[go: up one dir, main page]

WO2023082220A1 - Sensibilisateur de cellules nk préparé au moyen d'une combinaison de nano-sélénium fonctionnel avec de la metformine et utilisation associée - Google Patents

Sensibilisateur de cellules nk préparé au moyen d'une combinaison de nano-sélénium fonctionnel avec de la metformine et utilisation associée Download PDF

Info

Publication number
WO2023082220A1
WO2023082220A1 PCT/CN2021/130493 CN2021130493W WO2023082220A1 WO 2023082220 A1 WO2023082220 A1 WO 2023082220A1 CN 2021130493 W CN2021130493 W CN 2021130493W WO 2023082220 A1 WO2023082220 A1 WO 2023082220A1
Authority
WO
WIPO (PCT)
Prior art keywords
selenium
cells
nano
metformin
seo
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/CN2021/130493
Other languages
English (en)
Chinese (zh)
Inventor
李晓玲
陈填烽
刘婷
陈义康
李海伟
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Guangdong Jinan Selenium Source Nanotechnology Research Institute Co Ltd
Original Assignee
Guangdong Jinan Selenium Source Nanotechnology Research Institute Co Ltd
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Guangdong Jinan Selenium Source Nanotechnology Research Institute Co Ltd filed Critical Guangdong Jinan Selenium Source Nanotechnology Research Institute Co Ltd
Priority to PCT/CN2021/130493 priority Critical patent/WO2023082220A1/fr
Publication of WO2023082220A1 publication Critical patent/WO2023082220A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Images

Classifications

    • 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
    • A61K31/13—Amines
    • A61K31/155—Amidines (), e.g. guanidine (H2N—C(=NH)—NH2), isourea (N=C(OH)—NH2), isothiourea (—N=C(SH)—NH2)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K33/00—Medicinal preparations containing inorganic active ingredients
    • A61K33/04—Sulfur, selenium or tellurium; Compounds thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
    • A61K47/58—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained by reactions only involving carbon-to-carbon unsaturated bonds, e.g. poly[meth]acrylate, polyacrylamide, polystyrene, polyvinylpyrrolidone, polyvinylalcohol or polystyrene sulfonic acid resin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
    • A61K47/61—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule the organic macromolecular compound being a polysaccharide or a derivative thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
    • A61K47/66—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid the modifying agent being a pre-targeting system involving a peptide or protein for targeting specific cells

