Composite preparation based on NK cell extract and anticancer application thereof

Abstract

The invention discloses a compound preparation based on an NK cell extract and anti-cancer application of the compound preparation, and relates to the technical field of biological medicines. On the basis of researching the anti-cancer effect of the NK cell extract, the NK cell extract is wrapped and protected by a nano material, and meanwhile, the nano material is compounded with two plant extracts of oldenlandia diffusa and sculellaria barbata, vitamin B1 and sodium dichloroacetate to form a broad-spectrum powerful anti-cancer preparation.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine technology, and in particular to a composite preparation based on NK cell extract and its anti-cancer application. Background Art

[0002] NK cells are key elements of innate immunity, able to mediate powerful antitumor and antiviral responses, directly killing cancer or infected cells and indirectly improving antibody- and T-cell-mediated responses. Indeed, NK cell-mediated cytotoxicity and cytokine release can influence the activity of additional innate immune cells (dendritic cells, macrophages and neutrophils) and confer regulatory functions on NK cells, able to influence subsequent antigen-specific, HLA-restricted T and B-cell responses. The expression of a single self-iKIR by NK cells confers an advantage to the host, as it allows sensing loss or peptide-induced alterations of a single HLA class I allotype, leading to killing of compromised cells by a mechanism of “loss of self-recognition”.

[0003] Some studies have suggested that NK cells are interleukin-2 (IL-2) dependent, that is, IL-2 can promote their proliferation or activation. IL-2 is a cytokine secreted by activated T lymphocytes, composed of 133 amino acids. The IL-2 gene is located on chromosome 4, and the disulfide bond formed by cysteine ​​at positions 58 and 105 plays an important role in maintaining its activity. The discovery of IL-2 began in the mid-1960s. People found that the supernatant of leukocyte culture activated by antigen or mitogen contained a factor that could stimulate lymphocyte division; this factor could be used to maintain T cell culture for more than 9 months. This technology was quickly adapted to the culture of tumor-reactive T cells with sustained cytotoxic potential, and it was later named IL-2. The sequencing of the human IL-2 gene was reported in 1983, and then human recombinant IL-2 was developed in 1985 and used for the treatment of malignant tumors, opening the way for IL-2 immunotherapy and achieving satisfactory therapeutic effects. Currently, IL-2 has been used in the treatment of melanoma, lung metastasis, renal cell carcinoma, colon cancer, lung adenocarcinoma and other tumors. IL-2 can promote the proliferation of NK cells in vitro. IL-2, IL-15, IL-7 or feeder cell treatment have been proven to increase NK cell toxicity. The infusion of activated NK cells and IL-2 push have also been proven to be safe. However, the activity of natural IL-2 still needs to be improved, so researchers began to pay attention to the modification of the IL-2 sequence. For example, EP0200280A2 proposed replacing the methionine residue at position 104 in IL-2 with a conservative amino acid to improve the antioxidant activity; CN111018961A proposed introducing a cysteine ​​residue to form a disulfide bond to improve the binding activity of IL-2 to its receptor; CN113667004A proposed that site-directed mutagenesis at position 42 / 44 of IL-2 can improve anti-tumor activity; CN101426916A proposed a C125S mutation, and the mutant protein provides maintenance or enhancement of human NK cell-mediated natural killer cytotoxicity, lymphokine-activated killer (LAK) cytotoxicity, or ADCC-mediated cytotoxicity.

[0004] CD20 is one of the star targets of anti-tumor drugs. It is a transmembrane phosphoprotein composed of 279 amino acids. It is also a unique marker present on the surface of human B lymphocytes. The relative molecular mass is 33,000-37,000. It has 4 transmembrane domains, of which the main antigen epitope is located between the third and fourth transmembrane regions on the inner side of the cytoplasmic membrane. The main antigen epitope has a very important regulatory effect on the differentiation and proliferation of B lymphocytes. In the past 30 years, the US FDA has approved a total of 6 CD20 target antibody drugs for marketing, including 2 mouse antibody-conjugated isotope drugs. Among them, rituximab has gained a large market share, with sales of nearly US$7 billion in 2019. Ofatumumab, obinutuzumab, and ocrelizumab were also launched in 2009, 2013, and 2017, respectively, for the treatment of leukemia and multiple myeloma.

