A traditional Chinese medicine composition for treating lung cancer and a preparation method thereof

Abstract

The application discloses a traditional Chinese medicine composition for treating lung cancer and a preparation method thereof. The traditional Chinese medicine composition comprises the following raw medicinal materials in parts by weight: Fritillaria thunbergii Miq. 5-8 parts, Caesalpinia sappan 5-8 parts, Polygonatum odoratum 5-8 parts, Iridaceae 5-8 parts, Platycodon grandiflorum 5-8 parts, Inula japonica 5-8 parts, Sasa veitchii 22-25 parts, Humulus scandens 9-12 parts, Bidens pilosa 22-25 parts and Glycyrrhiza uralensis 2-5 parts. The traditional Chinese medicine composition is developed on the basis of the phlegm and blood stasis blocking lung pathogenesis of lung cancer in traditional Chinese medicine diagnosis and treatment, combines the phlegm and blood stasis treatment method of traditional Chinese medicine and laboratory research, and has the effects of resolving phlegm and removing blood stasis and inhibiting tumor growth, and has a good effect on the treatment of lung cancer. Experiments prove that the traditional Chinese medicine composition significantly inhibits lung cancer cell proliferation in vitro experiments and improves the lung cancer pathological process of lung cancer mice in vivo experiments, and is a great innovation of a lung cancer treatment drug.

Description

Technical Field

[0001] The present invention relates to a traditional Chinese medicine composition for treating lung cancer and a preparation method thereof, belonging to the technical field of traditional Chinese medicine. Background Art

[0002] Surgery, chemotherapy, and immune checkpoint inhibitors are commonly used clinically to treat lung cancer and can significantly prolong patient survival. However, these treatments are slow, have significant side effects, and many patients develop drug resistance or experience immune escape from tumor cells during treatment, which impacts treatment outcomes and increases the economic burden on patients and society. Therefore, there is an urgent need to further explore the mechanisms of lung cancer progression and identify novel therapeutic targets and safe and effective drugs.

[0003] Traditional Chinese medicine has great potential in the treatment of cancer. It has the effects of enhancing the effects of radiotherapy and chemotherapy, improving the quality of life of patients, and prolonging survival. Therefore, it is of great significance to strengthen the research of Chinese medicine compositions in the treatment of lung cancer. There is no name for lung cancer in traditional Chinese medicine. Based on literature records and clinical manifestations, scholars currently believe that lung cancer belongs to the category of lung diseases such as "cough", "phlegm and fluid retention", "lung abscess", and "lung accumulation" in traditional Chinese medicine. The lung is the organ that stores phlegm. When the lung qi fails to ascend and descend properly, the movement of qi in the meridians is blocked, and the distribution of body fluids is abnormal, which will accumulate as phlegm; when qi flows, blood flows, and when qi stagnates, blood stasis occurs. Phlegm and blood stasis fight against each other and accumulate in the chest to form a mass. Therefore, many famous traditional Chinese medicine practitioners in modern times believe that phlegm and blood stasis blocking the lungs is the key pathogenesis of lung cancer, and resolving phlegm and removing blood stasis is an important treatment method in the whole process of clinical diagnosis and treatment of lung cancer in traditional Chinese medicine. However, although there are traditional Chinese medicine compositions for lung cancer that are formulated with resolving phlegm and removing blood stasis as the main treatment method, some of them have problems such as complex formulation, high cost, and failure to achieve the expected treatment effect. Therefore, strengthening the research and development of new Chinese herbal prescriptions for resolving phlegm and removing blood stasis for lung cancer is of great significance for improving the level of clinical lung cancer prevention and treatment, reducing patient mortality, and improving patients' quality of life. Summary of the Invention

[0004] In view of the deficiencies in the prior art, the present invention aims to provide a Chinese medicine composition for treating lung cancer, a preparation method thereof, and an application thereof, thereby improving the quality of life of patients, prolonging their survival time, and reducing their mortality rate during the clinical prevention and treatment of lung cancer.

[0005] In order to achieve the above-mentioned purpose, the technical solution of the present invention is, on the one hand, to provide a traditional Chinese medicine composition for treating lung cancer, comprising the following raw materials in parts by weight: 5-8 parts of Fritillaria thunbergii, 5-8 parts of Sappan wood, 5-8 parts of Aster tataricus, 5-8 parts of Bletilla striata, 5-8 parts of Platycodon grandiflorum, 5-8 parts of Inula japonica, 22-25 parts of Cephalotaxus chinensis, 9-12 parts of Polygonum cuspidatum, 22-25 parts of Bidens pilosa, and 2-5 parts of Licorice.

