WO2014154925A1 - Neurogenic compounds comprising melatonin and the efficacy thereof in in vivo experiments for use in the treatment of diseases of the nervous system - Google Patents

Neurogenic compounds comprising melatonin and the efficacy thereof in in vivo experiments for use in the treatment of diseases of the nervous system Download PDF

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WO2014154925A1
WO2014154925A1 PCT/ES2014/070224 ES2014070224W WO2014154925A1 WO 2014154925 A1 WO2014154925 A1 WO 2014154925A1 ES 2014070224 W ES2014070224 W ES 2014070224W WO 2014154925 A1 WO2014154925 A1 WO 2014154925A1
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indol
methyl
methoxy
compound
formula
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PCT/ES2014/070224
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Spanish (es)
French (fr)
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María Isabel RODRIGUEZ FRANCO
Mario DE LA FUENTE REVENGA
Concepción PEREZ MARTIN
Ana María PEREZ CASTILLO
José Ángel MORALES GARCÍA
Sandra ALONSO GIL
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Consejo Superior De Investigaciones Científicas (Csic)
Universidad Autónoma de Madrid
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/4245—Oxadiazoles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/40—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil
    • A61K31/403—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil condensed with carbocyclic rings, e.g. carbazole
    • A61K31/404—Indoles, e.g. pindolol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/4196—1,2,4-Triazoles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/42—Oxazoles
    • A61K31/422—Oxazoles not condensed and containing further heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca

Definitions

  • the present invention which is included in the field of research and pharmaceutical industry, refers to new chemical entities derived from melatonin with neurogenic properties, modulators of melatonin and / or serotonin receptors, antioxidants and / or cholinergic. Experiments in mice have shown that they are capable of regenerating damaged neuronal populations in vivo, without causing the appearance of tumors.
  • This invention also relates to the processes for the preparation of these new compounds, the pharmaceutical compositions containing them and their use for the manufacture of a medicament for the treatment of diseases of the nervous system related to neuronal degeneration, depression, psychiatric disorders and Cognitive, trauma or cell injury, or other related neurological disorder, treatment of daytime fatigue, sleep disorders, loss of mental efficacy, weakness and irritability and symptoms related to hourly decompensation (jet-lag effect or transoceanic syndrome).
  • Neurogenesis is a vital process in the brains of vertebrates, in which new nerve cells are generated throughout the life of the individual. Although it was thought for a long time that the formation of nerve tissue was restricted to the early stages of embryonic development, this concept changed from 1962 when Altman demonstrated the formation of new neurons in the brain of adult rats (Altman, J. Are new neurons formed in the brains of adult mammals? Science 1962, 135, 1 127-1 128). In the following years, this same process was found in different species of vertebrates, including man (Eriksson, PS; Profileieva, E .; Bjork-Eriksson, T .; Alborn, AM; Nordborg, C; Peterson, DA; Gage, FH Neurogenesis in the adult human hippocampus. Nal Med. 1998, 4 , 1313-1317).
  • the complete neurogenesis process comprises four main stages: proliferation of stem or progenitor cells, migration to different areas of the central nervous system (CNS), differentiation and maturation in a specific cell type, and integration into neuronal circuits.
  • CNS central nervous system
  • differentiation and maturation in a specific cell type
  • integration into neuronal circuits.
  • SGZ subgranular zone
  • SVZ subventricular zone
  • multipotent progenitor neural cells continue to divide giving rise to new functional neurons and glial cells throughout the life of the individual (Tenth, I .; Bifari, F .; Krampera, M .; Fumagalli, G. Neural stem cell niches in health and diseases. Curr. Pharm. Des. 2012, 18, 1755-1783).
  • neurogenic drugs capable of promoting the formation of new neural populations and replacing those damaged by healthy and functionally competent cells, would imply a potential curative treatment for different neurodegenerative diseases and ischemic episodes, in which a loss of nerve cells occurs (Abdipranoto, A .; Wu, S .; Stayte, S .; Vissel, B. The role of neurogenesis in neurodegenerative diseases and its implications for therapeutic development. CNS Neurol. Disord. Drug Targets 2008, 7, 187-210).
  • Melatonin is an endogenous hormone produced in different tissues of the body, such as the pineal gland, the retina and the gastrointestinal tract. It participates in a wide variety of pathophysiological processes, including the regulation of the circadian cycle, the immune system and the endogenous antioxidant system. It is a proven fact that melatonin levels decrease with age and that this deficiency is associated with insomnia and depression. Recent research has also linked low levels of melatonin with aging and the development of neurodegenerative diseases, such as Alzheimer's (Bubenik, GA; Konturek, SJ Melatonin and aging: prospects for human treatment. J. Physiol. Pharmacol. 201 1, 62, 13-19).
  • melatonin has a very short half-life due to its rapid metabolism, which makes it advisable to have other ligands of the melatoninergic system that are metabolically more stable, in order to achieve a therapeutic effect superior to that of melatonin itself (Boutin, JA; Audinot, V .; Ferry, G .; Delagrange, P. Molecular tools to study melatonin pathways and actions. Trends Pharmacol. Sci. 2005, 26, 412-419).
  • melatonin receptor ligands have interesting properties in the CNS, mainly anxiolytic, antipsychotic and analgesic. Therapeutic applications have also been found in some types of cancer, in the regulation of ovulation, in diabetes and in the treatment of obesity (Zlotos, DP Recent progress in the development of agonists and antagonists for melatonin receptors. Curr. Med. Chem. 2012, 19, 3532-3549).
  • melatonin stimulates the synthesis of several endogenous antioxidant proteins, such as glutathione peroxidase that eliminates toxic radicals of the peroxide type.
  • mice with the double APP / Ps1 mutation the effects of melatonin on cerebral mitochondrial function have been studied, finding that the administration of this substance decreases between 2 and 4 times the levels of beta-amyloid peptide in different regions of the brain. And what is more interesting, this effect is accompanied by a complete recovery of mitochondrial respiratory rate, membrane potential and ATP levels in isolated mitochondria of the hippocampus, cortex and striatum (Dragicevic, N.
  • melatonin treatment mitochondrial restores function in Alzheimer's mice: a mitochondrial protective role of melatonin membrane receptor signaling. J. Pineal Res. 2011, 51, 75-86). Therefore, melatoninergic system ligands improve mitochondrial energy metabolism and can be considered innovative drugs for the treatment of neurodegenerative diseases, such as Alzheimer's.
  • melatonin has been shown to increase the survival of animals and improve neuronal function, even when administered 2 hours after the stroke. vascular. Melatonin increases neurogenesis and cell proliferation in the twilight regions of the infarcted regions, these effects being counteracted by melatonin receptor antagonists.
  • melatonin receptors by melatoninergic ligands could be an innovative treatment for diseases that occur with neuronal loss, such as neurodegenerations and accidents Cerebrovascular, among others (Chern, CM; Liao, JF; Wang, YH; Shen, YC Melatonin ameliorates neural function by promoting endogenous neurogenesis through the MT2 melatonin receptor in ischemic-stroke mice. Free Radie. Biol. Med. 2012, 52, 1634-1647).
  • a neurogenic compound based on melatonin and of general formula (I) constitutes a first object of the present invention:
  • X is O or N, with the particularity that when X is O, Y is N, L1 is absent and L2 is alkenyl or alkynyl
  • Y is O or N, with the particularity that when Y is O, X is N and L1 and L2 give rise to a 5-membered aromatic heterocycle comprising up to 3 heteroatoms and which can be substituted by hydrogen, (C1-C6) alkyl ), hydroxyl, (C1-C6) alkoxy, thiol or mercaptoalkyl (C1-C6) L2 is alkenyl, alkynyl (when X is O) or together with L1 forms a 5-membered aromatic heterocycle comprising up to 3 heteroatoms and which may be substituted by hydrogen, (C1-C6) alkyl, hydroxyl, (C1-C6) alkoxy , thiol or mercaptoalkyl (C1-C6)
  • X is O
  • Y N
  • n is equal to 1 or 2
  • L2 is allyl (prop-2-en- 1-yl) or prop-2-in-1-yl
  • X is N
  • Y is O
  • n is equal to 1 or 2
  • L1 and L2 form a 5-membered and three heteroatom aromatic heterocycle substituted with a radical R2 which is methyl, hydroxyl or thiol according to formula (V):
  • the compound is selected from the following group:
  • a process for obtaining a compound of general formula (II) consisting of the treatment of the corresponding carboxylic acid derived from 1 H-indole-3-yl in dry acetonitrile with allylamine or propargilamine in the presence of 1,1'-carbonyldiimidazole (CDI) and 4- (dimethylamino) pyridine (DMAP) .
  • CDI 1,1'-carbonyldiimidazole
  • DMAP 4- (dimethylamino) pyridine
  • the compound of general formula (III) is obtained by a procedure consisting in the treatment of the corresponding N- (prop-2-inyl) alkylamide derived from 1 H-indole-3-yl in dry dichloromethane with chloride of gold (III).
  • the compound of general formula (IV) is obtained by a process consisting of the reaction of an alkyl ester of the corresponding carboxylic acid derived from 1 H-indole-3-yl with acetamidoxime in the presence of sodium hydride.
  • the compound of general formula (V) is obtained by a procedure consisting of the reaction of the corresponding 1 H-indole-3-yl carboxylic acid in dry acetonitrile with the corresponding substituted hydrazide in the presence of 1, 1 '-carbonyldiimidazole (CDI) and 4- (dimethylamino) pyridine (DMAP), followed by treatment with phosphorus oxychloride.
  • CDI 1, 1 '-carbonyldiimidazole
  • DMAP 4- (dimethylamino) pyridine
  • the compound of general formula (VI) is obtained by a procedure consisting of reacting the hydrazide of the corresponding carboxylic acid derived from 1 H-indole-3-yl with an isocyanate or substituted isothiocyanate in a mixture of acetic acid and water, followed by treatment with sodium hydroxide in ethanol.
  • a further object of the present invention is a pharmaceutical composition
  • a pharmaceutical composition comprising a compound of formula (I) to (VI), which may also include another active ingredient.
  • the pharmaceutical composition is suitable for oral administration.
  • neurogenic compounds based on melatonin according to any one of the general formulas (I) to (VI) or those selected from compounds (1) to (24) in the preparation of a medicament or a pharmaceutical composition for the treatment of diseases of the nervous system that are selected among neurodegenerative diseases, the cognitive disorders, psychiatric diseases or disorders, trauma or cell injuries, neurological conditions and diseases or disorders related to the circadian cycle.
  • neurodegenerative disease is selected from Alzheimer's disease, Creutzfeld-Jacob disease, Parkinson's disease, systemic amyloidosis, amyotrophic lateral sclerosis, degenerative retinal disease, cerebral palsy, as well as a combination thereof. .
  • the cognitive disorder is selected from memory impairment, memory loss separate from dementia, mild cognitive impairment, age-related cognitive decline, memory loss due to attention deficit, cognitive impairment as a result of the use of anesthetics.
  • chemotherapy or radiotherapy cognitive impairment associated with post-surgical trauma or therapeutic intervention, cognitive decline associated with Alzheimer's disease or with epilepsy, senile dementia, vascular dementia, delirium, as well as related diseases or a combination thereof .
  • the psychiatric illness or disorder is selected from depression, major depression, neurotic depression, depression caused by drug or alcohol use, post-traumatic depression, postpartum depression, anxiety, obsessive-compulsive disorder, disorder bipolar, social phobia, seasonal mood disorder, as well as related diseases or a combination thereof.
  • the trauma or cell injury is selected from trauma or neurological injury, brain or spinal cord surgery, retinal injury, epilepsy-related injuries, brain or brain injuries. spinal cord in relation to the treatment of cancer, brain or spinal cord injuries related to infections, inflammatory processes, environmental toxins, ischemic episodes, as well as related diseases or a combination thereof.
  • the neurological condition is selected from learning disorder, autism, attention deficit disorder, narcolepsy, sleep disorder, epilepsy, temporal lobe epilepsy, as well as related diseases or a combination thereof.
  • the disease or disorder related to the circadian cycle is selected from sleep disorders, daytime fatigue, loss of mental efficacy, weakness and irritability and transoceanic syndrome, as well as related diseases or a combination thereof.
  • neurogenic compounds based on melatonin according to any one of the general formulas (I) to (VI) or those selected from the compounds (1) to (24), as a reagent in biological tests.
  • the invention relates to the synthesis of new products of general formula (I) and their use in the treatment of diseases of the nervous system related to neuronal degeneration, psychiatric and cognitive disorders, mood disorders, trauma or tissue injury, or other related neurological disorder, in which increased neurogenesis, modulation of melatonin and / or serotonin receptors, antioxidant properties and / or cholinergic properties cure or relieve the condition or disorder.
  • the invention relates to a compound of general formula (I):
  • X is O or N, with the particularity that when X is O, Y is N, L1 is absent and L2 is alkenyl or alkynyl
  • Y is O or N, with the particularity that when Y is O, X is N and L1 and L2 give rise to a 5-membered aromatic heterocycle comprising up to 3 heteroatoms and which can be substituted by hydrogen, (C1-C6) alkyl ), hydroxyl, (C1-C6) alkoxy, thiol or mercaptoalkyl (C1-C6)
  • L2 is alkenyl, alkynyl (when X is O) or together with L1 forms a 5-membered aromatic heterocycle comprising up to 3 heteroatoms and which may be substituted by hydrogen, (C1-C6) alkyl, hydroxyl, (C1-C6) alkoxy , thiol or mercaptoalkyl (C1-C6) or an isomer, a solvate, a pro-drug or a pharmaceutically acceptable salt thereof.
  • alkyl refers to hydrocarbon chains, linear or branched radicals, which consist of carbon and hydrogen atoms, which do not have unsaturation, which have 1 to 6 carbon atoms and which are attached to the rest of the molecule by a single bond, for example, methyl, ethyl, n-propyl, / - propyl, ⁇ -butyl, ferc-butyl, sec- butyl, n-pentyl or n-hexyl.
  • alkoxy refers to a radical of formula O-R where R is an alkyl group as defined above, for example, methoxy, ethoxy, propoxy, etc.
  • alkenyl refers to linear or branched hydrocarbon chain radicals containing one or more double carbon-carbon bonds, for example, vinyl, 1-propenyl, allyl, isoprenyl, 2-butenyl, 1, 3- butadienyl, etc. .
  • Alkenyl radicals may be optionally substituted by one or more substituents such as halogen, hydroxy, alkoxy, carboxyl, cyano, carbonyl, acyl, alkoxycarbonyl, amino, nitro, mercapto and alkylthio.
  • alkynyl refers to radicals of straight or branched hydrocarbon chains containing at least one triple carbon-carbon bond, for example, ethynyl and propynyl.
  • Alkynyl radicals can be optionally substituted by one or more substituents such as halogen, hydroxy, alkoxy, carboxyl, cyano, carbonyl, acyl, alkoxycarbonyl, amino, nitro, mercapto and alkylthio.
  • heterocycle refers to a stable 5-membered monocyclic radical, which is unsaturated, saturated or partially saturated, and which is formed by carbon atoms and at least two heteroatoms selected from the group consisting of nitrogen, oxygen and sulfur.
  • Halogen or “halo” refers to fluorine, chlorine, bromine or iodine.
  • mercaptoalkyl refers to a radical of formula SR where R is an alkyl group as defined above, for example, mercaptomethyl, mercaptoethyl, mercaptopropyl, etc.
  • R is an alkyl group as defined above, for example, mercaptomethyl, mercaptoethyl, mercaptopropyl, etc.
  • the compounds covered by this invention include the compounds defined in the general formulas, in the claims and those described in the examples.
  • the invention also includes those compounds that differ only in the presence of one or more isotopically enriched atoms.
  • the compounds having the present structures except for the replacement of a hydrogen with a deuterium or tritium, or the replacement of a carbon with a carbon enriched in 13 C or 14 C or a nitrogen enriched in 15 N, are within the scope of this invention.
  • the compounds of the present invention represented by formula (I) may include isomers, depending on the presence of multiple bonds, including optical isomers or enantiomers, depending on the presence of chiral centers.
  • the individual isomers, enantiomers or diastereoisomers and mixtures thereof fall within the scope of the present invention, that is, the term isomer also refers to any mixture of isomers, such as diastereomers, racemic, etc., even their optically isomers. assets or mixtures in different proportions thereof.
  • the individual enantiomers or diastereoisomers, as well as mixtures thereof, can be separated by conventional techniques.
  • prodrug or “pro-drug” as used herein includes any compound derived from a compound of formula (I), such as and not limited to: esters (including carboxylic acid esters, amino acid esters, amino acid esters). phosphate, sulphonate esters of metal salts, etc.), carbamates, amides, etc., which when administered to an individual can be transformed directly or indirectly into said compound of formula (I) in said individual.
  • said derivative is a compound that increases the bioavailability of the compound of formula (I) when administered to an individual or that enhances the release of the compound of formula (I) in a biological compartment.
  • the nature of said derivative is not critical as long as it can be administered to an individual and provides the compound of formula (I) in a biological compartment of an individual.
  • the preparation of said pro-drug can be carried out by conventional methods known to those skilled in the art. For example, pharmaceutically acceptable salts of compounds provided herein are synthesized by conventional chemical methods from an original compound containing a basic or acidic moiety.
  • such salts are prepared, for example, by reacting the free acid or base forms of the compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent or in a mixture of the two.
  • non-aqueous media such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile are preferred.
  • acid addition salts include mineral acid salts such as, for example, hydrochloride, hydrobromide, iodhydrate, sulfate, nitrate, phosphate and organic acid addition salts such as, for example, acetate, maleate, fumarate, citrate, oxalate, succinate, tartrate, malate, mandelate, methanesulfonate and p-toluenesulfonate.
  • base addition salts include inorganic salts such as, for example, sodium, potassium, calcium, ammonium, magnesium, aluminum and lithium salts, and salts of organic bases such as, for example, ethylenediamine, ethanolamine, N, N - dialkylene ethanolamine, triethanolamine, glucamine and basic amino acid salts.
  • Particularly preferred derivatives or pro-drugs are those that increase the bioavailability of the compounds of this invention when such compounds are administered to a patient (for example, by making a compound administered orally more easily absorbed by blood), or which enhances the release of the original compound in a biological compartment (for example, the brain or lymphatic system) in relation to the original species.
  • Any compound that is a pro-drug of a compound of general formula (I) is within the scope of the invention.
  • pro-drug is used in its broadest sense and encompasses those derivatives that are converted in vivo into the compounds of the invention.
  • esters amino acid esters, phosphate esters, metal salt sulphonate esters , carbamates and amides.
  • the compounds of general formula (I) may be in crystalline form as free compounds or as solvates and are both forms within the scope of the present invention.
  • Solvation methods are generally known within the art. Suitable solvates are pharmaceutically acceptable solvates. In a particular embodiment, the solvate is a hydrate.
  • the compounds of formula (I), their salts, prodrugs or solvates will preferably be in a pharmaceutically acceptable or substantially pure form, that is, having a pharmaceutically acceptable level of purity excluding normal pharmaceutical additives such as diluents and carriers, and not including material considered toxic at normal dosage levels.
  • the purity levels for the active ingredient are preferably greater than 50%, plus preferably, greater than 70%, and even more preferably, greater than 90%. In a preferred embodiment, they are greater than 95% of the compound of formula (I), or of its salts, solvates or pro-drugs.
  • the compounds of formula (I), their pharmaceutically acceptable salts, prodrugs or solvates thereof can therefore be used in the prevention and / or treatment of a disease or condition, in which the increase in neurogenesis, the Modulation of the melatonin and / or serotonin receptors, the antioxidant properties and the anticholinergic properties of the compounds of the invention prevent, cure or alleviate said disease or condition.
  • Pharmaceutical compositions containing a therapeutically effective amount of a compound of formula (I), its pharmaceutically acceptable salts, prodrugs or solvates thereof, together with pharmaceutically acceptable excipients constitute a further aspect of the present invention.
  • the amount of compound of formula (I), its pharmaceutically acceptable salts, prodrugs or solvates thereof, therapeutically effective to be administered as well as its dosage to treat a pathological state with said compounds will depend on numerous factors, among which is age , the patient's condition, the severity of the disease, the route and frequency of administration, the modulator compound to be used, etc.
  • the present invention further relates to the compounds of formulas (I) to (VI) for the manufacture of a medicament.
  • a further aspect of the present invention relates to a pharmaceutical composition
  • a pharmaceutical composition comprising the compounds of formulas (I) to (VI) as previously defined and at least one pharmaceutically acceptable excipient, adjuvant and / or carrier.
  • compositions may be administered by any appropriate route of administration, for example, oral, parenteral (subcutaneous, intraperitoneal, intravenous, intramuscular, etc.), rectal, etc.
  • said pharmaceutical compositions may be in a pharmaceutical form for oral administration, either in solid or liquid form.
  • Illustrative examples of pharmaceutical forms of oral administration include tablets, capsules, granules, solutions, suspensions, etc., and may contain conventional excipients, such as binders, diluents, disintegrants, lubricants, humectants, etc., and may be prepared. by conventional methods.
  • the pharmaceutical compositions can also be adapted for parenteral administration, in the form of, for example, sterile lyophilized solutions, suspensions or products, in the appropriate dosage form; in this case, said pharmaceutical compositions will include suitable excipients, such as buffers, surfactants, etc. In any case, the excipients will be chosen based on the pharmaceutical form of administration selected.
  • suitable excipients such as buffers, surfactants, etc.
  • the excipients will be chosen based on the pharmaceutical form of administration selected.
  • the compounds of the present invention result in increased neurogenesis, modulate the melatonin and / or serotonin receptors and exhibit antioxidant and / or cholinergic properties, said compounds may be useful for the preparation of a medicament or a composition.
  • Neural degeneration includes a neurodegenerative disorder, a neural stem cell disorder, a neural progenitor cell disorder, an ischemic episode or a combination thereof.
  • neurodegenerative disorders include, although non-limitingly, Alzheimer's disease, prion pathologies (for example, Creutzfeld-Jacob disease), Parkinson's disease, systemic amyloidosis, amyotrophic lateral sclerosis, degenerative retinal disease, cerebral palsy or a combination of same.
  • Cognitive disorders include senile dementia, vascular dementia, cognitive impairment, attention deficit disorder, as well as related disorders or a combination thereof.
  • Psychiatric conditions include, but are not limited to, depression, neurotic depression, depression caused by drug and / or alcohol use, post-traumatic depression, postpartum depression, anxiety, obsessive-compulsive disorder, bipolar disorder, social phobia, seasonal mood disorder and its combinations.
  • Cognitive disorders include, but are not limited to, memory impairment, memory loss separate from dementia, mild cognitive impairment, age-related cognitive decline, cognitive impairment as a result of the use of general anesthetics, chemotherapy, radiation treatment, trauma post-surgical, therapeutic intervention, cognitive decline associated with Alzheimer's disease or epilepsy, dementia, delirium, or a combination thereof.
  • the compounds defined above and the compositions derived therefrom are used to treat trauma or cell injury, including neurological trauma or injury, brain or spinal cord surgery, retinal injury, epilepsy-related injuries, injuries brain or spinal cord, brain or spinal cord injuries in relation to cancer treatment, brain or spinal cord injuries related to infections, inflammatory processes, environmental toxins, ischemic episodes, or a combination thereof.
  • the compounds and compositions object of the present patent are used to treat neurological conditions, such as learning disorder, autism, attention deficit disorder, narcolepsy, disorder. of sleep, epilepsy, temporal lobe epilepsy, or a combination thereof.
  • the compounds and compositions object of the present patent are used to treat the symptoms related to the circadian cycle or the time alteration, such as: sleep disorders, daytime fatigue, loss of mental efficacy, weakness and irritability and the transoceanic syndrome.
  • the present invention also relates to a compound of formula (I) as defined above for use in the treatment and / or prevention of a disease related to neurogenesis, modulation of melatonin and / or serotonin receptors. , presence of antioxidants and / or cholinesterase inhibition.
  • the present invention also relates to a method for the prevention or treatment of a disease related to neurogenesis, modulation of melatonin and / or serotonin receptors, presence of antioxidants and / or cholinesterase inhibition, which comprises administration to a patient in need of a therapeutically effective amount of a compound of formula (I) as previously defined.
  • the compounds of the present invention can be used as reagents in biological assays. This includes its use for the study of biology and neurogenesis mechanisms, its use as ligands for melatonin and serotonin receptors, its use as antioxidant and cholinergic agents.
  • the compounds of the present invention of formula (I) can be obtained or produced by a chemical synthetic route or obtained from a natural material of different origin.
  • the following examples are given only as an additional illustration of the invention, they should not be taken as a definition of the limits of the invention.
  • Schemes 1-5 show the procedures for the preparation of the compounds of the invention of general formulas (II) - (VI).
  • the treatment of the corresponding carboxylic acid derived from 1 / - -indole-3-yl in Dry acetonitrile with allylamine or propargilamine in the presence of carbonyldiimidazole (CDI) and 4- (dimethylamino) pyridine (DMAP) led to obtaining the products of general formula (II) (Scheme 1).
  • the compounds of the present invention can be purified by conventional methods, such as crystallization or chromatography. When the chemical synthesis process leads to mixtures of isomers, they can be separated by conventional techniques such as preparative chromatography. In the case of stereogenic centers the compounds can be obtained in the form of racemic or the pure enantiomers can be prepared by stereoselective, stereospecific synthesis or by resolution.
  • FIGURES Figure 1 Neural stem cell cultures of adult Wistar rats treated with vehicle (basal), melatonin (endogenous ligand of MT receptors), luzindole (antagonist of MT receptors), 4, 1 1 and 14, all at a concentration of 10 micromolar for 48 hours.
  • the anti-beta-tubulin antibody (clone Tuj1) was used as a marker associated with early neurogenesis.
  • FIG. 1 Neuronal stem cell cultures of adult Wistar rats treated with vehicle (basal), melatonin (endogenous ligand of MT receptors), luzindole (antagonist of MT receptors), 4, 1, 1 and 14, all at a concentration of 10 micromolar for 48 hours.
  • vehicle basic
  • melatonin endogenous ligand of MT receptors
  • luzindole antagonist of MT receptors
  • 4, 1, 1 and 14 all at a concentration of 10 micromolar for 48 hours.
  • the MAP-2 antibody microtubule-associated protein 2 was used as a marker associated with neuronal maturity.
  • a -Alyl-2- (5-methoxy-1H-indole-3-yl) acetamide (1) On a solution of 2- (5-methoxy-1 / - -indole-3-yl) acetic acid (172 mg, 0.84 mmol) in 20 mL of acetonitrile dry, carbonyldiimidazole (CDI) (163 mg, 1 mmol) and 4- (dimethylamino) pyridine (DMAP) (10 mg, 0.08 mmol) were added. After stirring for 2 hours at 50 ° C, allylamine (92 ⁇ _, 1.26 mmol) was added, maintaining the reaction at room temperature for an additional 2 hours.
  • CDI carbonyldiimidazole
  • DMAP 4- (dimethylamino) pyridine
  • A-Alyl-3- (5-methoxy-1H-indole-3-yl) propanamide (2) According to the preparation of A / -alyl-2- (5-methoxy-1 H-indol-3-yl) acetamide (1), from 3- (5-methoxy-1 / - / - indole) 3-yl) propanoic acid (80 mg, 0.36 mmol), 2 (81 mg, 87%) was obtained as a brown solid.
  • HEK and CHO cell lines with stable expression of human la and MT2 type melatonin receptors were cultured in DMEM medium (Dulbecco's modified Eagle's medium) supplemented with 10% fetal bovine serum, 2 mM glutamine, 100 Ul / mL penicillin and 100 mg / mL streptomycin.
  • the cells were grown to confluence at 37 ° C (95% O2 / 5% CO2), suspended in PBS containing 2 mM EDTA and centrifuged at 1000 xg for 5 min at 4 ° C. The resulting sediment was suspended in 5 mM Tris buffer (pH 7.5), containing 2 mM EDTA, and homogenized.
  • the homogenate was then centrifuged (95,000 xg, 30 min, 4 ° C) and the resulting sediment was suspended in 75 mM Tris buffer (pH 7.5) with 12.5 mM MgCl 2 and 2 mM EDTA. Aliquots of the membrane preparations were stored at -80 ° C until use.
  • Binding assays were performed by adding membrane preparations of transfected cells of the HEK or CHO type diluted in Tris-HCI buffer (50 mM, pH 7.4, with 5 mM MgCl 2 ) at 2- [ 125 L] iodomelatonin (25 or 200 pM for ⁇ and MT 2 receptors, respectively, expressed in HEK cells or 20 pM for ⁇ and MT 2 receptors expressed in CHO cells) and the compound to be tested.
  • Non-specific binding was defined in the presence of 1 ⁇ of melatonin.
  • Intrinsic activity assays were performed using [ 35 S] GTPyS (5'-O- [gamma-thio] guanosine triphosphate).
  • Membrane preparations of transfected CHO cells expressing the ⁇ and MT 2 subtypes, together with the compound tested, were diluted in HEPES buffer (20mM, pH 7.4, 100 mM NaCI, 3 ⁇ GDP, 3 mM MgCl 2 , and 20 ⁇ g / mL of saponin).
  • HEPES buffer 20mM, pH 7.4, 100 mM NaCI, 3 ⁇ GDP, 3 mM MgCl 2 , and 20 ⁇ g / mL of saponin.
  • a 0.2 nM solution of [ 35 S] GTPyS was added to the preparation containing the membranes (20 ⁇ g / mL) and the product to be evaluated and the whole was incubated for 1 hour at room temperature.
  • the membranes were previously incubated with both melatonin (3 nM) and the product to be tested for 30 min before the addition of [ 35 S] GTPyS.
  • Non-specific binding was defined using cold GTPyS (10 ⁇ ). All reactions were stopped by rapid filtration through GF / B filters, followed by three successive washes with ice-cold buffer.
  • the usual levels of [ 35 S] GTPyS (expressed in dpm) at the junction with the CHO-MT 2 membranes were: 2000 for baseline activity, 8000 in the presence of 1 ⁇ melatonin and 180 in the presence of 10 mM GTPyS defining non-specific binding.
  • the value The max (maximum inhibitory effect) was expressed as the percentage of the effect observed with melatonin, 30 nM or 3nM ([ago]) for the hMTi and hMT2 receptors, respectively.
  • Max is greater than 80%, the product is an agonist; when it is less than 20%, antagonist; and between these two figures, partial agonist.
  • the results of the intrinsic activity of a selection of the products of the invention are shown in Table 2.
  • the experiment was carried out on a Polarstar Galaxy fluorometer (BMG Labtechnologies GmbH, Offenburg, Germany) with 96-well plate reader, with excitation and emission filters at 485-P and 520-P, respectively.
  • the equipment was controlled by the Fluorostar Galaxy software (version 4.1 1 -0) for fluorescence measurement.
  • 2,2'-Azobis- (amidinopropane) dihydrochloride (AAPH), ( ⁇ ) -6-hydroxy-2, 5,7,8-tetramethylchroman-2-carboxylic acid (trolox) and fluorescein (FL) are acquired in Sigma-Aldrich.
  • the reaction was carried out in phosphate buffer (75 mM, pH 7.4), with a total volume of 200 ⁇ .
  • trolox in accordance with the value of 2.0 equiv. of trolox, previously described by Sofic et al. (Sofic, E .; Rimpapa, Z .; Kundurovic, Z .; Sapcanin, A .; Tahirovic, I .; Rustembegovic, A .; Cao, G. Antioxidant capacity of the neurohormone melatonin. J. Neural Transm. 2005, 112 , 349-358).
  • the products of the invention have good antioxidant properties, close to melatonin and up to 2.72 times more potent than trolox, the aromatic and active fragment of vitamin E and responsible for the capture of free radicals. Therefore, the products of the invention behave as useful antioxidants to counteract the oxidative stress caused by an excess of free radicals.
  • Table 3 Free radical capture (ORAC-FL), inhibition of human acetylcholinesterase (h-AChE) and human butyrylcholinesterase (h-BuChE) data.
