).

The investigator reviews each fact and evaluates its relationship with drug treatment (ie, unrelated, unlikely to be related, possibly related, probably related, related to almost total safety). Each sign or symptom is scored on a 3-point severity scale (mild, moderate or severe), and the date and time of the occurrence, temporal relationship with the dosage of the drug, duration and outcome of each event are noted. The following definitions are used to classify severity: (1) Mild: the adverse event is easily tolerated and does not interfere with daily activity; (2) Moderate: the adverse event interferes with daily activity, but the subject can still continue; (3) Serious: the adverse event incapacitates the subject and requires a medical intervention.

If any of the above adverse events is serious, special procedures are followed. All serious adverse events are reported to the sponsor at 24 hours, information that is followed by reports at 48 hours regardless of whether the serious facts are considered related to the drug or not.

A serious adverse event (HAG) is any harmful medical event that, at any dose, causes death, poses a threat to life, causes a disability or permanent incapacitation, requires hospitalization of the patient, prolongs hospitalization of the patient, is a congenital anomaly, may endanger the subject or may require intervention to avoid one or more of the other results listed above.

SAW. Pharmacokinetics

The following pharmacokinetic parameters are calculated from the individual plasma concentrations of the modified antihistamine compound using a compartment-free approach and an appropriate validated pharmacokinetic computer program (eg, WinNonlin Professional). The concentration values ​​published as BLQ are set to zero. If concentration data is available, if possible, intermediate calculations (non QC.d data) are made between periods. The dose increase does not depend on pharmacokinetic stones.

For each pharmacokinetic parameter per dose group, descriptive statistical data are calculated, including the mean, standard deviation, coefficient of variation, geometric mean, median, minimum and maximum. For each dose level, descriptive statistical data for ABC (0 – t), ABC (0 – inf) and Cmax transformed by natural logarithms are provided. In addition, the average and median concentration versus the time graphs are provided.

The proportionality of the dose after the study medication is examined by analyzing the pharmacokinetic variables ABC (0 – t), ABC (0 – inf) and Cmax transformed by natural logarithms with a linear model that includes the dose transformed by natural logarithms as it would cova It is concluded that there is proportionality of the dose if the 95% confidence interval for the covariant slope includes the value 1. The linearity of the dose for ABC (0 – t), ABC (0 – inf) and Cmax is also examined using a linear model.

VII. Safety assessment

A list of adverse events data arising as a result of treatment per patient is provided, including the literal term, the preferred term, the treatment, the severity and the relationship with the treatment.

A summary is made of the number of subjects experiencing adverse events and the number of adverse events per dose level using frequency counts.

Safety data, including laboratory evaluations and vital signs evaluations, by dose level and time point of collection are summarized. Descriptive statistical data is calculated for quantitative safety data and frequency counts are collected to classify quantitative safety data. In addition, an average change of the reference values ​​table for vital signs and a change table describing the changes of normal intervals for clinical laboratory results are provided.

ECG results are classified as normal and abnormal, and are summarized using frequency counts by dose group and time point of collection. Descriptive statistical data for the PR, QRS, QT and QTc intervals are calculated.

Changes in physical examinations are described in the text of the final report.

Heart rate data is summarized by treatment group and time point using descriptive statistical data, as individual changes occur with respect to the reference values. The results of the mean change with respect to the reference are used to compare the active dose groups with the placebo at each time point. Data from six subjects completed per dose level would provide 80% accuracy in detecting a difference of 20 beats per minute. After each period, an intermediate analysis is completed.

VIII. Effectiveness evaluation

VAS sedation scores are summarized by time point of collection of each dose level using descriptive statistical data.

Example 8: Preclinical evaluation of loxapine analogues

Before the clinical test of the compounds in humans, a preclinical test is performed. The preclinical evaluation includes the following tests:

i. Preclinical absorption, distribution, metabolism and excretion

The compound is administered to rats, dogs and macaques at a dose of approximately 3 mg / kg orally and intravenously. Plasma samples of all species are collected for pharmacokinetic analysis. Tmax and half-life (in hours) are measured in the rat, the dog and the macaque. The percentage of bound protein in rat and human plasma is also measured.

The brains of the rats are collected after oral administration to determine the brain levels of the precursor drug.

Cytochrome P450 inhibition is studied in vivo. In addition, the in vitro metabolism rate is determined in rat, dog, macaque and human hepatocyte cultures for each compound.

ii. Focus of cardiac effects

The main toxicological aspect studied during the clinical phase of candidate selection of the project is the prolongation of the QT interval. Historically, H1 antagonists have been associated with this effect. QT prolongation can rarely evolve into dangerous cardiac arrhythmias. To better analyze in vitro the likelihood of a compound causing a prolongation of QT, the hERG binding assay was chosen as a test system to study the potential of a compound to produce this effect. The human hERG channel, transfected to a stable cell line, is electrophysiologically studied and the percentage of channel current inhibition is published.

