WO2012014993A1 - Agent for increasing translation initiation factor or translation elongation factor, and use thereof for medical purposes - Google Patents

Agent for increasing translation initiation factor or translation elongation factor, and use thereof for medical purposes Download PDF

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WO2012014993A1
WO2012014993A1 PCT/JP2011/067315 JP2011067315W WO2012014993A1 WO 2012014993 A1 WO2012014993 A1 WO 2012014993A1 JP 2011067315 W JP2011067315 W JP 2011067315W WO 2012014993 A1 WO2012014993 A1 WO 2012014993A1
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diseases
translation
factor
translation initiation
disease
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French (fr)
Japanese (ja)
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謙二 橋本
環 石間
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国立大学法人 千葉大学
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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
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/47—Quinolines; Isoquinolines
    • A61K31/4709—Non-condensed quinolines and containing further heterocyclic rings
    • 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
    • A61K31/65—Tetracyclines
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/18—Antipsychotics, i.e. neuroleptics; Drugs for mania or schizophrenia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P5/00—Drugs for disorders of the endocrine system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P7/00—Drugs for disorders of the blood or the extracellular fluid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links

Definitions

  • the present invention prevents and / or prevents various diseases such as psychiatric diseases, neurological diseases, cerebrovascular diseases, cardiovascular diseases, respiratory diseases, endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors. It relates to a medicament for treatment. More specifically, the present invention relates to a pharmaceutical preparation containing a compound that increases a translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof.
  • Drug treatment is indispensable for the treatment of mental disorders such as depression and schizophrenia, such as antidepressants (tricyclic antidepressants, selective serotonin reuptake inhibitors, serotonin / norepinephrine reuptake inhibitors, etc.) ) And antipsychotic drugs (phenothiazine compounds, butyrophenone compounds, benzamide compounds, iminodibenzyl compounds, thiepine compounds, indole compounds, serotonin / dopamine receptor blockers, etc.).
  • antidepressants tricyclic antidepressants, selective serotonin reuptake inhibitors, serotonin / norepinephrine reuptake inhibitors, etc.
  • antipsychotic drugs phenothiazine compounds, butyrophenone compounds, benzamide compounds, iminodibenzyl compounds, thiepine compounds, indole compounds, serotonin / dopamine receptor blockers, etc.
  • tetracycline antibiotics are a general term for a group of broad-spectrum antibiotics, and are inexpensively prescribed for Chlamydia infection, rickettsia infection, mycoplasma infection, brucellosis, and spirochete infection (syphilis / Lyme disease). It is a drug.
  • Minocycline one of the tetracycline antibiotics
  • various actions such as neuroprotective action and anti-inflammatory action in addition to its action as an antibiotic.
  • Therapeutic effects on various diseases such as autoimmune diseases and human immunodeficiency virus (HIV) infection have been reported (Non-Patent Documents 1 to 7).
  • the inventors have reported that the second generation antibiotic minocycline or the antiplatelet drug cilostazol has an improving effect in studies such as model animals of schizophrenia and neuroplasticity using cell culture tests.
  • minocycline has an effect of improving cognitive dysfunction caused by administration of the NMDA receptor antagonist phencyclidine and prepulse depressive disorder caused by administration of the NMDA receptor antagonist MK-801 (Non-patent Documents 8 and 9). Furthermore, from a placebo-controlled double-blind study, it has been reported that minocycline significantly improves negative symptoms and cognitive impairment in schizophrenic patients (Non-patent Document 10). In addition, it has been reported that minocycline has an action to improve depressive symptoms in patients with bipolar disorder (Non-patent Document 11).
  • Non-patent Documents 12 and 13 the mechanism of action of minocycline and cilostazol is unclear.
  • PC12 cells which are pheochromocytomas derived from rat adrenal medulla, extend dendrites like nerves by the action of nerve growth factor (Nerve Growth Factor; NGF). Widely used. NGF-induced neurite outgrowth is an experimental system widely used as a treatment mechanism for various diseases (Non-Patent Documents 14 and 15).
  • the regulation of protein expression is closely related to cell growth and cell cycle progression. Therefore, it is expected that abnormalities in the translation process are related not only to the etiology / pathology of human diseases but also to the action of therapeutic agents.
  • Translation of mRNA consists of stages such as initiation, elongation and termination, and many eukaryotic translation initiation factors (eIFs) act specifically at the initiation stage. .
  • the translation initiation factor is also referred to as a polypeptide chain initiation factor, and is a protein group (eIF1, eIF2, eIF3, eIF4, eIF5, etc.) in which a liposome binds amino acids according to the base sequence of mRNA, that is, a translation process of protein synthesis.
  • the final purpose of initiating eIFs and translation is to form the 80S ribosome and place the mRNA start codon, Met-tRNA, on the ribosome P site (peptidyl tRNA binding site).
  • the translation elongation factor is also referred to as a polypeptide chain elongation factor, and is a protein group (eEF1, eEF2, eEF3, eEF4, eEF5, etc.) necessary for the translation process of polypeptide chain elongation that biosynthesizes a protein from mRNA.
  • Translation elongation factors are broadly divided into two types, functionally binding aminoacyl-tRNAs corresponding to the mRNA code to the A-site (aminoacyl-tRNA binding site) of the ribosome and present at the A-site immediately after peptidyl bond formation. It is a factor that translocates peptidyl tRNA to P site. Regulation of translation initiation factor and translation elongation factor is an important process in the sense that it regulates the expression level of the final protein, and changes in mRNA translation initiation and translation elongation control cause diseases in mammals including humans Understanding the control of the translation process is not only to elucidate the causes of various diseases, but also to develop new preventives, diagnostics, and therapeutics. It can be applied.
  • DomercqM Matute C. (2004). Neuroprotection by tetracyclines. Trends Pharmacol Sci 25l: 609-612. Yong VW, Wells J, Giuliani F, Casha S, Power C, Metz LM. (2004). The promise ofminocycline in neurology. Lancet Neurol 3: 744-751. Blum D, Chtarto A, Tenenbaum L, Brotchi J, Levivier M. (2004). Clinical potential ofminocycline for neurodegenerative disorders. Neurobiol Dis 217: 359-366. Zemke D, Majid A. (2004). The potential of minocycline for neuroprotection in humanneurologic diseases. Neurobiol Dis 217: 359-366. MiyaokaT. (2008).
  • Noble W Garwood CJ, JHanger DP.
  • OrsucciD Calsolaro V, Mancuso M, Siciliano G. (2009) .2009Neuroprotective effects oftetracyclines: molecular targets, animal models and human disease.
  • the subject of this invention is providing the pharmaceutical formulation effective in various diseases, especially a neurological disease.
  • the present inventors have conducted intensive studies to solve the above problems, and that minocycline or cilostazol has an action of significantly enhancing the NGF-induced neurite outgrowth action in PC12 cells (referred to as neurite outgrowth enhancement action).
  • minocycline or cilostazol has an action of significantly enhancing the NGF-induced neurite outgrowth action in PC12 cells (referred to as neurite outgrowth enhancement action).
  • Tecycline another second-generation antibiotic, has not been effective.
  • the main effect of cilostazol is phosphodiesterase (PDE) 3 inhibitory action, but the other PDE3 inhibitor cilostamide did not show NGF-induced neurite outgrowth effect. May not be related to the PDE3 inhibitory action of cilostazol.
  • the present inventors proceeded to elucidate the mechanism of action of minocycline or cilostazol on enhancing the NGF-induced neurite outgrowth, and the increase of translation initiation factor (eIF4A1 etc.) or translation elongation factor (eEF1A1 etc.) was involved in the action, respectively. It was found for the first time that the present invention was completed.
  • the present invention relates to a pharmaceutical preparation containing a compound having an action of increasing a translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof.
  • the present invention also relates to therapeutic agents for various diseases in which abnormalities of translation initiation factors or translation elongation factors are involved, and further to pharmaceutical formulations effective for neurological diseases.
  • the present invention relates to 1) a neurite outgrowth enhancer containing minocycline and / or cilostazol.
  • the present invention also relates to a neurite elongation enhancer characterized by increasing 2) a translation initiation factor and / or a translation elongation factor.
  • the present invention also relates to a neurite elongation enhancer characterized by increasing eIF4A isoform that is a translation initiation factor and / or eEF1A isoform that is a translation elongation factor.
  • the present invention also relates to 6) a translation initiation factor increasing agent and / or a translation elongation factor increasing agent comprising minocycline and / or cilostazol. Further, it relates to an agent for increasing 7) translation initiation factor eIF4A1 containing minocycline. Furthermore, it relates to an agent for increasing translation elongation factor eEF1A1 containing 8) cilostazol.
  • the present invention also relates to 9) a preventive and / or therapeutic agent for a disease caused by an abnormality in a translation initiation factor and / or translation elongation factor, containing minocycline and / or cilostazol.
  • Psychiatric diseases neurological diseases, cerebrovascular diseases, cardiovascular diseases derived from a decrease in eIF4A isoform which is a translation initiation factor and / or eEF1A isoform which is a translation elongation factor, containing minocycline and / or cilostazol.
  • an agent for preventing and / or treating a disease exhibiting one or more symptoms selected from the group consisting of endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors are examples of a preventive and / or therapeutic agent for a disease exhibiting one or more symptoms selected from the group consisting of respiratory diseases, endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors.
  • minocycline or cilostazol, or a pharmaceutically acceptable salt thereof significantly increased translation initiation factor (eIF4A1) or translation elongation factor (eEF1A1), respectively, compounds that increase translation initiation factor or translation elongation factor are It is effective for various diseases, diseases associated with abnormalities of translation initiation factors or translation elongation factors, and effective for the prevention and / or treatment thereof.
  • various diseases in which abnormalities of translation initiation factors or translation elongation factors have been pointed out and diseases associated with abnormal protein formation such as mental diseases, neurological diseases, cerebrovascular diseases, cardiovascular diseases, respiratory organs It is effective for the prevention and / or treatment of various diseases such as diseases, endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors.
  • FIG. 1 shows the effect of minocycline on nerve growth factor-induced neurite action.
  • Minocycline significantly enhanced NGF-induced neurite outgrowth in PC12 cells in a concentration-dependent manner, whereas other antibiotic tetracyclines did not affect the effect.
  • FIG. 2 is a diagram showing intracellular expression levels of eIF4A1 protein treated with minocycline and tetracycline. Minocycline significantly increased eIF4A1 protein expression.
  • FIG. 3 is a diagram showing the effect of siRNA treatment and the like on the increase in the expression level of intracellular translation initiation factor eIF4A1 protein of minocycline. The eIF4A1 siRNA treatment significantly decreased the protein expression level of eIF4A1. Negative RNA had no effect.
  • FIG. 1 shows the effect of minocycline on nerve growth factor-induced neurite action.
  • Minocycline significantly enhanced NGF-induced neurite outgrowth in PC12 cells in a concentration-dependent manner, whereas other antibiotic tetracyclines
  • FIG. 4 shows the effect of eIF4A1 on the NGF-induced neurite outgrowth action of minocycline.
  • FIG. 5 shows the effect of cilostazol on the NGF-induced neurite outgrowth action. Cilostazol significantly enhanced NGF-induced neurite outgrowth in PC12 cells in a concentration-dependent manner, whereas the other PDE3 inhibitor cilostamide did not affect the effect.
  • FIG. 6 is a graph showing the intracellular expression level of eEF1A1 protein treated with cilostazol and cilostamide.
  • FIG. 7 is a diagram showing the influence of siRNA treatment or the like on the increase in the expression level of cilostazol translation initiation factor eEF1A1 protein in cells.
  • the eEF1A1 siRNA treatment significantly decreased the protein expression level of eEF1A1.
