WO2021142920A1 - 一种治疗肺癌的中药组合物及其制剂和应用 - Google Patents

一种治疗肺癌的中药组合物及其制剂和应用 Download PDF

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WO2021142920A1
WO2021142920A1 PCT/CN2020/080112 CN2020080112W WO2021142920A1 WO 2021142920 A1 WO2021142920 A1 WO 2021142920A1 CN 2020080112 W CN2020080112 W CN 2020080112W WO 2021142920 A1 WO2021142920 A1 WO 2021142920A1
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chinese medicine
traditional chinese
cynomorium
medicine composition
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French (fr)
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李润
李伟煊
朱新燕
王宝儿
朱玲玲
沈敏莹
胡雪峰
舒加菊
李梦
谢桂巧
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东莞市东阳光冬虫夏草研发有限公司
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/06—Fungi, e.g. yeasts
    • A61K36/062—Ascomycota
    • A61K36/066—Clavicipitaceae
    • A61K36/068—Cordyceps
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/41—Crassulaceae (Stonecrop family)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/65—Paeoniaceae (Peony family), e.g. Chinese peony
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/71—Ranunculaceae (Buttercup family), e.g. larkspur, hepatica, hydrastis, columbine or goldenseal
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/04—Antineoplastic agents specific for metastasis

Definitions

  • the invention relates to the technical field of traditional Chinese medicine, in particular to a traditional Chinese medicine composition, as well as its application, and at the same time discloses a traditional Chinese medicine preparation.
  • Cordyceps sinensis also known as Cordyceps sinensis, is a complex of Cordyceps sinensis fungus and Hepidiaceae larvae.
  • Cordyceps sinensis is formed by connecting the worm body and the fungus seat that grows from the head of the worm.
  • the body shape is like a silkworm, 3-5cm long, 0.3-0.8cm in diameter, dark yellow to yellowish brown on the surface, with 20-30 ring patterns, the ring pattern near the head is thinner, and the head is reddish brown; 8 pairs of feet, middle part 4 pairs are more obvious; brittle, easy to break, slightly flat section, pale yellowish white.
  • the pedestal is slender and cylindrical, with a length of 4-7cm and a diameter of about 0.3cm.
  • the surface is dark brown to tan, with fine longitudinal wrinkles, and the upper part is slightly enlarged; the texture is flexible and the cross section is white.
  • Cordyceps has a slightly fishy smell and a slightly bitter taste. According to chemical analysis, Cordyceps contains 25%-30% crude protein, D-mannitol (cordycepic acid) 7%-29%, fat 8.4%, and also contains adenosine, ergosterol, vitamins, cordyceps polysaccharides, a variety of amino acids, etc. .
  • Cordyceps sinensis is one of China's three major tonics alongside ginseng and deer antler, and has a very long history of application in China.
  • the ancient Chinese medicine "Picture Book” records Cordyceps "clears lung heat, cures lung diseases, bacon disease”; “Notes on Golden Juice Manlu Aquarius” contains: “Cordyceps sinensis tastes sweet and warm. Nourishes kidney yin, nourishes lungs, cures lung diseases, bacon "Bacon” is a Vietnamese medical term meaning water and soil.
  • Bacon disease is a variety of allergic diseases caused by inconsistent water and soil, such as allergic rhinitis, asthma, allergic gastroenteritis, allergic dermatitis and other allergic diseases; Wu Yiluo of the Qing Dynasty "Ben Cao Congxin” records: protect lung qi, strengthen the body, nourish the kidney and nourish essence.
  • Cordyceps sinensis Many researches have been carried out on Cordyceps sinensis, and a variety of compatible pharmaceutical compositions of Cordyceps sinensis have been developed.
  • the authorized patent with the announcement number CN102716320B discloses a pharmaceutical composition for the treatment of non-small cell lung cancer, which includes 11 Chinese medicines such as Cordyceps sinensis and Schisandra.
  • the pharmaceutical composition has an effect on the growth of human lung cancer A549 cell transplanted tumors in nude mice. Antagonism.
  • the patent application with publication number CN104547525A also discloses a pharmaceutical composition for anti-non-small cell lung cancer, which is made up of 19 traditional Chinese medicines including astragalus, Radix Pseudostellariae, Ophiopogon japonicus, Schisandra, Cordyceps sinensis, etc.
  • the pharmaceutical composition is verified by experiments. It also has an inhibitory effect on the growth of human lung cancer A549 cell transplanted tumors in nude mice.
  • the existing medicinal compositions for treating lung cancer with Cordyceps participating in compatibility have complex formulations. On the one hand, too many components can easily increase toxicity, and on the other hand, too many components will increase the difficulty of subsequent preparation processes, and Increase production costs.
  • a technical solution adopted by the present invention is: a traditional Chinese medicine composition prepared from raw materials comprising the following parts by weight: 1-10 parts of Cordyceps sinensis, 4-20 parts of Rhodiola, and Cynomorium 3. ⁇ 15 parts, 2 ⁇ 10 parts of Cortex Moutan.
  • the traditional Chinese medicine composition is prepared from the following raw materials in parts by weight: 1-10 parts of Cordyceps sinensis, 4-20 parts of Rhodiola, 3-15 parts of Cynomorium, and 2-10 parts of Cortex Moutan.
  • the traditional Chinese medicine composition is prepared from the following raw materials in parts by weight: 1 to 4 parts of Cordyceps sinensis, 4 to 16 parts of Rhodiola, 3 to 9 parts of Cynomorium, and 2 to 6 parts of Cortex Moutan; more preferably It is prepared from the following raw materials in parts by weight: 3 to 4 parts of Cordyceps sinensis, 12 to 16 parts of Rhodiola, 6 to 9 parts of Cynomorium, and 4 to 6 parts of Cortex Moutan.
  • the present invention also provides a method for preparing the traditional Chinese medicine composition, which is prepared by solvent extraction after mixing the raw materials.
  • each of the raw materials is dry powder.
  • the traditional Chinese medicine composition is prepared by a method including the following steps: mixing the raw materials, soaking in a solvent, decocting, filtering to retain the filtrate, concentrating, and spray drying.
  • the solvent is an organic solvent or water, more preferably methanol, ethanol or water.
  • the present invention also provides a preparation made from the traditional Chinese medicine composition; preferably, the preparation is a powder.
  • the present invention further discloses the use of the traditional Chinese medicine composition and traditional Chinese medicine preparation.
  • the lung cancer is non-small cell lung cancer.
  • the use of the traditional Chinese medicine composition or the preparation in the preparation of a medicament for inhibiting the proliferation and/or migration of human large cell lung cancer cells is not limited.
  • the present invention also provides an inhibitor for inhibiting the proliferation and/or migration of human large cell lung cancer cells, comprising the traditional Chinese medicine composition or the preparation.
  • the invention provides a traditional Chinese medicine composition for treating lung cancer.
  • Tumor is a local manifestation of a systemic disease.
  • the TCM theory believes that the occurrence and development of tumors are caused by lack of righteousness in the human body, external evil and internal invasion, and poor emotions, resulting in imbalance of collective Yin and Yang, imbalance of internal organs, imbalance of Qi and blood, and blockage of meridians and collaterals. .