[0006] Preferably, the traditional Chinese medicine composition comprises the following raw materials in parts by weight: 5 parts of Fritillaria thunbergii, 5 parts of Sappan wood, 5 parts of Aster tataricus, 5 parts of Bletilla striata, 5 parts of Platycodon grandiflorum, 5 parts of Inula flower, 22 parts of Ligusticum chuanxiong, 9 parts of Polygonum cuspidatum, 22 parts of Bidens pilosa, and 2 parts of Licorice.

[0007] On the other hand, the technical solution of the present invention is to provide a method for preparing the Chinese medicine composition, comprising the following steps:

[0008] (1) Grind the fritillaria thunbergii, sappan wood, aster, white scutellaria, platycodon, cyperus rotundus, knotweed, bidens pilosa, and liquorice into powder, and add inula flower;

[0009] (2) adding ethanol solution, soaking, ultrasonic extraction, filtering, removing ethanol from the filtrate using a rotary evaporator, freeze-drying, and crushing into powder to obtain the product.

[0010] Furthermore, the volume concentration of the ethanol solution in step (2) is 70-90%, and the amount used each time is 8-12 times the weight of the raw material drug.

[0011] Furthermore, in step (2), the soaking temperature is room temperature and the soaking time is 30-60 minutes.

[0012] Furthermore, in step (2), ultrasonic extraction is performed multiple times, each time for 90-120 minutes.

[0013] Furthermore, the evaporation temperature of the rotary evaporator in step (2) is 48-55°C.

[0014] On the other hand, the technical solution of the present invention is to provide an application of the traditional Chinese medicine composition in the preparation of a drug for treating lung cancer.

[0015] On the other hand, the technical solution of the present invention is to provide a traditional Chinese medicine preparation for treating lung cancer, which is prepared by adding auxiliary materials required for preparation molding to the traditional Chinese medicine composition and following a conventional method for preparing the preparation.

[0016] Furthermore, the Chinese medicine preparation is in the form of pills, capsules, tablets, granules or oral liquid.

[0017] Pharmacological effects of each raw material drug in the present invention:

[0018] Fritillaria thunbergii is cold in nature and bitter in taste; it enters the lung and heart meridians. Its main functions are to clear away heat and resolve phlegm, dissipate stagnation and detoxify, and it is specialized in treating the lungs.

[0019] Su Mu (Sappan Wood) is neutral in nature, sweet and salty in flavor, and enters the Heart, Liver, and Spleen meridians. Its main functions are promoting blood circulation, dispersing blood stasis, reducing swelling, and relieving pain.

[0020] Aster root is warm in nature, pungent and bitter in flavor, and enters the Lung meridian. Its main effects: moistening the lungs, lowering qi, eliminating phlegm and relieving cough. It enters both the Qi and Blood components of the Lung meridian, promoting the flow of Qi and blood through the Lung meridian, and treating hematemesis, hemoptysis, and hemoptysis.

[0021] Baiqian is slightly warm in nature, pungent and bitter in taste, and enters the Lung meridian. Its main functions are to lower Qi, eliminate phlegm, and relieve cough; it can purge the lungs and lower Qi; it can eliminate phlegm and relieve cough.

[0022] Platycodon grandiflorum is neutral in nature, bitter and pungent in taste, and enters the Lung meridian. Its main effects: Bitterness dispels obstructions, promotes the flow of lung qi, removes stagnant qi, breaks up blood, and eliminates accumulated phlegm.

[0023] Inula flower is slightly warm in nature, with a bitter, pungent, and salty flavor. It enters the lung, spleen, stomach, and large intestine meridians. Its main effects include lowering qi, eliminating phlegm, promoting diuresis, and stopping vomiting.

[0024] Lithospermum officinale is neutral in nature and sweet in taste. It enters the lung and liver meridians. Its main functions are clearing away heat and detoxification, anti-cancer, and hemostasis.

[0025] Polygonum cuspidatum is slightly cold in nature and slightly bitter in taste; it enters the liver, gallbladder, and lung meridians. Its main functions are clearing away heat and detoxifying, dispersing blood stasis and relieving pain, and relieving cough and phlegm.

[0026] Bidens pilosa is neutral in nature, bitter in taste, and non-toxic; it enters the liver, lung, and large intestine meridians. Its main effects are clearing heat, detoxifying, dispersing blood stasis, and reducing swelling.

