WO2002031497A1 - Moyen pour examiner l'aptitude à l'angiogénèse - Google Patents

Moyen pour examiner l'aptitude à l'angiogénèse Download PDF

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Publication number
WO2002031497A1
WO2002031497A1 PCT/JP2001/008098 JP0108098W WO0231497A1 WO 2002031497 A1 WO2002031497 A1 WO 2002031497A1 JP 0108098 W JP0108098 W JP 0108098W WO 0231497 A1 WO0231497 A1 WO 0231497A1
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Prior art keywords
cancer
angiogenesis
vegf
endostatin
patient
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PCT/JP2001/008098
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English (en)
Japanese (ja)
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Akikuni Yagita
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Orient Cancer Therapy Co.,Ltd.
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Priority to JP2002534831A priority Critical patent/JPWO2002031497A1/ja
Priority to AU2001288058A priority patent/AU2001288058A1/en
Publication of WO2002031497A1 publication Critical patent/WO2002031497A1/fr

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/5005—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
    • G01N33/5091—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing the pathological state of an organism
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
    • G01N2333/475—Assays involving growth factors
    • G01N2333/50—Fibroblast growth factors [FGF]
    • G01N2333/503—Fibroblast growth factors [FGF] basic FGF [bFGF]
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
    • G01N2333/475—Assays involving growth factors
    • G01N2333/515—Angiogenesic factors; Angiogenin
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
    • G01N2333/705—Assays involving receptors, cell surface antigens or cell surface determinants
    • G01N2333/71—Assays involving receptors, cell surface antigens or cell surface determinants for growth factors; for growth regulators
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
    • G01N2333/78—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin, cold insoluble globulin [CIG]

