CN102210668B - Application of cantharidin and derivants thereof in preparation of tumor chemotherapy sensitivity enhancing medicine - Google Patents

Application of cantharidin and derivants thereof in preparation of tumor chemotherapy sensitivity enhancing medicine Download PDF

Info

Publication number
CN102210668B
CN102210668B CN2011101233823A CN201110123382A CN102210668B CN 102210668 B CN102210668 B CN 102210668B CN 2011101233823 A CN2011101233823 A CN 2011101233823A CN 201110123382 A CN201110123382 A CN 201110123382A CN 102210668 B CN102210668 B CN 102210668B
Authority
CN
China
Prior art keywords
cantharidin
cell
carcinoma
derivant
tumor
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Active
Application number
CN2011101233823A
Other languages
Chinese (zh)
Other versions
CN102210668A (en
Inventor
李玉新
鲍永利
郑丽华
乌垠
于春雷
孟祥颖
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Northeast Normal University
Original Assignee
INST OF GENETICS AND CYTOLOGY NORTHEAST NORMAL UNIV
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by INST OF GENETICS AND CYTOLOGY NORTHEAST NORMAL UNIV filed Critical INST OF GENETICS AND CYTOLOGY NORTHEAST NORMAL UNIV
Priority to CN2011101233823A priority Critical patent/CN102210668B/en
Publication of CN102210668A publication Critical patent/CN102210668A/en
Application granted granted Critical
Publication of CN102210668B publication Critical patent/CN102210668B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Images

Abstract

The invention discloses application of cantharidin and derivants thereof in preparation of a tumor chemotherapy sensitivity enhancing medicine for enhancing sensitivity of tumors to treatment medicines. Both a compound and derivants thereof related by the invention have the functions of reducing MDR (Multiple Medicament Resistance) genetic transcription and expression in various tumor cells and further inhibiting occurrence of MDR, so that relatively strong tumor chemotherapy sensitivity enhancing function is displayed.