  • the present invention adopts the method of replenishing the lungs and kidneys, promoting blood circulation and removing blood stasis according to the theory of traditional Chinese medicine, using rhodiola to nourish qi and activating blood as the monarch medicine, cynomorium tonifying the kidney and yang, nourishing essence and blood as the minister medicine, and using Mudanpi to clear heat and promote blood Resolving blood stasis and fresh Cordyceps (breeding products, that is, artificially breeding products) are used as adjuvants to invigorate the lungs and kidneys, and the traditional Chinese medicine anti-tumor compound preparation of the invention is prepared.
  • the raw materials used are introduced as follows.
  • Rhodiola is the dry root and rhizome of Rhodiola crenulata (Hook.f.et Thoms.)H.Ohba.), a plant of the Crassulaceae family. After the flower stems are withered in autumn, they are excavated, the rough bark is removed, washed, and dried. Rhodiola is fragrant in weather, slightly bitter and sweet afterwards. Nature and flavor return to classics: sweet, bitter, and flat. Return to the lung and heart meridian. Functions and indications: invigorating qi and activating blood circulation, clearing the pulse and relieving asthma. It is used for qi deficiency and blood stasis, chest pain, heartache, stroke and hemiplegia, fatigue and asthma.
  • Cynomorium is the dry fleshy stem of Cynomorium songaricum Rupr. Excavate in spring, remove the inflorescence, cut into sections, and dry. Cynomorium has a slight qi, sweet and astringent. Nature and flavor return to classics: sweet, warm. Return to the liver, kidney, and large intestine meridian. Functions and indications: invigorating kidney-yang, nourishing essence and blood, moistening the intestines and laxative. It is used for deficiency of kidney yang, deficiency of essence and blood, weak waist and knee atrophy, impotence smoothness, dry intestines and constipation.
  • Paeonia suffruticosa is the dried root bark of the peony (Paeonia suffruticosa Andr.) of the Ranunculaceae family.
  • the roots are excavated in autumn, fine roots and sand are removed, the root bark is peeled, the rough bark is sun-dried, the wood heart is removed, and the sun-dried.
  • the skin of peony has a fragrant, slightly bitter and astringent taste. Nature and flavor return to menstruation: bitter, pungent, slightly cold.
  • Fresh Cordyceps sinensis (breeding product) is a fresh or dried complex of the larvae of the ergot family fungus Ophicordyceps sinensis (Berk.) Sacc. parasitizing on the larvae of Hepidiaceae insect larvae and the larval carcass under simulated original ecological conditions. Slightly fishy smell and slightly bitter taste. Nature and flavor return to classics: sweet, flat. Return to the lung and kidney meridian. Functions and indications: nourish the lungs and kidneys, stop bleeding and resolve phlegm.
  • the cell activity experiment of the present invention proves that the traditional Chinese medicine composition can obviously inhibit the proliferation and migration of human large cell lung cancer cells (H460), and has anti-H460 tumor cell activity.
  • H460 human large cell lung cancer cells
  • the above experiments prove that the traditional Chinese medicine composition provided by the present invention has obvious synergistic effects.
  • Cordyceps sinensis combined with Rhodiola rosea, Cynomorium cynomorium and Moutan cortex with anti-tumor activity can exert a synergistic effect on the inhibition of the proliferation and migration of H460 tumor cells. Synergistic effect.
  • the Chinese medicine composition of the present invention is expected to be used as an alternative in the screening of Chinese patent medicines for tumors, and provides an excellent source of formulas for the development of new Chinese medicines.
  • the traditional Chinese medicine composition provided by the present invention can reduce the types of traditional Chinese medicines in the composition while achieving the effect of inhibiting tumor activity, has simple compatibility, avoids the toxic and side effects that may be caused by too many components, and can be further developed for clinical use.
  • the invention also provides a traditional Chinese medicine preparation, which is prepared by the preparation process of water extraction, concentration and spray drying.
  • the preparation process is as follows:
  • Purified water is preferred as the leaching solvent.
  • Water has good solubility for polar components such as alkaloids, glycosides, tannins, water-soluble organic acids, amino acids, inorganic salts, etc.; the preferred soaking time is 30 minutes, Facilitate the dissolution and leaching of active ingredients;
  • Fig. 1 is a comparative diagram of the effect of the dry powder of Cordyceps sinensis in different concentrations of Comparative Example 1 on the migration and scratching of human large cell lung cancer cells H460 in the present invention
  • Comparative Example 8 is a comparative diagram of the effects of the mixed dry powder of Rhodiola, Cynomorium, and Moutan Cortex provided by Comparative Example 8 with different concentrations in the present invention on the migration and scratching of human large cell lung cancer cells H460;
  • Figure 3 is a comparison diagram of the effect of the anti-tumor compound dry powder provided in Example 1 of the present invention at different concentrations on the migration and scratching of human large cell lung cancer cells H460;
  • Example 5 is a comparison diagram of the effect of the anti-tumor compound dry powder provided in Example 4 of the present invention at different concentrations on the migration and scratching of human large cell lung cancer cells H460;
  • Example 7 is a comparison diagram of the effect of the anti-tumor compound dry powder provided in Example 5 of the present invention at different concentrations on the migration and scratching of human large cell lung cancer cells H460;
  • Example 8 is a comparative diagram of the effect of the anti-tumor compound dry powder provided in Example 2 of the present invention on the migration and scratching of human large cell lung cancer cells H460;
  • Example 9 is a histogram of the scratch area change ratio of the anti-tumor compound dry powder provided in Example 2 of the present invention on the migration scratch of human large cell lung cancer cell H460;
  • Example 10 is a comparative diagram of the effect of the anti-tumor compound dry powder provided in Example 3 of the present invention on the migration and scratching of human large cell lung cancer cells H460;
  • Fig. 11 is a histogram of the scratch area change ratio of the anti-tumor compound dry powder provided in Example 3 of the present invention on the migration scratch of human large cell lung cancer cell H460.
  • This embodiment provides a traditional Chinese medicine composition, which is made of the following raw materials in parts by weight: 4 parts of Rhodiola, 3 parts of Cynomorium, 2 parts of Cortex Moutan, and 1 part of Cordyceps sinensis.
  • This embodiment provides a traditional Chinese medicine composition, which is made of the following raw materials in parts by weight: 10 parts of Rhodiola, 5 parts of Cynomorium, 4 parts of Cortex Moutan, and 2 parts of Cordyceps sinensis.
  • This embodiment provides a traditional Chinese medicine composition, which is made of the following raw materials by weight: 16 parts of Rhodiola, 7 parts of Cynomorium, 5 parts of Cortex Moutan, and 3 parts of Cordyceps sinensis.
  • This embodiment provides a traditional Chinese medicine composition, which is made of the following raw materials in parts by weight: 20 parts of Rhodiola, 15 parts of Cynomorium, 10 parts of Cortex Moutan, and 10 parts of Cordyceps sinensis.
  • This embodiment provides a traditional Chinese medicine composition, which is made of the following raw materials by weight: 10 parts of Rhodiola, 9 parts of Cynomorium, 6 parts of Cortex Moutan, and 5 parts of Cordyceps sinensis.