To determine if a compound can cause any change in the QT interval, the compound is studied in Beagle dogs telemetrically. Devices are implanted in dogs to continuously monitor ECG and arterial blood pressure. Dogs (groups of 4) are studied in a Latin square design, in which each dog receives 3 different doses and a placebo. Two studies are conducted with doses of 0.3, 1, 3, 10 and 30 mg / kg.

iii. Acute study in rats

The objective of this study is to evaluate the toxicity and the maximum tolerated dose (MDT) of the test articles administered orally through forced feeding to rats. Rats Crl: CD® (SD) IGS BR male (3 / group) are assigned to 5 groups. When starting the dosage, the animals are approximately 7 weeks old with body weights ranging from 172 to 206 g. Each group receives either 50, 100, 150, 200 or 250 mg / kg of the compound once a day for 5 days. All surviving animals are sacrificed on the 6th day.

The toxicity assessment is based on mortality, clinical observations and body weight data.

iv. Acute study in dogs

The objective of this study is to evaluate the toxicity and the maximum tolerated dose (MDT) of the compound administered at increasing doses orally by forced feeding to dogs. Two purebred male Beagle dogs are assigned to the study. When starting the dosage, the animals are at least 6 months old with body weights that vary from 8.0 to 10.9 kg. Dogs receive dose preparations containing the compound once a day for 5 days in increasing doses of 25, 50 or 75 mg / kg.

Dogs are observed at 0.25, 0.5, 0.75, 1.0 and 2.0 hours ± 5 minutes and at 4, 6, 8 and 24 hours ± 15 minutes after the dose. Days 1 and 6 are weighed.

Electrocardiograms are performed and blood pressures are taken before dosing and 1, 4 and 24 hours after the dose of 40 mg / kg on day 5.

Based on the interval and severity of the clinical signs observed, the MDT for the compound is calculated.

v. Study with 14-day rats with recovery study

The objective of this study is to evaluate the toxicity of the compound when it is administered orally to rats for at least 14 days and to evaluate the reversibility, persistence or delayed appearance of any effect after a recovery period of up to 14 days.

Rats Crl: CD® (SD) IGS BR male and female are assigned to seven groups, four main study groups and three groups for toxicokinetics. Each group receives dose preparations containing 0.25% 400 cps methylcellulose in 200mM acetate buffer, or 10, 30 or 150 mg of test article / kg body weight (mg / kg / day) at a dose volume 5 ml / kg

The toxicity assessment is based on mortality, clinical and ophthalmological observations, body weights, food consumption, clinical pathology, organ weights and macroscopic and microscopic findings. Blood samples are collected for toxicokinetic evaluation.

14-day study with dogs and recovery phase

Toxicity and toxicokinetics of a compound of the invention are determined when it is administered orally daily (Phase 1) or by capsules (Phase 2) to dogs for at least 14 days. It also evaluates the reversibility, persistence or delayed occurrence of observable effects after a recovery period of 7 days (Phase 1) or 14 days (Phase 2). The doses of 3, 10, 30 and 70 mg / kg / day are studied. All dogs in Phase 1 and Phase 2 survived until they were slaughtered as planned.

The compounds and the above protocols are useful in the pre-clinical evaluation of the loxapine compounds of the invention.

Example 9: Evaluation of analgesic activity

The analgesic activity of a loxapine analogue after oral administration is analyzed. Analgesic activity is evaluated by analysis of abdominal spasms in the rat and mouse. Analgesic activity is also evaluated using the tail clamp test in the mouse, the tail tail test in the rat, the Randall-Selitto test in the rat, and comparisons were made with the control vehicle group . Reference compounds of ASA (acetylsalicylic acid) and morphine are also included in the comparison.

The tail clamp and tail tail test provide useful information about the central analgesic activity of the test article. The Randall – Selitto test provides information on the ability of the compound to modify a hyperalgesic state, and the abdominal spasm test provides information on the peripheral analgesic activity of the test article. The test article is administered by forced oral feeding, this being the desired route of clinical administration. The dose levels used are expected to encompass the effective dose and provide an adequate safety margin.

Test article, reference compound and irritating formulation

All formulations are prepared every day of dosage. The test article is formulated in 0.25% MC (w / v) at the highest concentration required. The lower doses are obtained by serial dilution of the highest concentration, using 0.25% MC (w / v). The reference compound, acetylsalicylic acid, is formulated in 0.25% MC (w / v) at the required concentrations. Brewer's yeast is formulated in water for injection at the required concentration. Acetic acid is diluted with water for injection to provide the required administration concentration.

The dose levels are expressed in terms of the amount of the test compound / reference compound / irritant administered regardless of purity or active content.

Animals

An adequate number of Crl mice are acquired: male CD-I (ICR) BR and Wistar rats in Charles River (RU) Ltd., Margate, Kent. The mice are approximately 4 weeks old and weigh between 18 and 22 g upon arrival. Rats are approximately 5 weeks old and weigh between 150 and 170 g upon arrival. The age and weight of the animals at the beginning of the study are documented in the raw data and in the final report.