Definitions

  • An object of the present invention is to provide an optimal technique for a non-invasive treatment means in cancer treatment.
  • a novel immunotherapy established by the present inventors [induction of interleukin 12 (IL-12), activation of natural killer T cells (NKT cells), and biology based on inhibition of angiogenesis] Cancer therapy with biological response modifiers (BRM)) to provide a therapeutic means focusing on the inhibition of vascular neoplasia.
  • BRM biological response modifiers
  • IL-12 itself has an anticancer effect, but there is a fact that if IL-12 itself is directly administered into a living body, side effects occur and patients cannot tolerate treatment, so it can be used as an anticancer drug. I didn't come.
  • Yagida's reported preparations containing AHCC achieved significant healing and prolonging effects in the treatment of cancer.
  • Yagida is IL
  • Yagida aims to develop a new therapeutic agent for cancer that works by a mechanism different from the anticancer effect of AHCC, and is formulated with IL-X (a ratio of Shiitake 2, Suehiro mushroom 2, and Reishi mushroom 1). (Patent pending) and NK T cell activator (patent pending).
  • Yagida focused on the usefulness of shark cartilage as an angiogenesis inhibitor, and achieved optimal formulation (Patent No. 3103513), which has been applied to cancer patients since 1990.
  • Oral ingestion Beta-shark MCZO C: Seishin Co., Ltd.
  • VEGF vascular endothelial growth factor
  • b FGF basic fibroblast growth factor
  • Angiogenesis inhibitors have been reported.
  • VEGF Vascular Endothelial Cell Proliferation Factor
  • the problem to be solved by the present invention is to elucidate the in vivo kinetic relationship between an angiogenesis inhibitor and an angiogenesis promoting factor in the treatment with an angiogenesis inhibitor in the above-described neoimmunotherapy, It is to provide a more reliable means.
  • Yet another object of the present invention is to elucidate the relationship between vascular endothelial cell growth factor and cancer status.
  • the present inventors have performed immunological measurement and vascular endothelial growth factor (VEGF), basic fibroblast proliferation in a case where the new immunotherapy is being treated.
  • the present invention has been completed by measuring the blood kinetics of a factor (b FGF: basic fibroblast growth factor) and endostatin and analyzing the relationship with its therapeutic effect. .
  • the present invention provides
  • VEGF vascular endothelial growth factor
  • bFGF basic fibroblast growth factor
  • a test means for determining the dormancy of cancer characterized by measuring the vascular endothelial growth factor (VEGF) of the patient.
  • VEGF vascular endothelial growth factor
  • a test for compatibility between the angiogenesis inhibitor and / or the anticancer drug and the patient is performed by the test means described in 1 or 2 above.
  • a test or determination means for the effectiveness of the drug is performed by the test means described in 1 or 2 above.
  • FIG. 1 is a graph showing changes in blood concentrations of endostatin, VEGF (vascular endothelial growth factor) and bFGF (basic fibroblast growth factor) from the left side of the figure.
  • VEGF vascular endothelial growth factor
  • bFGF basic fibroblast growth factor
  • FIG. 2 is a graph showing the correlation between VEGF (vascular endothelial growth factor) and blood concentration of endostatin.
  • VEGF vascular endothelial growth factor
  • FIG. 3 is a diagram showing a VEGF / endostatin ratio.
  • a VEGF / endostatin ratio In the figure, before treatment (12 cases), after treatment (29 cases), response cases (CR, PR: 15 cases), effective cases (CR + PR + Long NC: 22 cases), and others (Short NC: 4 cases) , NC / Short NC + Long NC: 11 cases, PD: 3 cases).
  • FIG. 4 is a graph showing the correlation between the blood concentration of jVEGFI (vascular endothelial growth factor) and IL-11 induction.
  • jVEGFI vascular endothelial growth factor
  • FIG. 5 is a graph showing the correlation between the blood concentration of endostatin and induction of IL-12.
  • FIG. 6 is a graph showing the correlation between the blood concentration of VEGF (vascular endothelial growth factor) and IF Ny.
  • FIG. 7 Blood concentration of VEGF (vascular endothelial growth factor).
  • the outline of the new immunotherapy practiced by Asahikuni Yagida, a medical doctor, is as follows.
  • the pillar is immunological cancer treatment with biological response modifiers (BRM preparations), which has made IL-11 induction, activation of NKT cells, and inhibition of angiogenesis.
  • BRM preparations biological response modifiers
  • shark cartilage preparation (patent No.