[0027] Licorice root is neutral in nature and sweet in flavor. It enters the Heart, Lung, Spleen, and Stomach meridians. Its main functions are to tonify the spleen and replenish Qi, clear away heat and detoxify, dissipate phlegm and relieve cough, relieve pain, and harmonize various herbs.

[0028] Zhu Danxi, a famous physician in the Yuan Dynasty, proposed in "Danxi Xinfa": "If there are lumps in the upper, middle and lower parts of the body, they are mostly caused by phlegm." When the lung qi fails to ascend and descend properly, the movement of qi in the meridians is blocked, and the distribution of body fluids is abnormal, then the lumps will become phlegm; when qi flows, blood flows, and when qi stagnates, blood stagnates. When blood cannot flow normally, it stays and becomes blood stasis. Phlegm and blood stasis fight against each other and accumulate in the lungs to form lumps. This points out the relationship between phlegm and blood stasis and tumor occurrence, suggesting that phlegm and blood stasis blocking the lungs is the key pathogenesis of lung cancer, and that resolving phlegm and removing blood stasis is an important treatment method in the current clinical diagnosis and treatment of lung cancer in Traditional Chinese Medicine. The present invention adopts Fritillaria thunbergii and Sappan wood as monarch drugs, which can clear away heat and resolve phlegm, promote blood circulation and remove blood stasis, and relieve swelling and relieve pain; Radix Phellodendri and Radix Aster, which both enter the qi division of the lung meridian to purge the lung and descend qi, relieve phlegm and relieve cough, and enter the blood division to dredge the qi and blood of the lung meridian, and treat hematemesis, hemoptysis, and hemoptysis; Platycodon grandiflorum and Flos Inulae rise and fall respectively, and qi rises to dispel evil and expel it, and qi descends to descend water, so they can dredge and promote lung qi, descend qi and relieve phlegm, and the four are together ministerial drugs to enhance the monarch drug's effect of resolving phlegm, removing blood stasis and relieving pain; Ligusticum chuanxiong clears heat and detoxicates, has anti-cancer and hemostasis, Polygonum cuspidatum clears heat and detoxicates to relieve pain, and Bidens pilosa clears heat and detoxicates, promotes blood circulation and relieves swelling, and helps the monarch and minister drugs to resolve phlegm, remove blood stasis and relieve pain as adjuvants; Radix Glycyrrhizae is a harmonizing agent, and the combined prescription plays the effect of resolving phlegm, removing blood stasis and relieving pain.

[0029] Beneficial effects

[0030] The traditional Chinese medicine composition is developed based on the phlegm and blood stasis blocking lung pathogenesis of lung cancer in TCM diagnosis and treatment, combined with the phlegm and blood stasis removing treatment method and laboratory research, and has the effects of removing phlegm and blood stasis and inhibiting tumor growth, and has a good effect on the treatment of lung cancer. The experiment proves that the traditional Chinese medicine composition significantly inhibits lung cancer cell proliferation in vitro, improves the lung cancer pathological process of lung cancer mice in vivo, and is a great innovation of lung cancer drugs. BRIEF DESCRIPTION OF DRAWINGS

[0031] Figure 1 The influence of the traditional Chinese medicine composition on the epigenetic state of lung cancer cells.

[0032] Figure 2 The growth inhibition rate of the traditional Chinese medicine composition on lung cancer cells. DETAILED DESCRIPTION

[0033] The specific embodiments of the present application are further described in detail below in combination with examples.

[0034] Example 1

[0035] A traditional Chinese medicine composition for treating lung cancer comprises the following raw medicinal materials in parts by weight: Fritillaria thunbergii 5 parts, Sappan wood 5 parts, Aster 5 parts, Rehmannia glutinosa 5 parts, Platycodon grandiflorum 5 parts, Anemone vitifolia 5 parts, Bupleurum chinense 22 parts, Giant knotweed 9 parts, Bidens pilosa 22 parts, and Licorice 2 parts.

[0036] The preparation method is as follows:

[0037] (1) After Fritillaria thunbergii, Sappan wood, Aster, Rehmannia glutinosa, Platycodon grandiflorum, Anemone vitifolia, Bupleurum chinense, Giant knotweed, Bidens pilosa, and Licorice are powdered, Anemone vitifolia is added, and the mixture is placed in a beaker;

[0038] (2) 10 times the weight of the above raw medicinal materials and 70% (v / v) ethanol solution are added; the beaker is covered with plastic wrap, and pre-soaked for 30 minutes at room temperature; then ultrasonic extraction is performed every 30 minutes, and the mixture is stirred and shaken twice with a glass rod, and the ultrasonic extraction is performed for 90 minutes; then the plastic wrap is removed, the beaker is covered with nine layers of gauze, the ultrasonic liquid is poured out, and three layers of filter paper are used to perform reduced pressure filtration to obtain the filtrate; the remaining filter residue is added with 10 times the weight of the original medicinal materials and 70% (v / v) ethanol solution, and second ultrasonic extraction is performed; the filtrate is combined; the ethanol in the filtrate is removed by evaporation at 50°C using a rotary evaporator; and finally, the filtrate is dried into a solid state by a conventional freeze-drying method, and is pulverized into powder, which is stored at -20°C.