[0005] Although a large number of fruitful studies have been conducted on NK cell extracts, IL-2, anti-CD20 antibodies, etc. in the prior art, there is still the problem of how to enhance and improve the anti-tumor effect of NK cells. Based on the existing research, the present invention further regulates the expression levels of multiple factors and improves the tumor killing effect by compounding NK cell extracts with traditional Chinese medicine extracts, vitamin B1, and dichloroacetic acid. Summary of the invention

[0006] The main purpose of the present invention is to provide a composite preparation based on NK cell extract and its anti-cancer application, and to innovatively develop a product that is more efficient than the existing NK cell extract for treating cancer.

[0007] To achieve the above purpose, the present invention, based on the study of the anti-cancer effect of NK cell extracts, uses nanomaterials to wrap and protect NK cell extracts, and simultaneously uses two plant extracts of Hedyotis diffusa and Scutellaria barbata, vitamin B1, and sodium dichloroacetate to form a composite preparation. The composite preparation can be a capsule, tablet or any other suitable dosage form; it has a broad-spectrum and efficient anti-cancer function.

[0008] NK cell extracts are composed of NK cell exosomes and NK cell lysates, including a variety of cytokines such as IL-2 and IFN-γ, and have similar biological activities to NK cells. NK cell exosomes are prepared by ultrafiltration centrifugation. NK cell extracts protected by nanomaterials have the dual effects of improving immunity and directly killing cancer cells.

[0009] Both Hedyotis diffusa and Scutellaria barbata are both medicinal and edible plants. The active ingredients contained in their extracts can directly destroy the DNA structure of cancer cells, interfere with the signal transduction pathways during cancer cell proliferation, and thus exert an anti-cancer effect.

[0010] Vitamin B1, also known as thiamine, is the earliest discovered water-soluble B vitamin. It is very important for maintaining normal cell metabolism, cell growth and proliferation. High-dose vitamin B1 can inhibit the growth and proliferation of tumor cells, target and correct the aerobic glycolysis phenotype of tumors (Warburg effect), thereby inducing tumor cell apoptosis; it has no adverse effects on normal cells and has almost no dose-limiting toxic reactions. The anti-cancer mechanism of high-dose vitamin B1 may include the role of oxidative decarboxylase cofactors and PDK inhibitors.

[0011] Dichloroacetate can inhibit glycolysis in tumors by inhibiting pyruvate dehydrogenase kinase, increasing the flow of pyruvate into mitochondria, promoting sugar oxidation, and inhibiting glycolysis in tumors. This effect can reverse dysfunctional mitochondria, restore their function, and promote tumor cell apoptosis, thereby inhibiting the growth of tumor cells.

[0012] The present invention prepares a new composite preparation based on NK cell extracts, and the various components not only have a significant comprehensive anti-cancer effect, but are also very safe and will not cause toxic side effects such as cytotoxic anticancer drugs or targeted anticancer drugs. DETAILED DESCRIPTION

[0013] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0014] In an embodiment of the present invention, a composite preparation based on NK cell extract is provided, wherein the weight proportions of the composite preparation are: 0.7-1.2 (35-60 mg) parts of NK cell extract protected by nanomaterials, 2.1-3.6 (105-180 mg) parts of Hedyotis diffusa extract, 2.1-3.6 (105-180 mg) parts of Scutellaria barbata extract, 2.1-3.6 (105-180 mg) parts of vitamin B1, and 7-12 (350-600 mg) parts of sodium dichloroacetate.

[0015] in:

[0016] NK cell extracts contain a large number of cytokines including IL-2 and IFN-γ. NK cell extracts protected by nanomaterials have the dual effects of improving immunity and directly killing cancer cells.

[0017] IL-2 is a cytokine secreted by activated T lymphocytes. It is composed of 133 amino acids. The IL-2 gene is located on chromosome 4. The disulfide bond formed by cysteine ​​at positions 58 and 105 plays an important role in maintaining its activity. The discovery of IL-2 began in the mid-1960s. People found that the supernatant of leukocyte culture activated by antigen or mitogen contained a factor that could stimulate lymphocyte division; this factor could be used to maintain T cell culture for more than 9 months. This technology was quickly adapted to the culture of tumor-reactive T cells with sustained cytotoxic potential, and it was later named IL-2. The sequencing of the human IL-2 gene was reported in 1983, and human recombinant IL-2 was developed in 1985 and used in the treatment of malignant tumors, opening the way for IL-2 immunotherapy and achieving satisfactory therapeutic effects. At present, IL-2 has been used in the treatment of many tumors such as melanoma, lung metastasis, renal cell carcinoma, colon cancer, and lung adenocarcinoma.