  • Tacrine nd 0.04 ⁇ 0.002 0.010 ⁇ 0.004 a Average of 3 independent experiments ⁇ standard deviation. b Average of 3 independent experiments ⁇ SEM.
  • the compounds of the invention were evaluated as inhibitors of human acetyl- and butyrylcholinesterase (h-AChE and h-BuChE). Enzyme inhibition was measured using the Ellman method (Ellman, GL; Courtney, KD; Andrés, V., Jr .; Feather-Stone, RM A new and rapid colorimetric determination of acetylcholinesterase activity. Biochem. Pharmacol. 1961 , 7, 88-95).
  • h-AChE recombinant human acetylcholinesterase, Sigma Chemical Co.
  • h-BuChE human serum butyrylcholinesterase, Sigma Chemical Co.
  • the compounds to be evaluated were pre-incubated with the enzyme for 5 minutes at 30 ° C, the substrate was added and the absorbance changes were measured at 412 nm every 5 minutes in a Multiskan Spectrum UV / VIS spectrometer. The reaction rates were compared and the percentages of inhibition due to the presence of the compounds being analyzed were calculated. The enzymatic activity at each compound concentration is expressed as a percentage of activity with respect to the control in the absence of compound. IC50 is defined as the concentration of compound that inhibits 50% enzyme activity with respect to the control of untreated enzyme. The results are found in table 3, expressed as the average of three independent experiments ⁇ standard deviation.
  • results of inhibition of human cholinesterase indicate that most of the compounds evaluated are selective inhibitors of h-AChE with 50 inhibitory concentrations in the micromolar order. Therefore, they are able to moderately increase the levels of the neurotransmitter acetylcholine and improve the cognitive abilities of patients.
  • DMEM Dubecco's Modified Eagle's Medium
  • the cells obtained were cultured by established methods to achieve optimal proliferation in DMEM / F12 medium (1: 1, Invitrogen) and were supplemented with 10 ng / mL epidermal growth factor (EGF, Peprotech, London, United Kingdom), 10 ng / mL fibroblast growth factor (FGF, Peprotech) and medium B27 (Gibco) (Ferron, SR; Andreu-Agullo, C; Mira, H .; Sánchez, P .; Marqués-Torrejón, MA; Fari ⁇ as, I A combined ex / in vivo assay to detect effects of exogenously added factors in neural stem cells.
  • NS neurospheres
  • the neurospheres treated for 10 days on flotation were glued on a substrate (covers treated with 100 ⁇ g / mL of poly-L-lysine) and treated for 24, 48 and 96 more hours in the absence of exogenous and serum growth factors (Morales-Garc ⁇ a, JA; Luna-Medina, R .; Alonso-Gil, S .; Sanz-Sancristóbal, M .; Palomo, V .; Gil, C; Santos , A .; Mart ⁇ nez, A .; Pérez-Castillo, A.
  • Glycogen synthase kinase 3 inhibition promotes adult hippocampal neurogenesis in vitro and in vivo.
  • ACS Chem. Neurosci. 2012, 3, 963-971 After treatment, the crystals with the neurospheres were processed for immunocytochemistry with two types of neuronal markers associated with neurogenesis: anti-beta-tubulin antibody (clone Tuj1), related to early stages of neurogenesis and MAP-2 (microtubule-associated protein 2 ), marker of neuronal maturity.
  • the baseline values of the experiment were obtained under the same conditions, in the absence of product.
  • Melatonin (ligand) was used as controls endogenous to melatoninergic receptors) and luzindole (melatoninergic receptor antagonist).
  • the images were obtained using a Nikon 90i fluorescence microscope, coupled to a Qi digital camera. The microscope configuration was adjusted to produce the optimal signal-to-noise ratio.
  • Figures 1 and 2 show the neurogenic effect of the products of the invention on neuronal stem cell cultures of adult Wistar rats treated with vehicle (basal), melatonin (endogenous ligand of MT receptors), luzindole (antagonist of MT receptors) , 4, 11 and 14 all of them at a concentration of 10 micromolar.
  • Full neurosphere images are shown, as well as enlargements that show the inside (center of the neurosphere) and the outside of them (migration zone).
  • Two types of neuronal markers were used: anti-beta-tubulin antibody and MAP-2, respectively associated with early neurogenesis and neuronal maturity.
  • DAPI staining was used as a nuclear marker.
  • mice Animals. Neurogenesis studies in vivo were performed on C57BL / 6 age-matched male mice that do not express the transgene as wild-type animals. Once daily for 7 days, compound 11 (500 ⁇ g / kg) and BrdU (5-bromo-2-deoxyuridine) (50 mg / kg) were injected intraperitoneally to each mouse. The mice were divided into two groups and sacrificed under deep anesthesia, after 24 hours (group 1) or 21 days (group 2) since the last injection of BrdU. All animals were handled and treated according to the Directive of the European Parliament 2010/63 / EU, of September 22, 2010.
  • mice were deeply anesthetized with isophoran, perfused through the myocardium with 0.9% saline and their brains were immediately removed.
  • the tissues were then fixed in phosphate buffered solution pH 7.4 with 4% paraformaldehyde, at 4 ° C.
  • the fixed brains were cut in a vibratome (Leica Microsystems) at 30 ⁇ , and the tissue sections were collected in 0.1 M cold PBS and incubated at 4 ° C overnight with two primary antibodies, using mouse anti-BrdU ( 1: 20000, Hybridoma Bank) in both groups of mice.
  • BrdU is a modified nucleoside that is incorporated into DNA during the S phase of the cell cycle, so in these experiments it is used as a marker of newly formed cells.
  • DCX is a microtubule-associated protein expressed in immature neurons and NeuN is a marker of neuronal maturation.
  • BrdU + BrdU-positive cells
  • BrdU + cells were counted in each of the six sections from rostral (2 mm from the bregma) to flow rate (-4.3 mm from the bregma).
  • 100-150 BrdU + cells were analyzed through 4-6 sections per mouse by confocal microscopy for co-expression with DCX (group 1) or for co-expression with NeuN ( group 2).
  • BrdU + nuclei with DCX + colocalization in the cytoplasm were considered as newly born, still immature neurons, and those that co-localize BrdU + and NeuN + as mature neurons
  • the number of double-positive BrdlT-DCX + or BrdlT-NeuN + cells was expressed as a percentage against BrdlT cells.

Abstract

The invention, which is intended for use in the pharmaceutics and research field, relates to novel chemical entities derived from melatonin and having neurogenic properties, melatonin and/or serotonin receptor-modulating properties, and antioxidant and/or cholinergic properties. Experiments on mice have shown that they can be used for the in vivo regeneration of damaged neuronal populations, without leading to tumours. The invention also relates to methods for producing these novel compounds, to pharmaceutical compositions containing same and to the use thereof in the production of a drug for the treatment of diseases of the nervous system related to neuronal degeneration, depression, psychiatric and cognitive disorders, cell lesion or trauma and other related neurological disorders, treatment of daytime fatigue, sleep disorders, loss of mental acuity, debility and irritability and symptoms related with desynchronosis (jet lag or time zone change syndrome).

Description

COMPUESTOS NEUROGÉNICOS BASADOS EN MELATONINA Y SU EFICACIA EN EXPERIMENTOS IN VIVO PARA SU USO EN EL TRATAMIENTO DE ENFERMEDADES DEL SISTEMA NERVIOSO  NEELOGENIC COMPOUNDS BASED ON MELATONIN AND ITS EFFECTIVENESS IN IN VIVO EXPERIMENTS FOR USE IN THE TREATMENT OF NERVOUS SYSTEM DISEASES
SECTOR Y OBJETO DE LA INVENCION SECTOR AND OBJECT OF THE INVENTION
La presente invención, que se incluye en el campo de la investigación e industria farmacéutica, se refiere a nuevas entidades químicas derivadas de melatonina con propiedades neurogénicas, moduladoras de los receptores de melatonina y/o serotonina, antioxidantes y/o colinérgicas. Los experimentos en ratones han demostrado que son capaces de regenerar in vivo poblaciones neuronales dañadas, sin provocar la aparición de tumores. Esta invención también se refiere a los procedimientos para la preparación de estos nuevos compuestos, a las composiciones farmacéuticas que los contienen y a su uso para la fabricación de un medicamento para el tratamiento de enfermedades del sistema nervioso relacionadas con degeneración neuronal, depresión, trastornos psiquiátricos y cognitivos, trauma o lesión celular, u otro trastorno neurológico relacionado, tratamiento de fatiga diurna, trastornos del sueño, pérdida de eficacia mental, debilidad e irritabilidad y síntomas relacionados con la descompensación horaria (efecto jet-lag o síndrome transoceánico). The present invention, which is included in the field of research and pharmaceutical industry, refers to new chemical entities derived from melatonin with neurogenic properties, modulators of melatonin and / or serotonin receptors, antioxidants and / or cholinergic. Experiments in mice have shown that they are capable of regenerating damaged neuronal populations in vivo, without causing the appearance of tumors. This invention also relates to the processes for the preparation of these new compounds, the pharmaceutical compositions containing them and their use for the manufacture of a medicament for the treatment of diseases of the nervous system related to neuronal degeneration, depression, psychiatric disorders and Cognitive, trauma or cell injury, or other related neurological disorder, treatment of daytime fatigue, sleep disorders, loss of mental efficacy, weakness and irritability and symptoms related to hourly decompensation (jet-lag effect or transoceanic syndrome).
ESTADO DE LA TECNICA STATE OF THE TECHNIQUE
La neurogénesis es un proceso vital en el cerebro de los vertebrados, en el que se generan nuevas células nerviosas a lo largo de toda la vida del individuo. Aunque durante mucho tiempo se pensó que la formación de tejido nervioso estaba restringida a las primeras etapas del desarrollo embrionario, este concepto cambió a partir de 1962 cuando Altman demostró la formación de nuevas neuronas en el cerebro de ratas adultas (Altman, J. Are new neurons formed in the brains of adult mammals? Science 1962, 135, 1 127-1 128). En los años siguientes, se encontró este mismo proceso en diferentes especies de vertebrados, incluido el hombre (Eriksson, P. S.; Perfilieva, E.; Bjork-Eriksson, T.; Alborn, A. M.; Nordborg, C; Peterson, D. A.; Gage, F. H. Neurogenesis in the adult human hippocampus. Nal Med. 1998, 4, 1313-1317). Neurogenesis is a vital process in the brains of vertebrates, in which new nerve cells are generated throughout the life of the individual. Although it was thought for a long time that the formation of nerve tissue was restricted to the early stages of embryonic development, this concept changed from 1962 when Altman demonstrated the formation of new neurons in the brain of adult rats (Altman, J. Are new neurons formed in the brains of adult mammals? Science 1962, 135, 1 127-1 128). In the following years, this same process was found in different species of vertebrates, including man (Eriksson, PS; Profileieva, E .; Bjork-Eriksson, T .; Alborn, AM; Nordborg, C; Peterson, DA; Gage, FH Neurogenesis in the adult human hippocampus. Nal Med. 1998, 4 , 1313-1317).
El proceso completo de la neurogénesis comprende cuatro etapas principales: proliferación de las células madre o progenitoras, migración a diferentes áreas del sistema nervioso central (SNC), diferenciación y maduración en un tipo celular específico, e integración en los circuitos neuronales. En el cerebro de los mamíferos, incluido el hombre, existen dos principales depósitos de células madre neuronales: uno localizado en la zona subgranular (SGZ) del giro dentado del hipocampo y otro en la zona subventricular (SVZ) de los ventrículos laterales. En estas regiones, las células neurales progenitoras multipotentes continúan dividiéndose dando lugar a nuevas neuronas funcionales y células gliales a lo largo de la vida del individuo (Décimo, I.; Bifari, F.; Krampera, M.; Fumagalli, G. Neural stem cell niches in health and diseases. Curr. Pharm. Des. 2012, 18, 1755-1783). The complete neurogenesis process comprises four main stages: proliferation of stem or progenitor cells, migration to different areas of the central nervous system (CNS), differentiation and maturation in a specific cell type, and integration into neuronal circuits. In the mammalian brain, including man, there are two main neuronal stem cell deposits: one located in the subgranular zone (SGZ) of the dentate gyrus of the hippocampus and another in the subventricular zone (SVZ) of the lateral ventricles. In these regions, multipotent progenitor neural cells continue to divide giving rise to new functional neurons and glial cells throughout the life of the individual (Tenth, I .; Bifari, F .; Krampera, M .; Fumagalli, G. Neural stem cell niches in health and diseases. Curr. Pharm. Des. 2012, 18, 1755-1783).
Por lo tanto, los fármacos neurogénicos capaces de favorecer la formación de nuevas poblaciones neurales y reemplazar las dañadas por células sanas y funcionalmente competentes, supondrían un potencial tratamiento curativo para diferentes enfermedades neurodegenerativas y episodios isquémicos, en los que se produce una pérdida de células nerviosas (Abdipranoto, A.; Wu, S.; Stayte, S.; Vissel, B. The role of neurogenesis in neurodegenerative diseases and its implications for therapeutic development. CNS Neurol. Disord. Drug Targets 2008, 7, 187-210). Therefore, neurogenic drugs capable of promoting the formation of new neural populations and replacing those damaged by healthy and functionally competent cells, would imply a potential curative treatment for different neurodegenerative diseases and ischemic episodes, in which a loss of nerve cells occurs (Abdipranoto, A .; Wu, S .; Stayte, S .; Vissel, B. The role of neurogenesis in neurodegenerative diseases and its implications for therapeutic development. CNS Neurol. Disord. Drug Targets 2008, 7, 187-210).
Hasta el momento se han descubierto numerosas dianas biológicas implicadas en neurogénesis, como los receptores de serotonina, la presenilina 1 (PS1 ), la proteína precursora del beta-amiloide (APP) y sus metabolitos, la glucógeno sintasa quinasa 3 (GSK-3) y el sistema colinérgico, entre otros (Lazarov, O.; Marr, R. A. Neurogenesis and Alzheimer's disease: at the crossroads. Exp. Neurol. 2010, 223, 267-281 ). En lo que se refiere a la implicación de los receptores de serotonina en los mecanismos de neurogénesis, en estudios in vitro se ha observado que la estimulación de receptores 5-HTIA promueve la auto-renovación de las células precursoras neuronales y que la activación de los receptores 5-HT2c favorece la proliferación y diferenciación neuronal (Klempin, F.; Babu, H.; De Pietri Tonelli, D.; Alarcón, E.; Fabel, K.; Kempermann, G. Oppositional effects of serotonin receptors 5-HT1 a, 2, and 2c in the regulation of adult hippocampal neurogénesis. Front. Mol. Neurosci. 2010, 3, 14). So far, numerous biological targets involved in neurogenesis have been discovered, such as serotonin receptors, presenilin 1 (PS1), beta-amyloid precursor protein (APP) and its metabolites, glycogen synthase kinase 3 (GSK-3) and the cholinergic system, among others (Lazarov, O .; Marr, RA Neurogenesis and Alzheimer's disease: at the crossroads. Exp. Neurol. 2010, 223, 267-281). With regard to the involvement of serotonin receptors in neurogenesis mechanisms, in vitro studies it has been observed that the stimulation of 5-HTIA receptors promotes the self-renewal of neuronal precursor cells and that the activation of 5-HT 2 c receptors favor neuronal proliferation and differentiation (Klempin, F .; Babu, H .; De Pietri Tonelli, D .; Alarcón, E .; Fabel, K .; Kempermann, G. Oppositional effects of serotonin receptors 5 -HT1 a, 2, and 2c in the regulation of adult hippocampal neurogenesis. Front. Mol. Neurosci. 2010, 3, 14).
Además, estudios recientes indican la implicación de la mitocondria en la modulación de la neurogénesis, ya que el excesivo aumento de los niveles de radicales libres de oxígeno (ROS) y de óxido nítrico (NO), así como el aumento de citoquinas pro-inflamatorias inhiben la función mitocondrial y la proliferación de células madre neuronales (Voloboueva, L. A.; Giffard, R. G. Inflammation, mitochondria, and the inhibition of adult neurogénesis. J. Neurosci. Res. 2011 , 89, 1989-1996). In addition, recent studies indicate the involvement of mitochondria in the modulation of neurogenesis, as the excessive increase in levels of free oxygen radicals (ROS) and nitric oxide (NO), as well as the increase in pro-inflammatory cytokines inhibit mitochondrial function and proliferation of neuronal stem cells (Voloboueva, LA; Giffard, RG Inflammation, mitochondria, and the inhibition of adult neurogenesis. J. Neurosci. Res. 2011, 89, 1989-1996).
La melatonina es una hormona endógena producida en diferentes tejidos del organismo, como la glándula pineal, la retina y el tracto gastrointestinal. Participa en una gran variedad de procesos fisiopatológicos, entre los que se encuentra la regulación del ciclo circadiano, del sistema inmunitario y del sistema antioxidante endógeno. Es un hecho contrastado que los niveles de melatonina disminuyen con la edad y que esta deficiencia está asociada con insomnio y depresión. Investigaciones recientes también han relacionado los bajos niveles de melatonina con el envejecimiento y con el desarrollo de enfermedades neurodegenerativas, como por ejemplo la de Alzheimer (Bubenik, G. A.; Konturek, S. J. Melatonin and aging: prospects for human treatment. J. Physiol. Pharmacol. 201 1 , 62, 13-19). Melatonin is an endogenous hormone produced in different tissues of the body, such as the pineal gland, the retina and the gastrointestinal tract. It participates in a wide variety of pathophysiological processes, including the regulation of the circadian cycle, the immune system and the endogenous antioxidant system. It is a proven fact that melatonin levels decrease with age and that this deficiency is associated with insomnia and depression. Recent research has also linked low levels of melatonin with aging and the development of neurodegenerative diseases, such as Alzheimer's (Bubenik, GA; Konturek, SJ Melatonin and aging: prospects for human treatment. J. Physiol. Pharmacol. 201 1, 62, 13-19).
Muchos de los efectos farmacológicos de la melatonina se deben a su interacción directa con sus dos principales receptores, denominados ΜΤΊ y MT2, a su potente actividad antioxidante como captadora de radicales libres, o bien de manera indirecta a su interacción con receptores nucleares. Sin embargo, como fármaco la melatonina presenta una vida-media muy corta debido a su rápido metabolismo, lo que hace aconsejable disponer de otros ligandos del sistema melatoninérgico que sean metabólicamente más estables, con el fin de conseguir un efecto terapéutico superior al de la propia melatonina (Boutin, J. A.; Audinot, V.; Ferry, G.; Delagrange, P. Molecular tools to study melatonin pathways and actions. Trends Pharmacol. Sci. 2005, 26, 412-419). Many of the pharmacological effects of melatonin are due to its direct interaction with its two main receptors, called ΜΤΊ and MT 2 , to its potent antioxidant activity as a free radical scavenger, or indirectly to its interaction with nuclear receptors. Without However, as a drug, melatonin has a very short half-life due to its rapid metabolism, which makes it advisable to have other ligands of the melatoninergic system that are metabolically more stable, in order to achieve a therapeutic effect superior to that of melatonin itself (Boutin, JA; Audinot, V .; Ferry, G .; Delagrange, P. Molecular tools to study melatonin pathways and actions. Trends Pharmacol. Sci. 2005, 26, 412-419).
Además de los beneficios sobre el ciclo circadiano y los problemas de sueño, los ligandos de los receptores de melatonina presentan interesantes propiedades en el SNC, principalmente ansiolíticas, antipsicóticas y analgésicas. También se han encontrado aplicaciones terapéuticas en algunos tipos de cáncer, en la regulación de la ovulación, sobre la diabetes y en el tratamiento de la obesidad (Zlotos, D. P. Recent progress in the development of agonists and antagonists for melatonin receptors. Curr. Med. Chem. 2012, 19, 3532-3549). En el SNC, la melatonina estimula la síntesis de varias proteínas antioxidantes endógenas, como la glutatión peroxidasa que elimina radicales tóxicos de tipo peróxido. También se ha encontrado que produce efectos beneficiosos anti- amiloidogénicos y anti-apoptóticos, así como efectos neuroprotectores frente a la toxicidad provocada por el péptido beta-amiloide, marcador patológico característico de la enfermedad de Alzheimer (He, H.; Dong, W.; Huang, F. Anti-amyloidogenic and anti-apoptotic role of melatonin in Alzheimer disease. Curr. Neuropharmacol. 2010, 8, 21 1 -217). In addition to the benefits of the circadian cycle and sleep problems, melatonin receptor ligands have interesting properties in the CNS, mainly anxiolytic, antipsychotic and analgesic. Therapeutic applications have also been found in some types of cancer, in the regulation of ovulation, in diabetes and in the treatment of obesity (Zlotos, DP Recent progress in the development of agonists and antagonists for melatonin receptors. Curr. Med. Chem. 2012, 19, 3532-3549). In the CNS, melatonin stimulates the synthesis of several endogenous antioxidant proteins, such as glutathione peroxidase that eliminates toxic radicals of the peroxide type. It has also been found to produce beneficial anti-amyloidogenic and anti-apoptotic effects, as well as neuroprotective effects against toxicity caused by the beta-amyloid peptide, a pathological marker characteristic of Alzheimer's disease (He, H .; Dong, W. ; Huang, F. Anti-amyloidogenic and anti-apoptotic role of melatonin in Alzheimer disease. Curr. Neuropharmacol. 2010, 8, 21 1-217).
Recientemente se han estudiado los efectos in vivo de la melatonina, empleando diferentes modelos murinos de la enfermedad de Alzheimer, encontrándose resultados muy interesantes. En ratones transgénicos APP-695 se ha comprobado que esta molécula es capaz de aumentar la capacidad de aprendizaje y memoria (Feng, Z.; Chang, Y.; Cheng, Y.; Zhang, B. L; Qu, Z. W.; Qin, C; Zhang, J. T. Melatonin alleviates behavioral déficits associated with apoptosis and cholinergic system dysfunction in the APP 695 transgenic mouse model of Alzheimer's disease. J. Pineal. Res. 2004, 37, 129-136). Empleando ratones con la doble mutación APP/Ps1 , se han estudiado los efectos de la melatonina en la función mitocondrial cerebral, descubriéndose que la administración de esta sustancia disminuye entre 2 y 4 veces los niveles del péptido beta-amiloide en diferentes regiones del cerebro. Y lo que es más interesante, este efecto está acompañado por una completa recuperación de la frecuencia respiratoria mitocondrial, del potencial de membrana y de los niveles de ATP en las mitocondrias aisladas del hipocampo, de la corteza y del cuerpo estriado (Dragicevic, N.; Copes, N.; O'Neal-Moffitt, G.; Jin, J.; Buzzeo, R.; Mamcarz, M.; Tan, J.; Cao, C; Olcese, J. M.; Arendash, G. W.; Bradshaw, P. C. Melatonin treatment restores mitochondrial function in Alzheimer's mice: a mitochondrial protective role of melatonin membrane receptor signaling. J. Pineal Res. 2011 , 51, 75-86). Por lo tanto, los ligandos del sistema melatoninérgico mejoran el metabolismo energético mitocondrial y pueden considerarse fármacos innovadores para el tratamiento de enfermedades neurodegenerativas, como es la de Alzheimer. Recently, the in vivo effects of melatonin have been studied, using different murine models of Alzheimer's disease, finding very interesting results. In APP-695 transgenic mice it has been proven that this molecule is capable of increasing learning and memory capacity (Feng, Z .; Chang, Y .; Cheng, Y .; Zhang, B. L; Qu, ZW; Qin, C; Zhang, JT Melatonin alleviates behavioral deficits associated with apoptosis and cholinergic system dysfunction in the APP 695 transgenic mouse model of Alzheimer's disease. J. Pineal. Res. 2004, 37, 129-136). Using mice with the double APP / Ps1 mutation, the effects of melatonin on cerebral mitochondrial function have been studied, finding that the administration of this substance decreases between 2 and 4 times the levels of beta-amyloid peptide in different regions of the brain. And what is more interesting, this effect is accompanied by a complete recovery of mitochondrial respiratory rate, membrane potential and ATP levels in isolated mitochondria of the hippocampus, cortex and striatum (Dragicevic, N. ; Copes, N .; O'Neal-Moffitt, G .; Jin, J .; Buzzeo, R .; Mamcarz, M .; Tan, J .; Cao, C; Olcese, JM; Arendash, GW; Bradshaw, PC Melatonin treatment mitochondrial restores function in Alzheimer's mice: a mitochondrial protective role of melatonin membrane receptor signaling. J. Pineal Res. 2011, 51, 75-86). Therefore, melatoninergic system ligands improve mitochondrial energy metabolism and can be considered innovative drugs for the treatment of neurodegenerative diseases, such as Alzheimer's.
Resultados aún más recientes, han demostrado que un suplemento de melatonina retrasa la normal disminución de la neurogénesis en el hipocampo durante el proceso de envejecimiento de ratones (Ramírez-Rodríguez, G.; Vega-Rivera, N. M.; Benítez-King, G.; Castro-García, M.; Ortiz-López, L. Melatonin supplementation delays the decline of adult hippocampal neurogénesis during normal aging of mice. Neurosci. Lett. 2012, 530, 53-58). Por otra parte, en ratones sometidos a un daño cerebral transitorio de isquemia-reperfusión como modelo de ictus, se ha observado que la melatonina aumenta la supervivencia de los animales y mejora la función neuronal, incluso cuando es administrada 2 horas después del accidente cerebro-vascular. La melatonina aumenta la neurogénesis y la proliferación celular en las regiones de penumbra de las regiones infartadas, siendo estos efectos contrarrestados por antagonistas de los receptores de melatonina. Por lo tanto, la activación de los receptores de melatonina por ligandos melatoninérgicos podría ser un tratamiento innovador para enfermedades que cursan con pérdida neuronal, como las neurodegeneraciones y los accidentes cerebro-vasculares, entre otros (Chern, C. M.; Liao, J. F.; Wang, Y. H.; Shen, Y. C. Melatonin ameliorates neural function by promoting endogenous neurogenesis through the MT2 melatonin receptor in ischemic-stroke mice. Free Radie. Biol. Med. 2012, 52, 1634-1647). Even more recent results have shown that a melatonin supplement delays the normal decrease of neurogenesis in the hippocampus during the aging process of mice (Ramírez-Rodríguez, G .; Vega-Rivera, NM; Benítez-King, G .; Castro-García, M .; Ortiz-López, L. Melatonin supplementation delays the decline of adult hippocampal neurogenesis during normal aging of mice. Neurosci. Lett. 2012, 530, 53-58). On the other hand, in mice subjected to transient cerebral ischemia-reperfusion damage as a stroke model, melatonin has been shown to increase the survival of animals and improve neuronal function, even when administered 2 hours after the stroke. vascular. Melatonin increases neurogenesis and cell proliferation in the twilight regions of the infarcted regions, these effects being counteracted by melatonin receptor antagonists. Therefore, the activation of melatonin receptors by melatoninergic ligands could be an innovative treatment for diseases that occur with neuronal loss, such as neurodegenerations and accidents Cerebrovascular, among others (Chern, CM; Liao, JF; Wang, YH; Shen, YC Melatonin ameliorates neural function by promoting endogenous neurogenesis through the MT2 melatonin receptor in ischemic-stroke mice. Free Radie. Biol. Med. 2012, 52, 1634-1647).