[0039] Example 2

[0040] A traditional Chinese medicine composition for treating lung cancer comprises the following raw medicinal materials in parts by weight: Fritillaria thunbergii 6 parts, Sappan wood 6 parts, Aster 6 parts, Rehmannia glutinosa 6 parts, Platycodon grandiflorum 6 parts, Anemone vitifolia 6 parts, Bupleurum chinense 23 parts, Giant knotweed 10 parts, Bidens pilosa 23 parts, and Licorice 3 parts.

[0041] Its preparation method is:

[0042] (1) Grind Fritillaria thunbergii, Aster tataricus, Radix Platycodonis ... Sappanwood, Cyperus rotundus, Polygonum cuspidatum, Herba Bidens pilosa, and Radix Glycyrrhizae into powder, add Inula japonica, and place in a beaker;

[0043] (2) Add 8 times the weight of the above-mentioned raw material drug and an 80% (v / v) ethanol solution; cover the beaker mouth with plastic wrap, pre-soak for 40 minutes at room temperature; then use an ultrasonic device to perform ultrasonication, stir and shake twice with a glass rod every 30 minutes, and ultrasonically extract for 100 minutes; then remove the plastic wrap, cover the beaker mouth with nine layers of gauze, pour out the ultrasonic liquid, use three layers of filter paper to filter under reduced pressure, and retain the filtrate; add 8 times the weight of the original medicinal material and an 80% (v / v) ethanol solution to the remaining filter residue, perform a second ultrasonic extraction, filter, and combine the two filtrates; and use a rotary evaporator to evaporate and remove the ethanol in the filtrate at 48°C, and finally use a conventional freeze-drying method to dry it into a solid state, crush it into powder, and store it at -20°C.

[0044] Example 3

[0045] A traditional Chinese medicine composition for treating lung cancer comprises the following raw materials in parts by weight: 7 parts of thunbergia thunbergii, 7 parts of sappan wood, 7 parts of aster, 7 parts of scutellaria baicalensis, 7 parts of platycodon, 7 parts of inula flower, 24 parts of cyperus rotundus, 11 parts of knotweed, 24 parts of bidens pilosa, and 4 parts of liquorice.

[0046] Its preparation method is:

[0047] (1) Grind Fritillaria thunbergii, Aster tataricus, Radix Platycodonis ... Sappanwood, Cyperus rotundus, Polygonum cuspidatum, Herba Bidens pilosa, and Radix Glycyrrhizae into powder, add Inula japonica, and place in a beaker;

[0048] (2) Add 12 times the weight of the above-mentioned raw material drug and a 90% (v / v) ethanol solution; cover the beaker mouth with plastic wrap, pre-soak for 50 minutes at room temperature; then use an ultrasonic device to perform ultrasonication, stir and shake twice with a glass rod every 30 minutes, and ultrasonically extract for 110 minutes; then remove the plastic wrap, cover the beaker mouth with nine layers of gauze, pour out the ultrasonic liquid, use three layers of filter paper to filter under reduced pressure, and retain the filtrate; add 12 times the weight of the original medicinal material and a 90% (v / v) ethanol solution to the remaining filter residue, perform a second ultrasonic extraction, filter, and combine the two filtrates; and use a rotary evaporator to evaporate and remove the ethanol in the filtrate at 55°C, and finally use a conventional freeze-drying method to dry it into a solid state, crush it into powder, and store it at -20°C.

[0049] Example 4

[0050] A traditional Chinese medicine composition for treating lung cancer, comprising the following raw medicinal materials in parts by weight: Fritillaria thunbergii 8 parts, Sappan 8 parts, Radix Pulsatillae 8 parts, Rehmannia glutinosa 8 parts, Platycodon grandiflorum 8 parts, Inula racemosa 8 parts, Sabina vulgaris 25 parts, Giant Knotweed 12 parts, Bidens pilosa 25 parts, and Glycyrrhiza 5 parts.