[0018] IFN-γ exerts its killing effects directly by inducing apoptosis or promoting non-apoptotic cell death, and indirectly by making tumor cells susceptible to apoptosis-inducing immune responses or chemotherapy. For example, IFN-γ induces IRF1, a tumor suppressor gene, which in turn decreases Bcl2 and increases Bak. These events promote the release of cytochrome c from mitochondria and the activation of caspases, leading to apoptosis. Low concentrations of reactive oxygen species (ROS) and reactive nitrogen intermediates (RNI) are associated with cell proliferation. However, tumor cells that respond to IFN-γ stimulation produce large amounts of RNI and ROS and tend to undergo apoptosis. IFN-γ can also induce non-apoptotic cell death by inducing autophagy in human hepatocellular carcinoma (HCC). IFN-γ-induced STAT1 activation enhances the expression of the death receptor FAS and its ligand FAS-L in liver and colon cancer cells, and the expression of TRAIL and its receptor death receptor 5 (DR5) in human tumor cell lines. Thus, activated STAT1 sensitizes tumor cells to FAS- or TRAIL-mediated apoptosis. Furthermore, activation of STAT1 by IFN-γ suppressed the expression of the p53 inhibitor mouse double minute 2 (MDM2), thereby enhancing doxorubicin- and cisplatin-induced p53-induced apoptosis.

[0019] Although the IFN-γ / STAT1 pathway induces apoptosis in tumor cells, activation of STAT1 can also inhibit tumor cell growth and establish a dormant state. In melanoma, activation of the IFN-γ / STAT1 pathway leads to downregulation of cyclin E and cyclin A, which results in tumor cell dormancy. Activated STAT1 can also interact with cyclins D1, D2, D3, and CDK4, leading to cell cycle arrest in fibrosarcoma cells. IFN-γ-induced STAT1 activation upregulates the miRNA-29 family and downregulates CDK6 in melanoma cells, resulting in tumor suppressor function. IFN-γ-mediated tumor dormancy can also be induced independently of STAT1 signaling. Tumor clones that highly express indoleamine 2,3-dioxygenase 1 (IDO1) and kynurenine (Kyn)-aryl hydrocarbon receptor (AhR) respond to IFN-γ by upregulating the cell cycle inhibitor p27, thereby preventing STAT1 signaling and inducing tumor dormancy. In fact, p21 and p27 promote hypophosphorylation of the tumor suppressor Rb, thereby inhibiting the activity of the E2F transcription factor and suppressing the activation of genes involved in cell proliferation. During T antigen (Tag)-induced pancreatic islet multistage carcinogenesis, IFN-γ-producing CD4+ T cells inhibit tumor cell proliferation and establish tumor dormancy without destroying malignant cells. It has also been reported that CD8+ T cells maintain mouse B-cell lymphoma (BCL1) in a dormant state by producing IFN-γ. Radiation-induced tumor dormancy in Balb / c neu transgenic mice is also mediated by IFN-γ, and therefore, neutralization of IFN-γ can reverse radiation-induced tumor dormancy and lead to tumor recurrence. It has been shown that the expression level of IFN-γRα on breast tumor cells determines whether IFN-γ eliminates tumors or establishes tumor dormancy. Although low expression of IFN-γRα in tumor cells leads to tumor dormancy, high levels of IFN-γRα expression in FVB mice lead to tumor elimination and neu-specific CD8+ T cell responses that produce IFN-γ.