EXPLICACION DE LA INVENCION EXPLANATION OF THE INVENTION
Constituye un primer objeto de la presente invención un compuesto neurogénico basado en la melatonina y de fórmula general (I): A neurogenic compound based on melatonin and of general formula (I) constitutes a first object of the present invention:
Figure imgf000007_0001
sus sales, pro-fármacos o solvatos farmacéuticamente aceptables,
Figure imgf000007_0001
its pharmaceutically acceptable salts, pro-drugs or solvates,
donde R1 es H, alquilo (C1 -C6) o alcoxilo (C1 -C6) n = 1 -6 where R1 is H, (C1-C6) alkyl or (C1-C6) alkoxy n = 1-6
X es O o N, con la particularidad de que cuando X es O, Y es N, L1 está ausente y L2 es alquenilo o alquinilo X is O or N, with the particularity that when X is O, Y is N, L1 is absent and L2 is alkenyl or alkynyl
Y es O o N, con la particularidad de que cuando Y es O, X es N y L1 y L2 dan lugar a un heterociclo aromático de 5 miembros que comprende hasta 3 heteroátomos y que puede estar sustituido por hidrógeno, alquilo (C1 -C6), hidroxilo, alcoxilo (C1 -C6), tiol o mercaptoalquilo (C1 -C6) L2 es alquenilo, alquinilo (cuando X es O) o forma junto con L1 un heterociclo aromático de 5 miembros que comprende hasta 3 heteroátomos y que puede estar sustituido por hidrógeno, alquilo (C1 -C6), hidroxilo, alcoxilo (C1 -C6), tiol o mercaptoalquilo (C1 -C6) En una realización preferida, en el compuesto de fórmula general (I), X es O, Y es N, n es igual a 1 o 2 y L2 es alilo (prop-2-en-1 -ilo) o prop-2-in-1 -ilo, según la fórmula (II): Y is O or N, with the particularity that when Y is O, X is N and L1 and L2 give rise to a 5-membered aromatic heterocycle comprising up to 3 heteroatoms and which can be substituted by hydrogen, (C1-C6) alkyl ), hydroxyl, (C1-C6) alkoxy, thiol or mercaptoalkyl (C1-C6) L2 is alkenyl, alkynyl (when X is O) or together with L1 forms a 5-membered aromatic heterocycle comprising up to 3 heteroatoms and which may be substituted by hydrogen, (C1-C6) alkyl, hydroxyl, (C1-C6) alkoxy , thiol or mercaptoalkyl (C1-C6) In a preferred embodiment, in the compound of general formula (I), X is O, Y is N, n is equal to 1 or 2 and L2 is allyl (prop-2-en- 1-yl) or prop-2-in-1-yl, according to formula (II):
Figure imgf000008_0001
Figure imgf000008_0001
Fórmula (II) En otra realización preferida, en el compuesto de fórmula general (I), X es N, Y es O, n es igual a 1 o 2 y L1 y L2 forman un heterociclo aromático de 5 miembros y dos heteroátomos sustituido con un metilo, según la fórmula (III): Formula (II) In another preferred embodiment, in the compound of general formula (I), X is N, Y is O, n is equal to 1 or 2 and L1 and L2 form a 5-membered aromatic heterocycle and two heteroatoms substituted with a methyl, according to formula (III):
Figure imgf000008_0002
Figure imgf000008_0002
Fórmula (III) En otra realización preferida, en el compuesto de fórmula general (I), X es N, Y es O, n es igual a 1 o 2 y L1 y L2 forman un heterociclo aromático de 5 miembros y tres heteroátomos sustituido con un metilo según la fórmula (IV): Formula (III) In another preferred embodiment, in the compound of general formula (I), X is N, Y is O, n is equal to 1 or 2 and L1 and L2 form a 5-membered aromatic heterocycle and three heteroatoms substituted with a methyl according to formula (IV):
Figure imgf000009_0001
Figure imgf000009_0001
Fórmula (IV) Formula (IV)
En otra realización preferida, en el compuesto de fórmula general (I), X es N, Y es O, n es igual a 1 o 2, y L1 y L2 forman un heterociclo aromático de 5 miembros y tres heteroátomos sustituido con un radical R2 que es metilo, hidroxilo o tiol según la fórmula (V): In another preferred embodiment, in the compound of general formula (I), X is N, Y is O, n is equal to 1 or 2, and L1 and L2 form a 5-membered and three heteroatom aromatic heterocycle substituted with a radical R2 which is methyl, hydroxyl or thiol according to formula (V):
Figure imgf000009_0002
Figure imgf000009_0002
Fórmula (V) Formula (V)
En otra realización preferida, en el compuesto de fórmula general (I), donde X es N, Y es N, n es igual a 1 o 2, y L1 y L2 forman un heterociclo aromático de 5 miembros y tres heteroátomos sustituido con un radical R2 que es hidroxilo o tiol y un radical R3 que es alquilo según la fórmula (VI): In another preferred embodiment, in the compound of general formula (I), where X is N, Y is N, n is equal to 1 or 2, and L1 and L2 form a 5-membered and three heteroatom aromatic heterocycle substituted with a radical R2 which is hydroxyl or thiol and a radical R3 which is alkyl according to formula (VI):
Figure imgf000009_0003
Figure imgf000009_0003
Fórmula (VI) En otras realizaciones preferidas, el compuesto se selecciona de entre el siguiente grupo: Formula (VI) In other preferred embodiments, the compound is selected from the following group:
N-Alil-2-(5-metoxi-1 H-indol-3-il)acetamida (1 ) N-Alyl-2- (5-methoxy-1 H-indol-3-yl) acetamide (1)
N-Alil-3-(5-metoxi-1 H-indol-3-il)propanamida (2)  N-Alyl-3- (5-methoxy-1 H-indol-3-yl) propanamide (2)
· 2-(5-Metoxi-1 H-indol-3-il)-N-(prop-2-in-1 -il)acetamida (3) 2- (5-Methoxy-1 H-indol-3-yl) -N- (prop-2-in-1-yl) acetamide (3)
3- (5-Metoxi-1 H-indol-3-il)-N-(prop-2-in-1 -il)propanamida (4)  3- (5-Methoxy-1 H-indol-3-yl) -N- (prop-2-in-1-yl) propanamide (4)
2-(2-(1 H-lndol-3-il)etil)-5-metiloxazol (5)  2- (2- (1 H-lndol-3-yl) ethyl) -5-methylxazole (5)
2-((1 H-lndol-3-il)metil)-5-metiloxazol (6)  2 - ((1 H-lndol-3-yl) methyl) -5-methylxazole (6)
2-((5-Metoxi-1 H-indol-3-il)metil)-5-metiloxazol (7)  2 - ((5-Methoxy-1 H-indol-3-yl) methyl) -5-methylxazole (7)
· 2-(2-(5-Metoxi-1 H-indol-3-il)etil)-5-metiloxazol (8) 2- (2- (5-Methoxy-1 H-indol-3-yl) ethyl) -5-methylxazole (8)
5-((5-Metoxi-1 H-indol-3-il)metil)-3-metil-1 ,2,4-oxadiazol (9)  5 - ((5-Methoxy-1 H-indol-3-yl) methyl) -3-methyl-1, 2,4-oxadiazole (9)
5-(2-(5-Metoxi-1 H-indol-3-il)etil)-3-metil-1 ,2,4-oxadiazol (10)  5- (2- (5-Methoxy-1 H-indol-3-yl) ethyl) -3-methyl-1, 2,4-oxadiazole (10)
2-(2-(5-Metoxi-1 H-indol-3-il)etil)-5-metil-1 ,3,4-oxadiazol (1 1 )  2- (2- (5-Methoxy-1 H-indol-3-yl) ethyl) -5-methyl-1, 3,4-oxadiazole (1 1)
5-((5-Metoxi-1 H-indol-3-il)metil)-1 ,3,4-oxadiazol-2-ol (12)  5 - ((5-Methoxy-1 H-indol-3-yl) methyl) -1, 3,4-oxadiazol-2-ol (12)
· 5-(2-(1 H-lndol-3-il)etil)-1 ,3,4-oxadiazol-2-ol (13) 5- (2- (1 H-lndol-3-yl) ethyl) -1, 3,4-oxadiazol-2-ol (13)
5-((5-Metoxi-1 H-indol-3-il)metilo)-1 ,3,4-oxadiazol-2-tiol (14)  5 - ((5-Methoxy-1 H-indol-3-yl) methyl) -1, 3,4-oxadiazol-2-thiol (14)
5-(2-(1 H-lndol-3-il)etil)-1 ,3,4-oxadiazol-2-tiol (15)  5- (2- (1 H-lndol-3-yl) ethyl) -1, 3,4-oxadiazol-2-thiol (15)
5-((5-Metoxi-1 H-indol-3-il)metil)-4-metil-4H-1 ,2,4-triazol-3-ol (16)  5 - ((5-Methoxy-1 H-indol-3-yl) methyl) -4-methyl-4H-1, 2,4-triazol-3-ol (16)
5-((1 H-lndol-3-il)metil)-4-metil-4H-1 ,2,4-triazol-3-ol (17)  5 - ((1 H-lndol-3-yl) methyl) -4-methyl-4H-1, 2,4-triazol-3-ol (17)
· 4-Etil-5-((5-metoxi-1 H-indol-3-il)metil)-4H-1 ,2,4-triazol-3-tiol (18) 4-Ethyl-5 - ((5-methoxy-1 H-indol-3-yl) methyl) -4H-1, 2,4-triazol-3-thiol (18)
5-((1 H-lndol-3-il)metil)-4-etil-4H-1 ,2,4-triazol-3-tiol (19)  5 - ((1 H-lndol-3-yl) methyl) -4-ethyl-4H-1, 2,4-triazol-3-thiol (19)
2-((5-Metoxi-1 H-indol-3-il)metil)-5-metil-1 ,3,4-oxadiazol (20)  2 - ((5-Methoxy-1 H-indol-3-yl) methyl) -5-methyl-1, 3,4-oxadiazole (20)
5-(2-(5-Metoxi-1 H-indol-3-il)etil)-1 ,3,4-oxadiazol-2-ol (21 )  5- (2- (5-Methoxy-1 H-indol-3-yl) ethyl) -1, 3,4-oxadiazol-2-ol (21)
5-(2-(5-Metoxi-1 H-indol-3-il)etil)-1 ,3,4-oxadiazol-2-tiol (22)  5- (2- (5-Methoxy-1 H-indol-3-yl) ethyl) -1, 3,4-oxadiazol-2-thiol (22)
· 5-(2-(5-Metoxi-1 H-indol-3-il)etil)-4-metil-4H-1 ,2,4-triazol-3-ol (23) 5- (2- (5-Methoxy-1 H-indol-3-yl) ethyl) -4-methyl-4H-1, 2,4-triazol-3-ol (23)
4- Etil-5-(2-(5-metoxi-1 H-indol-3-il)etil)-4H-1 ,2,4-triazol-3-tiol (24) o un isómero, un solvato, un pro-fármaco o una sal farmacéuticamente aceptable del mismo.  4- Ethyl-5- (2- (5-methoxy-1 H-indol-3-yl) ethyl) -4H-1, 2,4-triazol-3-thiol (24) or an isomer, a solvate, a pro-drug or a pharmaceutically acceptable salt thereof.
Constituye igualmente un objeto de la presente invención, un procedimiento de obtención de un compuesto de fórmula general (II) según las reivindicaciones 1 y 2, consistente en el tratamiento del correspondiente ácido carboxílico derivado de 1 H-indol-3-ilo en acetonitrilo seco con alilamina o propargilamina en presencia de 1 ,1 '-carbonildiimidazol (CDI) y 4-(dimetilamino)piridina (DMAP). En una realización preferida, el compuesto de fórmula general (III) se obtiene mediante un procedimiento consistente en el tratamiento de la correspondiente N-(prop-2-inil) alquilamida derivada de 1 H-indol-3-ilo en diclorometano seco con cloruro de oro (III). It is also an object of the present invention, a process for obtaining a compound of general formula (II) according to claims 1 and 2, consisting of the treatment of the corresponding carboxylic acid derived from 1 H-indole-3-yl in dry acetonitrile with allylamine or propargilamine in the presence of 1,1'-carbonyldiimidazole (CDI) and 4- (dimethylamino) pyridine (DMAP) . In a preferred embodiment, the compound of general formula (III) is obtained by a procedure consisting in the treatment of the corresponding N- (prop-2-inyl) alkylamide derived from 1 H-indole-3-yl in dry dichloromethane with chloride of gold (III).
En otra realización preferida, el compuesto de fórmula general (IV) se obtiene mediante un procedimiento consistente en la reacción de un alquil éster del correspondiente ácido carboxílico derivado de 1 H-indol-3-ilo con acetamidoxima en presencia de hidruro sódico. In another preferred embodiment, the compound of general formula (IV) is obtained by a process consisting of the reaction of an alkyl ester of the corresponding carboxylic acid derived from 1 H-indole-3-yl with acetamidoxime in the presence of sodium hydride.
En otra realización preferida, el compuesto de fórmula general (V) se obtiene mediante un procedimiento consistente en la reacción del correspondiente ácido carboxílico derivado de 1 H-indol-3-ilo en acetonitrilo seco con la correspondiente hidrazida sustituida en presencia de 1 ,1 '-carbonildiimidazol (CDI) y 4-(dimetilamino)piridina (DMAP), seguida del tratamiento con oxicloruro de fósforo. In another preferred embodiment, the compound of general formula (V) is obtained by a procedure consisting of the reaction of the corresponding 1 H-indole-3-yl carboxylic acid in dry acetonitrile with the corresponding substituted hydrazide in the presence of 1, 1 '-carbonyldiimidazole (CDI) and 4- (dimethylamino) pyridine (DMAP), followed by treatment with phosphorus oxychloride.
En otra realización preferida, el compuesto de fórmula general (VI) se obtiene mediante un procedimiento consistente en reacción de la hidrazida del correspondiente ácido carboxílico derivado de 1 H-indol-3-ilo con un isocianato o isotiocianato sustituido en una mezcla de ácido acético y agua, seguida del tratamiento con hidróxido sódico en etanol. In another preferred embodiment, the compound of general formula (VI) is obtained by a procedure consisting of reacting the hydrazide of the corresponding carboxylic acid derived from 1 H-indole-3-yl with an isocyanate or substituted isothiocyanate in a mixture of acetic acid and water, followed by treatment with sodium hydroxide in ethanol.
Constituye un ulterior objeto de la presente invención una composición farmacéutica que comprende un compuesto de fórmula (I) a (VI), pudiendo incluir además otro principio activo. En una realización preferida, la composición farmacéutica es adecuada para la administración oral. A further object of the present invention is a pharmaceutical composition comprising a compound of formula (I) to (VI), which may also include another active ingredient. In a preferred embodiment, the pharmaceutical composition is suitable for oral administration.
Constituye igualmente un objeto de la presente invención el uso de los compuestos neurogénicos basados en melatonina según una cualquiera de las fórmulas generales (I) a (VI) o de los seleccionados entre los compuestos (1 ) a (24) en la preparación de un medicamento o una composición farmacéutica para el tratamiento de enfermedades del sistema nervioso que se seleccionan entre las enfermedades neurodegenerativas, los trastornos cognitivos, las enfermedades o trastornos psiquiátricos, los traumas o lesiones celulares, las afecciones neurológicas y las enfermedades o trastornos relacionados con el ciclo circadiano. It is also an object of the present invention to use the neurogenic compounds based on melatonin according to any one of the general formulas (I) to (VI) or those selected from compounds (1) to (24) in the preparation of a medicament or a pharmaceutical composition for the treatment of diseases of the nervous system that are selected among neurodegenerative diseases, the cognitive disorders, psychiatric diseases or disorders, trauma or cell injuries, neurological conditions and diseases or disorders related to the circadian cycle.
En una realización preferida, la enfermedad neurodegenerativa se selecciona entre la enfermedad de Alzheimer, enfermedad de Creutzfeld-Jacob, enfermedad de Parkinson, amiloidosis sistémica, esclerosis lateral amiotrófica, enfermedad degenerativa de la retina, la parálisis cerebral, así como una combinación de las mismas. In a preferred embodiment, neurodegenerative disease is selected from Alzheimer's disease, Creutzfeld-Jacob disease, Parkinson's disease, systemic amyloidosis, amyotrophic lateral sclerosis, degenerative retinal disease, cerebral palsy, as well as a combination thereof. .
En otra realización preferida, el trastorno cognitivo se selecciona entre alteración de memoria, pérdida de memoria separada de la demencia, deterioro cognitivo leve, disminución cognitiva relacionada con la edad, pérdida de memoria por déficit de atención, deterioro cognitivo como consecuencia del uso de anestésicos generales, de quimioterapia o de radioterapia, deterioro cognitivo asociado a trauma post-quirúrgico o a intervención terapéutica, declive cognitivo asociado con la enfermedad de Alzheimer o con epilepsia, demencia senil, demencia vascular, delirio, así como enfermedades relacionadas o una combinación de las mismas. In another preferred embodiment, the cognitive disorder is selected from memory impairment, memory loss separate from dementia, mild cognitive impairment, age-related cognitive decline, memory loss due to attention deficit, cognitive impairment as a result of the use of anesthetics. general, chemotherapy or radiotherapy, cognitive impairment associated with post-surgical trauma or therapeutic intervention, cognitive decline associated with Alzheimer's disease or with epilepsy, senile dementia, vascular dementia, delirium, as well as related diseases or a combination thereof .
En otra realización preferida, la enfermedad o el trastorno psiquiátrico se selecciona entre depresión, depresión mayor, depresión neurótica, depresión provocada por el consumo de drogas o alcohol, depresión post-traumática, depresión post-parto, ansiedad, trastorno obsesivo-compulsivo, trastorno bipolar, fobia social, trastorno del estado de ánimo estacional, así como enfermedades relacionadas o una combinación de las mismas. In another preferred embodiment, the psychiatric illness or disorder is selected from depression, major depression, neurotic depression, depression caused by drug or alcohol use, post-traumatic depression, postpartum depression, anxiety, obsessive-compulsive disorder, disorder bipolar, social phobia, seasonal mood disorder, as well as related diseases or a combination thereof.
En otra realización preferida, el trauma o lesión celular se selecciona entre trauma o lesión neurológica, cirugía cerebral o de la médula espinal, lesión de la retina, lesiones relacionadas con epilepsia, lesiones cerebrales o de la médula espinal en relación con el tratamiento del cáncer, lesiones cerebrales o de la médula espinal relacionadas con infecciones, procesos inflamatorios, toxinas ambientales, episodios isquémicos, así como enfermedades relacionadas o una combinación de las mismas. En otra realización preferida, la afección neurológica se selecciona entre el trastorno del aprendizaje, autismo, trastorno por déficit de atención, narcolepsia, trastorno del sueño, epilepsia, epilepsia del lóbulo temporal, así como enfermedades relacionadas o una combinación de las mismas. In another preferred embodiment, the trauma or cell injury is selected from trauma or neurological injury, brain or spinal cord surgery, retinal injury, epilepsy-related injuries, brain or brain injuries. spinal cord in relation to the treatment of cancer, brain or spinal cord injuries related to infections, inflammatory processes, environmental toxins, ischemic episodes, as well as related diseases or a combination thereof. In another preferred embodiment, the neurological condition is selected from learning disorder, autism, attention deficit disorder, narcolepsy, sleep disorder, epilepsy, temporal lobe epilepsy, as well as related diseases or a combination thereof.
En otra realización preferida, la enfermedad o trastorno relacionado con el ciclo circadiano se selecciona entre los trastornos del sueño, fatiga diurna, pérdida de eficacia mental, debilidad e irritabilidad y el síndrome transoceánico, así como enfermedades relacionadas o una combinación de las mismas. In another preferred embodiment, the disease or disorder related to the circadian cycle is selected from sleep disorders, daytime fatigue, loss of mental efficacy, weakness and irritability and transoceanic syndrome, as well as related diseases or a combination thereof.
Por último, constituye igualmente un objeto de la presente invención el uso de los compuestos neurogénicos basados en melatonina, según una cualquiera de las fórmulas generales (I) a (VI) o de los seleccionados entre los compuestos (1 ) a (24), como reactivo en ensayos biológicos. Finally, it is also an object of the present invention to use the neurogenic compounds based on melatonin, according to any one of the general formulas (I) to (VI) or those selected from the compounds (1) to (24), as a reagent in biological tests.
DESCRIPCION DETALLADA DE LA INVENCION DETAILED DESCRIPTION OF THE INVENTION
La invención se relaciona con la síntesis de nuevos productos de fórmula general (I) y su empleo en el tratamiento de enfermedades del sistema nervioso relacionadas con degeneración neuronal, trastornos psiquiátricos y cognitivos, trastornos del estado de ánimo, trauma o lesión tisular, u otro trastorno neurológico relacionado, en el que el aumento de la neurogénesis, la modulación de los receptores de melatonina y/o de serotonina, las propiedades antioxidantes y/o las propiedades colinérgicas curen o alivien la afección o trastorno. The invention relates to the synthesis of new products of general formula (I) and their use in the treatment of diseases of the nervous system related to neuronal degeneration, psychiatric and cognitive disorders, mood disorders, trauma or tissue injury, or other related neurological disorder, in which increased neurogenesis, modulation of melatonin and / or serotonin receptors, antioxidant properties and / or cholinergic properties cure or relieve the condition or disorder.
En un primer aspecto, la invención se refiere a un compuesto de fórmula general (I): In a first aspect, the invention relates to a compound of general formula (I):
Figure imgf000014_0001
donde
Figure imgf000014_0001
where
R1 es H, alquilo (C1 -C6) o alcoxilo (C1 -C6) n = 1 -6 X es O o N, con la particularidad de que cuando X es O, Y es N, L1 está ausente y L2 es alquenilo o alquinilo R1 is H, (C1-C6) alkyl or (C1-C6) alkoxy n = 1-6 X is O or N, with the particularity that when X is O, Y is N, L1 is absent and L2 is alkenyl or alkynyl
Y es O o N, con la particularidad de que cuando Y es O, X es N y L1 y L2 dan lugar a un heterociclo aromático de 5 miembros que comprende hasta 3 heteroátomos y que puede estar sustituido por hidrógeno, alquilo (C1 -C6), hidroxilo, alcoxilo (C1 -C6), tiol o mercaptoalquilo (C1 -C6) Y is O or N, with the particularity that when Y is O, X is N and L1 and L2 give rise to a 5-membered aromatic heterocycle comprising up to 3 heteroatoms and which can be substituted by hydrogen, (C1-C6) alkyl ), hydroxyl, (C1-C6) alkoxy, thiol or mercaptoalkyl (C1-C6)
L2 es alquenilo, alquinilo (cuando X es O) o forma junto con L1 un heterociclo aromático de 5 miembros que comprende hasta 3 heteroátomos y que puede estar sustituido por hidrógeno, alquilo (C1 -C6), hidroxilo, alcoxilo (C1 -C6), tiol o mercaptoalquilo (C1 -C6) o un isómero, un solvato, un pro-fármaco o una sal farmacéuticamente aceptable del mismo. L2 is alkenyl, alkynyl (when X is O) or together with L1 forms a 5-membered aromatic heterocycle comprising up to 3 heteroatoms and which may be substituted by hydrogen, (C1-C6) alkyl, hydroxyl, (C1-C6) alkoxy , thiol or mercaptoalkyl (C1-C6) or an isomer, a solvate, a pro-drug or a pharmaceutically acceptable salt thereof.
En la definición anterior de los compuestos de fórmula general (I), los siguientes términos tienen el significado indicado: In the above definition of the compounds of general formula (I), the following terms have the indicated meaning:
El término "alquilo" se refiere a radicales de cadenas hidrocarbonadas, lineales o ramificadas, que consisten en átomos de carbono e hidrógeno, que no tienen insaturación, que tienen de 1 a 6 átomos de carbono y que se unen al resto de la molécula mediante un enlace sencillo, por ejemplo, metilo, etilo, n-propilo, /- propilo, π-butilo, ferc-butilo, sec-butilo, n-pentilo o n-hexilo. The term "alkyl" refers to hydrocarbon chains, linear or branched radicals, which consist of carbon and hydrogen atoms, which do not have unsaturation, which have 1 to 6 carbon atoms and which are attached to the rest of the molecule by a single bond, for example, methyl, ethyl, n-propyl, / - propyl, π-butyl, ferc-butyl, sec- butyl, n-pentyl or n-hexyl.
El término "alcoxilo" se refiere a un radical de fórmula O-R donde R es un grupo alquilo como definido anteriormente, por ejemplo, metoxilo, etoxilo, propoxilo, etc. The term "alkoxy" refers to a radical of formula O-R where R is an alkyl group as defined above, for example, methoxy, ethoxy, propoxy, etc.
El término "alquenilo" se refiere a radicales de cadenas hidrocarbonadas lineales o ramificadas que contienen uno o más enlaces carbono-carbono dobles, por ejemplo, vinilo, 1 -propenilo, alilo, isoprenilo, 2-butenilo, 1 ,3- butadienilo, etc. Los radicales alquenilos pueden estar opcionalmente sustituidos por uno o más sustituyentes tales como halógeno, hidroxilo, alcoxilo, carboxilo, ciano, carbonilo, acilo, alcoxicarbonilo, amino, nitro, mercapto y alquiltio. The term "alkenyl" refers to linear or branched hydrocarbon chain radicals containing one or more double carbon-carbon bonds, for example, vinyl, 1-propenyl, allyl, isoprenyl, 2-butenyl, 1, 3- butadienyl, etc. . Alkenyl radicals may be optionally substituted by one or more substituents such as halogen, hydroxy, alkoxy, carboxyl, cyano, carbonyl, acyl, alkoxycarbonyl, amino, nitro, mercapto and alkylthio.
El término "alquinilo" se refiere a radicales de cadenas hidrocarbonadas lineales o ramificadas que contienen al menos un enlace carbono-carbono triple, por ejemplo, etinil y propinil. Los radicales alquinilo pueden estar opcionalmente sustituidos por uno o más sustituyentes tales como halógeno, hidroxilo, alcoxilo, carboxilo, ciano, carbonilo, acilo, alcoxicarbonilo, amino, nitro, mercapto y alquiltio. El término "heterociclo" se refiere a un radical estable monocíclico de 5 miembros, que está insaturado, saturado o parcialmente saturado, y que está formado por átomos de carbono y al menos dos heteroátomos seleccionados del grupo que consiste en nitrógeno, oxígeno y azufre. The term "alkynyl" refers to radicals of straight or branched hydrocarbon chains containing at least one triple carbon-carbon bond, for example, ethynyl and propynyl. Alkynyl radicals can be optionally substituted by one or more substituents such as halogen, hydroxy, alkoxy, carboxyl, cyano, carbonyl, acyl, alkoxycarbonyl, amino, nitro, mercapto and alkylthio. The term "heterocycle" refers to a stable 5-membered monocyclic radical, which is unsaturated, saturated or partially saturated, and which is formed by carbon atoms and at least two heteroatoms selected from the group consisting of nitrogen, oxygen and sulfur.
"Halógeno" o "halo" se refiere a flúor, cloro, bromo o yodo. El término "mercaptoalquilo" se refiere a un radical de fórmula S-R donde R es un grupo alquilo como se ha definido anteriormente, por ejemplo, mercaptometilo, mercaptoetilo, mercaptopropilo, etc. Los compuestos cubiertos por esta invención incluyen los compuestos definidos en las fórmulas generales, en las reivindicaciones y los descritos en los ejemplos. "Halogen" or "halo" refers to fluorine, chlorine, bromine or iodine. The term "mercaptoalkyl" refers to a radical of formula SR where R is an alkyl group as defined above, for example, mercaptomethyl, mercaptoethyl, mercaptopropyl, etc. The compounds covered by this invention include the compounds defined in the general formulas, in the claims and those described in the examples.
La invención también incluye aquellos compuestos que difieren sólo en la presencia de uno o más átomos isotópicamente enriquecidos. Por ejemplo, los compuestos que tienen las presentes estructuras, a excepción de la sustitución de un hidrógeno por un deuterio o por tritio, o la sustitución de un carbono por un carbono enriquecido en 13C o 14C o un nitrógeno enriquecido en 15N, están dentro del alcance de esta invención. Los compuestos de la presente invención representados por la fórmula (I) pueden incluir isómeros, dependiendo de la presencia de enlaces múltiples, incluyendo isómeros ópticos o enantiómeros, dependiendo de la presencia de centros quirales. Los isómeros, enantiómeros o diastereoisómeros individuales y las mezclas de los mismos caen dentro del alcance de la presente invención, es decir, el término isómero también se refiere a cualquier mezcla de isómeros, como diastereómeros, racémicos, etc., incluso a sus isómeros ópticamente activos o las mezclas en distintas proporciones de los mismos. Los enantiómeros o diastereoisómeros individuales, así como sus mezclas, pueden separarse mediante técnicas convencionales. Asimismo, dentro del alcance de esta invención se encuentran las sales, solvatos y pro-fármacos farmacéuticamente aceptables de los compuestos de fórmula (I) o cualquier otro compuesto que, cuando se administra a un paciente es capaz de proporcionar (directamente o indirectamente) un compuesto según se describe en el presente documento. Sin embargo, se apreciará que las sales farmacéuticamente no aceptables también están dentro del alcance de la invención ya que éstas pueden ser útiles en la preparación de sales farmacéuticamente aceptables. La preparación de sales, solvatos, profármacos y derivados puede llevarse a cabo mediante métodos conocidos en la técnica. El término "prodroga" o "pro-fármaco" tal como aquí se utiliza incluye cualquier compuesto derivado de un compuesto de fórmula (I), como por ejemplo y no limitativamente: ásteres (incluyendo ásteres de ácidos carboxílicos, ásteres de aminoácidos, ásteres de fosfato, ásteres de sulfonato de sales metálicas, etc.), carbamatos, amidas, etc., que al ser administrado a un individuo puede ser transformado directa o indirectamente en dicho compuesto de fórmula (I) en el mencionado individuo. Ventajosamente, dicho derivado es un compuesto que aumenta la biodisponibilidad del compuesto de fórmula (I) cuando se administra a un individuo o que potencia la liberación del compuesto de fórmula (I) en un compartimento biológico. La naturaleza de dicho derivado no es crítica siempre y cuando pueda ser administrado a un individuo y proporcione el compuesto de fórmula (I) en un compartimento biológico de un individuo. La preparación de dicho pro-fármaco puede llevarse a cabo mediante métodos convencionales conocidos por los expertos en la materia. Por ejemplo, sales farmacéuticamente aceptables de compuestos previstos en el presente documento, se sintetizan mediante métodos químicos convencionales a partir de un compuesto original que contiene un resto básico o ácido. Generalmente, tales sales se preparan, por ejemplo, haciendo reaccionar las formas de ácido o base libre de los compuestos con una cantidad estequiométrica de la base o ácido apropiado en agua o en un disolvente orgánico o en una mezcla de los dos. Generalmente, se prefieren medios no acuosos como éter, acetato de etilo, etanol, isopropanol o acetonitrilo. Ejemplos de sales de adición de ácidos incluyen sales de ácido mineral tales como, por ejemplo, clorhidrato, bromhidrato, yodhidrato, sulfato, nitrato, fosfato y sales de adición de ácido orgánico tales como, por ejemplo, acetato, maleato, fumarato, citrato, oxalato, succinato, tartrato, malato, mandelato, metanosulfonato y p-toluensulfonato. Ejemplos de sales de adición de bases incluyen sales inorgánicas tales como, por ejemplo, sales de sodio, potasio, calcio, amonio, magnesio, aluminio y litio, y sales de bases orgánicas tales como, por ejemplo, etilenodiamina, etanolamina, N,N- dialquilenetanolamina, trietanolamina, glucamina y sales de aminoácidos básicos. The invention also includes those compounds that differ only in the presence of one or more isotopically enriched atoms. For example, the compounds having the present structures, except for the replacement of a hydrogen with a deuterium or tritium, or the replacement of a carbon with a carbon enriched in 13 C or 14 C or a nitrogen enriched in 15 N, are within the scope of this invention. The compounds of the present invention represented by formula (I) may include isomers, depending on the presence of multiple bonds, including optical isomers or enantiomers, depending on the presence of chiral centers. The individual isomers, enantiomers or diastereoisomers and mixtures thereof fall within the scope of the present invention, that is, the term isomer also refers to any mixture of isomers, such as diastereomers, racemic, etc., even their optically isomers. assets or mixtures in different proportions thereof. The individual enantiomers or diastereoisomers, as well as mixtures thereof, can be separated by conventional techniques. Also, within the scope of this invention are pharmaceutically acceptable salts, solvates and pro-drugs of the compounds of formula (I) or any other compound which, when administered to a patient, is capable of providing (directly or indirectly) a compound as described herein. However, it will be appreciated that pharmaceutically acceptable salts are also within the scope of the invention since these may be useful in the preparation of pharmaceutically acceptable salts. The preparation of salts, solvates, prodrugs and derivatives can be carried out by methods known in the art. The term "prodrug" or "pro-drug" as used herein includes any compound derived from a compound of formula (I), such as and not limited to: esters (including carboxylic acid esters, amino acid esters, amino acid esters). phosphate, sulphonate esters of metal salts, etc.), carbamates, amides, etc., which when administered to an individual can be transformed directly or indirectly into said compound of formula (I) in said individual. Advantageously, said derivative is a compound that increases the bioavailability of the compound of formula (I) when administered to an individual or that enhances the release of the compound of formula (I) in a biological compartment. The nature of said derivative is not critical as long as it can be administered to an individual and provides the compound of formula (I) in a biological compartment of an individual. The preparation of said pro-drug can be carried out by conventional methods known to those skilled in the art. For example, pharmaceutically acceptable salts of compounds provided herein are synthesized by conventional chemical methods from an original compound containing a basic or acidic moiety. Generally, such salts are prepared, for example, by reacting the free acid or base forms of the compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent or in a mixture of the two. Generally, non-aqueous media such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile are preferred. Examples of acid addition salts include mineral acid salts such as, for example, hydrochloride, hydrobromide, iodhydrate, sulfate, nitrate, phosphate and organic acid addition salts such as, for example, acetate, maleate, fumarate, citrate, oxalate, succinate, tartrate, malate, mandelate, methanesulfonate and p-toluenesulfonate. Examples of base addition salts include inorganic salts such as, for example, sodium, potassium, calcium, ammonium, magnesium, aluminum and lithium salts, and salts of organic bases such as, for example, ethylenediamine, ethanolamine, N, N - dialkylene ethanolamine, triethanolamine, glucamine and basic amino acid salts.
Los derivados o pro-fármacos particularmente favoritos son aquellos que aumentan la biodisponibilidad de los compuestos de esta invención cuando se administran tales compuestos a un paciente (por ejemplo, haciendo que un compuesto administrado por vía oral se absorba más fácilmente por la sangre), o que potencia la liberación del compuesto original en un compartimento biológico (por ejemplo, el cerebro o el sistema linfático) con relación a la especie original. Cualquier compuesto que es un pro-fármaco de un compuesto de fórmula general (I) está dentro del alcance de la invención. El término "pro-fármaco" se usa en su sentido más amplio y abarca aquellos derivados que se convierten in vivo en los compuestos de la invención. Tales derivados serán evidentes para aquellos expertos en la técnica, e incluyen, dependiendo de los grupos funcionales presentes en la molécula y sin limitación, los siguientes derivados de los compuestos: ésteres, ésteres de aminoácido, ésteres de fosfato, ésteres de sulfonato de sales metálicas, carbamatos y amidas. Particularly preferred derivatives or pro-drugs are those that increase the bioavailability of the compounds of this invention when such compounds are administered to a patient (for example, by making a compound administered orally more easily absorbed by blood), or which enhances the release of the original compound in a biological compartment (for example, the brain or lymphatic system) in relation to the original species. Any compound that is a pro-drug of a compound of general formula (I) is within the scope of the invention. The term "pro-drug" is used in its broadest sense and encompasses those derivatives that are converted in vivo into the compounds of the invention. Such derivatives will be apparent to those skilled in the art, and include, depending on the functional groups present in the molecule and without limitation, the following derivatives of the compounds: esters, amino acid esters, phosphate esters, metal salt sulphonate esters , carbamates and amides.
Los compuestos de fórmula general (I) pueden estar en forma cristalina como compuestos libres o como solvatos y están ambas formas dentro del alcance de la presente invención. Los métodos de solvatación se conocen generalmente dentro de la técnica. Los solvatos adecuados son solvatos farmacéuticamente aceptables. En una realización particular, el solvato es un hidrato. The compounds of general formula (I) may be in crystalline form as free compounds or as solvates and are both forms within the scope of the present invention. Solvation methods are generally known within the art. Suitable solvates are pharmaceutically acceptable solvates. In a particular embodiment, the solvate is a hydrate.
