CN105998047A - Novel drug combination scheme for treating cancers - Google Patents
Novel drug combination scheme for treating cancers Download PDFInfo
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Abstract
The invention discloses a novel drug combination scheme for treating caners (particularly the liver cancer), namely a sorafenib and fludarabine phosphate combination drug scheme, and particularly comprises application of combination of sorafenib and fludarabine phosphate for preparing medicine for treating cancer and a medicine comprising sorafenib and fludarabine phosphate. A sorafenib and fludarabine phosphate combination drug shows the synergistic interaction, the curative effect of sorafenib for a patient suffering from the liver cancer and the survival benefit obtaining potential of the patient suffering from the liver cancer are improved, and the sorafenib and fludarabine phosphate combination drug treatment scheme can be adopted for a patient who has the limited curative effect or large side effect of sorafenib monotherapy so as to improve the curative effect of sorafenib, improve the life quality and prolong life time of the patient. The drug combination scheme provides the novel treatment thought and solution strategy for clinic treatment for the liver cancer, and the novel drug combination scheme has the huge development potential and application prospect for pushing clinic treatment for the liver cancer.
Description
Technical field
The invention belongs to pharmaceutical technology field.More particularly, to a kind of new scheme of combination drug therapy treating cancer.
Background technology
Cancer is typically to form by growing rapid cell, and they are likely to be of various abnormal chromosome, often in pleomorphism and in various degree between deform." cancer " this word occur so far for the first time from mankind's word dictionary, people are " talking cancer complexion changed ", obtain cancer, and are tantamount to pronounced death penalty.From initial fear, passive waiting, active treatment finally, Drug therapy, operative treatment etc., the mankind have started very long " anticancer battle ".Such as hepatocarcinoma, it is serious threat human life and healthy important diseases, is the fifth-largest kinds of tumor in the world, the third-largest tumor associated death reason.The global cancer statistical datas in 2012 issued according to American Cancer Society, 2012, whole world new cancer cases was about 782, and 500, death is about 745, and 500, and wherein 50% it both is from China.Although the traditional therapy of hepatocarcinoma is more, such as operation, intervention, radiotherapy, chemotherapy, liver transplantation etc., but the patient only less than 30% can undergo surgery or the curative therapy such as liver transplantation, and postoperative recurrence and the rate of transform the highest.Remaining most patients mostly then is patients with terminal, loses operation or the chance of liver transplantation.To this, systemic chemotherapy Drug therapy, it may be possible to the only effective Therapeutic Method.
At present, Sorafenib (Nexavar) is the standard care medicine that only one is approved by the fda in the United States for advanced liver cancer patient.Sorafenib is a kind of oral Mutiple Targets inhibitors of kinases, and its action target spot includes Raf-1 and B-Raf,
vascular endothelial growth factor receptor (VEGFR), platelet-derived growth
Factor receptor beta (PDGFR β), c-KIT and FLT3.The result of two extensive phase iii clinical trials shows, compared with placebo, Sorafenib can significantly improve clinical Benefit and the life span of hepatocarcinoma patient, delays the progression of disease of patient.It addition, research finds, Sorafenib applies also for tumor of kidney, to the no longer valid local recurrence of radioiodine therapy or transitivity, progressively differentiated thyroid etc..
But, result of study shows simultaneously, and Sorafenib makes patient's profit time extremely limited, life span extension probably only 2～3 months.Therefore, it is badly in need of finding new liver cancer treatment strategy, is effectively improved patient's life cycle and quality of life.A kind of brand-new medicine, from researching and developing, test, be registered to the whole process of listing, takes around time of 10～15 years and the input of 500 ten thousand to 20 hundred million dollars, and the cycle is long, costly, not in time, therefore, a lot of drugmakers are all unwilling to research and develop brand-new cancer therapy drug in return.In this process, a lot of patients are because the listing that can't wait new active drug is finally regretted dead.In consideration of it, re-use the cancer therapy drug that has been approved by of FDA, to find the new curative effect outside its original therapeutic effect, the medicine having been approved by FDA re-uses, and is of great immediate significance and exploitation advantages.First, these medicines have carried out strict pharmaceutical research, the existing clear and definite conclusion such as the activity of medicine, formula, dosage form and effective dose；Secondly, these medicines have carried out strict pharmacokinetics and pharmacodynamic study, Drug safety and toxicity strict test and appraisal and reference guide；3rd, the raw material of these medicines is readily available mostly, and processing technology is ripe, and has obtained the listing approval of Drug administrative organization, these advantages make to re-use the medicine being worth can enter clinical trial, faster faster for the new patient being suitable for disease.