Definitions

  • the invention belongs to the technical field of medical compounds, and in particular relates to an NK cell sensitizer prepared by functionalized nano-selenium combined with metformin and its application in anti-tumor.
  • Liver cancer is the sixth most common cancer in the world, accounting for 11% of cancer mortality.
  • Diabetes is a metabolic disease characterized by dysregulation of blood sugar and insulin.
  • Many studies have found that the longer the onset duration of diabetes, the greater the risk of liver cancer, which is 2 to 3 times higher than that of non-diabetic patients.
  • the possible carcinogenic mechanisms are the patient's own hyperglycemia, chronic inflammation, obesity and hyperinsulinemia.
  • Hyperglycemia can cause oxidative stress through glycosylation of hemoglobin to release ferrous ions (as a pro-oxidant), generating free radicals; autologous chronic inflammation induces tumor necrosis factor (TNF- ⁇ ) and interleukin-6 (IL-6) expression; obesity reduces the sensitivity of the human body to insulin.
  • TNF- ⁇ tumor necrosis factor
  • IL-6 interleukin-6
  • pancreatic ⁇ cells In order to maintain a normal insulin level, the pancreatic ⁇ cells will secrete more insulin, causing hyperinsulinemia, and further affecting liver cell damage and inflammation. , expression of fibrosis-associated matrix proteins; these complex conditions reduce immune system function and greatly increase the incidence of liver cancer.
  • Adoptive immune cell therapy uses living immunocompetent cells to expand in vitro and reintroduce them back into the patient's body. Such immunocompetent cells can directly or indirectly kill tumor cells.
  • Adoptive immune cells combined with chemotherapy drugs are used in the treatment of tumors, which can enhance the immune sensitivity of immune cells, increase the expression of immune cell sensitive receptors, and enhance the killing effect of immune cells on tumor cells.
  • Natural killer (NK) cells are important immune cells in the body. They are not only related to anti-tumor, anti-viral infection and immune regulation, but also participate in the occurrence of hypersensitivity and autoimmune diseases in some cases. They can recognize target cells, kill medium.
  • NK-92 cells are a kind of immune cells similar to NK cell activity. They have the main characteristics of NK cells and the main expression receptors. TNF- ⁇ , etc., and then kill tumor cells; and NK-92 cells can efficiently recognize and kill tumor cells without affecting the survival of normal cells. This special mechanism is not clear, but it is speculated that NK-92 cells do not express It is caused by immunoglobulin-like receptors (KIR) on NK cells.
  • KIR immunoglobulin-like receptors
  • cabozantinib can cooperate with NK-92 cells to cause a greater killing effect on kidney cancer, in which cabozantinib increases the expression of EGFR in tumor cells and reduces the expression of PD-L1 in tumor cells, reversing the immune response. inhibition. Therefore, it is a good treatment strategy to combine chemotherapy drugs with immune cells to enhance the immunotherapy effect on tumor cells.
  • SeNPs have the functions of anti-oxidation and scavenging free radicals in normal cells, and can reduce the oxidative stress caused by hyperglycemia; in terms of anti-tumor, they have the effect of killing tumors and can enhance the killing of immune cells tumor cells.
  • Metformin Metalformin
  • T2DM can lower blood sugar and has a certain anti-tumor effect. Therefore, how to compound metformin to enhance the immunotherapy effect of NK cells, in the new application field of compound combined with nano-selenium to improve the treatment of tumors, and to find nano-immunotherapy sensitizers for immunotherapy is a tumor treatment that needs further research.
  • the primary purpose of the present invention is to provide an application scheme of functionalized nano-selenium combined with metformin (Metformin) as an NK cell sensitizer, specifically functionalized nano-selenium combined with metformin as an NK cell sensitizer to assist NK cells to enhance the tumor cell sensitizer.
  • metformin metformin
  • Another object of the present invention is to provide a method for preparing an NK cell sensitizer, and to obtain an NK cell sensitizer prepared by combining functionalized nano-selenium with metformin.
  • the NK cell sensitizer may be an NK-92 cell sensitizer.
  • Another object of the present invention is to provide an antitumor drug.
  • the present inventors found that functionalized SeNPs combined with metformin has a synergistic anti-tumor effect, and further studied the immunotherapy effect of functionalized SeNPs combined with metformin to enhance NK cells, such as NK-92 cells, and preliminarily explored its mechanism of action through experiments.
  • the invention provides an application scheme of functionalized nano-selenium combined with metformin as an NK cell sensitizer.
  • the application of the functionalized nano-selenium combined with metformin as an NK cell sensitizer in anti-tumor can be that the functionalized nano-selenium combined with metformin is pre-incubated with NK cells for a period of time before treating tumor cells; or the functionalized nano-selenium combined with metformin is pretreated with tumor cells and then added to NK cells; or the functionalized nano-selenium Combining metformin and NK cells to treat tumor cells at the same time.
  • the tumors to be treated include any one of human liver cancer, human non-small cell lung cancer, human cervical cancer, melanoma, breast cancer and glioma.
  • the invention also provides a method for preparing an NK cell sensitizer by combining functionalized nano-selenium with metformin, comprising the steps of preparing the functionalized nano-selenium; adding metformin for mixed reaction to obtain the NK cell sensitizer.