Para su aplicación en terapia, los compuestos de fórmula (I), sus sales, pro- fármacos o solvatos, se encontrarán, preferentemente, en una forma farmacéuticamente aceptable o sustancialmente pura, es decir, que tiene un nivel de pureza farmacéuticamente aceptable excluyendo los aditivos farmacéuticos normales tales como diluyentes y portadores, y no incluyendo material considerado tóxico a niveles de dosificación normales. Los niveles de pureza para el principio activo son preferiblemente superiores al 50%, más preferiblemente, superiores al 70%, y todavía más preferiblemente, superiores al 90%. En una realización preferida, son superiores al 95% de compuesto de fórmula (I), o de sus sales, solvatos o pro-fármacos. For their application in therapy, the compounds of formula (I), their salts, prodrugs or solvates, will preferably be in a pharmaceutically acceptable or substantially pure form, that is, having a pharmaceutically acceptable level of purity excluding normal pharmaceutical additives such as diluents and carriers, and not including material considered toxic at normal dosage levels. The purity levels for the active ingredient are preferably greater than 50%, plus preferably, greater than 70%, and even more preferably, greater than 90%. In a preferred embodiment, they are greater than 95% of the compound of formula (I), or of its salts, solvates or pro-drugs.
Los compuestos de fórmula (I), sus sales farmacéuticamente aceptables, prodrogas o solvatos del mismo, pueden ser utilizados, por tanto, en la prevención y/o el tratamiento de una enfermedad o condición, en la que el aumento de la neurogénesis, la modulación de los receptores de melatonina y/o de serotonina, las propiedades antioxidantes y las propiedades anticolinérgicas de los compuestos de la invención prevengan, curen o alivien dicha enfermedad o afección. Las composiciones farmacéuticas que contienen una cantidad terapéuticamente eficaz de un compuesto de fórmula (I), sus sales farmacéuticamente aceptables, prodrogas o solvatos del mismo, junto con los excipientes farmacéuticamente aceptables, constituyen un aspecto adicional de la presente invención. La cantidad de compuesto de fórmula (I), sus sales farmacéuticamente aceptables, prodrogas o solvatos del mismo, terapéuticamente eficaz que debe administrarse así como su dosificación para tratar un estado patológico con dichos compuestos dependerá de numerosos factores, entre los que se encuentra la edad, el estado del paciente, la severidad de la enfermedad, la ruta y frecuencia de administración, el compuesto modulador a utilizar, etc. The compounds of formula (I), their pharmaceutically acceptable salts, prodrugs or solvates thereof, can therefore be used in the prevention and / or treatment of a disease or condition, in which the increase in neurogenesis, the Modulation of the melatonin and / or serotonin receptors, the antioxidant properties and the anticholinergic properties of the compounds of the invention prevent, cure or alleviate said disease or condition. Pharmaceutical compositions containing a therapeutically effective amount of a compound of formula (I), its pharmaceutically acceptable salts, prodrugs or solvates thereof, together with pharmaceutically acceptable excipients, constitute a further aspect of the present invention. The amount of compound of formula (I), its pharmaceutically acceptable salts, prodrugs or solvates thereof, therapeutically effective to be administered as well as its dosage to treat a pathological state with said compounds will depend on numerous factors, among which is age , the patient's condition, the severity of the disease, the route and frequency of administration, the modulator compound to be used, etc.
La presente invención se refiere además a los compuestos de fórmulas (I) a (VI) para la fabricación de un medicamento. The present invention further relates to the compounds of formulas (I) to (VI) for the manufacture of a medicament.
Un aspecto adicional de la presente invención se refiere a una composición farmacéutica que comprende a los compuestos de fórmulas (I) a (VI) como se ha definido previamente y al menos un excipiente, adyuvante y/o vehículos farmacéuticamente aceptables. A further aspect of the present invention relates to a pharmaceutical composition comprising the compounds of formulas (I) to (VI) as previously defined and at least one pharmaceutically acceptable excipient, adjuvant and / or carrier.
Las composiciones farmacéuticas se pueden administrar por cualquier vía de administración apropiada, por ejemplo, oral, parenteral (subcutánea, intraperitoneal, intravenosa, intramuscular, etc.), rectal, etc. En una realización particular, dichas composiciones farmacéuticas pueden estar en una forma farmacéutica de administración por vía oral, bien en forma sólida o líquida. Ejemplos ilustrativos de formas farmacéuticas de administración por vía oral incluyen comprimidos, cápsulas, granulados, soluciones, suspensiones, etc., y pueden contener los excipientes convencionales, tales como aglutinantes, diluyentes, desintegrantes, lubrificantes, humectantes, etc., y pueden ser preparadas por métodos convencionales. Las composiciones farmacéuticas también pueden ser adaptadas para su administración parenteral, en forma de, por ejemplo, soluciones, suspensiones o productos liofilizados, estériles, en la forma de dosificación apropiada; en este caso, dichas composiciones farmacéuticas incluirán los excipientes adecuados, tales como tampones, tensioactivos, etc. En cualquier caso, los excipientes se elegirán en función de la forma farmacéutica de administración seleccionada. Una revisión de las distintas formas farmacéuticas de administración de fármacos y de su preparación puede encontrarse en el libro "Tratado de Farmacia Galénica", de C. Faulí i Trillo, 10 Edición, 1993, Luzán 5, S.A. de Ediciones, o en cualquier libro de similares características que exista en cada país. The pharmaceutical compositions may be administered by any appropriate route of administration, for example, oral, parenteral (subcutaneous, intraperitoneal, intravenous, intramuscular, etc.), rectal, etc. In a particular embodiment, said pharmaceutical compositions may be in a pharmaceutical form for oral administration, either in solid or liquid form. Illustrative examples of pharmaceutical forms of oral administration include tablets, capsules, granules, solutions, suspensions, etc., and may contain conventional excipients, such as binders, diluents, disintegrants, lubricants, humectants, etc., and may be prepared. by conventional methods. The pharmaceutical compositions can also be adapted for parenteral administration, in the form of, for example, sterile lyophilized solutions, suspensions or products, in the appropriate dosage form; in this case, said pharmaceutical compositions will include suitable excipients, such as buffers, surfactants, etc. In any case, the excipients will be chosen based on the pharmaceutical form of administration selected. A review of the different pharmaceutical forms of drug administration and their preparation can be found in the book "Treaty of Galenic Pharmacy", by C. Faulí i Trillo, 10 Edition, 1993, Luzán 5, SA de Ediciones, or in any book of similar characteristics that exist in each country.
Dado que los compuestos de la presente invención dan lugar al aumento de la neurogénesis, modulan los receptores de melatonina y/o de serotonina y presentan propiedades antioxidantes y/o propiedades colinérgicas, dichos compuestos pueden ser útiles para la preparación de un medicamento o una composición farmacéutica para el tratamiento de enfermedades del sistema nervioso relacionadas con degeneración neuronal, trastornos psiquiátricos y cognitivos, trauma o lesión celular, u otro trastorno neurológico relacionado. También para el tratamiento de enfermedades o trastornos relacionados con el ciclo circadiano. Since the compounds of the present invention result in increased neurogenesis, modulate the melatonin and / or serotonin receptors and exhibit antioxidant and / or cholinergic properties, said compounds may be useful for the preparation of a medicament or a composition. Pharmaceutical for the treatment of diseases of the nervous system related to neuronal degeneration, psychiatric and cognitive disorders, trauma or cell injury, or other related neurological disorder. Also for the treatment of diseases or disorders related to the circadian cycle.
La degeneración neuronal incluye un trastorno neurodegenerativo, un trastorno de las células madre neurales, un trastorno de las células progenitoras neurales, un episodio isquémico o una combinación de los mismos. En realizaciones adicionales, los trastornos neurodegenerativos incluyen, aunque de manera no limitante, la enfermedad de Alzheimer, patologías priónicas (por ejemplo, enfermedad de Creutzfeld-Jacob), la enfermedad de Parkinson, amiloidosis sistémica, esclerosis lateral amiotrófica, la enfermedad degenerativa de la retina, la parálisis cerebral o una combinación de las mismas. Los trastornos cognitivos incluyen demencia senil, demencia vascular, deterioro cognitivo, trastorno por déficit de atención, así como trastornos relacionados o una combinación de los mismos. Neural degeneration includes a neurodegenerative disorder, a neural stem cell disorder, a neural progenitor cell disorder, an ischemic episode or a combination thereof. In further embodiments, neurodegenerative disorders include, although non-limitingly, Alzheimer's disease, prion pathologies (for example, Creutzfeld-Jacob disease), Parkinson's disease, systemic amyloidosis, amyotrophic lateral sclerosis, degenerative retinal disease, cerebral palsy or a combination of same. Cognitive disorders include senile dementia, vascular dementia, cognitive impairment, attention deficit disorder, as well as related disorders or a combination thereof.
Las condiciones psiquiátricas incluyen, de manera no limitante, depresión, depresión neurótica, depresión provocada por el consumo de drogas y/o alcohol, depresión post-traumática, depresión post-parto, ansiedad, trastorno obsesivo-compulsivo, trastorno bipolar, fobia social, trastorno del estado de ánimo estacional y sus combinaciones. Psychiatric conditions include, but are not limited to, depression, neurotic depression, depression caused by drug and / or alcohol use, post-traumatic depression, postpartum depression, anxiety, obsessive-compulsive disorder, bipolar disorder, social phobia, seasonal mood disorder and its combinations.
Los trastornos cognitivos incluyen, de manera no limitante, alteración de memoria, pérdida de memoria separada de la demencia, deterioro cognitivo leve, disminución cognitiva relacionada con la edad, deterioro cognitivo como consecuencia del uso de anestésicos generales, quimioterapia, tratamiento por radiación, trauma post-quirúrgica, intervención terapéutica, declive cognitivo asociado con la enfermedad de Alzheimer o la epilepsia, la demencia, el delirio, o una combinación de los mismos. En otro aspecto, los compuestos anteriormente definidos y las composiciones derivadas de los mismos se utilizan para tratar un trauma o lesión celular, incluyendo trauma neurológico o lesión, cirugía cerebral o de la médula espinal, lesión de la retina, lesiones relacionadas con epilepsia, lesiones cerebrales o de la médula espinal, lesiones cerebrales o de la médula espinal en relación con el tratamiento del cáncer, lesiones cerebrales o de la médula espinal relacionadas con infecciones, procesos inflamatorios, toxinas ambientales, episodios isquémicos, o una combinación de los mismos. Cognitive disorders include, but are not limited to, memory impairment, memory loss separate from dementia, mild cognitive impairment, age-related cognitive decline, cognitive impairment as a result of the use of general anesthetics, chemotherapy, radiation treatment, trauma post-surgical, therapeutic intervention, cognitive decline associated with Alzheimer's disease or epilepsy, dementia, delirium, or a combination thereof. In another aspect, the compounds defined above and the compositions derived therefrom are used to treat trauma or cell injury, including neurological trauma or injury, brain or spinal cord surgery, retinal injury, epilepsy-related injuries, injuries brain or spinal cord, brain or spinal cord injuries in relation to cancer treatment, brain or spinal cord injuries related to infections, inflammatory processes, environmental toxins, ischemic episodes, or a combination thereof.
En otro aspecto, los compuestos y las composiciones objeto de la presente patente se utilizan para tratar afecciones neurológicas, como trastorno del aprendizaje, autismo, trastorno por déficit de atención, narcolepsia, trastorno del sueño, epilepsia, epilepsia del lóbulo temporal, o una combinación de las mismas. In another aspect, the compounds and compositions object of the present patent are used to treat neurological conditions, such as learning disorder, autism, attention deficit disorder, narcolepsy, disorder. of sleep, epilepsy, temporal lobe epilepsy, or a combination thereof.
En otro aspecto, los compuestos y las composiciones objeto de la presente patente se utilizan para tratar los síntomas relacionados con el ciclo circadiano o la alteración horaria, como son: los trastornos del sueño, fatiga diurna, pérdida de eficacia mental, debilidad e irritabilidad y el síndrome transoceánico. In another aspect, the compounds and compositions object of the present patent are used to treat the symptoms related to the circadian cycle or the time alteration, such as: sleep disorders, daytime fatigue, loss of mental efficacy, weakness and irritability and the transoceanic syndrome.
La presente invención se relaciona asimismo con un compuesto de fórmula (I) tal como se ha definido anteriormente para su uso en el tratamiento y/o prevención de una enfermedad relacionada con la neurogénesis, la modulación de los receptores de melatonina y/o de serotonina, presencia de antioxidantes y/o la inhibición de colinesterasas. The present invention also relates to a compound of formula (I) as defined above for use in the treatment and / or prevention of a disease related to neurogenesis, modulation of melatonin and / or serotonin receptors. , presence of antioxidants and / or cholinesterase inhibition.
La presente invención se relaciona igualmente con un método para la prevención o tratamiento de una enfermedad relacionada con la neurogénesis, la modulación de los receptores de melatonina y/o de serotonina, presencia de antioxidantes y/o la inhibición de colinesterasas, que comprende la administración a un paciente que lo necesita de una cantidad terapéuticamente efectiva de un compuesto de fórmula (I) como se ha definido previamente. The present invention also relates to a method for the prevention or treatment of a disease related to neurogenesis, modulation of melatonin and / or serotonin receptors, presence of antioxidants and / or cholinesterase inhibition, which comprises administration to a patient in need of a therapeutically effective amount of a compound of formula (I) as previously defined.
Los compuestos de la presente invención pueden ser utilizados como reactivos en ensayos biológicos. Esto incluye su uso para el estudio de la biología y los mecanismos de neurogénesis, su uso como ligandos de los receptores de melatonina y serotonina, su uso como agentes antioxidantes y colinérgicos. The compounds of the present invention can be used as reagents in biological assays. This includes its use for the study of biology and neurogenesis mechanisms, its use as ligands for melatonin and serotonin receptors, its use as antioxidant and cholinergic agents.
Los compuestos de la presente invención de formula (I) pueden ser obtenidos o producidos mediante una vía sintética química u obtenidos a partir de una materia natural de distinto origen. Los siguientes ejemplos se dan solo como una ilustración adicional de la invención, no deben tomarse como una definición de los límites de la invención. The compounds of the present invention of formula (I) can be obtained or produced by a chemical synthetic route or obtained from a natural material of different origin. The following examples are given only as an additional illustration of the invention, they should not be taken as a definition of the limits of the invention.
Los esquemas 1 -5 muestran los procedimientos para la preparación de los compuestos de la invención de fórmulas generales (II) - (VI). Por ejemplo, el tratamiento del correspondiente ácido carboxílico derivado de 1 /- -indol-3-ilo en acetonitrilo seco con alilamina o propargilamina en presencia de carbonildiimidazol (CDI) y 4-(dimetilamino)piridina (DMAP) condujo obtención de los productos de fórmula general (II) (Esquema 1 ). Schemes 1-5 show the procedures for the preparation of the compounds of the invention of general formulas (II) - (VI). For example, the treatment of the corresponding carboxylic acid derived from 1 / - -indole-3-yl in Dry acetonitrile with allylamine or propargilamine in the presence of carbonyldiimidazole (CDI) and 4- (dimethylamino) pyridine (DMAP) led to obtaining the products of general formula (II) (Scheme 1).
Figure imgf000023_0001
Esquema 1
Figure imgf000023_0001
Scheme 1
El tratamiento de la correspondiente A/-(prop-2-inil)alquilamida derivada de 1 H- indol-3-ilo en diclorometano seco con cloruro de oro (III) permitió obtener los compuestos de fórmula general (III) (Esquema 2). The treatment of the corresponding A / - (prop-2-inyl) alkylamide derived from 1 H-indole-3-yl in dry dichloromethane with gold chloride (III) allowed to obtain the compounds of general formula (III) (Scheme 2) .
Figure imgf000023_0002
Figure imgf000023_0002
Esquema 2 Scheme 2
La reacción de un alquil éster del correspondiente ácido carboxílico derivado de 1 /- -indol-3-ilo con acetamidoxima en presencia de hidruro sódico proporcionó los compuestos de fórmula general (IV) (Esquema 3). Reaction of an alkyl ester of the corresponding carboxylic acid derived from 1 / - -indole-3-yl with acetamidoxime in the presence of sodium hydride provided the compounds of general formula (IV) (Scheme 3).
Figure imgf000023_0003
Figure imgf000023_0003
Esquema 3 La reacción del correspondiente ácido carboxílico derivado de 1 /- -indol-3-ilo en acetonitrilo seco con la correspondiente hidrazida sustituida en presencia de 1 ,1 '-carbonildiimidazol (CDI) y 4-(dimetilamino)piridina (DMAP), seguida del tratamiento con oxicloruro de fósforo condujo a los compuestos de fórmula general (V) (Esquema 4). Scheme 3 The reaction of the corresponding carboxylic acid derived from 1 / - -indole-3-yl in dry acetonitrile with the corresponding substituted hydrazide in the presence of 1,1'-carbonyldiimidazole (CDI) and 4- (dimethylamino) pyridine (DMAP), followed by Phosphorus oxychloride treatment led to the compounds of general formula (V) (Scheme 4).
Figure imgf000024_0001
Figure imgf000024_0001
Esquema 4 Scheme 4
La reacción de la hidrazida del correspondiente ácido carboxílico derivado de 1 /- -indol-3-ilo con un isocianato o isotiocianato sustituido en una mezcla de ácido acético y agua, seguida del tratamiento con hidróxido sódico en etanol, condujo a los compuestos de fórmula general (VI) (Esquema 5). The reaction of the hydrazide of the corresponding carboxylic acid derived from 1 / - -indole-3-yl with a substituted isocyanate or isothiocyanate in a mixture of acetic acid and water, followed by treatment with sodium hydroxide in ethanol, led to the compounds of formula general (VI) (Scheme 5).
Figure imgf000024_0002
Esquema 5
Figure imgf000024_0002
Scheme 5
Los compuestos de la presente invención pueden ser purificados mediante métodos convencionales, tales como cristalización o cromatografía. Cuando el procedimiento de síntesis química conduce a mezclas de isómeros, éstos pueden ser separados por técnicas convencionales como cromatografía preparativa. En el caso de centros estereogénicos los compuestos pueden ser obtenidos en forma de racémico o bien los enantiomeros puros pueden ser preparados mediante síntesis estereoselectiva, estereoespecífica o mediante resolución. The compounds of the present invention can be purified by conventional methods, such as crystallization or chromatography. When the chemical synthesis process leads to mixtures of isomers, they can be separated by conventional techniques such as preparative chromatography. In the case of stereogenic centers the compounds can be obtained in the form of racemic or the pure enantiomers can be prepared by stereoselective, stereospecific synthesis or by resolution.
BREVE DESCRIPCION DE LAS FIGURAS Figura 1. Cultivos de células madre neuronales de ratas Wistar adultas tratadas con vehículo (basal), melatonina (ligando endógeno de los receptores MT), luzindol (antagonista de los receptores MT), 4, 1 1 y 14, todos a una concentración de 10 micromolar durante 48 horas. Se empleó el anticuerpo anti-beta-tubulina (clon Tuj1 ) como marcador asociado a neurogénesis temprana. BRIEF DESCRIPTION OF THE FIGURES Figure 1. Neural stem cell cultures of adult Wistar rats treated with vehicle (basal), melatonin (endogenous ligand of MT receptors), luzindole (antagonist of MT receptors), 4, 1 1 and 14, all at a concentration of 10 micromolar for 48 hours. The anti-beta-tubulin antibody (clone Tuj1) was used as a marker associated with early neurogenesis.
Figura 2. Cultivos de células madre neuronales de ratas Wistar adultas tratadas con vehículo (basal), melatonina (ligando endógeno de los receptores MT), luzindol (antagonista de los receptores MT), 4, 1 1 y 14, todos a una concentración de 10 micromolar durante 48 horas. Se empleó el anticuerpo MAP-2 (microtubule-associated protein 2) como marcador asociado a madurez neuronal. Figure 2. Neuronal stem cell cultures of adult Wistar rats treated with vehicle (basal), melatonin (endogenous ligand of MT receptors), luzindole (antagonist of MT receptors), 4, 1, 1 and 14, all at a concentration of 10 micromolar for 48 hours. The MAP-2 antibody (microtubule-associated protein 2) was used as a marker associated with neuronal maturity.
Figura 3. Efecto del compuesto 11 a corto tiempo sobre el número de células recién formadas marcadas con BrdlT en hipocampo de ratones, comparado con el control (sin producto). Los ratones fueron tratados una vez al día durante 7 días con el compuesto 11 (500 I kg) y BrdU (5-bromo-2-desoxiuridina) (50 mg / kg) por vía intraperitoneal. Después de 24 horas desde el último tratamiento los ratones fueron sacrificados y sus cerebros analizados, como se indica en el texto (n = 3). Figure 3. Effect of compound 11 in a short time on the number of newly formed cells labeled with BrdlT in hippocampus of mice, compared with the control (without product). Mice were treated once daily for 7 days with compound 11 (500 I kg) and BrdU (5-bromo-2-deoxyuridine) (50 mg / kg) intraperitoneally. After 24 hours since the last treatment the mice were sacrificed and their brains analyzed, as indicated in the text (n = 3).
Figura 4. Efecto del compuesto 11 a medio-largo tiempo sobre el número de células recién formadas marcadas con BrdlT en hipocampo de ratones, comparado con el control (sin producto). Los ratones fueron tratados una vez al día durante 7 días con el compuesto 11 (500 \g I kg) y BrdU (5-bromo-2- desoxiuridina) (50 mg / kg) por vía intraperitoneal. Después de 21 días desde el último tratamiento los ratones fueron sacrificados y sus cerebros analizados, como se indica en el texto (n = 3). Figure 4. Effect of compound 11 at medium-long time on the number of newly formed cells labeled with BrdlT in the hippocampus of mice, compared to the control (without product). Mice were treated once a day for 7 days with compound 11 (500 µg) and BrdU (5-bromo-2-deoxyuridine) (50 mg / kg) intraperitoneally. After 21 days from Last treatment the mice were sacrificed and their brains analyzed, as indicated in the text (n = 3).
Figura 5. Efecto del compuesto 11 a corto plazo sobre el número de neuronas inmaduras doblemente marcadas con BrdlT - DCX+ en hipocampo de ratones, comparado con el control (sin producto). Los ratones fueron tratados una vez al día durante 7 días con el compuesto 11 (500 I kg) y BrdU (5-bromo-2- desoxiuridina) (50 mg / kg) por vía intraperitoneal. Después de 24 horas desde el último día de tratamiento los ratones fueron sacrificados y sus cerebros analizados, como se indica en el texto (n = 6). Figura 6. Efecto del compuesto 11 a medio-largo plazo sobre el número de neuronas maduras doblemente marcadas con BrdlT - NeuN+ en hipocampo de ratones, comparado con el control (sin producto). Los ratones fueron tratados una vez al día durante 7 días con el compuesto 11 (500 \g I kg) y BrdU (5- bromo-2-desoxiuridina) (50 mg / kg) por vía intraperitoneal. Después de 21 días desde el último día de tratamiento los ratones fueron sacrificados y sus cerebros analizados, como se indica en el texto (n = 6). Figure 5. Short-term effect of compound 11 on the number of immature neurons doubly labeled with BrdlT-DCX + in the hippocampus of mice, compared to the control (without product). Mice were treated once daily for 7 days with compound 11 (500 I kg) and BrdU (5-bromo-2-deoxyuridine) (50 mg / kg) intraperitoneally. After 24 hours from the last day of treatment the mice were sacrificed and their brains analyzed, as indicated in the text (n = 6). Figure 6. Effect of compound 11 in the medium-long term on the number of mature neurons doubly labeled with BrdlT-NeuN + in the hippocampus of mice, compared to the control (without product). Mice were treated once daily for 7 days with compound 11 (500 µg kg) and BrdU (5- bromo-2-deoxyuridine) (50 mg / kg) intraperitoneally. After 21 days from the last day of treatment the mice were sacrificed and their brains analyzed, as indicated in the text (n = 6).
Figura 7. Microimágenes representativas de hipocampo de ratones tratados una vez al día durante 7 días con vehículo (basal) o compuesto 11 (500 \g I kg) y BrdU (5-bromo-2-desoxiuridina) (50 mg / kg) por vía intraperitoneal. Después de 21 días desde el último tratamiento los ratones fueron sacrificados y sus cerebros marcados con BrdU y NeuN, como se indica en el texto (n = 6). Se muestran ejemplos de neuronas maduras doblemente marcadas con BrdU y NeuN. Figure 7. Representative hippocampal micro images of mice treated once a day for 7 days with vehicle (baseline) or compound 11 (500 µg) and BrdU (5-bromo-2-deoxyuridine) (50 mg / kg) per intraperitoneal route. After 21 days since the last treatment the mice were sacrificed and their brains marked with BrdU and NeuN, as indicated in the text (n = 6). Examples of mature neurons doubly labeled with BrdU and NeuN are shown.
MODO DE REALIZACION DE LA INVENCION 1. Síntesis de los compuestos de la invención EMBODIMENT OF THE INVENTION 1. Synthesis of the compounds of the invention
A -Alil-2-(5-metoxi-1H-indol-3-il)acetamida (1). Sobre una disolución de ácido 2-(5-metoxi-1 /- -indol-3-il)acético (172 mg, 0.84 mmol) en 20 mL de acetonitrilo seco, se añadieron carbonildiimidazol (CDI) (163 mg, 1 mmol) y 4- (dimetilamino)piridina (DMAP) (10 mg, 0.08 mmol). Después de agitar durante 2 horas a 50 °C se añadió alilamina (92 μΙ_, 1.26 mmol), manteniendo la reacción a temperatura ambiente durante 2 horas adicionales. Tras la eliminación del disolvente a presión reducida, se obtuvo un crudo que fue tratado con HCI 1 M (20 mL) y la suspensión resultante fue extraída con acetato de etilo (3 x 20 mi). La fase orgánica se lavó con 2 M de NaOH (3 x 20 mL), se secó con sulfato de magnesio anhidro y el disolvente se eliminó a vacío, dando lugar a 1 (178 mg, 86%) como un aceite amarillo que cristalizó al reposar.1H NMR(300 MHz, DMSO-d6) δ 10.71 - 10.66 (s, 1H), 8.05-7.97 (m, 1H), 7.24- 7.19 (d, J = 8.7 Hz, 1H), 7.14-7.12 (d, J = 2.2 Hz, 1H), 7.07-7.04 (d, J = 2.3 Hz, 1H), 6.74 - 6.67 (dd, J = 8.7, 2.4 Hz, 1H), 5.85- 5.70 (ddt, J= 17.1, 10.3, 5.2 Hz, 1H), 5.12 - 5.05 (dd, J = 17.2, 1.8 Hz, 1H), 5.03 - 4.97 (dd, J = 10.3, 1.7 Hz, 1H), 3.74 - 3.72 (s, 3H), 3.71 - 3.66 (m, 2H), 3.48 - 3.48 (s, 2H). 13C NMR (75 MHz, DMSO-d6) δ 170.51, 152.96, 135.49, 131.21, 127.49, 124.42, 114.85, 111.90, 111.04, 108.61, 100.53, 55.30, 40.90, 32.70. HPLC-MS (m/z) [MH+] 245. A -Alyl-2- (5-methoxy-1H-indole-3-yl) acetamide (1). On a solution of 2- (5-methoxy-1 / - -indole-3-yl) acetic acid (172 mg, 0.84 mmol) in 20 mL of acetonitrile dry, carbonyldiimidazole (CDI) (163 mg, 1 mmol) and 4- (dimethylamino) pyridine (DMAP) (10 mg, 0.08 mmol) were added. After stirring for 2 hours at 50 ° C, allylamine (92 µΙ_, 1.26 mmol) was added, maintaining the reaction at room temperature for an additional 2 hours. After removal of the solvent under reduced pressure, a crude was obtained which was treated with 1 M HCI (20 mL) and the resulting suspension was extracted with ethyl acetate (3 x 20 mL). The organic phase was washed with 2 M NaOH (3 x 20 mL), dried with anhydrous magnesium sulfate and the solvent was removed in vacuo, yielding 1 (178 mg, 86%) as a yellow oil that crystallized on rest. 1 H NMR (300 MHz, DMSO-d 6 ) δ 10.71 - 10.66 (s, 1H), 8.05-7.97 (m, 1H), 7.24- 7.19 (d, J = 8.7 Hz, 1H), 7.14-7.12 (d , J = 2.2 Hz, 1H), 7.07-7.04 (d, J = 2.3 Hz, 1H), 6.74 - 6.67 (dd, J = 8.7, 2.4 Hz, 1H), 5.85- 5.70 (ddt, J = 17.1, 10.3 , 5.2 Hz, 1H), 5.12 - 5.05 (dd, J = 17.2, 1.8 Hz, 1H), 5.03 - 4.97 (dd, J = 10.3, 1.7 Hz, 1H), 3.74 - 3.72 (s, 3H), 3.71 - 3.66 (m, 2H), 3.48-3.48 (s, 2H). 13 C NMR (75 MHz, DMSO-d 6 ) δ 170.51, 152.96, 135.49, 131.21, 127.49, 124.42, 114.85, 111.90, 111.04, 108.61, 100.53, 55.30, 40.90, 32.70. HPLC-MS (m / z) [MH + ] 245.
A-Alil-3-(5-metoxi-1H-indol-3-il)propanamida (2). De acuerdo con la preparación de A/-alil-2-(5-metoxi-1 H-indol-3-il) acetamida (1), a partir del ácido 3-(5-metoxi-1/-/-indol-3-il)propanoico (80 mg, 0.36 mmol), se obtuvo 2 (81 mg, 87%) como un sólido marrón.1H NMR (300 MHz, DMSO-d6) δ 10.62 - 10.54 (s, 1 H), 8.03 - 7.93 (t, J = 5.5 Hz, 1 H), 7.22 - 7.16 (d, J = 8.7 Hz, 1 H), 7.05 - 7.01 (d, J = 2.3 Hz, 1 H), 6.99 - 6.97 (d, J = 2.4 Hz, 1 H), 6.72 - 6.66 (dd, J = 8.7, 2.4 Hz, 1H), 5.83-5.69 (ddt, J= 17.2, 10.3, 5.2 Hz, 1H), 5.11 -4.98 (m, 2H), 3.76 - 3.74 (s, 3H), 3.71 - 3.65 (tt, J = 5.6, 1.7 Hz, 2H), 2.92 - 2.83 (t, J = 7.7 Hz, 2H), 2.48 - 2.41 (t, J = 7.7 Hz, 2H). 13C NMR (75 MHz, DMSO-d6) δ 172.08, 153.25, 135.87, 131.72, 127.66, 123.14, 115.30, 113.98, 112.25, 111.34, 100.62, 55.72, 41.18, 36.48, 21.44. HPLC-MS (m/z) [MH+] 259. A-Alyl-3- (5-methoxy-1H-indole-3-yl) propanamide (2). According to the preparation of A / -alyl-2- (5-methoxy-1 H-indol-3-yl) acetamide (1), from 3- (5-methoxy-1 / - / - indole) 3-yl) propanoic acid (80 mg, 0.36 mmol), 2 (81 mg, 87%) was obtained as a brown solid. 1 H NMR (300 MHz, DMSO-d 6 ) δ 10.62 - 10.54 (s, 1 H), 8.03 - 7.93 (t, J = 5.5 Hz, 1 H), 7.22 - 7.16 (d, J = 8.7 Hz, 1 H), 7.05 - 7.01 (d, J = 2.3 Hz, 1 H), 6.99 - 6.97 (d, J = 2.4 Hz, 1 H), 6.72 - 6.66 (dd, J = 8.7, 2.4 Hz, 1H), 5.83 -5.69 (ddt, J = 17.2, 10.3, 5.2 Hz, 1H), 5.11 -4.98 (m, 2H), 3.76 - 3.74 (s, 3H), 3.71 - 3.65 (tt, J = 5.6, 1.7 Hz, 2H) , 2.92 - 2.83 (t, J = 7.7 Hz, 2H), 2.48 - 2.41 (t, J = 7.7 Hz, 2H). 13 C NMR (75 MHz, DMSO-d6) δ 172.08, 153.25, 135.87, 131.72, 127.66, 123.14, 115.30, 113.98, 112.25, 111.34, 100.62, 55.72, 41.18, 36.48, 21.44. HPLC-MS (m / z) [MH + ] 259.
