A Chinese medicine preparation for lung cancer and its preparation method

Abstract

The present invention provides a Chinese medicine preparation for lung cancer and a preparation method thereof, and relates to the technical field of Chinese medicine pharmaceutical manufacturing. The Chinese medicine preparation is composed of cat's claw, ginseng, aconite, rhubarb, platycodon, peucedanum, asarum, peach kernel, calamus, gypsum, cinnamon bark, dried ginger, and liquorice, and has the effects of strengthening the body and eliminating evil, promoting yang and purging turbidity, resolving phlegm and removing blood stasis, and dispersing nodules and eliminating accumulation. The Chinese medicine preparation of the present invention has a significant effect of inhibiting proliferation of Lewis lung cancer cells and A549 cells in mice, and has a significant effect of inhibiting tumor growth, improving tumor tissue pathological morphology, and improving the quality of life of mice in Lewis lung cancer model mice. Clinical application data show that the Chinese medicine preparation of the present invention can significantly improve the clinical discomfort of lung cancer patients, improve the immunity and quality of life of patients, and prolong the survival of patients. The Chinese medicine preparation provided by the present invention provides a new idea and treatment method for the treatment of lung cancer.

Description

Technical Field

[0001] The present invention relates to the technical field of Chinese medicine preparations, and in particular to a Chinese medicine preparation for lung cancer and a preparation method thereof. Background Art

[0002] Lung cancer is composed of a group of molecular and histologically heterogeneous subtypes, which are divided into small cell lung cancer and non-small cell lung cancer. It is one of the most common malignant tumors in clinical practice. It is a malignant tumor that originates from the epidermal cells of the bronchial mucosal epithelium and alveolar wall at all levels and has the characteristics of unlimited proliferation. Its morbidity and mortality rate are among the highest among all malignant tumors in the world. It is a great hidden danger to people's life safety and physical health, and has long become a difficulty and hot spot for clinical researchers today. The main treatments for lung cancer currently include surgery, radiotherapy and chemotherapy, targeted therapy, and immunotherapy. Due to the limitations of surgical treatment and the cytotoxicity and tolerance of radiotherapy and chemotherapy, the quality of life of lung cancer patients is reduced, affecting the efficacy of treatment. In the past 20 years, with the emergence and discovery of various targeted therapies, lung cancer has made great progress in research, screening and personalized treatment. Although the above methods can specifically kill tumor cells, reduce toxic effects, prolong the patient's progression-free survival, and improve the quality of life, most patients with effective treatment will also face problems such as drug resistance, and then tumor recurrence or metastasis will occur, which will lead to a bottleneck in the treatment of lung cancer. Therefore, the development of new anti-lung cancer drugs is of great significance.

[0003] Due to its multi-target and multi-pathway characteristics, many Chinese medicine scholars have developed many Chinese medicine compound prescriptions for tumor treatment in recent years. Studies have shown that Chinese medicine has significant advantages in treating lung cancer, and has certain efficacy and advantages in enhancing patients' immune function, reducing the toxic and side effects of radiotherapy and chemotherapy, inhibiting lung cancer invasion and metastasis, and improving patients' quality of life. In addition, the country has vigorously advocated the research and development of Chinese medicine in recent years. Therefore, in line with the mission of inheriting and innovating Chinese medicine, it is urgent to find safe and effective Chinese medicine preparations for the treatment of lung cancer. Summary of the invention

[0004] 1. Technical issues to be resolved

[0005] In view of the deficiencies of the prior art, the present invention provides a traditional Chinese medicine preparation for lung cancer and a preparation method thereof.

[0006] (II) Technical solution

[0007] To achieve the above objectives, the present invention is implemented through the following technical solutions:

[0008] The invention provides a traditional Chinese medicine preparation for lung cancer. The traditional Chinese medicine preparation comprises the following raw materials in parts by weight: 5-25 parts of cat's claw, 5-25 parts of ginseng, 1-15 parts of aconite, 1-15 parts of rhubarb, 1-10 parts of platycodon, 1-10 parts of peucedanum, 1-10 parts of asarum, 1-10 parts of peach kernel, 1-10 parts of calamus, 1-10 parts of gypsum, 1-10 parts of cinnamon, 1-10 parts of dried ginger and 5-25 parts of liquorice.

[0009] Preferably, the Chinese medicine preparation comprises the following raw materials in parts by weight: 10-20 parts of cat's claw, 10-20 parts of ginseng, 5-10 parts of aconite, 5-10 parts of rhubarb, 1-5 parts of platycodon, 1-5 parts of peucedanum, 1-5 parts of asarum, 1-5 parts of peach kernel, 1-5 parts of calamus, 1-5 parts of gypsum, 1-5 parts of cinnamon, 1-5 parts of dried ginger, and 10-20 parts of licorice.

[0010] Preferably, the Chinese medicine preparation comprises the following raw materials in parts by weight: 10 parts of cat's claw, 10 parts of ginseng, 7 parts of aconite, 7 parts of rhubarb, 3 parts of platycodon, 3 parts of peucedanum, 3 parts of asarum, 3 parts of peach kernel, 3 parts of calamus, 3 parts of gypsum, 3 parts of cinnamon, 5 parts of dried ginger, and 10 parts of licorice.

[0011] The Chinese medicine preparation for lung cancer provided by the present invention is any oral dosage form acceptable to human body, including oral liquid, granules, powders, decoctions, pills and freeze-dried powders.

[0012] The Chinese medicine preparation is prepared into pills according to the following method: weigh the raw materials according to the above-mentioned weight portions, wash, dry, and grind into powder, mix them thoroughly and evenly, and then mix them with heated honey in a weight ratio of 100:60-70 to form honey pills with a particle size of 4-6mm and a weight of 1-3g / 30 pills, thereby obtaining the Chinese medicine pills for lung cancer.