But, the scheme of combination drug therapy between medicine and medicine is an extremely complex process, needs to consider the dynamics of cell proliferation of two medicines, mechanism of action, drug toxicity and toxicity, antitumor spectra, dosage etc. aspect.Therefore, the research and utilization of drug combination new departure of old medicine, is the research topic of a bulky complex.
Summary of the invention
The technical problem to be solved in the present invention is defect and the deficiency overcoming existing cancer treatment drugs, the scheme of combination drug therapy of a kind for the treatment of of cancer is provided, will Sorafenib and fludarabine phosphate use in conjunction, the effect of Synergistic can be realized, significantly improve the curative effect of Sorafenib treatment cancer, especially treat the curative effect of hepatocarcinoma.
It is an object of the invention to provide Sorafenib and fludarabine phosphate combination application in terms of preparation cancer treatment drugs.
The present invention another object is that a kind of cancer treatment drugs comprising Sorafenib and fludarabine phosphate.
Above-mentioned purpose of the present invention is achieved through the following technical solutions:
Sorafenib and fludarabine phosphate combination application in terms of preparation cancer treatment drugs.
Preferably, described cancer is hepatocarcinoma.
Sorafenib and fludarabine phosphate combination application in terms of the medicine preparing anticancer.
Preferably, described cancerous cell is hepatoma carcinoma cell.
Preferably, the medicine of above-mentioned anticancer is growth and/or the medicine of transfer of anticancer.
It is highly preferred that described hepatoma carcinoma cell is hepatoma carcinoma cell kind Hep3B, SMMC-7721, BEL-7402, Huh-7, PLC/PRF/5 or SK-Hep-1.
A kind of cancer treatment drugs, described pharmaceutical pack contains Sorafenib and fludarabine phosphate.
Preferably, described cancer is hepatocarcinoma.
Preferably, the ratio of described Sorafenib and fludarabine phosphate is 1～12:1～20.
It is highly preferred that the ratio of described Sorafenib and fludarabine phosphate is 3:10.
It is highly preferred that described medicine is suppression liver cancer cell growth and/or the medicine of transfer.
The medicine of Synergistic anti-cancer effect can be produced with Sorafenib use in conjunction to find, we have used the oncotherapy related drugs of tens kinds of U.S. FDA approveds, it is united and applied in hepatoma carcinoma cell Hep3B with Sorafenib, cell proliferation is detected, by calculating drug interaction coefficient CDI (The coefficient by the method for MTS
Of drug interaction), scheme of combination drug therapy associated with different pharmaceutical and Sorafenib is divided into synergism (CDI ＜ 1), summation action (CDI=1), antagonism (CDI ＞ 1) three classes, found that, Sorafenib and fludarabine phosphate drug combination can produce synergism (CDI ＜ 1), are obviously enhanced anti-cancer effectiveness.
Further, we are again in the hepatoma carcinoma cell of this five type of SMMC-7721, BEL-7402, Huh-7, PLC/PRF/5 and SK-Hep-1, the scheme for combining of Sorafenib and fludarabine phosphate is verified further, finds scheme of combination drug therapy four kinds of cells wherein of Sorafenib and fludarabine phosphate all demonstrate synergism (CDI ＜ 1).Additionally, we are also by the fludarabine phosphate bought from pharmaceutical companies and Sorafenib use in conjunction, again verify in six kinds of hepatoma carcinoma cell kinds of Hep3B, SMMC-7721, BEL-7402, Huh-7, PLC/PRF/5 and SK-Hep-1, calculate CDI value, also same conclusions is obtained, Sorafenib and fludarabine phosphate scheme of combination drug therapy are significantly better than Sorafenib list regimen, and hepatoma carcinoma cell is had coordinate repression.
