CN104857018A - Application of raspberry polysaccharide and chemotherapeutics in preparation of anti-tumor combined drug - Google Patents
Application of raspberry polysaccharide and chemotherapeutics in preparation of anti-tumor combined drug Download PDFInfo
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Abstract
The invention provides application of raspberry polysaccharide and chemotherapeutics in the preparation of an anti-tumor combined drug. Pharmacodynamic results show that raspberry polysaccharide has the advantages of improving immunity, enhancing the anti-tumor effect of chemical medicament, and reducing toxic and side effects.
Description
Technical field
The invention belongs to medical sci-tech field, be specifically related to the drug combination of Fructus Rubi polysaccharide and chemotherapeutics.
Technical background
Fructus Rubi (Rubus idaeus L.) has another name called Rubus corchorifolius Linn. f., Fructus Rubi corchorifolii Immaturus, Fructus Rubi etc., belongs to Rosaceae rubus.Fructus Rubi is mainly distributed in temperate zone, the Northern Hemisphere and frigid zone, and its fruit is the little aggregate fruit of many slurries, the sweet micro-acid of taste.Fructus Rubi contains polysaccharide, organic acid, flavone, anthocyanidin, and the various active compositions such as polyphenol, vitamin, mineral element, volatile oil, tannin, have high nutritive value and medical care effect, be described as " gold fruit ".
In Fructus Rubi, polyoses content is high, and polysaccharide yield can, up to 12%, be a kind of desirable Polyose extraction raw material.Research finds, polysaccharide has and regulates the multiple physiologically active such as immunity of organisms, antitumor, antioxidation, antibacterial, radioprotective, resisting fatigue, reducing blood sugar and blood fat.Fructus Rubi product is in the market mainly food, as taken Fructus Rubi as the fruit juice, fruit jam, fruit wine etc. of main component.Extract in Fructus Rubi and have the active polysaccharide be worth, and the example being applied to health product is considerably less, and with the buccal tablet researches that Fructus Rubi polysaccharide is active component.
Malignant tumor remains the major disease of serious threat human life.Nearest statistics shows, China annual newfound cancer patient about 2,000 ten thousand people, wherein 1,500,000 people die from cancer.The death toll of cancer accounts for 1/5 of total death toll.
Current, the most frequently used method of Therapeutic cancer is still chemotherapy.But, due to the nonspecific cytotoxicity of chemicals, the side effect that bone marrow depression, mucositis, dermatitis, diarrhoea etc. are serious can be produced.In addition, tumor patient easily develops immunity to drugs after repeatedly chemotherapy, thus the effect of chemotherapy is had a greatly reduced quality.The maximum drawback of the embolic chemotherapy applied at present is which inhibits the immune system of body, makes cancerous cell can escape the immune surveillance of body easily thus become and have more menace.Therefore, safer and more effective Therapeutic Method is found extremely urgent.In recent years, the immunotherapy of tumor has become the new focus of oncotherapy.So-called immunization therapy, broadly refers to a kind of Therapeutic Method coming anti-malignant tumor by transferring body self immune system.At present, this type of medicine existing enters clinical investigation phase.
Polysaccharide is that one is extensively present in animals and plants, has the biomacromolecule of multiple physiologically active.In recent years, due to natural, nontoxic characteristic and good curative effect, polysaccharide has become the new midicinal resources of a class great exploitation potential for its.Immunomodulating is the topmost physiologically active of polysaccharide.Research shows, lentinan, ganoderan, sea grass polysaccharide, flat beautiful snail polysaccharide etc. all have good immunological enhancement.But make the example for treating tumor also fewer polysaccharide and chemotherapy drugs in combination.
Summary of the invention
The object of the present invention is to provide the novelty teabag of Fructus Rubi polysaccharide.
Particularly, the invention provides Fructus Rubi polysaccharide and chemotherapeutics is preparing the purposes in antineoplastic combination medicine.
Wherein, described chemotherapeutics is selected from paclitaxel, cisplatin or/and docetaxel.
Wherein, described tumor is selected from melanoma, pulmonary carcinoma, hepatocarcinoma, gastric cancer, breast carcinoma, renal carcinoma, carcinoma of endometrium or colon cancer.
Wherein, described chemotherapeutics adopts injection type.Wherein, described injection type can be selected from the usual way in the various tumor therapeutic procedures such as subcutaneous injection, intramuscular injection or intravenous injection.
Wherein, described Fructus Rubi polysaccharide adopts peroral dosage form.Wherein, described peroral dosage form is selected from tablet, capsule, pill, granule, powder, extractum or oral liquid.
Further, Fructus Rubi polysaccharide is 20 ~ 40: 1w/w with the using dosage ratio of chemotherapeutics, and certainly, this ratio is the confirmation carried out with zoopery, may occur the deviation of dosage usage ratio in the actual use procedure of human body.
Wherein, in described Fructus Rubi polysaccharide, purity of polysaccharide is more than 50%; Further, purity of polysaccharide is more than 80%; Further, purity of polysaccharide is more than 85%; Preferably, purity of polysaccharide is 85 ~ 95%w/w, if moderate purity of the present invention is 90 ~ 92%.Above-mentioned purity adopts Phenol-sulphate acid method to measure.