  • the first pillar As the first pillar, angiogenesis inhibitors, select and improve the best shark cartilage in terms of efficacy, safety, ease of use, and economic efficiency from various basic studies In addition, Better Shear MC or LO was developed. At present, 20 g per day is orally administered on an empty stomach in 2-3 divided doses.
  • the second pillar, IL-12 inducer includes IL-X (formerly AHCC), which acts on mice or humans whose Th1 immune response is mainly functioning, in early cancer and For postoperative cases, for example, about 3 g days, for advanced cancer For example, it is administered orally in about 6 days.
  • krestin which responds to advanced cancer, terminal cancer, and Th2 mouse human, was administered, for example, at about 3 g / day or every other day. Based on these, various natural forms of total vitamin, urso (30 mg or 600 mg / day), and active vitamin D3 (1.0 mg Og Z days) are co-administered. .
  • PSK, OK432 or live BCG vaccines as needed.
  • NKT cells in the patients treated with the above two pillars, complete healing despite no IL-12 production
  • NK T cells are derived from IL-12 (CR: Complete Response) or partial healing (PR: Partial Response) in the same proportion as those in which IL-12 was produced and treatment was effective. Therefore, NK T cells are derived from IL-12 (CR: Complete Response) or partial healing (PR: Partial Response) in the same proportion as those in which IL-12 was produced and treatment was effective. Therefore, NK T cells are derived from IL-12 (CR: Complete Response) or partial healing (PR: Partial Response) were observed in the same proportion as those in which IL-12 was produced and treatment was effective. Therefore, NK T cells are derived from IL-12 (CR: Complete Response) or partial healing (PR: Partial Response) in the same proportion as those in which IL-12 was produced and treatment was effective. Therefore, NK T cells are derived from IL-12 (PR: Partial Response).
  • the T cell receptor Va24 Vj31 1 of NKT cells and the NK cell receptor NKR-P1 was studied.
  • the NKR-P1 receptor activated by saccharides having a 1-3 structure, such as Nig mouth sugar and oligosaccharides showed a positive correlation with IFNy and IL-12.
  • 3 11 activated by lipid showed a negative correlation. It also showed a high correlation with NKR-P1 in CR and PR cases.
  • CTLs cytotoxic T cells
  • another NKT cell might work as effector cells.
  • NITC new immunotherapy
  • NITC new immunotherapy
  • VEGF vascular endothelial growth factor
  • b FGF basic fibroblast growth factor
  • angiogenesis inhibitor endostatin enzyme linked immunosorbent assay
  • gastric cancer 2 prostatic cancer 1 pharyngeal cancer 1 uterine body cancer ovarian cancer 1 ovarian cancer 1 uterine body cancer 1
  • CR and PR may depend on how the angiogenesis promoting factor is suppressed.
  • bFGF which is an angiogenesis promoting factor. Therefore, by measuring the angiogenesis-promoting factor in the serum of the patient, it is possible to examine the angiogenesis of the patient and to test the effect of treating cancer.
  • FIG. 7 shows the results of measuring VEGF in NITC-treated patients.
  • NC 77 cases) 2 9 1 pg Zml decreased (p ⁇ 0.001)
  • PR 55 cases
  • CR 48 cases
  • p ⁇ 0.01 the values were significantly lower in NC, PR and CR.
  • measurement of VEGF is considered to be an important evaluation method for not only PR and CR but also NC dormancy.
  • VEGF assay was performed by ELISA (kit name: Quant). ikinehuman VEGF, R & D Systems).
  • Endostatin has a similar concept to that of angiostatin, that is, the anti-angiogenesis inhibitor produced by tumors in mice. It was separated from Ma. It is a protein with a molecular weight of 20 KDa and is a C-terminal fragment of type XVIII collagen from amino acid analysis. Endostatin production in 12 patients before treatment and 6 CRs in NITC showed significantly lower CR (p ⁇ 0.01). In addition, a significant difference was observed with PD cases (p ⁇ 0.05).
  • pro-angiogenic factors bFGF and VEGF also tend to decrease when they become CR, which is a suitable purpose when considered as an effect of shark cartilage. It is interesting, however, that endostatin production is also decreasing ( Figure 1). Furthermore, when judging the effectiveness of shark cartilage or the usefulness of immunotherapy, it should be judged based on the decrease in endostatin production. More preferably, it would be more reliable to judge based on a decrease in both the angiogenesis promoting factors VEGF and endostatin. In addition, and b FGF instead of VEGF, it may be subjected to judgment Ri by measuring the and E down Dosutachin c
  • VEGF endostatin
  • endostatin The correlation between VEGF and endostatin was examined in 12 cases before treatment, 29 cases during treatment, and 15 responders with CR and PR in NITC (Fig. 2). It can be seen that as the efficacy of NITC increases during the course of treatment, the production of VEGF decreases as compared to endostatin.
  • the ratio of VEGF / endostatin was examined in patients undergoing NITC (Fig. 3), the former significantly decreased after treatment and before treatment.
