WO2006070874A1 - Agent favorisant une réponse hypoxique - Google Patents

Agent favorisant une réponse hypoxique Download PDF

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Publication number
WO2006070874A1
WO2006070874A1 PCT/JP2005/024083 JP2005024083W WO2006070874A1 WO 2006070874 A1 WO2006070874 A1 WO 2006070874A1 JP 2005024083 W JP2005024083 W JP 2005024083W WO 2006070874 A1 WO2006070874 A1 WO 2006070874A1
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amino acid
leucine
isoleucine
hif
hypoxic response
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PCT/JP2005/024083
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English (en)
Japanese (ja)
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Hirotoshi Tanaka
Kenji Takehana
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Toudai Tlo, Ltd.
Ajinomoto Co., Inc.
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Publication of WO2006070874A1 publication Critical patent/WO2006070874A1/fr

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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
    • A61K31/185—Acids; Anhydrides, halides or salts thereof, e.g. sulfur acids, imidic, hydrazonic or hydroximic acids
    • A61K31/19—Carboxylic acids, e.g. valproic acid
    • A61K31/195—Carboxylic acids, e.g. valproic acid having an amino group
    • A61K31/197—Carboxylic acids, e.g. valproic acid having an amino group the amino and the carboxyl groups being attached to the same acyclic carbon chain, e.g. gamma-aminobutyric acid [GABA], beta-alanine, epsilon-aminocaproic acid or pantothenic acid
    • A61K31/198—Alpha-amino acids, e.g. alanine or edetic acid [EDTA]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/04—Drugs for disorders of the alimentary tract or the digestive system for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
    • 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
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A61P27/02—Ophthalmic agents
    • 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
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
    • 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
    • A61P31/12—Antivirals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P33/00—Antiparasitic 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
    • A61P37/02—Immunomodulators
    • A61P37/04—Immunostimulants
    • 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
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/04—Inotropic agents, i.e. stimulants of cardiac contraction; Drugs for heart failure
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis

Definitions

  • the present invention relates to a hypoxic response promoter and a pharmaceutical composition containing the same, and in particular, the drug is preferably used in the form of a pharmaceutical product and a beverage, or in the form of a food or drink. Rukoto can.
  • HIF-1a Hypoxia-inducible factor-1 ⁇
  • HIF-1a Hypoxia-inducible factor-1 ⁇
  • HIF-1 a translocated to the nucleus binds to the hypoxia responsive region of the response gene cluster and induces gene expression at the transcriptional level.
  • the increase in the amount of HIF-1 ⁇ protein is regulated not only by increasing stability but also by promoting translation, and mTOR (mammalian target of rap amycin) is thought to be involved in the upstream of the signal cascade. ing.
  • HIF-1a has been found as a transcriptional activity factor of erythropoietin that regulates erythropoiesis.
  • HIF-1a was later found to be a promoter of vascular endothelial growth factor VEGF, enolase 1, transferrin, aldolase A, etc. The binding site of ⁇ was strong (Non-patent Document 4).
  • Non-patent text 5 It is widely known that amino acids promote protein synthesis in cells by activating mTOR as a signal that only becomes a protein substrate (Non-patent Document 6, etc.).
  • Non-patent Document 7 the low molecular weight compound that activates mTOR such as amino acids 1S through the activation of HIF-1a in cells, whether or not the expression of HIF-1a target genes such as VEGF is enhanced. Is known and obscene.
  • angiogenesis is recognized as a result of physiological hypoxic response in organs and tissues that have become ischemic due to causes such as arteriosclerosis and blood clots.
  • it is considered that the induction of angiogenesis by using some drug that is often inadequate is a treatment for hypoxic disorders such as ischemia.
  • treatment with a protein preparation that induces angiogenesis such as VEGF and gene therapy for VE GF and HIF-1 ⁇ have been studied (Non-patent Document 8).