Wherein, described Fructus Rubi polysaccharide adopts decoction and alcohol sedimentation technique to prepare.
Further, described decoction and alcohol sedimentation technique concrete operations are as follows: get dry Fructus Rubi, after petroleum ether or ether defatting, with concentration 75 ~ 95%v/v ethanol extraction, filter, filtering residue extracting in water, after water intaking extract is concentrated, add ethanol and reach 75% ~ 85% to alcohol content, cool, leave standstill, get solid content and be Fructus Rubi polysaccharide.
Wherein, Fructus Rubi removes seed or does not remove seed before extracting.
Wherein, described Fructus Rubi is the fruit of Rosaceae rubus.
Further, described Fructus Rubi is selected from red raspberry or black raspberry.
The present invention studies discovery, Fructus Rubi polysaccharide and chemotherapy drugs in combination is used, and for the treatment of malignant tumor, has raising immunity of organisms, strengthens chemicals antitumous effect, reduces the advantages such as its toxic and side effects.
Accompanying drawing illustrates:
Accompanying drawing 1 chemotherapeutics ~ Fructus Rubi polysaccharide combines the impact on tumor-bearing mice liver organization form;
Accompanying drawing 2 chemotherapeutics ~ Fructus Rubi polysaccharide combines the impact on tumor-bearing mice renal tissue form;
Accompanying drawing 3 chemotherapeutics ~ Fructus Rubi polysaccharide combines the impact on C57BL/6 mouse spleen index;
Accompanying drawing 4 Fructus Rubi polysaccharide is on the impact of tumor-bearing mice splenocyte in-vitro multiplication;
Accompanying drawing 5 chemotherapeutics ~ Fructus Rubi polysaccharide combines the impact on Related Immune Enzyme Activity in tumor-bearing mice splenocyte;
Accompanying drawing 6 chemotherapeutics ~ Fructus Rubi polysaccharide combines the impact on CD4, CD8, IL ~ 2 in tumor-bearing mice serum, INF ~ γ and TNF ~ alpha content.
Detailed description of the invention
The Fructus Rubi polysaccharide used in the specific embodiment of the invention adopts conventional Polyose extraction, way of purification prepares, and concrete operations are as follows in the present invention:
Prepared by Fructus Rubi polysaccharide: get dry Fructus Rubi, after seed is removed in pulverizing, add 5 times of petroleum ether (boiling range 60 ~ 90 DEG C) defat, 75% ~ 95%v/v ethanol is added in 60 DEG C of backflows after eliminating petroleum ether, filtering residue after filtering is added 10 times of pure water supersound extraction (power 60W, temperature: 80 DEG C, processing time: 100min), centrifuging and taking supernatant, filtering residue extracts twice again by old terms, merging filtrate, be evaporated to 1/4 of original volume, adding 95% ethanol makes solution alcohol content reach 75%v/v, in 4 degrees Celsius of standing 24h, collecting by filtration filtering residue, filtering residue dries to obtain Fructus Rubi polysaccharide.
After measured, Fructus Rubi polysaccharide yield prepared by the present invention is 10.4% ~ 11.6%, and obtaining purity of polysaccharide through Phenol-sulphate acid method detection is 90.8% ~ 91.2%.
The present invention has carried out the pharmacodynamic study of adjuvant for chemotherapy of tumour from tumor-inhibiting action, attenuation synergistic and immunomodulating three aspects main component Fructus Rubi polysaccharide to Fructus Rubi polysaccharide buccal tablet, and in each embodiment, Fructus Rubi polysaccharide dosage is all in polysaccharide.
Embodiment 1
Docetaxel ~ Fructus Rubi polysaccharide combines the inhibitory action to the growth of melanin tumour b16 F10 mouse xenograft tumor
Take the logarithm the melanoma cell B16F10 of trophophase, is aseptically prepared into 5 × 10
7/ ml cell suspension, is inoculated in C57BL/6 right side of mice axillary fossa with 0.1ml subcutaneous.Use vernier caliper measurement mice-transplanted tumor diameter after 5 days, treat tumor growth to 60 ~ 90mm
3rear animal random packet.Use the method measuring tumor footpath, dynamically observe the antitumous effect of tested medicine.The pendulous frequency of diameter of tumor is every 2 days 1 time, and each measurement also needs weighing Mus heavy simultaneously.Grouping and administering mode as follows: docetaxel group adopt subcutaneous injection, 10mg/kg, administration 1 time in every three days.High administering drug combinations group in docetaxel ~ Fructus Rubi polysaccharide, docetaxel adopts subcutaneous injection, 10mg/kg, administration 1 time in every three days, and Fructus Rubi polysaccharide adopts gavage, and dosage is respectively 200mg/kg, 400mg/kg, administration every day 1 time.Negative control group injection normal saline, every day 1 time.Gross tumor volume computing formula: TV=0.52 × a × b
2, wherein a, b represent length and width respectively.Result according to measuring calculates relative tumour volume.The evaluation index of anti-tumor activity is Relative tumor rate of increase T/C (%), computing formula: T/C (%)=T
rTV/ C
rTV× 100%, T
rTV: treatment group RTV; C
rTV: negative control group RTV.