  • the numerical value of the ratio was further reduced. This suggested that NITC treatment reduced VEGF earlier and more abundantly than endostatin. Therefore, examining the ratio of VEGF / endostatin would allow a more accurate grasp of the state of angiogenesis inhibition.
  • IL-12 has also been found to exhibit an anti-angiogenic effect in the presence of IFNy.
  • a positive correlation was observed in 12 patients before treatment, but a negative correlation was observed after treatment.
  • IL-12 acting as an angiogenesis inhibitory agent or as a result of exerting an immunological antitumor effect. It is inferred that the result of 12 shows an antitumor effect.
  • IFN y and VEGF in patients treated with new immunotherapy The correlation with was examined. As can be seen from the correlation with IL-112, the correlation between IFN and VEGF was inversely correlated even in the treated patients compared to before treatment (Fig. 6). This suggests that 1-12 and 1] ⁇ 0 act together to suppress angiogenesis and that CTLs and NKT cells cooperatively exert antitumor effects.
  • At least one selected from mushroom mycelium processed products is the active ingredient.
  • Specific examples include SPG (sizofiran: Kaken Pharmaceutical Co., Ltd.) (a polysaccharide obtained from the oral liquid of the culture of Suehiro mushroom mycelium), SCP (oral preparation of a processed product of Suehiro mushroom mycelium) (limited Schizophyllum Commune Fries (Science name: Schizophyllum Commune Fries), Mycelium-derived product, PARA (Krestin), etc., Straw mushroom mycelium, AH CC and L EM (Noda edible bacteria) A processed product of mycelium of Shiitake mushrooms such as Kogyo Co., Ltd.
  • Suehiro mushroom fungi SPG sizofiran: Kaken Pharmaceutical Co., Ltd.
  • a polysaccharide derived from mycelium culture mouth liquid is used as an anticancer drug for some limited cancers (Taito Co., Ltd., Kaken Pharmaceutical Co., Ltd.) .
  • PSK Shinkokin
  • Substances that have the ability to activate NKT cells by selectively acting on NKR-P1 (natural killer P1) of NK T cells can be expressed in the form of —-1,3 and / or ⁇ -1,4 darcoside bonds.
  • NKR-P1 natural killer P1
  • the substance having the activity of activating cells may be a polysaccharide having this structure and / or a substance containing 2 to 10 oligosaccharides.
  • NK T cells NKR- P l natural killer PI
  • Other substances sugars Ru substance der of alpha 1 ⁇ 3 conformation selectively acts with the ability to activate NKT cells, Nigeroori pentasaccharide (3 — ⁇ — ⁇ —D—Dalcovirano Silose D—A saccharide containing glucose as a constituent unit)
  • Nigeroori pentasaccharide 3 — ⁇ — ⁇ —D—Dalcovirano Silose D—A saccharide containing glucose as a constituent unit
  • Glucose 1 Nigerose ⁇ -D-Glcp.
  • F- fucoidan / sulfated fucan derived from gagome kelp a sugar consisting solely of fucose (Formula 2)
  • G-fucoidan / sulfated fucogalatatan from Gagome kelp sugar consisting of galactose and fucose (Formula 3)
  • oligosaccharides for example, Sales Company / Shiroko Co., Ltd .: Extract from sea squirrel laver
  • the main components are at least one selected from ⁇ 1 ⁇ 3 linked galactan sulfate oligosaccharides and ⁇ 1 ⁇ 3 bond and galactan sulfate oligosaccharides composed of
  • the compound is not limited to these, and is a saccharide substance having an ⁇ 1 ⁇ 3 steric structure (a saccharide component having a 1 ⁇ 3 glucosidic bond structure), and furthermore, NK R—P 1 (natural killer) of ⁇ ⁇ ⁇ cells
  • NK R—P 1 natural killer of ⁇ ⁇ ⁇ cells
  • the dose of these compositions is about 1 to 200 mg / Kg body weight per day, and is taken orally in 10 to 12 months, 1 to several times a day.
  • parenteral ingestion is also possible by reducing the dosage and preparing a parenteral therapeutic agent for cancer.
  • Examples of a composition containing a substance derived from a mushroom mycelium used in a new immunotherapy include a spleen mouth such as an SCP, a processed mushroom mycelium, and a spleen mushroom mycelium culture port such as an SPG.
  • a spleen mouth such as an SCP
  • a processed mushroom mycelium and a spleen mushroom mycelium culture port such as an SPG.
  • a spleen mouth such as an SCP
  • a processed mushroom mycelium a processed mushroom mycelium
  • a spleen mushroom mycelium culture port such as an SPG.
  • Contains at least two types selected from liquid-derived polysaccharides, processed shiitake mycelia such as AH CC and LEM, and perennial mushroom mycelium additives such as MAK (Noda Shokubai Kogyo Co., Ltd.) and SM. It is widely known that most of these have been used as immunostimulants for anticancer effects.
  • NITC new immunotherapy
  • these combinations and IL-1 We found a relationship between the ability to induce production and the ability of NKT cells to selectively act on NKR-P1 (natural killer P1) to activate NKT cells. It has established an advantage that is incomparable with the anticancer effect (20% effective rate) of anticancer drugs.