2-(5-Metoxi-1H-indol-3-il)-W-(prop-2-in-1-il)acetamida (3). Siguiendo un método similar al anterior para la obtención de 1, a partir del ácido 2-(5-metoxi- 1/--indol-3-il)acético (200 mg, 1 mmol) y propargilamina (0.64 mi, 1 mmol) se obtuvo 3 (146 mg, 61%) como un sólido blanco.1H NMR (500 MHz, DMSO-d6) δ 10.70 (s, 1H), 8.33 (t, J = 5.5 Hz, 1H), 7.22 (d, J = 8.7 Hz, 1H), 7.13 (d, J = 2.4 Hz, 1 H), 7.04 (d, J = 2.4 Hz, 1 H), 6.71 (dd, J = 8.7, 2.5 Hz, 1 H), 3.85 (dd, J = 5.6, 2.6 Hz, 2H), 3.75 (s, 3H), 3.47 (s, 2H), 3.08 (t, J = 2.5 Hz, 1H).13C NMR (126 MHz, DMSO-d6) δ 170.91, 153.45, 131.65, 127.93, 124.87, 112.36, 111.53, 108.73, 100.99, 81.81, 73.26, 55.79, 32.93, 28.39. HPLC-MS (m/z) [MH+] 243. 2- (5-Methoxy-1H-indol-3-yl) -W- (prop-2-in-1-yl) acetamide (3). Following a method similar to the previous one to obtain 1, from 2- (5-methoxy-1 / -indol-3-yl) acetic acid (200 mg, 1 mmol) and propargilamine (0.64 ml, 1 mmol) be obtained 3 (146 mg, 61%) as a white solid. 1 H NMR (500 MHz, DMSO-d6) δ 10.70 (s, 1H), 8.33 (t, J = 5.5 Hz, 1H), 7.22 (d, J = 8.7 Hz, 1H), 7.13 (d, J = 2.4 Hz, 1 H), 7.04 (d, J = 2.4 Hz, 1 H), 6.71 (dd, J = 8.7, 2.5 Hz, 1 H), 3.85 (dd, J = 5.6, 2.6 Hz, 2H), 3.75 ( s, 3H), 3.47 (s, 2H), 3.08 (t, J = 2.5 Hz, 1H). 13 C NMR (126 MHz, DMSO-d6) δ 170.91, 153.45, 131.65, 127.93, 124.87, 112.36, 111.53, 108.73, 100.99, 81.81, 73.26, 55.79, 32.93, 28.39. HPLC-MS (m / z) [MH + ] 243.
3-(5-Metoxi-1H-indol-3-il)-W-(prop-2-in-1-il)propanamida (4). Emplean-do el método descrito para la obtención de 1, a partir del ácido 2-(5-metoxi-1/--indol- 3-il) propanoico (192 mg, 1 mmol) y propargilamina (0.71 mi, 1.1 mmol) se obtuvo 4 (160 mg, 65%) como un sólido blanco.1H NMR (300 MHz, DMSO-d6) δ 10.57 (s, 1H), 8.29 (s, 1H), 7.18 (d, J= 8.7 Hz, 1H), 7.02 (s, 1H), 6.97 (s, 1H), 6.68 (d, J = 8.6 Hz, 1 H), 3.90 - 3.80 (m, 2H), 3.74 (s, 3H), 3.09 - 3.07 (t, J = 2.7 Hz , 2H), 2.85 (t, J = 7.7 Hz, 2H), 2.42 (t, J = 6.8 Hz, 2H). 13C NMR (75 MHz, Acetone-d6)6173.16, 155.26, 133.31, 129.32, 124.06, 115.76, 113.29, 112.93, 101.79, 82.19, 72.44, 56.54, 37.93, 29.38, 22.53. HPLC-MS (m/z) [MH+] 257. 3- (5-Methoxy-1H-indol-3-yl) -W- (prop-2-in-1-yl) propanamide (4). Using the method described to obtain 1, from 2- (5-methoxy-1 / - indol-3-yl) propanoic acid (192 mg, 1 mmol) and propargilamine (0.71 ml, 1.1 mmol) 4 (160 mg, 65%) was obtained as a white solid. 1 H NMR (300 MHz, DMSO-d6) δ 10.57 (s, 1H), 8.29 (s, 1H), 7.18 (d, J = 8.7 Hz, 1H), 7.02 (s, 1H), 6.97 (s, 1H ), 6.68 (d, J = 8.6 Hz, 1 H), 3.90 - 3.80 (m, 2H), 3.74 (s, 3H), 3.09 - 3.07 (t, J = 2.7 Hz, 2H), 2.85 (t, J = 7.7 Hz, 2H), 2.42 (t, J = 6.8 Hz, 2H). 13 C NMR (75 MHz, Acetone-d6) 6173.16, 155.26, 133.31, 129.32, 124.06, 115.76, 113.29, 112.93, 101.79, 82.19, 72.44, 56.54, 37.93, 29.38, 22.53. HPLC-MS (m / z) [MH + ] 257.
2-(2-(1H-lndol-3-il)etil)-5-metiloxazol (5). A una solución de 3-(1 H-indol-3-il)- A/-(prop-2-inil)propanamida (377 mg, 1.67 mmol) en 20 mL de diclorometano seco se añadió cloruro de oro (III) (50 mg, 0.167 mmol) y la reacción se agitó a temperatura ambiente bajo una atmósfera inerte durante 18 h. Después de este tiempo, se añadió trietilamina (0.5 mL) y la mezcla resultante se filtró sobre una columna corta de gel de sílice eluyendo con acetato de etilo. El disolvente se evaporó a vacío y el aceite resultante se cromatografió sobre gel de sílice para dar 5 (23 mg, 7%).1H NMR (300 MHz, Metanol-d4) δ 10.14 (s, 1H), 7.45 (dt, J = 7.8, 1.0 Hz, 1H), 7.30 (dt, J = 8.1, 1.0 z, 1H), 7.07 (ddd, J = 8.2, 7.0, 1.3 Hz, 1 H), 7.04 - 6.91 (t, J = 7.8, 1 H; s, 1 H), 6.57 (s, 1 H), 3.12 (t, J = 8.1 z, 2H), 3.02 (t, J = 8.1 Hz, 2H), 2.19 (s, 3H).13C-NMR (75 MHz, Metanol-d4) δ 164.4, 148.6, 138.3, 127.0, 126.0, 123.8, 121.7, 120.0, 119.1, 113.5, 109.8, 117.3, 111.0, 31.8, 30.2, 22.5. HPLC-MS (m/z) [MH+] 227. 2-((1H-lndol-3-il)metil)-5-metiloxazol (6). A partir de 2-(1H-indol-3-il)-A/-(prop- 2-inil)acetamida (268 mg, 1.26 mmol), el compuesto 6 se obtuvo como un polvo blanco (76 mg, 29%), de acuerdo con la síntesis descrita para 5.1H NMR (300 MHz, DMSO-de) δ 10.94 (s, 1H), 7.46 (d, J = 7.8 Hz, 1H), 7.33 (d, J = 8.1 Hz, 1 H), 7.24 (d, J = 2.3 Hz, 1 H), 7.06 (t, J = 7.5 Hz, 1 H), 6.96 (t, J = 7.5 Hz, 1 H), 6.68 (s, 1H), 4.11 (s, 2H), 2.19 (s, 3H). HPLC-MS (m/z) [MH+] 241. 2- (2- (1H-lndol-3-yl) ethyl) -5-methylxazole (5). To a solution of 3- (1 H-indole-3-yl) - A / - (prop-2-inyl) propanamide (377 mg, 1.67 mmol) in 20 mL of dry dichloromethane was added gold chloride (III) ( 50 mg, 0.167 mmol) and the reaction was stirred at room temperature under an inert atmosphere for 18 h. After this time, triethylamine (0.5 mL) was added and the resulting mixture was filtered on a short column of silica gel eluting with ethyl acetate. The solvent was evaporated in vacuo and the resulting oil was chromatographed on silica gel to give 5 (23 mg, 7%). 1 H NMR (300 MHz, Methanol-d 4 ) δ 10.14 (s, 1H), 7.45 (dt, J = 7.8, 1.0 Hz, 1H), 7.30 (dt, J = 8.1, 1.0 z, 1H), 7.07 ( ddd, J = 8.2, 7.0, 1.3 Hz, 1 H), 7.04 - 6.91 (t, J = 7.8, 1 H; s, 1 H), 6.57 (s, 1 H), 3.12 (t, J = 8.1 z , 2H), 3.02 (t, J = 8.1 Hz, 2H), 2.19 (s, 3H). 13 C-NMR (75 MHz, Methanol-d 4 ) δ 164.4, 148.6, 138.3, 127.0, 126.0, 123.8, 121.7, 120.0, 119.1, 113.5, 109.8, 117.3, 111.0, 31.8, 30.2, 22.5. HPLC-MS (m / z) [MH + ] 227. 2 - ((1H-lndol-3-yl) methyl) -5-methylxazole (6). From 2- (1H-indole-3-yl) -A / - (prop-2-inyl) acetamide (268 mg, 1.26 mmol), compound 6 was obtained as a white powder (76 mg, 29%) , according to the synthesis described for 5. 1 H NMR (300 MHz, DMSO-de) δ 10.94 (s, 1H), 7.46 (d, J = 7.8 Hz, 1H), 7.33 (d, J = 8.1 Hz, 1 H), 7.24 (d, J = 2.3 Hz, 1 H), 7.06 (t, J = 7.5 Hz, 1 H), 6.96 (t, J = 7.5 Hz, 1 H), 6.68 (s, 1H), 4.11 (s, 2H), 2.19 (s, 3H). HPLC-MS (m / z) [MH + ] 241.
2-((5-Metoxi-1H-indol-3-il)metil)-5-metiloxazol (7). De acuerdo con la síntesis descrita para 5, a partir de 3 (100 mg, 0.41 mmol), el compuesto 7 se obtuvo como un sólido de color pálido (18 mg, 18%).1H NMR (300 MHz, CDCI3) δ 8.35 (s, 1H), 7.25 (d, J = 8.81H), 7.17 - 7.05 (m, 2H), 6.85 (dd, J= 8.7, 2.5 Hz, 1H), 6.66 (s, 1H), 4.21 (s, 2H), 3.85 (s, 3H), 2.26 (s, 3H).13C NMR (75 MHz, CDCI3) δ 161.52, 155.48, 144.90, 133.81, 128.38, 129.21, 124.04, 112.82, 112.72, 109.16, 106.68, 56.87, 28.43, 13.14. HPLC-MS (m/z) [MH+] 243. 2 - ((5-Methoxy-1H-indol-3-yl) methyl) -5-methylxazole (7). According to the synthesis described for 5, from 3 (100 mg, 0.41 mmol), compound 7 was obtained as a pale solid (18 mg, 18%). 1 H NMR (300 MHz, CDCI 3 ) δ 8.35 (s, 1H), 7.25 (d, J = 8.81H), 7.17 - 7.05 (m, 2H), 6.85 (dd, J = 8.7, 2.5 Hz, 1H) , 6.66 (s, 1H), 4.21 (s, 2H), 3.85 (s, 3H), 2.26 (s, 3H). 13 C NMR (75 MHz, CDCI 3 ) δ 161.52, 155.48, 144.90, 133.81, 128.38, 129.21, 124.04, 112.82, 112.72, 109.16, 106.68, 56.87, 28.43, 13.14. HPLC-MS (m / z) [MH + ] 243.
2-(2-(5-Metoxi-1H-indol-3-il)etil)-5-metiloxazol (8). De acuerdo con la síntesis descrita para 5, a partir de 4 (30 mg, 0.11 mmol) se obtuvo 8 como un sólido marrón claro (11 mg, 38%).1H NMR (300 MHz, CDCI3) δ 7.94 (s, 1H), 7.25 (d, J = 8.8 Hz, 1H), 7.03 (d, J = 2.3 Hz, 1H), 7.00 (s, 1H), 6.86 (dd, J = 8.8, 2.4 Hz, 1H), 6.64 (s, 1H), 3.87 (s, 3H), 3.24 - 3.17 (m, 2H), 3.11 (ddd, J = 9.4, 6.1, 2.0 Hz, 2H) 2.29 (s, 3H). 13C NMR (75 MHz, CDCI3) δ 166.32, 155.90, 145.49, 131.08, 127.71, 128.10, 123.63, 112.61, 111.72, 110.01, 105.79, 55.46, 28.28, 24.97, 12.43. HPLC-MS (m/z) [MH+] 257. 2- (2- (5-Methoxy-1H-indol-3-yl) ethyl) -5-methylxazole (8). According to the synthesis described for 5, from 4 (30 mg, 0.11 mmol) 8 was obtained as a light brown solid (11 mg, 38%). 1 H NMR (300 MHz, CDCI 3 ) δ 7.94 (s, 1H), 7.25 (d, J = 8.8 Hz, 1H), 7.03 (d, J = 2.3 Hz, 1H), 7.00 (s, 1H), 6.86 (dd, J = 8.8, 2.4 Hz, 1H), 6.64 (s, 1H), 3.87 (s, 3H), 3.24 - 3.17 (m, 2H), 3.11 (ddd, J = 9.4, 6.1, 2.0 Hz, 2H ) 2.29 (s, 3H). 13 C NMR (75 MHz, CDCI 3 ) δ 166.32, 155.90, 145.49, 131.08, 127.71, 128.10, 123.63, 112.61, 111.72, 110.01, 105.79, 55.46, 28.28, 24.97, 12.43. HPLC-MS (m / z) [MH + ] 257.
5-((5-Metoxi-1H-indol-3-il)metil)-3-metil-1,2,4-oxadiazol (9). 2-(5-Metoxi-1H- indol-3-il)acetato de etilo (150 mg, 0.64 mmol) es adicionado sobre una mezcla de acetamidoxima (150 mg, 0.64 mmol), hidruro sódico (61 mg, 2.5 mmol) y tamiz molecular (1.5 mg) en THF anhidro (5 mL) previamente agitada y calentada a 50°C durante 2 horas. Tras la adición la agitación se mantiene durante 2 horas a 80°C tras las cuales la mezcla se filtra y se evapora el disolvente. El crudo resultante se purifica por cromatografía para dar 9 (87 mg, 56%) como un sólido blanco. 1H NMR (300 MHz, CDCI3) δ 8.01 (s, 1H), 7.30 - 7.23 (m, 1H), 7.20 (d, J = 2.6 Hz, 1H), 7.05 (d, J = 2.4 Hz, 1H), 6.88 (dd, J = 8.8, 2.4 Hz, 1H), 4.31 (s, 2H), 3.86 (s, 2H), 2.37 (s, 3H). 13C NMR (75 MHz, CDCI3) δ 178.23, 167.24, 154.38, 131.23, 127.15, 123.74, 112.89, 112.06, 107.76, 100.33, 55.88, 23.26, 11.60. HPLC-MS (m/z) [MH+] 244. 5 - ((5-Methoxy-1H-indol-3-yl) methyl) -3-methyl-1,2,4-oxadiazole (9). 2- (5-Methoxy-1H-indole-3-yl) ethyl acetate (150 mg, 0.64 mmol) is added over a mixture of acetamidoxime (150 mg, 0.64 mmol), sodium hydride (61 mg, 2.5 mmol) and Molecular sieve (1.5 mg) in anhydrous THF (5 mL) previously stirred and heated at 50 ° C for 2 hours. After the addition the stirring is maintained for 2 hours at 80 ° C after which the mixture is filtered and the solvent is evaporated. The resulting crude is purified by chromatography to give 9 (87 mg, 56%) as a white solid. 1 H NMR (300 MHz, CDCI 3 ) δ 8.01 (s, 1H), 7.30 - 7.23 (m, 1H), 7.20 (d, J = 2.6 Hz, 1H), 7.05 (d, J = 2.4 Hz, 1H) , 6.88 (dd, J = 8.8, 2.4 Hz, 1H), 4.31 (s, 2H), 3.86 (s, 2H), 2.37 (s, 3H). 13 C NMR (75 MHz, CDCI 3 ) δ 178.23, 167.24, 154.38, 131.23, 127.15, 123.74, 112.89, 112.06, 107.76, 100.33, 55.88, 23.26, 11.60. HPLC-MS (m / z) [MH + ] 244.
5-(2-(5-Metoxi-1H-indol-3-il)etil)-3-metil-1,2,4-oxadiazol (10). Siguiendo un procedimiento semejante al anterior, a partir de3-(5-metoxi-1/-/-indol-3- il)propanato de etilo (173 mg, 0.7 mmol), se obtuvo 10 (80 mg, 45%) como un sólido blanco. 1H NMR (300 MHz, DMSO-d6) δ 10.64 (s, 1H), 7.20 (d, J = 8.7 Hz, 1 H), 7.07 (d, J = 2.2 Hz, 1 H), 6.97 (s, 1 H), 6.74 - 6.60 (m, 1 H), 3.74 (s, 3H), 3.25-3.18 (m, 2H), 3.15-3.08 (m, 2H), 2.28 (t, J= 1.2 Hz, 3H).13C NMR (101 MHz, DMSO-d6) δ 180.04, 167.35, 153.71, 131.95, 127.73, 123.90, 112.82, 112.73, 111.95, 100.54, 55.99, 27.40, 22.58, 11.80. HPLC-MS (m/z) [MH+] 258. 5- (2- (5-Methoxy-1H-indol-3-yl) ethyl) -3-methyl-1,2,4-oxadiazole (10). Following a procedure similar to the previous one, from ethyl 3- (5-methoxy-1 / - / - indol-3-) propanate (173 mg, 0.7 mmol), 10 (80 mg, 45%) was obtained as a white solid 1 H NMR (300 MHz, DMSO-d 6 ) δ 10.64 (s, 1H), 7.20 (d, J = 8.7 Hz, 1 H), 7.07 (d, J = 2.2 Hz, 1 H), 6.97 (s, 1 H), 6.74 - 6.60 (m, 1 H), 3.74 (s, 3H), 3.25-3.18 (m, 2H), 3.15-3.08 (m, 2H), 2.28 (t, J = 1.2 Hz, 3H) . 13 C NMR (101 MHz, DMSO-d6) δ 180.04, 167.35, 153.71, 131.95, 127.73, 123.90, 112.82, 112.73, 111.95, 100.54, 55.99, 27.40, 22.58, 11.80. HPLC-MS (m / z) [MH + ] 258.
2-(2-(5-Metoxi-1H-indol-3-il)etil)-5-metil-1,3,4-oxadiazol (11). Sobre una solución del ácido 3-(5-metoxi-1/-/-indol-3-il)propanoico (234 mg, 1 mmol) en 20 ml_ de acetonitrilo seco se añadieron 1 ,1'-carbonildiimidazol (CDI) (163 mg, 1 mmol) y DMAP (10 mg, 0.08 mmol). Después de agitar durante 1 hora a temperatura ambiente, se añadió acetilhidrazida (87 mg, 1.17 mmol) y la agitación se mantuvo 18 horas adicionales. Tras la eliminación del disolvente a vacío se obtuvo un crudo que fue tratado con HCI 1M (20 ml_) y la suspensión resultante fue extraída con acetato de etilo (3 x 20 ml_). Las capas orgánicas se juntaron, se lavaron con 2 M de NaOH (3 x 20 mi) y se secaron con sulfato de magnesio anhidro. Tras la eliminación del disolvente orgánico a presión reducida se obtuvo N-acetil-3-(5-metoxi-1H-indol-3-il)propanhidrazida (207 mg, 70%) como un sólido blanco, que se empleó en el siguiente paso sin purificar. 1H NMR (300 MHz, DMSO-d6) δ 10.60 (s, 1H), 9.75 (s, 1H), 9.73 (s, 1H), 7.19 (d, J = 8.7 Hz, 1 H), 7.08 (d, J = 2.3 Hz, 1 H), 6.98 (d, J = 2.5 Hz, 1 H), 6.69 (dd, J = 8.8, 2.4 Hz, 1H), 3.75 (s, 3H), 2.88 (t, J = 7.7 Hz, 2H), 2.52 - 2.37 (m, 2H), 1.83 (s, 3H). 13C NMR(101 MHz, DMSO-d6): δ (ppm) 171.4, 168.6, 153.6, 132.0, 127.9, 123.6, 113.9, 112.6, 111.7, 100.8, 56.0, 34.6, 21.4, 21.2. HPLC- MS (m/z) [MH+] 276. Sobre una solución de la anterior hidrazida intermedia (107 mg, 0.38 mmol) en acetonitrilo (2 mL) se añadió oxicloruro de fósforo (0.04 mL, 0.42 mmol) y la mezcla se calentó en un horno microondas a 180 °C durante 30 min. El disolvente se eliminó a vacío y el residuo se purificó por cromatografía en columna para dar 11 (30 mg, 30%) como un sólido blanco.1H NMR (300 MHz, CDCI3) δ 7.91 (s, 1 H), 7.26 (d, J = 8.8 Hz, 1 H), 7.03 (s, 1 H), 6.99 (d, J = 2.4 Hz, 1H), 6.87 (dd, J = 8.8, 2.4 Hz, 1H), 3.87 (s, 3H), 3.21 (m, J = 4.7 Hz, 4H), 2.48 (s, 3H). 13C NMR (75 MHz, CDCI3) δ 166.79, 163.64, 154.05, 131.37, 127.39, 122.40, 113.78, 112.34, 111.95, 100.28, 55.93, 26.17, 22.34, 10.90. HPLC-MS (m/z) [MH+] 258. 2- (2- (5-Methoxy-1H-indol-3-yl) ethyl) -5-methyl-1,3,4-oxadiazole (11). On a solution of 3- (5-methoxy-1 / - / - indol-3-yl) propanoic acid (234 mg, 1 mmol) in 20 ml_ of dry acetonitrile, 1,1'-carbonyldiimidazole (CDI) (163) mg, 1 mmol) and DMAP (10 mg, 0.08 mmol). After stirring for 1 hour at room temperature, acetylhydrazide (87 mg, 1.17 mmol) was added and stirring was maintained an additional 18 hours. After removal of the solvent in vacuo, a crude was obtained which was treated with 1M HCI (20 ml_) and the resulting suspension was extracted with ethyl acetate (3 x 20 ml_). The organic layers were combined, washed with 2 M NaOH (3 x 20 mL) and dried with anhydrous magnesium sulfate. After removal of the organic solvent under reduced pressure, N-acetyl-3- (5-methoxy-1H-indol-3-yl) propanhydrazide (207 mg, 70%) was obtained as a white solid, which was used in the next step without purifying. 1 H NMR (300 MHz, DMSO-d6) δ 10.60 (s, 1H), 9.75 (s, 1H), 9.73 (s, 1H), 7.19 (d, J = 8.7 Hz, 1 H), 7.08 (d, J = 2.3 Hz, 1 H), 6.98 (d, J = 2.5 Hz, 1 H), 6.69 (dd, J = 8.8, 2.4 Hz, 1H), 3.75 (s, 3H), 2.88 (t, J = 7.7 Hz, 2H), 2.52 - 2.37 (m, 2H), 1.83 (s, 3H). 13 C NMR (101 MHz, DMSO-d6): δ (ppm) 171.4, 168.6, 153.6, 132.0, 127.9, 123.6, 113.9, 112.6, 111.7, 100.8, 56.0, 34.6, 21.4, 21.2. HPLC-MS (m / z) [MH + ] 276. On a solution of the above intermediate hydrazide (107 mg, 0.38 mmol) in acetonitrile (2 mL) phosphorus oxychloride (0.04 mL, 0.42 mmol) was added and the mixture was heated in a microwave oven at 180 ° C for 30 min. The solvent was removed in vacuo and the residue was purified by column chromatography to give 11 (30 mg, 30%) as a white solid. 1 H NMR (300 MHz, CDCI 3 ) δ 7.91 (s, 1 H), 7.26 (d, J = 8.8 Hz, 1 H), 7.03 (s, 1 H), 6.99 (d, J = 2.4 Hz, 1H ), 6.87 (dd, J = 8.8, 2.4 Hz, 1H), 3.87 (s, 3H), 3.21 (m, J = 4.7 Hz, 4H), 2.48 (s, 3H). 13 C NMR (75 MHz, CDCI 3 ) δ 166.79, 163.64, 154.05, 131.37, 127.39, 122.40, 113.78, 112.34, 111.95, 100.28, 55.93, 26.17, 22.34, 10.90. HPLC-MS (m / z) [MH + ] 258.
5-((5-Metoxi-1H-indol-3-il)metil)-1,3,4-oxadiazol-2-ol (12). Una mezcla de 2- (5-metoxi-1H-indol-3-il)acetohidrazida (86 mg, 0,4 mmol), CDI (76 mg, 0.47 mmol) y trietilamina (0.11 mi, 0.8 mmol) en THF se calentó a 1000 C durante 15 min. Se evaporó el disolvente y el residuo se re-suspendió en una pequeña cantidad de agua (2 mi), que se puso a pH 12-13 con NaOH (conc.) y se filtró. El filtrado se recogió y se acidificó con HCI (conc), precipitando 12 como un sólido blanco nacarado, que se aisló por filtración (72 mg, 73%). 1H NMR (300 MHz, DMSO-d6) δ 12.12 - 12.01 (s, 1H), 10.90 - 10.84 (s, 1H), 7.28 - 7.25 (d, J = 6.1 Hz, 1 H), 7.25 - 7.24 (s, 1 H), 7.01 - 6.99 (d, J = 2.3 Hz, 1 H), 6.77 - 6.72 (dd, J = 8.8, 2.4 Hz, 1 H), 4.01 - 3.94 (s, 2H), 3.74 - 3.70 (s, 3H).13C NMR (75 MHz, DMSO-d6) δ 156.46, 155.07, 153.18, 131.24, 127.03, 124.78, 112.22, 111.27, 105.87, 100.11, 55.30, 22.54. HPLC-MS (m/z) [MH+] 246. 5 - ((5-Methoxy-1H-indol-3-yl) methyl) -1,3,4-oxadiazol-2-ol (12). A mixture of 2- (5-methoxy-1H-indole-3-yl) acetohydrazide (86 mg, 0.4 mmol), CDI (76 mg, 0.47 mmol) and triethylamine (0.11 mL, 0.8 mmol) in THF was heated at 100 0 C for 15 min. The solvent was evaporated and the residue was re-suspended in a small amount of water (2 ml), which was placed at pH 12-13 with NaOH (conc.) And filtered. The filtrate was collected and acidified with HCI (conc), precipitating 12 as a pearly white solid, which was isolated by filtration (72 mg, 73%). 1 H NMR (300 MHz, DMSO-d6) δ 12.12 - 12.01 (s, 1H), 10.90 - 10.84 (s, 1H), 7.28 - 7.25 (d, J = 6.1 Hz, 1 H), 7.25 - 7.24 (s , 1 H), 7.01 - 6.99 (d, J = 2.3 Hz, 1 H), 6.77 - 6.72 (dd, J = 8.8, 2.4 Hz, 1 H), 4.01 - 3.94 (s, 2H), 3.74 - 3.70 ( s, 3H). 13 C NMR (75 MHz, DMSO-d6) δ 156.46, 155.07, 153.18, 131.24, 127.03, 124.78, 112.22, 111.27, 105.87, 100.11, 55.30, 22.54. HPLC-MS (m / z) [MH + ] 246.
5-(2-(1H-lndol-3-il)etil)-1,3,4-oxadiazol-2-ol (13). Siguiendo el procedimiento del producto anterior, a partir de 3-(1/--indol-3-il)propanhidrazida (56 mg, 0.27 mmol) se obtuvo 13 (22 mg, 35%) como un sólido blanco nacarado. 1H NMR (300 MHz, CDCI3) δ 8.98 - 8.94 (s, 1 H), 8.07 - 8.00 (s, 1 H), 7.61 - 7.56 (d, J = 7.7 Hz, 1H), 7.39-7.35 (d, J= 8.7 Hz, 1H), 7.25-7.19 (t, J= 7.5 Hz, 1H), 7.17 - 7.11 (d, J = 8.7 Hz, 1 H), 7.04 - 7.02 (s, 1 H), 3.23 - 3.14 (t, J = 7.5 Hz, 2H), 3.00-2.93 (t, J=7.5 Hz, 2H).13C NMR (75 MHz, DMSO-d6) δ 158.14, 156.24, 136.52, 127.10, 124.35, 121.51, 118.94, 118.63, 111.87, 107.30, 27.38, 19.20. HPLC-MS (m/z) [MH+] 230. 5-((5-Metoxi-1H-indol-3-il)metilo)-1,3,4-oxadiazol-2-tiol (14). Una mezcla de 2-(5-metoxi-1H-indol-3-il)acetohidrazida (comercial) (83 mg, 0.37 mmol), KOH (70 mg, 1.24 mmol) y un exceso de disulfuro de carbono (1.35 ml_, 38 mmol) en EtOH se calentó a 155 °C durante 10 min bajo irradiación de microondas. El disolvente y el exceso de disulfuro de carbono se evaporaron bajo presión reducida y el residuo se resuspendió en agua. La solución se acidificó con HCI conc., precipitando 14 como un sólido amarillo que se separó por filtración (91 mg, 95%).1H NMR (300 MHz, DMSO-d6) δ 14.37 - 14.29 (s, 1H), 10.95- 10.88 (s, 1 H), 7.29 - 7.24 (d, J = 11.8 Hz, 1 H), 7,28—7.27 (s, 1 H), 7.02 - 6.99 (d, J = 2.3 Hz, 1H), 6.77 - 6.72 (dd, J = 8.8, 2.4 Hz, 1H), 4.18 - 4.15 (s, 2H), 3.74 - 3.72 (s, 3H). 13C NMR (75 MHz, DMSO-d6) δ 177.73, 163.44, 153.25, 131.23, 126.94, 125.01, 112.28, 111.38, 105.39, 100.08, 55.31, 21.74. HPLC-MS (m/z) [MH+] 262. 5- (2- (1H-lndol-3-yl) ethyl) -1,3,4-oxadiazol-2-ol (13). Following the procedure of the previous product, from 3- (1 / - indole-3-yl) propanhydrazide (56 mg, 0.27 mmol), 13 (22 mg, 35%) was obtained as a pearly white solid. 1 H NMR (300 MHz, CDCI3) δ 8.98 - 8.94 (s, 1 H), 8.07 - 8.00 (s, 1 H), 7.61 - 7.56 (d, J = 7.7 Hz, 1H), 7.39-7.35 (d, J = 8.7 Hz, 1H), 7.25-7.19 (t, J = 7.5 Hz, 1H), 7.17 - 7.11 (d, J = 8.7 Hz, 1 H), 7.04 - 7.02 (s, 1 H), 3.23 - 3.14 (t, J = 7.5 Hz, 2H), 3.00-2.93 (t, J = 7.5 Hz, 2H). 13 C NMR (75 MHz, DMSO-d6) δ 158.14, 156.24, 136.52, 127.10, 124.35, 121.51, 118.94, 118.63, 111.87, 107.30, 27.38, 19.20. HPLC-MS (m / z) [MH + ] 230. 5 - ((5-Methoxy-1H-indol-3-yl) methyl) -1,3,4-oxadiazol-2-thiol (14). A mixture of 2- (5-methoxy-1H-indole-3-yl) acetohydrazide (commercial) (83 mg, 0.37 mmol), KOH (70 mg, 1.24 mmol) and an excess of carbon disulfide (1.35 ml_, 38 mmol) in EtOH was heated at 155 ° C for 10 min under microwave irradiation. The solvent and excess carbon disulfide were evaporated under reduced pressure and the residue was resuspended in water. The solution was acidified with conc. HCI, precipitating 14 as a yellow solid that was filtered off (91 mg, 95%). 1 H NMR (300 MHz, DMSO-d6) δ 14.37 - 14.29 (s, 1H), 10.95-10.88 (s, 1 H), 7.29 - 7.24 (d, J = 11.8 Hz, 1 H), 7.28— 7.27 (s, 1 H), 7.02 - 6.99 (d, J = 2.3 Hz, 1H), 6.77 - 6.72 (dd, J = 8.8, 2.4 Hz, 1H), 4.18 - 4.15 (s, 2H), 3.74 - 3.72 (s, 3H). 13 C NMR (75 MHz, DMSO-d6) δ 177.73, 163.44, 153.25, 131.23, 126.94, 125.01, 112.28, 111.38, 105.39, 100.08, 55.31, 21.74. HPLC-MS (m / z) [MH + ] 262.