[0013] The Chinese medicine preparation of the present invention is obtained by reasonable compatibility according to the principle of TCM syndrome differentiation and treatment. In the prescription, cat's claw and ginseng are used as the main ingredients, which can eliminate and tonify, dispel evil and strengthen the body to disperse knots and eliminate accumulation; cat's claw is sweet, pungent and warm, enters the liver and lung meridians, and can dissolve phlegm and dissolve knots, detoxify and reduce swelling; ginseng is sweet and slightly bitter, warm, enters the spleen meridian and lung meridian, can greatly replenish vital energy, consolidate and produce body fluid, and calm the mind. According to the "Ben Jing", it can mainly replenish the five internal organs, calm the spirit, stop palpitations, eliminate evil spirits, improve eyesight, and improve happiness and intelligence.

[0014] With aconite and rhubarb as auxiliary drugs, it can be used together with cold and heat to promote the flow of yang, eliminate turbidity and break up various accumulations; aconite is pungent, sweet, hot and poisonous, and enters the heart, kidney and spleen meridians, restores yang and rescues adverse conditions, replenishes fire and assists yang, dispels cold and removes dampness, and is mainly used to treat "wind-cold cough and adverse evil qi, warms the middle, cold and dampness, cramps and knee pain, inability to walk, and breaking up hard accumulations and blood masses" (Ben Jing), and "treats three yin meridian syndromes and yin poison typhoid fever" (Compendium of Materia Medica); rhubarb is bitter and cold, and enters the stomach, large intestine and liver meridians, and can "remove blood stasis, blood stasis, cold and heat, break up accumulations of masses, retain fluid and old food, cleanse the intestines and stomach, remove the old and bring in the new, promote the flow of water and food, regulate the middle and digest food, and calm the five internal organs" (Ben Jing), and can also "benefit edema, break up phlegm and cold and heat accumulation" (Yao Xing Lun).

[0015] With asarum, cinnamon, dried ginger, gypsum and liquorice as adjuvants, warming and clearing together, helping the monarch and ministers to eliminate Yin and control evil heat. Asarum is pungent, warming and slightly toxic. It enters the lung, kidney, heart, liver, gallbladder and spleen meridians, and has the functions of warming and transforming cold phlegm, clearing the lung orifices, warming the meridians and relieving pain; cinnamon is pungent, sweet and hot. It enters the kidney, spleen, and bladder meridians, replenishes the original yang, warms the spleen and stomach, eliminates accumulated cold, and dredges the blood vessels. It also "treats all kinds of wind, replenishes the five kinds of fatigue and seven kinds of injuries, dredges the nine orifices, benefits the joints, benefits the essence, improves the eyesight, warms the waist and knees, breaks up lumps and tumors, eliminates blood stasis, treats rheumatism and joint contracture, repairs tendons and bones, and grows muscles ("Rihuazi Bencao")"; dried ginger is pungent and hot, and enters the spleen, stomach, kidney, heart, and lung meridians. It warms the middle and expels cold, restores yang and dredges the meridians. It treats cold pain in the heart and abdomen, vomiting and diarrhea, cold limbs and weak pulse, asthma and cough caused by cold drinking, etc. "Secret of Main Treatments" says: It dredges the heart qi, aids yang, removes deep coldness in the internal organs, and releases the coldness in all meridians; asarum, cinnamon, and dried ginger are used together, which is pungent and warm and aids yang, and is assisted by aconite to warm the meridians and dispel cold, and assists cat's claw to eliminate cold stagnation; gypsum is pungent and sweet, and is cold. Licorice root is sweet and neutral, and enters the spleen meridian, stomach meridian, and lung meridian. It “controls the cold and heat evil qi of the five internal organs and six organs” (Ben Jing), harmonizes the middle and relieves urgency, moistens the lungs, detoxifies, and harmonizes various medicines.

[0016] Peucedanum, Platycodon grandiflorum, Prunus persica, and Acorus calamus are used as the messengers to regulate qi and blood, resolve phlegm and remove blood stasis to regulate the lungs and eliminate accumulation. Peucedanum grandiflorum is bitter and pungent in taste, slightly cold in nature, and enters the lung, spleen, and liver meridians. It can dissipate wind-heat, lower qi, and eliminate phlegm. It can "cure all kinds of fatigue, lower all kinds of qi, stop coughs, and break up lumps; Platycodon grandiflorum is bitter and pungent in taste, neutral in nature, and enters the lung and stomach meridians, open and spread lung qi, and remove phlegm and pus. "Materia Medica" says: it can treat dysentery, break up blood, remove accumulated qi, eliminate accumulation, and phlegm. Li Gao believes that it can benefit the chest and diaphragm, treat throat qi congestion and pain, break up stagnant qi and accumulation, (Removes) lung wind-heat, clears the head and eyes, and benefits the orifices; peach kernel is bitter and sweet, and is neutral. It enters the heart meridian, liver meridian, and large intestine meridian, and can promote blood circulation and remove blood stasis, moisten dryness and lubricate the intestines. It "treats blood stasis, blood stasis, tumors, evil spirits, and kills small insects" (Ben Jing), and can "break up tumors, dredge meridians, and relieve pain" (Bie Lu); Acorus calamus tastes pungent and bitter, and is slightly warm in nature. It enters the heart, liver, and spleen meridians, and can open the orifices, resolve phlegm, regulate qi, promote blood circulation, dispel wind, and remove dampness.