  • VEGF vascular endothelial growth factor-1 ⁇
  • HIF-1a hypoxia-inducible factor-1a
  • VEGF vascular endothelial growth factor-1a
  • various other factors to react to form more normal vasculature than when only VEGF alone acts.
  • edema, inflammation, ulcers and tumors that occur when angiogenic factors are directly administered alone do not occur, and increasing the activity of HIF-1 ⁇ can induce physiological angiogenesis.
  • Non-patent document 9 Non-patent document 9).
  • the power that athletes often perform high altitude training in athletics is training aimed at increasing the oxygen supply to each body tissue. If the living body can be guided to a state similar to high oxygen at low altitudes even on flat ground, high altitude training and altitude training It is considered that the same effect can be expected.
  • Non-patent literature 1 Semenza GL. Hypoxia-inducible factor 1: oxygen homeostasis and dis ease pathophysiology. Trends Mol Med. 2001 Aug; 7 (8): 345-50
  • Non-Patent Document 2 Humar R, Kiefer FN, Berns H, Resink TJ, Battegay EJ.Hypoxia enhan ces cell blood cell and angiogenesis in vitro via rapamycin, m fOR) -dep endent signaling.FASEB J. 2002 Jun; 16 (8 ): 771- 80
  • Non-Patent Document 3 Treins C, Giorgett Peraldi S, Murdaca J, Semenza GL, Van Obbergh en E. Insulin stimulates hypoxia-inducible factor 1 through a phosphatidylinositol 3 -kinase / target of rapamycin- dependent signaling pathway.J Biol Chem. 2002 Aug 2; 277 (31): 27975-81.
  • Non-Patent Document 4 Wenger RH, Gassmann M. Oxygen (es) and the hypoxia-inducible fact or- 1. Biol Chem. 1997 Jul; 378 (7): 609-16
  • Non-Patent Document 5 Nat Med. 2004; 10 (6): 594-601
  • Non-Patent Document 6 Biochem Biophys Res Commun. 2004 Jan 9; 313 (2): 429-36
  • Non-Patent Document 7 Earle KA, Pancholi S, Vernon P, Yudkin JS. Amino acid depletion mo dulates vascular endothelial growth factor production during the life span of human v ascular smooth muscle cells.J Cell Physiol. 1998 Aug; 176 (2) : 359-64
  • Non-Patent Document 8 Nature Reviews Drug Discovery 2, 863 -872 (2003), Curr Opin Mol T Her. 2004 Apr; 6 (2): 151-9.
  • Non-Patent Document 9 Elson DA, Thurston G, Huang LE, Ginzinger DG, McDonald DM, Joh nson RS, naval JM.Induction of hypervascularity without leakage or inflammation in transgenic mice over expressing hypoxia-inducible factor— 1 alpha. Genes Dev. 2001 Oct l; 15 (19): 2520-32
  • the object of the present invention is to provide a disease treatment Z preventive agent or food having excellent hypoxic response promoting activity.
  • Another object of the present invention is to provide a therapeutic and / or preventive agent for ischemic disease and anemia, and / or an immunostimulatory agent.
  • a specific amino acid preferably a specific branched-chain amino acid has an excellent hypoxic response promoting activity, particularly the transcription factor HIF- 1 It has been found that it has an ⁇ -active activity, and further induces gene expression of VEGF and ADM, which is one of the hypoxia-responsive genes and is known to be activated by HIF-1a. Was completed.
  • the present invention provides a hypoxic response promoter characterized by containing an amino acid that also has a leucine, isoleucine, norin, or mixture power selected.
  • the present invention also provides a therapeutic and / or preventive agent for ischemic disease and / or an immunostimulatory agent characterized by comprising the hypoxic response promoter.
  • the present invention also provides a therapeutic agent for highland life hypoxia-induced disorder or a highland training aid, characterized by containing an amino acid for which leucine, isoleucine, norin, or a mixture thereof is also selected.
  • the present invention also provides a method for screening a hypoxic response promoter characterized by including the following steps.