Hepatoma carcinoma cell is utilized to carry out cell growth curve experiment, plate clone formation experiment, apoptosis test experience, scratch experiment, migration experiment and Western experiment further, confirm that the two drug combination can work in coordination with suppression growth of tumour cell and propagation, promotion apoptosis of tumor cells, and suppress Tumor Cell Migration and transfer ability.Utilize hepatocarcinoma subcutaneous one-tenth tumor animal model and hepatocarcinoma Lung metastases animal model, also confirm that the two drug combination is better than the independent medication of Sorafenib, it is possible to work in coordination with the growth in Mice Body of the suppression tumor and transfer, and there are the potentiality extending mice with tumor life span.The machine-processed explorative experiment of drug combination confirms, Sorafenib and fludarabine phosphate scheme of combination drug therapy are better than Sorafenib list regimen, it is because the two and p-Erk is had coordinate repression, simultaneously, fludarabine phosphate can reverse the rising of the p-Akt that the independent medication of Sorafenib causes, and this demonstrates drug combination can produce synergistic molecular mechanism.
Therefore, Sorafenib and fludarabine phosphate are united and applied in the treatment of liver cancer patient by the present invention, demonstrate more preferable hepatocarcinoma inhibition, for the patient that Sorafenib single therapy offer limited effectiveness or side effect are bigger, can consider to carry out Sorafenib and fludarabine phosphate drug combination therapeutic scheme, to improve the curative effect of Sorafenib, improve life quality and the life span of patient.
Sorafenib and fludarabine phosphate drug combination have the potentiality that the curative effect of liver cancer patient and the existence of liver cancer patient are benefited by raising Sorafenib further, have huge potentiality to be exploited and the application prospect pushing hepatocarcinoma clinical treatment to.
The method have the advantages that
The invention discloses one and treat the new scheme of combination drug therapy of cancer (especially hepatocarcinoma), i.e. Sorafenib and fludarabine phosphate scheme of combination drug therapy.This scheme of combination drug therapy shows the result of Synergistic, has the effect improving Sorafenib independent medication suppression hepatocarcinoma, and the clinical treatment for hepatocarcinoma provides new treatment thoughts and scheme policies.
Sorafenib and fludarabine phosphate drug combination have the potentiality that the curative effect of liver cancer patient and the existence of liver cancer patient are benefited by raising Sorafenib further, for the patient that Sorafenib single therapy offer limited effectiveness or side effect are bigger, can consider to carry out Sorafenib and fludarabine phosphate drug combination therapeutic scheme, to improve the curative effect of Sorafenib, improving life quality and the life span of patient, this scheme of combination drug therapy has huge potentiality to be exploited and the application prospect pushing hepatocarcinoma clinical treatment to.
Accompanying drawing explanation
Fig. 1 is six kinds of hepatoma carcinoma cell and a kind of liver immortalized cells L02 to the drug susceptibility of Sorafenib and IC50 value.
Fig. 2 is that Sorafenib ratifies antitumor drug drug combination the selection result in Hep3B cell with FDA.
Fig. 3 is that Sorafenib ratifies antitumor drug drug combination the result in SMMC-7721, five kinds of hepatoma carcinoma cell of BEL-7402, Huh-7, PLC/PRF/5, SK-Hep-1 with FDA.
Fig. 4 is fludarabine phosphate and Sorafenib is combined at Hep3B, the result again verified in six kinds of hepatoma carcinoma cell of SMMC-7721, BEL-7402, Huh-7, PLC/PRF/5, SK-Hep-1.
Fig. 5 be Sorafenib and fludarabine phosphate scheme of combination drug therapy liver cancer cell growth is bred, Clone formation and apoptotic effect.
Fig. 5 is the cell growth curve figure that Hep3B, SMMC-7721 hepatoma carcinoma cell accepts after variant group of process.