[0017] (III) Beneficial effects

[0018] The present invention provides a Chinese medicine preparation for lung cancer and a preparation method thereof. The Chinese medicine preparation is obtained by rationally combining the principles of TCM syndrome differentiation and treatment, and has the effects of strengthening the body and eliminating evil, promoting yang and purging turbidity, resolving phlegm and removing blood stasis, and dispersing nodules and eliminating accumulation. The Chinese medicine preparation of the present invention has a significant effect of inhibiting proliferation of Lewis lung cancer cells and A549 cells in mice, and has a significant effect of inhibiting tumor growth, improving tumor tissue pathological morphology, and improving the quality of life of mice in Lewis lung cancer model mice. Clinical application data show that the Chinese medicine preparation of the present invention can significantly improve the clinical discomfort of lung cancer patients, improve the immunity and quality of life of patients, and prolong the survival of patients. The Chinese medicine preparation provided by the present invention provides a new idea and treatment method for the treatment of lung cancer. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 The weight changes and net body weight of mice in each group during the experiment were compared with those in the model group, *p<0.05.

[0020] Figure 2 The in vivo imaging system was used to observe the tumor growth in each group of mice (A) and the integrated optical density value (B); compared with the model group, *p<0.05, **p<0.01.

[0021] Figure 3 Figure 2 shows the pathological morphological changes of tumor tissues in mice of each group.

[0022] Figure 4 Comparison of cell clone formation (A) and clone formation rate (B) in each group; compared with the control group, *p<0.05, **p<0.01.

[0023] Figure 5 Comparison of the 24h migration status (A) and migration rate (B) of cells in each group; compared with the control group, *p<0.05, **p<0.01.

[0024] Figure 6 These are the changes in chest CT images of Case 1 before and 8 months after treatment; A: Tumor lesions in the left upper lobe before treatment; B: Tumor lesions in the left upper lobe after treatment were smaller than before; C, E, G: Before treatment, there were mostly small patchy shadows in the left lower lobe; D, F, H: After treatment, there were mostly small patchy shadows in the left lower lobe, and the range of the lesions was smaller than before.

[0025] Figure 7 The CT image changes of case 2 before treatment and 5 months after treatment; A: the tumor lesion in the left upper lobe before treatment; B: the tumor lesion in the left upper lobe after treatment was smaller than before.

[0026] Figure 8The chest CT image changes of case 3 before treatment and 3 months after treatment; A: the tumor lesion in the right lower lobe before treatment; B: the tumor lesion in the right lower lobe after treatment was significantly smaller than before; C: the calcified nodule in the right upper lobe before treatment; D: the calcified nodule in the right upper lobe after treatment was smaller than before. DETAILED DESCRIPTION

[0027] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention will be clearly and completely described below in combination with the embodiments of the present invention. Obviously, the described embodiments are 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.

[0028] Example 1

[0029] A traditional Chinese medicine preparation for lung cancer, comprising the following raw materials in parts by weight: 10 parts of cat's claw, 10 parts of ginseng, 7 parts of aconite, 7 parts of rhubarb, 3 parts of platycodon, 3 parts of peucedanum, 3 parts of asarum, 3 parts of peach kernel, 3 parts of calamus, 3 parts of gypsum, 3 parts of cinnamon, 5 parts of dried ginger, and 10 parts of liquorice.

[0030] The Chinese medicine preparation for lung cancer is any oral dosage form acceptable to human body, including oral liquid, granules, powder, decoction, pills and freeze-dried powder.

[0031] The Chinese medicine preparation is prepared into pills according to the following method: weigh the raw materials according to the above-mentioned weight portions, wash, dry, and powder them, mix them thoroughly and evenly, and then mix them with heated honey in a ratio of 100g:65g to make honey pills with a particle size of 4-6mm and a weight of 1-3g / 30 pills, thereby obtaining the Chinese medicine pills for lung cancer.

[0032] The dosage is: take with warm water 3 times a day after meals. On the first day, take 3 pills each time; on the second day, take 4 pills each time; on the third day, take 5 pills each time; increase the dosage daily in this way until the 8th day, take 10 pills each time. After that, maintain 10 pills each time and do not increase the dosage.

[0033] Example 2

[0034] The difference between this embodiment and embodiment 1 is that the Chinese medicine preparation includes the following raw materials in parts by weight: 5 parts of cat's claw, 5 parts of ginseng, 1 part of aconite, 1 part of rhubarb, 1 part of platycodon, 1 part of peucedanum, 1 part of asarum, 1 part of peach kernel, 1 part of calamus, 1 part of gypsum, 1 part of cinnamon, 1 part of dried ginger, and 5 parts of licorice.

[0035] Example 3

[0036] The difference between this embodiment and embodiment 1 is that the Chinese medicine preparation includes the following raw materials in parts by weight: 25 parts of cat's claw, 25 parts of ginseng, 15 parts of aconite, 15 parts of rhubarb, 10 parts of platycodon, 10 parts of peucedanum, 10 parts of asarum, 10 parts of peach kernel, 10 parts of calamus, 10 parts of gypsum, 10 parts of cinnamon, 10 parts of dried ginger, and 25 parts of licorice.

[0037] Example 4

[0038] The difference between this embodiment and embodiment 1 is that the Chinese medicine preparation includes the following raw materials in parts by weight: 22 parts of cat's claw, 20 parts of ginseng, 12 parts of aconite, 12 parts of rhubarb, 6 parts of platycodon, 8 parts of peucedanum, 6 parts of asarum, 8 parts of peach kernel, 6 parts of calamus, 6 parts of gypsum, 6 parts of cinnamon, 8 parts of dried ginger, and 22 parts of licorice.

[0039] Example 5

[0040] The difference between this embodiment and embodiment 1 is that the Chinese medicine preparation includes the following raw materials in parts by weight: 8 parts of cat's claw, 9 parts of ginseng, 4 parts of aconite, 3 parts of rhubarb, 2 parts of platycodon, 2 parts of peucedanum, 2 parts of asarum, 2 parts of peach kernel, 2 parts of calamus, 2 parts of gypsum, 2 parts of cinnamon, 2 parts of dried ginger, and 10 parts of licorice.