The inhibitory action that table 1. docetaxel ~ Fructus Rubi polysaccharide drug combination grows C57BL/6 mouse xenograft tumor
Result: in table 1, the tumour inhibiting rate of docetaxel 10mg/kg group to melanin tumour b16 F10 mice-transplanted tumor is that the tumour inhibiting rate of 66.49% administering drug combinations group to melanin tumour b16 F10 mice-transplanted tumor is respectively 70.90% and 74.30%.During conbined usage, melanomatous inhibition is used alone higher than docetaxel.And docetaxel toxicity is comparatively large, the weight of animals declines obviously, and in experimentation, animal has death.After both conbined usage, without obvious decline, and there is not animal dead in experimentation in laboratory animal body weight.
Therefore, melanin tumour b16 F10 mice-transplanted tumor growth inhibition test result shows, compared with negative control group, each administration group group growth to melanin tumour b16 F10 mice-transplanted tumor has the inhibitory action (* P < 0.05, * * P < 0.01) of significance.In docetaxel ~ Fructus Rubi polysaccharide, the growth of high administering drug combinations group to melanin tumour b16 F10 mice-transplanted tumor has the inhibitory action of significance, and the inhibition of docetaxel ~ Fructus Rubi polysaccharide height administering drug combinations group higher than the independent medication group of docetaxel (
Δp < 0.05).Illustrate that Fructus Rubi polysaccharide can strengthen the anti-tumor capacity of docetaxel.In addition, compared with docetaxel, the body weight of Fructus Rubi polysaccharide to laboratory animal has no significant effect, and has no obvious toxicity.
Embodiment 2
Cisplatin ~ Fructus Rubi polysaccharide is combined people pulmonary carcinoma H460 nude mouse xenograft tumor growth inhibition test
To take the logarithm the tumor cell of trophophase, be aseptically prepared into 5 × 10
7/ ml cell suspension, is inoculated in axillary fossa on the right side of nude mice with 0.1ml subcutaneous.With vernier caliper measurement transplanted tumor in nude mice diameter, treat tumor growth to 100 ~ 200mm
3rear animal random packet.Use the method measuring tumor footpath, dynamically observe antitumous effect.The pendulous frequency of diameter of tumor is every 2 days 1 time, and each measurement also needs weighing Mus heavy simultaneously.Administering mode is as follows: negative control group injection normal saline, every day 1 time; Cisplatin group 10mg/kg, Per-Hop behavior 1 time; High administering drug combinations group in cisplatin ~ Fructus Rubi polysaccharide, cisplatin adopts subcutaneous injection, 10mg/kg, Per-Hop behavior 1 time, and Fructus Rubi polysaccharide adopts gavage, and dosage is respectively 200mg/kg, 400mg/kg, administration every day 1 time.Gross tumor volume computing formula:
TV＝0.52×a×b
2
Wherein a, b represent length and width respectively.Result according to measuring calculates relative tumour volume.The evaluation index of anti-tumor activity is Relative tumor rate of increase T/C (%), and computing formula is as follows:
T/C(％)＝T
RTV/C
RTV×100％
T
rTV: treatment group RTV; C
rTV: negative control group RTV
The inhibitory action that table 2. cisplatin ~ Fructus Rubi polysaccharide drug combination grows people pulmonary carcinoma H460 nude mouse xenograft tumor
Result: in table 2, the tumour inhibiting rate of cisplatin 10mg/kg group to people pulmonary carcinoma H460 transplanted tumor in nude mice is 61.49%; Cisplatin ~ Fructus Rubi polysaccharide is combined the tumour inhibiting rate of middle and high dosage group to people pulmonary carcinoma H460 transplanted tumor in nude mice and is respectively 65.90%, 72.30%.Wherein, the inhibition of cisplatin ~ Fructus Rubi polysaccharide height administering drug combinations group is higher than the independent medication group of cisplatin.In addition, toxicity of cisplatin is comparatively large, and the weight of animals declines obviously, and in experimentation, animal has death.After both conbined usage, without obvious decline, and there is not animal dead in experimentation in laboratory animal body weight.
Therefore, people pulmonary carcinoma H460 transplanted tumor in nude mice growth inhibition test result is shown, compared with negative control group, cisplatin 10mg/kg group, the cisplatin ~ Fructus Rubi polysaccharide senior middle school growth of administering drug combinations group to people pulmonary carcinoma H460 transplanted tumor has the inhibitory action (* P < 0.05, * * P < 0.01) of pole significance.Compared with positive control cisplatin, the cisplatin ~ growth of Fructus Rubi polysaccharide height administering drug combinations group to H460 transplanted tumor have significance inhibitory action (
Δp < 0.05), and the body weight of laboratory animal has no significant effect, and has no obvious toxicity.