  • Negative RNA had no effect.
  • FIG. 8 is a graph showing the effect of eIF1A1 on the NGF-induced neurite outgrowth action of cilostazol. Under the conditions shown in FIG. 7, when the protein expression level of the translation initiation factor eEF1A1 was decreased by siRNA treatment, the NGF-induced neurite outgrowth action of cilostazol was suppressed.
  • FIG. 7 is a diagram showing the influence of siRNA treatment or the like on the increase in the expression level of cilostazol translation initiation factor eEF1A1 protein in cells.
  • the eEF1A1 siRNA treatment significantly decreased the protein expression level of eEF1A1.
  • Negative RNA had no effect.
  • FIG. 9 shows the effects of tetracycline and minocycline on eEF1A1 protein expression, and the effects of cilostamide and cilostazol on eIF4A1 protein expression.
  • Tetracycline and minocycline did not affect eEF1A1 protein expression, and cilostamide and cilostazol did not affect eIF4A1 protein expression.
  • FIG. 10 is a diagram showing the nucleotide sequence of siRNA of eIF4A1 as an antisense.
  • FIG. 11 is a diagram showing the nucleotide sequence of eEF1A1 siRNA as antisense.
  • the translation initiation factor is a protein group that initiates the process of ribosome binding amino acids according to the base sequence of mRNA, that is, the translation process of protein synthesis.
  • Translation elongation factors are a group of proteins that control the process involved in peptide chain elongation, which advances the translation process of protein synthesis from mRNA.
  • a protein called eIF4A is known to have several isoforms (or subclasses) such as eIF4A1, eIF4A2, and eIF4A3, their functions are considered to be the same, and the effects of the present invention can be achieved.
  • the invention can be implemented for one or more isoforms.
  • eIF4A isoforms.
  • eIF4A1 can be mentioned.
  • the protein called eEF1A among the translation elongation factors is known to have several isoforms (or subclasses) such as eEF1A1, eEF1A2, and eEF1A3, and their functions are considered to be the same.
  • the invention can be implemented for one or more isoforms.
  • these proteins are referred to, they are referred to as eEF1A isoforms.
  • eEF1A1 can be mentioned as a preferred eEF1A isoform.
  • the neurite outgrowth action refers to the action of causing neurite formation in nerve cells, and the enhancement action (neurite outgrowth enhancement action) is, for example, the effect of performing a quantification test as described in the Examples. Can be determined.
  • a compound having a neurite outgrowth enhancing action and having the effect sufficient as a medicine is called a neurite outgrowth enhancer.
  • the present invention relates to a neurite elongation enhancer characterized by increasing a translation initiation factor and / or a translation elongation factor. More preferably, the present invention relates to a neurite extension enhancer characterized by increasing eIF4A isoform which is a translation initiation factor and / or eEF1A isoform which is a translation elongation factor. More preferably, the present invention relates to a neurite outgrowth enhancer characterized by increasing eIF4A which is a translation initiation factor and / or eEF1A which is a translation elongation factor.
  • Minocycline can be used as an enhancer of a translation initiation factor, preferably an eIF4A isoform, more preferably eIF4A1. It can be used as an increaser for cilostazol, a translation elongation factor, preferably an eEF1A isoform, more preferably eEF1A1.
  • the pharmaceutical preparation of the present invention containing a compound that increases a translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof, such as a prophylactic agent and / or therapeutic agent, is administered orally or parenterally.
  • a pharmaceutically acceptable salt thereof such as a prophylactic agent and / or therapeutic agent
  • parenteral administration may be performed by rectal administration using suppositories, transdermal administration using patches, liniments, gels, etc., transmucosal administration using sprays, aerosols, etc. It may be administered internally, intramuscularly, subcutaneously or intraventricularly.
  • the compound that increases the translation initiation factor or translation elongation factor can be used in the form of both a free base or a pharmaceutically acceptable salt thereof.
  • the pharmaceutically acceptable salt is preferably a pharmaceutically acceptable acid addition salt, more preferably a hydrochloride.
  • the preventive agent and / or therapeutic agent of the present invention may contain other medicinal ingredients in addition to a compound that increases a translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof.
  • an antioxidant in addition to these medicinal ingredients, an antioxidant, a stabilizer, an antiseptic, a corrigent, a coloring agent, a solubilizer, a solubilizer, a surfactant, an emulsifier, and an antifoaming agent depending on the dosage form as appropriate.
  • suitable physiologically acceptable substances well known to those skilled in the art, such as viscosity modifiers, gelling agents, absorption enhancers, dispersants, excipients, and pH adjusters.
  • the preventive agent and / or therapeutic agent of the present invention is produced by appropriately combining a compound that increases a translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof, and these substances.
  • a compound that increases a translation initiation factor or translation elongation factor or a pharmaceutically acceptable salt thereof, and these substances.
  • the prophylactic and / or therapeutic agent of the present invention is prepared as a preparation for injection (subcutaneous injection, intramuscular injection, or intravenous injection), the form of a solution or suspension is preferred, and for vaginal or rectal administration
  • Semi-solid dosage forms such as creams or suppositories are preferred, and for nasal administration, powder, nasal drops, or aerosol dosage forms are preferred.
  • Injectable preparations can contain, for example, plasma-derived proteins such as albumin, amino acids such as glycine, and sugars such as mannitol as carriers, and buffers, solubilizers, isotonic agents, and the like can also be added. it can.
  • plasma-derived proteins such as albumin
  • amino acids such as glycine
  • sugars such as mannitol as carriers
  • buffers solubilizers, isotonic agents, and the like can also be added. it can.
  • surfactants such as Tween (trademark) 80 and Tween (trademark) 20, in order to prevent aggregation.
  • parenteral dosage forms other than injectable preparations may contain distilled water or physiological saline, polyalkylene glycols such as polyethylene glycol, oils of plant origin, hydrogenated naphthalene, and the like.
  • preparations for vaginal or rectal administration such as suppositories contain, for example, polyalkylene glycol, petrolatum, cacao oil and the like as common excipients.
  • Vaginal preparations may contain absorption enhancers such as bile salts, ethylenediamine salts, and citrate salts.
  • Inhalation formulations may be solid, may contain, for example, lactose as an excipient, and nasal drops may be water or oil solutions.
  • the exact dose and administration schedule of the preventive agent and / or therapeutic agent of the present invention can be adjusted depending on the required amount, treatment method, disease or degree of necessity for each individual treatment target.
  • the dosage can be determined according to age, weight, general health condition, sex, diet, administration time, administration method, excretion rate, combination of drugs, patient condition, etc. It may be determined in consideration of these factors.
  • a compound that increases translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof, has no problem in pharmacokinetics in humans, and can be used safely.
  • a compound (free base) that increases translation initiation factor or translation elongation factor parenterally, subcutaneously, intravenously, intramuscularly or rectally, depending on the condition, body weight, type of compound, administration route, etc. It is administered at about 0.01 to 1000 mg / person / day, preferably 0.1 to 500 mg / person / day, and orally about 0.01 to 500 mg / person / day, preferably 0.1 to 100 mg. It is desirable to be administered per person / day.
  • the present invention relates to a method for enhancing neurite outgrowth, which comprises administering an agent for increasing a translation initiation factor and / or an agent for increasing a translation elongation factor to a subject.
  • the present invention also relates to the use of an agent for increasing translation initiation factor and / or an agent for increasing translation elongation factor in the method for enhancing neurite elongation and the production of neurite elongation enhancer.
  • the present invention relates to a method for preventing and / or treating a mental illness characterized by enhancing neurite outgrowth by administering to a subject an agent for increasing a translation initiation factor and / or an agent for increasing a translation elongation factor.
  • the present invention also relates to the use of an agent for increasing a translation initiation factor and / or an agent for increasing a translation elongation factor in the production of a method for preventing and / or treating mental illness and a prophylactic and / or therapeutic agent for mental illness.
  • morphometric analysis was performed using digital images of live cells taken with a phase contrast inverted microscope equipped with a camera. . Three fields of view were taken per well. The average number of cells per field was 100. The number of differentiated cells in each visual field was counted by visual recognition. Only cells having one or more neurites with a length equal to the cell body diameter were counted and expressed as a percentage of the total number of cells in the field of view. Measurements were performed blind.
  • Rabbit anti-eIF4A antibody (Abcam) and mouse anti- ⁇ -actin antibody (Sigma Aldrich) diluted at 1: 250 and 1: 10000 in the blocking buffer were used as primary antibodies.
  • an anti-rabbit IgG horseradish peroxidase labeled antibody (GE Healthcare) diluted 25,000 times in TBST containing 5% skim milk and an anti-mouse IgG horseradish peroxidase labeled antibody (GE Health) diluted 25000 times in TBST containing 5% skim milk. Care) was used as the secondary antibody.
  • Antigen proteins on the membrane were detected with ECL TM Plus Western Blotting Detection System (GE Healthcare).
  • PC12 cells were cultured and transfected with the above siRNA using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) and OPTI-MEM (Invitrogen, Carlsbad, CA, USA). 6 hours later, NGF (2.5 ng / ml), and further exchange with DMEM medium containing 0.5% FBA, 1% penicillin / streptomycin with or without the addition of various drugs, to examine the neurite outgrowth enhancing effect And subjected to Western blot analysis. (5) Statistical analysis Data are shown as mean ⁇ standard deviation. Statistical analysis was performed using one-way analysis of variance and post-hoc Bonferroni test. A p value of 0.05 or less was considered statistically significant.
  • siRNA Minocycline or cilostazol significantly increased eIF4A1 or eEF1A1 in PC12 cells, respectively, and when gene was knocked down by the respective siRNA, an increase in eIF4A1 or eEF1A1 in PC12 cells was seen There wasn't. Furthermore, even if siRNA (negative RNA) that had nothing to do with the target was allowed to act, the increase in eIF4A1 or eEF1A1 in PC12 cells was not affected (FIGS. 3 and 7).
  • minocycline or cilostazol significantly increased the number of PC12 cells that showed neurite outgrowth by (2.5 ng / mL) in a concentration-dependent manner, but neurite outgrowth when the gene was knocked down by siRNA treatment Did not increase the number of cells that showed. Furthermore, even when siRNA (negative RNA) that had nothing to do with the target was allowed to act, the increase in the number of cells showing neurite outgrowth was not affected (FIGS. 4 and 8). On the other hand, tetracycline or minocycline did not affect the amount of eEF1A1 protein (FIG. 9). Moreover, cilostamide or cilostazol did not affect the amount of eIF4A1 protein (FIG. 9).
  • minocycline or cilostazol showed a significant enhancing action in the NGF-induced neurite elongation action model, and that the translation initiation factor or translation elongation factor was involved in the enhancement action. It will be appreciated that compounds that increase translation elongation factors will be useful as new therapeutic agents in the future.
  • a compound of the present invention containing a compound that increases a translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof is used in mental diseases, neurological diseases, cerebrovascular diseases, cardiovascular diseases, respiratory organs It is effective as a prophylactic and / or therapeutic agent for various diseases including diseases, endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors.
  • minocycline and / or cilostazol can be used as neurite outgrowth enhancers.
  • a neurite outgrowth enhancer is accompanied by a specific increase in translation initiation factors, particularly eIF4A1, in minocycline, and a specific increase in translation elongation factors, particularly eEF1A1, in cilostazol. It was a thing. Furthermore, from these, it is possible to use minocycline as a translation initiation factor increasing agent, particularly eIF4A1 increasing agent, and cilostazol as a translation elongation factor increasing agent, particularly eEF1A1 increasing agent. It was.
  • minocycline and / or cilostazol can be used as a preventive and / or therapeutic agent for various diseases caused by abnormal translation initiation factors and / or translation elongation factors, particularly neurological diseases.