  • the present invention also provides an amino acid-containing drug or food containing an amino acid that is selected for leucine, isoleucine, norin, or a mixture thereof and packaged in a package that describes the effect of assisting in training at high altitude.
  • the present invention also includes an mTOR activator, a HIF-1 activator or a vascular endothelial growth factor (VEGF) characterized in that it comprises an amino acid that is also selected for leucine, isoleucine, norrin or a mixture thereof.
  • VEGF vascular endothelial growth factor
  • Gene and Z or adrenomodullin (ADM) gene expression inducers are provided.
  • Ischemic disease therapeutic agent and / or anemia treatment agent and / or immunostimulatory agent and / or high altitude life hypoxia-induced disorder therapeutic agent and / or highland containing the hypoxic response promoter of the present invention In the case of training aids or other pharmaceutical or food compositions, mTOR activators per formulation, e.g. leucine, isoleucine, norin or mixtures thereof. It is preferable to contain about 0.000001-50g, for example. More preferably, 1 to: about LOg is preferably contained.
  • the L-form is preferably used as the amino acid.
  • oral administration intravenous administration, subcutaneous administration, or intramuscular administration can be performed, but oral administration is also preferred for convenience.
  • the dose varies depending on the symptom, age and administration method of the patient to be administered, but is usually 0.1 to 30 gZkgZ days.
  • the therapeutic / preventive pharmaceutical or food of the present invention can be formulated by a conventional method.
  • the dosage form include injections, tablets, granules, fine granules, powders, capsules, creams, suppositories, etc.
  • the carrier for the preparation include lactose, glucose, D-mannthol.
  • the L-form is also desirable from the viewpoint that it exists in nature.
  • the amino acid in the hypoxic response promoter of the present invention avoids amino acid imbalance in the body at the time of taking, so that leucine, isoleucine, and parin are mixed in an appropriate blending ratio rather than leucine alone, for example, 100 mass of leucine.
  • a branched chain amino acid in which 10 to 100 parts by mass of isoleucine and 10 to 100 parts by mass of valine are mixed per part is desirable.
  • the hypoxic response promoter of the present invention may be an amino acid alone or may be contained with a general food.
  • the pharmaceutical agent used for the purpose of treatment or prevention of various disease abnormalities targeted by the present invention is used.
  • the above-mentioned calculation range is determined as an active ingredient, for other purposes, such as the need for a normal diet, or for the treatment of other diseases, amino acids that are ingested or administered, This need not be included in the calculation.
  • hypoxic response promoter of the present invention improves ischemic disease and its complications caused by hypoxia through the activity of HIF-1 ⁇ , which is a major regulator of hypoxic response. Can be relaxed, recovered.
  • medical agent of this invention can be used as a pharmaceutical, it can also be used as a foodstuff. Therefore, the present invention can be widely used particularly in the fields of pharmaceuticals, foods and the like, and therefore the present invention is very useful.
  • Ischemic diseases include myocardial infarction, heart failure, cerebral infarction, cerebral hyperemia, diabetic vascular disorder, obstructive arteriosclerosis colitis, ulcer, spinal cord disorder? Examples include sight neuropathy or neuropathy.
  • Examples of immunity that can be activated by an immunostimulator containing a hypoxic response promoter include tumor immunity, bacterial infection immunity, parasitic infection immunity, and viral infection immunity. It is.
  • the present invention also provides a method for screening a hypoxic response promoter characterized by including the following steps.
  • the amino acids preferably contain leucine, isoleucine, norin or a mixture of those amino acids, and it is particularly preferable to use only these amino acids.
  • cultured cells established cultured cells such as ⁇ 4 ⁇ cells can be used, but normal human-derived cells that are physiological VEGF-producing cells such as vascular endothelial cells and vascular smooth muscle cells are used. For example, it is preferable to culture at a temperature of 30 to 37.
  • Example 1 Induction of VEGF gene expression by leucine and BCAA
  • PRMI-AA a medium in which glucose was adjusted to 4.5 g / L except for all amino acids based on RPMI1640 was used.