Embodiment 3
Paclitaxel ~ Fructus Rubi polysaccharide combines the inhibition test to the growth of people hepatocarcinoma Bel ~ 7402 nude mouse xenograft tumor
Specific embodiments is with reference to embodiment 2.Dosage regimen is as follows: negative control group injection normal saline, every day 1 time; Paclitaxel group 10mg/kg, Per-Hop behavior 1 time; High administering drug combinations group in paclitaxel ~ Fructus Rubi polysaccharide, paclitaxel adopts subcutaneous injection, 10mg/kg, Per-Hop behavior 1 time, and Fructus Rubi polysaccharide adopts gavage, and dosage is respectively 200mg/kg, 400mg/kg, administration every day 1 time.
The inhibitory action that table 3. paclitaxel ~ Fructus Rubi polysaccharide grows people hepatocarcinoma Bel ~ 7402 nude mouse xenograft tumor
Result: in table 3, the tumour inhibiting rate of paclitaxel 10mg/kg group to people hepatocarcinoma Bel ~ 7402 transplanted tumor in nude mice is 68.55%; Paclitaxel ~ Fructus Rubi polysaccharide is combined the tumour inhibiting rate of middle and high dosage group to people hepatocarcinoma Bel ~ 7402 transplanted tumor in nude mice and is reached 72.47% respectively, 76.47%.Wherein, the inhibition of paclitaxel ~ Fructus Rubi polysaccharide senior middle school administering drug combinations group higher than the independent medication group of paclitaxel (
Δp < 0.05).And paclitaxel toxicity is comparatively large, the weight of animals declines obviously, and in experimentation, animal has death.After both conbined usage, without obvious decline, and there is not animal dead in experimentation in laboratory animal body weight.
Therefore, paclitaxel ~ Fructus Rubi polysaccharide shows people hepatocarcinoma Bel ~ 7402 transplanted tumor in nude mice growth inhibition test result, compared with negative control group, paclitaxel 10mg/kg group, the paclitaxel ~ Fructus Rubi polysaccharide senior middle school growth of administering drug combinations group to people hepatocarcinoma Bel ~ 7402 transplanted tumor has the inhibitory action (* P < 0.05, * * P < 0.01) of pole significance.Compared with positive controls paclitaxel, the paclitaxel ~ body weight of Fructus Rubi polysaccharide senior middle school administering drug combinations to laboratory animal does not have a significant effect, and has no obvious toxicity.
Embodiment 4
Paclitaxel ~ Fructus Rubi polysaccharide is combined people Gastric Cancer MGC ~ 803 nude mouse xenograft tumor growth inhibition test
Specific embodiments is with reference to embodiment 2.Dosage regimen is as follows: negative control group injection normal saline, every day 1 time; Paclitaxel group 10mg/kg, Per-Hop behavior 1 time; High administering drug combinations group in paclitaxel ~ Fructus Rubi polysaccharide, paclitaxel adopts subcutaneous injection, 10mg/kg, Per-Hop behavior 1 time, and Fructus Rubi polysaccharide adopts gavage, and dosage is respectively 200mg/kg, 400mg/kg, administration every day 1 time.
The inhibitory action that table 4. paclitaxel ~ Fructus Rubi polysaccharide grows people Gastric Cancer MGC ~ 803 nude mouse xenograft tumor
Result: in table 4, the tumour inhibiting rate of paclitaxel 10mg/kg group to people Gastric Cancer MGC ~ 803 transplanted tumor in nude mice is 65.26%; Paclitaxel ~ Fructus Rubi polysaccharide is combined the tumour inhibiting rate of middle and high dosage group to people Gastric Cancer MGC ~ 803 transplanted tumor in nude mice and is reached 70.35% respectively, 72.48%.Wherein, the inhibition of paclitaxel ~ Fructus Rubi polysaccharide senior middle school administering drug combinations group higher than the independent medication group of paclitaxel (
Δp < 0.05).And paclitaxel toxicity is comparatively large, the weight of animals declines obviously, and in experimentation, animal has death.After both conbined usage, without obvious decline, and there is not animal dead in experimentation in laboratory animal body weight.
Therefore, paclitaxel ~ Fructus Rubi polysaccharide shows people Gastric Cancer MGC ~ 803 transplanted tumor in nude mice growth inhibition test result, compared with negative control group, paclitaxel 10mg/kg group, the paclitaxel ~ Fructus Rubi polysaccharide senior middle school growth of administering drug combinations group to people Gastric Cancer MGC ~ 803 transplanted tumor has the inhibitory action (* P < 0.05, * * P < 0.01) of pole significance.Compared with positive controls paclitaxel, the paclitaxel ~ body weight of Fructus Rubi polysaccharide senior middle school administering drug combinations to laboratory animal does not have a significant effect, and has no obvious toxicity.
Embodiment 5
Paclitaxel ~ Fructus Rubi polysaccharide is combined human breast carcinoma MDA ~ MB ~ 231 nude mouse xenograft tumor growth inhibition test
Specific embodiments is with reference to embodiment 2.Dosage regimen is as follows: negative control group injection normal saline, every day 1 time; Paclitaxel group 10mg/kg, Per-Hop behavior 1 time; Paclitaxel ~ Fructus Rubi polysaccharide administering drug combinations group, paclitaxel adopts subcutaneous injection, 10mg/kg, Per-Hop behavior 1 time, and Fructus Rubi polysaccharide adopts gavage, and dosage is 400mg/kg, administration every day 1 time.