  • the compound that increases the translation initiation factor or translation elongation factor of the present invention is a mental disease, neurological disease, cerebrovascular disease, cardiovascular disease, respiratory disease, endocrine disease, metabolic disease. It can be used as a novel pharmaceutical effective for various diseases such as blood diseases, immune diseases, infectious diseases, and malignant tumors.
  • SEQ ID NO: 1 designed oligonucleotide.
  • SEQ ID NO: 2 designed oligonucleotide.
  • SEQ ID NO: 3 Designed oligonucleotide.
  • SEQ ID NO: 4 Designed oligonucleotide.
  • SEQ ID NO: 5 designed oligonucleotide.
  • SEQ ID NO: 6 designed oligonucleotide.
  • SEQ ID NO: 7 designed oligonucleotide.
  • SEQ ID NO: 8 designed oligonucleotide.
  • SEQ ID NO: 9 designed oligonucleotide.
  • SEQ ID NO: 10 designed oligonucleotide.
  • SEQ ID NO: 11 designed oligonucleotide.
  • SEQ ID NO: 12 designed oligonucleotide.

Abstract

[Problem] The purpose of the present invention is to provide a pharmaceutical preparation for preventing and/or treating various diseases, which comprises a compound capable of increasing a translation initiation factor or a transcription elongation factor or a pharmaceutically acceptable salt thereof as an active ingredient. [Solution] The present inventors made extensive studies for the purpose of solving the problem, and it is found for the first time that the increase in a translation initiation factor (e.g., eIF4A1) or a translation elongation factor (e.g., eEF1A1) is involved in the NGF-induced neurite elongation potentiating activity of minocycline or cilostazol, and the invention has been accomplished.

Description

翻訳開始因子または翻訳伸長因子の増加剤、およびその医薬用途Translation initiation factor or translation elongation factor increasing agent and pharmaceutical use thereof
　本発明は、精神疾患、神経疾患、脳血管疾患、循環器疾患、呼吸器疾患、内分泌疾患、代謝疾患、血液疾患、免疫疾患、感染症、および悪性腫瘍などの様々な疾患の予防および／または治療のための医薬に関する。さらに詳しくは、翻訳開始因子もしくは翻訳伸長因子を増加させるような化合物、またはその薬学的に許容し得る塩を含有する、医薬製剤に関する。 The present invention prevents and / or prevents various diseases such as psychiatric diseases, neurological diseases, cerebrovascular diseases, cardiovascular diseases, respiratory diseases, endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors. It relates to a medicament for treatment. More specifically, the present invention relates to a pharmaceutical preparation containing a compound that increases a translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof.
　社会生活様式の変化や社会の高齢化に伴い、精神疾患、神経疾患、脳血管疾患、循環器疾患、呼吸器疾患、内分泌疾患、代謝疾患、血液疾患、免疫疾患、感染症、悪性腫瘍などの様々な疾患は、全体として増加する傾向にある。例えば、代表的な精神疾患であるうつ病や統合失調症は発症率が高く、医療経済という点からも大きな問題となっている。うつ病や統合失調症などの精神疾患の治療には、薬物治療が不可欠であり、抗うつ薬（三環系抗うつ薬、選択的セロトニン再取り込み阻害薬、およびセロトニン・ノルエピネフリン再取り込み阻害薬など）や抗精神病薬（フェノチアジン系化合物、ブチロフェノン系化合物、ベンズアミド系化合物、イミノジベンジル系化合物、チエピン系化合物、インドール系化合物およびセロトニン・ドーパミン受容体遮断薬など）などが投与されている。しかしながら、臨床の場で実際に使用されているこれらの薬剤は、一部の患者や一部の症状には有効であるが、これらの薬剤では効果が無い、いわゆる治療抵抗性の患者が存在する事も知られており、新しい治療薬の開発が切望されている。既存の薬物では、これらの精神疾患に対する十分な治療効果が出ているとは言い難く、現実的には有効な予防法や治療法はほとんど無いのが現状である。
　さらに、精神疾患以外の疾患、例えば、神経疾患、脳血管疾患、循環器疾患、呼吸器疾患、内分泌疾患、代謝疾患、血液疾患、免疫疾患、感染症、および悪性腫瘍などの様々な他の疾患においても、既存の薬物では、十分な治療効果が出ているとは言い難く、新しい治療薬の開発が切望されている。 With changes in social lifestyles and aging of society, mental diseases, neurological diseases, cerebrovascular diseases, cardiovascular diseases, respiratory diseases, endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, malignant tumors, etc. Various diseases tend to increase as a whole. For example, depression and schizophrenia, which are typical mental illnesses, have a high incidence and are a major problem in terms of medical economy. Drug treatment is indispensable for the treatment of mental disorders such as depression and schizophrenia, such as antidepressants (tricyclic antidepressants, selective serotonin reuptake inhibitors, serotonin / norepinephrine reuptake inhibitors, etc.) ) And antipsychotic drugs (phenothiazine compounds, butyrophenone compounds, benzamide compounds, iminodibenzyl compounds, thiepine compounds, indole compounds, serotonin / dopamine receptor blockers, etc.). However, these drugs that are actually used in clinical settings are effective for some patients and some symptoms, but there are so-called treatment-resistant patients that are ineffective with these drugs. Things are also known, and the development of new therapeutic agents is eagerly desired. It is difficult to say that existing drugs have a sufficient therapeutic effect on these psychiatric disorders, and there are practically no effective preventive or therapeutic methods in reality.
In addition, various other diseases such as diseases other than mental diseases such as neurological diseases, cerebrovascular diseases, cardiovascular diseases, respiratory diseases, endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors However, it is difficult to say that existing drugs have sufficient therapeutic effects, and the development of new therapeutic drugs is eagerly desired.
　このように、うつ病や統合失調症などの精神疾患、さらには神経疾患、脳血管疾患、循環器疾患、呼吸器疾患、内分泌疾患、代謝疾患、血液疾患、免疫疾患、感染症、および悪性腫瘍などの様々な他の疾患の予防や治療に使用されている薬剤では十分な治療効果が出ているとは言い難い状況がある。
　ところで、テトラサイクリン系抗生物質は一群の広域スペクトラム抗生物質の総称であり、クラミジア感染症、リケッチア感染症、マイコプラズマ感染症、ブルセラ症、およびスピロヘータ感染症（梅毒・ライム病）などで処方される安価な薬剤である。近年、テトラサイクリン系抗生物質の一つであるミノサイクリンが、抗生物質としての作用以外に神経保護作用および抗炎症作用などの様々な作用を有する事が知られるようになり、また精神疾患、神経変性疾患、自己免疫疾患、ヒト免疫不全ウイルス（ＨＩＶ）感染などの様々な疾患に対する治療効果が報告されている（非特許文献１から７）。
　最近、発明者らは、第二世代抗生物質ミノサイクリンまたは抗血小板薬シロスタゾールが統合失調症のモデル動物や細胞培養試験を用いた神経可塑性等の試験において改善作用があることを報告した。例えば、ミノサイクリンが、ＮＭＤＡ受容体拮抗薬フェンサイクリジン投与による認知機能障害やＮＭＤＡ受容体拮抗薬ＭＫ－８０１投与によるプレパルス抑性障害に対して改善作用を有する事を報告した（非特許文献８および９）。
　さらに、プラセボ対照二重盲検試験より、ミノサイクリンが統合失調症患者の陰性症状および認知機能障害を有意に改善する事が報告されている（非特許文献１０）。また、ミノサイクリンは双極性障害患者のうつ症状を改善する作用があることが報告されている（非特許文献１１）。
　さらに、発明者らは、シロスタゾールがフェンサイクリジン投与による認知機能障害やＮＭＤＡ受容体拮抗薬ＭＫ－８０１投与によるプレパルス抑性障害に対して改善作用を有する事を報告し、統合失調症などの精神疾患の治療薬としての可能性を報告した（非特許文献１２および１３）。
　しかしながら、現時点ではミノサイクリンおよびシロスタゾールの作用メカニズムは不明である。 Thus, mental disorders such as depression and schizophrenia, as well as neurological diseases, cerebrovascular diseases, cardiovascular diseases, respiratory diseases, endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors There are situations where it is difficult to say that drugs used for the prevention and treatment of various other diseases such as these have sufficient therapeutic effects.
By the way, tetracycline antibiotics are a general term for a group of broad-spectrum antibiotics, and are inexpensively prescribed for Chlamydia infection, rickettsia infection, mycoplasma infection, brucellosis, and spirochete infection (syphilis / Lyme disease). It is a drug. In recent years, it has been known that minocycline, one of the tetracycline antibiotics, has various actions such as neuroprotective action and anti-inflammatory action in addition to its action as an antibiotic. Therapeutic effects on various diseases such as autoimmune diseases and human immunodeficiency virus (HIV) infection have been reported (Non-Patent Documents 1 to 7).
Recently, the inventors have reported that the second generation antibiotic minocycline or the antiplatelet drug cilostazol has an improving effect in studies such as model animals of schizophrenia and neuroplasticity using cell culture tests. For example, it has been reported that minocycline has an effect of improving cognitive dysfunction caused by administration of the NMDA receptor antagonist phencyclidine and prepulse depressive disorder caused by administration of the NMDA receptor antagonist MK-801 (Non-patent Documents 8 and 9).
Furthermore, from a placebo-controlled double-blind study, it has been reported that minocycline significantly improves negative symptoms and cognitive impairment in schizophrenic patients (Non-patent Document 10). In addition, it has been reported that minocycline has an action to improve depressive symptoms in patients with bipolar disorder (Non-patent Document 11).
Furthermore, the inventors have reported that cilostazol has an improving effect on cognitive dysfunction caused by phencyclidine administration and prepulse depressive disorder caused by NMDA receptor antagonist MK-801 administration. The possibility as a therapeutic agent for diseases was reported (Non-patent Documents 12 and 13).
However, at present, the mechanism of action of minocycline and cilostazol is unclear.
　一方、ラット副腎髄質由来の褐色細胞腫であるＰＣ１２細胞は、神経成長因子（Ｎｅｒｖｅ　Ｇｒｏｗｔｈ　Ｆａｃｔｏｒ；ＮＧＦ）を作用させることにより、神経のように樹状突起を伸ばすことから、神経細胞分化のモデルとして幅広く使用されている。またＮＧＦ誘発の神経突起伸長作用は、様々な疾患の治療メカニズムとして、幅広く使用されている実験系である（非特許文献１４および１５）。 On the other hand, PC12 cells, which are pheochromocytomas derived from rat adrenal medulla, extend dendrites like nerves by the action of nerve growth factor (Nerve Growth Factor; NGF). Widely used. NGF-induced neurite outgrowth is an experimental system widely used as a treatment mechanism for various diseases (Non-Patent Documents 14 and 15).