  • the optimal combination is a composition that combines three types of processed products of Suehi or Mushroom mycelium, or a polysaccharide derived from oral liquid of Sue or Mushroom mycelium, Shiitake mushroom mycelium, and Perennial mushroom mycelium.
  • the dosage may be adjusted depending on the amount of IL-11 induced to produce and the degree of activation of NKT cells by selective action of NKT or NKT cells on NKR-P1 (natural killer P1).
  • the administration period is 10 days to 12 months, and the frequency of administration is one to several times a day.
  • Polysaccharides obtained from mycelium cultured port solution of Sue human B mushrooms, is already SPG (si Z o ⁇ iran) and to Kaken and Taiwan Sugar Corporation and c and their preparation are commercialized, JP-B
  • the method described in JP-A No. 524-26434, JP-B No. 52-444634, etc. is exemplified.
  • mycelium of shiitake mushrooms such as L EM and reishi (manne mushrooms) mycelium such as MAK
  • the product has already been commercialized by Noda Shokubai Kogyo Co., Ltd.
  • One example of these production methods is to add rice bran to bagasse (sugar cane juice residue), mix well, adjust the water content, fill in a certain container, create a solid culture medium, and perform high-pressure steam sterilization. Inoculate the hypha of each bacterium, which has been cultivated beforehand, into the culture medium, and culture the hypha in a culturing room at 23 ° C for four months.
  • this oral preparation [SCP (Tozai Pharmaceutical Laboratory Co., Ltd.)] was used.
  • Other mushroom mycelium processed products can be prepared by the same treatment according to their solubility characteristics (water-soluble / oil-soluble).
  • Oral preparations are prepared into tablets, powders, capsules, syrups and the like.
  • the preparation can also be prepared by blending known additives such as excipients, disintegrants, binders, and lubricants, and using conventional means. If necessary, flavoring agents, coloring agents, fragrances, stabilizers, bactericides, preservatives and the like can be added.
  • the present invention focuses on angiogenesis inhibitory and angiogenesis-promoting functions in new immunotherapy (NITC), and promotes angiogenesis-promoting and angiogenesis-inhibiting factors such as VEGF, bFGF, and endostatin. Blood movement It clarifies the relationship between the condition and the condition of cancer and the therapeutic effect. That is, the present invention provides a means for testing angiogenesis, which comprises simultaneously measuring the blood levels of VEGF or bFGF and endostatin in a patient. From the values of the concentrations of VEGF or bFGF and endostatin in blood obtained by the detection means of the present invention, the effect of the substance used for treatment, for example, an angiogenesis inhibitor can be assayed.
  • NITC new immunotherapy
  • the blood of a patient is collected before and after the administration of the angiogenesis inhibitor, and the concentrations of VEGF or bFGF and endstatin in the serum are measured. If these concentrations after administration are lower than before administration, it is determined that the angiogenic ability has decreased. If the concentrations of these factors do not change or increase, it is determined that the angiogenesis inhibitor used is not effective, and it can be determined that another angiogenesis inhibitor is selected. That is, the present invention 'provides a means for assaying an angiogenesis inhibitor, in which an assay for compatibility between an angiogenesis inhibitor and a patient is performed by the above-described examination means.
  • angiogenesis-promoting factors and angiogenesis-inhibiting factors are measured in animals. For example, using a tumor-bearing mouse as described above, administering the compound thereto, collecting blood before and after administration, preparing serum, and preparing angiogenic factors and angiogenesis in the serum. The concentration of the inhibitor is measured and compounds that can reduce these concentrations after administration compared to before administration are selected. The obtained compound is a compound that can reduce the angiogenesis ability, and can be used for diseases related to angiogenesis such as solid cancer.
  • TNF tumor necrosis factor
  • ⁇ GF transforming growth factor
  • interleukin-11] 3 type III prostaglandin and the like.
  • VEGF or bFGF is measured. It is preferable to measure endostatin as an angiogenesis inhibitor.
  • the animal used for the screening may be an animal in which angiogenesis has been induced, and for example, a mouse transplanted with cancer can be suitably used.
  • the present invention focuses on angiogenesis inhibition and angiogenesis promotion functions in new immunotherapy (NITC), and is a factor relating to angiogenesis promotion and angiogenesis inhibition.