Description

The application in the preparation sensitization medicament for tumour chemotherapy of cantharidin and derivant thereof
Technical field
The invention discloses the application in the preparation sensitization medicament for tumour chemotherapy of cantharidin new medical way, especially cantharidin and derivant thereof, belong to biomedicine field.
Background technology
Tumor cell meeting in chemotherapy process does not have many medicines generation drug resistance of dependency to a big class formation and function; This phenomenon is called " multidrug resistance "; The acquisition of tumor multidrug-resistance combines the proteic expression of boxlike carrier closely related with the various ATP of cell surface; In human chromosomal, about 50 kinds of ABC vector genes are arranged, the drug resistance that wherein in tumor cell, mediates with the P-glycoprotein of MDR1 coded by said gene is the most extensive.Thinking at present has two mechanism to participate in the rise of MDR1 gene in the malignant tumor.One is that the promoter of MDR1 gene can be activated through environmental factors, and another main mechanism is the methylating and demethylation of CpG site on the MDR1 promoter.
P-gp avoids the infringement of heterotoxin to protecting body, drains its metabolite, and conversion endogenous harmful substance etc. has the important physiological meaning.Yet because the overexpression of P-gp makes cytotoxic drug be pumped out the extracellular, intracellular drug level descends in tumor cell, thereby produces drug resistance, and the overexpression of MDR degree and MDR1 is proportionate.
At present, the P-gp protein function suppressive drug of the reverse multiple drug resistance of tumor of having found is a sensitization agent for tumour chemotherapy, mainly contains calcium channel blocker, antisense RNA, ribozyme etc.But they all have fatal weakness, and the medicine of the blocking-up P-gp pump of having found because toxic and side effects is big, is used (Sharma V et al. than difficulty clinically Chem Rev.1999,99:2545～2560.).Will be in vivo an antisense RNA and a ribozyme transfection are given tumor cell and do not got into other tissue, technical have a suitable difficulty.Therefore fully understand the relation of MDR1 gene and drug resistance of tumor in the human tumor cells, filter out efficiently, the chemotherapeutic sensitizer and the reversal of drug resistance agent of low toxicity be problem demanding prompt solution in the present oncotherapy.
(cantharidin is extensively to be present in more than the 1500 kind of intravital a kind of natural defensive toxin of Mylabris CA) to cantharidin.The overgrown with weeds blue or green section of Mylabris insecticide, acrid in the mouth cold in nature for hypertoxic Chinese crude drug, except that having active anticancer, still has various active such as antiviral, tonifying YANG, leukocyte increasing.
The activity research situation of cantharidin derivative shows (Ceng Wennan; Lu Yi. the synthetic and activity research progress of cantharidin and derivant thereof. organic chemistry. 2006; 26 (5): 579 ~ 591), the structure activity relationship of cantharidin (see figure 1) as: (1) dicyclo [2.2.1] heptane is the basic framework ring of cantharidin, and the methyl of removing on 2-C or the 3-C can not influence its activity; On this site, can introduce other bigger substituent group, therefore very big structure of modification space is arranged on this site; L-C or 4-C go up the introducing substituent group can make its active reduction, but nearest research shows that the cantharidin derivative that has absolute steric configuration in the l-c position has good inhibition selectivity to PP2B; Can introduce substituent group on 5-c or the 6-C, also can be two keys between 5-C and the 6-C; 7-oxo bridge key is must be obligato, possibly be the cause that oxygen atom ability and PP1 and PP2A form hydrogen bond.(2) after the oxygen atom on the anhydride rings is replaced by nitrogen-atoms, the inhibitory action of PP1 and PP2A is significantly reduced; Oxygen atom can not influence its inhibitory action to PP1 and PP2A after being replaced by sulphur atom; The anhydride open loop obviously changes its inhibitory action to PP1 and PP2A, and anhydride moiety also principle can lose the inhibitory action to PP1 and PP2A.In a series of cantharidin derivatives, that is relatively paid close attention at present has following several kinds.
(Norcantharidin is the synthesis of derivatives of cantharidin NCTD) to cantharidin, is the chemical constitution according to the anticancer effective component cantharidin of coleoptera Meloidae insecticide Mylabris; Remove 1; 2 methyl synthetic and getting, its characteristics can obviously alleviate the intensive urinary system zest of cantharidin, keep stronger anti-tumor activity and unique function of increasing leukocyte; Mainly be applicable to the treatment of primary hepatocarcinoma, effect is better.
N-methylcantharidimide (N-methylcantharidimide) and N-hydroxycantharidin (N-hydroxycantharidimide) are a kind of imide derivant of cantharidin.The toxic and side effects of N-methylcantharidimide and N-hydroxycantharidin reduces greatly.Zoopery shows that the antitumaous effect of N-hydroxycantharidin is similar with cantharidin, but its toxicity is merely one of five percentages of cantharidin.
Disodium cantharidinate be cantharidin and sodium hydroxide when hot altogether hydrolysis generate (Wang Zemin. contemporary structure medicament complete or collected works [M] Beijing: Beijing science tech publishing house, 1993:2125～2126).Disodium cantharidinate has not only kept the distinctive active anticancer of cantharidin, and toxic and side effects is littler than cantharidin, but also can improve tumor patient
Immunologic function (Liang Feng, Wang Mingyan. the progress of disodium cantharidinate. Jiangxi College of Traditional Chinese Medicine journal, 2006,18 (1): 67-68).
In a word; Understanding for cantharidin and derivant pharmacological action thereof mainly concentrated on its antitumor action in the past; But the present invention finds cantharidin and derivant thereof and has the multi-medicine tolerant reversal effect, can reduce medicine and pump to extracellular, thereby has the chemotherapy sensitizing effect.
Summary of the invention
The present invention discloses a kind of cantharidin and the purposes of derivant in the preparation sensitization medicament for tumour chemotherapy thereof, is used to strengthen the sensitivity of tumor to medicine.
Cantharidin involved in the present invention and derivant thereof have following general formula:
Figure 977537DEST_PATH_IMAGE001
R wherein 1Be CH 3, R 2Be CH 2, X is O, Y is O (cantharidin);
Or R 1Be CH 3, R 2Be CH 2, X is N-CH 3, Y is O (N-methylcantharidimide)
Or R 1Be CH 3, R 2Be CH 2, X is N-OH, Y is O (N-hydroxycantharidin)
Or R 1Be CH 3, R 2Be CH 2, X is (ONa) 2, Y is O (disodium cantharidinate)
Or R 1Be CH 3, R 2Be CH 2, X is (OH) 2, Y is O (Cantharidic acid .);
Or R 1Be CH 3, R 2Be CH 2, X is (OH) 2, Y is H 2(deoxidation Cantharidic acid .)
Or R 1Be CH 3, R 2Be CH 2, X is S, Y is O (a thia cantharidin).
Or R 1Be H, R 2Be CH 2, X is O, Y is O (norcantharidin);
Or R 1Be H, R 2Be CH, X is O, and Y is O (dehydronorcantharidiimide is plain);
Or R 1Be H, R 2Be CH 2, X is O, Y is H 2(nor-deoxidation cantharidin);
Or R 1Be H, R 2Be CH 2, X is N-CH 3, Y is O (a demethyl cantharidimide);
Or R 1Be H, R 2Be CH 2, X is N-CH 2CH 2OH, Y are O (the nor-cantharidimide of hydroxyethyl);
Or R 1Be H, R 2Be CH 2, X is N-CH 2CH 2OCH 3, Y is O (the nor-cantharidimide of methoxy ethyl);
Or R 1Be H, R 2Be CH 2, X is N-CH 2CH 2OCH 2CH 2OH, Y are O (the nor-cantharidimide of hydroxyl ethylene glycol ethyl);
Or R 1Be H, R 2Be CH 2, X is N-CH 2CH 2OCH 2CH 2OCH 3, Y is O (the nor-cantharidimide of Propylene Glycol ethyl);
Or R 1Be H, R 2Be CH 2, X is HN-CH 2CH 2OCH 2CH 2OCH 3(OH), Y is O (the nor-Mylabris amic acid of Propylene Glycol ethyl);
Or R 1Be H, R 2Be CH 2, X is N-CH 2CO-NHCH 2COOH, Y are O (the nor-cantharidimide of glycine dipeptidase);
Or R 1Be H, R 2Be CH 2, X is HN-CH 2CO-NHCH 2CO (OH) 2, Y is O (the nor-Mylabris amino acid of glycine dipeptidase);
Or R 1Be H, R 2Be CH 2, X is (OH) 2, Y is H 2(nor-deoxidation Cantharidic acid .);
Or R 1Be H, R 2Be CH 2, X is N-CH (R)-COOH, Y is O (an aminoacid demethyl cantharidimide);
Or R 1Be H, R 2Be CH 2, X is (ONa) 2, Y is O (Injectio natarii norcantharidatis).
