CN103479933A - Traditional Chinese medicine composition used for treating cancer, and preparation method thereof - Google Patents
Traditional Chinese medicine composition used for treating cancer, and preparation method thereof Download PDFInfo
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Abstract
The invention relates to a traditional Chinese medicine composition used for treating cancer. The traditional Chinese medicine composition comprises following raw materials, by weight, 15 portions of herba sarcandrae, 20 portions of radix rehmanniae, 15 portions of parched concha arcae, 30 portions of concha meretricis seu cyclinae, 10 portions of haematitum, 10 portions of radix clematidis, 12 portions of the root of Chinese pulsatilla, 12 portions of ficus carica, 20 portions of dark malt, 10 portions of herba schizonepetae, 30 portions of raw coix seed, 60 portions of willow root, 15 portions of roasted endothelium corneum gigeriae galli, 12 portions of spica prunellae, 40 portions of scouring rush, 20 portions of common macrocarpium fruit pulp, 10 portions of radix saposhnikoviae and 15 portions of semen vaccariae. It is shown by the results of in vivo and in vitro experiments that the traditional Chinese medicine composition possesses obvious inhibition effects on growth of human lung cancer A549 cell line and human lung cancer NCI-H460 cells. The traditional Chinese medicine composition can be used for treating cancer, and especially for treating of lung cancer.
Description
Technical field
The present invention relates to the field of Chinese medicines, be specifically related to a kind of Chinese medicine composition for the treatment of cancer and preparation method thereof.
Background technology
Malignant tumor, also claim cancer, is that phalangeal cell is out of hand, and abnormality proliferation continues growth and then invades and close on normal structure, but and intravascular, lymphatic vessel and body cavity transfer to other positions of health, finally jeopardize human life's a class serious disease.Come second in China's malignant tumor in the various causes of the death, at urban, be discharged to first place; And nearly 20 years, the sickness rate of tumor and mortality rate were obvious ascendant trend, and wherein the lung cancer morbidity rate ranks first.At present, the clinical treatment of tumor is mainly applied the methods such as excision, radiotherapy, chemotherapy, Biotherapeutics and treatment by Chinese herbs.Because the grade malignancy of tumor is high, current treatment means is difficult to obtain good efficacy, leads oncogenic recurrence and transfer, and has the deficiency of some treatment meanss, as the toxic and side effects of chemotherapy usually causes bone marrow depression.Therefore, develop new antitumor drug to the treatment tumor significant.The traditional Chinese medicine of China has very important value on oncotherapy, has accumulated the experience of many preciousnesses, and for performing the operation and postoperative patient, and the aspect such as toxic and side effects has auxiliary therapeutic action in reducing radiotherapy, chemotherapy and Biotherapeutics.Therefore, effectively Chinese medicine compound has been brought into play important effect in clinical treatment tumour.
Strengthening vital QI to eliminate pathogenic factors is the universal rule of Chinese medicine treatment disease, can be used for prevention and the treatment of tumor.Chinese medicine composition of the present invention is comprised of 18 medicines, and mainly for the loss of pulmonary carcinoma lung yin, spleen the moon is depressed, accumulation and obstruction of sputum, qi depression to blood stasis, and the weak gold of soil declines and waits the side's of establishing legislation, is used for the treatment of cancer, especially for treating pulmonary carcinoma.
Summary of the invention
For the deficiencies in the prior art, the invention provides a kind of Chinese medicine composition.This Chinese medicine composition has obvious inhibitory action to the growth of people's lung cancer A549 cell system and people's pulmonary carcinoma NCI-H460 cell, can be used for treating cancer, especially for treatment pulmonary carcinoma.
The present invention relates to a kind of Chinese medicine composition, it is comprised of by corresponding weight portion proportioning following raw material: 15 parts of Herba Sarcandraes, 20 parts, the Radix Rehmanniae, 15 parts of burnt Concha Arcaes, 30 parts of Concha Meretricis seu Cyclinaes, 10 parts of Haematitum, 10 parts of Radix Clematidis, 12 parts of the Radix Pulsatillaes, 12 parts of Fructus Ficis, 20 parts of Fructus Hordei Germinatus (parched to brown)s, 10 parts of Herba Schizonepetaes, 30 parts of Semen Coicis, 60 parts of willow root, process 15 parts of Endothelium corneum, 12 parts of Spica Prunellaes, 40 parts of Herba Equiseti Hiemalis, 20 parts of Fructus Corni, 10 parts of Radix Saposhnikoviaes, 15 parts of Semen Vaccariae.Preferably, this Chinese medicine composition can be comprised of the raw material of following weight: Herba Sarcandrae 15 grams, the Radix Rehmanniae 20 grams, burnt Concha Arcae 15 grams, Concha Meretricis seu Cyclinae 30 grams, Haematitum 10 grams, Radix Clematidis 10 grams, the Radix Pulsatillae 12 grams, Fructus Fici 12 grams, Fructus Hordei Germinatus (parched to brown) 20 grams, Herba Schizonepetae 10 grams, Semen Coicis 30 grams, willow root 60 grams, process Endothelium corneum 15 grams, Spica Prunellae 12 grams, Herba Equiseti Hiemalis 40 grams, Fructus Corni 20 grams, Radix Saposhnikoviae 10 grams, Semen Vaccariae 15 grams.