  • Rhodiola and Cynomorium mixed dry powder Prepare Rhodiola and Cynomorium mixed dry powder.
  • the weight parts of Rhodiola and Cynomorium are 4 parts of Rhodiola and 3 parts of Cynomorium.
  • Rhodiola rosea and tree peony bark A mixed dry powder of Rhodiola rosea and tree peony bark is prepared. Among them, the weight parts of Rhodiola and Moutan bark are 4 parts of Rhodiola rosea and 2 parts of Moutan bark.
  • a mixed dry powder of Rhodiola rosea and Cordyceps sinensis is prepared.
  • the parts by weight of Rhodiola and Cordyceps are 4 parts of Rhodiola and 1 part of Cordyceps.
  • Cynomorium and Cordyceps A mixed dry powder of Cynomorium and Cordyceps was prepared. Among them, the weight parts of Cynomorium and Cordyceps are 3 parts of Cynomorium and 1 part of Cordyceps.
  • Rhodiola rosea A mixed dry powder of Rhodiola rosea, Cynomorium cynomorium, and Paeonia suffruticosa bark was prepared.
  • the weight parts of Rhodiola, Cynomorium, and Moutan Cortex are 4 parts of Rhodiola, 3 parts of Cynomorium, and 2 parts of Cortex Moutan.
  • Rhodiola rosea A mixed dry powder of Rhodiola rosea, Cynomorium cynomorium, and Paeonia suffruticosa bark was prepared.
  • the weight parts of Rhodiola, Cynomorium, and Moutan Cortex are 10 parts of Rhodiola, 5 parts of Cynomorium, and 4 parts of Cortex Moutan.
  • Rhodiola rosea A mixed dry powder of Rhodiola rosea, Cynomorium cynomorium, and Paeonia suffruticosa bark was prepared.
  • the weight parts of Rhodiola, Cynomorium and Moutan Cortex are 16 parts of Rhodiola, 7 parts of Cynomorium, and 5 parts of Cortex Moutan.
  • Rhodiola rosea A mixed dry powder of Rhodiola rosea, Cynomorium cynomorium, and Paeonia suffruticosa bark was prepared.
  • the weight parts of Rhodiola, Cynomorium, and Moutan Cortex are 20 parts of Rhodiola, 15 parts of Cynomorium, and 10 parts of Cortex Moutan.
  • Rhodiola rosea A mixed dry powder of Rhodiola rosea, Cynomorium cynomorium, and Paeonia suffruticosa bark was prepared.
  • the weight parts of Rhodiola, Cynomorium, and Moutan Cortex are 10 parts of Rhodiola, 9 parts of Cynomorium, and 6 parts of Cortex Moutan.
  • the cell viability test was performed on the anti-tumor compound dry powder provided in the above examples.
  • the specific experiment process is as follows.
  • the cells in the culture flask grow to 85%-95% confluence, the cells are subcultured; discard the old culture medium, wash twice with PBS, add 2ml 0.25% trypsin digestion solution, after the cells become round and float, add 4ml culture solution Terminate the digestion, then transfer to a 15ml sterile centrifuge tube, centrifuge at 1000r/min for 4min, discard the supernatant, and transfer to a new sterile culture flask at a ratio of 1:4.
  • H460 cells in logarithmic growth phase were collected, and counted with RPMI-1640 complete medium and resuspended cells, cells are seeded seeding density of 104 cells / well, a total of 100 L; 24 incubated at 37 °C, 5% CO 2 conditions Hour.
  • Co-cultured cell growth inhibition rate % [(Ac-As)/(Ac-Ab)] ⁇ 100%;
  • Table 1 shows the inhibitory rate and IC 50 value of different concentrations of Comparative Example 1 Cordyceps dry powder on the proliferation of human large cell lung cancer cells H460.
  • Table 2 shows the Rhodiola ⁇ Cynomorium mixed dry powder provided by Comparative Example 2 at different concentrations, the Rhodiola rosea ⁇ Cordyceps sinensis mixed dry powder provided by Comparative Example 3, the Rhodiola rosea ⁇ Cordyceps sinensis mixed dry powder provided by Comparative Example 4, and Comparative Example 5
  • Table 3 shows the inhibitory rate and IC 50 value of the mixed dry powder of Rhodiola, Cynomorium, and Moutan Cortex provided by Comparative Example 8 at different concentrations on the proliferation of human large cell lung cancer cells H460.
  • Table 4 shows the inhibition rate and IC 50 value of the anti-tumor compound dry powder provided in Example 1 at different concentrations on the proliferation of human large cell lung cancer cells H460.
  • Table 5 shows the inhibitory rate and IC 50 value of the mixed dry powder of Rhodiola, Cynomorium and Moutan bark provided by Comparative Example 11 at different concentrations on the proliferation of human large cell lung cancer cells H460.
  • Table 6 shows the inhibition rate and IC 50 value of the anti-tumor compound dry powder provided in Example 4 at different concentrations on the proliferation of human large cell lung cancer cells H460.
  • Table 7 shows the inhibitory rate and IC 50 value of the mixed dry powder of Rhodiola, Cynomorium, and Moutan Cortex provided by Comparative Example 12 at different concentrations on the proliferation of human large cell lung cancer cells H460.
  • Table 8 shows the inhibition rate and IC 50 value of the anti-tumor compound dry powder provided in Example 5 at different concentrations on the proliferation of human large cell lung cancer cells H460.
  • the dry powder made by mixing any two or three of Cordyceps sinensis has a significant increase in the inhibition rate of H460 cell proliferation and a significant decrease in IC 50 value, indicating that the compatibility of Rhodiola rosea, Cynomorium sylvestris, Paeonia suffruticosa and Cordyceps sinensis has achieved synergistic effect.
  • the mixed dry powder made has an inhibitory effect on the proliferation of lung cancer cells H460. Among them, the anti-tumor compound of Example 1 has the best effect.
  • the H460 cells in the logarithmic growth phase were digested with 0.25%-EDTA trypsin and pipetted into a single cell suspension.
  • the cell concentration was adjusted to 5 ⁇ 10 5 cells/mL, and 1 mL/well was seeded in a 6-well plate. Incubate for 24 hours in a 37°C, 5% CO 2 incubator.
  • the comparison diagram of the effect of the anti-tumor compound dry powder provided in Example 2 on the migration and scratch of human large cell lung cancer cells H460 is shown in Figure 8, the histogram of the change in scratch area is shown in Figure 9, and the data of the change in the area of migration and scratch is shown in the table. 16.
  • the sample concentrations of Comparative Example 1, Comparative Example 9 and Example 2 used in Table 16 are all 3 mg/mL.
  • control group is a blank control, cultured with RPMI-1640 complete medium without adding any drugs.
  • the comparative diagram of the effect of the anti-tumor compound dry powder provided in Example 3 on the migration and scratch of human large cell lung cancer cells H460 is shown in Figure 10
  • the histogram of the change in scratch area is shown in Figure 11
  • the data of the change in the area of migration and scratch is shown in the table. 17.
  • the sample concentrations of Comparative Example 1, Comparative Example 10, and Example 3 used in Table 17 are all 3 mg/mL.