  • the functionalized nano-selenium is nano-selenium (S-SeNPs), polyallylamine hydrochloride (Aladdin) modified nano-selenium (PAH-SeNPs), polyvinylpyrrolidone-modified nano-selenium (PVP-SeNPs ), TW80-modified nano-selenium (TW80-SeNPs), polysaccharide-modified nano-selenium, folic acid-modified nano-selenium (folate-modified tumor-targeting nano-selenium), transferrin-modified nano-selenium (transferrin-modified tumor-targeting Targeted nano-selenium) at least one.
  • S-SeNPs nano-selenium
  • Align polyallylamine hydrochloride
  • PVP-SeNPs polyvinylpyrrolidone-modified nano-selenium
  • TW80-SeNPs TW80
  • the preparation of functionalized nano-selenium is prepared by any of the following methods:
  • the concentration of the H 2 SeO 3 solution or the Na 2 SeO 3 solution is 8-10 mmol/L; the amount of the H 2 SeO 3 solution or the Na 2 SeO 3 solution is calculated based on the final concentration of 1-5 mmol/L;
  • the amount of substance A added is calculated based on the final concentration of 0.5-10 mg/ml;
  • Vc and H 2 SeO 3 are mixed in a molar ratio of 3 to 10:1; or the Vc and Na 2 SeO 3 are mixed in a molar ratio of 3 to 10:1; the reaction temperature is 4 °C ⁇ 5°C, the time is 0.5 ⁇ 24 hours;
  • the dialysis method is to use a dialysis bag for dialysis, and the dialysis time is 12 to 48 hours.
  • the present invention also provides an NK cell sensitizer, which is an NK cell sensitizer obtained by the above method, based on functionalized nano-selenium combined with metformin as an NK cell sensitizer, which can be composed of nano-selenium and a modifier allowed by the drug, And made into various forms such as injections.
  • anti-tumor drugs comprising the NK cell sensitizer obtained by the above method, and NK cells.
  • anti-tumor drugs can be composed of functionalized nano-selenium, metformin and NK cells; or as another embodiment, anti-tumor drugs can be composed of functionalized nano-selenium, metformin, NK cells and other existing anti-tumor drugs Compound composition.
  • NK-92 cells natural killer cells of human malignant non-Hodgkin's lymphoma patients
  • NK cells obtained by conventional methods such as purchased through commercial channels, or obtained by using human peripheral Blood, bone marrow or umbilical cord blood are obtained by separating mononuclear cells and inducing culture with IL-2 and IL-15 in vitro.
  • the tumors that the antitumor drugs are used to treat include any one of human liver cancer, human non-small cell lung cancer, human cervical cancer, melanoma, breast cancer and brain glioma. Preferred is the treatment of human liver cancer.
  • the functionalized nano-selenium and metformin in the present invention can effectively inhibit the proliferation of liver cancer cells HepG2 through NK sensitization, that is, they can be used to treat liver cancer.
  • the present invention has the following advantages and effects:
  • the present invention discloses the application of functionalized nano-selenium combined with metformin as a sensitizer for NK cell immunotherapy, which may be that the functionalized nano-selenium combined with metformin is incubated with NK cells for a period of time in advance to treat tumor cells; or the described The functionalized nano-selenium combined with metformin pretreats the tumor cells and then adds NK cells; or the functionalized nano-selenium combined with metformin and NK cells simultaneously treats the tumor cells.
  • the modified tumor-targeting nano-selenium has obvious NK cell sensitization characteristics, and has a high-efficiency inhibitory effect on liver cancer cell HepG2 in vitro cell experiments, and can also significantly inhibit human non-small cell lung cancer cells A549, human cervical Proliferation of cancer cell HeLa, melanoma cell A375, breast cancer cell MCF-7 and brain glioma cell C6. Its mechanism is mainly through regulating the production of NKG2D ligand on HepG2 cells, thereby inducing cell cycle arrest and tumor cell apoptosis.
  • the test results suggest that functionalized nano-selenium combined with metformin can be developed as a new immunotherapy sensitizer, which can significantly reduce the survival rate of tumor cells.
  • the method for preparing the NK cell sensitizer of the present invention is simple in operation and strong in applicability, and vigorously developing the treatment of nano-selenium combined with metformin to sensitize NK cells has broad application prospects.
  • the present invention has developed an antitumor drug with high efficiency and low toxicity, including the NK cell sensitizer and NK cells obtained by the above method, and functionalized nano-selenium combined with metformin as a sensitizer for NK cell immunotherapy, is
  • the antitumor drug with new ingredients has certain curative effect.
  • the present invention uses vitamin C to reduce sodium selenite to prepare functionalized nano-selenium.
  • Functionalized nano-selenium combined with metformin can indirectly kill tumor cells by regulating the reactivity of the body's immune system, thereby greatly inhibiting the proliferation of human liver cancer cells HepG2.
  • SeNPs, metformin, and NK cells in the control group did not have this significant characteristic.
  • TW80-SeNPs combined with metformin can significantly improve the killing effect of NK-92 cells on HepG2 cells.
  • the modified tumor-targeting nano-selenium and metformin synergistically stimulate HepG2 cells to recruit more NK cells and make tumor cells more sensitive to NK cells, thereby killing more tumor cells.
  • the test results suggest that functionalized nano-selenium combined with metformin can be used as an effective new sensitizer to improve tumors, for example, it has a good therapeutic effect on liver cancer.