  • the PRMI-AA medium was adjusted as follows. In advance, 4X Inorganic Slats Soln. And lOOXvitamin Soln. Were adjusted and stored as follows, and other components and 4X Inorganic Salts Soln. Were added to double-stage distilled water to a final concentration of IX. Depending on the experimental conditions, PRMI-AA medium supplemented with a final concentration of 800 M leucine was used.
  • 4XRPMI base Medium (glucose concentration: 4.5g / l) Created by AA
  • oral isine (800 M: Fig. 1) or BCAA final concentrations 800 ⁇ leucine, 400 ⁇ isoleucine, 480 ⁇ ⁇ parin: Fig.
  • the cDNA synthesis of the template used for Taqman PCR was performed using Superscript First-Strand Synthesis System for RT-PCR (manufactured by GIBCO BRL). 500 ng of total RNA, 0.5 ⁇ / ⁇ ⁇ 01igo (dT) 12-18 1 ⁇ 1, lOmMdNTPmixl ⁇ 1 was dissolved in DEPC-treated water to a total volume of 10 ⁇ 1. After 5 minutes of reaction at 65 ° C, cool down, 10XRT Buffer 21, 1, 25 mM MgCl 4, 1, 0
  • Primers were designed for the human VEGF gene and human 13-actin as a housekeeping gene as a control.
  • the external database Primer3 shown below was used for the design.
  • Table 1 shows the GenBank numbers, gene names, and primer base sequences of human VEGF gene and human j8 actin gene.
  • the reaction solution having the composition shown in Table 2 was mixed in a PCR tube for Taqman, and PCR reaction was performed in ABI7700 Prism Sequence Dtector.
  • the reaction conditions were as follows.
  • Reaction conditions 50 ° C 2 minutes ⁇ 95 ° C 10 minutes ⁇ (95 ° C 15 seconds ⁇ 60 ° C 1 minute) 40 cycles.
  • VEGF gene expression was induced by leucine, and the VEGF mRNA level was increased approximately 3-fold in 8 hours (Fig. 1). Similar results were obtained with leucine, isoleucine, and amino acid combinations of phosphorous (Fig. 2). ⁇ Amino acid composition without leucine Such an increase in VEGF expression was not observed (Fig. 3). The increase in VEGF expression by leucine was inhibited by treatment with the mTOR inhibitor rapamycin ( Figure 4).
  • VEGF ability to increase the production of VE GF in vivo by administering VEGF as a protein preparation or gene therapy of the VEGF gene
  • various ischemic diseases such as obstructive arteriosclerosis and heart failure, high treatment Because it is effective (Nature Reviews Drug Discovery 2, 863 -872 (2003), Curr Opin Mol Ther. 2004 Apr; 6 (2): 151— 9.)
  • it is as safe as an amino acid.
  • the expression of endogenous VEGF can be increased in vivo by a drug with excellent oral absorbability, it promotes hypoxic stress response to prevent cell death and improve energy metabolism. It is clear that a therapeutic effect can be obtained.
  • Example 2 Increase in the amount of HIF-1 ⁇ protein under hypoxic stress by amino acids
  • PBMC peripheral blood mononuclear cells
  • the composition of the IX medium was lOOmg / L Ca (NO) ⁇ 4 ⁇ 0, 400mg / L, 48.84mg / L MgSO, 6000mg / L NaCl, 800mg / L NaH
  • the degree was defined as IX, and 1 / 2X, 1 / 4X, 2X, 3X, 4X media were prepared.
  • the amino acid removal medium (-AA) is a medium excluding all the above amino acids.
  • BCAA-free medium (-BCAA) is a medium obtained by removing Valine, Leucine and Isoleucine from the above IX medium, and leucine-free medium (-LEU) is a medium obtained by removing Leucine from the above IX medium.