  • control group is a blank control, cultured with RPMI-1640 complete medium without adding any drugs.
  • the experimental animals were SPF-grade C57BL/6 male mice from Slack, Hunan, weighing 20-22g. After the animals are purchased, they are placed in the quarantine room for observation for a week, and then transferred to the SPF laboratory for breeding.
  • Example 1 Take the anti-tumor compound dry powder of Example 1 and use 0.9% normal saline to prepare drug concentrations of 30, 50, and 100 mg/mL, respectively, and administer by gavage.
  • the gavage volume is 10 mL/kg, and the drug is administered once a day.
  • Cisplatin is administered by intraperitoneal injection every other day, the injection volume is 1 mL/kg, and the administration dose is 2 mg/kg.
  • A, B, C, D, and E representing the model group
  • the cisplatin group (2mg/kg)
  • the low-dose Chinese medicine antitumor compound group 300mg/kg
  • the Chinese medicine The medium-dose group of anti-tumor compound preparation (500 mg/kg) and the high-dose group of anti-tumor compound preparation of Chinese medicine (1000 mg/kg), 15 animals in each group, a total of 75 animals.
  • mouse Lewis lung cancer cells were cultured with DMEM medium containing 10% fresh fetal bovine serum. In the logarithmic growth phase, the cells were routinely digested with 0.25% pancreatin, and the tumor density was adjusted to 1 ⁇ 10 7 /mL cell suspension.
  • mice in the 8-part group was subcutaneously inoculated with 1 ⁇ 10 7 Lewis lung cancer cells under the armpit of the left extremity, and was administered according to the above-mentioned administration method and dosage at the same time.
  • each mouse of group B was injected with 2mg/kg cisplatin solution, which was administered every other day for a total of 20 days. The administration mode and dosage of the other groups were unchanged, and the rats were given by intragastric administration for 20 days.
  • mice in groups C, D, and E were weighed every day and their mental status was observed. On the 7th day, observe whether the mice in the above 5 groups are dead. On days 1, 3, 13, 15, 17, and 20 after inoculation, the animal tumor measurement and tracking system was used to measure the longest diameter and shortest diameter of the tumor (cm) with an electronic digital caliper.
  • Tumor volume longest tumor diameter ⁇ shortest diameter ⁇ The shortest path/2.
  • the mice in each group were weighed, and the mice were sacrificed by removing the cervical spine. The tumor mass of the mice was completely removed and weighed, and the tumor inhibition rate was calculated.
  • Tumor inhibition rate (%) (average tumor weight of the control group-average tumor weight of the administration group) ⁇ 100%/average tumor weight of the control group. After standardization of all experimental data, GraphPad Prism 7 was used for statistical analysis and graphing.
  • Example 1 the anti-tumor compound of Example 1 at different concentrations did not affect the normal body weight and growth of the mice.
  • the mice in each group were in good mental state and did not die.
  • the anti-tumor compound had no obvious toxic and side effects.
  • the high-dose Example 1 anti-tumor compound significantly inhibited the tumor volume and weight of tumor-bearing mice, and the tumor-inhibiting effect was equivalent to that of the positive control drug cisplatin.

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Abstract

一种治疗肺癌的中药组合物及其制剂和应用，所述组合物由冬虫夏草1～10份，红景天4～20份，锁阳3～15份，牡丹皮2～10份制成。

Description

一种中药组合物、应用及中药制剂 技术领域
本发明涉及中医药技术领域，特别是涉及一种中药组合物，同时还涉及其应用，并同时公开了一种中药制剂。
背景技术
冬虫夏草，别称冬虫草，是冬虫夏草菌和蝙蝠蛾科幼虫的复合体。冬虫夏草由虫体与从虫头部长出的真菌子座相连而成。虫体形如蚕，长3-5cm，直径0.3-0.8cm，表面深黄色至黄棕色，有环纹20-30个，近头部的环纹较细，头部红棕色；足8对，中部4对较明显；质脆，易折断，断面略平坦，淡黄白色。子座细长圆柱形，长4-7cm，直径约0.3cm，表面深棕色至棕褐色，有细纵皱纹，上部稍膨大；质柔韧，断面类白色。冬虫夏草气微腥，味微苦。经化学分析，冬虫夏草含粗蛋白25％～30％，D-甘露醇(即虫草酸)7％～29％，脂肪8.4％，还含有腺苷、麦角甾醇、维生素、虫草多糖、多种氨基酸等。