[0020] Many chemical components contained in Hedyotis diffusa have inhibitory effects on various types of cancer, such as polysaccharides on lymphosarcoma No. 1 ascites type and Ehrlich ascites carcinoma subcutaneous type; coumarins on uterine cancer type 14, sarcoma 180, and solid liver cancer; triterpenes have inhibitory effects on lymphosarcoma No. 1 ascites type, cervical cancer type 14, sarcoma 180, and solid liver cancer. Triterpenes, coumarins, polysaccharides and sterols are isolated from the ethanol extract of Hedyotis diffusa. The obtained components are studied in animal experiments. The results show that triterpenes have the best anti-tumor effect. Clinical studies have shown that stigmasterol, β-sitosterol, oleanolic acid, ursolic acid, and para-coumaric acid can stimulate the proliferation of the reticuloendothelial system and enhance the ability of phagocytic cells. Ursolic acid and oleanolic acid have antibacterial, anti-inflammatory, anti-cancer, and enzyme-lowering effects, inhibit the growth of sarcoma 180 beads, and are effective anti-tumor ingredients. Zhou Jianbo et al. conducted qualitative and semi-quantitative analysis of the trace elements in this product and found that this product contains seven elements, including iron (Fe), manganese (Mn), titanium (Ti), magnesium (Mg), aluminum (Ai), silicon (Si), and calcium (Ca). In addition, the Ti element can stimulate phagocytes and enhance the body's immune ability.

[0021] White flower oldenlandia diffusa has the effect of inhibiting the killing of liver cancer cells in vitro. In vitro, it can not only inhibit the growth of transplanted S-180 solid tumors in mice, but also significantly improve the atrophy of immune organs and damage to the hematopoietic system caused by cyclophosphamide when used in combination with cyclophosphamide. Microscopic observation found that lymphocytes, neutrophils, lymph nodes, spleen, and reticuloendothelial cells in the liver also proliferated significantly around the cancer tissue. The above shows that white flower oldenlandia diffusa has anti-cancer effects both in vivo and in vitro.

[0022] Influencing the function of T cells The role of T lymphocyte-mediated cellular immunity in the body's anti-tumor has long been known. It can directly kill cancer cells and other foreign molecules while preserving itself; it can enhance the body's humoral immune function by assisting B lymphocytes in producing antibodies; after being activated by antigens or non-specific mitogens, it can produce various lymphokines with immune enhancement and anti-cancer effects. White flower oldenlandia diffusa extract can activate T lymphocytes, causing them to release interferon and kill tumor cells. It has the effects of anti-mutation, anti-damage to lymphocyte DNA caused by tobacco tar, and enhancing the non-specific immune function of the human body.

[0023] The ethanol extract of Hedyotis diffusa (SCD) can significantly enhance the proliferation of PBMC induced by concanavalin (Con A) or lipopolysaccharide (LPS). When combined with Con A or LPS, the proliferation effect of SCD is stronger than when added alone, and it shows a strong ability to promote proliferation at a low concentration of 0.125 μg / L. The proliferation rate reached 344.13% and 152.18%. SCD showed an inhibitory effect on the proliferation of human oral epidermoid carcinoma cells (KB), human poorly differentiated gastric adenocarcinoma cells (BGC), human myeloid leukemia cells (HL-60), mouse melanoma cells (B16), human hepatoma cells (SMMC-7721), human cervical cancer cells (HELA), and human lung cancer cells (A549). The proliferation inhibition rates at high concentrations (100 μg / L) were 30.53%, 42.82%, 23.77%, 63.93%, 39.40%, 46.33%, and 42.56%, respectively.

[0024] The water extract of Hedyotis diffusa has a proliferation inhibition effect on human intestinal adenocarcinoma cell line SW620 at low concentrations, with a proliferation inhibition rate of 30%; at high concentrations, the proliferation inhibition rates of human intestinal cancer cell line SW620, breast cancer cell line MCF7, liver cancer cell line HepG2, and intestinal adenocarcinoma cell line LS180 are 41%, 19%, 25%, and 40%, respectively. Zhao Haoru et al. have proved through experiments that the water-soluble polysaccharides and lipophilic extracts of Hedyotis diffusa (the extracts were identified by thin layer chromatography, and the results showed that they contained glucose, galactose, arabinose, and rhamnose) have a significant inhibitory effect on sarcoma 180 tumor cells. Oral administration (62.5 / 31.25 mg / kg) can significantly inhibit the growth of mouse transplanted S-180 solid tumors (P<0.05), and the inhibition rate is more than 40%. While inhibiting tumors, the polysaccharide part has a protective effect against chemical damage.