The present invention provides a kind of method for preparing above-mentioned Chinese medicine composition simultaneously, the method comprises following steps: aforesaid raw material is mixed by corresponding weight proportion, comprise 15 parts of Herba Sarcandraes, 20 parts, the Radix Rehmanniae, 15 parts of burnt Concha Arcaes, 30 parts of Concha Meretricis seu Cyclinaes, 10 parts of Haematitum, 10 parts of Radix Clematidis, 12 parts of the Radix Pulsatillaes, 12 parts of Fructus Ficis, 20 parts of Fructus Hordei Germinatus (parched to brown)s, 10 parts of Herba Schizonepetaes, 30 parts of Semen Coicis, 60 parts of willow root, process 15 parts of Endothelium corneum, 15 parts of 12 parts of Spica Prunellaes, 40 parts of Herba Equiseti Hiemalis, 20 parts of Fructus Corni, 10 parts of Radix Saposhnikoviaes and Semen Vaccariae; First with clear water, rinse twice, residual to remove dust, pesticide, chemical fertilizer; Then it is used to warm water soaking, decoct two times for 100 ℃, become flowing soaking paste, then dilute three times, shake up to obtain extractum; Finally by described extractum cryogenic seal or storage.This extractum can be done medicinal, also can be used for as the experiment of animal tumor suppression.
Particularly, this Chinese medicine composition can be mixed by corresponding weight by following raw materials according, comprises Herba Sarcandrae 15 grams, the Radix Rehmanniae 20 grams, burnt Concha Arcae 15 grams, Concha Meretricis seu Cyclinae 30 grams, Haematitum 10 grams, Radix Clematidis 10 grams, the Radix Pulsatillae 12 grams, Fructus Fici 12 grams, Fructus Hordei Germinatus (parched to brown) 20 grams, Herba Schizonepetae 10 grams, Semen Coicis 30 grams, willow root 60 grams, processs Endothelium corneum 15 grams, Spica Prunellae 12 grams, Herba Equiseti Hiemalis 40 grams, Fructus Corni 20 grams, Radix Saposhnikoviae 10 grams and Semen Vaccariae 15 grams; First with clear water, rinse twice, residual to remove dust, pesticide, chemical fertilizer; Then it is used to warm water soaking, decoct two times for 100 ℃, become flowing soaking paste, then dilute three times, shake up to obtain extractum; Finally by described extractum cryogenic seal or storage.This extractum can be done medicinal, also can be used for as the experiment of animal tumor suppression.
Further, the present invention also provides the working solution be made into by aforementioned extractum.The distilled water that aforementioned extractum is added to certain volume, be mixed with the mother solution that concentration is 100mg/ml, after autoclaving, this mother solution is filtered to 2-3 time with filter, and dilution can be made into certain gradient concentration working solution.But this working solution cryogenic seal or storage, can do medicinal, also can be used for research, as the research of cellular level.
Specifically, described Chinese medicine composition provided by the invention routinely technique add adjuvant to make tablet, capsule, granule, drop pill, micropill, dispersible tablet, oral cavity disintegration tablet, pill, oral liquid; Described adjuvant comprises one or more in solvent, disintegrating agent, correctives, antiseptic, coloring agent, binding agent, lubricant and substrate.
In addition, the invention still further relates to the application of described Chinese medicine composition in preparation treatment cancer drug.More specifically, the present invention relates to the application of described Chinese medicine composition in preparation treatment lung-cancer medicament.
Chinese medicine constituent in the present invention is comprised of 18 flavor Chinese medicines, and mainly for the loss of pulmonary carcinoma lung yin, spleen the moon is depressed, accumulation and obstruction of sputum, qi depression to blood stasis, the weak gold of soil declines and waits the side's of establishing legislation, and its side separates as follows: " lung governing qi; towards hundred arteries and veins ", energy and blood of human body body fluid normally moves the promotion of entirely bad gas.The patients with lung cancer stagnation of QI-blood, must affect the normal defeated cloth in lung Tianjin, lung not cloth Tianjin body fluid stop gathering, smoulder not all rightly, and then it is turbid to be converted into expectorant.Because we have no report, only in the other side, the therapeutic efficiency of each Chinese medicine is expressed as follows:
(1) Herba Sarcandrae: the nature and flavor toil is flat.Enter heart channel, Liver Channel.But clearing away heat and cooling blood, the speckle removing of invigorating blood circulation, dispelling wind and removing obstruction in the collateral, have antitumaous effect;
(2) Radix Rehmanniae: sweet in flavor and cold in property.Enter heart channel, Liver Channel, kidney channel.Heat clearing and blood cooling, YIN nourishing and the production of body fluid promoting, removing blood stasis to promote tissue regeneration, moisturize laxation, pain relieving.Multiplex in treatment yin asthenia generating intrinsic heat, hectic fever due to YIN-deficiency consumptive fever, crimson tongue excessive thirst, a speckle dermexanthesis etc.;
(3) burnt Concha Arcae: nature and flavor are salty flat.Return lung meridian, stomach warp, Liver Channel.The expectorant blood stasis dispelling, hard masses softening and resolving, relieving gastric hyperacidity to alleviate stomachache.For the long-pending knot of pertinacious phlegm, the thickness difficulty is coughed up, goiter, scrofula, abdominal mass mass in the abdomen, stomachache pantothenic acid etc.;
(4) Concha Meretricis seu Cyclinae: nature and flavor are salty flat.Enter heart channel, kidney channel.But clearing away heat and promoting diuresis, softening and eliminating sputum.Be used for the treatment of heat-phlegm and breathe heavily and cough, edema, goiter is gathered, blood agglomeration chest pain etc.;
(5) Haematitum: the property bitter but sweet flavor is flat.Enter Liver Channel, stomach warp, heart born of the same parents warp.But suppressing the hyperactive liver and subsiding YANG, the sending down the abnormal ascending QI hemostasis.For vertigo and tinnitus, pant, eructation, vomiting, singultus, spit blood, epistaxis etc.;