Animals are introduced into groups appropriate for the size of the cage used, in cages that comply with the code of practice of confinement and care of animals used in the Law of Scientific Procedures (Law of the Ministry of Interior on Scientific Procedures with Animals, 1986) . Aspen wood shavings for beds are renewed weekly in each cage (Dates and Ltd, Manchester, UK). The specific contaminants of the beds are analyzed and the results are recorded in a file in Covance. The cages are cleaned and dried before use. Aspen chewable blocks are placed in the cages as a form of environmental enrichment. Periodically, confinement rooms are maintained in acceptable limits of temperature and relative humidity (nominally 19 to 25 ° C and 40 to 70%, respectively). These rooms are illuminated with fluorescent light for 1.2 of the 24 hours of the cycle and are designed to receive at least 15 changes of pure air per hour.

RM1 diet (E) .SQC., (Special Diets Services Ltd., Witham, UK) and main tap water will be provided at will, unless otherwise specified below. Specific constituents are analyzed periodically and are not found to contain any biological or chemical entity that may interfere with the analysis system. The treatment groups used for the study are those shown in Table 9:

Table 9. Treatment groups

<dl><dt>Group </dt><dd>Treatment Dose level (mg / kg) Conc. (Mg / ml) No. of animals </dd></dl>

<dl><dt>1 2 3 4 5 </dt><dd>Loxapine analog vehicle Loxapine analog Loxapine analog Morphine - 3 10 30 100 - 0.3 1.0 3.0 10.0 8 8 8 8 8 </dd></dl>

Pressure measurements are taken from the left and right hind legs of each animal immediately before the administration of the vehicle, the test article or the reference compound and 30, 60, 120 and 240 5 minutes after oral administration . The order of the pressure measurements is that of the left leg followed by the right leg.

Rat abdominal spasm test

Each animal receives a single administration of vehicle, test article or reference compound by forced oral feeding, using a constant dose volume of 10 mg / kg. Individual dose volumes 10 are based on individual body weights obtained on the day of dosing. In Table 10, the treatment groups are shown.

Table 10. Treatment groups

<dl><dt>Group </dt><dd>Treatment Dose level (mg / kg) Conc. (Mg / ml) No. of animals </dd></dl>

<dl><dt>1 2 3 4 5 </dt><dd>Loxapine analogue vehicle Loxapine analogue Loxapine analogue ASA - 3 10 30 100 - 0.3 1.0 3.0 10.0 6 6 6 6 6 </dd></dl>

Forty-five minutes after oral administration, each animal receives an intraperitoneal injection of 1 ml of 1% acetic acid. The animals are immediately placed in individual observation chambers and the number of abdominal spasms caused in a subsequent period of 25 minutes is recorded.

Abdominal mouse spasm test

Each animal receives a single administration of vehicle, test article or reference compound by forced oral feeding, using a constant dose volume of 10 ml / kg. Individual dose volumes are based on the individual body weights obtained on the day of dosing. In Table 11, the treatment groups are shown.

Table 11. Treatment groups

<dl><dt>Group </dt><dd>Treatment Dose level (mg / kg) Conc. (Mg / ml) No. of animals </dd></dl>

<dl><dt>1 2 3 4 5 </dt><dd>Loxapine analogue vehicle Loxapine analogue Loxapine analogue ASA - 3 10 30 100 - 0.3 1.0 3.0 10.0 6 6 6 6 6 </dd></dl>

Forty-five minutes after oral administration, each animal receives an intraperitoneal injection of 0.25

25 ml of 0.5% acetic acid. The animals are immediately placed in individual observation chambers and the number of abdominal spasms caused in a subsequent period of 25 minutes is recorded.

Terminal procedures

At the end of each test, the animals will be sacrificed without causing them suffering through a Program 1 compound (eg, exposure to carbon dioxide gas at an increasing concentration followed by neck dislocation) and

30 will withdraw without performing the autopsy. If an animal shows any sign of serious discomfort during the study, it will be sacrificed immediately without causing suffering. Any animal found dead or who dies prematurely during the study will undergo a necropsy. A macroscopic examination is performed, after opening the thoracic and abdominal cavity by observing the appearance of the tissues in situ. Any anomaly is recorded.

EXAMPLE 10: Synthesis of compound 1

35 The synthesis of compound 1 is summarized in Scheme I.

Scheme I

Treatment of 10H – Dibenzo [b, f] [1,4] tricyclic oxazepin-11-one (3) with phosphorus oxychloride in the presence of N, N-dimethylaniline in amidine (5) with 2-carbomethoxy-2-methyl -Propionaldehyde gave alkylated piperazine (6), which was purified on silica gel. The basic hydrolysis of methyl ester of (5) in aqueous ethanol followed by acidification gave the carboxylic acid (7) corresponding to compound 1.