Cantharidin of the present invention and derivant thereof comprise cantharidin; N-methylcantharidimide; N-hydroxycantharidin; Disodium cantharidinate; Norcantharidin; Dehydronorcantharidiimide is plain; Nor-deoxidation cantharidin; Cantharidic acid.; The deoxidation Cantharidic acid.; Nor-deoxidation Cantharidic acid.; The thia cantharidin; Nor-cantharidimide; The nor-cantharidimide of hydroxyethyl; The nor-cantharidimide of methoxy ethyl; The nor-cantharidimide of aminoacid; The nor-cantharidimide of hydroxyl ethylene glycol ethyl; The nor-cantharidimide of Propylene Glycol ethyl; The nor-Mylabris amino acid of glycine dipeptidase; Injectio natarii norcantharidatis; The nor-cantharidimide of glycine dipeptidase etc.
Thereby cantharidin involved in the present invention and derivant thereof are all through suppressing the expression performance chemotherapy sensitizing effect of MDRG MDR1.
The chemotherapy sensitizing purposes and the Mechanism Study thereof of cantharidin of the present invention and derivant thereof are carried out through following method:
1. the acquisition of cantharidin and derivant thereof
Cantharidin and derivant thereof can prepare through the commodity purchasing acquisition or through following method:
The cantharidin method for preparing is seen patent 200610040991.1.
The N-methylcantharidimide method for preparing is seen patent 200410067589.3.
The disodium cantharidinate method for preparing is seen patent 200410052938.4.
The method for preparing of norcantharidin is seen Diels, O., Alder, K.Chem.Ber., 1929,62,554.
The Injectio natarii norcantharidatis method for preparing is seen patent 200510045564.8.
Cantharidimide and the nor-cantharidimide of nor-cantharidimide derivant hydroxyethyl, the nor-cantharidimide of methoxy ethyl, the nor-cantharidimide of aminoacid, the nor-cantharidimide of hydroxyl ethylene glycol ethyl, the nor-cantharidimide of Propylene Glycol ethyl, the nor-Mylabris amino acid of glycine dipeptidase, the nor-cantharidimide method for preparing of glycine dipeptidase are seen patent 200310116882.X.
The method for preparing of dehydronorcantharidiimide element, nor-deoxidation cantharidin, Cantharidic acid. or thia cantharidin is seen patent 200510033897.9.
2. multidrug resistance cell strain induces
Respectively human liver cancer cell HepG2, human leukemia cell K562, human colon cancer cell LoVo cell, human breast cancer cell MCF-7, human lung cancer cell A549, gastric carcinoma cells SGC7901, ovarian cancer SKOV3, cervical cancer Hela, renal carcinoma 786-0 or the carcinoma of prostate PC-3 of debita spissitudo is inoculated in 6 orifice plates and cultivates; Add the ADM that final concentration is 0.05 μ g/ml next day; Every liquid that changed at a distance from 2-3 days, the while increases the concentration of ADM gradually; Mortality can appear in cell after dosing 2-3 time, and still visible small amounts of cells is adherent, continues to increase progressively gradually the concentration of ADM, forms one cell clone until the attached cell of remnants; Treating that cell forms big clone, with pancreatin cell dissociation is got off, be uniformly dispersed, continue to add ADM and induce, can be among the ADM of 2 μ g/ml at final concentration until cell, go down to posterity normally, frozen and the recovery.Induce more than 8 months,, keep the drug resistance character of cell with the ADM of 1 μ g/ml.
3. the evaluation of multidrug resistance cell strain
Inoculate sensitive cells and mdr cell respectively in 96 orifice plates, cell density is 5 * 10 4Individual/ml;
Dosing next day, the Concentraton gradient of amycin is: 30,3,0.3,0.03,0.003 μ g/ml; The Concentraton gradient of vincristine is: 10,1,0.1,0.01,0.001 μ g/ml; The Concentraton gradient of paclitaxel is: 10,1,0.1,0.01,0.001 μ g/ml; The Concentraton gradient of 5-FU is: 10,1,0.1,0.01,0.001 μ g/ml, and medicine all carries out doubling dilution with the DMEM culture medium that contains 3%FBS, and negative group adds the DMEM culture medium that contains 3%FBS, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ lMTT (5mg/ml), continues to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ lDMSO, and on ELIASA, vibration 600s detects the OD value at 570nm place, and calculates suppression ratio, suppression ratio=(the absorbance average of the absorbance average/matched group of 1-experimental group) * 100%; More than the independent experiment triplicate, experimental data is all calculated IC with the SPSS statistical software 50Value, and calculate the drug resistance multiple (IC of drug resistance multiple=persister 50The IC of value/sensitive strain 50Value).The result is shown in table 1-10, and the multidrug resistance cell strain of various tumor cell lines is induced successfully.
Figure 364656DEST_PATH_IMAGE002
Figure 937720DEST_PATH_IMAGE003
Figure 562736DEST_PATH_IMAGE004
Figure 359791DEST_PATH_IMAGE005
Figure 437469DEST_PATH_IMAGE006
Figure 79803DEST_PATH_IMAGE007
Figure 559325DEST_PATH_IMAGE008
Figure 527281DEST_PATH_IMAGE009
Figure 7121DEST_PATH_IMAGE011
? 4. Cantharidin and its derivatives nontoxic doses of detection
Inoculate L02, HepG2, HepG2/ADM and 293T cell respectively in 96 orifice plates, cell density is 5 * 10 4Individual/ml; Dosing next day; The Concentraton gradient of cantharidin and derivant thereof is: 10,1,0.1,0.01,0.001 μ g/ml, and medicine all carries out doubling dilution with the DMEM culture medium that contains 3%FBS, and negative group adds the DMEM culture medium that contains 3%FBS; Establish 3 multiple holes for every group, the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT (5mg/ml, PBS joins), continues to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on ELIASA, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate, experimental data is all calculated IC with the SPSS statistical software 10Value, the drug level that the cell survival more than 90% is arranged is nontoxic or low toxicity dosage, confirms the working concentration of medicine for subsequent experimental.The result sees table 11.
Figure 615519DEST_PATH_IMAGE012
5. cantharidin and derivant thereof are to the chemotherapy sensitizing effect of multidrug resistance tumor cells strain
(1) to the chemotherapy sensitizing effect of drug-resistant cell strain
Inoculate sensitive cells and mdr cell respectively in 96 orifice plates, cell density is 5 * 10 4Individual/ml; Dosing next day, the Concentraton gradient of amycin is: 30,3,0.3,0.03,0.003 μ g/ml; It is high, medium and low three dose groups that cantharidin and derivant thereof are got 2 μ g/ml, 1 μ g/ml, 0.5 μ g/ml respectively; Medicine all carries out doubling dilution with the DMEM culture medium that contains 3%FBS; Negative group adds the DMEM culture medium that contains 3%FBS, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT (5mg/ml), continues to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on ELIASA, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate, experimental data is all calculated IC with the SPSS statistical software 50Value, and calculate drug resistance reversal fold, reverse the IC of the preceding persister of multiple=administration 50The IC of persister after the value/administration 50Value.The result shows; Cantharidin that the present invention mentioned and derivant thereof comprise that to the kinds of tumors drug-resistant cell strain human liver cancer cell HepG2, human leukemia cell K562, human colon cancer cell LoVo cell, human breast cancer cell MCF-7, human lung cancer cell A549, gastric carcinoma cells SGC7901, ovarian cancer SKOV3, cervical cancer Hela, renal carcinoma 786-0 or carcinoma of prostate PC-3 all have drug resistance inversion effect in various degree.Sensitizer when therefore, above-claimed cpd can be used as tumor pharmacother uses.
Figure 754376DEST_PATH_IMAGE013
Figure 806646DEST_PATH_IMAGE014
Figure 525203DEST_PATH_IMAGE015
Figure 510477DEST_PATH_IMAGE016
Figure 289077DEST_PATH_IMAGE017
Figure 828643DEST_PATH_IMAGE018
Figure 350891DEST_PATH_IMAGE019
Figure 901455DEST_PATH_IMAGE022
(2) chemotherapy sensitization in the body
With 5 * 10 6Individual drug-resistant tumor cell inoculation is subcutaneous in the right axil of BALB/c (nu/nu) nude mice.3 days began the lumbar injection amycin 2 mg/kg/ days behind tumor inoculation, cantharidin group intravenous injection cantharidin or derivatives thereof, and dosage is 1.5 mg/kg, every day 1 time, totally 10 times, experimental group is then injected amycin and cantharidin or derivatives thereof simultaneously.Put to death animal after 30 days, strip tumor and weigh, calculate tumour inhibiting rate, tumour inhibiting rate (the %)=average tumor of (it is heavy that average tumor is organized in the average tumor weight-treatment of tumor matched group)/tumor matched group heavy * 100.The result shows that the tumour inhibiting rate of injecting amycin and cantharidin group simultaneously is far longer than amycin group and cantharidin group tumour inhibiting rate sum.