  • the raw materials of the nano-selenium sensitizer obtained in the present invention are cheap and easy to obtain, the synthesis and purification steps are highly operable, and the preparation steps are simple.
  • the synthesis scale can be appropriately expanded by optimizing the process to realize the commercialization and application of the drug.
  • Figure 1 is the evaluation of the cell viability of functionalized SeNPs combined with metformin on HepG-2 cells.
  • FIG. 1-1A is the evaluation of functionalized SeNPs on the survival rate of liver cancer HepG-2 cells
  • Fig. 1-1B is the evaluation of metformin on the survival rate of liver cancer HepG-2 cells
  • Fig. 1-1C is the evaluation chart of the cell viability of functionalized SeNPs combined with metformin on HepG-2.
  • Figure 2 is the evaluation of the cell viability of HepG-2 cells in combination with NK-92 cells.
  • FIG. 2-1 is the evaluation of the survival rate of NK-92 cells by functionalized SeNPs and metformin
  • Figure 2-2A and Figure 2-2B is the effect of functionalized SeNPs and metformin combined with NK-92 cells on HepG-2 cells Survival assessment.
  • FIG. 3-1 ( Figure 3-2) is the survival rate of functionalized SeNPs combined with metformin and NK-92 cells on HepG-2 cells;
  • Figure 3-3) is the effect of polymer PAH, PVP, TW80 on HepG -2 Cell viability assessment.
  • Figure 4 is the evaluation of the cell viability of different forms of selenium including (A) Na 2 SeO 3 , (B) TW80-SeNPs and (C) selenocystine SeC combined with metformin and NK-92 on HepG-2 cells.
  • Figure 5 shows the enhancement factor (Figure 5A) of TW80-SeNPs combined with metformin and live NK cells on HepG-2 cells and (Figure 5B) the cell survival rates of Case 2 and Case 6.
  • Figure 6 shows the expression of receptor signals on the surface of HepG-2 cells by TW80-SeNPs combined with metformin.
  • Figure 6A the expression of ULBP-1/2/3/4;
  • Figure 6B the expression of PD-L1 and MICA;
  • Figure 6C the fluorescence value of each receptor.
  • Figure 7 shows the expression level of ROS in HepG-2 cells by TW80-SeNPs combined with metformin.
  • Figure 8 is the evaluation of the survival rate of SSB nanoemulsion combined with NK cells on MBA-MD-231 cells.
  • the invention provides functionalized nano-selenium combined with metformin as an NK cell sensitizer.
  • the tumor includes any one of human liver cancer, human non-small cell lung cancer, human cervical cancer, melanoma, breast cancer and glioma, and is especially effective for treating liver cancer.
  • the present invention also provides a method for preparing an NK cell sensitizer by combining functionalized nano-selenium with metformin, comprising steps
  • the first implementation of functionalized nano-selenium it is to mix H 2 SeO 3 solution or Na 2 SeO 3 solution and Vc evenly, react at a temperature of 4° C. for 0.5 to 24 hours, and dialyze to obtain nano-selenium (S -SeNPs).
  • concentration of the H 2 SeO 3 solution or Na 2 SeO 3 solution is 10mmol/L; the addition amount of the H 2 SeO 3 solution or Na 2 SeO 3 solution is calculated based on the final concentration of 1-5mmol/L.
  • the Vc and H 2 SeO 3 are mixed in a molar ratio of 3-10:1; or the Vc and Na 2 SeO 3 are mixed in a molar ratio of 3-10:1.
  • the dialysis method is to use a dialysis bag for dialysis, and the dialysis time is 12 to 48 hours.
  • H 2 SeO 3 solution or Na 2 SeO 3 solution and substance A are uniformly mixed, and Vc is added to react at a temperature of 4°C-5°C for 0.5-24 hours, Dialysis to obtain different functionalized nano-selenium.
  • the substance A is any one of PAH, PVP, TW80, polysaccharide, folic acid, and transferrin.
  • the concentration of the H 2 SeO 3 solution or the Na 2 SeO 3 solution is 8-10 mmol/L; the amount of the H 2 SeO 3 solution or the Na 2 SeO 3 solution is calculated based on the final concentration of 1-5 mmol/L; The amount of substance A added is calculated according to the final concentration of 0.5-10 mg/ml; the Vc and H 2 SeO 3 are mixed in a molar ratio of 3-10:1; or the Vc and Na 2 SeO 3 are molar The ratio is 3 to 10:1 for proportioning.
  • the dialysis method is to use a dialysis bag for dialysis, and the dialysis time is 12 to 48 hours.
  • the third embodiment of preparing functionalized nano-selenium mix substance A and Vc evenly, then add H 2 SeO 3 solution or Na 2 SeO 3 solution at a temperature of 4°C to 5°C for 0.5 to 24 hours, and dialyze , to obtain different functionalized nanoselenium.
  • the substance A is any one of PAH, PVP, TW80, folic acid and transferrin.
  • the concentration of the H 2 SeO 3 solution or the Na 2 SeO 3 solution is 8-10 mmol/L; the amount of the H 2 SeO 3 solution or the Na 2 SeO 3 solution is calculated based on the final concentration of 1-5 mmol/L;
  • the amount of substance A added is calculated according to the final concentration of 0.5-10 mg/ml.
  • the Vc and H 2 SeO 3 are mixed in a molar ratio of 3-10:1; or the Vc and Na 2 SeO 3 are mixed in a molar ratio of 3-10:1.
  • the dialysis method is to use a dialysis bag for dialysis, and the dialysis time is 12 to 48 hours.
  • the functionalized nano-selenium prepared by the above step (1) is nano-selenium (S-SeNPs), polyallylamine hydrochloride (Aladdin) modified nano-selenium (PAH-SeNPs), polyvinylpyrrolidone modified nano-selenium ( PVP-SeNPs), TW80-modified nano-selenium (TW80-SeNPs), polysaccharide-modified nano-selenium, folic acid-modified nano-selenium (folate-modified tumor-targeting nano-selenium), transferrin-modified nano-selenium (transferrin At least one of modified tumor-targeting nano-selenium).
  • S-SeNPs nano-selenium
  • Align polyallylamine hydrochloride
  • PVP-SeNPs polyvinylpyrrolidone modified nano-selenium
  • TW80-SeNPs TW80-modified nano