  • NITC new immunotherapy
  • the present invention provides a mechanism for implementing the new immunotherapy (NITC) with the above-mentioned examination means and / or the above-mentioned examination means, a mechanism for judging the test results, and a mechanism for performing a therapeutic procedure based on the judgment.
  • NITC new immunotherapy
  • a system to provide cancer immunotherapy in cooperation with an immune function testing organization, an immune function evaluation organization, a treatment and / or prescription organization will be formed.
  • an immune function testing organization for example, when blood of a cancer patient is transferred to an immune function testing laboratory after collection, at least VEGF or bFGF and endostatin are collected at the testing laboratory. Are measured, and the obtained measurement results are sent to the affiliated immune function evaluation organization and evaluated based on the test results. Further, it is preferable to conduct a test for immune functions such as IL-112, NKT activity, IFN ⁇ and the like.
  • therapeutic agents and treatment methods that are compatible with the blood kinetics, immune function, and stage of cancer progression of factors related to angiogenesis of the cancer patient are selected from the above-mentioned respective prescriptions, and the information is used as the treatment and / or prescription. It is transmitted to an institution (medical institution), and performs cancer immunotherapy based on the notification. Such a new cancer immunotherapy system is extremely useful. In addition, the cancer immunotherapy using each of the above-mentioned prescriptions also has extremely high utility.
  • a shark cartilage preparation (Patent No. 3135131), Bettashark MC (Seishin K.K.), was orally ingested 20 g / day.
  • patients with Th2 were ingested PSK (Sankyo) 3.Og / day, and combined with IL-X. For ingestion. The number of patients was 29, and the results are shown in Table 2 below.
  • Period treatment The method for measuring each factor shown in the table is shown below.
  • the enzyme-linked immunosorbent assay (ELISA) of commercially available kits was used for the measurement (ACCUCYTE Human VEGF, ACCUCYTE Human bFGF, ACCUCYTE Human Endostatin: CYTIMMUNE Sciences Inc.). (Measurement of NK T cells)
  • the measurement of the activation by the action on NKR-1 is based on the determination of the cell surface antigens (CD3 and CD161) which are specifically present on the cell surface of the ⁇ KT ⁇ cells.
  • the measurement can be performed by examining the increase in the number of cells. Specifically, lymphocytes in peripheral blood are assayed for CD3-positive cells and CD166-positive cells. That is, CD3 and CD161, which are cell surface antigens of ⁇ ⁇ cells, are measured by Two Co1 or test using flow cytometry using a monoclonal antibody. I do.
  • “Activated NKT cells” means that the percentage of NKT cells in lymphocytes is 10% or more.
  • the NKT cell activating ability means a function of increasing the ratio of NKT cells to 10% or more, or a function of further increasing the ratio of NKT cells before administration of a certain substance.
  • the percentage of cells that are positive for the cell surface antigens CD3 and CD161 in blood cells was determined by Two Co1 or flow cytometry. It was measured as usual.
  • CD 3 and C As the monoclonal antibody against D161, CD3-PC5 manufactured by Coulter and CD161 manufactured by Becton Dickinson were used, respectively.
  • Va24 and V ⁇ 1 which are cell surface antigens of NKT cells, are conjugated to monoclonal antibodies (TCR-VQ: 24PE, TCR-Vi311 FITC; Beckman Cou lter). was measured by Two Co1 or inspection using flow cytometry. (Preparation of sample for measuring site force-in)
  • a mononuclear cell fraction is separated and prepared from the blood of a cancer patient.
  • Heparin-added peripheral blood obtained from a cancer patient was diluted 2-fold with phosphate buffered saline (PBS) and mixed, and then F £: 011—.
  • the mononuclear cell fraction was collected after layering on 0 & 1 & 7 solution (specific gravity 1.077), centrifuging at 400 G for 20 minutes. After washing, RPMI-164 medium supplemented with 10% fetal bovine serum (FBS) was added to adjust the cell number to 1 ⁇ 10 6.
  • PBS phosphate buffered saline
  • IL-12 was measured by the ELISA method using a kit made by R & D SYSTEMS.
  • 50 ⁇ l of the assay diluent Assay Diluent RD 1F, 200 ⁇ l of the standard solution or the sample prepared in Example 1 was placed in each well of the 96-well microplate. After each dispensing, the mixture was allowed to stand at room temperature and reacted for 2 hours. Thereafter, horseradishpelexidase (hereinafter, abbreviated as HRP) -labeled anti-IL-11 antibody was dispensed in a quantity of 200 ⁇ l each, and allowed to stand at room temperature for 2 hours.