Figure 227394DEST_PATH_IMAGE023
Figure 969085DEST_PATH_IMAGE028
Figure 261843DEST_PATH_IMAGE030
Figure 664006DEST_PATH_IMAGE031
Conclusion:
In view of cantharidin, N-methylcantharidimide, disodium cantharidinate, N-hydroxycantharidin and nor-cantharidimide all have drug resistance inversion effect preferably; Be in their structures common ground the performance this effect; Other has the cantharidin derivative of this common structure; It is active to have identical drug resistance inversion, can be used as sensitization medicament for tumour chemotherapy.
Figure 332884DEST_PATH_IMAGE001
6. cantharidin and derivant thereof are to the Mechanism Study of multi-medicine tolerant reversal effect
(1) cantharidin and derivant thereof are to the influence of MDR1 promoter activity
Press the said method of document (Maitra R; Halpin PA; Karlson KH. et al. Differential effects of mitomycin C and doxorubicin on P-glycoprotein expression. Biochem. J. 2001; 355,617-624.) angle and get the MDR1 promoter and be cloned among the pGL3-Basic, thereby construct the pGL3-MDR1-pro reporter plasmid.
In transfection the previous day, mdr cell is inoculated in 24 orifice plates, inoculum density is about 50-60%; Treat to begin when cell reaches 70-80% transfection.During transfection; Change liquid (every hole adds 0.5ml 10% DMEM) at first for the cell in 24 orifice plates; With calcium chloride solution mixing DNA, be about among the calcium chloride 12.5 μ l of 0.375 μ g DNA (0.25 μ g purpose plasmid+0.125 μ g is with reference to plasmid) adding respective volume mixing then.Calcium chloride behind the mixing-DNA mixed liquor is added dropwise in the 12.5 μ l BBS solution abundant mixing, room temperature held 20 minutes.At last BBS-calcium chloride-DNA mixture 25 μ l are added in 24 orifice plates, front and back vibrate gently, make its uniform distribution, and place 37 ℃, 5%CO2 incubator continuation cultivation.After cultivating 6h, inhale and remove culture fluid in the hole, add the fresh complete medium of 2ml, with cell mixing, counting, be laid in 96 orifice plates after the overnight incubation.Inhale behind the 24h and go culture medium, add the DMEM culture medium that contains 3%FBS, in this culture medium, add the cantharidin or derivatives thereof again, making its final concentration is 1 μ g/ml, does contrast with solvent DMSO simultaneously.After continuing to cultivate 24h, cell lysis is surveyed uciferase activity.
Utilize Luciferase Reporter Assay System and FluoStar Optima to detect the uciferase activity of transfectional cell lysate.Get 45 μ L cell pyrolysis liquids; The Assay Cocktail reagent that adds 5 μ L; Add 100 μ Luciferin reactant liquors behind the mixing; Utilize FluoStar Optima instrument to survey the fluorescence counting of 2s, this counting has reflected the activity level of research plasmid reporter gene expression LUC Photinus pyralis LUC Photinus pyralis FL (firefly luciferase).Other gets 20 μ L cell pyrolysis liquids; Add 37.5 μ L β-gal Buffer, add 12.5 μ L ONPG (6mg/mL) again, place 30min for 37 ℃ behind the mixing; Utilize ELIASA to survey the absorption value under the 450nm, this value has reflected that confidential reference items plasmid pCMV-β expresses the activity level of beta galactosidase.(ratio of the activity of β-gal) has been rejected the difference of sample transfection efficiency, more objectively the difference of promoter activity between reflected sample for the activity of LUC Photinus pyralis LUC Photinus pyralis FL (firefly luciferase) and beta galactosidase.The luciferase relative activity of different samples calculates with following formula.
Figure 60669DEST_PATH_IMAGE032
The result is as shown in Figure 2, and cantharidin that the present invention is mentioned and derivant thereof all can suppress the activity of MDRG MDR1 promoter to some extent.
(2) cantharidin and derivant thereof are to the influence of MDR1 gene encoding production P-gp expression
Sensitive cells and mdr cell are laid on 6 orifice plates respectively, and adding final concentration after 24 hours is cantharidin and the derivant thereof of 1 μ g/ml, continuation effect 24 hours.With PBS washing 3 times, every hole (6 orifice plate) adds 200 μ l lysate (50mM Tris-HCl (pH7.5), 150mM NaCl, 1mM NaF, 0.5% NP-40, the Aprotinin of 2 μ g/ml with cell; 1mM PMSF), acts on 30 minutes on ice, whenever rocked cell plates, so that the abundant cracking of cell at a distance from 5 minutes.Behind centrifugal 10 minutes of the 15000rpm, supernatant is moved into EP pipe, be placed in after packing-80 ℃ frozen subsequent use.Take out a pipe during electrophoresis and add 4 * albumen sample-loading buffer, boiled 10 minutes, refrigeration on ice, instantaneous centrifugal, last appearance electrophoresis.
Preparation is changeed the film buffer and in 4 ℃ of pre-coolings.Pvdf membrane is soaked 40sec with methanol, and being soaked in after the taking-up changes subsequent use in the film buffer.Change the film buffer with putting in the square plate, under liquid level, carry out the installation of membrane-transferring device.And membrane-transferring device put into electrophoresis tank, and pouring into and change the film buffer, 55V takes out pvdf membrane after changeing film 3h, with membrane marker positive and negative and electrophoresis direction.Wash pvdf membrane 5 minutes 2 times with TBST (0.2% Tween20), 4 ℃ of sealings of the defatted milk powder of reuse 5% are spent the night.Use TBST (0.2% Tween20) to get express developed afterwards 2 times, wash again 15 minutes 1 time, 5 minutes 4 times (all being put in when washing film on the shaking table, as follows).1G2 (2.5mg/ml) with after TBST (0.2% Tween20) the 1:500 dilution, is together packed in the hybridization bag with pvdf membrane, and 37 ℃ of shaking tables are hatched 2h.Take out pvdf membrane, get express developed 2 times, wash 5 minutes 4 times again 15 minutes 1 time with TBST (0.2% Tween20).Two anti-HRP-goat anti-mouse (0.8mg/ml) with after TBST (0.2% Tween20) the 1:2000 dilution, are together packed in the hybridization bag with pvdf membrane, and 37 ℃ of shaking tables are hatched 1h.Take out pvdf membrane, get express developed 2 times, wash 10 minutes 4 times again 15 minutes 1 time with TBST (0.2% Tween20).Pvdf membrane is taken into the darkroom ECL colour developing.The result is as shown in Figure 3, and cantharidin that the present invention mentioned and derivant thereof all can reduce the expression of P-gp in the persister to some extent.
(3) cantharidin and derivant thereof influence to accumulating in the amycin cell
Because amycin has autofluorescence, therefore can utilize this character directly to detect the content of amycin in the cell with flow cytometer.
Sensitive cells and mdr cell are laid on 6 orifice plates respectively, and adding final concentration after 24 hours is the cantharidin or derivatives thereof of 2 μ g/ml, cultivates 1h for 37 ℃, washes cell with PBS, trypsinization, and collecting cell, and analyze with flow cytometer.The result is as shown in Figure 4, and cantharidin can increase amycin accumulating in mdr cell, thereby performance chemotherapy sensitizing effect infers that other cantharidin derivatives also bring into play the chemotherapy sensitizing effect through same mechanism.
Pharmaceutical composition and Therapeutic Method
The method of pharmaceutical composition and treatment people's MDR gene-correlation disease such as tumor multi-medicine drug-resistant etc. is also within scope of the present invention.Said pharmaceutical composition comprises cantharidin of the present invention and the derivant and the pharmaceutically suitable carrier of treating effective dose." pharmaceutically suitable carrier " comprises solvent, dispersant (dispersion medium), coating (a coating), antibacterium and antifungal and isotonic agent (isotonic agent) and absorption delay agent (absorption delaying agent) etc.
Pharmaceutical composition of the present invention can be processed the various pharmaceutical dosage forms that are adapted to different way of administration through traditional method.For example, it can be made into oral capsule, gel seal or tablet.Capsule can comprise pharmaceutically acceptable material such as gelatin, the cellulose etc. of any standard.Tablet can be about to pharmaceutical composition and solid phase carrier and lubricant compression by traditional method and make.Said solid phase carrier comprises starch and sugared bentonite (sugar bentonite).Pharmaceutical composition of the present invention also can be made into duricrust tablet (hard shell tablet) or comprises the capsule of binding agent (binder) like lactose or mannitol, conventional filler and tableting agent.Pharmaceutical composition of the present invention also can pass through the parenteral route administration.The parenteral route form of administration comprise pharmaceutical composition of the present invention water preparation, etc. open the sugar juice of saline solution or 5% and the preparation that forms with other pharmaceutically acceptable excipient well known in the art.Cyclodextrin or other chaotropic agents known in those skilled in the art all can be used as pharmaceutical excipient and come submission pharmaceutical composition of the present invention.