[0051] The preparation method is as follows:

[0052] (1) After Fritillaria thunbergii, Radix Pulsatillae, Rehmannia glutinosa, Platycodon grandiflorum, Sappan, Sabina vulgaris, Giant Knotweed, Bidens pilosa, and Glycyrrhiza are powdered, Inula racemosa is added and placed in a beaker;

[0053] (2) 10 times the weight of the above raw medicinal materials and 70% (v / v) ethanol solution are added; the beaker opening is covered with cling film, and pre-soaked for 60 minutes at room temperature; then ultrasonic extraction is performed every 30 minutes, and the beaker is stirred and shaken twice with a glass rod; the ultrasonic extraction is performed for 120 minutes; then the cling film is removed, the beaker opening is covered with nine layers of gauze, and the ultrasonic liquid is poured out; three layers of filter paper are used to perform reduced pressure filtration, and the filtrate is retained; the remaining filter residue is added with 10 times the weight of the original medicinal materials and 70% (v / v) ethanol solution, and second ultrasonic extraction is performed; the filtrate is combined; the ethanol in the filtrate is evaporated at 50°C using a rotary evaporator; finally, the solid state is dried by a conventional freeze-drying method, and the powder is obtained by pulverization; and the powder is stored at -20°C.

[0054] Control example

[0055] A control traditional Chinese medicine composition, comprising the following raw medicinal materials in parts by weight: Sinopodophyllum emodi 1 part, Saposhnikovia divaricata 1 part, Artemisia annua 1 part, Radix Angelicae Dahuricae 1 part, Chrysanthemum indicum 1 part, Rabdosia rubescens 1 part, Artemisia rupestris 1 part, Citrus aurantium 1 part, and Cuscuta chinensis 1 part.

[0056] The preparation method is as follows:

[0057] (1) After Sinopodophyllum emodi, Saposhnikovia divaricata, Artemisia annua, Radix Angelicae Dahuricae, Chrysanthemum indicum, Rabdosia rubescens, Artemisia rupestris, Citrus aurantium, and Cuscuta chinensis are powdered, the beaker is placed in a beaker;

[0058] (2) 10 times the weight of the above raw medicinal materials and 70% (v / v) ethanol solution are added; the beaker opening is covered with cling film, and pre-soaked for 30 minutes at room temperature; then ultrasonic extraction is performed every 30 minutes, and the beaker is stirred and shaken twice with a glass rod; the ultrasonic extraction is performed for 90 minutes; then the cling film is removed, the beaker opening is covered with nine layers of gauze, and the ultrasonic liquid is poured out; three layers of filter paper are used to perform reduced pressure filtration, and the filtrate is retained; the remaining filter residue is added with 10 times the weight of the original medicinal materials and 70% (v / v) ethanol solution, and second ultrasonic extraction is performed; the filtrate is combined; the ethanol in the filtrate is evaporated at 50°C using a rotary evaporator; finally, the solid state is dried by a conventional freeze-drying method, and the powder is obtained by pulverization; and the powder is stored at -20°C.

[0059] 1Material

[0060] 1.1 Cell lines

[0061] Human lung adenocarcinoma cell line A549 cells and mouse lung cancer cell line LLC cells were purchased from the Cell Center of the Chinese Academy of Medical Sciences.

[0062] 1.2 Animals

[0063] C57BL / 6 mice, 4-6 weeks old, 14-16 g, were purchased from Spefox (Beijing) Biotechnology Co., Ltd.

[0064] 2 Methods

[0065] 2.1 Cell culture

[0066] A549 cells were grown in DMEM complete medium containing 10% (v / v) fetal bovine serum in a 37° C., 5% CO 2 cell culture incubator and subcultured every 2-3 days.

[0067] 2.2 Mouse husbandry

[0068] The animal room is SPF-grade constant temperature, the feeding room is exposed to ultraviolet light once a day, and the personnel entering and leaving are strictly controlled and must change sterile work clothes before each entry and exit; the mice are fed with vacuum-packed irradiated feed, and corn cobs sterilized by irradiation Co60 are used as mouse bedding. Distilled water is used as drinking water. The feed, bedding and drinking water are changed every 3 days. There is no restriction on diet, and the feeding environment is kept quiet, clean and comfortable.