The inhibitory action that table 5. paclitaxel ~ Fructus Rubi polysaccharide grows human breast carcinoma MDA ~ MB ~ 231 nude mouse xenograft tumor
Result: in table 5, the tumour inhibiting rate of paclitaxel 10mg/kg group to human breast carcinoma MDA ~ MB ~ 231 transplanted tumor in nude mice is 70.87%; Paclitaxel ~ Fructus Rubi polysaccharide combines the inhibitory rate 75.36% of group to human breast carcinoma MDA ~ MB ~ 231 transplanted tumor in nude mice.Wherein, the inhibition of paclitaxel ~ Fructus Rubi polysaccharide administering drug combinations group higher than the independent medication group of paclitaxel (
Δp < 0.05).And paclitaxel toxicity is comparatively large, the weight of animals declines obviously, and in experimentation, animal has death.After both conbined usage, without obvious decline, and there is not animal dead in experimentation in laboratory animal body weight.
Therefore, paclitaxel ~ Fructus Rubi polysaccharide shows human breast carcinoma MDA ~ MB ~ 231 transplanted tumor in nude mice growth inhibition test result, compared with negative control group, paclitaxel 10mg/kg group, the paclitaxel ~ growth of Fructus Rubi polysaccharide administering drug combinations group to human breast carcinoma MDA ~ MB ~ 231 transplanted tumor has the inhibitory action (* P < 0.05, * * P < 0.01) of pole significance.Compared with positive controls paclitaxel, the paclitaxel ~ body weight of Fructus Rubi polysaccharide administering drug combinations to laboratory animal does not have a significant effect, and has no obvious toxicity.
Embodiment 6
Paclitaxel ~ Fructus Rubi polysaccharide is combined people renal carcinoma A498 nude mouse xenograft tumor growth inhibition test
Specific embodiments is with reference to embodiment 2.Dosage regimen is as follows: negative control group injection normal saline, every day 1 time; Paclitaxel group 10mg/kg, Per-Hop behavior 1 time; High administering drug combinations group in paclitaxel ~ Fructus Rubi polysaccharide, paclitaxel adopts subcutaneous injection, 10mg/kg, Per-Hop behavior 1 time, and Fructus Rubi polysaccharide adopts gavage, and dosage is respectively 200mg/kg, 400mg/kg, administration every day 1 time.
The inhibitory action that table 6. paclitaxel ~ Fructus Rubi polysaccharide grows people renal carcinoma A498 nude mouse xenograft tumor
Result: in table 6, the tumour inhibiting rate of paclitaxel 10mg/kg group to people renal carcinoma A498 transplanted tumor in nude mice is 60.21%; Paclitaxel ~ Fructus Rubi polysaccharide is combined the tumour inhibiting rate of middle and high dosage group to people renal carcinoma A498 transplanted tumor in nude mice and is reached 69.55% respectively, 72.19%.Wherein, in paclitaxel ~ Fructus Rubi polysaccharide high administering drug combinations group inhibition higher than the independent medication group of paclitaxel (
Δp < 0.05).And paclitaxel toxicity is comparatively large, the weight of animals declines obviously, and in experimentation, animal has death.After both conbined usage, without obvious decline, and there is not animal dead in experimentation in laboratory animal body weight.
Therefore, paclitaxel ~ Fructus Rubi polysaccharide shows people renal carcinoma A498 transplanted tumor in nude mice growth inhibition test result, compared with negative control group, paclitaxel 10mg/kg group, the paclitaxel ~ Fructus Rubi polysaccharide senior middle school growth of administering drug combinations group to people renal carcinoma A498 transplanted tumor has the inhibitory action (* P < 0.05, * * P < 0.01) of pole significance.Compared with positive controls paclitaxel, the paclitaxel ~ body weight of Fructus Rubi polysaccharide senior middle school administering drug combinations to laboratory animal does not have a significant effect, and has no obvious toxicity.
Embodiment 7
Paclitaxel ~ Fructus Rubi polysaccharide combines the inhibition test to the growth of human colon carcinoma HT ~ 29 nude mouse xenograft tumor
Specific embodiments is with reference to embodiment 2.Dosage regimen is as follows: negative control group injection normal saline, every day 1 time; Paclitaxel group 10mg/kg, Per-Hop behavior 1 time; Paclitaxel ~ Fructus Rubi polysaccharide administering drug combinations group, paclitaxel adopts subcutaneous injection, 10mg/kg, Per-Hop behavior 1 time, and Fructus Rubi polysaccharide adopts gavage, and dosage is 400mg/kg, administration every day 1 time.