(6) Radix Clematidis: the hot salty temperature of nature and flavor, poisonous.Enter urinary bladder channel.But expelling wind and removing dampness, removing obstruction in the collateral to relieve pain, expectorant water, the loose hypochondriac lump;
(7) Radix Pulsatillae: nature and flavor bitter cold; Return stomach warp, large intestine channel.There is heat-clearing and toxic substances removing, eliminating pathogenic heat from blood to cure dysentery, the effect of removing dampness and killing parasites.Can treat the carbuncle skin ulcer, scrofula etc.;
(8) Fructus Fici: the sweet micro-suffering of nature and flavor is flat, slightly poisonous.Cure mainly swelling and pain of hemorrhoid etc.;
(9) Fructus Hordei Germinatus (parched to brown): nature and flavor are sweet flat.Return spleen channel, stomach warp.But strengthening the spleen and nourishing the stomach, the food stagnation removing food stagnation;
(10) Herba Schizonepetae carbon: the hot puckery slight fever of nature and flavor.Return lung meridian, Liver Channel.But hemostasis with astringents;
(11) Semen Coicis: nature and flavor are cool sweet light.But the spleen invigorating eliminating dampness by diuresis, eliminating impediment antidiarrheal.Can be used for treating edema, dysuria, arthralgia chiefly caused by damp pathogen contracture, diarrhea due to hypofunction of the spleen;
(12) willow root: but expelling wind and removing dampness, reducing swelling and alleviating pain;
(13) process Endothelium corneum: property is hidden sweet cold.Return spleen channel, stomach warp, small intestine meridian, urinary bladder channel.But promoting digestion and invigorating the stomach is aid digestion, arresting seminal emission.Be used for the treatment of dyspepsia, shallow complexion, do not think Na Gu, the disease such as infantile malnutrition, emaciated physique, enlarged abdomen abdominal distention, weakness of the spleen and stomach, food stagnation distension;
(14) Spica Prunellae: the nature and flavor micro-hardship of sweet suffering of trembling with fear, there is the effect of letting out clearly liver-fire, mass dissipating and swelling eliminating, heat-clearing and toxic substances removing, expelling phlegm for arresting cough, cooling blood for hemostasis, be applicable to the diseases such as lymphoid tuberculosis, goiter, acute mastitis.Modern pharmacological research shows, Spica Prunellae has the effect of anticancer;
(15) Herba Equiseti Hiemalis: nature and flavor sweetness and bitterness is flat, nontoxic, is slightly cold.Enter heart channel, Liver Channel, stomach warp, urinary bladder channel.Can stop blooding, liver heat removing and eyesight improving, inducing diuresis for treating stranguria syndrome, be used for the treatment of the diseases such as cough, conjunctival congestion and swelling pain, discharging fresh blood stool;
(16) Fructus Corni: the sour and astringent tepor of nature and flavor.As Liver Channel, kidney channel.But liver and kidney tonifying, restrain astringent or styptic treatment for spontaneous sweating, controlling nocturnal emission with astringent drugs reducing urination, and hidroschesis promotes the production of body fluid.Can be used for soreness of waist and knee joint, profuse sweating collapse, the interior-heat disease such as quench one's thirst;
(17) Radix Saposhnikoviae: property acrid-sweet flavor tepor.Enter urinary bladder channel, lung meridian, spleen channel, Liver Channel.But expelling pathogenic wind from the body surface, removing dampness to relieve pain, spasmolytic are antipruritic.Be used for the treatment of affection of exogenous wind-cold, the headache general pain, rheumatic arthralgia, joint is ached, the stomachache disease such as have loose bowels;
(18) Semen Vaccariae: nature and flavor are bitter flat.Enter Liver Channel, the stomach warp.But promoting the circulation of blood regulating menstruation, reducing swelling and alleviating pain, the lactogenesis of invigorating blood circulation, expedite the emergence of the thymus dredging of stimulating milk secretion, puerperal.Be used for the treatment of the blood stasis amenorrhea, dysmenorrhea, difficult labour, the acute mastitis disease such as swell and ache.
Can increase other adjuvant drugs and/or make medicine on the basis of this prescription, further to improve curative effect.
Advantage of the present invention is that the Chinese medicine composition that provides is in the treatment cancer, particularly treat aspect pulmonary carcinoma and have significant curative effect.The inventor found through experiments, and said composition reaches 70% to the suppression ratio of people's immortalized hepatocyte LO2, to the suppression ratio of nude mice tumor, can reach 75%, and the symptom to patient's pulmonary carcinoma in clinical trial improves significantly and therapeutical effect.
The accompanying drawing explanation
Fig. 1 Chinese medicine composition acts on the morphology photo of people's immortalized hepatocyte LO2 after 48 hours.
Fig. 2 Chinese medicine composition acts on the morphology photo of human lung cancer cell A549 after 48 hours.
Fig. 3 Chinese medicine composition acts on the morphology photo of human lung carcinoma cell NCI-H460 after 48 hours.
Fig. 4 Chinese medicine composition is made the Trypan Blue result (p of employment immortalized hepatocyte LO2 after 48 hours
1=0.0417,
*p<0.05,
*p<0.01,
* *p<0.001).
Fig. 5 Chinese medicine composition effect human lung carcinoma cell trypan blue coloration result (p after 48 hours
1=0.0053, p
2=0.0013; p
3=0.0048).
Trypan Blue (the p of Fig. 6 Chinese medicine composition effect human lung carcinoma cell NCI-H460 after 48 hours
1=0.0075, p
2=0.0158, p
3=0.0167).
MTT testing result (the p of Fig. 7 Chinese medicine composition effect immortalized hepatocyte LO2 after 48 hours
1=0.0163).
Fig. 8 Chinese medicine composition effect human lung cancer cell A549 MTT testing result (p after 48 hours
1=0.017, p
2=0.0070, p
3=0.0061).