  • the present invention also provides an NK cell sensitizer, which is an NK cell sensitizer obtained by the above method, based on functionalized nano-selenium combined with metformin as an NK cell sensitizer, which can be composed of nano-selenium and a modifier allowed by the drug, And made into various forms such as injections.
  • the present invention also provides an antitumor drug, comprising the NK cell sensitizer obtained by the above method, and NK cells.
  • the anti-tumor drug can be composed of functionalized nano-selenium, metformin and NK cells; or as another embodiment, the anti-tumor drug can be composed of functionalized nano-selenium, metformin, NK cells and other existing anti-cancer drugs Composition of tumor compounds.
  • NK-92 cells natural killer cells of human malignant non-Hodgkin's lymphoma patients
  • NK cells obtained by conventional methods such as purchased through commercial channels, or obtained by using human peripheral Blood, bone marrow or umbilical cord blood are obtained by separating mononuclear cells and inducing culture with IL-2 and IL-15 in vitro.
  • the tumors that the antitumor drugs are used to treat include any one of human liver cancer, human non-small cell lung cancer, human cervical cancer, melanoma, breast cancer and brain glioma. Preferred is the treatment of human liver cancer.
  • Example 1 Functionalized SeNPs combined with metformin enhances the immunotherapy effect of NK-92 cells.
  • HepG2 cells and NK-92 cells were purchased from the American Standard Culture Collection (ATCC), HepG2 was cultured with complete medium (DMEM, Gibco), and NK-92 cells were cultured with minimum essential medium (MEM, Gibco) culture, and add 10% (v/v) fetal bovine serum (FBS, Gibco) and 1% (v/v) double antibody (penicillin and streptomycin mixed solution, Gibco), cultured at 37°C in a humidified incubator with 5% (v/v) carbon dioxide.
  • ATCC American Standard Culture Collection
  • DMEM complete medium
  • MEM minimum essential medium
  • FBS fetal bovine serum
  • FBS fetal bovine serum
  • v/v double antibody
  • Example 2 the in vitro anti-hepatic tumor activity of different forms of selenium (Na 2 SeO 3 , TW80-SeNPs and SeC) combined with metformin and NK-92 cells.
  • NK cells were cultured (20000/mL) for 24 hours, NK cells, metformin and nano-selenium were not added as blank control, and NK cells (5 times the number of HepG2 cells were added) and metformin at a final concentration of 5 mM were added to the experimental group.
  • NK cells 5 times the number of HepG2 cells were added
  • metformin at a final concentration of 5 mM were added to the experimental group.
  • the final concentration of 5 mM metformin combined with 2.5 ⁇ M of different forms of selenium (Na 2 SeO 3 , TW80-SeNPs and SeC) to treat HepG2 first
  • NK-92 cells were added to treat HepG2 cells for 24 hours.
  • TW80-SeNPs combined with metformin enhanced the anti-tumor activity of patient-derived NK cells.
  • Drug combination can enhance the immunotherapy effect of NK-92 cells. Then, whether the combination of drugs can enhance the therapeutic effect of NK cells from patients. Therefore, we tested the immunotherapeutic effect of TW80-SeNPs combined with metformin on NK cells from patients in 6 cases. As shown in the experimental results shown in Figure 5A, it can be seen that in 6 patients with NK, TW80-SeNPs combined with metformin enhanced the immunotherapy effect of NK cells in patients by 0.8-10 times, and there were some differences in the enhancement times for different living NK cells.
  • the enhancement factor of case 1 was between 0.8 and 1.9 times; the enhancement factor of case 2 was between 1.1 and 1.5 times; the enhancement factor of case 3 was between 0.8 and 5.2 times; the enhancement factor of case 4 was between 2.8 and 4.6 times The enhancement factor of case 5 was between 1.1 and 1.5 times; the enhancement factor of case 6 was between 4.4 and 10.6 times. From Figure 5B in the cell survival rate of case 2 and case 6, it can be seen that the cell survival rate of TW80-SeNPs (5 ⁇ M)+metformin+NK group was 32% and 19%, respectively, compared to the NK group alone (65% and 95% %) and TW80-SeNPs (5 ⁇ M)+metformin group (63% and 51%) had significant differences. Therefore, TW80-SeNPs combined with metformin can significantly enhance the immunotherapeutic effect of NK cells from patients.
  • Example 4 regulation of TW80-SeNPs combined with metformin on tumor cell surface receptor signal expression.
  • Soluble MHC class I molecule-associated protein (MICA/B), ULBPs family proteins (ULBP-1/2/3/4) are functional ligands for tumor cells to bind to NK-92 cell surface signaling receptors (NKG2D), which The increase in the expression level can affect the recognition of tumor cells by NK-92 cells, thereby affecting the immunotherapy effect of NK-92 cells.
  • the programmed death ligand (PD-L1) in tumor cells can bind to the programmed death receptor 1 (PD-1) on the surface of immune cells to enhance the immune escape of tumor cells, so inhibiting their binding can enhance the immune cell function. The effect of immunotherapy.
  • TW80-SeNPs combined with metformin induced an increase in ROS levels in HepG-2 cells.
  • Example 1 Compared with Example 1, the nanoemulsion (SSB) loaded with SeC and TGF- ⁇ inhibitor enhances the anti-breast cancer effect of NK cells.
  • SSB nanoemulsion loaded with SeC and TGF- ⁇ inhibitor enhances the anti-breast cancer effect of NK cells.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Public Health (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Epidemiology (AREA)
  • Medicinal Chemistry (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Engineering & Computer Science (AREA)
  • Inorganic Chemistry (AREA)
  • Cell Biology (AREA)
  • Molecular Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Medicines Containing Material From Animals Or Micro-Organisms (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Abstract