  • FCS heat-inactivated fetal calf serum
  • FCS heat-inactivated fetal calf serum
  • T cells cultured at a concentration of 2xl0 6 cells / ml with RPMI 1640 were collected, centrifuged, and the supernatant was removed. The supernatant was further washed once with PBS, and adjusted with a medium having a different amino acid composition.
  • the whole cell extract was prepared, and HIF-1a specific antibody (Ab463: Abcam), phosphorylated S6K specific antibody, phosphorylated eIF2a antibody, phosphorylated Akt Immunoblotting was performed using various antibodies (Cell Signaling Technology) such as specific antibodies.
  • HIF-1a specific antibody Ab463: Abcam
  • phosphorylated S6K specific antibody phosphorylated S6K specific antibody
  • phosphorylated eIF2a antibody phosphorylated Akt Immunoblotting
  • the total RNA extract was prepared and detected by RT-PCR according to the method described in J. Immunology, 171: 6534-6540 (2003) using primers specific for each mRNA.
  • FIG. 5 shows the effect of amino acid concentration in the medium on the expression level of HIF-1a protein under low oxygen concentration.
  • normal oxygen concentration is reduced to low oxygen concentration.
  • the power of increasing HIF-1a protein after 2 hours. This effect was lost when all amino acids in the medium were removed.
  • phosphorylation of S6K1 existing downstream of signal transduction by amino acids was lost, and phosphorylation of eIF2a present downstream of signal transduction in response to amino acid starvation was enhanced.
  • the amino acid concentration in the medium was varied, an increase in the amount of HIF-1a protein was observed depending on the amino acid concentration. This indicates that amino acids are essential for the expression of HIF-1a protein.
  • Fig. 6 shows the amount of HIF-1 ⁇ protein 2 hours after removal of BCAA alone (-BCAA) from RPMI1640 medium containing all amino acids under low oxygen (1% oxygen partial pressure) It is.
  • the removal of BCAA alone showed the same effect as the removal of all amino acids, indicating that BCAA is particularly important among amino acids for the expression of HIF-1a protein.
  • S6 K1 phosphorylation was also reduced Akt's phosphorylation had no effect.
  • Figure 7 shows the intracellular HIF-1 a protein 2 hours after removal of oral isine (-LEU) from RPMI1640 medium containing all amino acids under hypoxia (1% oxygen partial pressure). The amount was examined.
  • BCAA is important among the amino acids, and leucine is the most important for the expression of HIF-1a protein.
  • RNA was extracted 18 hours after hypoxia treatment, and various mRNA levels were examined by RT-PCR.
  • ADM adrenom edullin
  • Transcriptional induction is impaired in leucine-removing medium (-LEU)! / This example shows that leucine or a combination of leucine, isoleucine, and parin-containing amino acid (BCAA) is HIF in cells under hypoxia.
  • ADM adrenomedullin
  • Leucine increases the expression of HIF-1 ⁇ and subsequently induces various ischemic diseases and hypertension through induction of ADM expression It is clear that it is useful in the treatment of
  • HIF-1a the activity of HIF-1a is thought to play an extremely important role in maintaining the survival of activated T cells in peripheral tissues (J. Immunology, 171: 6534-6540 (2003)).
  • T cells In order for T cells to perform biological defense functions such as tumor immunity and immunity against bacterial infection, it is necessary to infiltrate the thread and tissue from the blood vessels and survive locally for a certain period of time. From this, it is clear that leucine or BCAA is indispensable for the function of T cells in vivo, especially in the local area of tissues. Therefore, leucine or BCAA activates T cell immunity through activation of HIF-1a, and enhances tumor immunity, bacterial infection immunity, parasitic immunity immunity, and viral infection immunity. Can improve disease and pathology.
  • Example 3 Enhancement of HIF-1 ⁇ expression in peripheral blood mononuclear cells by oral administration of BCAA in healthy individuals
  • HIF-1 expression in peripheral blood mononuclear cells after oral administration of BCAA was examined by immunoblotting.