[0069] 2.3 Observation of the epigenetic status of A549 cells in the normal group and the Chinese medicine combination

[0070] When A549 cells were cultured to cover more than 80% of the culture dish area, they were digested with trypsin solution (0.02% ethylenediaminetetraacetic acid disodium salt (EDTA) solution and 0.25% trypsin solution in a volume ratio of 1:1) and the cell pellets were collected and resuspended in DMEM complete medium. 5 The cells were seeded at a density of 100 μL / well in a 12-well flat-bottom cell culture plate. The cells were incubated at 37°C, 5% CO2 in an incubator for 24 h. 2 mL of new DMEM complete medium was replaced in each well. A blank group, an experimental group (the Chinese medicine composition of Example 1), and a control group (the Chinese medicine composition of the reference example) were set up. The blank group was directly added with DMEM complete medium at 20 μL / well. The experimental group was screened according to the IC values ​​of 2.4. 30(drug concentration when the inhibition rate reaches 30%), DMEM complete medium containing 36.7 mg / mL of the Chinese medicine composition of the present invention was added at 20 μL / well, so that the working concentration of the Chinese medicine composition was 367 μg / mL. The control group was added with DMEM complete medium containing the control Chinese medicine composition, and the dosage was consistent with that of the experimental group; the cells were cultured in an incubator at 37°C and 5% CO2 for 48 hours, observed under an inverted microscope, and photographed and recorded.

[0071] 2.4 MTT assay to determine the dose-effect relationship of the inhibitory effect of the Chinese herbal medicine composition on A549 cell growth

[0072] A549 cells were seeded at 3000 cells / 100 μL / well in 96-well flat-bottom cell culture plates and cultured at 37°C, 5% CO2 for 24 h. At this time, the cells were evenly dispersed, completely adhered to the wall and in good condition. 100 μL of new DMEM complete medium was replaced and added. A blank group and 5 groups of Chinese medicine compositions were set up, with 6 replicates in each group. 10 μL of DMEM complete medium was added to the blank group, and 10 μL of Chinese medicine extract solutions with concentrations of 312, 625, 1250, 2500, and 5000 μg / mL (the Chinese medicine composition in Example 1, the solvent was DMEM complete medium) were added to the 5 groups of Chinese medicine compositions, so that the working concentrations of the Chinese medicine compositions were 31.2, 62.5, 125, 250, and 500 μg / mL. Continue to culture in a 37°C, 5% CO2 incubator for 48 hours, aspirate the culture medium, add 100 μL of 0.5 mg / mL thiazolyl blue (MTT) solution to each well, continue to culture in a 37°C, 5% CO2 incubator for 4 hours, carefully aspirate the supernatant, add 100 μL of dimethyl sulfoxide (DMSO), and incubate in a 37°C, 5% CO2 incubator in the dark on a shaking table for 10 minutes to completely dissolve the purple crystalline precipitate. Use a microplate reader to measure the corresponding absorbance (OD value) of each well at wavelengths of 490 nm and 570 nm. According to the formula:

[0073] Inhibition rate (%) = (1-average A 490 / 570值 / Average A of blank wells 490 / 570值 )×100%

[0074] Chinese medicine composition IC 30 Calculation of IC values: Repeat the above experiment three times, import the obtained inhibition rate into Excel, insert the scatter plot to generate a trend line, and calculate IC according to the trend line generation formula. 30 value.

[0075] 2.5 Effects of Chinese herbal medicine combination on LLC lung cancer cell transplanted mice

[0076] LLC cells in the logarithmic growth phase were obtained, digested with trypsin, centrifuged, and precipitated. The cell pellet was resuspended in PBS buffer (phosphate buffer) to a concentration of 1×10 7 / mL. 6 mice were respectively injected with 0.15mL of PBS buffer solution subcutaneously in the right anterior armpit and set as the blank group. The other mice were injected with 0.15mL of LLC cell suspension subcutaneously in the right anterior armpit to construct an LLC lung cancer transplant mouse model. After 7 days, the modeling was successful when a tumor the size of a mung bean was observed or touched under the right armpit of the modeling mouse. After successful modeling, the modeling mice were divided into a model group, a Chinese medicine composition experimental group (Chinese medicine composition of Example 1), a Chinese medicine composition control group (reference example Chinese medicine composition), and a cisplatin group, with 6 mice in each group. The Chinese medicine composition experimental group and the Chinese medicine composition control group were gavaged once a day, and the cisplatin group was intraperitoneally injected once every other day. Dosage Calculation: Based on the body surface area method, the equivalent dose of the Chinese herbal medicine in mice was 17.55 g / kg, corresponding to a powdered dose of 3.91 g / kg of the Chinese herbal medicine composition, administered in a volume of 0.1 mL / 10 g (the experimental and control groups received the same dose). The cisplatin group received an intraperitoneal injection of 1 mg / kg (both solvents were sterile saline), at a volume of 0.1 mL / 10 g. The model and blank groups were gavage-administered with an equal volume of saline as controls. Tumor volume was measured every two days. After two weeks of treatment, mice were sacrificed by cervical dislocation, the tumors were removed, and the final tumor volume was measured and the tumor inhibition rate was calculated.