Briefly say; Cantharidin of the present invention and derivant thereof can be hanged and be dissolved in pharmaceutically suitable carrier (like physiological solution); Administered through oral or venous transfusion, or through under subcutaneous, the flesh, in intrathoracic, the intraperitoneal, internal rectum, intravaginal, intranasal, gastric, air flue, administrations such as pulmonary injection or transfusion.
The influence that the selection of dosage form receives route of administration, preparation type, patient's (sick kind, the state of an illness, the bodily form, body weight, body surface area, age, sex), medicine influences each other and accept many factors such as diagnosis of doctor for medical treatment.The preparation amount ranges that is suitable for is 0.01～100.00mg/kg.Amount ranges can be done corresponding adjustment with patient and route of administration different.It will depend primarily on the diagnosis of accepting the doctor for medical treatment.For example, oral dose generally will be higher than intravenous injection dosage.Said dosage can be adjusted through experience optimization method well known in the art.Pharmaceutical composition of the present invention is wrapped in appropriate drug submission carrier (like polymer particle body or input equipment) can improves the efficient of administration, particularly oral administration.
The activity of pharmaceutical composition of the present invention can through external ( In vitro) and body in ( In vivo) test and estimate.In brief, the pharmacologically active of pharmaceutical composition of the present invention is reflected on the ability of its mediator MDR activity of gene expression.In the experiment, said pharmaceutical composition is injected in animal (like the mouse model) body to estimate its pharmacologically active in vivo.On this basis, suitable dosage ranges and route of administration are able to confirm then.
For the ease of understanding the present invention, the spy enumerates following examples.Its effect should be understood that it is to explaination of the present invention but not to any type of restriction of the present invention.
Description of drawings
Fig. 1: cantharidin structure chart;
Fig. 2: cantharidin is to the influence of MDR1 promoter activity;
Fig. 3: the influence that cantharidin is expressed MDR1 gene encoding production P-gp;
Fig. 4: the influence of cantharidin to accumulating in the amycin cell;
Fig. 4 A:HepG2 blank control group;
Fig. 4 B:HepG2 adds the ADM group;
Fig. 4 C:HepG2/ADM adds the ADM group;
Fig. 4 D:HepG2/ADM adds ADM and adds the cantharidin group.
The specific embodiment
Below in conjunction with the three experiments specific embodiment, it does not limit the present invention, and scope of the present invention is limited claim.
Embodiment 1
The acquisition of cantharidin and derivant thereof
Cantharidin and derivant thereof can prepare through the commodity purchasing acquisition or through following method:
The cantharidin method for preparing is seen patent 200610040991.1.
The N-methylcantharidimide method for preparing is seen patent 200410067589.3.
The disodium cantharidinate method for preparing is seen patent 200410052938.4.
The method for preparing of norcantharidin is seen Diels, O., Alder, K.Chem.Ber., 1929,62,554.
The Injectio natarii norcantharidatis method for preparing is seen patent 200510045564.8.
Cantharidimide and the nor-cantharidimide of nor-cantharidimide derivant hydroxyethyl, the nor-cantharidimide of methoxy ethyl, the nor-cantharidimide of aminoacid, the nor-cantharidimide of hydroxyl ethylene glycol ethyl, the nor-cantharidimide of Propylene Glycol ethyl, the nor-Mylabris amino acid of glycine dipeptidase, the nor-cantharidimide method for preparing of glycine dipeptidase are seen patent 200310116882.X.
The method for preparing of dehydronorcantharidiimide element, nor-deoxidation cantharidin, Cantharidic acid. or thia cantharidin is seen patent 200510033897.9.
Synthesizing of Cantharidic acid.: in the 50ml round-bottomed flask, place cantharidin 196.2mg (1.0mmol), add distilled water 20.0ml, be heated to and refluxed 0.5 hour, filtration, washing, vacuum drying, column chromatography purification get the 172.4mg white solid.Productive rate is 80.5%.The product warp 1H NMR composes mensuration.Analysis result is following:
1H NMR composes (300MHz, CDCl 3), d:1.24 (s, 6H), 1.73 ~ 1.82 (m, 4H), 4.71 (t, J=2.4 Hz, 2H).
Embodiment 2
Inducing of multidrug resistance cell strain
Respectively human liver cancer cell HepG2, human leukemia cell K562, human colon cancer cell LoVo cell, human breast cancer cell MCF-7, human lung cancer cell A549, gastric carcinoma cells SGC7901, ovarian cancer SKOV3, cervical cancer Hela, renal carcinoma 786-0 or the carcinoma of prostate PC-3 of debita spissitudo is inoculated in 6 orifice plates and cultivates; Add the ADM that final concentration is 0.05 μ g/ml next day; Every liquid that changed at a distance from 2-3 days, the while increases the concentration of ADM gradually; Mortality can appear in cell after dosing 2-3 time, and still visible small amounts of cells is adherent, continues to increase progressively gradually the concentration of ADM, forms one cell clone until the attached cell of remnants; Treating that cell forms big clone, with pancreatin cell dissociation is got off, be uniformly dispersed, continue to add ADM and induce, can be among the ADM of 2 μ g/ml at final concentration until cell, go down to posterity normally, frozen and the recovery.Induce more than 8 months,, keep the drug resistance character of cell with the ADM of 1 μ g/ml.
Embodiment 3
The evaluation of multidrug resistance cell strain
Inoculate sensitive cells and mdr cell respectively in 96 orifice plates, cell density is 5 * 10 4Individual/ml;
Dosing next day, the Concentraton gradient of amycin is: 30,3,0.3,0.03,0.003 μ g/ml; The Concentraton gradient of vincristine is: 10,1,0.1,0.01,0.001 μ g/ml; The Concentraton gradient of paclitaxel is: 10,1,0.1,0.01,0.001 μ g/ml; The Concentraton gradient of 5-FU is: 10,1,0.1,0.01,0.001 μ g/ml, and medicine all carries out doubling dilution with the DMEM culture medium that contains 3%FBS, and negative group adds the DMEM culture medium that contains 3%FBS, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ lMTT (5mg/ml), continues to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ lDMSO, on ELIASA, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate, experimental data is all calculated IC with the SPSS statistical software 50Value, and calculate the drug resistance multiple (IC of drug resistance multiple=persister 50The IC of value/sensitive strain 50Value).The result shows that the multidrug resistance cell strain of various tumor cell lines is induced successfully.
Embodiment 4
The detection of cantharidin and derivant non-toxic thereof
Inoculate L02, HepG2, HepG2/ADM cell respectively in 96 orifice plates, cell density is 5 * 10 4Individual/ml; Dosing next day; The Concentraton gradient of cantharidin and derivant thereof is: 10,1,0.1,0.01,0.001 μ g/ml, and medicine all carries out doubling dilution with the DMEM culture medium that contains 3%FBS, and negative group adds the DMEM culture medium that contains 3%FBS; Establish 3 multiple holes for every group, the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT (5mg/ml, PBS joins), continues to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ l DMSO, on ELIASA, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate, experimental data is all calculated IC with the SPSS statistical software 10Value, the drug level that the cell survival more than 90% is arranged is nontoxic or low toxicity dosage, confirms the working concentration of medicine for subsequent experimental.
Embodiment 5
Cantharidin and derivant thereof are to the chemotherapy sensitizing effect of multidrug resistance tumor cells strain
(1) to the chemotherapy sensitizing effect of drug-resistant cell strain