冬虫夏草作为与人参、鹿茸并列的中国三大补药之一，在中国具有十分久远的应用历史。例如，中医药古籍《图鉴》有记载：冬虫夏草“清肺热，治肺病、培根病”；《金汁甘露宝瓶札记》载：“冬虫夏草味甘性温。滋补肾阴，润肺，治肺病、培根病。”，“培根”为藏医术语，意为水土，培根病即水土不服引起的各种过敏性疾病如过敏性鼻炎、哮喘、过敏性肠胃炎、过敏性皮炎等过敏性疾病；清朝吴仪洛《本草从新》记载：保肺气，实腠理，补肾益精。现代医学关于冬虫夏草的功效，在《中华人民共和国药典一部(2005版)》冬虫夏草项下的“功能与主治”记载为：补肺益肾，止血化痰；用于久咳虚喘，劳嗽咯血，阳痿遗精、腰膝酸痛。
针对冬虫夏草人们进行了多方面研究，目前已经研发出了多种冬虫夏草配伍的药物组合物。例如公告号为CN102716320B的授权专利 公开了一种治疗非小细胞肺癌的药物组合物，其中包括冬虫夏草、五味子等11味中药，实验发现该药物组合物对人肺癌A549细胞裸鼠移植瘤的生长有拮抗作用。公开号为CN104547525A的专利申请也公开了一种抗非小细胞肺癌的药物组合物，由黄芪、太子参、麦冬、五味子、冬虫夏草等19味中药配伍而成，经试验验证，该药物组合物对人肺癌A549细胞裸鼠移植瘤的生长也有抑制作用。但是，现有的冬虫夏草参与配伍的治疗肺癌的药物组合物，组方复杂，一方面组分太多容易造成毒性的增加，另一方面组分过多也会增大后续制剂工艺的难度，且使生产成本升高。
发明内容
为解决上述技术问题，本发明采用的一个技术方案是：一种中药组合物，由包括以下重量份的原料药制备而成：冬虫夏草1～10份，红景天4～20份，锁阳3～15份，牡丹皮2～10份。
优选地，所述中药组合物由以下重量份的原料药制备而成：冬虫夏草1～10份，红景天4～20份，锁阳3～15份，牡丹皮2～10份。
进一步优选地，所述中药组合物由以下重量份的原料药制备而成：冬虫夏草1～4份，红景天4～16份，锁阳3～9份，牡丹皮2～6份；更优选地，由以下重量份的原料药制备而成：冬虫夏草3～4份，红景天12～16份，锁阳6～9份，牡丹皮4～6份。
本发明还提供了所述中药组合物的制备方法，所述中药组合物是将各原料药混合后，经溶剂提取制得。优选地，所述各原料药均为干燥药粉。
优选地，所述中药组合物由包括以下步骤的方法制备而成：将各原料药混合，在溶剂中浸泡、煎煮，过滤留取滤液，浓缩、喷雾干燥。
优选地，所述溶剂为有机溶剂或水，进一步优选为甲醇、乙醇或水。
本发明还提供了一种由所述中药组合物制成的制剂；优选地，所述制剂为粉剂。
本发明还进一步公开了所述中药组合物和中药制剂的用途。
第一方面，所述中药组合物或所述制剂在制备治疗肺癌的药物中的用途；
优选地，所述肺癌为非小细胞肺癌。
第二方面，所述中药组合物或所述制剂在制备用于抑制人大细胞肺癌细胞增殖和/或迁移药物中的用途。
本发明还提供了一种抑制人大细胞肺癌细胞增殖和/或迁移的抑制剂，包含所述中药组合物或所述制剂。
本发明提供了一种用于治疗肺癌的中药组合物。肿瘤是全身疾病的局部表现，中医理论认为肿瘤的发生发展是由于人体正气不足，外邪内侵，加之情志不畅，致使集体阴阳失衡，脏腑失和，气血失调，经络阻滞所致。本发明根据中医“扶正祛邪”理论，采用补益肺肾、活血化瘀法，以红景天益气活血为君药，以锁阳补肾阳、益精血为臣药，以牡丹皮清热活血化瘀、鲜冬虫夏草(繁育品，即人工繁育品)补肺肾为佐使药，制成了本发明的中药抗肿瘤复方制剂。
采用的各原料药分别介绍如下。
红景天为景天科植物大花红景天(Rhodiola crenulata(Hook.f.et Thoms.)H.Ohba.)的干燥根和根茎。秋季花茎凋枯后采挖，除去粗皮，洗净，晒干。红景天气芳香，味微苦涩、后甜。性味归经：甘、苦，平。归肺、心经。功能主治：益气活血，通脉平喘。用于气虚血瘀，胸痹心痛，中风偏瘫，倦怠气喘。
锁阳为锁阳科植物锁阳(Cynomorium songaricum Rupr.)的干燥肉质茎。春季采挖，除去花序，切段，晒干。锁阳气微，味甘而涩。性味归经：甘，温。归肝、肾、大肠经。功能主治：补肾阳，益精血，润肠通便。用于肾阳不足，精血亏虚，腰膝痿软，阳痿滑精，肠燥便秘。
牡丹皮为毛莨科植物牡丹(Paeonia suffruticosa Andr.)的干燥根皮。秋季采挖根部，除去细根和泥沙，剥取根皮，晒干粗皮，除去木心，晒干。牡丹皮气芳香，味微苦而涩。性味归经：苦、辛，微寒。归心、肝、肾经。功能主治：清热凉血，活血化瘀。用于热入营血， 温病发斑，吐血衄血，夜热早凉，无汗骨蒸，经闭痛经，跌扑伤痛，痈肿疮毒。
鲜冬虫夏草(繁育品)为模拟原生态条件下麦角菌科真菌冬虫夏草(Ophicordyceps sinensis(Berk.)Sacc.)寄生在蝙蝠蛾科昆虫幼虫上的子座和幼虫尸体的新鲜或干燥的复合体。气微腥，味微苦。性味归经：甘，平。归肺、肾经。功能主治：补肺益肾，止血化痰。用于肾虚精亏，阳痿遗精，腰膝酸痛，久咳虚喘，劳嗽咯血。长期以来野生冬虫夏草资源匮乏，价格昂贵，质量参差不齐。随着东阳光人工培育冬虫夏草的成功和产能的不断扩大，并且冬虫夏草(繁育品)已获得中检院、药监局的权威认可，还被收录在广东省地方中药材标准，这能为中药新药开发提供稳定可靠的原料来源。
本发明经细胞活性实验证明，该中药组合物能够明显抑制人大细胞肺癌细胞(H460)的增殖及迁移，具有抗H460肿瘤细胞的活性。以上实验证明，本发明提供的该中药组合物具有明显的协同增效作用，冬虫夏草联用具有抗肿瘤活性的红景天、锁阳及牡丹皮，能对H460肿瘤细胞增殖及迁移的抑制发挥协同增效的作用。本发明的中药组合物有望在肿瘤中成药的筛选中作为备选方案，为中药新药的研发提供优秀的配方来源。
本发明提供的该中药组合物在达到具有抑制肿瘤活性效果的同时，减少了组合物中中药的种类，配伍简单，避免了组分太多可能带来的毒副作用，可进一步开发成用于临床治疗非小细胞肺癌的药物。
本发明还提供了一种中药制剂，是采用水提、浓缩、喷雾干燥的制备工艺制得的中药制剂。制备过程如下：
(1)对鲜冬虫夏草、红景天、锁阳、牡丹皮原药材饮片进行鉴定，挑选和整理；
(2)分别把以上四味药材粉碎成粗粉，使药材的粒径减小到1-2mm，这样可以增大药材的表面积，提高浸出提取时有效成分的浸出效率，但也不能过度粉碎，否则容易吸附，不利于浸出提取时的扩散；
(3)优选纯净水作为浸出溶剂，水对极性成分如生物碱、甙、鞣质、水溶性有机酸、氨基酸、无机盐等都有较好的溶解性；优选的浸泡时间为30min，以利于有效成分的溶解和浸出；
(4)浸泡之后开始煎煮，在煎煮过程中，每隔3min搅拌一次，这样可以提高扩散速率，对浸出有利；煎煮结束后过滤，留取滤液；
(5)将滤液减压低温浓缩，浓缩去除水后得到浓药液；
(6)浓缩液经喷雾干燥制成药粉。
附图说明
图1是本发明中不同浓度的对比例1冬虫夏草干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图；
图2是本发明中不同浓度的对比例8提供的红景天、锁阳、牡丹皮混合干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图；
图3是本发明中不同浓度的实施例1提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图；
图4是本发明中不同浓度的对比例11提供的红景天、锁阳、牡丹皮混合干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图；
图5是本发明中不同浓度的实施例4提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图；
图6是本发明中不同浓度的对比例12提供的红景天、锁阳、牡丹皮混合干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图；
图7是本发明中不同浓度的实施例5提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图；
图8是本发明中实施例2提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图；
图9是本发明中实施例2提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460迁移划痕的划痕面积变化比柱状图；
图10是本发明中实施例3提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图；
图11是本发明中实施例3提供的抗肿瘤复方干粉对人大细胞肺 癌细胞H460迁移划痕的划痕面积变化比柱状图。
具体实施方式
下面通过具体实施例对本发明的技术方案进行详细说明。
实施例1
本实施例提供了一种中药组合物，由以下重量份的原料药制成：红景天4份，锁阳3份，牡丹皮2份，冬虫夏草1份。
具体如下：
(1)取鲜冬虫夏草3g、红景天12g、锁阳9g、牡丹皮6g，均为原药材饮片；