The inhibitory action that table 7. paclitaxel ~ Fructus Rubi polysaccharide grows human colon carcinoma HT ~ 29 nude mouse xenograft tumor
Result: in table 7, the tumour inhibiting rate of paclitaxel 10mg/kg group to human colon carcinoma HT ~ 29 transplanted tumor in nude mice is 62.88%; Paclitaxel ~ Fructus Rubi polysaccharide combines the inhibitory rate 68.23% of group to human colon carcinoma HT ~ 29 transplanted tumor in nude mice.Wherein, the inhibition of paclitaxel ~ Fructus Rubi polysaccharide administering drug combinations group higher than the independent medication group of paclitaxel (
Δp < 0.05).And paclitaxel toxicity is comparatively large, the weight of animals declines obviously, and in experimentation, animal has death.After both conbined usage, without obvious decline, and there is not animal dead in experimentation in laboratory animal body weight.
Therefore, paclitaxel ~ Fructus Rubi polysaccharide shows human colon carcinoma HT ~ 29 transplanted tumor in nude mice growth inhibition test result, compared with negative control group, paclitaxel 10mg/kg group, the paclitaxel ~ growth of Fructus Rubi polysaccharide administering drug combinations group to human colon carcinoma HT ~ 29 transplanted tumor has the inhibitory action (* P < 0.05, * * P < 0.01) of pole significance.Compared with positive controls paclitaxel, the paclitaxel ~ body weight of Fructus Rubi polysaccharide administering drug combinations to laboratory animal does not have a significant effect, and has no obvious toxicity.
Embodiment 8
Paclitaxel ~ Fructus Rubi polysaccharide combines the inhibition test to the growth of people's carcinoma of endometrium HHUA nude mouse xenograft tumor
Specific embodiments is with reference to embodiment 2.Dosage regimen is as follows: negative control group injection normal saline, every day 1 time; Paclitaxel group 10mg/kg, Per-Hop behavior 1 time; High administering drug combinations group in paclitaxel ~ Fructus Rubi polysaccharide, paclitaxel adopts subcutaneous injection, 10mg/kg, Per-Hop behavior 1 time, and Fructus Rubi polysaccharide adopts gavage, and dosage is respectively 200mg/kg, 400mg/kg, administration every day 1 time.
The inhibitory action that table 8. paclitaxel ~ Fructus Rubi polysaccharide grows people's carcinoma of endometrium HHUA nude mouse xenograft tumor
Result: in table 8, the tumour inhibiting rate of paclitaxel 10mg/kg group to people's carcinoma of endometrium HHUA transplanted tumor in nude mice is 58.77%; Paclitaxel ~ Fructus Rubi polysaccharide is combined the tumour inhibiting rate of middle and high dosage group to people's carcinoma of endometrium HHUA transplanted tumor in nude mice and is reached 67.21% respectively, 69.34%.Wherein, in paclitaxel ~ Fructus Rubi polysaccharide high administering drug combinations group inhibition higher than the independent medication group of paclitaxel (
Δp < 0.05).And paclitaxel toxicity is comparatively large, the weight of animals declines obviously, and in experimentation, animal has death.After both conbined usage, without obvious decline, and there is not animal dead in experimentation in laboratory animal body weight.
Therefore, paclitaxel ~ Fructus Rubi polysaccharide shows people's carcinoma of endometrium HHUA transplanted tumor in nude mice growth inhibition test result, compared with negative control group, paclitaxel 10mg/kg group, the paclitaxel ~ growth of Fructus Rubi polysaccharide senior middle school administering drug combinations group to people's carcinoma of endometrium HHUA transplanted tumor has the inhibitory action (* P < 0.05, * * P < 0.01) of pole significance.Compared with positive controls paclitaxel, in paclitaxel ~ Fructus Rubi polysaccharide, the body weight of high administering drug combinations to laboratory animal does not have a significant effect, and has no obvious toxicity.
Embodiment 9
Fructus Rubi polysaccharide is on the impact of tumor-bearing mice liver, renal tissue form
After each group of tumor-bearing mice anesthesia in embodiment 1 ~ 8 anti-tumor experiment, be separated liver and kidney.Liver, kidney are soaked in the formalin of 10% fixing.By the tissue that fixes through ethanol dehydration, dimethylbenzene is transparent, paraffin embedding, section paster, dimethylbenzene dewaxes, and HE dyes, and dewaters transparent, takes pictures after sealing in optical microphotograph Microscopic observation.
As shown in Figure 1, the hepatocyte edge clear of normal mouse, Cytoplasm fills the hepatic tissue section of each group of mice, and nucleus is clear, and cell arrangement is neat.By contrast, the hepatocyte height swelling of chemotherapeutics group mice, subregion hepatic tissue focal necrosis, interstitial stove cell infiltration.And the hepatocyte that chemotherapeutics ~ Fructus Rubi polysaccharide combines group mice has obvious recovery, the hepatocyte that wherein chemotherapeutics ~ Fructus Rubi polysaccharide high dose combines group mice is similar to normal mouse hepatocyte.Visible, Fructus Rubi polysaccharide has certain repair to the tumor-bearing mice hepatic injury that chemotherapeutics causes.