  • HRP horseradishpelexidase
  • the chromogenic substrate solution was dispensed one by one, and allowed to stand at room temperature for 20 minutes, and then the enzyme reaction stop solution was dispensed in 50 ⁇ l portions.
  • the absorbance of each well at 450 nm was measured by Etn ax (Wako Pure Chemical Industries, Ltd.).
  • the IL-l-l2 amount is expressed as pg Zml.
  • the ability to induce IL-11 production is a function that enhances the amount of IL-11 produced by peripheral blood mononuclear cells upon stimulation to 7.8 pg Zml or more (7.8 is the measurement limit value), Alternatively, it means a function of enhancing the production amount of IL-112 before administering a certain substance.
  • IF N y The measurement of IF N y is based on the Io of Bio SouracEuroeS.
  • IL-10 was measured by a solid-phase enzyme immunoassay (ELISA method) using an IL-10 EASIA kit from BioSourceEuropeS. The method was performed according to the method for measuring IF ⁇ except that an anti-IL 110 antibody was used. The IL 110 amount is expressed as pg / ml.
  • Th1 / Th2 cell ratio was assayed by a routine method by a helper T (Th) cell line Threcco1or analysis by flow cytometry.
  • Th1 / Th2 is the ratio of helper T cells that have the cell surface antigen CD4 that produce IFN ⁇ (Th1) to cells that produce IL-4 (Th2). And expressed as CD 4 XIFNT // IL—4.
  • the blood of a cancer patient was treated with phorbol 1 2 —myristate 13 —acetate (phorbol 12 2 — myristate 13 A aceetate) and ionomycin (Ionomycin) for 37 hours for 4 hours.
  • Cells in the blood were stimulated to produce cytokines.
  • the production reaction was stopped by adding preferdin A (B referdin A), and CD4, an anti-CD4 antibody, was used as a cell surface marker, using CD5-PC5 (Beckman Couter). After staining the cells and fixing the cells, FACSL ysing Solution (Nippon Vector (Kinson).
  • FACSP ermeabi 1 izing Solution Naippon Vector Dickinson Company
  • FASTI MMU NEIFN y FITC / IL-4PE Intracellular cytokins were stained with Betaton Dickinson (Japan) and measured and analyzed using a flow cytometer (FACSC alibur, Becton Diekinson).
  • Immune function testing institutions medical institutions, and immunological competence testing and treatment policy guidance agencies.
  • a medical institution collects blood from a patient by a known collection method corresponding to a target test measurement item in the blood component.
  • the medical institution separates serum and blood cell components from the blood, if necessary, depending on the intended test measurement items.
  • the medical institution will identify the blood, serum, and / or blood cell components and the patients from which they are derived, and immediately send them to the affiliated immunological testing laboratory.
  • Immune function testing institutes conduct tests for specific immune-related functions determined by the guidance of an institution that conducts immunological ability tests and guidance on treatment policies (hereinafter referred to as “instruction bodies”) (for example, IL-11 production ability, NKT cell activity).
  • IFN y value, TNF a value, etc. are performed on blood, serum, and / or blood cell components sent from medical institutions. If necessary, serum and blood cell components are separated from the blood.
  • the immunological function testing organization conducts tests for cancer markers and tests for cancer-related factors (for example, angiogenesis promoting factors and angiogenesis inhibitory factors) if necessary. Inform the instructor of the test results I do. Based on the notified test results, the instructor should at least assay the subject's IL-12 production ability, NKT cell activity, IFNy level, TNFa level, and promote angiogenesis factors (such as VEGF and bFGF), angiogenesis inhibitors (eg endostatin), and various cancer markers as needed.
  • angiogenesis factors such as VEGF and bFGF
  • angiogenesis inhibitors eg endostatin
  • the instructing institution shall apply the assay results based on the prescription examples of IL-12 inducer, NKT cell activator, other BRM preparations, shark cartilage preparations, etc. To identify.
  • the test results and treatment examples are immediately transmitted to the affiliated medical institutions.
  • the medical institution treats the target patient with cancer by referring to the specified prescription example.
  • the guidance organization determines the effectiveness and ineffectiveness of the treatment by examining the results of various tests performed on the patient's blood components in the same manner as above during the course of treatment, and determines whether the treatment is effective or ineffective.
  • Guidance agencies collect guidance fees by presenting the test results and prescription examples to medical institutions.
  • VEGF vascular fibroblast growth factor
  • VEGF basic fibroblast growth factor
  • endostatin endostatin