  • the subject fasted from 8:00 pm the previous day and took 1 branched-chain amino acid preparation (Libatato Granule (Ajinomoto Co., Inc.) 1 pm the following morning. Blood was collected at 2 pm 6 hours later. In the same way, blood was collected at 2 pm after an overnight fast, and the amount of blood collected at one time was approximately 80 ml, and peripheral blood mononuclear cells were isolated from this blood as follows. did.
  • the supernatant was drained, and 40 ml of PBS was added to each tube, the precipitate was washed, and centrifuged at 1800 rpm lOmin 20 ° C (accelerator fast, brake fast).
  • the supernatant was discarded, suspended in 6 ml of RPMI medium (FCS 10%), transferred to a 15 ml falcon tube, and placed in two 10 cm dishes.
  • HIF-1 alpha specific antibody After adjustment electrophoresis of whole cell extracts, HIF-1 alpha specific antibody: it was Imunoburotto using (Ab463 Abcam Inc.).
  • FIG. 8 shows the results of immunoblotting of HIF-1a.
  • the leftmost lane 1 (PC) is a positive control for HIF-1a protein.
  • HIF-l a protein of peripheral blood mononuclear cells isolated from blood after fasting overnight is hardly detected in hypoxic (1% oxygen partial pressure) culture (lane 2), but in blood treated with BCAA A small amount of HIF-1a protein was detected in peripheral blood mononuclear cells (lane 3).
  • PMA treatment was performed during culture, a large amount of HIF-1 ⁇ protein was detected regardless of the in vivo application of BCAA (lanes 4 and 5). Therefore, it became clear that administration of ⁇ CAA increases the expression level of HIF-1 ⁇ in peripheral blood mononuclear cells.
  • FIG. 1 shows the effect of leucine on VEGF expression.
  • FIG. 2 shows the effect of BCAA on VEGF expression.
  • FIG. 3 shows the effect of essential amino acids without BCAA on VEGF expression.
  • FIG. 4 shows the influence of rapamycin on leucine-induced VEGF expression.
  • FIG. 5 shows the effect of amino acid concentration in the medium on the expression level of HIF-1 a protein under low oxygen concentration.
  • FIG. 6 shows the amount of HIF-1 a protein after 2 hours when only BCAA is removed from RPMI1640 medium containing all amino acids under low oxygen (1% oxygen partial pressure).
  • FIG. 8 shows the results of immunoblotting of HIF-1a.
  • the arrow indicates the band of HIF-1a.

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Abstract

La présente invention a pour objet un agent favorisant une réponse hypoxique, un agent thérapeutique pour les troubles liés à l'hypoxie et survenant à haute altitude, ainsi qu'un complément alimentaire pour entraînement à haute altitude contenant un acide aminé sélectionné parmi la leucine, l'isoleucine, la valine ou un mélange de ces acides aminés, et un agent prophylactique ou thérapeutique destiné à traiter les maladies ischémiques ou un agent immunostimulant contenant ledit agent favorisant une réponse hypoxique.