La présente invention concerne un sensibilisateur de cellules NK préparé au moyen d'une combinaison d'un nano-sélénium fonctionnel avec de la metformine, le nano-sélénium fonctionnel étant au moins l'un parmi le nano-sélénium unique, le nano-sélénium modifié avec du chlorhydrate de polyacrylamide, le nano-sélénium modifié avec de la polyvinylpyrrolidone, le nano-sélénium modifié avec du Tween 80, le nano-sélénium modifié avec un polysaccharide, le nano-sélénium de ciblage des tumeurs modifié avec de l'acide folique et le nano-sélénium de ciblage des tumeurs modifié avec de la transferrine, et les cellules NK qui en font partie sont des cellules NK92. L'invention concerne également un sensibilisateur de cellules NK préparé au moyen d'une combinaison d'un nano-sélénium fonctionnel avec de la metformine, le nano-sélénium fonctionnel étant au moins l'un parmi le nano-sélénium unique, le nano-sélénium modifié avec du chlorhydrate de polyacrylamide, le nano-sélénium modifié avec de la polyvinylpyrrolidone, le nano-sélénium modifié avec du Tween 80, le nano-sélénium modifié par un polysaccharide, un nano-sélénium de ciblage des tumeurs modifié avec de l'acide folique et un nano-sélénium de ciblage des tumeurs modifié avec de la transferrine, et les cellules NK qui en font partie sont des cellules NK92.
PCT/CN2021/130493 2021-11-13 2021-11-13 Sensibilisateur de cellules nk préparé au moyen d'une combinaison de nano-sélénium fonctionnel avec de la metformine et utilisation associée Ceased WO2023082220A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
PCT/CN2021/130493 WO2023082220A1 (fr) 2021-11-13 2021-11-13 Sensibilisateur de cellules nk préparé au moyen d'une combinaison de nano-sélénium fonctionnel avec de la metformine et utilisation associée