Inoculate sensitive cells and mdr cell respectively in 96 orifice plates, cell density is 5 * 10 4Individual/ml; Dosing next day, the Concentraton gradient of amycin is: 30,3,0.3,0.03,0.003 μ g/ml; It is high, medium and low three dose groups that cantharidin and derivant thereof are got 2 μ g/ml, 1 μ g/ml, 0.5 μ g/ml respectively; Medicine all carries out doubling dilution with the DMEM culture medium that contains 3%FBS; Negative group adds the DMEM culture medium that contains 3%FBS, establishes 3 multiple holes for every group, and the administration volume is 100 μ l/ holes; Cell is containing 5% CO 237 ℃ of incubators continue to cultivate 44h after, every hole adds 20 μ l MTT (5mg/ml), continues to cultivate 4h; Supernatant is abandoned in suction, and every hole adds 100 μ lDMSO, on ELIASA, and vibration 600s, the OD value at detection 570nm place; More than the independent experiment triplicate, experimental data is all calculated IC with the SPSS statistical software 50Value, and calculate drug resistance reversal fold.The result shows; Cantharidin that the present invention mentioned and derivant thereof comprise that to the kinds of tumors drug-resistant cell strain human liver cancer cell HepG2, human leukemia cell K562, human colon cancer cell LoVo cell, human breast cancer cell MCF-7, human lung cancer cell A549, gastric carcinoma cells SGC7901, ovarian cancer SKOV3, cervical cancer Hela, renal carcinoma 786-0 or carcinoma of prostate PC-3 all have drug resistance inversion effect in various degree.Sensitizer when therefore, above-claimed cpd can be used as tumor pharmacother uses.
(2) chemotherapy sensitization in the body
With 5 * 10 6Individual tumor cell inoculation is subcutaneous in the right axil of BALB/c (nu/nu) nude mice.3 days began the lumbar injection amycin 2 mg/kg/ days behind tumor inoculation, while experimental group intravenous injection cantharidin or derivatives thereof, and dosage is 1.5 mg/kg, every day 1 time, totally 10 times.Measure tumor major diameter a (cm) and perpendicular minor axis b (cm) during the treatment weekly for 2 times and be calculated as follows the heavy W (g) of tumor: W=(a * b 2) * 1/2.After 30 days, put to death animal, strip tumor and weigh, calculate tumour inhibiting rate.The result shows that the tumour inhibiting rate of injecting amycin and cantharidin group simultaneously is far longer than amycin group and cantharidin group tumour inhibiting rate sum.
Embodiment 6
Cantharidin and derivant thereof are to the Mechanism Study of multi-medicine tolerant reversal effect
(1) cantharidin and derivant thereof are to the influence of MDR1 promoter activity
Press the said method of document (Maitra R; Halpin PA; Karlson KH. et al. Differential effects of mitomycin C and doxorubicin on P-glycoprotein expression. Biochem. J. 2001; 355,617-624.) angle and get the MDR1 promoter and be cloned among the pGL3-Basic, thereby construct the pGL3-MDR1-pro reporter plasmid.
In transfection the previous day, mdr cell is inoculated in 24 orifice plates, inoculum density is about 50-60%; Treat to begin when cell reaches 70-80% transfection.During transfection; Change liquid (every hole adds 0.5ml 10% DMEM) at first for the cell in 24 orifice plates; With calcium chloride solution mixing DNA, be about among the calcium chloride 12.5 μ l of 0.375 μ g DNA (0.25 μ g purpose plasmid+0.125 μ g is with reference to plasmid) adding respective volume mixing then.Calcium chloride behind the mixing-DNA mixed liquor is added dropwise in the 12.5 μ l BBS solution abundant mixing, room temperature held 20 minutes.At last BBS-calcium chloride-DNA mixture 25 μ l are added in 24 orifice plates, front and back vibrate gently, make its uniform distribution, and place 37 ℃, 5%CO2 incubator continuation cultivation.After cultivating 6h, inhale and remove culture fluid in the hole, add the fresh complete medium of 2ml, with cell mixing, counting, be laid in 96 orifice plates after the overnight incubation.Inhale behind the 24h and go culture medium, add the DMEM culture medium that contains 3%FBS, in this culture medium, add the cantharidin or derivatives thereof again, making its final concentration is 1 μ g/ml.After continuing to cultivate 24h, cell lysis is surveyed uciferase activity.
Utilize Luciferase Reporter Assay System and FluoStar Optima to detect the uciferase activity of transfectional cell lysate.Get 45 μ L cell pyrolysis liquids; The Assay Cocktail reagent that adds 5 μ L; Add 100 μ Luciferin reactant liquors behind the mixing; Utilize FluoStar Optima instrument to survey the fluorescence counting of 2s, this counting has reflected the activity level of research plasmid reporter gene expression LUC Photinus pyralis LUC Photinus pyralis FL (firefly luciferase).Other gets 20 μ L cell pyrolysis liquids; Add 37.5 μ L β-gal Buffer, add 12.5 μ L ONPG (6mg/mL) again, place 30min for 37 ℃ behind the mixing; Utilize ELIASA to survey the absorption value under the 450nm, this value has reflected that confidential reference items plasmid pCMV-β expresses the activity level of beta galactosidase.(ratio of the activity of β-gal) has been rejected the difference of sample transfection efficiency, more objectively the difference of promoter activity between reflected sample for the activity of LUC Photinus pyralis LUC Photinus pyralis FL (firefly luciferase) and beta galactosidase. ?
(2) cantharidin and derivant thereof are to the influence of MDR1 coded product P-gp expression
Sensitive cells and mdr cell are laid on 6 orifice plates respectively, and adding final concentration after 24 hours is cantharidin and the derivant thereof of 1 μ g/ml, continuation effect 24 hours.With PBS washing 3 times, every hole (6 orifice plate) adds 200 μ l lysate (50mM Tris-HCl (pH7.5), 150mM NaCl, 1mM NaF, 0.5% NP-40, the Aprotinin of 2 μ g/ml with cell; 1mM PMSF), acts on 30 minutes on ice, whenever rocked cell plates, so that the abundant cracking of cell at a distance from 5 minutes.Behind centrifugal 10 minutes of the 15000rpm, supernatant is moved into EP pipe, be placed in after packing-80 ℃ frozen subsequent use.Take out a pipe during electrophoresis and add 4 * albumen sample-loading buffer, boiled 10 minutes, refrigeration on ice, instantaneous centrifugal, last appearance electrophoresis.
Preparation is changeed the film buffer and in 4 ℃ of pre-coolings.Pvdf membrane is soaked 40sec with methanol, and being soaked in after the taking-up changes subsequent use in the film buffer.Change the film buffer with putting in the square plate, under liquid level, carry out the installation of membrane-transferring device.And membrane-transferring device put into electrophoresis tank, and pouring into and change the film buffer, 55V takes out pvdf membrane after changeing film 3h, with membrane marker positive and negative and electrophoresis direction.Wash pvdf membrane 5 minutes 2 times with TBST (0.2% Tween20), 4 ℃ of sealings of the defatted milk powder of reuse 5% are spent the night.Use TBST (0.2% Tween20) to get express developed afterwards 2 times, wash again 15 minutes 1 time, 5 minutes 4 times (all being put in when washing film on the shaking table, as follows).1G2 (2.5mg/ml) with after TBST (0.2% Tween20) the 1:500 dilution, is together packed in the hybridization bag with pvdf membrane, and 37 ℃ of shaking tables are hatched 2h.Take out pvdf membrane, get express developed 2 times, wash 5 minutes 4 times again 15 minutes 1 time with TBST (0.2% Tween20).Two anti-HRP-goat anti-mouse (0.8mg/ml) with after TBST (0.2% Tween20) the 1:2000 dilution, are together packed in the hybridization bag with pvdf membrane, and 37 ℃ of shaking tables are hatched 1h.Take out pvdf membrane, get express developed 2 times, wash 10 minutes 4 times again 15 minutes 1 time with TBST (0.2% Tween20).Pvdf membrane is taken into the darkroom ECL colour developing.The result shows that cantharidin that the present invention mentioned and derivant thereof all can reduce the expression of P-gp in the persister to some extent.
(3) cantharidin and derivant thereof influence to accumulating in the amycin cell
Because amycin has autofluorescence, therefore can utilize this character directly to detect the content of amycin in the cell with flow cytometer.
Sensitive cells and mdr cell are laid on 6 orifice plates respectively, and adding final concentration after 24 hours is the cantharidin or derivatives thereof of 2 μ g/ml, cultivates 1h for 37 ℃, washes cell with PBS, trypsinization, and collecting cell, and analyze with flow cytometer.The result shows that cantharidin that the present invention mentioned and derivant thereof all can increase amycin accumulating in mdr cell to some extent, thus the effect of performance chemotherapy sensitizing.