　細胞成長や細胞周期の進展には、タンパク質発現の調節が密接に関わっている。したがって、翻訳過程の異常が、ヒトの病気の病因・病態だけでなく治療薬の作用にも関わっている事が予想される。ｍＲＮＡの翻訳は、ｉｎｉｔｉａｔｉｏｎ（開始）・ｅｌｏｎｇａｔｉｏｎ（伸長）・ｔｅｒｍｉｎａｔｉｏｎ（終結）といった段階からなり、多くの真核生物の翻訳開始因子（ｅＩＦｓ；ｅｕｋａｒｙｏｔｉｃ　ｉｎｉｔｉａｔｉｏｎ　ｆａｃｔｏｒｓ）は、開始段階で特異的に作用する。翻訳開始因子は、ポリペプチド鎖開始因子とも称され、リポゾームがｍＲＮＡの塩基配列に従ってアミノ酸を結合させる過程、すなわちタンパク質合成の翻訳過程を開始させるタンパク質群（ｅＩＦ１、ｅＩＦ２、ｅＩＦ３、ｅＩＦ４、ｅＩＦ５など）をいう。ｅＩＦｓや 翻訳開始の最終目的は、８０Ｓリボソームの形成とリボソームのＰ部位（ぺプチジルｔＲＮＡ結合部位）上にｍＲＮＡの開始コドン、Ｍｅｔ－ｔＲＮＡの配置をすることである。翻訳伸長因子は、ポリペプチド鎖伸長因子とも称され、ｍＲＮＡからタンパク質を生合成させるポリペプチド鎖伸長の翻訳過程に必要なタンパク質群（ｅＥＦ１、ｅＥＦ２、ｅＥＦ３、ｅＥＦ４、ｅＥＦ５など）である。翻訳伸長因子は、機能的に大別して２種類に分けられ、ｍＲＮＡの暗号に対応したアミノアシルｔＲＮＡをリボソームのＡ部位（アミノアシルｔＲＮＡ結合部位）へ結合させる因子と、ぺプチジル結合形成直後Ａ部位に存在するペプチジルｔＲＮＡをＰ部位へトランスロケーションさせる因子である。翻訳開始因子および翻訳伸長因子の調節は、最終産物であるタンパク質の発現量を調節するという意味で重要な過程であり、ｍＲＮＡ翻訳開始および翻訳伸長の制御の変化はヒトを含む哺乳類の病気を発症させる原因の一つと考えられているので（非特許文献１６から２０）、翻訳過程の制御を理解する事は、様々な病気の原因解明だけなく、新しい予防薬、診断薬、治療薬の開発に応用できると考えられる。 The regulation of protein expression is closely related to cell growth and cell cycle progression. Therefore, it is expected that abnormalities in the translation process are related not only to the etiology / pathology of human diseases but also to the action of therapeutic agents. Translation of mRNA consists of stages such as initiation, elongation and termination, and many eukaryotic translation initiation factors (eIFs) act specifically at the initiation stage. . The translation initiation factor is also referred to as a polypeptide chain initiation factor, and is a protein group (eIF1, eIF2, eIF3, eIF4, eIF5, etc.) in which a liposome binds amino acids according to the base sequence of mRNA, that is, a translation process of protein synthesis. Say. The final purpose of initiating eIFs and translation is to form the 80S ribosome and place the mRNA start codon, Met-tRNA, on the ribosome P site (peptidyl tRNA binding site). The translation elongation factor is also referred to as a polypeptide chain elongation factor, and is a protein group (eEF1, eEF2, eEF3, eEF4, eEF5, etc.) necessary for the translation process of polypeptide chain elongation that biosynthesizes a protein from mRNA. Translation elongation factors are broadly divided into two types, functionally binding aminoacyl-tRNAs corresponding to the mRNA code to the A-site (aminoacyl-tRNA binding site) of the ribosome and present at the A-site immediately after peptidyl bond formation. It is a factor that translocates peptidyl tRNA to P site. Regulation of translation initiation factor and translation elongation factor is an important process in the sense that it regulates the expression level of the final protein, and changes in mRNA translation initiation and translation elongation control cause diseases in mammals including humans Understanding the control of the translation process is not only to elucidate the causes of various diseases, but also to develop new preventives, diagnostics, and therapeutics. It can be applied.
　ミノサイクリンやシロスタゾールの詳細な作用メカニズムを解明する事が、様々な疾患に対する新規な予防薬、診断薬、および治療薬等の開発に資することになる。本発明の課題は、様々な疾患、特に神経疾患に有効な医薬製剤を提供することにある。 Elucidation of the detailed mechanism of action of minocycline and cilostazol will contribute to the development of new preventives, diagnostics, and therapeutics for various diseases. The subject of this invention is providing the pharmaceutical formulation effective in various diseases, especially a neurological disease.
　本発明者らは、上記課題を解決すべく鋭意検討を行う中で、ミノサイクリンまたはシロスタゾールが、ＰＣ１２細胞におけるＮＧＦ誘発神経突起伸長作用を有意に増強させる作用を有する事（神経突起伸長増強作用という）を発見した。他の第二世代抗生物質であるテトラサイクリンには、その効果が認められなかった。また、シロスタゾールの主作用は、フォスフォジエステラーゼ（ＰＤＥ）３阻害作用であるが、他のＰＤＥ３阻害薬シロスタミドはＮＧＦ誘発神経突起伸長効果を示さなかったことから、ＮＧＦ誘発神経突起伸長増強作用には、シロスタゾールのＰＤＥ３阻害作用は関係していない可能性があった。本発明者らは、ミノサイクリンまたはシロスタゾールのＮＧＦ誘発神経突起伸長増強作用の作用メカニズムの解明を進め、当該作用にそれぞれ翻訳開始因子（ｅＩＦ４Ａ１など）または翻訳伸長因子（ｅＥＦ１Ａ１など）の増加が関与していることを初めて見出し、本発明を完成させた。 The present inventors have conducted intensive studies to solve the above problems, and that minocycline or cilostazol has an action of significantly enhancing the NGF-induced neurite outgrowth action in PC12 cells (referred to as neurite outgrowth enhancement action). I found Tecycline, another second-generation antibiotic, has not been effective. The main effect of cilostazol is phosphodiesterase (PDE) 3 inhibitory action, but the other PDE3 inhibitor cilostamide did not show NGF-induced neurite outgrowth effect. May not be related to the PDE3 inhibitory action of cilostazol. The present inventors proceeded to elucidate the mechanism of action of minocycline or cilostazol on enhancing the NGF-induced neurite outgrowth, and the increase of translation initiation factor (eIF4A1 etc.) or translation elongation factor (eEF1A1 etc.) was involved in the action, respectively. It was found for the first time that the present invention was completed.
　すなわち、本発明は、翻訳開始因子もしくは翻訳伸長因子を増加させる作用を有する化合物、またはその薬学的に許容し得る塩を含有する、医薬製剤に関する。
　また、本発明は、翻訳開始因子または翻訳伸長因子の異常が関わっているような様々な疾患の治療薬に関し、さらに、神経疾患に有効な医薬製剤に関する。 That is, the present invention relates to a pharmaceutical preparation containing a compound having an action of increasing a translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof.
The present invention also relates to therapeutic agents for various diseases in which abnormalities of translation initiation factors or translation elongation factors are involved, and further to pharmaceutical formulations effective for neurological diseases.
　本発明は、１）ミノサイクリンおよび／またはシロスタゾールを含有する神経突起伸長増強剤に関する。
　また、２）翻訳開始因子および／または翻訳伸長因子を増加させることを特徴とする神経突起伸長増強剤に関する。
　また、３）翻訳開始因子であるｅＩＦ４Ａアイソフォームおよび／または翻訳伸長因子であるｅＥＦ１Ａアイソフォームを増加させることを特徴とする神経突起伸長増強剤に関する。
　さらに、４）翻訳開始因子であるｅＩＦ４Ａおよび／または翻訳伸長因子であるｅＥＦ１Ａを増加させることを特徴とする神経突起伸長増強剤に関する。
　並びに、５）上述の１）から４）のいずれかに記載の神経突起伸長増強剤の精神疾患治療への使用に関する。
　また、本発明は、６）ミノサイクリンおよび／またはシロスタゾールを含有する、翻訳開始因子増加剤および／または翻訳伸長因子増加剤に関する。
　さらに、７）ミノサイクリンを含有する翻訳開始因子ｅＩＦ４Ａ１の増加剤に関する。
　さらに、８）シロスタゾールを含有する翻訳伸長因子ｅＥＦ１Ａ１の増加剤に関する。
　また、９）ミノサイクリンおよび／またはシロスタゾールを含有する、翻訳開始因子および／または翻訳伸長因子の異常に起因する疾患の予防剤および／または治療剤に関する。
　また、１０）ミノサイクリンおよび／またはシロスタゾールを含有する、翻訳開始因子であるｅＩＦ４Ａアイソフォームおよび／または翻訳伸長因子であるｅＥＦ１Ａアイソフォームの減少に由来する精神疾患、神経疾患、脳血管疾患、循環器疾患、呼吸器疾患、内分泌疾患、代謝疾患、血液疾患、免疫疾患、感染症、および悪性腫瘍からなる群から選択される１種または２種以上の症状を呈する疾患の予防剤および／または治療剤に関する。
　さらに、１１）ミノサイクリンおよび／またはシロスタゾールを含有する、翻訳開始因子であるｅＩＦ４Ａ１および／または翻訳伸長因子であるｅＥＦ１Ａ１の減少に由来する精神疾患、神経疾患、脳血管疾患、循環器疾患、呼吸器疾患、内分泌疾患、代謝疾患、血液疾患、免疫疾患、感染症、および悪性腫瘍からなる群から選択される１種または２種以上の症状を呈する疾患の予防剤および／または治療剤に関する。 The present invention relates to 1) a neurite outgrowth enhancer containing minocycline and / or cilostazol.
The present invention also relates to a neurite elongation enhancer characterized by increasing 2) a translation initiation factor and / or a translation elongation factor.
The present invention also relates to a neurite elongation enhancer characterized by increasing eIF4A isoform that is a translation initiation factor and / or eEF1A isoform that is a translation elongation factor.
And 4) a neurite elongation enhancer characterized by increasing eIF4A as a translation initiation factor and / or eEF1A as a translation elongation factor.
And 5) the use of the neurite outgrowth enhancer according to any one of 1) to 4) for treating psychiatric disorders.
The present invention also relates to 6) a translation initiation factor increasing agent and / or a translation elongation factor increasing agent comprising minocycline and / or cilostazol.
Further, it relates to an agent for increasing 7) translation initiation factor eIF4A1 containing minocycline.
Furthermore, it relates to an agent for increasing translation elongation factor eEF1A1 containing 8) cilostazol.
The present invention also relates to 9) a preventive and / or therapeutic agent for a disease caused by an abnormality in a translation initiation factor and / or translation elongation factor, containing minocycline and / or cilostazol.
10) Psychiatric diseases, neurological diseases, cerebrovascular diseases, cardiovascular diseases derived from a decrease in eIF4A isoform which is a translation initiation factor and / or eEF1A isoform which is a translation elongation factor, containing minocycline and / or cilostazol. A preventive and / or therapeutic agent for a disease exhibiting one or more symptoms selected from the group consisting of respiratory diseases, endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors .
11) Psychiatric diseases, neurological diseases, cerebrovascular diseases, cardiovascular diseases, respiratory diseases derived from a decrease in eIF4A1 which is a translation initiation factor and / or eEF1A1 which is a translation elongation factor, containing minocycline and / or cilostazol And an agent for preventing and / or treating a disease exhibiting one or more symptoms selected from the group consisting of endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors.
　ミノサイクリンもしくはシロスタゾール、またはその薬学的に許容し得る塩は、それぞれ翻訳開始因子（ｅＩＦ４Ａ１）または翻訳伸長因子（ｅＥＦ１Ａ１）を有意に増加させたことから、翻訳開始因子または翻訳伸長因子を増加させる化合物は、様々な疾患、翻訳開始因子または翻訳伸長因子の異常が関係するような疾患に対して効果を奏し、その予防および／または治療に有効である。
　例えば、翻訳開始因子または翻訳伸長因子の異常が指摘されている様々な疾患やタンパクの異常形成が関連する疾患、具体的には、精神疾患、神経疾患、脳血管疾患、循環器疾患、呼吸器疾患、内分泌疾患、代謝疾患、血液疾患、免疫疾患、感染症、および悪性腫瘍などの様々な疾患の予防および／または治療に有効である。 Since minocycline or cilostazol, or a pharmaceutically acceptable salt thereof, significantly increased translation initiation factor (eIF4A1) or translation elongation factor (eEF1A1), respectively, compounds that increase translation initiation factor or translation elongation factor are It is effective for various diseases, diseases associated with abnormalities of translation initiation factors or translation elongation factors, and effective for the prevention and / or treatment thereof.