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Immunology (AREA)
  • Engineering & Computer Science (AREA)
  • Urology & Nephrology (AREA)
  • Chemical & Material Sciences (AREA)
  • Biomedical Technology (AREA)
  • Molecular Biology (AREA)
  • Hematology (AREA)
  • Medicinal Chemistry (AREA)
  • Analytical Chemistry (AREA)
  • Biotechnology (AREA)
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  • Tropical Medicine & Parasitology (AREA)
  • Food Science & Technology (AREA)
  • Microbiology (AREA)
  • Physics & Mathematics (AREA)
  • Cell Biology (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Medicines Containing Material From Animals Or Micro-Organisms (AREA)

Abstract

L'invention concerne un moyen pour examiner l'aptitude d'un patient à l'angiogénèse, comprenant la mesure simultanée des concentrations du facteur de croissance de l'endothélium vasculaire (VEGF) ou du facteur de croissance fibroblastique basique (bFGF) et de l'endostatine dans le sang du patient.
PCT/JP2001/008098 2000-10-11 2001-09-18 Moyen pour examiner l'aptitude à l'angiogénèse WO2002031497A1 (fr)

Priority Applications (2)

Application Number Priority Date Filing Date Title
JP2002534831A JPWO2002031497A1 (ja) 2000-10-11 2001-09-18 血管新生能の検査手段
AU2001288058A AU2001288058A1 (en) 2000-10-11 2001-09-18 Means of examining ability of angiogenesis

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
JP2000310349 2000-10-11
JP2000-310349 2000-10-11
JP2001067473 2001-03-09
JP2001-67473 2001-03-09

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WO2002031497A1 true WO2002031497A1 (fr) 2002-04-18

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PCT/JP2001/008098 WO2002031497A1 (fr) 2000-10-11 2001-09-18 Moyen pour examiner l'aptitude à l'angiogénèse

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JP (1) JPWO2002031497A1 (fr)
AU (1) AU2001288058A1 (fr)
WO (1) WO2002031497A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2009512860A (ja) * 2005-10-21 2009-03-26 バイエル ヘルスケア エルエルシー がんの予測及び予後の検査方法、並びにがん治療のモニタリング

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH10123131A (ja) * 1996-10-16 1998-05-15 Toagosei Co Ltd 予後検査方法
WO1998022616A1 (fr) * 1996-11-21 1998-05-28 Kyowa Hakko Kogyo Co., Ltd. Anticorps monoclonal avec recepteur f1t-1 vegf anti-humain
JPH11201966A (ja) * 1998-01-12 1999-07-30 Toagosei Co Ltd Ｖｅｇｆ／ｖｐｆ拮抗阻害剤スクリーニング方法及びスクリーニング用キット

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH10123131A (ja) * 1996-10-16 1998-05-15 Toagosei Co Ltd 予後検査方法
WO1998022616A1 (fr) * 1996-11-21 1998-05-28 Kyowa Hakko Kogyo Co., Ltd. Anticorps monoclonal avec recepteur f1t-1 vegf anti-humain
JPH11201966A (ja) * 1998-01-12 1999-07-30 Toagosei Co Ltd Ｖｅｇｆ／ｖｐｆ拮抗阻害剤スクリーニング方法及びスクリーニング用キット

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
FELDMAN ET AL.: "Correlation between serum endostatin and vascular endothelial growth factor(VEGP) levels in patents with renal cancer", PROCEEDINGS OF THE ANNUAL MEETING -AMERICAN ASSOCIATION FOR CANCER RESEARCH, vol. 41, March 2000 (2000-03-01), pages 272, ABSTRACT 1734, XP002949795 *
YOKOMIZO ET AL.: "Jin gan to zenritsusen gan ni okeru kekkan shinsei", MOLECULAR MEDICINE, vol. 37, no. 3, February 2000 (2000-02-01), pages 330 - 337, XP002949796 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2009512860A (ja) * 2005-10-21 2009-03-26 バイエル ヘルスケア エルエルシー がんの予測及び予後の検査方法、並びにがん治療のモニタリング

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JPWO2002031497A1 (ja) 2004-02-19

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