[0025] Scutellaria barbata extract is a dry whole herb extract of Scutellaria barbata D.Don, a plant of the Labiatae family. Flavonoids are its main active ingredients. It has anti-inflammatory, anti-tumor, anti-chemical book oxidation, anti-atherosclerosis and other biological activities, and has great application value in the prevention and treatment of cardiovascular and cerebrovascular diseases. It is commonly used in the clinic to treat various tumors such as liver cancer, lung cancer, gastric cancer, rectal cancer, gynecological tumors and leukemia.

[0026] The antioxidant activity of the ethanol extract of Scutellaria barbata and the extracts with different polar solvents in vitro was studied. The ethanol extract of Scutellaria barbata had significant antioxidant activity, and each extract could reduce its antioxidant activity. The total alkaloid extract of Scutellaria barbata had antibacterial effects on Staphylococcus aureus, Escherichia coli, Enterococcus faecalis, and Pseudomonas aeruginosa. The total flavonoids of Scutellaria barbata showed a certain therapeutic or (and) antagonistic effect on the infection of mice caused by influenza A (H1N1) virus and the death caused by it, and its efficacy showed a dose-related characteristic. Three flavonoid compounds were isolated from Scutellaria barbata, namely kaempferol-3-O-rhamnoside, kaempferol-3-O-glucoside, and kaempferol-3,7-dirhamnoside. They were found to have good anti-Helicobacter pylori activity and could reduce the minimum inhibitory concentration of ampicillin against Helicobacter pylori.

[0027] Scutellaria barbata ethanol extract has a certain anti-fibrosis effect on rat liver, can affect the apoptosis of activated hepatic stellate cells, and improve the degree of fibrosis of liver tissue. Scutellaria barbata polysaccharide has the effect of resisting and scavenging free radicals, can improve the body's antioxidant capacity, and has a certain protective effect on carbon tetrachloride-induced liver injury in mice. Scutellaria barbata polysaccharide has a significant inhibitory effect on CCL4 liver injury mouse liver homogenate and in vitro induced lipid peroxidation, and can significantly reduce the level of MDA in its liver, and has the effect of scavenging oxygen free radicals and resisting lipid peroxidation.

[0028] Scutellaria barbata extract has the effects of inhibiting the proliferation of gastric cancer SGC7901 cells and inducing apoptosis of gastric cancer cells.

[0029] Vitamin B1, also known as thiamine, is the earliest discovered water-soluble B vitamin. It is very important for maintaining normal cell metabolism, cell growth and proliferation. High-dose vitamin B1 can inhibit the growth and proliferation of tumor cells, target and correct the aerobic glycolysis phenotype of tumors (Warburg effect), thereby inducing tumor cell apoptosis; it has no adverse effects on normal cells and has almost no dose-limiting toxic reactions. The anti-cancer mechanism of high-dose vitamin B1 may include the role of oxidative decarboxylase cofactors and PDK inhibitors.

[0030] Dichloroacetate can be said to be the earliest metabolic anticancer drug discovered. It has low toxicity, is relatively cheap, has a broad spectrum of anticancer effects, and can alleviate the effects of excessive lactic acid caused by advanced cancer.

[0031] Dichloroacetate can activate pyruvate dehydrogenase kinase, thereby promoting the entry of pyruvate into mitochondria, increasing sugar aerobic metabolism, mediating mitochondrial depolarization, reversing the reconstructed mitochondria, and restoring the mitochondrial membrane potential level close to that of non-tumor cells. This process does not affect normal non-tumor cells. Relying on the above process, ROS generation increases, MTP opens, c-ymc and apoptosis-inducing factors are released from mitochondria, thereby mediating cell apoptosis.

[0032] Mitochondria also regulate some important K + , Ca 2+ Plasma channels, such as Kv channels in some tumor cells, are inhibited due to mitochondrial remodeling. If Kv channels are inhibited, cells may resist apoptosis. Studies have shown that dichloroacetate can reduce mitochondrial membrane potential, restore mitochondrial function, increase mitochondrial H2O2, activate Kv channels in tumor cells, and do not affect normal cells, further mediate cell apoptosis, and inhibit tumor growth.