PCT/JP2005/024083 2004-12-28 2005-12-28 Agent favorisant une réponse hypoxique WO2006070874A1 (fr)

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Citations (13)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH01301619A (ja) * 1988-05-30 1989-12-05 Otsuka Pharmaceut Factory Inc 癌用アミノ酸製剤
JPH02243621A (ja) * 1989-03-16 1990-09-27 Ajinomoto Co Inc 虚血性脳障害治療薬
JPH04264024A (ja) * 1990-10-30 1992-09-18 Nb Internatl Technol 虚血性心臓組織の保護および代謝回復のための方法および組成物
JP2000026290A (ja) * 1998-07-07 2000-01-25 Crescendo Corporation:Kk 分岐鎖アミノ酸による筋力維持
JP2000026289A (ja) * 1998-07-01 2000-01-25 Crescendo Corporation:Kk 分岐鎖アミノ酸による筋肉痛・筋肉のこり、はりへの効果
JP2001169752A (ja) * 1999-12-15 2001-06-26 Fancl Corp 食品組成物
JP2002003372A (ja) * 2000-06-20 2002-01-09 Ajinomoto Co Inc 造血および栄養状態改善用アミノ酸組成物
JP2002223732A (ja) * 2001-01-29 2002-08-13 Ajinomoto Co Inc 液体流動食
JP2003238401A (ja) * 2002-02-20 2003-08-27 Ajinomoto Co Inc 疾患の治療、改善又は予防用医薬品及び飲食品
WO2003084545A1 (fr) * 2002-04-09 2003-10-16 Eisai Co., Ltd. Medicament contenant un compose a base de riboflavine
JP2004501968A (ja) * 2000-07-04 2004-01-22 プロフェッショナル・ダイエテティクス・ソシエタ・ア・レスポンサビリタ・リミタータ アミノ酸を基本成分とする心不全の処置に適当な組成物
JP2004262924A (ja) * 2003-02-10 2004-09-24 Masami Moriyama インフルエンザウイルス感染予防剤
JP2004534073A (ja) * 2001-06-15 2004-11-11 プロフェッショナル・ダイエテティクス・ソシエタ・ア・レスポンサビリタ・リミタータ 糖尿病患者における心室心筋機能を改善するためのアミノ酸を主成分とする組成物

Patent Citations (13)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH01301619A (ja) * 1988-05-30 1989-12-05 Otsuka Pharmaceut Factory Inc 癌用アミノ酸製剤
JPH02243621A (ja) * 1989-03-16 1990-09-27 Ajinomoto Co Inc 虚血性脳障害治療薬
JPH04264024A (ja) * 1990-10-30 1992-09-18 Nb Internatl Technol 虚血性心臓組織の保護および代謝回復のための方法および組成物
JP2000026289A (ja) * 1998-07-01 2000-01-25 Crescendo Corporation:Kk 分岐鎖アミノ酸による筋肉痛・筋肉のこり、はりへの効果
JP2000026290A (ja) * 1998-07-07 2000-01-25 Crescendo Corporation:Kk 分岐鎖アミノ酸による筋力維持
JP2001169752A (ja) * 1999-12-15 2001-06-26 Fancl Corp 食品組成物
JP2002003372A (ja) * 2000-06-20 2002-01-09 Ajinomoto Co Inc 造血および栄養状態改善用アミノ酸組成物
JP2004501968A (ja) * 2000-07-04 2004-01-22 プロフェッショナル・ダイエテティクス・ソシエタ・ア・レスポンサビリタ・リミタータ アミノ酸を基本成分とする心不全の処置に適当な組成物
JP2002223732A (ja) * 2001-01-29 2002-08-13 Ajinomoto Co Inc 液体流動食
JP2004534073A (ja) * 2001-06-15 2004-11-11 プロフェッショナル・ダイエテティクス・ソシエタ・ア・レスポンサビリタ・リミタータ 糖尿病患者における心室心筋機能を改善するためのアミノ酸を主成分とする組成物
JP2003238401A (ja) * 2002-02-20 2003-08-27 Ajinomoto Co Inc 疾患の治療、改善又は予防用医薬品及び飲食品
WO2003084545A1 (fr) * 2002-04-09 2003-10-16 Eisai Co., Ltd. Medicament contenant un compose a base de riboflavine
JP2004262924A (ja) * 2003-02-10 2004-09-24 Masami Moriyama インフルエンザウイルス感染予防剤

Non-Patent Citations (15)

* Cited by examiner, † Cited by third party
Title
ALAM H. ET AL.: "Follicle-stimulating Hormone Activation of Hypoxia.inducible Factor-1 by the Phosphatidylinositol 3-Kinase/AKT/RAS Homolog Enriched in Brain (Rheb)/Mammalian Target of Rapamycin (mTOR) Pathway Is Necessary for Induction of Select Protein Markers of Follicular Differentiation", J. BIOL. CHEM., vol. 279, no. 19, 2004, pages 19431 - 19440, XP003004549 *