[0041] Example 6

[0042] The difference between this embodiment and embodiment 1 is that the Chinese medicine preparation includes the following raw materials in parts by weight: 10 parts of cat's claw, 10 parts of ginseng, 5 parts of aconite, 5 parts of rhubarb, 1 part of platycodon, 1 part of peucedanum, 1 part of asarum, 1 part of peach kernel, 1 part of calamus, 1 part of gypsum, 1 part of cinnamon, 1 part of dried ginger, and 10 parts of licorice.

[0043] Example 7

[0044] The difference between this embodiment and embodiment 1 is that the Chinese medicine preparation includes the following raw materials in parts by weight: 20 parts of cat's claw, 20 parts of ginseng, 10 parts of aconite, 10 parts of rhubarb, 5 parts of platycodon, 5 parts of peucedanum, 5 parts of asarum, 5 parts of peach kernel, 5 parts of calamus, 5 parts of gypsum, 5 parts of cinnamon, 5 parts of dried ginger, and 20 parts of licorice.

[0045] Example 8

[0046] The difference between this embodiment and embodiment 1 is that the Chinese medicine preparation includes the following raw materials in parts by weight: 14 parts of cat's claw, 14 parts of ginseng, 6 parts of aconite, 8 parts of rhubarb, 4 parts of platycodon, 3 parts of peucedanum, 4 parts of asarum, 2 parts of peach kernel, 3 parts of calamus, 3 parts of gypsum, 4 parts of cinnamon, 2 parts of dried ginger, and 12 parts of licorice.

[0047] Test Example 1

[0048] Inhibitory effect of traditional Chinese medicine preparation on proliferation of Lewis lung cancer cells in mice

[0049] 1 Materials and methods

[0050] 1.1 Experimental animals and Lewis lung cancer cell lines

[0051] Thirty-five SD rats, 6-8 weeks old, male, weighing (200-230) g, were purchased from Zhengzhou Huaxing Experimental Animal Farm.

[0052] Mouse Lewis lung cancer cell line was provided by the Cell Bank of the Chinese Academy of Sciences.

[0053] 1.2 Experimental drugs

[0054] Medicine 1: 10 parts of cat's claw, 10 parts of ginseng, 7 parts of aconite, 7 parts of rhubarb, 3 parts of platycodon, 3 parts of peucedanum, 3 parts of asarum, 3 parts of peach kernel, 3 parts of calamus, 3 parts of gypsum, 3 parts of cinnamon, 5 parts of dried ginger, and 10 parts of licorice.

[0055] Medicine 2: 7 parts of aconite, 7 parts of rhubarb, 3 parts of platycodon, 3 parts of peucedanum, 3 parts of asarum, 3 parts of peach kernel, 3 parts of calamus, 3 parts of gypsum, 3 parts of cinnamon, 5 parts of dried ginger, and 10 parts of licorice.

[0056] Medicine 3: 10 parts of cat's claw, 10 parts of ginseng, 3 parts of platycodon, 3 parts of peucedanum, 3 parts of asarum, 3 parts of peach kernel, 3 parts of calamus, 3 parts of gypsum, 3 parts of cinnamon, 5 parts of dried ginger, and 10 parts of licorice.

[0057] Medicine 4: 10 parts of cat's claw, 10 parts of ginseng, 7 parts of aconite, 7 parts of rhubarb, 3 parts of platycodon, 3 parts of peucedanum, 3 parts of peach kernel, 3 parts of calamus, 5 parts of dried ginger, and 10 parts of licorice.

[0058] Drug 5: 10 parts of cat's claw, 10 parts of ginseng, 7 parts of aconite, 7 parts of rhubarb, 3 parts of platycodon, 3 parts of peucedanum, 3 parts of asarum, 3 parts of peach kernel, 3 parts of calamus, 3 parts of gypsum, and 3 parts of cinnamon.

[0059] Medicine 6: 10 parts of cat's claw, 10 parts of ginseng, 7 parts of aconite, 7 parts of rhubarb, 3 parts of asarum, 3 parts of gypsum, 3 parts of cinnamon, 5 parts of dried ginger, and 10 parts of licorice.

[0060] Weigh each raw material according to the above weight proportions, wash, dry and crush it, and prepare the Chinese medicine stock solution according to the conventional decoction method, and concentrate it to a concentration of 2 mL / g (each mL is equivalent to 2 g of the original medicinal material) to obtain the Chinese medicine stock solution 1-6, which is stored in a refrigerator at 0-4°C for standby use.

[0061] 1.3 Methods

[0062] 1.3.1 Preparation of drug-containing serum

[0063] 35 SD rats were randomly divided into 7 groups, 5 rats in each group, including blank control group and experimental groups 1-6; the blank control group was given normal saline, and the experimental groups 1-6 were given Chinese medicine concentrates 1-6 respectively, by gavage, twice a day, 0.5 mL each time. Each group was given continuous gavage for 7 days. 2 hours after the last administration, 10% chloral hydrate was used for ip anesthesia, and blood was collected from the abdominal aorta under a sterile environment, and the blood was kept at 4℃ for 3 hours, and the serum was separated by centrifugation. The serum of the same group was combined, inactivated in a 56℃ water bath for 30 minutes, filtered and sterilized with a 0.22μm syringe filter, and aseptically divided into 1ml EP tubes, and stored in a -20℃ refrigerator for later use.