[0033] Embodiment 1:

[0034] The composite preparation is a capsule, and the weight parts are: 0.9 parts of NK cell extract wrapped and protected by nanomaterials, 2.7 parts of Hedyotis diffusa extract, 2.7 parts of Scutellaria barbata extract, 2.7 parts of vitamin B1, and 9 parts of sodium dichloroacetate.

[0035] Embodiment 2:

[0036] The composite preparation is a capsule, and the weight parts are: 1 part of NK cell extract wrapped and protected by nanomaterials, 3 parts of Hedyotis diffusa extract, 3 parts of Scutellaria barbata extract, 2.4 parts of vitamin B1, and 10 parts of sodium dichloroacetate.

[0037] Embodiment 3:

[0038] The composite preparation is a capsule, and the weight parts are: 0.9 parts of NK cell extract wrapped and protected by nanomaterials, 2.7 parts of Hedyotis diffusa extract, 2.7 parts of Scutellaria barbata extract, 2.7 parts of vitamin B1, and 9 parts of sodium dichloroacetate.

[0039] Embodiment 4:

[0040] The composite preparation is a capsule, and the weight parts are: 1.1 parts of NK cell extract wrapped and protected by nanomaterials, 3.7 parts of Hedyotis diffusa extract, 3 parts of Scutellaria barbata extract, 3 parts of vitamin B1, and 9.9 parts of sodium dichloroacetate.

[0041] Embodiment 5:

[0042] The composite preparation is a capsule, and the weight parts are: 1 part of NK cell extract wrapped and protected by nanomaterials, 3.3 parts of Hedyotis diffusa extract, 3.3 parts of Scutellaria barbata extract, 2.7 parts of vitamin B1, and 10 parts of sodium dichloroacetate.

[0043] Test Example 1: Animal Experiment

[0044] 1.1 Anti-tumor effect

[0045] 1.1.1 Experimental subjects: tumor-bearing mice (H22 and S180) weighing 19-23 g were selected, of which H22 and S180 were obtained from the Institute of Materia Medica, Chinese Academy of Medical Sciences;

[0046] 1.1.2 Experimental methods and results: 0.2 mL of ascites of the breeding mice was added, and the breeding was subcutaneously transferred once a week. The breeding was carried out in the right axilla, and the drug was administered by intraperitoneal injection on the second day. The negative control group was given NS, and the experimental group was given capsules prepared in Examples 1, 2, 3, 4, and 5 (solid preparations were dissolved in water and liquid preparations were concentrated before use to prepare equal volume of drug solution). The positive control group was given cyclophosphamide 0.4 mg / mouse, once a day; H22 mice were given the drug for 10 days, and the tumor weight was measured the next day; S180 mice were given the drug for 8 days, and the tumor weight was measured the next day, and the tumor inhibition rate was calculated. The tumor inhibition effects on H22 mice and H180 mice are shown in Table 1:

[0047] Table 1:

[0048]

[0049] Note: Compared with the negative group, *P<0.05, **P<0.01

[0050] 1.2 Effects on the phagocytic function of mouse macrophages

[0051] BALB / c mice, weighing 16-20g, both male and female, were randomly divided into 6 groups. Group 1 was given NS injection, and groups 2-6 were given capsules made from Examples 1, 2, 3, 4, and 5 (solid preparations were dissolved in water and liquid preparations were concentrated before use to make equal volume of liquid medicine). The dosage was 0.4ml / mouse, and the mice were injected once a day. On the 4th day, 0.4ml / mouse of 0.5% starch NS solution was injected. 30 minutes after administration on the 6th day, 1ml / mouse of 2% chicken red blood cell suspension was injected. After 30 minutes, the mice were killed, the peritoneal cavity was flushed with 2mlNS, the peritoneal fluid was smeared, and after drying, it was stained with Wright's stain, and 100 macrophages were examined under a microscope to calculate the phagocytic rate and phagocytic index. The results are shown in Table 2.

[0052] Table 2

[0053]

[0054] Note: Compared with NS group, *P<0.05, **P<0.01

[0055] The above results show that: 1. The composite capsule of the present invention has a significant anti-tumor effect on both H22 mice and S180 mice, and the effect is better than cyclophosphamide; 2. The composite capsule of the present invention can significantly increase BALB / C Phagocytic function of mouse phagocytes; 3. The high prescription amount of the composite capsule of the present invention has an anti-tumor effect, but the low prescription amount has an unsatisfactory effect, which indicates that the present invention can play a better role within the prescription range.