For example, various diseases in which abnormalities of translation initiation factors or translation elongation factors have been pointed out and diseases associated with abnormal protein formation, such as mental diseases, neurological diseases, cerebrovascular diseases, cardiovascular diseases, respiratory organs It is effective for the prevention and / or treatment of various diseases such as diseases, endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors.
図１は、神経成長因子誘発神経突起作用に対するミノサイクリンの効果を示す図である。ミノサイクリンは、濃度依存的にＰＣ１２細胞におけるＮＧＦ誘発神経突起伸長作用を有意に増強させたが、他の抗生物質テトラサイクリンは該作用に影響を与えなかった。FIG. 1 shows the effect of minocycline on nerve growth factor-induced neurite action. Minocycline significantly enhanced NGF-induced neurite outgrowth in PC12 cells in a concentration-dependent manner, whereas other antibiotic tetracyclines did not affect the effect. 図２は、ミノサイクリンおよびテトラサイクリン処置による細胞内のｅＩＦ４Ａ１タンパク発現量を示す図である。ミノサイクリンは、ｅＩＦ４Ａ１タンパク発現量を有意に増加させた。FIG. 2 is a diagram showing intracellular expression levels of eIF4A1 protein treated with minocycline and tetracycline. Minocycline significantly increased eIF4A1 protein expression. 図３は、ミノサイクリンの細胞内の翻訳開始因子ｅＩＦ４Ａ１タンパク発現量増加に対するｓｉＲＮＡ処置等の影響を示す図である。ｅＩＦ４Ａ１のｓｉＲＮＡ処置によりｅＩＦ４Ａ１のタンパク発現量は有意に減少した。ネガティブＲＮＡでは、影響はなかった。FIG. 3 is a diagram showing the effect of siRNA treatment and the like on the increase in the expression level of intracellular translation initiation factor eIF4A1 protein of minocycline. The eIF4A1 siRNA treatment significantly decreased the protein expression level of eIF4A1. Negative RNA had no effect. 図４は、ミノサイクリンのＮＧＦ誘発神経突起伸長作用におけるｅＩＦ４Ａ１の効果を示す図である。図２に示した条件下で、ｓｉＲＮＡ処置により翻訳開始因子ｅＩＦ４Ａ１のタンパク発現量の低下させると、ミノサイクリンのＮＧＦ誘発神経突起伸長作用を抑制した。FIG. 4 shows the effect of eIF4A1 on the NGF-induced neurite outgrowth action of minocycline. Under the conditions shown in FIG. 2, when the protein expression level of the translation initiation factor eIF4A1 was decreased by siRNA treatment, the NGF-induced neurite outgrowth action of minocycline was suppressed. 図５は、ＮＧＦ誘発神経突起伸長作用に対するシロスタゾールの効果を示す図である。シロスタゾールは濃度依存的にＰＣ１２細胞におけるＮＧＦ誘発神経突起伸長作用を有意に増強させたが、他のＰＤＥ３阻害薬シロスタミドは該作用に影響を与えなかった。FIG. 5 shows the effect of cilostazol on the NGF-induced neurite outgrowth action. Cilostazol significantly enhanced NGF-induced neurite outgrowth in PC12 cells in a concentration-dependent manner, whereas the other PDE3 inhibitor cilostamide did not affect the effect. 図６は、シロスタゾールおよびシロスタミド処置による細胞内のｅＥＦ１Ａ１タンパク発現量を示す図である。シロスタゾールは、ｅＥＦ１Ａ１タンパク発現量を有意に増加させた。FIG. 6 is a graph showing the intracellular expression level of eEF1A1 protein treated with cilostazol and cilostamide. Cilostazol significantly increased the expression level of eEF1A1 protein. 図７は、シロスタゾールの細胞内の翻訳開始因子ｅＥＦ１Ａ１タンパク発現量増加に対するｓｉＲＮＡ処置等の影響を示す図である。ｅＥＦ１Ａ１のｓｉＲＮＡ処置によりｅＥＦ１Ａ１のタンパク発現量は有意に減少した。ネガティブＲＮＡでは、影響はなかった。FIG. 7 is a diagram showing the influence of siRNA treatment or the like on the increase in the expression level of cilostazol translation initiation factor eEF1A1 protein in cells. The eEF1A1 siRNA treatment significantly decreased the protein expression level of eEF1A1. Negative RNA had no effect. 図８は、シロスタゾールのＮＧＦ誘発神経突起伸長作用におけるｅＩＦ１Ａ１の効果を示す図である。図７に示した条件下で、ｓｉＲＮＡ処置により翻訳開始因子ｅＥＦ１Ａ１のタンパク発現量の低下させると、シロスタゾールのＮＧＦ誘発神経突起伸長作用を抑制した。FIG. 8 is a graph showing the effect of eIF1A1 on the NGF-induced neurite outgrowth action of cilostazol. Under the conditions shown in FIG. 7, when the protein expression level of the translation initiation factor eEF1A1 was decreased by siRNA treatment, the NGF-induced neurite outgrowth action of cilostazol was suppressed. 図９は、ｅＥＦ１Ａ１タンパク発現に及ぼすテトラサイクリンおよびミノサイクリンの効果、並びにｅＩＦ４Ａ１タンパク発現に及ぼすシロスタミドおよびシロスタゾールの効果を示した図である。テトラサイクリンおよびミノサイクリンは、ｅＥＦ１Ａ１タンパク発現には影響を与えず、並びにシロスタミドおよびシロスタゾールは、ｅＩＦ４Ａ１タンパク発現には影響を与えなかった。FIG. 9 shows the effects of tetracycline and minocycline on eEF1A1 protein expression, and the effects of cilostamide and cilostazol on eIF4A1 protein expression. Tetracycline and minocycline did not affect eEF1A1 protein expression, and cilostamide and cilostazol did not affect eIF4A1 protein expression. 図１０は、ｅＩＦ４Ａ１のｓｉＲＮＡの塩基配列を、アンチセンスとして示した図である。FIG. 10 is a diagram showing the nucleotide sequence of siRNA of eIF4A1 as an antisense. 図１１は、ｅＥＦ１Ａ１のｓｉＲＮＡの塩基配列を、アンチセンスとして示した図である。FIG. 11 is a diagram showing the nucleotide sequence of eEF1A1 siRNA as antisense.
　まず、本明細書における用語の意味あるいは定義について述べる。翻訳開始因子とは、リボソームがｍＲＮＡの塩基配列に従ってアミノ酸を結合させる過程、すなわちタンパク質合成の翻訳過程を開始させるタンパク質群である。翻訳伸長因子とは、ｍＲＮＡからタンパク質合成の翻訳過程を進行させる、ペプチド鎖伸長に関わる過程を制御するタンパク質群である。
　翻訳開始因子のうちｅＩＦ４Ａというタンパク質は、ｅＩＦ４Ａ１、ｅＩＦ４Ａ２、ｅＩＦ４Ａ３など幾つかのアイソフォーム（またはサブクラス）が知られており、それらの機能は同じものと考えられ、本発明の効果を奏するものであれば、１種または２種以上のアイソフォームを対象として発明を実施することができる。これらのタンパク質を示す場合、ｅＩＦ４Ａアイソフォームという。本発明において好ましいｅＩＦ４ＡアイソフォームとしてｅＩＦ４Ａ１を挙げることができる。
　また、同様に、翻訳伸長因子についても、翻訳伸長因子のうちｅＥＦ１Ａというタンパク質は、ｅＥＦ１Ａ１、ｅＥＦ１Ａ２、ｅＥＦ１Ａ３など幾つかのアイソフォーム（またはサブクラス）が知られており、それらの機能は同じものと考えられ、本発明の効果を奏するものであれば、１種または２種以上のアイソフォームを対象として発明を実施することができる。これらのタンパク質を示す場合、ｅＥＦ１Ａアイソフォームという。本発明において好ましいｅＥＦ１ＡアイソフォームとしてｅＥＦ１Ａ１を挙げることができる。
　神経突起伸長作用とは、神経細胞の神経突起形成を起こさせる作用をいい、その増強作用（神経突起伸長増強作用）は、例えば、実施例中で述べたような定量化試験を行えばその効果が判定できる。さらに、神経突起伸長増強作用を有し、医薬として充分な該効果を有する化合物を神経突起伸長増強剤という。 First, the meaning or definition of terms in this specification will be described. The translation initiation factor is a protein group that initiates the process of ribosome binding amino acids according to the base sequence of mRNA, that is, the translation process of protein synthesis. Translation elongation factors are a group of proteins that control the process involved in peptide chain elongation, which advances the translation process of protein synthesis from mRNA.
Among the translation initiation factors, a protein called eIF4A is known to have several isoforms (or subclasses) such as eIF4A1, eIF4A2, and eIF4A3, their functions are considered to be the same, and the effects of the present invention can be achieved. For example, the invention can be implemented for one or more isoforms. When referring to these proteins, they are referred to as eIF4A isoforms. As a preferred eIF4A isoform in the present invention, eIF4A1 can be mentioned.
Similarly, as for the translation elongation factor, the protein called eEF1A among the translation elongation factors is known to have several isoforms (or subclasses) such as eEF1A1, eEF1A2, and eEF1A3, and their functions are considered to be the same. As long as the effects of the present invention are exhibited, the invention can be implemented for one or more isoforms. When these proteins are referred to, they are referred to as eEF1A isoforms. In the present invention, eEF1A1 can be mentioned as a preferred eEF1A isoform.
The neurite outgrowth action refers to the action of causing neurite formation in nerve cells, and the enhancement action (neurite outgrowth enhancement action) is, for example, the effect of performing a quantification test as described in the Examples. Can be determined. Furthermore, a compound having a neurite outgrowth enhancing action and having the effect sufficient as a medicine is called a neurite outgrowth enhancer.
　本発明は、翻訳開始因子および／または翻訳伸長因子を増加させることを特徴とする神経突起伸長増強剤に関する。より好ましくは本発明は、翻訳開始因子であるｅＩＦ４Ａアイソフォームおよび／または翻訳伸長因子であるｅＥＦ１Ａアイソフォームを増加させることを特徴とする神経突起伸長増強剤に関する。さらに好ましくは本発明は、翻訳開始因子であるｅＩＦ４Ａおよび／または翻訳伸長因子であるｅＥＦ１Ａを増加させることを特徴とする神経突起伸長増強剤に関する。 The present invention relates to a neurite elongation enhancer characterized by increasing a translation initiation factor and / or a translation elongation factor. More preferably, the present invention relates to a neurite extension enhancer characterized by increasing eIF4A isoform which is a translation initiation factor and / or eEF1A isoform which is a translation elongation factor. More preferably, the present invention relates to a neurite outgrowth enhancer characterized by increasing eIF4A which is a translation initiation factor and / or eEF1A which is a translation elongation factor.
　翻訳開始因子もしくは翻訳伸長因子を増加させる化合物として、ミノサイクリンおよびシロスタゾールを好ましく例示できる。ミノサイクリンは、翻訳開始因子、好ましくはｅＩＦ４Ａアイソフォーム、より好ましくはｅＩＦ４Ａ１の増加剤として使用できる。シロスタゾール、翻訳伸長因子、好ましくはｅＥＦ１Ａアイソフォーム、より好ましくはｅＥＦ１Ａ１の増加剤として使用できる。 Preferred examples of compounds that increase the translation initiation factor or translation elongation factor include minocycline and cilostazol. Minocycline can be used as an enhancer of a translation initiation factor, preferably an eIF4A isoform, more preferably eIF4A1. It can be used as an increaser for cilostazol, a translation elongation factor, preferably an eEF1A isoform, more preferably eEF1A1.