MTT result (the p of Fig. 9 Chinese medicine composition effect human lung carcinoma cell NCI-H460 after 48 hours
1=0.0125, p
2=0.0358, p
3=0.0167).
Figure 10 Chinese medicine composition is made the streaming result of employment immortalized hepatocyte LO2 after 48 hours.
The streaming result of Figure 11 Chinese medicine composition effect lung cell A549 after 48 hours.
Figure 12 BALB/C nude mice weight is relation over time.
Figure 13 BALB/C nude mice gross tumor volume is relation over time.
The tumor mass photo (6 every group) of Figure 14 BALB/C nude mice gastric infusion after 45 days.
The tumor mass weight block diagram of Figure 15 BALB/C nude mice gastric infusion after 45 days.
Figure 16 is for the RNA quality analysis result of genechip detection.
The specific embodiment
[1] Chinese medicine composition composition and preparation thereof
Chinese medicine composition by the following raw materials in parts by weight proportioning is: 15 parts of Herba Sarcandraes, 20 parts, the Radix Rehmanniae, 15 parts of burnt Concha Arcaes, 30 parts of Concha Meretricis seu Cyclinaes, 10 parts of Haematitum, 10 parts of Radix Clematidis, 12 parts of the Radix Pulsatillaes, 12 parts of Fructus Ficis, 20 parts of Fructus Hordei Germinatus (parched to brown)s, 10 parts of Herba Schizonepetaes, 30 parts of Semen Coicis, 60 parts of willow root, process 15 parts of Endothelium corneum, 12 parts of Spica Prunellaes, 40 parts of Herba Equiseti Hiemalis, 20 parts of Fructus Corni, 10 parts of Radix Saposhnikoviaes, 15 parts of Semen Vaccariae.
In the present embodiment, described Chinese medicine composition is by Herba Sarcandrae 15 grams, the Radix Rehmanniae 20 grams, burnt Concha Arcae 15 grams, Concha Meretricis seu Cyclinae 30 grams, Haematitum 10 grams, Radix Clematidis 10 grams, the Radix Pulsatillae 12 grams, Fructus Fici 12 grams, Fructus Hordei Germinatus (parched to brown) 20 grams, Herba Schizonepetae 10 grams, Semen Coicis 30 grams, willow root 60 grams, process Endothelium corneum 15 grams, Spica Prunellae 12 grams, Herba Equiseti Hiemalis 40 grams, Fructus Corni 20 grams, Radix Saposhnikoviae 10 grams and Semen Vaccariae 15 grams form.
The preparation of this Chinese medicine composition comprises the following steps:
A) raw material of aforesaid kind and component first rinses twice with clear water, residual to remove dust, pesticide, chemical fertilizer;
B) use warm water soaking, decoct two times for 100 ℃, become flowing soaking paste, dilute three times, shake up to obtain extractum;
C) by described extractum cryogenic seal or storage.
This extractum has therapeutic effect, can, for the experiment of animal tumor suppression, also can do medicinal.
Aforementioned extractum is added to the distilled water of certain volume, be mixed with mother solution, concentration is 100mg/ml, after autoclaving, with filter, filters 2-3 time, can be diluted to certain gradient concentration working solution.But this work is cryogenic seal or storage also, can be used for research, for example research of cellular level.
[2] process for preparing raw material
1. please medical material base supply business supply medicine by prescription.After medicine is bought, at first check into the formality of medicine channel, understand the place of production; Whether next side's of checking medicine conforms to; Screening, determine that whether medicine has impurity, the adulterated and medicine that goes mouldy, and could put processing in storage after errorless again.
2. according to quantity medicine is dropped into to reactor, adding water cleans two times, purpose is to remove ash, keep the medicine cleaning, reduce agriculture chemical residual simultaneously, stir and also can reduce content of beary metal in the fine laundering process in addition, make the various content of beary metal such as the rear lead of medicine processing, arsenic meet enterprise, national standard fully, ensure drug quality.
3. add medicine to ready after, adopt 100 ℃ of high temperature to decoct secondaries, decoct two hours at every turn, add for the first time 10 times of decoctings and boil, add for the second time six times of decoctings and boil, then extract decoction liquor and pour in cylinder, by the liquid spray drying, be powder again, in the sealing bag of packing into after finally xeraphium being harvested and stored.
[3] tablet machining process
1. by dry semi-finished product powder, add adjuvant pelletize, the tablettings such as corn starch, stearic acid U.S., microcrystalline Cellulose, silicon dioxide.
2. after medicine processing, every drug weight is 550 milligrams, and wherein the crude drug powder is that 420 milligrams, corn starch are 90 milligrams, 5 milligrams of silica 1s, U.S. 5 milligrams of stearic acid.
[4] dry capsule prilling process
1. get 300 milligrams of spray drying powder extract powders, 60 milligrams of microcrystalline Cellulose, 5 milligrams of silica 1s, U.S. 5 milligrams of stearic acid, add up to 400 milligrams, the capsule of packing into after pelletize No. 0, by extract powder and microcrystalline Cellulose mix homogeneously; Add 80% ethanol to make suitable soft material, 16 order nylon mesh are granulated; 50～60 ℃ of dryings, 18 order nylon mesh granulate, add methylol ingot powder sodium, silicon dioxide, stearic acid U.S., mix homogeneously during granulate; The capsule of packing into No. 0, refill and airtight.
[5] suspension and grain ball processing method
The method of suspension processing is that after the secondary decoction liquor is concentrated, then, in the after heat loading sealing Packaging Bottle of heating, then heating, vacuum is processed and got final product
[6] drop pill processing method
Drop pill be by spray drying after centrifuge is processed, the impurity such as two oxycarbides are removed, remaining micropowder adds adjuvant, can be processed into drop pill.