As shown in Figure 2, the glomerule regular shape of normal mouse, glomerular capsule gap is little, and glomerular basement membrane is complete in the nephridial tissue section of each group of mice.By contrast, there is certain degeneration and damage in the glomerule of chemotherapeutics group mice, is in particular in glomerular basement membrane distortion, and renal capsule gap is obviously widened.Chemotherapeutics ~ Fructus Rubi polysaccharide combine group mice glomerule recover clearly, be mainly manifested in renal capsule gap comparatively chemotherapeutics group obviously diminish.Visible, Fructus Rubi polysaccharide also has certain repair to the tumor-bearing mice injury of kidney that chemotherapeutics causes.
These results suggest that, the lesions of liver and kidney that Fructus Rubi polysaccharide causes after repairing chemotherapeutic drug therapy, after chemotherapeutics and Fructus Rubi polysaccharide conbined usage, toxic and side effects reduces to some extent.
Embodiment 10
Fructus Rubi polysaccharide is on the impact of tumor-bearing mice index and spleen index
After embodiment 1 ~ 8 anti-tumor experiment terminates, by mouse anesthesia, be separated tumor tissues and spleen, weigh.Index and spleen index is calculated according to weighing results.Computing formula: index and spleen index=spleen weight (g)/clean body weight (kg), wherein, clean body weight=body weight ~ tumor weight.Test the result obtained to represent with mean ± SD, and carry out statistics T inspection, P < 0.05 is significant difference, and P < 0.01 is pole significant difference (*: compared with negative control group, # are compared with chemotherapeutics group).
The results are shown in Figure 3.Compared with chemotherapeutics group, the index and spleen index of chemotherapeutics ~ Fructus Rubi polysaccharide administering drug combinations group all significantly increases.
Embodiment 11
The impact of Fructus Rubi Polysaccharides on Mice splenocyte in-vitro multiplication
Cervical dislocation puts to death mice, soaks 1 ~ 2 minute, taking out spleen rapidly, filling in the plate of ice-cold PBS as being equipped with 400 order nylon screens in advance in aseptic operating platform in 75% ethanol.Screen cloth gently twists with the fingers spleen tissue with aseptic nook closing member, makes individual cells filter screen cloth and enter plate.Collecting cell suspension after screen cloth is rinsed, in 4 DEG C centrifugal (1000r/min) 5 minutes with PBS.Abandon supernatant, also count under the microscope with the RPMI ~ 1640 culture medium re-suspended cell containing 2%FBS, cell concentration is adjusted to 1 × 10
7individual/ml, by cell suspension inoculation in 96 orifice plates, 100 μ l/ holes, and in hole, add the ConA of 5 μ g/ml, and in 37 DEG C, 5%CO
2overnight incubation in incubator.Next day, add respectively after Fructus Rubi polysaccharide culture fluid is diluted to each predetermined concentration in (100 μ l/ hole) in 96 orifice plates.Using do not add ConA as blank group, do not add the group of Fructus Rubi polysaccharide for positive controls only to add ConA.At 37 DEG C, 5%CO
2incubator hatches 48 hours.In 96 orifice plates, add the MTT of 5mg/ml, every hole 20 μ l, continue to cultivate 4h.Sop up culture medium, every hole adds 150 μ l DMSO and dissolves, and shaking table mixes for 10 minutes gently.With microplate reader mensuration wavelength be 570nm, reference wavelength be 630nm place mensuration light absorption value.Test the result obtained to represent with mean ± SD, and carry out statistics T inspection, P < 0.05 is significant difference, and P < 0.01 is pole significant difference (*: compared with negative control group, # are compared with positive controls).
As shown in Figure 4, the light absorption value that Fructus Rubi polysaccharide is respectively organized all is significantly higher than negative control group to result, and presents dose-dependence.In addition, from 64 μ g/ml, positive controls is significantly higher than.Illustrate thus, the propagation of Fructus Rubi polysaccharide energy significant stimulation mouse spleen lymphocyte.
Embodiment 12
Fructus Rubi polysaccharide is on the impact of Related Immune Enzyme Activity in tumor-bearing mice splenocyte
Spleen embodiment 1 ~ 8 respectively being organized tumor-bearing mice uses cell crushing instrument fragmentation (400 amperes, each 5 seconds, repeat 3 times) under condition of ice bath, crushing medium is pH7.4,0.01mol/L Tris ~ HCl, 0.0001mol/L EDTA ~ 2Na, 0.01mol/L sucrose, 0.8%NaCl.Collect supernatant in 4 DEG C centrifugal (5000r/min) after 10 minutes, to detect in splenocyte the activity of four kinds of immune relevant enzyme AKP, ACP, LDH and SOD with testing cassete respectively.Test the result obtained to represent with mean ± SD, and carry out statistics T inspection, P < 0.05 is significant difference, and P < 0.01 is pole significant difference (*: compared with negative control group, # are compared with positive controls).
As shown in Figure 5, compared with negative control group, Fructus Rubi polysaccharide significantly can promote the activity of 4 kinds of enzymes to result, and presents dose-dependence.Compared with chemotherapeutics group, chemotherapeutics ~ Fructus Rubi polysaccharide combines the activity that improve ACP of group significance.Illustrate thus, after chemotherapeutics and Fructus Rubi polysaccharide conbined usage, it obtains enhancing to the raising ability of ACP activity.