　翻訳開始因子もしくは翻訳伸長因子を増加させる化合物、またはその薬学的に許容しうる塩を含有する本発明の医薬製剤、例えば予防剤および／または治療剤は、経口的または非経口的に投与することができる。経口投与には、錠剤、カプセル、コーティング錠、トローチ、溶液または懸濁液などの液剤といった既知の投与用剤形を用いることができる。また、非経口投与は、坐剤などを用いた直腸投与、パッチやリニメント、ゲルなどを用いた経皮投与、スプレーやエアロゾルなどを用いた経粘膜投与などによって行われてもよく、注射により静脈内、筋肉内、皮下または脳室内等に投与されてもよい。 The pharmaceutical preparation of the present invention containing a compound that increases a translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof, such as a prophylactic agent and / or therapeutic agent, is administered orally or parenterally. Can do. For oral administration, known dosage forms for administration such as tablets, capsules, coated tablets, troches, solutions such as solutions or suspensions can be used. Parenteral administration may be performed by rectal administration using suppositories, transdermal administration using patches, liniments, gels, etc., transmucosal administration using sprays, aerosols, etc. It may be administered internally, intramuscularly, subcutaneously or intraventricularly.
　本発明の予防剤および／または治療剤は、翻訳開始因子もしくは翻訳伸長因子を増加させる化合物は遊離塩基、またはその薬学的に許容し得る塩の両方の形態で用いることができる。薬学的に許容し得る塩としては、薬学的に許容し得る酸の付加塩が好ましく、より好ましくは塩酸塩である。
　本発明の予防剤および／または治療剤は、翻訳開始因子もしくは翻訳伸長因子を増加させる化合物、またはその薬学的に許容しうる塩の他に、他の薬効成分を含有していてもよい。また、これら薬効成分の他に、適宜、投与形態などに応じて、抗酸化剤、安定剤、防腐剤、矯味剤、着色料、溶解剤、可溶化剤、界面活性剤、乳化剤、消泡剤、粘度調整剤、ゲル化剤、吸収促進剤、分散剤、賦形剤、およびｐＨ調整剤等の当業者によく知られた適切な生理学的に許容される物質を含有していてもよい。 In the prophylactic and / or therapeutic agent of the present invention, the compound that increases the translation initiation factor or translation elongation factor can be used in the form of both a free base or a pharmaceutically acceptable salt thereof. The pharmaceutically acceptable salt is preferably a pharmaceutically acceptable acid addition salt, more preferably a hydrochloride.
The preventive agent and / or therapeutic agent of the present invention may contain other medicinal ingredients in addition to a compound that increases a translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof. In addition to these medicinal ingredients, an antioxidant, a stabilizer, an antiseptic, a corrigent, a coloring agent, a solubilizer, a solubilizer, a surfactant, an emulsifier, and an antifoaming agent depending on the dosage form as appropriate. And suitable physiologically acceptable substances well known to those skilled in the art, such as viscosity modifiers, gelling agents, absorption enhancers, dispersants, excipients, and pH adjusters.
　したがって、本発明の予防剤および／または治療剤は、翻訳開始因子もしくは翻訳伸長因子を増加させる化合物、またはその薬学的に許容しうる塩と、これらの物質とを適宜配合することにより製造することができる。本発明の予防剤および／または治療剤を注射（皮下注射、筋肉注射、または静脈注射）用製剤として調製する場合、溶液剤または懸濁剤の製剤の形態が好ましく、膣または直腸投与用の場合、クリ－ムまたは坐薬のような半固形剤の製剤の形態が好ましく、経鼻腔投与用の場合、粉末、鼻用滴剤、またはエアロゾル剤の製剤の形態が好ましい。これらの製剤はいずれも、例えばレミントンの製薬科学（マック・パブリッシング・カンパニー、イーストン、ＰＡ、１９７０年）に記載されているような製薬技術上当業者に知られているいずれかの方法によって調製することができる。注射用製剤は担体として、例えば、アルブミン等の血漿由来タンパク、グリシン等のアミノ酸、およびマンニトール等の糖を加えることができ、さらに緩衝剤、溶解補助剤、および等張剤等を添加することもできる。また、水溶製剤または凍結乾燥製剤として使用する場合、凝集を防ぐためにＴｗｅｅｎ（登録商標）８０、Ｔｗｅｅｎ（登録商標）２０などの界面活性剤を添加するのが好ましい。さらに、注射用製剤以外の非経口投与剤形は、蒸留水または生理食塩液、ポリエチレングリコ－ルのようなポリアルキレングリコ－ル、植物起源の油、および水素化したナフタレン等を含有してもよい。例えば、坐薬のような膣または直腸投与用の製剤は、一般的な賦形剤として、例えば、ポリアキレングリコ－ル、ワセリン、およびカカオ油脂等を含有する。膣用製剤では、胆汁塩、エチレンジアミン塩、およびクエン酸塩等の吸収促進剤を含有してもよい。吸入用製剤は固体でもよく、賦形剤として、例えば、ラクト－スを含有してもよく、さらに、経鼻腔滴剤は水または油溶液であってもよい。 Therefore, the preventive agent and / or therapeutic agent of the present invention is produced by appropriately combining a compound that increases a translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof, and these substances. Can do. When preparing the prophylactic and / or therapeutic agent of the present invention as a preparation for injection (subcutaneous injection, intramuscular injection, or intravenous injection), the form of a solution or suspension is preferred, and for vaginal or rectal administration Semi-solid dosage forms such as creams or suppositories are preferred, and for nasal administration, powder, nasal drops, or aerosol dosage forms are preferred. All of these preparations should be prepared by any method known to those skilled in the pharmaceutical arts, as described, for example, in Remington's Pharmaceutical Sciences (Mac Publishing Company, Easton, PA, 1970). Can do. Injectable preparations can contain, for example, plasma-derived proteins such as albumin, amino acids such as glycine, and sugars such as mannitol as carriers, and buffers, solubilizers, isotonic agents, and the like can also be added. it can. Moreover, when using as a water-soluble formulation or a freeze-dried formulation, it is preferable to add surfactants, such as Tween (trademark) 80 and Tween (trademark) 20, in order to prevent aggregation. Further, parenteral dosage forms other than injectable preparations may contain distilled water or physiological saline, polyalkylene glycols such as polyethylene glycol, oils of plant origin, hydrogenated naphthalene, and the like. Good. For example, preparations for vaginal or rectal administration such as suppositories contain, for example, polyalkylene glycol, petrolatum, cacao oil and the like as common excipients. Vaginal preparations may contain absorption enhancers such as bile salts, ethylenediamine salts, and citrate salts. Inhalation formulations may be solid, may contain, for example, lactose as an excipient, and nasal drops may be water or oil solutions.
　本発明の予防剤および／または治療剤の正確な投与量および投与計画は、個々の治療対象毎の所要量、治療方法、疾病または必要性の程度などに依存して調整できる。投与量は、具体的には年齢、体重、一般的健康状態、性別、食事、投与時間、投与方法、***速度、薬物の組合せ、および患者の病状などに応じて決めることができ、さらに、その他の要因を考慮して決定してもよい。翻訳開始因子あるいは翻訳伸長因子を増加させる化合物、またはその薬学的に許容しうる塩は、ヒトにおける薬物動態に問題がなく、安全に使用することができ、その１日の投与量は、患者の状態や体重、化合物の種類、投与経路などによって異なるが、例えば、非経口的には皮下、静脈内、筋肉内または直腸内に、翻訳開始因子または翻訳伸長因子を増加させる化合物（遊離塩基）として約０．０１～１０００ｍｇ／人／日、好ましくは０．１～５００ｍｇ／人／日で投与され、また、経口的には約０．０１～５００ｍｇ／人／日、好ましくは０．１～１００ｍｇ／人／日で投与されることが望ましい。 The exact dose and administration schedule of the preventive agent and / or therapeutic agent of the present invention can be adjusted depending on the required amount, treatment method, disease or degree of necessity for each individual treatment target. Specifically, the dosage can be determined according to age, weight, general health condition, sex, diet, administration time, administration method, excretion rate, combination of drugs, patient condition, etc. It may be determined in consideration of these factors. A compound that increases translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof, has no problem in pharmacokinetics in humans, and can be used safely. As a compound (free base) that increases translation initiation factor or translation elongation factor parenterally, subcutaneously, intravenously, intramuscularly or rectally, depending on the condition, body weight, type of compound, administration route, etc. It is administered at about 0.01 to 1000 mg / person / day, preferably 0.1 to 500 mg / person / day, and orally about 0.01 to 500 mg / person / day, preferably 0.1 to 100 mg. It is desirable to be administered per person / day.
　さらに本発明は、翻訳開始因子の増加剤および／または翻訳伸長因子の増加剤を対象に投与することを特徴とする神経突起伸長増強方法に関する。また本発明は、神経突起伸長増強方法や神経突起伸長増強剤の製造における、翻訳開始因子の増加剤および／または翻訳伸長因子の増加剤の使用に関する。さらにまた本発明は、翻訳開始因子の増加剤および／または翻訳伸長因子の増加剤を対象に投与して神経突起伸長を増強することを特徴とする精神疾患の予防および／または治療方法に関する。また本発明は、精神疾患の予防および／または治療方法や精神疾患の予防および／または治療剤の製造における、翻訳開始因子の増加剤および／または翻訳伸長因子の増加剤の使用に関する。 Furthermore, the present invention relates to a method for enhancing neurite outgrowth, which comprises administering an agent for increasing a translation initiation factor and / or an agent for increasing a translation elongation factor to a subject. The present invention also relates to the use of an agent for increasing translation initiation factor and / or an agent for increasing translation elongation factor in the method for enhancing neurite elongation and the production of neurite elongation enhancer. Furthermore, the present invention relates to a method for preventing and / or treating a mental illness characterized by enhancing neurite outgrowth by administering to a subject an agent for increasing a translation initiation factor and / or an agent for increasing a translation elongation factor. The present invention also relates to the use of an agent for increasing a translation initiation factor and / or an agent for increasing a translation elongation factor in the production of a method for preventing and / or treating mental illness and a prophylactic and / or therapeutic agent for mental illness.
　以下、実施例にて、本発明をさらに具体的に説明するが、本発明はこの実施例に限定されない。また、本発明の技術的思想を逸脱しない範囲で種々の変更が可能である。 Hereinafter, the present invention will be described more specifically with reference to examples, but the present invention is not limited to these examples. Various modifications can be made without departing from the technical idea of the present invention.