Embodiment 2: pharmacodynamic study
[1] experiment material and instrument and equipment
100% ethanol (Tianjin chemical reagent wholesaling firm), hyclone (Invitrogen, USA), RPMI1640(Invitrogen, USA), pancreatin (giving birth to work biological engineering (Shanghai) limited company), trypan blue (the prosperous Bioisystech Co., Ltd of Beijing ancient cooking vessel state), the MTT(full name is 3-(4, 5)-dimethylthiahiazo (z-y1)-3, 5-di-phenytetrazoliumromide, 3-(4, 5-dimethylthiazole-2)-2, 5-diphenyl tetrazole bromine salt, the trade name tetrazolium bromide, Thermo, USA), the DMSO(dimethyl sulfoxide, the prosperous Bioisystech Co., Ltd of Beijing ancient cooking vessel state), propidium iodide (Sigma, USA), blood counting chamber (Shanghai refinement biochemical reagents company limited), low speed desk centrifuge (Hunan, Changsha instrument centrifuge instrument company limited), enzyme-linked immunosorbent assay instrument (BIO-RAD, USA), flow cytometer (BD FACSDiVa, USA), decolorization swinging table (the development in science and technology company limited is helped in moral sky, Beijing), CO2 incubator (sanyo, Japan), inverted microscope (OLYMPUS, Japan).
[2] cell culture
Non-small cell lung carcinoma cell line A549, the sclc cell line NCI-H460 of the National People's Congress and people's immortalized hepatocyte strain LO2 are purchased from Nanjing KaiJi Biology Science Development Co., Ltd.The application of above-mentioned cell strain is the 5%CO at 37 ℃ containing the RPMI1640 culture medium (Gibco, USA) of 10% hyclone (HyClone, Australia)
2in incubator, cultivate.
[3] statistical method
Adopt Student ' s t check to carry out statistical analysis to data, P<0.05 and P<0.01 is significant difference.
[4] morphological observation
1. prepare the Chinese medicine mother solution of certain density Chinese medicine composition, autoclaving, filtering and impurity removing 2～3 times, be diluted to a series of gradient concentrations (0.01,0.1,1.0,5.0 and 10.0mg/ml) standby; Cell strain adopts people's lung cancer A549 cell system, people's pulmonary carcinoma NCI-H460 cell and people's immortalized hepatocyte strain LO2;
2. the cell that will be grown in logarithmic (log) phase spreads 6 orifice plates, when putting into incubator and being cultured to cell and growing to 80%～90%, adds respectively the Chinese medicine 200 μ l of variable concentrations, and wherein matched group adds 200 μ l PBS;
3. incubator is cultivated 48 hours, the impact of micro-Microscopic observation variable concentrations medicine (0.01,0.1,1.0,5.0 and 10.0mg/ml) on cell, and take pictures;
4. observed result:
After the Chinese medicine composition of 1mg/ml concentration acts on immortalized hepatocyte LO2 cell, form, observation of cell is without significant change, and when drug level increases to 5mg/ml, the LO2 cell occurs dead and comes off; People's lung cancer A549 cell about 1/3 cell survival under the drug level of 1mg/ml, and seldom cell survival is almost arranged under the drug level of 5mg/ml and 10mg/ml; People's pulmonary carcinoma NCI-H460 cell and A549 cell situation are similar, even more responsive than A549 cell when low concentration.
[5] Trypan Blue
1. normal living cells, the after birth structural integrity, can repel trypan blue, makes it to enter in born of the same parents; And the incomplete cell of loss of activity or cell membrane, the permeability of after birth increases, and can be dyed blueness by trypan blue.It has been generally acknowledged that the cell membrane integrity forfeiture, can think that cell is dead, this is contrary with the dimethyl diaminophenazine chloride effect.Therefore, can distinguish very easy, rapidly living cells and dead cell by Trypan Blue;
2. the cell that will be grown in logarithmic (log) phase spreads 6 orifice plates, when putting into incubator and being cultured to cell and growing to 80%～90%, adds respectively the Chinese medicine 200 μ l(0.01 of variable concentrations, 0.1,1.0,5.0 and 10.0mg/ml), wherein matched group adds 200 μ l PBS;
3. incubator is cultivated 48 hours, the impact of micro-Microscopic observation variable concentrations medicine on cell.Collect respectively supernatant, pancreatin digests respectively the cell that different pharmaceutical concentration is processed, the cell that in merging, cleer and peaceful digestion is got off; Centrifugal 10 minutes of 800rpm, abandon supernatant; Add 5ml PBS resuspended, centrifugal 10 minutes of 800rpm, abandon supernatant; Add 1ml PBS resuspended;
4. the trypan blue solution of getting respectively 10 μ l sample solutions and 0.4% mixes dyeing 3～6 minutes, then carries out blood counting chamber living cell counting and dead cell number, and painted is dead cell, and not having painted is living cells; Every strain cell line at least repeats 3 times;
5. experimental result:
Table 1 people immortalized hepatocyte is LO2 trypan blue count results
| Concentration mg/ |
0 | 0.01 | 0.1 | 1 | 5 | 10 |
| |
48.5 | 52 | 53 | 53.5 | 12 | 3 |
Table 2 people lung cancer A549 cell is the trypan blue count results
| Concentration mg/ |
0 | 0.01 | 0.1 | 1 | 5 | 10 |
| |
55.4 | 47.8 | 43.1 | 14.7 | 9.3 | 6.3 |
Table 3 people pulmonary carcinoma NCI-H460 cell line trypan blue count results
| Concentration mg/ |
0 | 0.01 | 0.1 | 1 | 5 | 10 |
| |
103.5 | 100.5 | 58.5 | 26 | 6.5 | 3.5 |
6. adopt the Logit method to calculate the medium effective concentration IC of Chinese medicine to the A549 cell
50value, IC
50=0.369mg/ml.