Claims (3)

1. cantharidin structural modification thing is in the purposes of preparation in the sensitization agent for tumour chemotherapy, and wherein said cantharidin structural modification thing is the following compounds that meets following general structure:
Figure 2011101233823100001DEST_PATH_IMAGE001
Wherein R is CH 3, X 1, X 2Be OH, Y is H 2, then this chemical compound is the deoxidation Cantharidic acid.;
Or R is H, X 1Be HN-CH 2CH 2OCH 2CH 2OCH 3, X 2Be OH, or X 1Be OH, X 2Be HN-CH 2CH 2OCH 2CH 2OCH 3, Y is O, then this chemical compound is the nor-Mylabris amino acid of Propylene Glycol ethyl;
Or R is H, X 1Be HN-CH 2CO-NHCH 2COOH, X 2Be OH, or X 1Be OH, X 2Be HN-CH 2CO-NHCH 2COOH, Y are O, and then this chemical compound is the nor-Mylabris amino acid of glycine dipeptidase;
Or R is H, X 1, X 2Be OH, Y is H 2, then this chemical compound is nor-deoxidation Cantharidic acid.;
Or R is H, X 1, X 2Be ONa, Y is O, and then this chemical compound is an Injectio natarii norcantharidatis.
2. the purposes of the said cantharidin structural modification of claim 1 thing in preparation hepatocarcinoma, colon cancer, pulmonary carcinoma, breast carcinoma, gastric cancer, ovarian cancer, cervical cancer, renal carcinoma, carcinoma of prostate chemotherapeutic sensitizer.
3. the purposes of cantharidin structural modification thing in preparation hepatocarcinoma, colon cancer, pulmonary carcinoma, breast carcinoma, gastric cancer, ovarian cancer, cervical cancer, renal carcinoma, carcinoma of prostate chemotherapeutic sensitizer, wherein said cantharidin structural modification thing is the following compounds that meets following general structure:
Figure 624508DEST_PATH_IMAGE001
Wherein R is CH 3, X 1, X 2Be OH, Y is O, and then this chemical compound is a Cantharidic acid..
CN2011101233823A 2008-07-23 2008-07-23 Application of cantharidin and derivants thereof in preparation of tumor chemotherapy sensitivity enhancing medicine Active CN102210668B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN2011101233823A CN102210668B (en) 2008-07-23 2008-07-23 Application of cantharidin and derivants thereof in preparation of tumor chemotherapy sensitivity enhancing medicine