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
PCT/CN2021/130493 WO2023082220A1 (fr) 2021-11-13 2021-11-13 Sensibilisateur de cellules nk préparé au moyen d'une combinaison de nano-sélénium fonctionnel avec de la metformine et utilisation associée

Publications (1)

Publication Number Publication Date
WO2023082220A1 true WO2023082220A1 (fr) 2023-05-19

Family

ID=86334886

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/CN2021/130493 Ceased WO2023082220A1 (fr) 2021-11-13 2021-11-13 Sensibilisateur de cellules nk préparé au moyen d'une combinaison de nano-sélénium fonctionnel avec de la metformine et utilisation associée

Country Status (1)

Country Link
WO (1) WO2023082220A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN117982531A (zh) * 2024-03-13 2024-05-07 广东海洋大学 羊栖菜多糖纳米硒的制备方法及在制备抗肿瘤药物中的应用

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102327620A (zh) * 2011-07-29 2012-01-25 暨南大学 纳米硒在抗肿瘤药物载体中的应用
CN106692181A (zh) * 2017-01-23 2017-05-24 暨南大学 纳米硒作为cik细胞增敏剂的应用

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102327620A (zh) * 2011-07-29 2012-01-25 暨南大学 纳米硒在抗肿瘤药物载体中的应用
CN106692181A (zh) * 2017-01-23 2017-05-24 暨南大学 纳米硒作为cik细胞增敏剂的应用