[6] mtt assay detects
1.MTT method claims again the MTT colorimetry, is a kind of method that detects cell survival and growth.Its detection principle is that the succinate dehydrogenase in the living cells mitochondrion can make exogenous MTT be reduced to water-insoluble bluish violet crystallization first a ceremonial jade-ladle, used in libation and be deposited in cell, and dead cell is without this function.First a ceremonial jade-ladle, used in libation in dimethyl sulfoxide (DMSO) energy dissolved cell, measure its absorbance value with enzyme-linked immunosorbent assay instrument at 490nm wavelength place, can indirectly reflect living cells quantity.In certain cell number scope, the amount that the MTT crystallization forms is directly proportional to cell number;
2. the exponential phase cell is spread to 6 orifice plates, when putting into incubator and being cultured to cell and growing to 80%～90%, adds respectively the Chinese medicine 200 μ l(0.01 of variable concentrations, 0.1,1.0,5.0 and 10.0mg/ml), wherein matched group adds 200 μ l PBS;
3. incubator is cultivated 48 hours, the impact of micro-Microscopic observation variable concentrations medicine on cell; Collect respectively supernatant, pancreatin digests respectively the cell that different pharmaceutical concentration is processed, the cell that in merging, cleer and peaceful digestion is got off; Centrifugal 10 minutes of 800rpm, abandon supernatant; Add 5ml PBS resuspended, centrifugal 10 minutes of 800rpm, abandon supernatant; Repeating 2～3 times does not have color to resuspended liquid, adds 1ml PBS resuspended;
4. spread 96 orifice plates, every hole adds 100 μ l culture medium, 6～8 repetitions of each concentration; Add 15 μ l MTT(5mg/ml again in every hole), put incubator and cultivate 3.5～4 hours; Suck supernatant with the 1ml syringe, in every hole, add 150 μ l DMSO, on shaking table, the 150rpm vibration is 15 minutes, surveys OD
490-620;
5. experimental result:
Table 4 people immortalized hepatocyte is the MTT testing result of LO2
| Concentration mg/ |
0 | 0.01 | 0.1 | 1 | 5 | 10 |
| OD 490-620 | 0.252 | 0.270 | 0.269 | 0.274 | 0.071 | -0.104 |
Table 5 people lung cancer A549 cell is the MTT testing result
| Concentration mg/ |
0 | 0.01 | 0.1 | 1 | 5 | 10 |
| OD 490-620 | 1.078 | 1.047 | 0.857 | 0.210 | 0.147 | 0.079 |
Table 6 people pulmonary carcinoma NCI-H460 cell line MTT result
| Concentration mg/ |
0 | 0.01 | 0.1 | 1 | 5 | 10 |
| OD 490-620 | 1.073 | 0.832 | 0.722 | 0.594 | 0.399 | 0.038 |
[7] fluidic cell is learned and is detected
1. the synchronization process cell, process the A549 cell with the Chinese medicine composition of 0mg/ml, 1mg/ml concentration respectively; Process NCI-H460 cell, drug effect 48 hours with the Chinese medicine composition of 0mg/ml, 0.1mg/ml concentration;
2. trypsinization attached cell, PBS dispels separation, collects (15ml centrifuge tube) in centrifuge tube; Centrifugal 10 minutes of 800rpm, abandon supernatant; Add 5ml PBS resuspended, centrifugal 5 minutes of 800rpm, abandon supernatant; Add 5ml PBS resuspended, centrifugal 5 minutes of 800rpm, abandon supernatant; The ethanol that adds rapidly 1ml pre-cooling (20 ℃) 75%, piping and druming cell dispersion (when Stirring prevents conglomeration, be very slow add ethanol); 4 ℃ are fixedly spent the night, and the resuspended precipitation of 900 μ l PBS adds 100 μ l RNase A(not add), hatch 30 minutes to 1 hour for 37 ℃; Add 10 μ l propidium iodides (concentration is 5mg/ml), 20 minutes on ice, lucifuge, room temperature 5-10 minute, flow cytometer showed (filtering with 400 order nylon membranes);
3. experimental result:
With 0mg/ml drug treating immortalized hepatocyte (LO2), S phase LO2 cell proportion is 31.14%; By the drug treating of 1mg/ml, S phase LO2 cell proportion is 24.52%, and S phase cell proportion reduces compared with the control, and the medicine of 1mg/ml has no significant effect thin LO2 born of the same parents; With 0mg/ml drug treating human lung cancer cell line A549, S phase cell proportion is 29.94%; By the drug treating of 0.1mg/ml, S phase cell proportion is 8.95%, and S phase cell proportion reduces a lot compared with the control; Therefore, the medicine of 0.1mg/ml has obvious inhibitory action to the propagation of A549 cell.
[8] animal nude mice heteroplastic transplantation model
1. experimental technique: by 1 * 10
7a549 cell inoculation nude mice, get the tumor tissue piece after becoming tumor, tumor tissue is cut into to big or small about 1.5mm
3, with the trocar be inoculated in animal one side or bilateral axillary regions subcutaneous, totally 12 of tieback nude mices, be divided into two groups (experimental group and matched groups), 6 every group.When the tumor BOB(beginning of block) increases, (approximately after 1 week), start to apply the irrigation stomach device medicine feed, administration every day 1 time, and each 0.3ml, measure Mouse Weight and tumor block size record every day.After administration 45 days, put to death mice.Get the tumor measurement volumes, weigh and take pictures.
2. experimental result
The therapeutic evaluation formula: in tumor control rate=(C-T)/C * 100% formula, T is the average tumor weight of administration group; C is the average tumor weight of matched group.The demonstration of table 7 result, tumour inhibiting rate is 75%.