5-(2-(1H-lndol-3-il)etil)-1,3,4-oxadiazol-2-tiol (15). Siguiendo el procedimiento anterior, a partir de 3-(1/--indol-3-il)propanohidrazida (67 mg, 0.33 mmol) se obtuvo 15 (74 mg, 97%) como un sólido naranja. 1H NMR (300 MHz, DMSO- d6) δ 14.31 - 14.23 (s, 1H), 10.88 - 10.80 (s, 1H), 7.53 - 7.48 (d, J = 7.7 Hz, 1H), 7.35-7.30 (d, J = 7.4 Hz, 1H), 7.16-7.12 (d, J = 2.2 Hz, 1H), 7.10-7.02 (t, J = 7.5 Hz, 1 H), 7.00 - 6.93 (t, J = 7.4 Hz, 1 H), 3.08 - 3.07 (s, 4H).13C NMR (75 MHz, DMSO-d6)5177.76, 163.43, 136.15, 126.55, 124.46, 121.35, 118.82, 118.10, 111.63, 105.70, 26.9, 19.91. HPLC-MS (m/z) [MH+] 246. 5- (2- (1H-lndol-3-yl) ethyl) -1,3,4-oxadiazol-2-thiol (15). Following the above procedure, starting from 3- (1 / - indole-3-yl) propanohydrazide (67 mg, 0.33 mmol) 15 (74 mg, 97%) was obtained as an orange solid. 1 H NMR (300 MHz, DMSO-d6) δ 14.31 - 14.23 (s, 1H), 10.88 - 10.80 (s, 1H), 7.53 - 7.48 (d, J = 7.7 Hz, 1H), 7.35-7.30 (d, J = 7.4 Hz, 1H), 7.16-7.12 (d, J = 2.2 Hz, 1H), 7.10-7.02 (t, J = 7.5 Hz, 1 H), 7.00 - 6.93 (t, J = 7.4 Hz, 1 H ), 3.08-3.07 (s, 4H). 13 C NMR (75 MHz, DMSO-d6) 5177.76, 163.43, 136.15, 126.55, 124.46, 121.35, 118.82, 118.10, 111.63, 105.70, 26.9, 19.91. HPLC-MS (m / z) [MH + ] 246.
5-((5-Metoxi-1H-indol-3-il)metil)-4-metil-4H-1,2,4-triazol-3-ol (16). Sobre una solución de 2-(5-metoxi-1/--indol-3-il)acetohidrazida (comercial) (66 mg, 0.3 mmol) en una mezcla de agua (2.5 mL) y ácido acético (6 gotas) se añadió isocianato de metilo (0.60 mL, 0.97 mmol) y la mezcla se agitó durante 1.5 h a temperatura ambiente. El disolvente se evaporó a presión reducida y el residuo se disolvió en EtOH (3 mL), tratándose a continuación con NaOH 2M (2.25 mL). La mezcla se calienta bajo irradiación de microondas a 100 °C durante 15 min. Se evapora el disolvente y el crudo se suspendió en agua (1 mL), se llevó a pH 2 mediante la adición de HCI conc, precipitando 16 como un sólido blanco que se separó por filtración (58 mg, 75%). 1H NMR (300 MHz, DMSO-d6) δ 11.41 (s, 1H), 10.82 (s, 1H), 7.25 (d, J = 8.8 Hz, 1H), 7.20 (d, J = 2.3 Hz, 1H), 7.01 (d, J = 2.4 Hz, 1H), 6.74 (dd, J = 8.8, 2.4 Hz, 1H), 3.95 (s, 2H), 3.72 (s, 3H), 2.98 (s, 3H). 13C NMR (75 MHz, DMS0-d6) δ 155.65, 153.43, 147.20, 131.77, 127.50, 124.77, 112.51, 111.54, 107.40, 100.68, 55.65, 26.88, 22.86. HPLC-MS (m/z) [MH+] 259. 5 - ((5-Methoxy-1H-indole-3-yl) methyl) -4-methyl-4H-1,2,4-triazol-3-ol (16). On a solution of 2- (5-methoxy-1 / - indol-3-yl) acetohydrazide (commercial) (66 mg, 0.3 mmol) in a mixture of water (2.5 mL) and acetic acid (6 drops) was added methyl isocyanate (0.60 mL, 0.97 mmol) and the mixture was stirred for 1.5 h at room temperature. The solvent was evaporated under reduced pressure and the residue was dissolved in EtOH (3 mL), then treated with 2M NaOH (2.25 mL). The mixture is heated under microwave irradiation at 100 ° C for 15 min. The solvent is evaporated and the crude is suspended in water (1 mL), brought to pH 2 by the addition of conc HCI, precipitating 16 as a white solid that is filtered off (58 mg, 75%). 1 H NMR (300 MHz, DMSO-d6) δ 11.41 (s, 1H), 10.82 (s, 1H), 7.25 (d, J = 8.8 Hz, 1H), 7.20 (d, J = 2.3 Hz, 1H), 7.01 (d, J = 2.4 Hz, 1H), 6.74 (dd, J = 8.8, 2.4 Hz, 1H), 3.95 (s, 2H), 3.72 (s, 3H), 2.98 (s, 3H). 13 C NMR (75 MHz, DMS0-d6) δ 155.65, 153.43, 147.20, 131.77, 127.50, 124.77, 112.51, 111.54, 107.40, 100.68, 55.65, 26.88, 22.86. HPLC-MS (m / z) [MH + ] 259.
5-((1H-lndol-3-il)metil)-4-metil-4H-1,2,4-triazol-3-ol (17). A partir de 2-(1H- indol-3-il)acetohidrazida (19 mg, 0.1 mmol), de acuerdo con el procedimiento anterior, se obtuvo 17 como un sólido cristalino de color blanco (14 mg, 62%). 1H NMR (300 MHz, DMSO-d6) δ 11.39 (s, 1H), 10.97 (s, 1H), 7.49 (d, J = 7.9 Hz, 1H), 7.34 (dt, J = 8.1, 0.9 Hz, 1H), 7.25 (d, J = 2.4 Hz, 1H), 7.07 (ddd, J = 8.2, 7.1, 1.2 Hz, 1H), 6.96 (ddd, J= 8.0, 7.0, 1.1 Hz, 1H), 3.97 (s, 2H), 2.96 (s, 3H).13C NMR (126 MHz, DMSO-d6) δ 155.71, 147.25, 136.71, 127.21, 124.26, 121.65, 119.00, 118.77, 111.96, 107.82, 26.97, 22.94. HPLC-MS (m/z) [MH+] 229. 4-Etil-5-((5-metoxi-1H-indol-3-il)metil)-4H-1,2,4-triazol-3-tiol (18). Sobre una solución de 2-(5-metoxi-1/--indol-3-il)acetohidrazida (74 mg, 0.33 mmol) en tetrahidrofurano (THF, 10 mL) se añadió tioisocianato de etilo (0.035 mL, 0.4 mmol) y la mezcla se agitó durante 18 horas a temperatura ambiente. La aciltiosemicarbazida intermedia se separó mediante filtración (53 mg, 60%). Siguiendo el procedimiento anterior para la ciclación (ver síntesis de 16), a partir de 40 mg (0.13 mmol) de este intermedio se obtuvo 18 (30 mg, 80%) como un sólido amarillo. 1H NMR (300 MHz, DMSO-d6) δ 13.51 (s, 1H), 10.88 (s, 1H), 7.27 (d, J = 3.2 Hz, 1H), 7.25 (d, J = 3.2 Hz, 1H), 6.99 (d, J = 2.3 Hz, 1H), 6.74 (dd, J = 8.8, 2.4 Hz, 1H), 4.16 (s, 2H), 3.90 (q, J = 7.0 Hz, 2H), 3.72 (s, 3H), 0.94 (t, J = 7.1 Hz, 3H). 13C NMR (75 MHz, DMSO-d6) δ 166.63, 153.49, 151.61, 131.74, 127.43, 125.08, 112.60, 111.58, 107.29, 100.64, 55.65, 38.62, 22.22, 13.23. HPLC-MS (m/z) [MH+] 289. 5 - ((1H-lndol-3-yl) methyl) -4-methyl-4H-1,2,4-triazol-3-ol (17). From 2- (1H-indole-3-yl) acetohydrazide (19 mg, 0.1 mmol), according to the above procedure, 17 was obtained as a white crystalline solid (14 mg, 62%). 1 H NMR (300 MHz, DMSO-d6) δ 11.39 (s, 1H), 10.97 (s, 1H), 7.49 (d, J = 7.9 Hz, 1H), 7.34 (dt, J = 8.1, 0.9 Hz, 1H ), 7.25 (d, J = 2.4 Hz, 1H), 7.07 (ddd, J = 8.2, 7.1, 1.2 Hz, 1H), 6.96 (ddd, J = 8.0, 7.0, 1.1 Hz, 1H), 3.97 (s, 2H), 2.96 (s, 3H). 13 C NMR (126 MHz, DMSO-d6) δ 155.71, 147.25, 136.71, 127.21, 124.26, 121.65, 119.00, 118.77, 111.96, 107.82, 26.97, 22.94. HPLC-MS (m / z) [MH + ] 229. 4-Ethyl-5 - ((5-methoxy-1H-indol-3-yl) methyl) -4H-1,2,4-triazol-3-thiol (18). To a solution of 2- (5-methoxy-1 / - indol-3-yl) acetohydrazide (74 mg, 0.33 mmol) in tetrahydrofuran (THF, 10 mL) was added ethyl thioisocyanate (0.035 mL, 0.4 mmol) and The mixture was stirred for 18 hours at room temperature. The intermediate acylthiosemicarbazide was filtered off (53 mg, 60%). Following the above procedure for cyclization (see synthesis of 16), from 40 mg (0.13 mmol) of this intermediate, 18 (30 mg, 80%) was obtained as a yellow solid. 1 H NMR (300 MHz, DMSO-d6) δ 13.51 (s, 1H), 10.88 (s, 1H), 7.27 (d, J = 3.2 Hz, 1H), 7.25 (d, J = 3.2 Hz, 1H), 6.99 (d, J = 2.3 Hz, 1H), 6.74 (dd, J = 8.8, 2.4 Hz, 1H), 4.16 (s, 2H), 3.90 (q, J = 7.0 Hz, 2H), 3.72 (s, 3H ), 0.94 (t, J = 7.1 Hz, 3H). 13 C NMR (75 MHz, DMSO-d6) δ 166.63, 153.49, 151.61, 131.74, 127.43, 125.08, 112.60, 111.58, 107.29, 100.64, 55.65, 38.62, 22.22, 13.23. HPLC-MS (m / z) [MH + ] 289.
5-((1H-lndol-3-il)metil)-4-etil-4H-1,2,4-triazol-3-tiol (19). A partir de 2-(1H- indol-3-il)acetohidrazida (19 mg, 0.1 mmol), de acuerdo con el procedimiento descrito para 16, se obtuvo 19 como un sólido blanco perlado (16 mg, 62%).1H NMR (300 MHz, DMSO-d6) δ 1 1 .06 (s, 1 H), 7.47 (d, J = 7.7 Hz, 1 H), 7.35 (d, J = 8.1 Hz, 1 H), 7.31 (d, J = 2.3 Hz, 1 H), 7.07 (t, J = 7.5 Hz, 1 H), 6.95 (t, J = 7.5 Hz, 1 H), 4.18 (s, 2H), 3.89 (q, J = 7.0 Hz, 2H), 0.90 (t, J = 7.1 Hz, 3H). 13C NMR (126 MHz, DMSO-de) δ 166.65, 151 .68, 136.68, 127.12, 124.60, 121 .73, 1 19.13, 1 18.71 , 1 12.04, 107.67, 38.71 , 22.33, 13.31 . HPLC-MS (m/z) [MH+] 259. 5 - ((1H-lndol-3-yl) methyl) -4-ethyl-4H-1,2,4-triazol-3-thiol (19). From 2- (1H-indole-3-yl) acetohydrazide (19 mg, 0.1 mmol), according to the procedure described for 16, 19 was obtained as a pearl white solid (16 mg, 62%). 1 h NMR (300 MHz, DMSO-d6) δ 1 1 .06 (s, 1 H), 7.47 (d, J = 7.7 Hz, 1 H), 7.35 (d, J = 8.1 Hz, 1 H), 7.31 (d , J = 2.3 Hz, 1 H), 7.07 (t, J = 7.5 Hz, 1 H), 6.95 (t, J = 7.5 Hz, 1 H), 4.18 (s, 2H), 3.89 (q, J = 7.0 Hz, 2H), 0.90 (t, J = 7.1 Hz, 3H). 13 C NMR (126 MHz, DMSO-de) δ 166.65, 151 .68, 136.68, 127.12, 124.60, 121 .73, 1 19.13, 1 18.71, 1 12.04, 107.67, 38.71, 22.33, 13.31. HPLC-MS (m / z) [MH + ] 259.
2-((5-Metoxi-1H-indol-3-il)metil)-5-metil-1 ,3,4-oxadiazol (20). De acuerdo al procedimiento general para la preparación de 1 ,3,4-oxadiazoles, a partir de 2- (5-metoxi-1 /- -indol-3-il) acetohidrazida (100 mg, 0.45 mmol), ortoacetato de trietilo (1 ml_) y una cantidad catalítica de ácido acético, se obtuvo 20 (71 mg, 64%) como un sólido de mp: 139 - 140°C. 1H NMR (300 MHz, CDCI3) δ 8.33 (s, 1 H), 7.25 (d, J = 8.8 Hz, 1 H), 7.14 (s, 1 H), 7.06 (d, J = 2.2 Hz, 1 H), 6.86 (dd, J = 8.8, 2.4 Hz, 1 H), 4.27 (s, 2H), 3.84 (s, 3H), 2.44 (s, 3H). 13C NMR (75 MHz, CDCI3) δ 165.95, 163.93, 154.25, 131 .26, 127.15, 123.78, 1 12.75, 1 12.09, 107.86, 100.24, 55.85, 22.09, 10.94. HRMS (ESI+): m/z caled para C13H13N3O2 (M)+ 243.1008, encontrado: 243.1017. Pureza por HPLC 100% (230 - 400 nm) 2 - ((5-Methoxy-1H-indol-3-yl) methyl) -5-methyl-1, 3,4-oxadiazole (20). According to the general procedure for the preparation of 1, 3,4-oxadiazoles, from 2- (5-methoxy-1 / - -indole-3-yl) acetohydrazide (100 mg, 0.45 mmol), triethyl orthoacetate ( 1 ml_) and a catalytic amount of acetic acid, 20 (71 mg, 64%) was obtained as a solid of mp: 139-140 ° C. 1 H NMR (300 MHz, CDCI 3 ) δ 8.33 (s, 1 H), 7.25 (d, J = 8.8 Hz, 1 H), 7.14 (s, 1 H), 7.06 (d, J = 2.2 Hz, 1 H), 6.86 (dd, J = 8.8, 2.4 Hz, 1 H), 4.27 (s, 2H), 3.84 (s, 3H), 2.44 (s, 3H). 13 C NMR (75 MHz, CDCI 3 ) δ 165.95, 163.93, 154.25, 131 .26, 127.15, 123.78, 1 12.75, 1 12.09, 107.86, 100.24, 55.85, 22.09, 10.94. HRMS (ESI + ): m / z caled for C13H13N3O2 (M) + 243.1008, found: 243.1017. 100% HPLC purity (230-400 nm)
5-(2-(5-Metoxi-1H-indol-3-il)etil)-1 ,3,4-oxadiazol-2-ol (21). Siguiendo el procedimiento seguido para la obtención del compuesto 12, a partir de (5- metoxi-1 /- -indol-3-il) propanhidrazida (146 mg, 0.62 mmol) se obtuvo 22 (138 mg, 86%)como un sólido blanco de mp: 140 - 144°C. 1H NMR (400 MHz, DMSO) δ 12.00 (s, 1 H), 10.66 (s, 1 H), 7.20 (d, J = 8.7 Hz, 1 H), 7.09 (d, J = 2.3 Hz, 1 H), 6.95 (d, J = 2.4 Hz, 1 H), 6.69 (dd, J = 8.7, 2.4 Hz, 1 H), 3.74 (s, 3H), 2.97 (t, J = 7.4 Hz, 2H), 2.85 (t, J = 7.7 Hz, 2H). 13C NMR (101 MHz, DMSO) δ 157.65, 155.77, 153.70, 131 .92, 127.77, 124.00, 1 12.76, 1 12.72, 1 1 1 .93, 100.36, 55.92, 27.55, 21 .54 HRMS (ESI+): m/z caled para C13H13N3O3 (M)+ 259.0957, encontrado 259.0966. Pureza por HPLC 100% (230 - 400 nm). 5- (2- (5-Methoxy-1H-indol-3-yl) ethyl) -1, 3,4-oxadiazol-2-ol (21). Following the procedure followed to obtain compound 12, from (5- methoxy-1 / - -indole-3-yl) propanhydrazide (146 mg, 0.62 mmol) 22 (138 mg, 86%) was obtained as a solid mp white: 140-144 ° C. 1 H NMR (400 MHz, DMSO) δ 12.00 (s, 1 H), 10.66 (s, 1 H), 7.20 (d, J = 8.7 Hz, 1 H), 7.09 (d, J = 2.3 Hz, 1 H ), 6.95 (d, J = 2.4 Hz, 1 H), 6.69 (dd, J = 8.7, 2.4 Hz, 1 H), 3.74 (s, 3H), 2.97 (t, J = 7.4 Hz, 2H), 2.85 (t, J = 7.7 Hz, 2H). 13 C NMR (101 MHz, DMSO) δ 157.65, 155.77, 153.70, 131 .92, 127.77, 124.00, 1 12.76, 1 12.72, 1 1 1 .93, 100.36, 55.92, 27.55, 21 .54 HRMS (ESI + ) : m / z caled for C13H13N3O3 (M) + 259.0957, found 259.0966. 100% HPLC purity (230-400 nm).
5-(2-(5-Metoxi-1H-indol-3-il)etil)-1 ,3,4-oxadiazol-2-tiol (22). Siguiendo el procedimiento seguido para la obtención del compuesto 12, a partir de (5- metoxi-1 /- -indol-3-il) propanhidrazida (1 16 mg, 0.5 mmol) se obtuvo 23 (56 mg, 41 %) como un sólido blanco de mp: 99 - 101 °C. 1H NMR (400 MHz, DMSO) δ 14.29 (s, 1H), 10.70 (d, J = 2.6 Hz, 1H), 7.21 (d, J = 8.7 Hz, 1H), 7.11 (d, J = 2.4 Hz, 1H), 6.97 (d, J = 2.4 Hz, 1H), 6.70 (dd, J = 8.7, 2.4 Hz, 1H), 3.76 (s, 3H), 3.10 - 2.97 (m, 4H). 13C NMR (101 MHz, DMSO) δ 177.69, 164.05, 153.08, 131.25, 127.06, 123.38, 112.12, 111.84, 111.37, 99.70, 55.31, 26.14, 21.05. HRMS (ESI+): m/z caled para C13H13N3O2S (M)+ 275.0728, encontrado 275.0726. Pureza por HPLC 95% (230 - 400 nm). 5- (2- (5-Methoxy-1H-indol-3-yl) ethyl) -1, 3,4-oxadiazol-2-thiol (22). Following the procedure followed to obtain compound 12, starting from (5- methoxy-1 / - -indole-3-yl) propanhydrazide (1 16 mg, 0.5 mmol) 23 (56 mg, 41%) was obtained as a mp white solid: 99 - 101 ° C. 1 H NMR (400 MHz, DMSO) δ 14.29 (s, 1H), 10.70 (d, J = 2.6 Hz, 1H), 7.21 (d, J = 8.7 Hz, 1H), 7.11 (d, J = 2.4 Hz, 1H), 6.97 (d, J = 2.4 Hz, 1H), 6.70 (dd, J = 8.7, 2.4 Hz, 1H), 3.76 (s, 3H), 3.10 - 2.97 (m, 4H). 13 C NMR (101 MHz, DMSO) δ 177.69, 164.05, 153.08, 131.25, 127.06, 123.38, 112.12, 111.84, 111.37, 99.70, 55.31, 26.14, 21.05. HRMS (ESI + ): m / z caled for C13H13N3O2S (M) + 275.0728, found 275.0726. HPLC purity 95% (230-400 nm).
5-(2-(5-Metoxi-1H-indol-3-il)etil)-4-metil-4H-1,2,4-triazol-3-ol (23). Siguiendo el procedimiento para la obtención del compuesto 16, a partir de (5-metoxi-1/-- indol-3-il) propanhidrazida (230 mg, 0.98 mmol) se obtuvo 24 (110 mg, 41%) como un sólido blanco de mp 201 - 202 °C.1H NMR (500 MHz, DMSO) δ 11.38 (s, 1H), 10.67 (d, J= 3.0 Hz, 1H), 7.22 (d, J= 8.7 Hz, 1H), 7.12 (d, J = 2.4 Hz, 1H), 6.94 (d, J = 2.4 Hz, 1H), 6.70 (dd, J = 8.7, 2.4 Hz, 1H), 3.75 (s, 3H), 3.01 (s, 3H), 2.98 (t, 2H), 2.82 (t, 2H). 13C NMR (126 MHz, DMSO-d6) δ 155.65, 153.39, 148.13, 131.70, 127.66, 123.70, 113.24, 112.45, 111.60, 100.26, 55.68, 26.75, 26.73, 21.74. HRMS (ESI+): Caled. Para C14H16N4O2 (M)+ 272.1283, encontrado m/z 272.1273. Pureza por HPLC 100% (230-400 nm). 5- (2- (5-Methoxy-1H-indole-3-yl) ethyl) -4-methyl-4H-1,2,4-triazol-3-ol (23). Following the procedure for obtaining compound 16, from (5-methoxy-1 / - indol-3-yl) propanhydrazide (230 mg, 0.98 mmol) 24 (110 mg, 41%) was obtained as a white solid mp 201-202 ° C. 1 H NMR (500 MHz, DMSO) δ 11.38 (s, 1H), 10.67 (d, J = 3.0 Hz, 1H), 7.22 (d, J = 8.7 Hz, 1H), 7.12 (d, J = 2.4 Hz, 1H), 6.94 (d, J = 2.4 Hz, 1H), 6.70 (dd, J = 8.7, 2.4 Hz, 1H), 3.75 (s, 3H), 3.01 (s, 3H), 2.98 (t, 2H), 2.82 (t, 2H). 13 C NMR (126 MHz, DMSO-d 6 ) δ 155.65, 153.39, 148.13, 131.70, 127.66, 123.70, 113.24, 112.45, 111.60, 100.26, 55.68, 26.75, 26.73, 21.74. HRMS (ESI + ): Caled. For C 14 H 16 N 4 O 2 (M) + 272.1283, found m / z 272.1273. 100% HPLC purity (230-400 nm).
4-Etil-5-(2-(5-metoxi-1H-indol-3-il)etil)-4H-1,2,4-triazol-3-tiol (24). Siguiendo el procedimiento empleado para la obtención del compuesto 18, a partir de (5- metoxi-1/--indol-3-il) propanhidrazida se obtuvo 25 (140 mg, 84%) como un sólido blanco de mp: 214 -217°C.1H NMR (500 MHz, DMSO) δ 13.52 (s, 1H,), 10.68 (ó,J = 2.7 Hz, 1H), 7.22 (d, J=8.7 Hz, 1 H), 7.14 (d, J = 2.4 Hz, 1H), 6.96 (d, J = 2.4 Hz, 1 H), 6.70 (dd, J = 8.7, 2.4 Hz, 1 H), 3.91 (q, J = 7.2 Hz, 2H), 3.75 (s, 3H), 3.06 - 3.02 (m, 2H), 3.02 - 2.97 (m, 2H), 1.14 (t, J = 7.2 Hz, 3H).13C NMR (126 MHz, DMSO) δ 166.26, 153.42, 152.45, 131.69, 127.60, 123.77, 113.04, 112.48, 111.66, 100.24, 55.69, 38.40, 26.15, 22.05, 13.77. HRMS (ESI+): Caled para C15H18N4OS (M)+ 302.1201, encontrado m/z 302.1207. HPLC-MS (230 - 400 nm) 98% (m/z) (M+) 303.28. 2. Evaluación biológica 4-Ethyl-5- (2- (5-methoxy-1H-indole-3-yl) ethyl) -4H-1,2,4-triazol-3-thiol (24). Following the procedure used to obtain compound 18, from (5- methoxy-1 / - indole-3-yl) propanhydrazide 25 (140 mg, 84%) was obtained as a white solid of mp: 214-217 ° C. 1 H NMR (500 MHz, DMSO) δ 13.52 (s, 1H,), 10.68 (or, J = 2.7 Hz, 1H), 7.22 (d, J = 8.7 Hz, 1 H), 7.14 (d, J = 2.4 Hz, 1H), 6.96 (d, J = 2.4 Hz, 1 H), 6.70 (dd, J = 8.7, 2.4 Hz, 1 H), 3.91 (q, J = 7.2 Hz, 2H), 3.75 (s, 3H ), 3.06 - 3.02 (m, 2H), 3.02 - 2.97 (m, 2H), 1.14 (t, J = 7.2 Hz, 3H). 13 C NMR (126 MHz, DMSO) δ 166.26, 153.42, 152.45, 131.69, 127.60, 123.77, 113.04, 112.48, 111.66, 100.24, 55.69, 38.40, 26.15, 22.05, 13.77. HRMS (ESI + ): Caled for C 15 H 18 N 4 OS (M) + 302.1201, found m / z 302.1207. HPLC-MS (230-400 nm) 98% (m / z) (M + ) 303.28. 2. Biological evaluation
2.1. Ensayos de unión a receptores de melatonina ΜΊΊ y T2 y estudio de la actividad intrínseca Los productos de la invención se evaluaron frente a receptores humanos de melatonina de tipo ΜΤΊ y MT2 transfectados en líneas celulares HEK y CHO, empleando 2-[125l]yodomelatonina como radioligando y siguiendo protocolos descritos (Ettaoussi, M.; Sabaouni, A.; Rami, M.; Boutin, J. A.; Delagrange, P.; Renard, P.; Spedding, M.; Caignard, D.-H.; Berthelot, P.; Yous, S. Design, synthesis and pharmacological evaluation of new series of naphthalenic analogues as melatoninergic (MT1/MT2) and serotoninergic 5-HT2C dual ligands (I). Eur. J. Med. Chem. 2012, 49, 310-323; Audinot, V.; Mailliet, F.; Lahaye-Brasseur, C; Bonnaud, A.; Le Gall, A.; Amosse, C; Dromaint, S.; Rodríguez, M.; Nagel, N.; Galizzi, J. P.; Malpaux, B.; Guillaumet, G.; Lesieur, D.; Lefoulon, F.; Renard, P.; Delagrange, P.; Boutin, J. A. New selective ligands of human cloned melatonin MT1 and MT2 receptors. Naunyn-Schmiedeberg's Arch. Pharmacol. 2003, 367, 553-561 ). 2.1. Melatonin ΜΊΊ and T 2 receptor binding assays and study of intrinsic activity The products of the invention were evaluated against human la and MT2 type melatonin receptors transfected into HEK and CHO cell lines, using 2- [ 125 l] iodomelatonin as radioligand and following described protocols (Ettaoussi, M .; Sabaouni, A .; Rami, M .; Boutin, JA; Delagrange, P .; Renard, P .; Spedding, M .; Caignard, D.-H .; Berthelot, P .; Yous, S. Design, synthesis and pharmacological evaluation of new series of naphthalenic analogues as melatoninergic (MT1 / MT2) and serotoninergic 5-HT2C dual ligands (I). Eur. J. Med. Chem. 2012, 49 , 310-323; Audinot, V .; Mailliet, F .; Lahaye-Brasseur, C; Bonnaud, A .; Le Gall, A .; Amosse, C; Dromaint, S .; Rodríguez, M .; Nagel, N. ; Galizzi, JP; Malpaux, B .; Guillaumet, G .; Lesieur, D .; Lefoulon, F .; Renard, P .; Delagrange, P .; Boutin, JA New selective ligands of human cloned melatonin MT1 and MT2 receptors. Naunyn-Schmiedeberg's Arch. Pharmacol 2003, 367, 553-561).
Las líneas celulares HEK y CHO con expresión estable de receptores humanos de melatonina de tipo ΜΤΊ y MT2 fueron cultivadas en medio DMEM (Dulbecco's modified Eagle's médium) suplementado con 10% de suero fetal bovino, 2 mM de glutamina, 100 Ul / mL de penicilina y 100 mg / mL de estreptomicina. Las células se cultivaron hasta confluencia a 37 °C (95% O2 / 5% de CO2), se suspendieron en PBS que contenía 2 mM de EDTA y se centrifugaron a 1000 x g durante 5 min a 4 °C. El sedimento resultante se suspendió en tampón Tris 5 mM (pH 7.5), que contenía 2 mM de EDTA, y se homogeneizó. A continuación, el homogeneizado se centrifugó (95.000 x g, 30 min, 4 °C) y el sedimento resultante se suspendió en tampón Tris 75 mM (pH 7.5) con 12.5 mM de MgCI2 y 2 mM de EDTA. Las alícuotas de las preparaciones de membrana se almacenaron a -80 °C hasta su uso. Los ensayos de unión se realizaron mediante la adición de las preparaciones de membrana de células transfectadas de tipo HEK o CHO diluidas en tampón Tris-HCI (50 mM, pH 7.4, con 5 mM de MgCI2) a 2-[125l]yodomelatonina (25 o 200 pM para receptores ΜΤΊ y MT2, respectivamente, expresados en células HEK o 20 pM para receptores ΜΤΊ y MT2 expresados en células CHO) y el compuesto a ensayar. La unión no específica se definió en presencia de 1 μΜ de melatonina. Después de 120 min de incubación a 37 °C, la reacción se detuvo por filtración rápida a través de filtros GF/B previamente empapados con una solución de polietilenimina al 0.5% (v / v). Los filtros se lavaron tres veces con 1 mL de tampón Tris-HCI (50 mM, pH 7.4) enfriado con hielo. Los datos de las curvas dosis-respuesta (siete concentraciones en duplicado) se analizaron utilizando el programa PRISM (Graph Pad Software Inc., San Diego, CA) para calcular la IC5o (concentración inhibitoria 50). Los resultados se encuentran en la tabla 1 y se expresan como K¡ = CI5o / 1 + ([L] / KD), donde [L] es la concentración de radioligando usado en el ensayo y KD, la constante de disociación del radioligando. HEK and CHO cell lines with stable expression of human la and MT2 type melatonin receptors were cultured in DMEM medium (Dulbecco's modified Eagle's medium) supplemented with 10% fetal bovine serum, 2 mM glutamine, 100 Ul / mL penicillin and 100 mg / mL streptomycin. The cells were grown to confluence at 37 ° C (95% O2 / 5% CO2), suspended in PBS containing 2 mM EDTA and centrifuged at 1000 xg for 5 min at 4 ° C. The resulting sediment was suspended in 5 mM Tris buffer (pH 7.5), containing 2 mM EDTA, and homogenized. The homogenate was then centrifuged (95,000 xg, 30 min, 4 ° C) and the resulting sediment was suspended in 75 mM Tris buffer (pH 7.5) with 12.5 mM MgCl 2 and 2 mM EDTA. Aliquots of the membrane preparations were stored at -80 ° C until use. Binding assays were performed by adding membrane preparations of transfected cells of the HEK or CHO type diluted in Tris-HCI buffer (50 mM, pH 7.4, with 5 mM MgCl 2 ) at 2- [ 125 L] iodomelatonin (25 or 200 pM for ΜΤΊ and MT 2 receptors, respectively, expressed in HEK cells or 20 pM for ΜΤΊ and MT 2 receptors expressed in CHO cells) and the compound to be tested. Non-specific binding was defined in the presence of 1 μΜ of melatonin. After 120 min incubation at 37 ° C, the reaction was stopped by rapid filtration through GF / B filters previously soaked with a 0.5% (v / v) polyethyleneimine solution. The filters were washed three times with 1 mL of Tris-HCI buffer (50 mM, pH 7.4) cooled with ice. Data from the dose-response curves (seven concentrations in duplicate) were analyzed using the PRISM program (Graph Pad Software Inc., San Diego, CA) to calculate the IC 5 or (50 inhibitory concentration). The results are found in Table 1 and are expressed as K¡ = CI 5 or / 1 + ([L] / K D ), where [L] is the concentration of radioligand used in the test and KD, the dissociation constant of radioligand.