（１）細胞培養
　ダルベッコ変法イーグル培地（ＤＭＥＭ）に５％熱不活化ウシ胎児血清（ＦＢＳ）、１０％熱不活化ウマ血清、１％ペニシリン／ストレプトマイシンを添加し、ＰＣ１２細胞（理化学研究所バイオリソースセンター細胞材料開発室、日本、つくば）をポリ－Ｄ－リジン／ラミニンコーティングした１００ｍｍ細胞培養ディッシュ（ベクトン・ディッキンソン、Ｆｒａｎｋｌｉｎ　Ｌａｋｅｓ，ＮＪ，ＵＳＡ）を用い、３７℃、５％ＣＯ２で培養した。週に２～３回培地を交換した。以下に述べる神経突起伸長増強効果を検討する試験、ウェスタンブロット法に供する際は、血清は様々な成長因子を含有することが知られているため、最低限の濃度の血清（０．５％ＦＢＳ）を含む培地を用いた。ＰＣ１２細胞分化誘導の為、ＮＧＦ（２．５ｎｇ／ｍＬ）の条件で、播種から２４時間後、０．５％ＦＢＳ、１％ペニシリン／ストレプトマイシンを含むＤＭＥＭ培地にさらに各種薬剤を添加したもの、しないものと培地を交換した。
（２）神経突起形成の定量化
　神経突起伸長増強の効果を検討する試験においては、ポリ－Ｄ－リジン／ラミニンコーティングした２４ウェル組織培養プレートに、比較的低密度で（０．２５×１０４個／ｃｍ２）ＰＣ１２細胞を播種した。ＮＧＦ（２．５ｎｇ／ｍｌ）を加え、さらに、各種薬剤の添加または不添加で４日間培養した後、カメラ付き位相差倒立顕微鏡で撮影した生細胞のデジタル画像を用いて形態計測分析を実施した。１ウェルにつき３視野の画像を撮影した。１視野当たりの細胞数は平均１００個であった。視認により各視野の分化細胞数を計測した。細胞体直径と等しい長さの神経突起が１本以上ある細胞のみを数え、当該視野内の総細胞数に対する割合として示した。計測は盲検下にて行われた。
（３）タンパク質発現定量化
　ウェスタンブロット分析では、ＮＧＦ（２．５ｎｇ／ｍｌ）を加え、さらに、各種薬剤の添加または不添加で４日間培養した後、細胞溶解物からのタンパク質（総３０μｇ）を７．５％ポリアクリルアミドゲル電気泳動（Ｂｉｏ－Ｒａｄ，Ｈｅｒｃｕｌｅｓ，ＣＡ，ＵＳＡ）にかけて分離した。Ｔａｎｋ－ｔｒａｎｓｆｅｒ装置（Ｂｉｏ－Ｒａｄ）を用いてタンパク質をポリビニリデンフルオライド膜（ＧＥヘルスケア）に移し、この膜を５％スキムミルク、２％ＢＳＡ、５％ＦＢＳを含むＴＢＳＴ（５０ｍＭ Ｔｒｉｓ／ＨＣｌ、ｐＨ７．８、０．１３Ｍ ＮａＣｌ、０．１％ Ｔｗｅｅｎ（登録商標）２０）でブロックした。ウサギ抗ｅＩＦ４Ａ抗体（Ａｂｃａｍ）およびマウス抗β‐アクチン抗体（シグマアルドリッチ）を上記ブロッキング緩衝液中でそれぞれ１：２５０、１：１００００に希釈したものを一次抗体として使用した。さらに、５％スキムミルク含有ＴＢＳＴ中で２５０００倍希釈の抗ウサギＩｇＧ西洋ワサビペルオキシダーゼ標識抗体（ＧＥヘルスケア）、および５％スキムミルク含有ＴＢＳＴ中で２５０００倍希釈の抗マウスＩｇＧ西洋ワサビペルオキシダーゼ標識抗体（ＧＥヘルスケア）を二次抗体として使用した。膜上の抗原タンパク質はＥＣＬＴＭ Ｐｌｕｓ Ｗｅｓｔｅｒｎ Ｂｌｏｔｔｉｎｇ Ｄｅｔｅｃｔｉｏｎ Ｓｙｓｔｅｍ（ＧＥヘルスケア）で検出した。冷却ＣＣＤカメラによるバンドの可視化をＬＡＳ３０００ｍｉｎｉ（富士写真フイルム社）により行い、解析ソフトＭｕｌｔｉ Ｇａｕｇｅ（富士写真フイルム社）によりタンパク質発現の定量化をおこなった。
（４）ｓｉＲＮＡによる遺伝子ノックダウン
　ｅＩＦ４Ａ１、あるいはｅＥＦ１Ａ１を標的とするｓｉＲＮＡを用いてｅＩＦ４Ａ１、あるいはｅＥＦ１Ａ１の発現を抑制した。化学合成されたｓｉＲＮＡは、試薬会社（Ｉｎｔｅｇｒａｔｅｄ　ＤＮＡ　Ｔｅｃｈｎｏｌｏｇｉｅｓ，Ｃｏｒａｌｖｉｌｌｅ，ＣＡ，ＵＳＡ）より購入した（図１０、図１１）。実験に供したｓｉＲＮＡのアンチセンス配列およびセンス配列はそれぞれ配列表の配列番号１から６および配列番号７から１２に示した。ＰＣ１２細胞を培養し、リポフェクタミン２０００（Ｉｎｖｉｔｒｏｇｅｎ，Ｃａｒｌｓｂａｄ，ＣＡ，ＵＳＡ）およびＯＰＴＩ－ＭＥＭ（Ｉｎｖｉｔｒｏｇｅｎ，Ｃａｒｌｓｂａｄ，ＣＡ，ＵＳＡ）を用いて上記のｓｉＲＮＡをトランスフェクションした。６時間後にＮＧＦ（２．５ｎｇ／ｍｌ）、さらに、各種薬剤の添加または不添加の０．５％ＦＢＡ、１％ペニシリン／ストレプトマイシンを含むＤＭＥＭ培地と交換し、神経突起伸長増強効果を検討する試験およびウェスタンブロット分析に供した。
（５）統計分析
　データは平均値±標準偏差で示した。統計分析は一元配置分散分析を用いて行ない、ｐｏｓｔ－ｈｏｃ　ボンフェロニ検定で行なった。０．０５以下のｐ値は統計的に有意とした。 (1) Cell culture 5% heat-inactivated fetal bovine serum (FBS), 10% heat-inactivated horse serum, 1% penicillin / streptomycin was added to Dulbecco's modified Eagle's medium (DMEM), and PC12 cells (RIKEN BioResources) The cells were cultured at 37 ° C. and 5% CO 2 using a poly-D-lysine / laminin coated 100 mm cell culture dish (Becton Dickinson, Franklin Lakes, NJ, USA). The medium was changed 2-3 times a week. When subjected to Western blotting, a test for examining the effect of enhancing neurite outgrowth described below, since serum is known to contain various growth factors, a minimum concentration of serum (0.5% FBS) ) Was used. For induction of PC12 cell differentiation, 24 hours after seeding under the condition of NGF (2.5 ng / mL), no addition of various drugs to DMEM medium containing 0.5% FBS, 1% penicillin / streptomycin The medium was replaced with the one.
(2) Quantification of neurite formation In a study examining the effect of enhancing neurite outgrowth, poly-D-lysine / laminin coated 24-well tissue culture plates were placed at a relatively low density (0.25 × 10 4). Cells / cm 2 ) PC12 cells were seeded. After adding NGF (2.5 ng / ml) and further culturing for 4 days with or without the addition of various drugs, morphometric analysis was performed using digital images of live cells taken with a phase contrast inverted microscope equipped with a camera. . Three fields of view were taken per well. The average number of cells per field was 100. The number of differentiated cells in each visual field was counted by visual recognition. Only cells having one or more neurites with a length equal to the cell body diameter were counted and expressed as a percentage of the total number of cells in the field of view. Measurements were performed blind.
(3) Quantification of protein expression In Western blot analysis, NGF (2.5 ng / ml) was added, and after further culturing for 4 days with or without the addition of various drugs, proteins from cell lysates (total 30 μg) were added. Separation was performed by 7.5% polyacrylamide gel electrophoresis (Bio-Rad, Hercules, CA, USA). The protein was transferred to a polyvinylidene fluoride membrane (GE Healthcare) using a Tank-transfer apparatus (Bio-Rad), and this membrane was transferred to TBST (50 mM Tris / HCl, 5% FBS, 5% skim milk, 2% BSA, 5% FBS). Blocked with pH 7.8, 0.13 M NaCl, 0.1% Tween® 20). Rabbit anti-eIF4A antibody (Abcam) and mouse anti-β-actin antibody (Sigma Aldrich) diluted at 1: 250 and 1: 10000 in the blocking buffer were used as primary antibodies. Furthermore, an anti-rabbit IgG horseradish peroxidase labeled antibody (GE Healthcare) diluted 25,000 times in TBST containing 5% skim milk and an anti-mouse IgG horseradish peroxidase labeled antibody (GE Health) diluted 25000 times in TBST containing 5% skim milk. Care) was used as the secondary antibody. Antigen proteins on the membrane were detected with ECL ™ Plus Western Blotting Detection System (GE Healthcare). Band visualization with a cooled CCD camera was performed with LAS3000mini (Fuji Photo Film), and protein expression was quantified with analysis software Multi Gauge (Fuji Photo Film).
(4) Gene knockdown by siRNA The expression of eIF4A1 or eEF1A1 was suppressed using siRNA targeting eIF4A1 or eEF1A1. The chemically synthesized siRNA was purchased from a reagent company (Integrated DNA Technologies, Coralville, CA, USA) (FIGS. 10 and 11). The antisense sequence and the sense sequence of siRNA subjected to the experiment are shown in SEQ ID NO: 1 to 6 and SEQ ID NO: 7 to 12, respectively. PC12 cells were cultured and transfected with the above siRNA using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) and OPTI-MEM (Invitrogen, Carlsbad, CA, USA). 6 hours later, NGF (2.5 ng / ml), and further exchange with DMEM medium containing 0.5% FBA, 1% penicillin / streptomycin with or without the addition of various drugs, to examine the neurite outgrowth enhancing effect And subjected to Western blot analysis.
(5) Statistical analysis Data are shown as mean ± standard deviation. Statistical analysis was performed using one-way analysis of variance and post-hoc Bonferroni test. A p value of 0.05 or less was considered statistically significant.
［実験結果］
（１）ＮＧＦ誘発神経突起伸長に対するミノサイクリン、シロスタゾールの作用
　ミノサイクリン、シロスタゾールは、ＮＧＦ（２．５ｎｇ／ｍＬ）による神経突起伸長を示したＰＣ１２細胞の数を濃度依存的に有意に増加させた（図１、図５）。一方、テトラサイクリン、シロスタミドでは、神経突起伸長を示した細胞数の増加は見られなかった（図１、図５）。
（２） ｅＩＦ４Ａ１およびｅＥＦ１Ａ１タンパク質発現定量化
　ミノサイクリンは、ＰＣ１２細胞中のｅＩＦ４Ａ１を有意に増加させた（図２）。シロスタゾールは、ＰＣ１２細胞中のｅＥＦ１Ａ１を有意に増加させた（図６）。
（３）ｓｉＲＮＡによる遺伝子ノックダウン
　ミノサイクリンまたはシロスタゾールは、それぞれＰＣ１２細胞中のｅＩＦ４Ａ１またはｅＥＦ１Ａ１を有意に増加させ、それぞれのｓｉＲＮＡにより遺伝子をノックダウンさせると、ＰＣ１２細胞中のｅＩＦ４Ａ１またはｅＥＦ１Ａ１の増加は見られなかった。さらに、ターゲットとは全く関係のないｓｉＲＮＡ（ネガティブＲＮＡ）を作用させてもＰＣ１２細胞中のｅＩＦ４Ａ１またはｅＥＦ１Ａ１の増加に影響はなかった （図３、図７）。
　さらに、ミノサイクリンまたはシロスタゾールは、（２．５ｎｇ／ｍＬ）による神経突起伸長を示したＰＣ１２細胞の数を濃度依存的に有意に増加させたが、ｓｉＲＮＡ処理により遺伝子をノックダウンさせると、神経突起伸長を示した細胞数を増加させなかった。さらに、ターゲットとは全く関係のないｓｉＲＮＡ（ネガティブＲＮＡ）を作用させても神経突起伸長を示した細胞数の増加に影響はなかった （図４、図８）。
　一方、テトラサイクリンまたはミノサイクリンは、ｅＥＦ１Ａ１タンパク量には影響を与えなかった（図９）。また、シロスタミドまたはシロスタゾールは、ｅＩＦ４Ａ１タンパク量には影響を与えなかった（図９）。 [Experimental result]
(1) Effects of minocycline and cilostazol on NGF-induced neurite outgrowth Minocycline and cilostazol significantly increased the number of PC12 cells that showed neurite outgrowth by NGF (2.5 ng / mL) in a concentration-dependent manner (Fig. 1, FIG. 5). On the other hand, tetracycline and cilostamide did not show an increase in the number of cells showing neurite outgrowth (FIGS. 1 and 5).