[0064] 1.3.2 Cell culture and drug intervention

[0065] Lewis lung cancer cell suspension was prepared and cultured according to conventional cell culture methods. Lewis lung cancer cells with good growth at the 6th to 8th generation were amplified for experiments. The cells were divided into 7 groups, with 6 replicate wells in each group and 1×10 6 Each well was inoculated in a 96-well plate, and the cells were cultured in BPMI1640 culture medium containing 10% fetal bovine serum for 24 hours. The culture medium was aspirated, and 200 μL of serum-free RPMI1640 culture medium was added to each culture well, and cultured in the incubator for 24 hours to synchronize Lewis lung cancer cells. Lewis lung cancer cells were then cultured in RPMI1640 containing 10% blank control group mouse serum and test 1-6 group mouse drug-containing serum, 200 μL per well. The effect was 24 hours and 48 hours respectively.

[0066] 1.3.3CCK8 detection of Lewis lung cancer cell proliferation ability

[0067] After the drug intervention, 10 μL CCK8 was added and cultured for 1-4 hours, then the absorbance at 450 nm was measured and the cell proliferation inhibition rate was calculated. Cell proliferation inhibition rate = (absorbance of the test group - absorbance of the blank control group) / absorbance of the blank control group × 100%.

[0068] 1.4 Statistical methods

[0069] SPSS 18.0 statistical analysis software was used for processing. ± standard deviation (s), and one-way analysis of variance was used for comparison between groups. Bonferroni test was used for analysis when variance was equal, and Dunnett's T3 test was used for analysis when variance was unequal. The test level was α = 0.05.

[0070] 2 Results

[0071] The results of the inhibitory effects of different Chinese medicine preparations on the proliferation of Lewis lung cancer cells in mice are shown in Table 1. Compared with the blank group, test groups 1-6 can significantly inhibit the proliferation ability of Lewis lung cancer cells in mice (p<0.05, p<0.01). Among them, the inhibition rate of Lewis lung cancer cell proliferation in test group 1 was significantly higher than that in other test groups (p<0.05, p<0.01). The results show that the inhibitory effect of the Chinese medicine preparation of the present invention (and test group 1) on Lewis lung cancer cells in mice is significantly better than that of other Chinese medicine preparations, especially the Chinese medicine preparation lacking cat's claw, ginseng or rhubarb, and aconite (test group 2 or test group 3) on lung cancer cells is significantly lower than that of the other groups. Each medicinal material of the Chinese medicine preparation of the present invention is indispensable, and the Chinese medicine preparation lacking some of the medicinal materials cannot achieve a good inhibitory effect on the proliferation of Lewis lung cancer cells.

[0072] Table 1 Effects of different Chinese herbal preparations on the proliferation of Lewis lung cancer cells in mice

[0073]

[0074] Compared with the blank control group, *p<0.05, **p<0.01; compared with the experimental group 1, # p<0.05, ## p<0.01Test Example 2

[0075] 1 Materials and methods

[0076] 1.1 Experimental animals and Lewis lung cancer cell lines

[0077] Thirty SPF grade C57BL / 6 mice, 6-8 weeks old, male, weighing (20±3) g, were purchased from Yusibeifu (Beijing) Biotechnology Co., Ltd.

[0078] Twenty SPF SD rats, half male and half female, weighing (200-230) g, were purchased from Zhengzhou Huaxing Experimental Animal Farm.

[0079] Lewis lung cancer cell line labeled with luciferase (Lewis-luc) was provided by Shanghai Fuheng Biotechnology Co., Ltd.

[0080] A549 lung cancer cells were provided by the Cell Bank of the Chinese Academy of Sciences.

[0081] 1.2 Trial Drugs

[0082] The Chinese medicine pill of the present invention comprises the following raw materials: 10 parts of cat's claw, 10 parts of ginseng, 7 parts of aconite, 7 parts of rhubarb, 3 parts of platycodon, 3 parts of peucedanum, 3 parts of asarum, 3 parts of peach kernel, 3 parts of calamus, 3 parts of gypsum, 3 parts of cinnamon bark, 5 parts of dried ginger and 10 parts of liquorice, which are washed, dried and powdered, fully mixed and then fully mixed with heated honey in a ratio of 100 g:65 g to prepare honey pills with a particle size of 5-6 mm, and the weight of the pills is 1.58 g / 30.

[0083] Cisplatin injection, product of Qilu Pharmaceutical Co., Ltd., product batch number EA1A9004B.

[0084] 1.3 Establishment of animal model and grouping treatment

[0085] Thirty mice were randomly divided into a model group, a Chinese medicine group, and a cisplatin group, with 10 mice in each group. A mouse model of lung cancer was established by subcutaneous inoculation: Lewis-luc cells cultured in DMEM high-glucose medium (containing 100 mL / L fetal bovine serum) were digested with trypsin, counted, and prepared into 1×10 7 / mL of cell suspension. The above cell suspension was inoculated subcutaneously in the right axilla of each group of C57BL / 6 mice, with an inoculation volume of 0.2mL / mouse. After inoculation, the mice were returned to the breeding cage for continued feeding. One day after inoculation, the following interventions were performed on each group of mice. Chinese medicine group: 240μg / (g·d) of the Chinese medicine pills of the present invention were given by gavage + intraperitoneal injection of an equal volume of normal saline; cisplatin group: an equal volume of normal saline was given by gavage + intraperitoneal injection of cisplatin 0.5μg / (g·d); model group: an equal volume of normal saline was given by gavage + intraperitoneal injection. Gavage and intraperitoneal injection were given once a day for 14 days.