Los ensayos de actividad intrínseca se realizaron empleando [35S] GTPyS (5'- O-[gamma-tio]trifosfato de guanosina). Las preparaciones de membranas de células CHO transfectadas que expresan los subtipos ΜΤΊ y MT2, junto con el compuesto ensayado, fueron diluidas en tampón HEPES (20mM, pH 7.4, 100 mM de NaCI, 3 μΜ de GDP, 3 mM de MgCI2, y 20 μg/mL de saponina). Para medir la actividad agonista, sobre la preparación conteniendo las membranas (20 μg / mL) y el producto a evaluar se añadió una solución 0.2 nM de [35S] GTPyS y el conjunto se incubó durante 1 hora a temperatura ambiente. Para evaluar la actividad antagonista, las membranas se incubaron previamente tanto con melatonina (3 nM) como con el producto a ensayar durante 30 min antes de la adición de [35S] GTPyS. La unión no específica se definió utilizando GTPyS frío (10 μΜ). Todas las reacciones se detuvieron mediante filtración rápida a través de filtros GF / B, seguido de tres lavados sucesivos con tampón enfriado con hielo. Los niveles habituales de [35S] GTPyS (expresados en dpm) en la unión con las membranas CHO-MT2 fueron: 2000 para la actividad basal, 8000 en presencia de melatonina 1 μΜ y 180 en la presencia de 10 mM GTPyS que define la unión no específica. Los datos de la curvas dosis-respuesta (siete concentraciones por duplicado) fueron analizados utilizando el programa PRISM (Graph Pad Software Inc., San Diego, CA) para calcular la EC5o (concentración efectiva al 50%) y £max (efecto máximo, expresado como el porcentaje con relación al observado con 1 μΜ de melatonina definido como 100%) para los agonistas. La potencia de los antagonistas se expresaron como KB = IC5o / 1 + ([Ago] / EC50 ago), donde IC50 es la concentración de antagonista que proporciona el 50% de inhibición de la unión de [35S] GTPyS en presencia de una concentración fija de la melatonina ([Ago]) y EC5o ago es la EC50 del agonista cuando se probó solo. El valor Elmax (efecto inhibidor máximo) se expresó como el porcentaje del efecto observado con melatonina, 30 nM o 3nM ([ago]) para los receptores hMTi y hMT2, respectivamente. Cuando Elmax es superior al 80%, el producto es un agonista; cuando es inferior al 20%, antagonista; y entre estas dos cifras, agonista parcial. Los resultados de la actividad intrínseca de una selección de los productos de la invención se encuentran recogidos en la tabla 2. Intrinsic activity assays were performed using [ 35 S] GTPyS (5'-O- [gamma-thio] guanosine triphosphate). Membrane preparations of transfected CHO cells expressing the ΜΤΊ and MT 2 subtypes, together with the compound tested, were diluted in HEPES buffer (20mM, pH 7.4, 100 mM NaCI, 3 μΜ GDP, 3 mM MgCl 2 , and 20 μg / mL of saponin). To measure the agonist activity, a 0.2 nM solution of [ 35 S] GTPyS was added to the preparation containing the membranes (20 μg / mL) and the product to be evaluated and the whole was incubated for 1 hour at room temperature. To evaluate the antagonistic activity, the membranes were previously incubated with both melatonin (3 nM) and the product to be tested for 30 min before the addition of [ 35 S] GTPyS. Non-specific binding was defined using cold GTPyS (10 μΜ). All reactions were stopped by rapid filtration through GF / B filters, followed by three successive washes with ice-cold buffer. The usual levels of [ 35 S] GTPyS (expressed in dpm) at the junction with the CHO-MT 2 membranes were: 2000 for baseline activity, 8000 in the presence of 1 μΜ melatonin and 180 in the presence of 10 mM GTPyS defining non-specific binding. Data from the dose-response curves (seven concentrations in duplicate) were analyzed using the PRISM program (Graph Pad Software Inc., San Diego, CA) to calculate EC 5 or (50% effective concentration) and £ max (effect maximum, expressed as the percentage in relation to that observed with 1 μΜ of melatonin defined as 100%) for agonists. The potency of the antagonists were expressed as K B = IC 5 or / 1 + ([Aug] / EC50 ago), where IC50 is the concentration of antagonist that provides 50% inhibition of the binding of [ 35 S] GTPyS in Presence of a fixed concentration of melatonin ([Aug]) and EC 5 or Aug is the EC 5 0 of the agonist when tested alone. The value The max (maximum inhibitory effect) was expressed as the percentage of the effect observed with melatonin, 30 nM or 3nM ([ago]) for the hMTi and hMT2 receptors, respectively. When Max is greater than 80%, the product is an agonist; when it is less than 20%, antagonist; and between these two figures, partial agonist. The results of the intrinsic activity of a selection of the products of the invention are shown in Table 2.
Tabla 1. Datos de unión (binding) a receptores de melatonina ΜΤΊ y MT2 ( ¡, molar) de los productos de la invención. Table 1. Binding data to melatonin ΜΤΊ and MT2 (¡, molar) receptors of the products of the invention.
Comp. MTi MT2 Comp. MTi MT 2
1 1 .7 10"8 4.9 10"9 1 1 .7 10 "8 4.9 10 " 9
2 2.5 10"9 4.5 10"1C 2 2.5 10 "9 4.5 10 " 1C
3 1 .4 10"8 3.8 10"9 3 1 .4 10 "8 3.8 10 " 9
4 5.0 10"9 1 .0 10"9 4 5.0 10 "9 1 .0 10 " 9
5 > 10"5 > 10"5 5> 10 "5 > 10 " 5
6 > 10"5 > 10"5 6> 10 "5 > 10 " 5
7 > 10"5 2.8 10"7 7> 10 "5 2.8 10 " 7
8 > 10"5 5.0 10"7 9 1 .7 10"7 4.4 10"8 8> 10 "5 5.0 10 " 7 9 1 .7 10 "7 4.4 10 " 8
10 > 10"5 > 10"5 10> 10 "5 > 10 " 5
11 7.0 10"7 1 .9 10"7 11 7.0 10 "7 1 .9 10 " 7
12 > 10"5 1 .6 10"7 12> 10 "5 1 .6 10 " 7
13 > 10"5 > 10"5 13> 10 "5 > 10 " 5
14 > 10"5 5.3 10"7 14> 10 "5 5.3 10 " 7
15 > 10"5 > 10"5 15> 10 "5 > 10 " 5
16 > 10"5 5.6 10"7 16> 10 "5 5.6 10 " 7
17 > 10"5 > 10"5 17> 10 "5 > 10 " 5
18 > 10"5 2.1 10"7 18> 10 "5 2.1 10 " 7
19 > 10"5 > 10"5 19> 10 "5 > 10 " 5
20 2.1 10"7 1 .6 10"8 20 2.1 10 "7 1 .6 10 " 8
21 3.5 10"8 4.0 10"9 21 3.5 10 "8 4.0 10 " 9
22 > 10"5 5.3 10"7 22> 10 "5 5.3 10 " 7
23 > 10"5 > 10"5 23> 10 "5 > 10 " 5
24 > 10"5 > 10"5 24> 10 "5 > 10 " 5
Tabla 2. Actividad intrínseca de los productos de la invención sobre los Table 2. Intrinsic activity of the products of the invention on the
receptores de melatonina MTi y MT2  MTi and MT2 melatonin receptors
MTi MT2 MTi MT 2
Comp EC50 (M) E lmax (%) Carácter EC50 (M) Elmax (%) Carácter Comp EC 50 (M) E lmax (%) Character EC 50 (M) Elmax (%) Character
Agonista AgonistaAgonist Agonist
1 5.3 10"7 64 1 .3 10"7 70 1 5.3 10 "7 64 1 .3 10 " 7 70
parcial parcial  partial partial
2 2.0 10"7 92 Agonista 1 .1 10"8 88 Agonista 2 2.0 10 "7 92 Agonist 1 .1 10 " 8 88 Agonist
Agonista Agonist
3 3.0 10"7 67 8.0 10"8 82 Agonista parcial 3 3.0 10 "7 67 8.0 10 " 8 82 Partial Agonist
4 4.0 10"8 100 Agonista 3.0 10"9 99 Agonista 4 4.0 10 "8 100 Agonist 3.0 10 " 9 99 Agonist
AgonistaAgonist
7 nd nd nd 9.9 10"7 23 7 nd nd nd 9.9 10 "7 23
parcial partial
AgonistaAgonist
8 nd nd nd 5.0 10"7 32 8 nd nd nd 5.0 10 "7 32
parcial partial
AgonistaAgonist
9 7.1 10"7 16 Antagonista 3.0 10"7 48 9 7.1 10 "7 16 Antagonist 3.0 10 " 7 48
parcial partial
AgonistaAgonist
11 9.6 10"7 19 Antagonista 8.8 10"7 32 11 9.6 10 "7 19 Antagonist 8.8 10 " 7 32
parcial partial
AgonistaAgonist
20 nd nd nd 7.1 10"8 24 20 nd nd nd 7.1 10 "8 24
parcial partial
Agonista AgonistaAgonist Agonist
21 3.3 10"7 26 2.9 10"8 29 21 3.3 10 "7 26 2.9 10 " 8 29
parcial parcial  partial partial
Agonista Agonist
22 nd nd nd 1 .1 10"6 32 22 nd nd nd 1 .1 10 "6 32
parcial 2.2. Determinación de propiedades antioxidantes partial 2.2. Determination of antioxidant properties
La capacidad de los productos de la invención para capturar radicales libres de oxígeno, como una medida de sus propiedades antioxidantes, se evaluó siguiendo el método ORAC-FL (Oxygen Radical Absorbance Capacity Assay by Fluorescence), descrito por Ou y col. (Ou, B.; Hampsch-Woodill, M.; Prior, R. L. Development and validation of an improved oxygen radical absorbance capacity assay using fluorescein as the fluorescent probé. J. Agrie. Food Chem. 2001 , 49, 4619-4626) y parcialmente modificado por Dávalos y col. (Dávalos, A.; Gómez-Cordovés, C; Bartolomé, B. Extending applicability of the oxygen radical absorbance capacity (ORAC-fluorescein) assay. J. Agrie. Food Chem. 2004, 52, 48-54). The ability of the products of the invention to capture oxygen free radicals, as a measure of their antioxidant properties, was evaluated following the ORAC-FL method (Oxygen Radical Absorbance Capacity Assay by Fluorescence), described by Ou et al. (Ou, B .; Hampsch-Woodill, M .; Prior, RL Development and validation of an improved oxygen radical absorbance capacity assay using fluorescein as the fluorescent tested. J. Agrie. Food Chem. 2001, 49, 4619-4626) and partially modified by Dávalos et al. (Dávalos, A .; Gómez-Cordovés, C; Bartolomé, B. Extending applicability of the oxygen radical absorbance capacity (ORAC-fluorescein) assay. J. Agrie. Food Chem. 2004, 52, 48-54).
El experimento se llevó a cabo en un fluorímetro Polarstar Galaxy (BMG Labtechnologies GmbH, Offenburg, Alemania) con lector de placas de 96- pocilios, con filtros de excitación y de emisión a 485-P y 520-P, respectivamente. El equipo fue controlado mediante el software Fluorostar Galaxy (versión 4.1 1 -0) para la medición de fluorescencia. El dihidrocloruro de 2,2'-azobis-(amidinopropano) (AAPH), el ácido (±)-6-hidroxi-2, 5,7,8- tetrametilcroman-2-carboxílico (trolox) y la fluoresceína (FL) se adquirieron en Sigma-Aldrich. La reacción se llevó a cabo en tampón fosfato (75 mM, pH 7.4), con un volumen total de 200 μί. Diferentes soluciones del producto a ensayar (20 μί) y de FL (120 μί, concentración final de 70 mM) se colocaron en una microplaca de 96 pocilios de color negro (Nunc, 96F sin tratar). La mezcla se preincubó durante 15 min a 37 °C, y luego se añadió rápidamente una solución de AAPH (60 μί, concentración final de 12 mM) usando una pipeta multicanal. La microplaca se colocó inmediatamente en el lector y se registró la fluorescencia cada minuto durante 80 min, agitando automáticamente la microplaca antes de cada lectura. Cada uno de los productos ensayados se midió en ocho concentraciones diferentes (0.1 -1 μΜ). En cada uno de los experimentos, se emplearon cuatro pocilios para el blanco (FL y AAPH en fosfato tampón, sin muestra) y ocho soluciones de calibración utilizando trolox (1 -8 μΜ). Todas las mezclas de reacción se prepararon por duplicado y cada producto se midió en tres ensayos independientes como mínimo. Los datos de fluorescencia fueron exportados desde el software Galaxy Fluostar a una hoja de Excel para cálculos posteriores. Las curvas de fluorescencia frente al tiempo fueron normalizadas con el blanco, calculándose a continuación el área bajo la curva (AUC) de cada muestra. Para cada uno de los productos ensayados, la representación gráfica de la AUC frente a la concentración de producto proporcionó rectas, que se ajustaron por regresión lineal. Los valores de ORAC-FL de los productos de la invención se encuentran recogidos en la tabla 3 y se expresan como equivalentes de trolox (μιηοΙ de trolox / μΐΎΐοΙ de producto) en una escala relativa, en la que al trolox se le asignó el valor de la unidad. La melatonina también se evaluó como patrón positivo, dando un valor de ORAC-FL de 2.3 equiv. de trolox, en concordancia con el valor de 2.0 equiv. de trolox, previamente descrito por Sofic et al. (Sofic, E.; Rimpapa, Z.; Kundurovic, Z.; Sapcanin, A.; Tahirovic, I.; Rustembegovic, A.; Cao, G. Antioxidant capacity of the neurohormone melatonin. J. Neural Transm. 2005, 112, 349-358). The experiment was carried out on a Polarstar Galaxy fluorometer (BMG Labtechnologies GmbH, Offenburg, Germany) with 96-well plate reader, with excitation and emission filters at 485-P and 520-P, respectively. The equipment was controlled by the Fluorostar Galaxy software (version 4.1 1 -0) for fluorescence measurement. 2,2'-Azobis- (amidinopropane) dihydrochloride (AAPH), (±) -6-hydroxy-2, 5,7,8-tetramethylchroman-2-carboxylic acid (trolox) and fluorescein (FL) are acquired in Sigma-Aldrich. The reaction was carried out in phosphate buffer (75 mM, pH 7.4), with a total volume of 200 μί. Different solutions of the product to be tested (20 μί) and FL (120 μί, final concentration of 70 mM) were placed in a black 96-well microplate (Nunc, 96F untreated). The mixture was pre-incubated for 15 min at 37 ° C, and then a solution of AAPH (60 μί, final concentration of 12 mM) was quickly added using a multichannel pipette. The microplate was immediately placed in the reader and fluorescence was recorded every minute for 80 min, automatically shaking the microplate before each reading. Each of the products tested was measured in eight different concentrations (0.1 -1 μΜ). In each of the experiments, four wells were used for the target (FL and AAPH in phosphate buffer, without sample) and eight calibration solutions using trolox (1-8 μΜ). All reaction mixtures were prepared in duplicate and each product was measured in at least three independent tests. Fluorescence data was exported from the Galaxy Fluostar software to an Excel sheet for later calculations. The fluorescence versus time curves were normalized with the blank, then the area under the curve (AUC) of each sample was calculated. For each of the products tested, the graphic representation of the AUC versus the product concentration provided straight lines, which were adjusted by linear regression. The ORAC-FL values of the products of the invention are listed in Table 3 and are expressed as equivalents of trolox (μιηοΙ of trolox / μΐΎΐοΙ of product) on a relative scale, in which the trolox was assigned the value of the unit. Melatonin was also evaluated as a positive standard, giving an ORAC-FL value of 2.3 equiv. of trolox, in accordance with the value of 2.0 equiv. of trolox, previously described by Sofic et al. (Sofic, E .; Rimpapa, Z .; Kundurovic, Z .; Sapcanin, A .; Tahirovic, I .; Rustembegovic, A .; Cao, G. Antioxidant capacity of the neurohormone melatonin. J. Neural Transm. 2005, 112 , 349-358).
Como puede observarse en la tabla 3, los productos de la invención presentan buenas propiedades antioxidantes, próximas a la melatonina y hasta 2.72 veces más potentes que trolox, el fragmento aromático y activo de la vitamina E y el responsable de la captura de radicales libres. Por lo tanto, los productos de la invención se comportan como antioxidantes útiles para contrarrestar el estrés oxidativo producido por un exceso de radicales libres. Tabla 3. Datos de captura de radicales libres (ORAC-FL), de inhibición de acetilcolinesterasa humana (h-AChE) y de butirilcolinesterasa humana (h- BuChE). As can be seen in Table 3, the products of the invention have good antioxidant properties, close to melatonin and up to 2.72 times more potent than trolox, the aromatic and active fragment of vitamin E and responsible for the capture of free radicals. Therefore, the products of the invention behave as useful antioxidants to counteract the oxidative stress caused by an excess of free radicals. Table 3. Free radical capture (ORAC-FL), inhibition of human acetylcholinesterase (h-AChE) and human butyrylcholinesterase (h-BuChE) data.
ORAC-FL ORAC-FL
h-AChE h-BuChE h-AChE h-BuChE
Comp. (μΐΎΐοΙ trolox / μιηοΙ Comp. (μΐΎΐοΙ trolox / μιηοΙ
(Clso, MM)b (Cl50, MM)b producto)3 (Clso, MM) b (Cl 50 , MM) b product) 3
1 1.65 ±0.08 6.95 ±0.34 >101 1.65 ± 0.08 6.95 ± 0.34> 10
2 2.05 ±0.31 7.59 ±0.30 >102 2.05 ± 0.31 7.59 ± 0.30> 10
3 1.63 ± 0.09 >10 >103 1.63 ± 0.09> 10> 10
4 1.99 ± 0.12 >10 >104 1.99 ± 0.12> 10> 10
5 1.62 ± 0.23 >10 >105 1.62 ± 0.23> 10> 10
6 1.78 ± 0.06 >10 >106 1.78 ± 0.06> 10> 10
10 1.61 ± 0.04 >10 >10 10 1.61 ± 0.04> 10> 10
11 1.66 ± 0.08 >10 >10  11 1.66 ± 0.08> 10> 10
12 1.97 ± 0.11 6.26 ±0.20 >10  12 1.97 ± 0.11 6.26 ± 0.20> 10
13 2.06 ± 0.22 6.99 ±0.26 >10  13 2.06 ± 0.22 6.99 ± 0.26> 10
14 1.94 ± 0.07 5.07 ±0.23 >10  14 1.94 ± 0.07 5.07 ± 0.23> 10
15 2.28 ± 0.22 6.24 ±0.27 >10  15 2.28 ± 0.22 6.24 ± 0.27> 10
16 1.78 ± 0.02 6.05 ±0.41 >10  16 1.78 ± 0.02 6.05 ± 0.41> 10
17 1.52 ± 0.03 4.86 ±0.24 >10  17 1.52 ± 0.03 4.86 ± 0.24> 10
18 1.55 ±0.01 6.58 ±0.33 >10  18 1.55 ± 0.01 6.58 ± 0.33> 10
19 1.29 ± 0.19 5.30 ±0.41 >10  19 1.29 ± 0.19 5.30 ± 0.41> 10
20 2.72 ± 0.33 8.08 ±0.69 >10  20 2.72 ± 0.33 8.08 ± 0.69> 10
21 2.57 ± 0.68 6.50 ±0.51 >10  21 2.57 ± 0.68 6.50 ± 0.51> 10
22 1.89 ± 0.12 7.12 ±0.32 >10  22 1.89 ± 0.12 7.12 ± 0.32> 10
23 1.92 ± 0.11 6.49 ±0.39 >10  23 1.92 ± 0.11 6.49 ± 0.39> 10
24 2.01 ± 0.17 8.93 ±0.48 >10  24 2.01 ± 0.17 8.93 ± 0.48> 10
Melatonina 2.30 ± 0.10 nd nd  Melatonin 2.30 ± 0.10 nd nd
Tacrina nd 0.04 ± 0.002 0.010 ±0.004 aMedia de 3 experimentos independientes ± desviación estándar. bMedia de 3 experimentos independientes ± SEM. Tacrine nd 0.04 ± 0.002 0.010 ± 0.004 a Average of 3 independent experiments ± standard deviation. b Average of 3 independent experiments ± SEM.
2.3. Inhibición de acetil- y butirilcolinesterasas humanas 2.3. Inhibition of human acetyl- and butyrylcholinesterase
Los compuestos de la invención fueron evaluados como inhibidores de acetil- y butirilcolinesterasa humanas (h-AChE y h-BuChE). La medida de la inhibición enzimática se realizó siguiendo el método de Ellman (Ellman, G. L.; Courtney, K. D.; Andrés, V., Jr.; Feather-Stone, R. M. A new and rapid colorimetric determination of acetylcholinesterase activity. Biochem. Pharmacol. 1961 , 7, 88-95). La disolución de ensayo estaba formada por tampón fosfato 0.1 M a pH=8, ácido 5,5'-ditiobis-2-nitrobenzoico (DTNB) 400 μΜ, las enzimas h-AChE (acetilcolinesterasa humana recombinante, Sigma Chemical Co.) 0.05 ud/mL o h-BuChE (butirilcolinesterasa de suero humano, Sigma Chemical Co.) 0.024 ud/mL y 800 μΜ de yoduro de acetiltiocolina o 500 μΜ de butiriltiocolina respectivamente, como sustratos de las reacciones enzimáticas. Los compuestos a evaluar se preincubaron con la enzima durante 5 minutos a 30°C, se añadió el sustrato y se midieron los cambios de absorbancia a 412 nm cada 5 minutos en un espectrómetro UV/VIS Multiskan Spectrum. Se compararon las velocidades de reacción y se calcularon los porcentajes de inhibición debidos a la presencia de los compuestos que se analizaban. La actividad enzimática a cada concentración de compuesto se expresa como porcentaje de actividad con respecto al control en ausencia de compuesto. La CI50 se define como la concentración de compuesto que inhibe la actividad enzimática un 50% con respecto al control de enzima sin tratar. Los resultados se encuentran en la tabla 3, expresándose como la media de tres experimentos independientes ± desviación estándar. The compounds of the invention were evaluated as inhibitors of human acetyl- and butyrylcholinesterase (h-AChE and h-BuChE). Enzyme inhibition was measured using the Ellman method (Ellman, GL; Courtney, KD; Andrés, V., Jr .; Feather-Stone, RM A new and rapid colorimetric determination of acetylcholinesterase activity. Biochem. Pharmacol. 1961 , 7, 88-95). The test solution was formed by 0.1 M phosphate buffer at pH = 8, 5,5'-dithiobis-2-nitrobenzoic acid (DTNB) 400 μΜ, the enzymes h-AChE (recombinant human acetylcholinesterase, Sigma Chemical Co.) 0.05 ud / mL or h-BuChE (human serum butyrylcholinesterase, Sigma Chemical Co.) 0.024 ud / mL and 800 μΜ of acetylthiocholine iodide or 500 μΜ of butyrylthiocholine respectively, as substrates for enzymatic reactions. The compounds to be evaluated were pre-incubated with the enzyme for 5 minutes at 30 ° C, the substrate was added and the absorbance changes were measured at 412 nm every 5 minutes in a Multiskan Spectrum UV / VIS spectrometer. The reaction rates were compared and the percentages of inhibition due to the presence of the compounds being analyzed were calculated. The enzymatic activity at each compound concentration is expressed as a percentage of activity with respect to the control in the absence of compound. IC50 is defined as the concentration of compound that inhibits 50% enzyme activity with respect to the control of untreated enzyme. The results are found in table 3, expressed as the average of three independent experiments ± standard deviation.
Los resultados de inhibición de colinesterasas humanas indican que la mayoría de los compuestos evaluados son inhibidores selectivos de h-AChE con concentraciones inhibitorias 50 en el orden micromolar. Por lo tanto, son capaces de aumentar moderadamente los niveles del neurotransmisor acetilcolina y mejorar las capacidades cognitivas de los pacientes. The results of inhibition of human cholinesterase indicate that most of the compounds evaluated are selective inhibitors of h-AChE with 50 inhibitory concentrations in the micromolar order. Therefore, they are able to moderately increase the levels of the neurotransmitter acetylcholine and improve the cognitive abilities of patients.
2.4. Ensayos de binding sobre receptores de serotonina Varios compuestos de la invención fueron ensayados frente a los siguientes subtipos de receptores humanos de serotonina 5-HTiA, 5-HTiB, 5-HTiD, 5-HTiE, 5-HT2A, 5-ΗΪ2Β, 5-HT2C, 5-HT3, 5-HT5A, 5-HT6 y 5-HT7 en la Universidad de Carolina del Norte de Chapel Hill USA (National Institute of Mental Health's Psychoactive Drug Screening Program), donde se siguieron los protocolos experimentales descritos en http://pdsp.med.unc.edu/. 2.4. Binding assays on serotonin receptors Several compounds of the invention were tested against the following subtypes of human serotonin receptors 5-HTi A , 5-HTi B , 5-HTi D , 5-HTi E , 5-HT 2A , 5 -ΗΪ2Β, 5-HT 2C , 5-HT 3 , 5-HT 5A , 5-HT 6 and 5-HT 7 at the University of North Carolina at Chapel Hill USA (National Institute of Mental Health's Psychoactive Drug Screening Program), where the experimental protocols described in http://pdsp.med.unc.edu/ were followed.
De los productos ensayados, se encontró que 12 y 15 son ligandos del receptor 5-HTIA recombinante humano con constantes de inhibición K¡ de 7768 y de 957 nanomolar, respectivamente, empleando como radioligando [3H]8-OH-DPAT. En el resto de los receptores ensayados no se observó unión usando una concentración de producto de 10 micromolar, lo que indica una clara selectividad por el subtipo 5-HTiA. Of the products tested, 12 and 15 were found to be ligands of the human recombinant 5-HTIA receptor with K68 inhibition constants of 7768 and 957 nanomolar, respectively, using as radioligand [ 3 H] 8-OH-DPAT. In the rest of the receptors tested no binding was observed using a product concentration of 10 micromolar, indicating a clear selectivity for the 5-HTi A subtype.
También se evaluó la afinidad del compuesto 11 por el sitio del agonista del receptor humano 5-HTIA en células transfectadas HEK-293, empleando [3H]8- OH-DPAT como radioligando (Cerep, www.cerep.com/). A una concentración de 10 micromolar, el producto 11 inhibió la unión del radioligando en un 94% y fue capaz de provocar un 38% de respuesta en comparación con el producto de referencia, serotonina. En un ensayo similar sobre el receptor 5-HT2C el compuesto 11 demostró no estimular ninguna respuesta significativa a dicha concentración. The affinity of compound 11 for the 5-HTIA human receptor agonist site in HEK-293 transfected cells was also evaluated, using [ 3 H] 8-OH-DPAT as radioligand (Cerep, www.cerep.com/). At a concentration of 10 micromolar, product 11 inhibited radioligand binding in 94% and was able to cause a 38% response compared to the reference product, serotonin. In a similar test on the 5-HT2C receptor, compound 11 demonstrated no stimulation of any significant response to said concentration.
2.5. Estudios in vitro de neurogénesis sobre células madre neurales de ratas adultas 2.5 In vitro studies of neurogenesis on neural stem cells of adult rats
Los estudios de neurogénesis in vitro se realizaron empleando cultivos de células madre neurales que se iniciaron a partir de las dos principales áreas neurogénicas: la zona subventricular (SVZ) y la zona subgranular del giro dentado del hipocampo de ratas Wistar adultas. Para ello se diseccionó el cerebro obteniéndose la SVZ y el hipocampo, los cuales fueron disgregados en medio DMEM (Dubecco's Modified Eagle's Médium) (Morales-García, J. A.; Luna-Medina, R.; Alfaro-Cervello, C; Cortes-Canteli, M.; Santos, A.; García- Verdugo, J. M.; Pérez-Castillo, A. Peroxisome proliferator-activated receptor gamma ligands regúlate neural stem cell proliferation and differentiation in vitro and in vivo. Glia 2011 , 59, 293-307). Las células obtenidas se cultivaron mediante métodos establecidos para conseguir una proliferación óptima en medio DMEM/F12 (1 :1 , Invitrogen) y se suplementaron con 10 ng / mL de factor de crecimiento epidérmico (EGF, Peprotech, Londres, Reino Unido), 10 ng / mL de factor de crecimiento de fibroblastos (FGF, Peprotech) y medio B27 (Gibco) (Ferron, S. R.; Andreu-Agullo, C; Mira, H.; Sánchez, P.; Marqués-Torrejón, M. A.; Fariñas, I. A combined ex/in vivo assay to detect effects of exogenously added factors in neural stem cells. Nat. Protoc. 2007, 2, 849-859). En estas condiciones las células proliferan en flotación y forman grupos esféricos llamados neuroesferas (NS). Tras 3 días en cultivo las NS se trataron con los productos 4, 11 y 14 a una concentración de 10 micromolar. Para determinar la capacidad de los compuestos para inducir la diferenciación, las neuroesferas tratadas durante 10 días en flotación, se pegaron sobre un sustrato (cubres tratados con 100 μg / mL de poli-L-lisina) y se trataron durante 24, 48 y 96 horas más en ausencia de factores de crecimiento exógenos y con suero (Morales-García, J. A.; Luna-Medina, R.; Alonso-Gil, S.; Sanz-Sancristóbal, M.; Palomo, V.; Gil, C; Santos, A.; Martínez, A.; Pérez-Castillo, A. Glycogen synthase kinase 3 inhibition promotes adult hippocampal neurogenesis in vitro and in vivo. ACS Chem. Neurosci. 2012, 3, 963-971 ). Tras el tratamiento, los cristales con las neuroesferas se procesaron para inmunocitoquímica con dos tipos de marcadores neuronales asociados a neurogénesis: anticuerpo anti- beta-tubulina (clon Tuj1 ), relacionado con estadios tempranos de neurogénesis y MAP-2 (microtubule-associated protein 2), marcador de madurez neuronal. Los valores básales del experimento se obtuvieron en las mismas condiciones, en ausencia de producto. Como controles se emplearon melatonina (ligando endógeno de los receptores melatoninérgicos) y luzindol (antagonista de los receptores melatoninérgicos). Las imágenes se obtuvieron utilizando un microscopio de fluorescencia Nikon 90i, acoplado a una cámara digital Qi. La configuración del microscopio se ajustó para producir la óptima relación señal- ruido. In vitro neurogenesis studies were performed using neural stem cell cultures that were initiated from the two main areas neurogenic: the subventricular zone (SVZ) and the subgranular zone of the dentate gyrus of the adult Wistar rat hippocampus. For this, the brain was dissected, obtaining the SVZ and the hippocampus, which were divided into DMEM (Dubecco's Modified Eagle's Medium) (Morales-García, JA; Luna-Medina, R .; Alfaro-Cervello, C; Cortes-Canteli, M .; Santos, A .; García-Verdugo, JM; Pérez-Castillo, A. Peroxisome proliferator-activated receptor gamma ligands regulate neural stem cell proliferation and differentiation in vitro and in vivo. Glia 2011, 59, 293-307). The cells obtained were cultured by established methods to achieve optimal proliferation in DMEM / F12 medium (1: 1, Invitrogen) and were supplemented with 10 ng / mL epidermal growth factor (EGF, Peprotech, London, United Kingdom), 10 ng / mL fibroblast growth factor (FGF, Peprotech) and medium B27 (Gibco) (Ferron, SR; Andreu-Agullo, C; Mira, H .; Sánchez, P .; Marqués-Torrejón, MA; Fariñas, I A combined ex / in vivo assay to detect effects of exogenously added factors in neural stem cells. Nat. Protoc. 2007, 2, 849-859). Under these conditions the cells proliferate in flotation and form spherical groups called neurospheres (NS). After 3 days in culture the NS were treated with products 4, 11 and 14 at a concentration of 10 micromolar. To determine the ability of the compounds to induce differentiation, the neurospheres treated for 10 days on flotation were glued on a substrate (covers treated with 100 μg / mL of poly-L-lysine) and treated for 24, 48 and 96 more hours in the absence of exogenous and serum growth factors (Morales-García, JA; Luna-Medina, R .; Alonso-Gil, S .; Sanz-Sancristóbal, M .; Palomo, V .; Gil, C; Santos , A .; Martínez, A .; Pérez-Castillo, A. Glycogen synthase kinase 3 inhibition promotes adult hippocampal neurogenesis in vitro and in vivo. ACS Chem. Neurosci. 2012, 3, 963-971). After treatment, the crystals with the neurospheres were processed for immunocytochemistry with two types of neuronal markers associated with neurogenesis: anti-beta-tubulin antibody (clone Tuj1), related to early stages of neurogenesis and MAP-2 (microtubule-associated protein 2 ), marker of neuronal maturity. The baseline values of the experiment were obtained under the same conditions, in the absence of product. Melatonin (ligand) was used as controls endogenous to melatoninergic receptors) and luzindole (melatoninergic receptor antagonist). The images were obtained using a Nikon 90i fluorescence microscope, coupled to a Qi digital camera. The microscope configuration was adjusted to produce the optimal signal-to-noise ratio.