Fig. 6 is the cell clonal formation result that Hep3B, SMMC-7721 hepatoma carcinoma cell accepts after variant group of process.
Fig. 7 is that Hep3B, SMMC-7721 hepatoma carcinoma cell accepts Apoptosis by Flow Cytometry result after variant group of process.
Fig. 8 is that Hep3B, SMMC-7721 hepatoma carcinoma cell accepts the western blot experiment detection significant molecule of apoptosis after variant group of process.
Fig. 9 is that Hep3B, SMMC-7721 hepatoma carcinoma cell accepts cell cut Healing Experiments result after variant group of process.
Figure 10 is that Hep3B, SMMC-7721 hepatoma carcinoma cell accepts Cell migration assay result after variant group of process.
Figure 11 is that Hep3B, SMMC-7721 hepatoma carcinoma cell accepts western blot experiment detection EMT developed by molecule situation of change after variant group of process.
Figure 12 be Sorafenib with fludarabine phosphate scheme of combination drug therapy to hepatocarcinoma subcutaneous become tumor mouse model effect；A figure is to carry out each group of mice subcutaneous tumors change in volume figure with administration；B figure is to strip each group of mice subcutaneous tumors to weigh the tumor multigraph obtained；C figure is taken pictures comparison diagram for stripping each group of mice subcutaneous tumors；D figure is to carry out each group of Mouse Weight variation diagram with administration.
Figure 13 be Sorafenib with fludarabine phosphate scheme of combination drug therapy to hepatocarcinoma subcutaneous become tumor mouse model effect, each group HE colored graph of mice subcutaneous tumors tissue slice, Ki-67 colored graph and TUNEL colored graph.
Figure 14 is Sorafenib and the effect to hepatocarcinoma Lung metastases mouse model of the fludarabine phosphate scheme of combination drug therapy, and A figure is each group of rat liver cancer Lung metastases situation；B figure is the HE colored graph of the hepatoma Metastasis tumor formed in each group of mice lungs；Figure C is the size and number cartogram of the hepatoma Metastasis tumor formed in each group of mice lungs.
Figure 15 is each group mouse survival curve chart of hepatocarcinoma Lung metastases mouse model.
Figure 16 is the Mechanism Study that Sorafenib and fludarabine phosphate scheme of combination drug therapy produce synergistic function.
Detailed description of the invention
Further illustrate the present invention below in conjunction with Figure of description and specific embodiment, but the present invention is not limited in any form by embodiment.Unless stated otherwise, the present invention uses reagent, method and apparatus are the art conventional reagent, method and apparatus.
Unless stated otherwise, agents useful for same of the present invention and material are commercial.
Embodiment
1
The cell Antibiotics resistance test to Sorafenib
First six kinds of hepatoma carcinoma cell Hep3B of detection, SMMC-7721, BEL-7402, Huh-7, PLC/PRF/5, SK-Hep-1 and a kind of liver immortalized cells L02 drug susceptibility to Sorafenib.
1, experimental technique
Cell spreads 96 orifice plates, adherent overnight after, prepare the Sorafenib of a series of concentration from high to low, and matched group be set, add in hand-hole, each concentration arranges 3 multiple holes.After hatching 48 hours in cell culture incubator, after adding MTS, 2 hours, utilize the cell OD value of multi-functional microplate reader detection 490nm wavelength, calculate Sorafenib and act on the IC50(half maximal of different cell
Inhibitory concentration) value.
2, result is as it is shown in figure 1, in Fig. 1, A figure is to process the amount effect curve after each hepatoma carcinoma cell with the Sorafenib of variable concentrations；The block diagram that B figure is drawn by the IC50 value that each hepatoma carcinoma cell is corresponding.
Result shows, Sk-Hep-1 cellular sensitivity is the highest, and BEL-7402 cell is minimum.
It addition, the Drug level of Sorafenib list medicine group in every kind of cell can be determined according to the IC50 value of different hepatoma carcinoma cell.
Embodiment
2
Scheme of combination drug therapy research
Tens kinds of tumor associated treatment medicines from the application of America NI H/NCI are utilized to carry out scheme of combination drug therapy screening Hep3B cell with Sorafenib.