[0077] Tumor volume calculation: The longest diameter L (mm) and the maximum transverse diameter W (mm) of the subcutaneous tumor of the mouse were measured with an electronic vernier caliper, and the tumor volume V (mm) was calculated according to the Lee method. 3 )=L×W 2 / 2.

[0078] Tumor inhibition rate of drugs: Tumor inhibition rate = (1-average tumor volume of drug-treated group / average tumor volume of model group) × 100%

[0079] 2.6 Effect of Chinese herbal medicine combination on body weight of LLC lung cancer transplanted mice

[0080] After administration, the animals were observed every two days for general condition and weighed using a small animal electronic scale. Observations included dietary status, activity status, hair sparseness and glossiness, and perception of the external environment.

[0081] 2.7 Statistical methods

[0082] The quantitative data were analyzed using one-way analysis of variance using GraphPad Prism 9.5.0 software, and the results were subjected to correlation analysis and curve fitting.

[0083] 3 Experimental results

[0084] 3.1 The influence of the traditional Chinese medicine composition on the growth and morphology of A549 lung cancer cells

[0085] Under the inverted microscope, the cells in the blank group adhered to the wall and grew, the cells were spindle-shaped, smooth, and the cell boundaries were clear, and some cells had pseudopods. Compared with the cells in the blank group, a small amount of cells in the control group grew pseudopods, but the overall cell state did not change significantly. In the traditional Chinese medicine composition group (the experimental group) in the application, the cell cytoplasm was extremely rough, filled with a large amount of granular material, and as the drug concentration increased, a large range of necrotic blank fields appeared in the cells (see Figure 1 ). This shows that the tumor cell state after treatment with the traditional Chinese medicine composition in the application is poor, and even a large area of death.

[0086] 3.2 The influence of the traditional Chinese medicine composition on the growth of A549 lung cancer cells (inhibition rate)

[0087] The traditional Chinese medicine composition can inhibit the growth of A549 lung cancer cells, and as the concentration of the traditional Chinese medicine composition increases, the growth inhibition of lung cancer cells shows a clear dose-effect dependence (see Table 1 and Figure 2 ), and the IC 30 value is 367 μg / mL.

[0088] Table 1: Relationship table between the growth inhibition rate of A549 lung cancer cells and the drug concentration of the traditional Chinese medicine composition (unit: μg / mL)

[0089]

[0090]

[0091] 3.3 The inhibitory effect of the traditional Chinese medicine composition on the proliferation of LLC lung cancer transplanted tumor mice

[0092] After 9 days of inoculation of LLC cells, 100% of the mice grew tumor bodies the size of soybeans subcutaneously, and the tumor bodies grew slowly at the beginning of inoculation. After about 12 days of inoculation, the tumor bodies of the model group mice grew rapidly. At the end of the experiment, compared with the model group mice, cisplatin significantly inhibited the tumor growth of the tumor mice; compared with the cisplatin group mice, the traditional Chinese medicine composition control group did not significantly inhibit the tumor growth, while the traditional Chinese medicine composition experimental group significantly inhibited the tumor growth of the tumor mice, although it was slightly weaker than the inhibitory effect of cisplatin on tumor growth, but there was no significant difference; compared with the traditional Chinese medicine composition control group, the traditional Chinese medicine composition in the application had a significant inhibitory effect on tumor growth (see Table 2).

[0093] Table 2: Influence of the traditional Chinese medicine composition on the tumor volume of lung cancer transplanted tumor mice (mm 3 )

[0094]

[0095] * P<0.01 vs. model group; P<0.05 vs. cisplatin group; $ P<0.01 vs. traditional Chinese medicine composition control group.