[0086] 1.4 Preparation of drug-containing serum

[0087] 20 SD rats were randomly divided into 2 groups, 10 in each group, respectively, a control group and a Chinese medicine group; the Chinese medicine group: the Chinese medicine pills of the present invention were given by intragastric gavage of 240 μg / (g·d) + intraperitoneal injection of an equal volume of normal saline; the control group: the same volume of normal saline was given by intragastric gavage + intraperitoneal injection. Each group was continuously gavaged for 7 days. 2 hours after the last administration, 10% chloral hydrate was used for ip anesthesia, and blood was collected from the abdominal aorta under a sterile environment, and the blood was kept at 4°C for 3 hours, and the serum was separated by centrifugation. The serum of the same group was combined, inactivated in a 56°C water bath for 30 minutes, filtered and sterilized with a 0.22μm syringe filter, aseptically packaged in 1ml EP tubes, and stored in a -20°C refrigerator for standby use.

[0088] 1.5 Cell culture and drug intervention

[0089] A549 lung cancer cell suspension was prepared and cultured according to conventional cell culture methods. A549 lung cancer cells with good growth at the 6th to 8th generation were amplified for experiments. The cells were divided into 3 groups, with 5 replicates in each group and 1×10 6 Each well was inoculated with 10% fetal bovine serum in BPMI1640 culture medium for 24 hours, the culture medium was aspirated, and 200 μL of serum-free RPMI1640 culture medium was added to each well, and the culture was continued in the incubator for 24 hours to synchronize Lewis lung cancer cells. A549 lung cancer cells were then cultured with RPMI1640 containing 10% control group mouse serum, Chinese medicine group serum and cisplatin (5 μg / mL), 200 μL per well.

[0090] 1.6 Detection indicators

[0091] 1.6.1 General

[0092] After the model was prepared, the mice's fur color, spirit, diet, activity, etc. were observed at fixed time every 2 days, the weight of each group of mice was recorded, and the net weight of each group of mice was calculated.

[0093] 1.6.2 Tumor growth

[0094] 24 hours after the last administration, the growth of tumor in mice was observed by small animal in vivo imaging system. Mice were anesthetized with 50 μg / g body weight of sodium pentobarbital and intraperitoneally injected with 125 μg / g body weight of fluorescein substrate. After 10 minutes, the fluorescence signal intensity was detected, photos were taken, and the integrated optical density value was analyzed by Image-Pro Plus 6.0 software.

[0095] 1.6.3 Tumor tissue pathological morphology

[0096] The mouse tumor tissues were fixed with 40 g / L paraformaldehyde, embedded in conventional paraffin, and 4 μm thick sections were prepared. The sections were stained with hematoxylin-eosin, and the morphological changes of the tumor tissues were observed under a microscope.

[0097] 1.6.4 A549 lung cancer cell clone formation

[0098] 400 A549 cells were inoculated in each well of a 6-well plate. After the cells grew overnight, the drug-containing serum prepared in 1.4 was used for drug intervention according to the grouping. The amount of drug-containing serum added was 10% of the culture medium. After 48 hours, all the culture medium was replaced with complete culture medium. After 10 days, the cell clone formation was observed, the supernatant was discarded, the cells were washed, 2 mL of 4% polymethanol was added to each well for fixation for 30 minutes, the fixative was discarded, and after washing, an appropriate amount of crystal violet stain was added to each well for staining for 30 minutes. The stain was carefully rinsed with tap water, and the number of cell clones was counted after drying to calculate the clone formation rate. The clone formation rate = (clone number / inoculated cell number) × 100%.

[0099] 1.6.5 Migration of A549 lung cancer cells

[0100] Use a marker pen to draw three evenly spaced horizontal lines on the back of the 6-well plate. Add about 1×10 5 Cells were cultured until the density reached more than 90%. A 200 μL pipette tip was used to measure the ruler and draw horizontal and vertical lines vertically from the back. PBS was used to wash away the damaged and detached cells. The drug-containing serum prepared in 1.4 was used for drug intervention according to the grouping. The amount of drug-containing serum added was 10% of the culture medium. The cells were returned to the incubator for 24 hours and photographed and sampled. The migration rate was calculated. The cell migration rate = (0h scratch width - 24h scratch width) / 0h scratch width × 100%.

[0101] 1.7 Statistical analysis

[0102] SPSS 18.0 statistical analysis software was used for processing. ± standard deviation (s), and one-way analysis of variance was used for comparison between groups. Bonferroni test was used for analysis when variance was equal, and Dunnett's T3 test was used for analysis when variance was unequal. The test level was α = 0.05.

[0103] 2 Results

[0104] 2.1 Comparison of general conditions among groups

[0105] The mice in the model control group showed dull fur, thin body, mental fatigue, reduced activity, and reduced food and water intake. The fur gloss, activity, food intake, and mental state of the mice in the cisplatin group and the Chinese medicine group improved, with the effect of the Chinese medicine group being more obvious. The mice in the Chinese medicine group achieved stable weight maintenance ( Figure 1 A), and the net body weight was also significantly higher than that of mice in the cisplatin group ( Figure 1 B), the results show that the Chinese medicine preparation of the present invention has more advantages in improving the quality of life of mice with lung cancer.

[0106] 2.2 Growth of tumor tissue in each group of mice

[0107] like Figure 2 As shown, the fluorescence signal in the model group mice was enhanced and the tumor proliferation was obvious. Compared with the model group, the intensity and area of ​​the tumor fluorescence signal in the Chinese medicine group and the cisplatin group mice were significantly reduced, and the tumor volume was significantly reduced. The difference was extremely significant (p<0.01), and the Chinese medicine preparation of the present invention had more advantages in inhibiting the growth of mouse tumor tissue.

[0108] 2.3 Pathological morphology of tumor tissues in each group of mice

[0109] like Figure 3As shown in the figure, the tumor cells in the model group were closely arranged and diffusely distributed, with obvious atypia, abundant cytoplasm, large and darkly stained nuclei, and frequent nuclear divisions. Compared with the model group, the Chinese medicine group and the cisplatin group significantly alleviated the phenomenon of closely arranged and diffusely distributed tumor cells, while reducing nuclear divisions and increasing the number of apoptotic cells.