1, experimental technique
Hep3B hepatoma carcinoma cell spreads 96 orifice plates, adherent overnight after, cell is divided into blank group, DMSO matched group, Sorafenib list medicine group, NCI mono-medicine group, and Sorafenib and NCI combination therapies group, the often multiple holes of group three, it is separately added into culture medium or the drug solution of correspondence, after hatching 48 hours, MTS method detection cell proliferative conditions, and the drug interaction coefficient CDI value of different scheme of combination drug therapy is calculated according to cell OD value.
CDI ＜ 1, represents that scheme of combination drug therapy demonstrates synergism；CDI=1, represents that scheme of combination drug therapy demonstrates summation action；CDI ＞ 1, represents that scheme of combination drug therapy demonstrates antagonism.
2, result is as in figure 2 it is shown, in Fig. 2, A figure is after Hep3B cell accepts different NCI medicine and the process of Sorafenib drug combination, the point and line chart that in each experimental group, the relative survival rate of cell obtains after Z-score converts；B figure is, after Hep3B cell accepts different NCI medicine and the process of Sorafenib drug combination, to show that synergistic drug combination combination CDI value and this combination CDI value in other five kinds of hepatoma carcinoma cell collect the CDI Distribution value thermal map obtained；C figure is that Hep3B cell accepts after different NCI medicine processes with Sorafenib drug combination, calculates the block diagram that the CDI value of scatterplot that the CDI of each scheme of combination drug therapy is worth to and the synergistic scheme of combination drug therapy of all displays is drawn.
Result shows, the CDI ＜ 1 of Sorafenib and fludarabine phosphate scheme of combination drug therapy, represents that Sorafenib and fludarabine phosphate scheme of combination drug therapy demonstrate the effect of obvious Synergistic.
Embodiment
3
Sorafenib and fludarabine phosphate scheme of combination drug therapy research
1, in order to verify result above, to SMMC-7721, five kinds of hepatoma carcinoma cell application Sorafenibs of BEL-7402, Huh-7, PLC/PRF/5, SK-Hep-1 and fludarabine phosphate scheme of combination drug therapy, method is with method for Hep3B in embodiment 2.
2, result is as it is shown on figure 3, in Fig. 3, A figure is the result of checking in SMMC-7721 cell；B figure is the result of checking in BEL-7402 cell；C figure is the result of checking in Huh-7 cell；D figure is the result of checking in PLC/PRF/5 cell；E figure is the result of checking in SK-Hep-1 cell.
Sorafenib and fludarabine phosphate scheme of combination drug therapy demonstrate the most significantly synergistic function, and four kinds of cells in five kinds of cells all demonstrate synergistic function (CDI ＜ 1).
Embodiment
4
Sorafenib and the checking research of fludarabine phosphate scheme of combination drug therapy
1, in order to verify result above further, by the fludarabine phosphate bought from pharmaceutical companies and Sorafenib use in conjunction, again at Hep3B, SMMC-7721, BEL-7402, Huh-7, PLC/PRF/5, verifying in six kinds of hepatoma carcinoma cell of SK-Hep-1, method is with method for Hep3B in embodiment 2.
2, result
As shown in Figure 4, also obtain same conclusions, Sorafenib and fludarabine phosphate scheme of combination drug therapy and be significantly better than Sorafenib list regimen, hepatoma carcinoma cell is demonstrated coordinate repression.
Embodiment
5
Sorafenib and fludarabine phosphate scheme of combination drug therapy are to liver cancer cell growth propagation, Clone formation and apoptotic effect
1, additionally, it is better than Sorafenib list regimen to more comprehensively prove Sorafenib and fludarabine phosphate scheme of combination drug therapy, hepatocarcinoma had coordinate repression, cell growth curve experiment, plate clone formation experiment, apoptosis test experience, scratch experiment, migration experiment and Western experiment are carried out respectively, hepatocarcinoma subcutaneous one-tenth tumor animal model and hepatocarcinoma Lung metastases animal model experiment, and the molecular mechanisms experiment of drug combination.