(2)分别把以上四味药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入120g水，先浸泡30min，然后放在电炉上，煮沸40min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在40℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成抗肿瘤复方干粉。
实施例2
本实施例提供了一种中药组合物，由以下重量份的原料药制成：红景天10份，锁阳5份，牡丹皮4份，冬虫夏草2份。
具体如下：
(1)取鲜冬虫夏草2g、红景天10g、锁阳5g、牡丹皮4g，均为原药材饮片；
(2)分别把以上四味药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入84g水，先浸泡30min，然后放在电炉上，煮沸50min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在60℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成抗肿瘤复方干粉。
实施例3
本实施例提供了一种中药组合物，由以下重量份的原料药制成：红景天16份，锁阳7份，牡丹皮5份，冬虫夏草3份。
具体如下：
(1)取鲜冬虫夏草3g、红景天16g、锁阳7g、牡丹皮5g，均为原药材饮片；
(2)分别把以上四味药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入124g水，先浸泡30min，然后放在电炉上，煮沸60min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在60℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成抗肿瘤复方干粉。
实施例4
本实施例提供了一种中药组合物，由以下重量份的原料药制成：红景天20份，锁阳15份，牡丹皮10份，冬虫夏草10份。
具体如下：
(1)取鲜冬虫夏草10g、红景天20g、锁阳15g、牡丹皮10g，均为原药材饮片；
(2)分别把以上四味药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入220g水，先浸泡30min，然后放在电炉上，煮沸40min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在40℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成抗肿瘤复方干粉。
实施例5
本实施例提供了一种中药组合物，由以下重量份的原料药制成：红景天10份，锁阳9份，牡丹皮6份，冬虫夏草5份。
具体如下：
(1)取鲜冬虫夏草5g、红景天10g、锁阳9g、牡丹皮6g，均为原药材饮片；
(2)分别把以上四味药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入120g水，先浸泡30min，然后放在电炉上，煮沸40min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在40℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成抗肿瘤复方干粉。
对比例1
制备冬虫夏草干粉。
具体如下：
(1)取鲜冬虫夏草饮片10g，粉碎至粒径减小到1-2mm，得到粗粉；
(3)将粗粉放入蒸煮罐内，加入80g水，先浸泡30min，然后放在电炉上，煮沸40min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在40℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成冬虫夏草干粉。
对比例2
制备红景天、锁阳混合干粉。其中红景天、锁阳的重量份数为红景天4份，锁阳3份。
具体如下：
(1)取红景天16g、锁阳12g，均为原药材饮片；
(2)分别把以上药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入75g水，先浸泡30min，然后放在电炉上，煮沸40min，在煎煮过程中，每隔3min 搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在55℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成混合干粉。
对比例3
制备红景天、牡丹皮混合干粉。其中红景天、牡丹皮的重量份数为红景天4份，牡丹皮2份。
具体如下：
(1)取红景天8g、牡丹皮4g，均为原药材饮片；
(2)分别把以上药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入93g水，先浸泡30min，然后放在电炉上，煮沸40min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在50℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成混合干粉。
对比例4
制备红景天、冬虫夏草混合干粉。其中红景天、冬虫夏草的重量份数为红景天4份，冬虫夏草1份。
具体如下：
(1)取红景天8g、冬虫夏草2g，均为原药材饮片；
(2)分别把以上药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入87g水，先浸泡30min，然后放在电炉上，煮沸40min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在60℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成混合干粉。
对比例5
制备锁阳、牡丹皮混合干粉。其中锁阳、牡丹皮的重量份数为锁 阳3份，牡丹皮2份。
具体如下：
(1)取锁阳12g、牡丹皮8g，均为原药材饮片；
(2)分别把以上药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入76g水，先浸泡30min，然后放在电炉上，煮沸40min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在65℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成混合干粉。
对比例6
制备锁阳、冬虫夏草混合干粉。其中锁阳、冬虫夏草的重量份数为锁阳3份，冬虫夏草1份。
具体如下：
(1)取锁阳9g、冬虫夏草3g，均为原药材饮片；
(2)分别把以上药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入68g水，先浸泡30min，然后放在电炉上，煮沸30min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在40℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成混合干粉。
对比例7
制备牡丹皮、冬虫夏草混合干粉。其中牡丹皮、冬虫夏草的重量份数为牡丹皮2份，冬虫夏草1份。
具体如下：
(1)取牡丹皮8g、冬虫夏草4g，均为原药材饮片；
(2)分别把以上药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入108g水，先浸泡30min，然后放在电炉上，煮沸40min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在40℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成混合干粉。
对比例8
制备红景天、锁阳、牡丹皮混合干粉。其中红景天、锁阳、牡丹皮的重量份数为红景天4份，锁阳3份，牡丹皮2份。
具体如下：
(1)取红景天12g、锁阳9g、牡丹皮6g，均为原药材饮片；
(2)分别把以上三味药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入108g水，先浸泡30min，然后放在电炉上，煮沸40min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在40℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成混合干粉。
对比例9
制备红景天、锁阳、牡丹皮混合干粉。其中红景天、锁阳、牡丹皮的重量份数为红景天10份，锁阳5份，牡丹皮4份。
具体如下：
(1)取红景天20g、锁阳10g、牡丹皮8g，均为原药材饮片；