[0110] 2.4 Inhibition of proliferation of A549 lung cancer cells

[0111] like Figure 4 As shown in A, the A549 cells in the control group had better cloning ability, and the number of A549 cells in the Chinese medicine group and cisplatin group was significantly less than that in the control group; compared with the control group, the cloning rate of the Chinese medicine group and cisplatin group was significantly lower than that of the control group (p<0.01), and the cloning rate of the Chinese medicine group was also significantly lower than that of the cisplatin group ( Figure 4 B); The above results show that the Chinese medicine preparation of the present invention has a significant inhibitory effect on the clone formation ability of A549 cells, and the inhibitory ability is better than cisplatin.

[0112] 2.5 Migration of A549 lung cancer cells

[0113] like Figure 5 As shown in A, the A549 cells in the control group had better migration ability than those in the Chinese medicine group and the cisplatin group; compared with the control group, the 24h migration rates of the Chinese medicine group and the cisplatin group were significantly lower than those of the control group (p<0.01), and the 24h migration rate of the Chinese medicine group was equivalent to that of the cisplatin group ( Figure 5 B); The above results show that the Chinese medicine preparation of the present invention has the ability to significantly inhibit the migration of A549 cells.

[0114] 3 Conclusion

[0115] The above test results can show that the Chinese medicine preparation of the present invention can significantly inhibit the tumor growth of Lewis lung cancer mice, improve the pathological morphology of tumor tissue, and improve the quality of life of mice; at the same time, it can also inhibit the proliferation and migration ability of A549 lung cancer cells. The Chinese medicine preparation provided by the present invention provides a new idea and treatment method for the treatment of lung cancer.

[0116] Clinical application cases

[0117] Case 1: Li, female, 57 years old. First visit on February 23, 2022. Symptoms include cough, coughing up white sticky sputum, aggravated by exposure to cold air and wind, and even accompanied by dull pain in the right intercostal space, fatigue, poor spirit, pale complexion, dark red tongue, thin white fur, and stringy pulse. Chest computed tomography (CT) showed: (1) lesions in the apex posterior segment and lower lobe of the left upper lobe, space-occupying lesions and surrounding inflammation, multiple irregular small nodules in the right lower lung, and metastasis to be excluded; (2) enlarged lymph nodes in the left hilar lung, irregular bone destruction of the third rib and right scapula on the right side, irregular high-density shadows and partial bone destruction of multiple thoracic and lumbar vertebrae, and metastasis is considered; (3) local bone destruction of the right femoral head, metastasis cannot be excluded. Positron emission tomography / computed tomography (PET / CT) showed: multiple foci of increased bone uptake throughout the body, suggesting multiple bone metastases. Lung puncture pathological biopsy showed: lung adenocarcinoma. Immunohistochemistry showed: TTF-1 (+), NapsinA (+), CK7 (+), CD56 (-), CK5 / 6 (-), P40 (-), Ki-67 (about 20%), PD-L1 (TPS about 1%). Lung cancer-related gene mutation detection showed: exon 19 deletion insertion mutation, BRAF gene V600E mutation.

[0118] Diagnosis: Lung adenocarcinoma with bilateral lung / lymph node / bone metastasis (T4N2M1c, stage IVb).

[0119] On March 8, 2022, he began to take osimertinib mesylate tablets (80 mg / time / day) for targeted anti-tumor treatment, and experienced adverse gastrointestinal reactions such as poor appetite and diarrhea. Starting from May 1, 2022, on the basis of the patient's targeted treatment, he took the Chinese medicine pills prepared in Example 1 orally. 3 times / day, 10 pills / time. During regular follow-up, the patient's cough and sputum were significantly improved compared with before, and the adverse reactions such as poor appetite, poor sleep, and diarrhea caused by the previous targeted drugs disappeared, and the general fatigue was relieved. After taking the medicine for 8 months, he was re-examined on January 9, 2023. He was in good spirits, with a pale red tongue, a thin white fur, and a stringy pulse. Chest CT images and tumor markers before and after treatment showed that the tumor lesions in the left upper lobe of the lung were reduced, and the multiple scattered small patchy shadows in both lungs were smaller than the previous lesions ( Figure 6 The levels of tumor markers carcinoembryonic antigen (CEA) and neuron-specific enolase (NSE) decreased significantly, while the level of cytokeratin 19 fragment (Cyfra21-1) remained basically unchanged (Table 2).

[0120] Case 2: Xu, female, 57 years old. First visit on August 26, 2022. Symptoms include cough, coughing up yellow sticky sputum, coughing up blood, no chest tightness, wheezing and other discomforts, poor appetite, loss of appetite, poor spirit, dark complexion, dark tongue, thick white fur, and deep and stringy pulse. Chest CT showed nodules in the left upper lobe of the lung, considering the possibility of Ca; lumbar MRI (Magnetic Resonance Imaging, MRI) showed: T11, T12 vertebral bodies and left pedicles, L1 vertebral bodies and bilateral pedicles, L2, L3 vertebral bodies with multiple abnormal signal shadows, considering metastasis. Bronchoscopy biopsy was performed, and the pathological results showed: adenocarcinoma. Immunohistochemistry showed: CK7 (+), NapsinA (+), TTF-1 (-), CK5 / 6 (-), P40 (-), Ki-67 (about 10% +). Genetic test results showed: EGFR gene mutation.

[0121] Diagnosis: left lung adenocarcinoma with bone metastasis (T2N1M1a, stage IVa).