2, Hep3B, SMMC-7721 hepatoma carcinoma cell spreads 96 orifice plates, adherent overnight after, cell is divided into matched group, Sorafenib list medicine group, NCI mono-medicine group, and Sorafenib and NCI combination therapies group, the often multiple hole of group three, is separately added into culture medium or the drug solution of correspondence, hatches 7 days, every 24 hours MTS method detection cell proliferative conditions, and draw cell growth curve according to cell OD value.
Result is as it is shown in figure 5, Fig. 5 is the cell growth curve figure that Hep3B, SMMC-7721 hepatoma carcinoma cell accepts after variant group of process.Compared with matched group and single medicine group, drug combination cell proliferation has obvious coordinate repression, and drug combination group cell growth rate is the slowest.
3, Hep3B, SMMC-7721 hepatoma carcinoma cell spreads 6 orifice plates, adherent overnight after, cell is divided into matched group, Sorafenib list medicine group, NCI mono-medicine group, and Sorafenib and NCI combination therapies group, the often multiple hole of group three, it is separately added into culture medium or the drug solution of correspondence, hatches 14 days, detect cell plates Clone formation situation.
As shown in Figure 6, Fig. 6 is the cell clonal formation figure that Hep3B, SMMC-7721 hepatoma carcinoma cell accepts after variant group of process to result.Compared with matched group and single medicine group, drug combination has obvious coordinate repression to cell clonal formation, and drug combination group cell clone quantity is minimum, and volume is minimum.
4, Hep3B, SMMC-7721 hepatoma carcinoma cell spreads 6 orifice plates, adherent overnight after, cell is divided into matched group, Sorafenib list medicine group, NCI mono-medicine group, and Sorafenib and NCI combination therapies group, the often multiple hole of group three, is separately added into culture medium or the drug solution of correspondence, hatches 24 hours, then collecting each group of cell, Flow cytometry respectively organizes apoptosis situation.
Result accepts Apoptosis by Flow Cytometry figure after variant group of process as it is shown in fig. 7, Fig. 7 is Hep3B, SMMC-7721 hepatoma carcinoma cell.Compared with matched group and single medicine group, drug combination has the most collaborative facilitation to apoptosis, and drug combination group apoptosis ratio is most.
5, Hep3B, SMMC-7721 hepatoma carcinoma cell spreads 6 orifice plates, adherent overnight after, cell is divided into matched group, Sorafenib list medicine group, NCI mono-medicine group, and Sorafenib and NCI combination therapies group, the often multiple holes of group three, it is separately added into culture medium or the drug solution of correspondence, hatch 24 hours, then collect each group of cell, extract cell protein, the shearing situation of western method detection apoptosis significant molecule caspase and PARP.
As shown in Figure 8, Fig. 8 is that Hep3B, SMMC-7721 hepatoma carcinoma cell accepts the western blot experiment detection significant molecule of apoptosis after variant group of process to result.Compared with matched group and single medicine group, drug combination has been obviously promoted apoptosis, caspase and the PARP showed increased of shear pattern.
Embodiment
6
Sorafenib and the effect to fucosylation ability of the fludarabine phosphate scheme of combination drug therapy
1, Hep3B, SMMC-7721 hepatoma carcinoma cell spreads 6 orifice plates, adherent overnight after, carry out cell scratch experiment, then cell be divided into matched group, Sorafenib list medicine group, NCI mono-medicine group, and Sorafenib and NCI combination therapies group, the often multiple holes of group three, it is separately added into culture medium or the drug solution of correspondence, the when of hatching 24 hours and 48 hours, take pictures respectively, observation of cell cut healing state.
Result accepts cell cut Healing Experiments after variant group of process as it is shown in figure 9, Fig. 9 is Hep3B, SMMC-7721 hepatoma carcinoma cell.Compared with matched group and single medicine group, drug combination has obvious coordinate repression to the healing of cell cut, and the healing of drug combination group cell cut is the slowest.