Table 7 model in nude mice administration tumor mass weight after 45 days
| Group | Tumor heavy (mg) | Tumour inhibiting rate |
| Matched group (CG) | 23.52 | - |
| Experimental group (EG) | 5.87 | 75% |
[1] total RNA sample preparation methods following (attached cell lung cancer A549 cell system):
1. the lung cancer A549 cell of cultivation is cultivated and grown to exponential phase, experimental group adds the Chinese medicine composition 10 μ l of 100mg/ml, and matched group adds PBS10 μ l; Put into incubator and continue to cultivate, drug effect 48 hours;
2. discard culture fluid, PBS cleans 2～3 times; Add 1.5ml TRLzol reagent respectively in culture bottle, repeatedly beat with 1ml rifle head, the culture bottle surface of TRLzol contact all cells is fully digested; Transfer in the test tube of RNase-free and repeatedly lash cell until cannot see agglomerating cell lump with disposable syringe, whole solution should be the refrigerant and state of thickness not;
3. put into ultra-low temp and preserve, then deliver to Boao Biological Co., Ltd and carry out the genechip detection experiment.
[2] experimental result:
Gene chip (Gene Chip, DNA Chip), claim again DNA microarray (DNA Microarray), refers to according to precalculated position and be fixed on solid phase carrier the microarray array that ten million nucleic acid molecules in small size forms very much.Under certain condition, the nucleic acid molecules on carrier can with the nucleic acid fragment hybridization of sequence complementation from sample.If the nucleic acid fragment in sample is carried out to labelling, on special-purpose chip reading instrument, just hybridization signal can be detected.
Biochip technology mainly comprises four key steps: chip preparation, sample preparation, hybridization and signal detection and interpretation of result.Chip gene expression profile is to adopt cDNA or the general acid fragment of few core to make probe, is solidificated on chip; The mRNA of testing sample (processed group) and control sample is carried out to labelling with two kinds of different fluorescence molecules, then with chip, hybridized simultaneously, the ratio of the fluorescence intensity by analyzing two kinds of samples and probe hybridization, carry out the variation of gene expression detection level.This chip gene expression profile detects and is completed by Boao Biological Co., Ltd.
The total RNA sample prepared previously is transported to Boao Biological Co., Ltd with liquid nitrogen, does the chip gene expression profile analysis, the sample quality inspection is described as follows:
Table 8 sample quality inspection result
| Sample | A 260/280 | A 260/230 | Concentration (μ g/ μ L) | Total amount (μ g) | Sample quality |
| 0.1mg/ml | 1.98 | 1.99 | 0.708 | 35.40 | RNA is more complete |
| 0mg/ml | 1.97 | 1.58 | 0.884 | 44.19 | RNA is more complete |
RNA purity: RNA sample A
260/280>1.70, meet rich chip of expression spectrum requirement of experiment difficult to understand.The RNA total amount: RNA sample total amount>8 μ g, total amount meets rich chip of expression spectrum requirement of experiment difficult to understand.The RNA integrity: through electrophoresis detection, the RNA sample strip is more clear, and the brightness of 28S ﹕ 18S rRNA electrophoretic band is greater than 1 ﹕ 1, meets rich chip of expression spectrum requirement of experiment difficult to understand.In sum: RNA sample purity, total amount and integrity meet rich chip of expression spectrum requirement of experiment difficult to understand, can continue follow-up chip experiment.The gene chip results that provide according to Boao Biological Co., Ltd, we apply the molecule annotation system MAS3.0 provided of Boao Biological Co., Ltd to data analysis.
The differential expression result shows: in administration group (0.1mg/ml) vs matched group, difference expression gene has 1103, wherein the upper gene of lowering that is in harmonious proportion is respectively 490 and 613, screening criteria is Ratio(administration group/matched group) be greater than 2.5, be less than 0.5, the P value is less than the gene that 0.05 gene is significant difference simultaneously.
The enrichment analysis result of Pathway shows: in administration group (0.1mg/ml) vs matched group, the signal path of significant difference mainly comprises oxidative phosphorylation (Oxidative phosphorylation), ribosome (Ribosome), cell cycle (Cell cycle), Heparan sulfate synthesizes (Heparan sulfate biosynthesis), the O-glycosyl synthesizes (O-Glycan biosynthesis), DNA polymerase(DNA polymerase), Wnt signal path (Wnt signaling pathway), MAPK signal path (MAPK signaling pathway), TGF-signal beta Signal Transduction Pathways (TGF-beta signaling pathway), closely connect (Tight junction), Adipocyte Factor signal transduction pathway (Adipocytokine signaling pathway), cell adhesion (Cell adhesion molecules), cytokine interphase interaction (Cytokine-cytokine receptor interaction), ECM acceptor interaction (ECM-receptor interaction), stick together speckle (Focal adhesion), gap connects (Gap junction), apoptosis (Apoptosis), Jak-STAT signal path (Jak-STAT signaling pathway), p53 signal path (p53signaling pathway), PPAR signal path (PPAR signaling pathway), glycosyl-phosphatidyl inositol anchor is synthetic (Glycosylphosphatidylinositol (GPI)-anchor biosynthesis) etc. surely.The related gene expression level changes, and for example: PCNA lowers 0.29 times, and WNT6 lowers 0.22 times, and SKP2 lowers 0.29 times, and TNF raises 4.16 times, etc.
In the present invention, select to meet the case of pulmonary cancer diagnosis standard, by every day, take pharmaceutical composition of the present invention three times.