(2)分别把以上三味药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入152g水，先浸泡30min，然后放在电炉上，煮沸50min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在60℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成混合干粉。
对比例10
制备红景天、锁阳、牡丹皮混合干粉。其中红景天、锁阳、牡丹皮的重量份数为红景天16份，锁阳7份，牡丹皮5份。
具体如下：
(1)取红景天32g、锁阳14g、牡丹皮10g，均为原药材饮片；
(2)分别把以上三味药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入224g水，先浸泡30min，然后放在电炉上，煮沸60min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在60℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成混合干粉。
对比例11
制备红景天、锁阳、牡丹皮混合干粉。其中红景天、锁阳、牡丹皮的重量份数为红景天20份，锁阳15份，牡丹皮10份。
具体如下：
(1)取红景天20g、锁阳15g、牡丹皮10g，均为原药材饮片；
(2)分别把以上三味药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入185g水，先浸泡30min，然后放在电炉上，煮沸50min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在40℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成混合干粉。
对比例12
制备红景天、锁阳、牡丹皮混合干粉。其中红景天、锁阳、牡丹皮的重量份数为红景天10份，锁阳9份，牡丹皮6份。
具体如下：
(1)取红景天20g、锁阳18g、牡丹皮12g，均为原药材饮片；
(2)分别把以上三味药材粉碎成粗粉，粉碎至药材的粒径减小到1-2mm；
(3)将粉碎后的药材混合一起放入蒸煮罐内，加入207g水，先浸泡30min，然后放在电炉上，煮沸60min，在煎煮过程中，每隔3min搅拌一次，煎煮结束后过滤，留取滤液；
(4)将滤液减压低温浓缩，在40℃减压浓缩，去除水后得到浓药液，将得到的浓缩液进行喷雾干燥，制成混合干粉。
实验例
对以上各实施例提供的抗肿瘤复方干粉进行了细胞活性检测。具体实验过程如下。
一、细胞培养
取H460细胞，采用RPMI-1640完全培养基培养，置于37℃、5％二氧化碳的培养箱中培养；
每日倒置显微镜观察细胞1次，隔2天换培养液一次；
培养瓶中的细胞生长到85％～95％融合时进行细胞传代；弃去旧培养液，PBS反复冲洗2次，加入0.25％胰酶消化液2ml，细胞变圆、上浮后，加入4ml培养液终止消化，再转移至15ml无菌离心管中，1000r/min离心4min，弃上清液，以1:4比例转移至新的无菌培养瓶中培养。
二、细胞增殖实验
分别准确称取上述实施例及对比例中得到的干粉，以3mg/mL为初始终浓度，3倍稀释成一系列梯度浓度，共设6个浓度点，溶于RPMI-1640完全培养基中，用无菌滤头过滤，制成给药样品。
收集对数生长期的H460细胞，计数，用RPMI-1640完全培养基重新悬浮细胞，接种细胞，接种密度为10 4个细胞/孔，共100μL；在37℃、5％CO 2条件下孵育24小时。
按100μL/孔加入给药样品，加完药之后，置于37℃、5％CO 2培养箱中，孵育24小时。
弃去培养基，用100μL无血清培养基加上10μL CCK-8溶液， 置于37℃培养箱中孵育1小时，酶标仪450nm处检测各孔的吸光度(A)值。按下式计算药物对肿瘤细胞生长的抑制率：
共培养细胞生长抑制率％＝[(Ac-As)/(Ac-Ab)]×100％；
其中：
As:样品的吸光度(肿瘤细胞+待测提取物+CCK-8)
Ac:阴性对照的吸光度(肿瘤细胞+CCK-8)
Ab:空白对照的吸光度(培养基+CCK-8)
并采用Graph Pad Prism5软件计算IC 50数值。
具体结果见表1-表3所示。
表1为不同浓度的对比例1冬虫夏草干粉对人大细胞肺癌细胞H460增殖的抑制率和IC 50数值。
表1
Figure PCTCN2020080112-appb-000001
表2为不同浓度的对比例2提供的红景天\锁阳混合干粉、对比例3提供的红景天\牡丹皮混合干粉、对比例4提供的红景天\冬虫夏草混合干粉、对比例5提供的锁阳\牡丹皮混合干粉、对比例6提供的锁阳\冬虫夏草混合干粉以及对比例7提供的牡丹皮\冬虫夏草混合干粉对人大细胞肺癌细胞H460增殖的抑制率和IC 50数值。
表2
Figure PCTCN2020080112-appb-000002
Figure PCTCN2020080112-appb-000003
表3为不同浓度的对比例8提供的红景天、锁阳、牡丹皮混合干粉对人大细胞肺癌细胞H460增殖的抑制率和IC 50数值。
表3
Figure PCTCN2020080112-appb-000004
表4为不同浓度的实施例1提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460增殖的抑制率和IC 50数值。
表4
Figure PCTCN2020080112-appb-000005
表5为不同浓度的对比例11提供的红景天、锁阳、牡丹皮混合干粉对人大细胞肺癌细胞H460增殖的抑制率和IC 50数值。
表5
Figure PCTCN2020080112-appb-000006
表6为不同浓度的实施例4提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460增殖的抑制率和IC 50数值。
表6
Figure PCTCN2020080112-appb-000007
表7为不同浓度的对比例12提供的红景天、锁阳、牡丹皮混合干粉对人大细胞肺癌细胞H460增殖的抑制率和IC 50数值。
表7
Figure PCTCN2020080112-appb-000008
表8为不同浓度的实施例5提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460增殖的抑制率和IC 50数值。
表8
Figure PCTCN2020080112-appb-000009
通过表1-表8数据可以看出，红景天、锁阳、牡丹皮与冬虫夏草配伍制成的混合干粉，相比冬虫夏草单味药制成的干粉以及由红景天、锁阳、牡丹皮、冬虫夏草中任意两种或三种混合制成的干粉，对H460细胞增殖的抑制率显著提升，IC 50值显著降低，说明红景天、锁阳、牡丹皮与冬虫夏草配伍后达到了协同增效的效果，制成的混合干粉对肺癌细胞H460增殖具有抑制作用。其中，尤以实施例1抗肿瘤复方效果最好。
三、细胞迁移实验
分别称取上述实施例及对比例中得到的干粉，溶于RPMI-1640完全培养基中，用无菌滤头过滤，制成给药样品。
将对数生长期的H460细胞用0.25％-EDTA的胰酶消化吹打成单细胞悬液，调整细胞浓度为5×10 5个/mL，按1mL/孔接种于6孔板中，置于37℃、5％CO 2培养箱中，孵育24小时。
当细胞单层长满时，用200μL移液枪枪头在每孔中央沿垂直方向划痕。
用PBS洗掉多余的细胞，并且加入含有不同浓度药物(0、0.012、0.333、3mg/mL)的无血清培养基1mL，拍照记录。24小时后，再拍照记录，与0小时划痕面积相比，计算划痕面积变化比。
其中，不同浓度的对比例1冬虫夏草干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图见图1所示，迁移划痕面积变化比数据 见表9。
表9
Figure PCTCN2020080112-appb-000010
不同浓度的对比例8提供的红景天、锁阳、牡丹皮混合干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图见图2所示，迁移划痕面积变化比数据见表10。
表10
Figure PCTCN2020080112-appb-000011
不同浓度的实施例1提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图见图3所示，迁移划痕面积变化比数据见表11。
表11
Figure PCTCN2020080112-appb-000012
不同浓度的对比例11提供的红景天、锁阳、牡丹皮混合干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图见图4所示，迁移划痕面积变化比数据见表12。
表12
Figure PCTCN2020080112-appb-000013
不同浓度的实施例4提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图见图5所示，迁移划痕面积变化比数 据见表13。
表13
Figure PCTCN2020080112-appb-000014