[0056] Test Example 2: Toxicity Test

[0057] 50 healthy mice were selected, weighing 80-120g, and randomly divided into 5 groups, each group of male and female half, and the composite capsules made of Examples 1, 2, 3, 4, and 5 were administered respectively, and the injection preparation was injected at a volume of 0.2mL / 10g each time, and the oral preparation was administered by gavage at a volume of 0.4ml / 10g, once in the morning and evening. After 7 days of administration, the mice of Examples 1, 2, 3, and 4 did not show any abnormal performance, diet, good activity, and no death. The mental state of the mice given Example 5 (high prescription) was slightly depressed, the hair was slightly dull, and the appetite was slightly poor; after 13 weeks of administration, each group of mice had no abnormal performance, lively behavior, smooth back hair, normal diet, average body weight of 340g, and no significant differences in blood picture, biochemical indexes, and organ coefficients. Considering factors such as whether body tolerance and long-term toxicity will accumulate, the high prescription dosage of Example 5 is not considered.

[0058] In addition, composite capsules prepared by the prescription amount of the composition of the present invention in Examples 2, 3, and 4 were subjected to general pharmacological tests on blood pressure, heart rate, electrocardiogram, respiration, etc. in rabbits after administration, as well as tests on the central nervous function of rats in awake and anesthetized states. It was found that the blood pressure, heart rate, electrocardiogram, and respiration of the animals after administration were not significantly changed compared with those before administration, and no excitement or inhibition was observed in the animals. This indicates that the medication is safe within the prescription range of the present invention.

[0059] The above description is only a preferred embodiment of the present invention, and does not limit the patent scope of the present invention. All equivalent structural changes made by using the contents of the present invention specification under the inventive concept of the present invention, or directly / indirectly applied in other related technical fields are included in the patent protection scope of the present invention.

Claims

1. A composite preparation based on NK cell extract, characterized in that: The composite preparation comprises the following parts by weight: 0.7-1.2 parts of NK cell extract protected by nanomaterials, 2.1-3.6 parts of oldenlandia diffusa extract, 2.1-3.6 parts of Scutellaria barbata extract, 2.1-3.6 parts of vitamin B1, and 7-12 parts of sodium dichloroacetate.

2. A composite preparation based on NK cell extract according to claim 1, characterized in that: The compound preparation can be in the form of capsules, tablets or other suitable dosage forms.

3. A composite preparation based on NK cell extract according to claim 1, characterized in that: The composite preparation comprises the following parts by weight: 0.8-1.1 parts of NK cell extract protected by nanomaterials, 2.4-3.3 parts of Hedyotis diffusa extract, 2.4-3.3 parts of Scutellaria barbata extract, 2.4-3.3 parts of vitamin B1, and 8-11 parts of sodium dichloroacetate.

4. The composite preparation based on NK cell extract according to claim 1, characterized in that: The weight proportions of the composite preparation are as follows: 0.9 parts of NK cell extract protected by nanomaterials, 2.7 parts of Hedyotis diffusa extract, 2.7 parts of Scutellaria barbata extract, 2.7 parts of vitamin B1, and 9 parts of sodium dichloroacetate.

5. The composite preparation based on NK cell extract according to claim 1, characterized in that: The weight proportions of the composite preparation are as follows: 1 part of NK cell extract protected by nanomaterials, 3 parts of Hedyotis diffusa extract, 3 parts of Scutellaria barbata extract, 2.4 parts of vitamin B1 and 10 parts of sodium dichloroacetate.

6. The composite capsule based on NK cell extract according to claim 1, characterized in that: The weight proportions of the composite preparation are as follows: 0.9 parts of NK cell extract protected by nanomaterials, 2.7 parts of Hedyotis diffusa extract, 2.4 parts of Scutellaria barbata extract, 2.7 parts of vitamin B1 and 9 parts of sodium dichloroacetate.

7. An application of a composite preparation based on NK cell extract in anti-cancer, characterized in that: Use the composite preparation based on NK cell extract as described in any one of claims 1 to 6 above.

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