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
"Master's Thesis", 30 June 2020, JINAN UNIVERSITY, CN, article ZHANG ZEHANG: "The Application and Action Mechanism of Functionalized SeNPs Combined with Metformin as Anti-tumor Therapy Agents", pages: 1 - 68, XP009545539, DOI: 10.27167/d.cnki.gjinu.2020.000978 *
EBOKAIWE AZUBUIKE P.; OKORI STEPHEN; NWANKWO JOSEPH O.; EJIKE CHUKWUNONSO E. C. C.; OSAWE SHARON O.: "Selenium nanoparticles and metformin ameliorate streptozotocin-instigated brain oxidative-inflammatory stress and neurobehavioral alterations in rats", NAUNYN-SCHMIEDEBERG'S ARCHIVES OF PHARMACOLOGY, SPRINGER, DE, vol. 394, no. 4, 1 January 1900 (1900-01-01), DE , pages 591 - 602, XP037403660, ISSN: 0028-1298, DOI: 10.1007/s00210-020-02000-2 *
YANG YU, ZHANG ZEHANG, CHEN QI, YOU YUANYUAN, LI XIAOLING, CHEN TIANFENG: "Functionalized Selenium Nanoparticles Synergizes With Metformin to Treat Breast Cancer Cells Through Regulation of Selenoproteins", FRONTIERS IN BIOENGINEERING AND BIOTECHNOLOGY, vol. 9, XP093065745, DOI: 10.3389/fbioe.2021.758482 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN117982531A (zh) * 2024-03-13 2024-05-07 广东海洋大学 羊栖菜多糖纳米硒的制备方法及在制备抗肿瘤药物中的应用

Similar Documents

Publication Publication Date Title
TW201842921A (zh) 治療性ｒｎａ
Park et al. Enhanced antitumor efficacy of cisplatin in combination with HemoHIM in tumor-bearing mice
JP2018513209A (ja) 遺伝子組換え細菌ｖｎｐ２０００９−ｍの、がん転移を予防および治療するための医薬品の調製における使用
CN107801379A (zh) 抗癌剂
Li et al. Current status and future challenges of CAR-T cell therapy for osteosarcoma
CN114286681A (zh) Nad+和/或nad+抑制剂和/或nad+激动剂的用途及其联合制剂
Shyyka et al. Anticancer activity evaluation of thieno [3, 2-e][1, 2, 3] triazolo [1, 5-a] pyrimidines and thieno [2, 3-e][1, 2, 3] triazolo [1, 5-a] pyrimidine derivatives
JPH09501411A (ja) 抗−新生物組成物及びその使用方法
CN106692181A (zh) 纳米硒作为cik细胞增敏剂的应用
US20200377545A1 (en) Novel anti-tumor compound and application thereof in preparing anti-tumor drugs
CN114177194A (zh) 功能化纳米硒联合二甲双胍制备的nk细胞增敏剂及应用
Yang et al. The killing effect of Tanshinol on breast cancer cells: insight into the reversion of TGF-𝜷1-mediated suppression of NK cell functions
WO2023082220A1 (fr) Sensibilisateur de cellules nk préparé au moyen d'une combinaison de nano-sélénium fonctionnel avec de la metformine et utilisation associée
Wang et al. Adoptive immunotherapy for stage IV renal cell carcinoma: a novel protocol utilizing periodate and interleukin-2-activated autologous leukocytes and continuous infusions of low-dose interleukin-2
WO2022116369A1 (fr) Préparation et composition d'un nouveau type de complexe d'agoniste de sting et utilisation de ce dernier dans un médicament antitumoral
US11534448B2 (en) Use of canagliflozin in preparation of antitumor drug
Liu et al. IDO Inhibitor and gallic acid cross-linked small molecule drug synergistic treatment of melanoma
TWI649328B (zh) 作為紅血球生成素及紅血球生成素受體之正變構調節子以治療紅血球生成素缺乏疾病之新化合物
CN111138508A (zh) 创新环二核苷酸金属配合物的合成及其在抗肿瘤药物中的应用
Lee et al. Isolinderalactone inhibits glioblastoma cell supernatant-induced angiogenesis
CN117447610A (zh) 一种嵌合抗原受体（car）及其在制备抗肿瘤药物中的应用
CN115925974A (zh) 一种针对实体瘤的通用性的ips来源的car-nk细胞的制备方法
WO1989011487A1 (fr) Derives d'amp cyclique pour traiter le cancer
EP3684355B1 (fr) Ovothiols pour le traitement d'une inflammation généralisée chronique de faible grade (clgsi) et de maladies associées
CN113855788A (zh) 一种抗体纳米粒子及其制备方法和应用

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 21963668

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 21963668

Country of ref document: EP

Kind code of ref document: A1