Embodiment 13
Fructus Rubi polysaccharide is on the impact of CD4, CD8, IL ~ 2 in tumor-bearing mice serum, INF ~ γ and TNF ~ alpha content
Blood is got by extracing eyeball after each group of tumor-bearing mice anesthesia in embodiment 1 ~ 8 anti-tumor experiment.Blood leaves standstill and collects serum in 4 DEG C centrifugal (2500r/min) after 10 minutes in a moment.CD4, CD8, IL in serum ~ 2, INF ~ γ and TNF ~ alpha content is detected respectively with enzyme-linked immunologic detecting kit.Test the result obtained to represent with mean ± SD, and carry out statistics T inspection, P < 0.05 is significant difference, and P < 0.01 is pole significant difference (*: compared with negative control group, # are compared with positive controls).
As shown in Figure 6, compared with positive controls, chemotherapeutics ~ Fructus Rubi polysaccharide combines group all can significantly or the content of raising 5 kinds of albumen of pole significance for result.Illustrate thus, after docetaxel and Fructus Rubi polysaccharide conbined usage, it obtains enhancing to the raising ability of these five kinds of protein contents.
Each immunization experiment result further illustrates above, and Fructus Rubi polysaccharide can strengthen the immunity of tumor-bearing mice, and substantially increases the therapeutic effect of chemotherapeutics.
Claims (10)
1. Fructus Rubi polysaccharide and chemotherapeutics are preparing the purposes in antineoplastic combination medicine.
2. purposes according to claim 1, is characterized in that: described chemotherapeutics is selected from paclitaxel, cisplatin or/and docetaxel.
3. purposes according to claim 1, is characterized in that: described tumor is selected from melanoma, pulmonary carcinoma, hepatocarcinoma, gastric cancer, breast carcinoma, renal carcinoma, carcinoma of endometrium or colon cancer.
4. purposes according to claim 1 and 2, is characterized in that: described chemotherapeutics adopts injection type.
5. purposes according to claim 1 and 2, is characterized in that: described Fructus Rubi polysaccharide adopts peroral dosage form.
6. the purposes according to Claims 1 to 5 any one, is characterized in that: Fructus Rubi polysaccharide is 20 ~ 40: 1w/w with the using dosage ratio of chemotherapeutics.
7. the purposes according to claim 1 ~ 6 any one, is characterized in that: in described Fructus Rubi polysaccharide, purity of polysaccharide is more than 50%; Further, purity of polysaccharide is more than 80%; Further, purity of polysaccharide is more than 85%; Preferably, purity of polysaccharide is 85 ~ 95%w/w.
8. the purposes according to claim 1 ~ 7 any one, is characterized in that: described Fructus Rubi polysaccharide adopts decoction and alcohol sedimentation technique to prepare; Further, described decoction and alcohol sedimentation technique concrete operations are as follows: get dry Fructus Rubi, after petroleum ether or ether defatting, with concentration 75 ~ 95%v/v ethanol extraction, filter, filtering residue extracting in water, after water intaking extract is concentrated, add ethanol and reach 75% ~ 85% to alcohol content, cool, leave standstill, get solid content and be Fructus Rubi polysaccharide.
9. purposes according to claim 8, is characterized in that: Fructus Rubi removes seed or do not remove seed before extracting.
10. the purposes according to claim 1 ~ 9 any one, is characterized in that: described Fructus Rubi is the fruit of Rosaceae rubus; Further, described Fructus Rubi is selected from red raspberry or black raspberry.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105254773A (en) * | 2015-10-27 | 2016-01-20 | 齐齐哈尔医学院 | Preparation method of antitumor polysaccharide |
| CN112358553A (en) * | 2020-12-09 | 2021-02-12 | 青海大学 | Polysaccharide SM-0.2M and anti-tumor product prepared from same |
| CN112538121A (en) * | 2020-12-09 | 2021-03-23 | 青海大学 | Polysaccharide SM-0.4M and anti-tumor product prepared from same |
| CN115414465A (en) * | 2022-10-17 | 2022-12-02 | 苏州明人医药生物科技有限公司 | Application of JWA polypeptide in preparation of antitumor drug synergist |
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| CN102731668A (en) * | 2011-04-12 | 2012-10-17 | 谭智丽 | Method for extracting and purifying Codonopsis pilosula polysaccharide |
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| CN102731668A (en) * | 2011-04-12 | 2012-10-17 | 谭智丽 | Method for extracting and purifying Codonopsis pilosula polysaccharide |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105254773A (en) * | 2015-10-27 | 2016-01-20 | 齐齐哈尔医学院 | Preparation method of antitumor polysaccharide |
| CN112358553A (en) * | 2020-12-09 | 2021-02-12 | 青海大学 | Polysaccharide SM-0.2M and anti-tumor product prepared from same |
| CN112538121A (en) * | 2020-12-09 | 2021-03-23 | 青海大学 | Polysaccharide SM-0.4M and anti-tumor product prepared from same |
| CN115414465A (en) * | 2022-10-17 | 2022-12-02 | 苏州明人医药生物科技有限公司 | Application of JWA polypeptide in preparation of antitumor drug synergist |
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