[0122] On September 3, 2022, chemotherapy with "pemetrexed disodium + nedaplatin + denosumab" was given, and on September 9, 2022, targeted anti-tumor treatment with osimertinib mesylate tablets (80 mg / time / day) was given, and adverse reactions such as rash and itching occurred. Starting from September 15, 2022, on the basis of targeted therapy, the Chinese medicine pills prepared in Example 1 were taken orally, 3 times / day, 10 pills / time. Regular follow-up showed that during the medication period, the patient's cough and sputum were improved compared with before, and the discomfort such as rash and itching were significantly relieved, and the appetite was good. After taking the medicine for 5 months, the patient was re-examined on February 6, 2023. He was in good spirits, with a dark tongue, a thin white fur, and a deep and stringy pulse. Chest CT images and tumor markers before and half a year after treatment showed that the tumor lesions in the left upper lobe of the lung were smaller than before treatment ( Figure 7 The level of tumor marker CEA decreased significantly, the level of NSE increased slightly, and the level of Cyfra21-1 remained basically unchanged (Table 2).

[0123] Case 3: Cao, male, 80 years old. First visit on February 20, 2022. Symptoms include cough, expectoration, large amount of white sputum, easy to cough, occasional blood in sputum, occasional chest tightness, wheezing, no fever, headache or other discomfort symptoms, poor spirit, sallow complexion, red tongue with teeth marks on the edges, and weak pulse.

[0124] Chest CT showed: (1) Central lung cancer in the posterior segment of the left lower lobe, with mediastinal lymph node metastasis; (2) Lesions in the posterior segment of the apex of the left upper lobe, focal inflammation, and intrapulmonary metastasis; (3) Atelectasis of the right middle lobe. Bronchoscopy and biopsy were performed, and the pathological results showed a tendency toward adenocarcinoma. Immunohistochemistry showed: TTF-1 (weakly +), P53 (+), CK5 / 6 (-), CD56 (-), NapsinA (-), P40 (-), Ki-67 (40%), PD-L1 (TPS < 1%). Genetic testing results showed: TP53 mutation rate was 14.4%.

[0125] Diagnosis: Lung adenocarcinoma with lung metastasis (T2aN1M1a, stage IVa).

[0126] On April 25, 2022, 48Gy CyberKnife treatment was performed. Starting from May 3, 2022, on the basis of Western medicine treatment, the Chinese medicine pills prepared in Example 1 were taken orally, 3 times / day, 10 pills / time. Regular follow-up showed that the cough and sputum during the medication period were improved compared with before, and the discomfort such as chest tightness and asthma were significantly alleviated. After taking the medicine for 3 months, the follow-up visit on August 2, 2022 showed that the patient was in good spirits, with a red tongue, a thin white fur, and a thin pulse; chest CT images and tumor markers before and 3 months after treatment showed that the tumor lesions in the dorsal segment of the left lower lobe and the left side of the hilum were smaller than before treatment ( Figure 8 The lesion in the posterior segment of the left upper lobe apex was significantly smaller than before; the levels of tumor markers CEA, NSE, and Cyfra21-1 were significantly decreased compared with before (Table 2).

[0127] Table 2 Changes in the levels of tumor markers CEA, NSE and Cyfra21-1 before and after treatment in cases 1-3

[0128]

[0129]

[0130] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit the same. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some of the technical features may be replaced by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A Chinese medicine preparation for treating lung cancer, characterized in that: The Chinese medicine preparation is prepared from the following Chinese medicine raw materials in parts by weight: 5-25 parts of cat's claw, 5-25 parts of ginseng, 1-15 parts of aconite, 1-15 parts of rhubarb, 1-10 parts of platycodon, 1-10 parts of peucedanum, 1-10 parts of asarum, 1-10 parts of peach kernel, 1-10 parts of calamus, 1-10 parts of gypsum, 1-10 parts of cinnamon, 1-10 parts of dried ginger, and 5-25 parts of liquorice.

2. A Chinese medicine preparation for treating lung cancer according to claim 1, characterized in that: The traditional Chinese medicine preparation is prepared from the following traditional Chinese medicine raw materials in parts by weight: 10-20 parts of cat's claw, 10-20 parts of ginseng, 5-10 parts of aconite, 5-10 parts of rhubarb, 1-5 parts of platycodon, 1-5 parts of peucedanum, 1-5 parts of asarum, 1-5 parts of peach kernel, 1-5 parts of calamus, 1-5 parts of gypsum, 1-5 parts of cinnamon, 1-5 parts of dried ginger, and 10-20 parts of licorice.

3. A Chinese medicine preparation for treating lung cancer according to claim 1, characterized in that: The Chinese medicine preparation is made of the following Chinese medicine raw materials in parts by weight: 10 parts of cat's claw, 10 parts of ginseng, 7 parts of aconite, 7 parts of rhubarb, 3 parts of platycodon, 3 parts of peucedanum, 3 parts of asarum, 3 parts of peach kernel, 3 parts of calamus, 3 parts of gypsum, 3 parts of cinnamon, 5 parts of dried ginger, and 10 parts of licorice.

4. The Chinese medicine preparation for treating lung cancer according to any one of claims 1 to 3, characterized in that: The Chinese medicine preparation is an oral dosage form, including oral liquid, granules, powders, decoctions, pills, and freeze-dried powders.

5. The Chinese medicine preparation for treating lung cancer according to claim 1, characterized in that: The Chinese medicine preparation is prepared into pills according to the following method: weigh each Chinese medicine raw material according to the above-mentioned weight proportions, wash, dry, grind into powder, fully mix them evenly, and then mix them with heated honey in a weight ratio of 100:60-70 to form honey pills with a particle size of 4-6 mm and a weight of 1-3g / 30 pills, thereby obtaining Chinese medicine pills for treating lung cancer.

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