(2) eIF4A1 and eEF1A1 protein expression quantification Minocycline significantly increased eIF4A1 in PC12 cells (FIG. 2). Cilostazol significantly increased eEF1A1 in PC12 cells (FIG. 6).
(3) Gene knockdown by siRNA Minocycline or cilostazol significantly increased eIF4A1 or eEF1A1 in PC12 cells, respectively, and when gene was knocked down by the respective siRNA, an increase in eIF4A1 or eEF1A1 in PC12 cells was seen There wasn't. Furthermore, even if siRNA (negative RNA) that had nothing to do with the target was allowed to act, the increase in eIF4A1 or eEF1A1 in PC12 cells was not affected (FIGS. 3 and 7).
Furthermore, minocycline or cilostazol significantly increased the number of PC12 cells that showed neurite outgrowth by (2.5 ng / mL) in a concentration-dependent manner, but neurite outgrowth when the gene was knocked down by siRNA treatment Did not increase the number of cells that showed. Furthermore, even when siRNA (negative RNA) that had nothing to do with the target was allowed to act, the increase in the number of cells showing neurite outgrowth was not affected (FIGS. 4 and 8).
On the other hand, tetracycline or minocycline did not affect the amount of eEF1A1 protein (FIG. 9). Moreover, cilostamide or cilostazol did not affect the amount of eIF4A1 protein (FIG. 9).
　上述した結果より、ＮＧＦ誘発神経突起伸長作用モデルにおいてミノサイクリンまたはシロスタゾールが有意に増強作用を示したこと、およびその増強作用に翻訳開始因子または翻訳伸長因子が関与していることから、翻訳開始因子または翻訳伸長因子を増加するような化合物が、将来新規治療薬として有用であることが理解される。したがって、翻訳開始因子もしくは翻訳伸長因子を増加するような化合物、またはその薬学的に許容し得る塩を含有する本発明の薬剤は、精神疾患、神経疾患、脳血管疾患、循環器疾患、呼吸器疾患、内分泌疾患、代謝疾患、血液疾患、免疫疾患、感染症、および悪性腫瘍などを含む様々な疾患の予防薬および／または治療薬として有効である。
　さらに、ミノサイクリンおよび／またはシロスタゾールが神経突起伸長増強剤として使用できる。神経突起伸長増強剤としての使用は、ミノサイクリンにおいては、翻訳開始因子、その中でもｅＩＦ４Ａ１の特異的増加を伴うものであり、また、シロスタゾールにおいては、翻訳伸長因子、その中でもｅＥＦ１Ａ１の特異的増加を伴うものであった。さらに、これらより、ミノサイクリンの翻訳開始因子増加剤、その中でもｅＩＦ４Ａ１の増加剤としての使用が可能であり、また、シロスタゾールの翻訳伸長因子増加剤、その中でもｅＥＦ１Ａ１の増加剤としての使用が可能となった。神経突起伸長増強剤の伸長増強作用は、翻訳開始因子または翻訳伸長因子にそれぞれ独立に係わっていることから、ミノサイクリンおよびシロスタゾールの併用効果も図れる。並びに、翻訳開始因子および／または翻訳伸長因子の異常に起因する種々の疾患、とりわけ神経疾患の予防剤および／または治療剤として、ミノサイクリンおよび／またはシロスタゾールが使えることが明らかとなった。 From the results described above, it was shown that minocycline or cilostazol showed a significant enhancing action in the NGF-induced neurite elongation action model, and that the translation initiation factor or translation elongation factor was involved in the enhancement action. It will be appreciated that compounds that increase translation elongation factors will be useful as new therapeutic agents in the future. Therefore, a compound of the present invention containing a compound that increases a translation initiation factor or translation elongation factor, or a pharmaceutically acceptable salt thereof, is used in mental diseases, neurological diseases, cerebrovascular diseases, cardiovascular diseases, respiratory organs It is effective as a prophylactic and / or therapeutic agent for various diseases including diseases, endocrine diseases, metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors.
In addition, minocycline and / or cilostazol can be used as neurite outgrowth enhancers. Use as a neurite outgrowth enhancer is accompanied by a specific increase in translation initiation factors, particularly eIF4A1, in minocycline, and a specific increase in translation elongation factors, particularly eEF1A1, in cilostazol. It was a thing. Furthermore, from these, it is possible to use minocycline as a translation initiation factor increasing agent, particularly eIF4A1 increasing agent, and cilostazol as a translation elongation factor increasing agent, particularly eEF1A1 increasing agent. It was. Since the elongation enhancing action of the neurite elongation enhancing agent is independently related to the translation initiation factor or translation elongation factor, the combined effect of minocycline and cilostazol can also be achieved. In addition, it has been revealed that minocycline and / or cilostazol can be used as a preventive and / or therapeutic agent for various diseases caused by abnormal translation initiation factors and / or translation elongation factors, particularly neurological diseases.
　上述の記載にて説明したとおり、本発明の翻訳開始因子または翻訳伸長因子を増加するような化合物は、精神疾患、神経疾患、脳血管疾患、循環器疾患、呼吸器疾患、内分泌疾患、代謝疾患、血液疾患、免疫疾患、感染症、および悪性腫瘍などの種々の疾患に有効な新規医薬品として用いることができる。 As explained in the above description, the compound that increases the translation initiation factor or translation elongation factor of the present invention is a mental disease, neurological disease, cerebrovascular disease, cardiovascular disease, respiratory disease, endocrine disease, metabolic disease. It can be used as a novel pharmaceutical effective for various diseases such as blood diseases, immune diseases, infectious diseases, and malignant tumors.
配列番号１：設計されたオリゴヌクレオチド。
配列番号２：設計されたオリゴヌクレオチド。
配列番号３：設計されたオリゴヌクレオチド。
配列番号４：設計されたオリゴヌクレオチド。
配列番号５：設計されたオリゴヌクレオチド。
配列番号６：設計されたオリゴヌクレオチド。
配列番号７：設計されたオリゴヌクレオチド。
配列番号８：設計されたオリゴヌクレオチド。
配列番号９：設計されたオリゴヌクレオチド。
配列番号１０：設計されたオリゴヌクレオチド。
配列番号１１：設計されたオリゴヌクレオチド。
配列番号１２：設計されたオリゴヌクレオチド。 SEQ ID NO: 1 designed oligonucleotide.
SEQ ID NO: 2: designed oligonucleotide.
SEQ ID NO: 3: Designed oligonucleotide.
SEQ ID NO: 4: Designed oligonucleotide.
SEQ ID NO: 5: designed oligonucleotide.
SEQ ID NO: 6: designed oligonucleotide.
SEQ ID NO: 7 designed oligonucleotide.
SEQ ID NO: 8 designed oligonucleotide.
SEQ ID NO: 9: designed oligonucleotide.
SEQ ID NO: 10 designed oligonucleotide.
SEQ ID NO: 11 designed oligonucleotide.
SEQ ID NO: 12: designed oligonucleotide.

Claims (11)

  1. 　ミノサイクリンおよび／またはシロスタゾールを含有する神経突起伸長増強剤。 A neurite outgrowth enhancer containing minocycline and / or cilostazol.
  2. 　翻訳開始因子および／または翻訳伸長因子を増加させることを特徴とする神経突起伸長増強剤。 A neurite elongation enhancer characterized by increasing a translation initiation factor and / or a translation elongation factor.
  3. 　翻訳開始因子であるｅＩＦ４Ａアイソフォームおよび／または翻訳伸長因子であるｅＥＦ１Ａアイソフォームを増加させることを特徴とする神経突起伸長増強剤。 A neurite elongation enhancer characterized by increasing eIF4A isoform as a translation initiation factor and / or eEF1A isoform as a translation elongation factor.
  4. 　翻訳開始因子であるｅＩＦ４Ａ１および／または翻訳伸長因子であるｅＥＦ１Ａ１を増加させることを特徴とする神経突起伸長増強剤。 A neurite elongation enhancer characterized by increasing eIF4A1 which is a translation initiation factor and / or eEF1A1 which is a translation elongation factor.
  5. 　請求項１から４のいずれか１項に記載の神経突起伸長増強剤の精神疾患治療への使用。 Use of the neurite outgrowth enhancing agent according to any one of claims 1 to 4 for treating psychiatric disorders.
  6. 　ミノサイクリンおよび／またはシロスタゾールを含有する、翻訳開始因子増加剤および／または翻訳伸長因子増加剤。 A translation initiation factor increasing agent and / or a translation elongation factor increasing agent containing minocycline and / or cilostazol.
  7. 　ミノサイクリンを含有する翻訳開始因子ｅＩＦ４Ａ１の増加剤。 Increaser of translation initiation factor eIF4A1 containing minocycline.
  8. 　シロスタゾールを含有する翻訳伸長因子ｅＥＦ１Ａ１の増加剤。 An agent for increasing translation elongation factor eEF1A1 containing cilostazol.
  9. 　ミノサイクリンおよび／またはシロスタゾールを含有する、翻訳開始因子および／または翻訳伸長因子の異常に起因する疾患の予防剤および／または治療剤。 An agent for preventing and / or treating a disease caused by an abnormality in a translation initiation factor and / or translation elongation factor, comprising minocycline and / or cilostazol.
  10. 　ミノサイクリンおよび／またはシロスタゾールを含有する、翻訳開始因子であるｅＩＦ４Ａ１アイソフォームおよび／または翻訳伸長因子であるｅＥＦ１Ａ１アイソフォームの減少に由来する精神疾患、神経疾患、脳血管疾患、循環器疾患、呼吸器疾患、内分泌疾患、代謝疾患、血液疾患、免疫疾患、感染症、および悪性腫瘍からなる群から選択される１種または２種以上の症状を呈する疾患の予防剤および／または治療剤。 Psychiatric disorder, neurological disorder, cerebrovascular disease, cardiovascular disease, respiratory disease derived from a decrease in eIF4A1 isoform which is translation initiation factor and / or eEF1A1 isoform which is translation elongation factor, containing minocycline and / or cilostazol An agent for preventing and / or treating a disease exhibiting one or more symptoms selected from the group consisting of endocrine disease, metabolic disease, blood disease, immune disease, infectious disease, and malignant tumor.
  11. 　ミノサイクリンおよび／またはシロスタゾールを含有する、翻訳開始因子であるｅＩＦ４Ａ１および／または翻訳伸長因子であるｅＥＦ１Ａ１の減少に由来する精神疾患、神経疾患、脳血管疾患、循環器疾患、呼吸器疾患、内分泌疾患、代謝疾患、血液疾患、免疫疾患、感染症、および悪性腫瘍からなる群から選択される１種または２種以上の症状を呈する疾患の予防剤および／または治療剤。 Psychiatric diseases, neurological diseases, cerebrovascular diseases, cardiovascular diseases, respiratory diseases, endocrine diseases derived from a decrease in eIF4A1 which is a translation initiation factor and / or eEF1A1 which is a translation elongation factor, containing minocycline and / or cilostazol, A preventive and / or therapeutic agent for a disease exhibiting one or more symptoms selected from the group consisting of metabolic diseases, blood diseases, immune diseases, infectious diseases, and malignant tumors.
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