Las figuras 1 y 2 muestran el efecto neurogénico de los productos de la invención sobre cultivos de células madre neuronales de ratas Wistar adultas tratadas con vehículo (basal), melatonina (ligando endógeno de los receptores MT), luzindol (antagonista de los receptores MT), 4, 11 y 14 todos ellos a una concentración de 10 micromolar. Se muestran imágenes de neuroesferas completas, así como ampliaciones que muestran el interior (centro de la neuroesfera) y el exterior de las mismas (zona de migración). Se emplearon dos tipos de marcadores neuronales: anticuerpo anti-beta-tubulina y MAP-2, asociados respectivamente a neurogénesis temprana y madurez neuronal. La tinción DAPI se utilizó como marcador nuclear. Todos los productos evaluados fueron capaces de aumentar la diferenciación de neuroesferas (marcador Tuj1 , neurogénesis temprana) (Figura 1 ). Además, 11 y 14 son potentes inductores de neurogénesis a todos los tiempos estudiados y con ambos marcadores (TuJ y MAP-2), siendo más potentes que la propia melatonina (Figura 2). Figures 1 and 2 show the neurogenic effect of the products of the invention on neuronal stem cell cultures of adult Wistar rats treated with vehicle (basal), melatonin (endogenous ligand of MT receptors), luzindole (antagonist of MT receptors) , 4, 11 and 14 all of them at a concentration of 10 micromolar. Full neurosphere images are shown, as well as enlargements that show the inside (center of the neurosphere) and the outside of them (migration zone). Two types of neuronal markers were used: anti-beta-tubulin antibody and MAP-2, respectively associated with early neurogenesis and neuronal maturity. DAPI staining was used as a nuclear marker. All the products evaluated were able to increase the differentiation of neurospheres (Tuj1 marker, early neurogenesis) (Figure 1). In addition, 11 and 14 are potent inducers of neurogenesis at all times studied and with both markers (TuJ and MAP-2), being more potent than melatonin itself (Figure 2).
2.6. Estudios in vivo de neurogénesis en ratones 2.6. In vivo studies of neurogenesis in mice
Animales. Los estudios de neurogénesis in vivo se realizaron sobre ratones macho C57BL/6 emparejados por edad que no expresan el transgén como animales de tipo salvaje. Una vez al día durante 7 días, el compuesto 11 (500 μg / kg) y BrdU (5-bromo-2-desoxiuridina) (50 mg / kg) se inyectaron por vía intraperitoneal a cada ratón. Los ratones fueron divididos en dos grupos y sacrificados bajo anestesia profunda, después de 24 horas (grupo 1 ) o de 21 días (grupo 2) desde la última inyección de BrdU. Todos los animales fueron manipulados y atendidos según la Directiva del Parlamento Europeo 2010/63/UE, de 22 de septiembre de 2010. Inmunohistoquímica. Los ratones fueron anestesiados profundamente con isoforano, perfundidos a través del miocardio con solución salina 0.9% y sus cerebros fueron extraídos inmediatamente. A continuación, los tejidos se fijaron en solución tamponada de fosfato pH 7.4 con paraformaldehído al 4%, a 4 °C. Los cerebros fijados se cortaron en un vibratomo (Leica Microsystems) a 30 μΐΎΐ, y las secciones de tejido se recogieron en PBS 0.1 M frío y se incubaron a 4 °C durante la noche con dos anticuerpos primarios, empleando anti - BrdU de ratón (1 :20000, Hybridoma Bank) en ambos grupos de ratones. En el primer grupo (3 animales sacrificados 24 horas después de la última inyección de BrdU) se empleó además anti - doublecortin de cabra (DCX, 1 :500, Santa Cruz Biotechnology), mientras que en el segundo (6 animales) se usó NeuN (1 :500, Millipore). Después de la incubación durante una noche, la tinción de los anticuerpos primarios se revelaron con los siguientes anticuerpos: anti - IgG de ratón 488 (FluoProbes®, Interchim) para BrdU, y Rojo Texas anti - IgG de conejo (Jackson Immunoresearch, West Grove) para DCX y NeuN. Animals. Neurogenesis studies in vivo were performed on C57BL / 6 age-matched male mice that do not express the transgene as wild-type animals. Once daily for 7 days, compound 11 (500 μg / kg) and BrdU (5-bromo-2-deoxyuridine) (50 mg / kg) were injected intraperitoneally to each mouse. The mice were divided into two groups and sacrificed under deep anesthesia, after 24 hours (group 1) or 21 days (group 2) since the last injection of BrdU. All animals were handled and treated according to the Directive of the European Parliament 2010/63 / EU, of September 22, 2010. Immunohistochemistry The mice were deeply anesthetized with isophoran, perfused through the myocardium with 0.9% saline and their brains were immediately removed. The tissues were then fixed in phosphate buffered solution pH 7.4 with 4% paraformaldehyde, at 4 ° C. The fixed brains were cut in a vibratome (Leica Microsystems) at 30 μΐΎΐ, and the tissue sections were collected in 0.1 M cold PBS and incubated at 4 ° C overnight with two primary antibodies, using mouse anti-BrdU ( 1: 20000, Hybridoma Bank) in both groups of mice. In the first group (3 animals slaughtered 24 hours after the last injection of BrdU) goat anti-doublecortin (DCX, 1: 500, Santa Cruz Biotechnology) was also used, while in the second (6 animals) NeuN was used (1: 500, Millipore). After overnight incubation, staining of the primary antibodies was revealed with the following antibodies: anti-mouse IgG 488 (FluoProbes®, Interchim) for BrdU, and Texas Red anti-rabbit IgG (Jackson Immunoresearch, West Grove ) for DCX and NeuN.
BrdU es un nucleósido modificado que se incorpora al ADN durante la fase S del ciclo celular, por lo que en estos experimentos se emplea como marcador de células recién formadas. DCX es una proteína asociada a microtúbulos expresada en neuronas inmaduras y NeuN es un marcador de maduración neuronal. BrdU is a modified nucleoside that is incorporated into DNA during the S phase of the cell cycle, so in these experiments it is used as a marker of newly formed cells. DCX is a microtubule-associated protein expressed in immature neurons and NeuN is a marker of neuronal maturation.
Cuantificación. Para estimar el número total de células BrdU-positivas (BrdU+) en todo el giro dentado, se contó el número de BrdU+ en la células granulares y en la capa de células subgranular del giro dentado. Se contaron las células BrdU+ en cada una de las seis secciones desde rostral (2 mm desde el bregma) a caudal (-4,3 mm desde el bregma). Para determinar el destino de las células que se dividen, se analizaron100-150 células BrdU+ a través de 4-6 secciones por ratón mediante microscopía confocal para la co-expresión con DCX (grupo 1 ) o para la co-expresión con NeuN (grupo 2). Aquellos núcleos BrdU+ con colocalización de DCX+ en el citoplasma se consideraron como neuronas recién nacidas, todavía inmaduras, y las que co-localizan BrdU+ y NeuN+ como neuronas maduras. El número de células doble-positivas BrdlT - DCX+ ó BrdlT - NeuN+ se expresó como porcentaje frente a las células BrdlT. Quantification. To estimate the total number of BrdU-positive cells (BrdU + ) in the entire dentate gyrus, the number of BrdU + in the granular cells and in the subgranular cell layer of the dentate gyrus was counted. BrdU + cells were counted in each of the six sections from rostral (2 mm from the bregma) to flow rate (-4.3 mm from the bregma). To determine the fate of the dividing cells, 100-150 BrdU + cells were analyzed through 4-6 sections per mouse by confocal microscopy for co-expression with DCX (group 1) or for co-expression with NeuN ( group 2). Those BrdU + nuclei with DCX + colocalization in the cytoplasm were considered as newly born, still immature neurons, and those that co-localize BrdU + and NeuN + as mature neurons The number of double-positive BrdlT-DCX + or BrdlT-NeuN + cells was expressed as a percentage against BrdlT cells.
Resultados. La proliferación celular no diferenciada se evaluó por la incorporación de BrdU en el ADN de las nuevas células, como se explica en el párrafo anterior. El número de células BrdU positivas, BrdlT (proliferación celular, sin diferenciación del fenotipo) en el giro dentado del hipocampo no cambió significativamente con respecto al basal, ni en el grupo 1 (animales sacrificados 24 horas después del último tratamiento) (Figura 3) ni en el grupo 2 (animales sacrificados 21 días después del último tratamiento) (Figura 4). Estos resultados indican que el compuesto 11 no altera significativamente la tasa de proliferación no diferenciada, ni a corto ni a medio-largo plazo. Results Undifferentiated cell proliferation was evaluated by the incorporation of BrdU into the DNA of the new cells, as explained in the previous paragraph. The number of BrdU positive cells, BrdlT (cell proliferation, without differentiation of the phenotype) in the dentate gyrus of the hippocampus did not change significantly from baseline, nor in group 1 (animals sacrificed 24 hours after the last treatment) (Figure 3) nor in group 2 (animals slaughtered 21 days after the last treatment) (Figure 4). These results indicate that compound 11 does not significantly alter the rate of undifferentiated proliferation, either in the short or medium-long term.
En el grupo 1 (animales sacrificados 24 horas después del último tratamiento) se observó un aumento significativo de los núcleos doblemente marcados con BrdU y DCX, de casi el doble (Figura 5). Estos resultados indican que en ratones tratados con el compuesto 11 durante 7 días, se duplicó el número de neuronas inmaduras al cabo de 24 horas desde la última administración del compuesto 11. In group 1 (animals slaughtered 24 hours after the last treatment) there was a significant increase in the doubly labeled nuclei with BrdU and DCX, almost double (Figure 5). These results indicate that in mice treated with compound 11 for 7 days, the number of immature neurons doubled after 24 hours since the last administration of compound 11.
En el grupo 2 (animales sacrificados 21 días después del último tratamiento) se observó un aumento significativo de los núcleos doblemente marcados con BrdU y NeuN, de seis veces (Figura 6). Estos resultados indican que en ratones tratados con el compuesto 11 durante 7 días, el número de neuronas maduras se multiplicó por seis al cabo de 21 días desde la última administración del compuesto 11. En la figura 7 se muestran microimágenes representativas de hipocampo de ratones, donde se observan células BrdU+ - NeuN+ que corresponden a neuronas maduras. In group 2 (animals slaughtered 21 days after the last treatment) there was a significant increase in the doubly labeled nuclei with BrdU and NeuN, six times (Figure 6). These results indicate that in mice treated with compound 11 for 7 days, the number of mature neurons was multiplied by six after 21 days since the last administration of compound 11. Representative micro hip images of mice are shown in Figure 7, where BrdU + - NeuN + cells corresponding to mature neurons are observed.
En su conjunto, estos experimentos con ratones muestran que el compuesto 11 es capaz de aumentar in vivo el número de progenitores neuronales y de neuronas maduras, sin alterar la tasa de la proliferación no diferenciada. Por lo tanto, se puede concluir que el derivado 11 es un compuesto con capacidad para regenerar in vivo poblaciones neuronales dañadas sin provocar la aparición de tumores. Together, these experiments with mice show that compound 11 is capable of increasing in vivo the number of neuronal progenitors and mature neurons, without altering the rate of undifferentiated proliferation. Therefore, it can be concluded that derivative 11 is a compound capable of regenerating damaged neuronal populations in vivo without causing the appearance of tumors.

Claims

REIVINDICACIONES
1.- Compuesto de fórmula general (I) 1.- Compound of general formula (I)
Figure imgf000051_0001
sus sales o solvatos farmacéuticamente aceptables, dónde R1 es H, alquilo (C1 -C6) o alcoxilo (C1 -C6) n = 1 -6
Figure imgf000051_0001
their pharmaceutically acceptable salts or solvates, where R1 is H, alkyl (C1 -C6) or alkoxyl (C1 -C6) n = 1 -6
X es O o N, con la particularidad de que cuando X es O, Y es N, L1 está ausente y L2 es alquenilo o alquinilo Y es O o N, con la particularidad de que cuando Y es O, X es N y L1 y L2 dan lugar a un heterociclo aromático de 5 miembros que comprende hasta 3 heteroátomos y que puede estar sustituido por hidrógeno, alquilo (C1 -C6), hidroxilo, alcoxilo (C1 -C6), tiol o mercaptoalquilo (C1 -C6) X is O or N, with the particularity that when and L2 give rise to a 5-membered aromatic heterocycle comprising up to 3 heteroatoms and which can be substituted by hydrogen, alkyl (C1 -C6), hydroxyl, alkoxy (C1 -C6), thiol or mercaptoalkyl (C1 -C6)
L2 es alquenilo, alquinilo (cuando X es O) o forma junto con L1 un heterociclo aromático de 5 miembros que comprende hasta 3 heteroátomos y que puede estar sustituido por hidrógeno, alquilo (C1 -C6), hidroxilo, alcoxilo (C1 -C6), tiol o mercaptoalquilo (C1 -C6) L2 is alkenyl, alkynyl (when , thiol or mercaptoalkyl (C1 -C6)
2.- Compuesto según la reivindicación 1 y la fórmula general (I), donde X es O, Y es N, n es igual a 1 o 2 y L2 es alilo o prop-2-in-1 -ilo, según la fórmula (II): 2.- Compound according to claim 1 and the general formula (I), where X is O, Y is N, n is equal to 1 or 2 and L2 is allyl or prop-2-in-1 -yl, according to the formula (II):
Figure imgf000052_0001
Figure imgf000052_0001
Fórmula (II) Formula (II)
3.- Compuesto según la reivindicación 1 y la fórmula general (I), donde X es N, Y es O, n es igual a 1 o 2 y L1 y L2 forman un heterociclo aromático de 5 miembros y dos heteroátomos sustituido con un metilo, según la fórmula 3.- Compound according to claim 1 and the general formula (I), where X is N, Y is O, n is equal to 1 or 2 and L1 and L2 form a 5-membered aromatic heterocycle and two heteroatoms substituted with a methyl , according to the formula
Figure imgf000052_0002
Figure imgf000052_0002
FórmulaFormula
4.- Compuesto según la reivindicación 1 y la fórmula general (I), donde X es N, Y es O, n es igual a 1 o 2 y L1 y L2 forman un heterociclo aromático de 5 miembros y tres heteroátomos, sustituido con un metilo según la fórmula (IV): 4.- Compound according to claim 1 and the general formula (I), where X is N, Y is O, n is equal to 1 or 2 and L1 and L2 form an aromatic heterocycle with 5 members and three heteroatoms, substituted with a methyl according to formula (IV):
Figure imgf000052_0003
Figure imgf000052_0003
Fórmula (IV) Formula (IV)
5.- Compuesto según la reivindicación 1 y la fórmula general (I), donde X es N, Y es O, n es igual a 1 o 2, y L1 y L2 forman un heterociclo aromático de 5 miembros y tres heteroátomos, sustituido con un radical R2 que es metilo, hidroxilo o tiol según la fórmula (V): 5.- Compound according to claim 1 and the general formula (I), where X is N, Y is O, n is equal to 1 or 2, and L1 and L2 form a 5-membered aromatic heterocycle and three heteroatoms, substituted with a radical R2 that is methyl, hydroxyl or thiol according to formula (V):
Figure imgf000053_0001
Figure imgf000053_0001
Fórmula (V) Formula (V)
6.- Compuesto según la reivindicación 1 y la fórmula general (I), donde X es N, Y es N, n es igual a 1 o 2, y L1 y L2 forman un heterociclo aromático de 5 miembros y tres heteroátomos, sustituido con un radical R2 que es hidroxilo o tiol y un radical R3 que es alquilo según la fórmula (VI): 6.- Compound according to claim 1 and the general formula (I), where X is N, Y is N, n is equal to 1 or 2, and L1 and L2 form a 5-membered aromatic heterocycle and three heteroatoms, substituted with a radical R2 which is hydroxyl or thiol and a radical R3 which is alkyl according to formula (VI):
Figure imgf000053_0002
Figure imgf000053_0002
Fórmula (VI) Formula (VI)
7.- Compuesto según la reivindicación 1 donde dicho compuesto se selecciona de entre el siguiente grupo: · N-Alil-2-(5-metoxi-1 H-indol-3-il)acetamida (1 ) 7.- Compound according to claim 1 wherein said compound is selected from the following group: N-Allyl-2-(5-methoxy-1 H-indol-3-yl)acetamide (1)
N-Alil-3-(5-metoxi-1 H-indol-3-il)propanamida (2) N-Allyl-3-(5-methoxy-1 H-indol-3-yl)propanamide (2)
2- (5-Metoxi-1 H-indol-3-il)-N-(prop-2-in-1 -il)acetamida (3) 2- (5-Methoxy-1 H-indol-3-yl)-N-(prop-2-in-1 -yl)acetamide (3)
3- (5-Metoxi-1 H-indol-3-il)-N-(prop-2-in-1 -il)propanamida (4) 3- (5-Methoxy-1 H-indol-3-yl)-N-(prop-2-in-1 -yl)propanamide (4)
2-(2-(1 H-lndol-3-il)etil)-5-metiloxazol (5) 2-(2-(1 H-lndol-3-yl)ethyl)-5-methyloxazole (5)
· 2-((1 H-lndol-3-il)metil)-5-metiloxazol (6) 2-((5-Metoxi-1 H-indol-3-il)metil)-5-metiloxazol (7) · 2-((1 H-lndol-3-yl)methyl)-5-methyloxazole (6) 2-((5-Methoxy-1 H-indol-3-yl)methyl)-5-methyloxazole (7)
2-(2-(5-Metoxi-1 H-indol-3-il)etil)-5-metiloxazol (8) 2-(2-(5-Methoxy-1 H-indol-3-yl)ethyl)-5-methyloxazole (8)
5-((5-Metoxi-1 H-indol-3-il)metil)-3-metil-1 ,2,4-oxadiazol (9) 5-((5-Methoxy-1 H-indol-3-yl)methyl)-3-methyl-1,2,4-oxadiazole (9)
5-(2-(5-Metoxi-1 H-indol-3-il)etil)-3-metil-1 ,2,4-oxadiazol (10) 5-(2-(5-Methoxy-1 H-indol-3-yl)ethyl)-3-methyl-1,2,4-oxadiazole (10)
· 2-(2-(5-Metoxi-1 H-indol-3-il)etil)-5-metil-1 ,3,4-oxadiazol (1 1 ) · 2-(2-(5-Methoxy-1 H-indol-3-yl)ethyl)-5-methyl-1,3,4-oxadiazole (1 1)
5-((5-Metoxi-1 H-indol-3-il)metil)-1 ,3,4-oxadiazol-2-ol (12) 5-((5-Methoxy-1 H-indol-3-yl)methyl)-1,3,4-oxadiazol-2-ol (12)
5-(2-(1 H-lndol-3-il)etil)-1 ,3,4-oxadiazol-2-ol (13) 5-(2-(1 H-lndol-3-yl)ethyl)-1,3,4-oxadiazol-2-ol (13)
5-((5-Metoxi-1 H-indol-3-il)metilo)-1 ,3,4-oxadiazol-2-tiol (14) 5-((5-Methoxy-1 H-indol-3-yl)methyl)-1,3,4-oxadiazol-2-thiol (14)
5-(2-(1 H-lndol-3-il)etil)-1 ,3,4-oxadiazol-2-tiol (15) 5-(2-(1 H-lndol-3-yl)ethyl)-1,3,4-oxadiazol-2-thiol (15)
· 5-((5-Metoxi-1 H-indol-3-il)metil)-4-metil-4H-1 ,2,4-triazol-3-ol (16) · 5-((5-Methoxy-1 H-indol-3-yl)methyl)-4-methyl-4H-1,2,4-triazol-3-ol (16)
5-((1 H-lndol-3-il)metil)-4-metil-4H-1 ,2,4-triazol-3-ol (17) 5-((1H-lndol-3-yl)methyl)-4-methyl-4H-1,2,4-triazol-3-ol (17)
4- Etil-5-((5-metoxi-1 H-indol-3-il)metil)-4H-1 ,2,4-triazol-3-tiol (18) 4- Ethyl-5-((5-methoxy-1 H-indol-3-yl)methyl)-4H-1,2,4-triazol-3-thiol (18)
5- ((1 H-lndol-3-il)metil)-4-etil-4H-1 ,2,4-triazol-3-tiol (19) o un isómero, un solvato, o una sal farmacéuticamente aceptable del mismo. 5-((1H-lndol-3-yl)methyl)-4-ethyl-4H-1,2,4-triazol-3-thiol (19) or an isomer, a solvate, or a pharmaceutically acceptable salt thereof .
8. - Procedimiento de obtención de un compuesto de fórmula general (I I) según las reivindicaciones 1 y 2, consistente en el tratamiento del correspondiente ácido carboxílico derivado de 1 H-indol-3-ilo en acetonitrilo seco con alilamina o propargilamina en presencia de 1 ,1 '-carbonildiimidazol (CDI) y 4- (dimetilamino)piridina (DMAP). 8. - Procedure for obtaining a compound of general formula (I I) according to claims 1 and 2, consisting of the treatment of the corresponding carboxylic acid derived from 1 H-indol-3-yl in dry acetonitrile with allylamine or propargylamine in the presence of 1,1'-carbonyldiimidazole (CDI) and 4-(dimethylamino)pyridine (DMAP).
9. - Procedimiento de obtención de un compuesto de fórmula general (III) según las reivindicaciones 1 y 3, consistente en el tratamiento de la correspondiente N-(prop-2-inil) alquilamida derivada de 1 H-indol-3-ilo en diclorometano seco con cloruro de oro (III). 9. - Procedure for obtaining a compound of general formula (III) according to claims 1 and 3, consisting of the treatment of the corresponding N-(prop-2-ynyl) alkylamide derived from 1 H-indol-3-yl in dry dichloromethane with gold(III) chloride.
10. - Procedimiento de obtención de un compuesto de fórmula general (IV) según las reivindicaciones 1 y 4, consistente en la reacción de un alquil éster del correspondiente ácido carboxílico derivado de 1 H-indol-3-ilo con acetamidoxima en presencia de hidruro sódico. 10. - Procedure for obtaining a compound of general formula (IV) according to claims 1 and 4, consisting of the reaction of an alkyl ester of the corresponding carboxylic acid derived from 1 H-indol-3-yl with acetamidoxime in the presence of sodium hydride.
11. - Procedimiento de obtención de un compuesto de fórmula general (V) según las reivindicaciones 1 y 5, consistente en la reacción del correspondiente ácido carboxílico derivado de 1 H-indol-3-ilo en acetonitrilo seco con la correspondiente hidrazida sustituida en presencia de 1 ,1 '-carbonildiimidazol (CDI) y 4-(dimetilamino)piridina (DMAP), seguida del tratamiento con oxicloruro de fósforo. 11. - Procedure for obtaining a compound of general formula (V) according to claims 1 and 5, consisting of the reaction of the corresponding carboxylic acid derived from 1 H-indol-3-yl in dry acetonitrile with the corresponding substituted hydrazide in the presence of 1,1'-carbonyldiimidazole (CDI) and 4-(dimethylamino)pyridine (DMAP), followed by treatment with phosphorus oxychloride.
12. - Procedimiento de obtención de un compuesto de fórmula general (VI) según las reivindicaciones 1 y 6, consistente en reacción de la hidrazida del correspondiente ácido carboxílico derivado de 1 H-indol-3-ilo con un isocianato o isotiocianato sustituido en una mezcla de ácido acético y agua, seguida del tratamiento con hidróxido sódico en etanol. 12. - Procedure for obtaining a compound of general formula (VI) according to claims 1 and 6, consisting of the reaction of the hydrazide of the corresponding carboxylic acid derived from 1 H-indol-3-yl with an isocyanate or isothiocyanate substituted in a mixture of acetic acid and water, followed by treatment with sodium hydroxide in ethanol.
13. - Composición farmacéutica que comprende un compuesto de fórmula (I) a (VI) según cualquiera de las reivindicaciones de la 1 a la 7. 13. - Pharmaceutical composition comprising a compound of formula (I) to (VI) according to any of claims 1 to 7.
14.- Composición según la reivindicación 13 que comprende además otro principio activo. 14.- Composition according to claim 13 that also comprises another active ingredient.
15.- Composición farmacéutica según las reivindicaciones 13 y 14 adecuada para la administración oral. 15.- Pharmaceutical composition according to claims 13 and 14 suitable for oral administration.
16. - Uso de compuestos neurogénicos basados en melatonina según una cualquiera de las reivindicaciones 1 a 7 en la preparación de un medicamento o una composición farmacéutica para el tratamiento de enfermedades del sistema nervioso que se seleccionan entre las enfermedades neurodegenerativas, los trastornos cognitivos, las enfermedades o trastornos psiquiátricos, los traumas o lesiones celulares, las afecciones neurológicas y las enfermedades o trastornos relacionados con el ciclo circadiano. 16. - Use of neurogenic compounds based on melatonin according to any one of claims 1 to 7 in the preparation of a medicine or a pharmaceutical composition for the treatment of diseases of the nervous system that are selected from neurodegenerative diseases, cognitive disorders, psychiatric diseases or disorders, cellular trauma or injuries, neurological conditions and diseases or disorders related to the circadian cycle.
17. - Uso según la reivindicación 16, donde la enfermedad neurodegenerativa se selecciona entre la enfermedad de Alzheimer, enfermedad de Creutzfeld-17. - Use according to claim 16, wherein the neurodegenerative disease is selected from Alzheimer's disease, Creutzfeld's disease.
Jacob, enfermedad de Parkinson, amiloidosis sistémica, esclerosis lateral amiotrófica, enfermedad degenerativa de la retina, la parálisis cerebral o una combinación de las mismas. Jacob, Parkinson's disease, systemic amyloidosis, amyotrophic lateral sclerosis, retinal degenerative disease, cerebral palsy or a combination thereof.
18.- Uso según la reivindicación 16, donde el trastorno cognitivo se selecciona entre alteración de memoria, pérdida de memoria separada de la demencia, deterioro cognitivo leve, disminución cognitiva relacionada con la edad, pérdida de memoria por déficit de atención, deterioro cognitivo como consecuencia del uso de anestésicos generales, de quimioterapia o de radioterapia, deterioro cognitivo asociado a trauma post-quirúrgico o a intervención terapéutica, declive cognitivo asociado con la enfermedad de Alzheimer o con epilepsia, demencia senil, demencia vascular, delirio o una combinación de las mismas. 18.- Use according to claim 16, wherein the cognitive disorder is selected from memory impairment, memory loss separate from dementia, mild cognitive impairment, age-related cognitive decline, memory loss due to attention deficit, cognitive impairment such as consequence of the use of general anesthetics, chemotherapy or radiotherapy, cognitive impairment associated with post-surgical trauma or therapeutic intervention, cognitive decline associated with Alzheimer's disease or with epilepsy, senile dementia, vascular dementia, delirium or a combination thereof .
19.- Uso según la reivindicación 16, donde la enfermedad o el trastorno psiquiátrico se selecciona entre depresión, depresión mayor, depresión neurótica, depresión provocada por el consumo de drogas o alcohol, depresión post-traumática, depresión post-parto, ansiedad, trastorno obsesivo- compulsivo, trastorno bipolar, fobia social, trastorno del estado de ánimo estacional o una combinación de las mismas. 19.- Use according to claim 16, wherein the psychiatric disease or disorder is selected from depression, major depression, neurotic depression, depression caused by drug or alcohol consumption, post-traumatic depression, postpartum depression, anxiety, disorder obsessive-compulsive disorder, bipolar disorder, social phobia, seasonal mood disorder, or a combination thereof.
20. - Uso según la reivindicación 16, donde el trauma o lesión celular se selecciona entre trauma o lesión neurológica, cirugía cerebral o de la médula espinal, lesión de la retina, lesiones relacionadas con epilepsia, lesiones cerebrales o de la médula espinal en relación con el tratamiento del cáncer, lesiones cerebrales o de la médula espinal relacionadas con infecciones, procesos inflamatorios, toxinas ambientales, episodios isquémicos, o una combinación de las mismas. 20. - Use according to claim 16, wherein the cellular trauma or injury is selected from neurological trauma or injury, brain or spinal cord surgery, retinal injury, epilepsy-related injuries, brain or spinal cord injuries in relation to with the treatment of cancer, brain or spinal cord injuries related to infections, inflammatory processes, environmental toxins, ischemic events, or a combination thereof.
21. - Uso según la reivindicación 16, donde la afección neurológica se selecciona entre el trastorno del aprendizaje, autismo, trastorno por déficit de atención, narcolepsia, trastorno del sueño, epilepsia, epilepsia del lóbulo temporal o una combinación de las mismas. 21. - Use according to claim 16, wherein the neurological condition is selected from learning disorder, autism, attention deficit disorder, narcolepsy, sleep disorder, epilepsy, temporal lobe epilepsy or a combination thereof.
22. - Uso según la reivindicación 16, donde la enfermedad o trastorno relacionado con el ciclo circadiano se selecciona entre los trastornos del sueño, fatiga diurna, pérdida de eficacia mental, debilidad e irritabilidad y el síndrome transoceánico o una combinación de las mismas. 22. - Use according to claim 16, wherein the disease or disorder related to the circadian cycle is selected from sleep disorders, daytime fatigue, loss of mental efficiency, weakness and irritability and transoceanic syndrome or a combination thereof.
23. - Uso de un compuesto según una cualquiera de las reivindicaciones de la 1 a la 7 como reactivo en ensayos biológicos. 23. - Use of a compound according to any one of claims 1 to 7 as a reagent in biological assays.
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