ARSHAM A. M. ET AL.: "A Novel Hypoxia-inducible Factor-independent Hypoxic Response Regulating Mammalian Target of Rapamycin and Its Targets", J. BIOL. CHEM., vol. 278, no. 32, 2003, pages 29655 - 29660, XP003004556 *
EARLE K. A.: "Amino Acid Depletion Modulates Vascular Endothelial Growth Factor Production During the Life Span of Human Vascular Smooth Muscle Cells", JOURNAL OF CELLULAR PHYSIOLOGY, vol. 176, 1998, pages 359 - 364, XP003004555 *
HUDSON C. C. ET AL.: "Regulation of Hypoxia-Inducible Factor 1alpha Expression and Function by the Mammalian Target of Rapamycin", MOLECULAR AND CELLULAR BIOLOGY, vol. 22, no. 20, 2002, pages 7004 - 7014, XP001205925 *
KIMBALL S. R. ET AL.: "Leucine Regulates Translation of Specific mRNAs in L6 Myoblasts through mTOR-mediated Changes in Availability of eIF4E and Phosphorylation of Ribosomal Protein S6", J. BIOL. CHEM., vol. 274, no. 17, 1999, pages 11647 - 11652, XP003004548 *
MARU Y.: "Gan Ten'i to Kekkan Shinsei", JAPANESE JOURNAL OF CANCER AND CHEMOTHERAPY, vol. 26, no. 10, 1999, pages 1509 - 1519, XP003004554 *
MAYERHOFER M. ET AL.: "BCR/ABL induces expression of vascular endothelial growth factor and its transcriptional activator, hypoxia inducible factor-1alpha, through a pathway involving phosphoinositide 3-kinase and the mammalian target of rapamycin", BLOOD, vol. 100, no. 10, 2002, pages 3767 - 3775, XP003004550 *
MINEGISHI N.: "Teisanso Hannosei Ketsueki Kekkan Kyotsu Bunshi No Hatsugen Seigyo", KEKKAN IGAKU, vol. 2, no. 5, 2001, pages 451 - 459, XP003004553 *
MURATA K. ET AL.: "Isoleucine no Daichogan Kan Ten'i Yokusei Koka", ACTA HEPATOLOGICA JAPONICA, vol. 45 SUPPL, no. 1, 2004, pages A206, XP003004557 *
NISHITANI S.: "Bunkisa Amino Acid no Totaisha Kaizen Sayo", JAPANESE JOURNAL OF NUTRITIONAL ASSESSMENT, vol. 20, no. 5, 2003, pages 491 - 494, XP003004546 *
OSHIRO N. ET AL: "Amino acid to saibonai Signal Dentatsu -Tanpanushitsu Gosei o Chushin ni", JAPANSE JOURNAL OF NUTRITIONAL ASSESSMENT, vol. 20, no. 5, 2003, pages 478 - 482, XP003004545 *
TANAKA H. ET AL.: "Tensha Indhi HIF-1alpha", THE LUNG PERSPECTIVES, vol. 12, no. 4, 2004, pages 413 - 423, XP003004551 *
TSUCHIYA T.: "Hypoxic response and transcriptional regulation", THE LUNG PERPECTIVES, vol. 9, no. 3, 2001, pages 313 - 316, XP003004552 *
YONEZAWA K.: "mTOR/raptor-p70S6 Kinase", JAPANSE JOURNAL OF NUTRITIONAL ASSESSMENT, vol. 20, no. 2, 2003, pages 197 - 201, XP003004544 *
YOSHIZAWA T.: "Karadatanpakushitsu Gosei no Hon'yaku Dankai Chosetsu ni Kansuru Eiyo Seikagakuteki Kenkyu", JOURNAL OF JAPANESE SOCIETY OF NUTRITION, AND FOOD SCIENCE, vol. 56, no. 2, 2003, pages 117 - 125, XP003004547 *

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