Now enumerate some model cases as follows:
Zhang, man, 65 years old.Find pulmonary shadow during in May, 2009 health check-up, the prompting of check on March 3rd, 2010 breast CT, make a definite diagnosis pulmonary carcinoma, and carry out thoracoscope under general anesthesia on March 22 in 2010, lower inferior lobe of right lung excision, hilus pulumonis, mediastinal lymphadenectomy art, postoperatively do scorching anti symptom treatment, recover smoothly.Ordinary circumstance while leaving hospital, without uncomfortable main suit, without heating, otch is taken out stitches, II/ first healing, the check hemogram is normal, and rabat shows Postoperative changes, and postoperative do not have a chemotherapy.On April 7th, 2010 starts to take medicine, and takes medicine continuously 10 months, and every day three times, the rear obvious body constitution of having improved of taking medicine, to JIUYUE in 2011 1 day, check and be showed no recurrence for 5 times and shift.
Hu, man, 60 years old.Cough appears in June, 2012, and chest pain, tell a large amount of brown expectorant, dyspnea, and without appetite, the symptoms such as sleep difference.CT is in hospital, and the imaging diagnosis report: inferior lobe of right lung is not opened, right lung inflammation, right pleural effusion.Make a definite diagnosis pulmonary carcinoma.After reaffirm as pulmonary carcinoma, because of health and economic cause abandoning cure.On July 21st, 2012 starts to take medicine.Medication after 20 days the expectorant color transfer yellow sticky expectorant to, medication 40 days, without any discomfort, defecation is normal, sleep, appetite is good, appetite is normal, does not substantially cough, amount of expectoration reduces, the expectorant yellow skin glues, apoplexy due to phlegm is once in a while with any scarlet blood streak.
Experimental data shows: take Chinese medicine composition of the present invention, patient's the state of an illness is had and alleviates or therapeutical effect.Therefore, these experiments further illustrate the present invention has good pharmacodynamic action to treatment cancer, particularly pulmonary carcinoma, is the treatment cancer, particularly treats pulmonary carcinoma Chinese medicine preparation preferably.
Claims (6)
1. a Chinese medicine composition for the treatment of cancer, be comprised of by corresponding weight portion proportioning following raw material: 15 parts of Herba Sarcandraes, 20 parts, the Radix Rehmanniae, 15 parts of burnt Concha Arcaes, 30 parts of Concha Meretricis seu Cyclinaes, 10 parts of Haematitum, 10 parts of Radix Clematidis, 12 parts of the Radix Pulsatillaes, 12 parts of Fructus Ficis, 20 parts of Fructus Hordei Germinatus (parched to brown)s, 10 parts of Herba Schizonepetaes, 30 parts of Semen Coicis, 60 parts of willow root, process 15 parts of Endothelium corneum, 12 parts of Spica Prunellaes, 40 parts of Herba Equiseti Hiemalis, 20 parts of Fructus Corni, 10 parts of Radix Saposhnikoviaes, 15 parts of Semen Vaccariae.
2. Chinese medicine composition claimed in claim 1, described Chinese medicine composition is made by corresponding weight by following raw material: Herba Sarcandrae 15 grams, the Radix Rehmanniae 20 grams, burnt Concha Arcae 15 grams, Concha Meretricis seu Cyclinae 30 grams, Haematitum 10 grams, Radix Clematidis 10 grams, the Radix Pulsatillae 12 grams, Fructus Fici 12 grams, Fructus Hordei Germinatus (parched to brown) 20 grams, Herba Schizonepetae 10 grams, Semen Coicis 30 grams, willow root 60 grams, process Endothelium corneum 15 grams, Spica Prunellae 12 grams, Herba Equiseti Hiemalis 40 grams, Fructus Corni 20 grams, Radix Saposhnikoviae 10 grams, Semen Vaccariae 15 grams.
3. a method for preparing claim 1 or 2 described Chinese medicine composition comprises the following steps:
A) clear water for the raw material of the described kind of claim 1 and component is rinsed to twice, residual to remove dust, pesticide, chemical fertilizer;
B) use warm water soaking, decoct two times for 100 ℃, become flowing soaking paste, dilute three times, shake up to obtain extractum;
C) by described extractum cryogenic seal or storage.
4. the described Chinese medicine composition of claim 1 or 2, described pharmaceutical composition technique routinely adds adjuvant to make tablet, capsule, granule, drop pill, micropill, dispersible tablet, oral cavity disintegration tablet, pill or oral liquid, and described adjuvant comprises one or more in solvent, disintegrating agent, correctives, antiseptic, coloring agent, binding agent, lubricant and substrate.
5. the application of the described Chinese medicine composition of claim 1 or 2 in preparation treatment cancer drug.
6. the application of the described Chinese medicine composition of claim 1 or 2 in preparation treatment lung-cancer medicament.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1483423A (en) * | 2002-09-19 | 2004-03-24 | 于天德 | Traditional Chinese medicine for treating cancer and preparation method thereof |
| CN1927307A (en) * | 2006-08-29 | 2007-03-14 | 张龙标 | Chinese herbal medicine prescription for treating cancer |
| CN101670071A (en) * | 2009-09-25 | 2010-03-17 | 卢速江 | Traditional Chinese medicine preparation for treating cancer and preparation method thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1483423A (en) * | 2002-09-19 | 2004-03-24 | 于天德 | Traditional Chinese medicine for treating cancer and preparation method thereof |
| CN1927307A (en) * | 2006-08-29 | 2007-03-14 | 张龙标 | Chinese herbal medicine prescription for treating cancer |
| CN101670071A (en) * | 2009-09-25 | 2010-03-17 | 卢速江 | Traditional Chinese medicine preparation for treating cancer and preparation method thereof |
Non-Patent Citations (2)
| Title |
|---|
| 姚翔等: "中医药在肺癌综合治疗中的应用概况", 《甘肃中医》 * |
| 姜胜攀等: "杨新中治疗肺癌验案举隅", 《湖北中医杂志》 * |
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