不同浓度的对比例12提供的红景天、锁阳、牡丹皮混合干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图见图6所示，迁移划痕面积变化比数据见表14。
表14
Figure PCTCN2020080112-appb-000015
不同浓度的实施例5提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图见图7所示，迁移划痕面积变化比数据见表15。
表15
Figure PCTCN2020080112-appb-000016
实施例2提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图见图8所示，划痕面积变化比柱状图见图9所示，迁移划痕面积变化比数据见表16。表16中采用的对比例1、对比例9和实施例2的样品浓度均为3mg/mL。
表16
Figure PCTCN2020080112-appb-000017
Figure PCTCN2020080112-appb-000018
注：对照组为空白对照，用RPMI-1640完全培养基培养，且不加任何药物。
实施例3提供的抗肿瘤复方干粉对人大细胞肺癌细胞H460迁移划痕的影响对比图见图10所示，划痕面积变化比柱状图见图11所示，迁移划痕面积变化比数据见表17。表17中采用的对比例1、对比例10、实施例3样品浓度均为3mg/mL。
表17
Figure PCTCN2020080112-appb-000019
注：对照组为空白对照，用RPMI-1640完全培养基培养，且不加任何药物。
通过以上表9-表17数据可以看出，红景天、锁阳、牡丹皮与冬虫夏草配伍制成的混合干粉，相比冬虫夏草单味药制成的干粉以及由红景天、锁阳、牡丹皮制成的干粉，对H460细胞迁移的抑制率显著提升，说明红景天、锁阳、牡丹皮与冬虫夏草配伍后达到了协同增效的效果，制成的混合干粉对肺癌细胞H460迁移具有抑制作用。其中，尤以实施例3抗肿瘤复方效果最好。
四、中药抗肿瘤复方制剂对Lewis肺癌小鼠的抗肿瘤研究
(一)实验材料
实验动物选取来自湖南斯莱克的SPF级C57BL/6雄性小鼠，体重为20～22g。动物购进后，先放置在检疫室观察一周，然后转入SPF级实验室饲养。
取实施例1的抗肿瘤复方干粉，用0.9％生理盐水分别配制成30、50和100mg/mL的药物浓度，采用灌胃给药，灌胃体积为10mL\kg，每天给药一次。顺铂采用隔天腹腔注射给药，注射体积为1mL\kg，给药剂量为2mg/kg。
(二)实验设计
根据动物体重进行随机分为A、B、C、D、E共5个组，分别代表模型组、顺铂组(2mg/kg)、中药抗肿瘤复方制剂低剂量组(300mg/kg)、中药抗肿瘤复方制剂中剂量组(500mg/kg)、中药抗肿瘤复方制剂高剂量组(1000mg/kg)，每组15只动物，共75只。
(三)实验步骤
1、细胞培养
在37℃、体积分数为5％CO 2的培养箱中，用含有10％新鲜胎牛血清的DMEM培养基培养小鼠Lewis肺癌细胞。至对数生长期，用0.25％胰酶常规消化细胞，调整瘤株密度至1×10 7/mL的细胞悬液。
2、Lewis小鼠肺癌模型的建立、给药及肿瘤体积测量
实验开始前7天，A和B组每天灌胃给予0.9％生理盐水，C、D、E组每天灌胃给予相应剂量等体积的药物。第8天，8组分组中每只小鼠都在左肢腋下皮下接种1×10 7个Lewis肺癌细胞，同时按上述给药方式及剂量给药。第9天后，B组小鼠每只注射2mg/kg的顺铂溶液，隔天给药，共20天。其余组给药方式及剂量不变，连续灌胃给药20天。
3、数据测量
第1、3、7天，每天称量C、D、E组各小鼠体重并观察精神状态。第7天时，观察以上5组小鼠是否死亡。接种后第1、3、13、15、17、20天，采用动物肿瘤测量跟踪系统，电子数显卡尺测量肿瘤最长径、最短径(cm)，肿瘤体积＝肿瘤最长径×最短径×最短径/2。第29天称量各组小鼠体重后脱颈椎处死小鼠，完整剥离小鼠瘤块称重，计算抑瘤率。抑瘤率(％)＝(对照组平均瘤重-给药组平均瘤重)×100％/对照组平均瘤重。所有实验数据经标准化处理后，用GraphPad Prism 7进行统计学分析和作图。
动物实验结果如下：
不同浓度的实施例1的抗肿瘤复方干粉在第1、3、7天对Lewis肺癌小鼠体重的影响见表18。
表18
Figure PCTCN2020080112-appb-000020
不同浓度的实施例1的抗肿瘤复方干粉在接种后第1、3、13、15、17、20天对Lewis肺癌小鼠肿瘤体积的影响见表19。
表19
Figure PCTCN2020080112-appb-000021
注：与NC组相比 #p, *p<0.05或 ##p, **p<0.01。
不同浓度的实施例1的抗肿瘤复方干粉在接种后第20天对Lewis肺癌小鼠肿瘤抑制率的影响见表20。
表20
组别 肿瘤重量(g) 抑瘤率(％)
模型组 2.63±1.47 -
顺铂组 1.40±0.85* 46.79
低剂量组(300mg/kg) 2.03±0.35 22.81
中剂量组(500mg/kg) 1.81±0.57 31.00
高剂量组(1000mg/kg) 1.53±0.86 # 41.83
注：与NC组相比 #p, *p<0.05。
通过表18-表20数据可以看出，不同浓度的实施例1抗肿瘤复方未影响小鼠正常体重及生长，各组小鼠精神状态良好，未出现死亡，抗肿瘤复方无明显的毒副作用。高剂量实施例1抗肿瘤复方显著抑制荷瘤小鼠肿瘤体积和重量，且抑瘤效果与阳性对照药顺铂相当。
虽然，上文中已经用一般性说明、具体实施方式及试验，对本发明作了详尽的描述，但在本发明基础上，可以对之作一些修改或改进，这对本领域技术人员而言是显而易见的。因此，在不偏离本发明精神的基础上所做的这些修改或改进，均属于本发明要求保护的范围。

Claims (10)

  1. 一种中药组合物，其特征在于，由包括以下重量份的原料药制备而成：冬虫夏草1～10份，红景天4～20份，锁阳3～15份，牡丹皮2～10份。
  2. 根据权利要求1所述的中药组合物，其特征在于，由以下重量份的原料药制备而成：冬虫夏草1～10份，红景天4～20份，锁阳3～15份，牡丹皮2～10份。
  3. 根据权利要求2所述的中药组合物，其特征在于，由以下重量份的原料药制备而成：冬虫夏草1～4份，红景天4～16份，锁阳3～9份，牡丹皮2～6份；优选地，由以下重量份的原料药制备而成：冬虫夏草3～4份，红景天12～16份，锁阳6～9份，牡丹皮4～6份。
  4. 根据权利要求1～3任意一项所述的中药组合物，其特征在于，所述中药组合物是将各原料药混合后，经溶剂提取制得；优选地，所述各原料药均为干燥药粉。
  5. 根据权利要求4所述的中药组合物，其特征在于，所述中药组合物由包括以下步骤的方法制备而成：将各原料药混合，在溶剂中浸泡、煎煮，过滤留取滤液，浓缩、喷雾干燥。
  6. 根据权利要求5所述的中药组合物，其特征在于，所述溶剂为有机溶剂或水，优选为甲醇、乙醇或水。
  7. 包含权利要求1～6任意一项所述中药组合物的制剂；优选地，所述制剂为粉剂。
  8. 权利要求1～6任意一项所述中药组合物或权利要求7所述制剂在制备治疗肺癌的药物中的用途；
    优选地，所述肺癌为非小细胞肺癌。
  9. 权利要求1～6任意一项所述中药组合物或权利要求7所述制剂在制备用于抑制人大细胞肺癌细胞增殖和/或迁移药物中的用途。
  10. 一种抑制人大细胞肺癌细胞增殖和/或迁移的抑制剂，其特征在于，包含权利要求1～6任意一项所述中药组合物或权利要求7所述 制剂。
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN112057473A (zh) * 2020-09-22 2020-12-11 郭国康 一种关于肺病肺癌增强肺活量的组合物及其制备方法

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101301413A (zh) * 2007-07-31 2008-11-12 四川回春堂生物科技有限公司 虫草红景天胶囊及其制备方法
CN101380350A (zh) * 2008-09-30 2009-03-11 苟春虎 虫草救心复方胶囊

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101301413A (zh) * 2007-07-31 2008-11-12 四川回春堂生物科技有限公司 虫草红景天胶囊及其制备方法
CN101380350A (zh) * 2008-09-30 2009-03-11 苟春虎 虫草救心复方胶囊

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN112057473A (zh) * 2020-09-22 2020-12-11 郭国康 一种关于肺病肺癌增强肺活量的组合物及其制备方法

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