2, Hep3B, SMMC-7721 hepatoma carcinoma cell paving Transwell cell, it is divided into matched group, Sorafenib list medicine group, NCI mono-medicine group, and Sorafenib and NCI combination therapies group, the often multiple holes of group three, it is separately added into culture medium or the drug solution of correspondence, hatch 24 hours, observation of cell migration situation.
As shown in Figure 10, Figure 10 is that Hep3B, SMMC-7721 hepatoma carcinoma cell accepts Cell migration assay after variant group of process to result.Compared with matched group and single medicine group, drug combination cell migration ability has obvious coordinate repression, drug combination group cell migration minimum number.
3, Hep3B, SMMC-7721 hepatoma carcinoma cell spreads 6 orifice plates, adherent overnight after, cell is divided into matched group, Sorafenib list medicine group, NCI mono-medicine group, and Sorafenib and NCI combination therapies group, the often multiple holes of group three, it is separately added into culture medium or the drug solution of correspondence, hatch 24 hours, then collect each group of cell, extract cell protein, the expression of significant molecule E-cadherin, N-cadherin, β-catenin of western method detection EMT process.
As shown in figure 11, Figure 11 is that Hep3B, SMMC-7721 hepatoma carcinoma cell accepts western blot experiment detection EMT developed by molecule situation of change after variant group of process to result.Compared with matched group and single medicine group, drug combination substantially inhibits the EMT process of cell, and E-cadherin expresses and increases, and N-cadherin, β-catenin expresses minimizing.
Embodiment
7
Sorafenib and fludarabine phosphate scheme of combination drug therapy to hepatocarcinoma subcutaneous become tumor mouse model and the effect of hepatocarcinoma Lung metastases mouse model
1, method
Hepatocellular carcinoma in nude mice subcutaneous one-tenth tumor model is built by subcutaneous vaccination with Hep3B cell, hepatocellular carcinoma in nude mice Lung metastases model is built by tail vein injection with Sk-Hep-1 cell, after model construction success, mice is divided into four groups, matched group, Sorafenib list medicine group, NCI mono-medicine group, and Sorafenib and NCI combination therapies group, and it is administered process, Sorafenib oral administration gavage is administered, one day 1 time, 30mg/day/kg, fludarabine phosphate intraperitoneal injection, one day 1 time, 100mg/day/kg, it is administered 21 days altogether.Observe growth and the transfer case of mouse interior tumor.
2, result is as shown in Figure 12 ~ Figure 15,
(1) hepatocarcinoma subcutaneous one-tenth tumor result shows (as shown in A～D figure in Figure 12), and Tu12Zhong, A figure is to carry out each group of mice subcutaneous tumors change in volume figure with administration；B figure is to strip each group of mice subcutaneous tumors to weigh the tumor multigraph obtained；C figure is taken pictures comparison diagram for stripping each group of mice subcutaneous tumors；D figure is to carry out each group of Mouse Weight variation diagram with administration.Compared with matched group and single medicine group, drug combination has obvious inhibitory action to tumor growth, and drug combination group tumor growth rate is the slowest, and the gross tumor volume finally stripped is minimum and weight is the lightest.
(2) Ki-67 and TUNEL coloration result is as shown in figure 13, and Figure 13 is the HE colored graph of each group of mice subcutaneous tumors tissue slice, Ki-67 colored graph and TUNEL colored graph.Result shows, drug combination substantially inhibits tumor proliferation, promotes tumor death, drug combination group Ki-67 and TUNEL staining positive cells ratio and intensity is the highest.
(3) hepatocarcinoma Lung metastases model result is as shown in Figure 14 and Figure 15.In Figure 14, A figure is each group of rat liver cancer Lung metastases situation；B figure is the HE colored graph of the hepatoma Metastasis tumor formed in each group of mice lungs；Figure C is the size and number cartogram of the hepatoma Metastasis tumor formed in each group of mice lungs.Figure 15 is each group of mouse survival curve chart.
Result shows, compared with matched group and single medicine group, drug combination substantially inhibits the Lung metastases ability of hepatoma carcinoma cell, and the metastatic tumor minimum number, the area that are formed in drug combination group mouse lung are minimum.The life span of statistics mice, the life span of drug combination group mice extends.