[0096] 3.4 Effects of the traditional Chinese medicine composition on the general conditions and body weights of LLC lung cancer transplanted mice

[0097] No animal died during the whole experiment. Compared with the blank group, the model group mice had dry and dull hair, and sparse hair, and the mice were in a spirit of debauchery, and their eyes were dull, and they had a behavior of arching their backs. Although the mice showed general emaciation, the body weight of the model group mice was significantly increased due to the rapid growth of the tumor in the later stage. The traditional Chinese medicine composition experimental group had bright and smooth hair, and their eyes were bright, and they were active and had normal diet, and their body weight did not change significantly. The traditional Chinese medicine composition control group mice had sparse hair, and their eyes were dull, and their body weight increased due to the increase in tumor volume, but they showed general emaciation. The cisplatin group mice had dry and dull hair, and sparse hair, and their spirit was debauched, and their diet was reduced, and their body weight was significantly reduced (see Table 3). These results showed that, compared with the model group, cisplatin did not improve the survival state and quality of life of lung cancer mice; compared with the cisplatin group, the traditional Chinese medicine composition could improve the survival state and emaciation of lung cancer mice, and improve the quality of life, while the traditional Chinese medicine composition control group had no effect on the quality of life of lung cancer mice; at the same time, compared with the traditional Chinese medicine composition control group, the traditional Chinese medicine composition also significantly improved the quality of life and emaciation of lung cancer mice.

[0098] Table 3: Effects of the traditional Chinese medicine composition on the body weights of lung cancer transplanted mice (N=6)

[0099]

[0100] # P<0.01 vs. blank group; * P<0.01 vs. model group; P<0.01 vs. cisplatin group; $ P<0.01 vs. traditional Chinese medicine composition control group.

[0101] 4 Conclusion

[0102] 4.1 The anti-lung cancer traditional Chinese medicine composition showed good anti-lung cancer effect in vitro cell experiments, and had a dose-effect relationship.

[0103] 4.2 The anti-lung cancer traditional Chinese medicine composition could significantly inhibit the growth of LLC lung cancer cell transplanted tumors in mice in a dose-dependent manner, and could reduce the effects of lung cancer on spirit debauchery, activity reduction, general emaciation, diet, etc.; at the same time, it played a good effect in inhibiting tumor growth.

[0104] 4.3 Anti-lung cancer traditional Chinese medicine composition has good anti-lung cancer effect, and has small toxic and side effects, is the innovation of lung cancer drugs, and has significant clinical significance.

Claims

1. A Chinese medicine composition for treating lung cancer, characterized in that: The invention is composed of the following raw materials in parts by weight: 5-8 parts of thunbergii bulbs, 5-8 parts of sappan wood, 5-8 parts of aster, 5-8 parts of scutellaria baicalensis, 5-8 parts of platycodon, 5-8 parts of inula flower, 22-25 parts of cyperus rotundus, 9-12 parts of knotweed, 22-25 parts of bidens pilosa, and 2-5 parts of liquorice.

2. The Chinese medicine composition according to claim 1, characterized in that The invention is composed of the following raw materials in parts by weight: 5 parts of thunbergii bulbs, 5 parts of sappan wood, 5 parts of aster, 5 parts of scutellaria baicalensis, 5 parts of platycodon, 5 parts of inula flower, 22 parts of cyperus rotundus, 9 parts of knotweed, 22 parts of bidens pilosa, and 2 parts of liquorice.

3. A method for preparing the Chinese medicine composition according to claim 1 or 2, characterized in that: The following steps are involved: (1) Grind Fritillaria thunbergii, Sappan wood, Aster tataricus, Radix Bletillae, Platycodon grandiflorum, Cyperus rotundus, Polygonum cuspidatum, Bidens pilosa, and Licorice root into powder, and add Inula japonica flower; (2) Add ethanol solution, soak, perform ultrasonic extraction, filter, use a rotary evaporator to remove ethanol in the filtrate, freeze-dry, and grind into powder to obtain.

4. The preparation method according to claim 3, characterized in that The volume concentration of the ethanol solution in step (2) is 70-90%, and the amount used each time is 8-12 times the weight of the raw material drug.

5. The preparation method according to claim 3, characterized in that The soaking temperature in step (2) is room temperature and the soaking time is 30-60 minutes.

6. The preparation method according to claim 3, characterized in that In step (2), ultrasonic extraction is performed multiple times, each time for 90-120 minutes.

7. The preparation method according to claim 3, characterized in that The evaporation temperature of the rotary evaporator in step (2) is 48-55°C.

8. Use of the traditional Chinese medicine composition according to claim 1 or 2 in preparing medicine for treating lung cancer.

9. A Chinese medicine preparation for treating lung cancer, characterized in that: The auxiliary materials required for preparation molding are added to the traditional Chinese medicine composition according to claim 1 or 2, and the preparation is made according to a conventional method for preparing preparations.

10. The Chinese medicine preparation according to claim 9, characterized in that The traditional Chinese medicine preparation is in the form of pills, capsules, tablets, granules or oral liquids.

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