Embodiment
8
Sorafenib and fludarabine phosphate scheme of combination drug therapy Mechanism Study
1, having synergistic molecular mechanism to prove drug combination, Hep3B, SMMC-7721 hepatoma carcinoma cell spreads 6 orifice plates, adherent overnight after, cell is divided into matched group, Sorafenib list medicine group, NCI mono-medicine group, and Sorafenib and NCI combination therapies group, the often multiple hole of group three, it is separately added into culture medium or the drug solution of correspondence, hatches 24 hours, then collect each group of cell, extract cell protein, the expression of detection Erk/p-Erk and Akt/mTOR signal path correlation molecule.
2, result is as shown in Figure 16.In Figure 16, A figure is after Hep3B, SMMC-7721 hepatoma carcinoma cell accepts contrast solution, Sorafenib list drug solns, the mono-drug solns of NCI and Sorafenib and the process of NCI combination therapies solution respectively, the expression situation of change of western blot experiment detection Erk/p-Erk and Akt/mTOR signal path key molecule；B figure is after Hep3B, SMMC-7721 hepatoma carcinoma cell accepts contrast solution, Sorafenib list drug solns, MK2206 solution (AKT inhibitor), Sorafenib and the process of MK2206 drug combination solution respectively, the expression situation of change of western blot experiment detection AKT/p-AKT molecule；C figure is after Hep3B, SMMC-7721 hepatoma carcinoma cell accepts contrast solution, Sorafenib list drug solns, Rapamycin solution (mTOR inhibitors), Sorafenib and the process of Rapamycin drug combination solution respectively, the expression situation of change of western blot experiment detection mTOR/p-mTOR, S6/p-S6 molecule.
Result shows, Sorafenib and fludarabine phosphate drug combination have coordinate repression to p-Erk, meanwhile, fludarabine phosphate can reverse the rising of the Akt/mTOR signal path that the independent medication of Sorafenib causes, and this is that drug combination can produce synergistic molecular mechanism.
Claims (10)
1. Sorafenib and fludarabine phosphate combination application in terms of preparation cancer treatment drugs.
Application the most according to claim 1, it is characterised in that described cancer is hepatocarcinoma.
3. Sorafenib and fludarabine phosphate combination application in terms of the medicine preparing anticancer.
Application the most according to claim 3, it is characterised in that described cancerous cell is hepatoma carcinoma cell.
Application the most according to claim 3, it is characterised in that described suppression is growth and/or the transfer of anticancer.
Application the most according to claim 4, it is characterised in that described hepatoma carcinoma cell is hepatoma carcinoma cell kind Hep3B, SMMC-7721, BEL-7402, Huh-7, PLC/PRF/5 or SK-Hep-1.
7. a cancer treatment drugs, it is characterised in that described pharmaceutical pack contains Sorafenib and fludarabine phosphate.
Cancer treatment drugs the most according to claim 7, it is characterised in that described cancer is hepatocarcinoma.
Cancer treatment drugs the most according to claim 7, it is characterised in that the ratio of described Sorafenib and fludarabine phosphate is 1～12:1～20.
Cancer treatment drugs the most according to claim 7, it is characterised in that described medicine is suppression liver cancer cell growth and/or the medicine of transfer.
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| CN107929712A (en) * | 2017-12-12 | 2018-04-20 | 中山大学肿瘤防治中心 | A kind of combination medicine for treating liver cancer |
| CN111789847A (en) * | 2020-07-01 | 2020-10-20 | 佳木斯大学 | Application of combination of sorafenib and oleanolic acid in preparation of medicine for treating liver cancer |
| CN113662939A (en) * | 2021-09-29 | 2021-11-19 | 遵义医科大学 | Application of composition of magnesium demethylcantharidinate and sorafenib in preparation of anti-liver cancer drugs |
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| CN113662939A (en) * | 2021-09-29 | 2021-11-19 | 遵义医科大学 | Application of composition of magnesium demethylcantharidinate and sorafenib in preparation of anti-liver cancer drugs |
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