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN2011101233823A CN102210668B (en) 2008-07-23 2008-07-23 Application of cantharidin and derivants thereof in preparation of tumor chemotherapy sensitivity enhancing medicine

Related Parent Applications (1)

Application Number Title Priority Date Filing Date
CN2008100510174A Division CN101317835B (en) 2008-07-23 2008-07-23 Application of cantharidin and its derivant in preparing sensitization medicament for tumour chemotherapy

Publications (2)

Publication Number Publication Date
CN102210668A CN102210668A (en) 2011-10-12
CN102210668B true CN102210668B (en) 2012-11-07

Family

ID=44742356

Family Applications (1)

Application Number Title Priority Date Filing Date
CN2011101233823A Active CN102210668B (en) 2008-07-23 2008-07-23 Application of cantharidin and derivants thereof in preparation of tumor chemotherapy sensitivity enhancing medicine

Country Status (1)

Country Link
CN (1) CN102210668B (en)

Families Citing this family (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN106083879A (en) * 2016-06-21 2016-11-09 遵义医学院 Norcantharidin mono-acid monoester derivates and antitumor application thereof
CN106008545B (en) * 2016-06-21 2019-02-22 遵义医学院 The compound salt derivative of Norcantharidin and its antitumor application thereof
CN106008546A (en) * 2016-06-21 2016-10-12 遵义医学院 Norcantharidin monoester salt derivative and anti-tumor application thereof

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
王立新等.去甲基斑蝥酸钠脂质微球体内外评价.《药学学报》.2006,第41卷(第8期),784-788页. *
魏素菊等.斑蝥酸钠逆转K562/AO2细胞.《实用肿瘤杂志》.2007,第22卷(第5期),401-404页. *

Also Published As

Publication number Publication date
CN102210668A (en) 2011-10-12

Similar Documents

Publication Publication Date Title
CN103221373A (en) Methods and compositions for treating lung cancer
CN108358973A (en) Naphthalimide tetravalence platinum-like compounds, preparation method and its application in preparation of anti-tumor drugs
CN104163823B (en) A kind of camptothecine and Artesunate conjugate and preparation method thereof and application
CN106478569A (en) Isoalantolactone derivative and its salt
CN101624443A (en) Preparation method for amycin pro drug and application thereof
CN102475710B (en) Application of diosgenin and derivative thereof for preparing tumor chemotherapy sensitization medicines
CN102210668B (en) Application of cantharidin and derivants thereof in preparation of tumor chemotherapy sensitivity enhancing medicine
CN102516344B (en) Compound with antitumor activity and preparation method and application thereof
CN102526073A (en) Application of mogrol H9 for preparing antitumor drugs
CN110123809A (en) 5- methyl-dihydro benzofuran-application of the imidazole salt compound in pharmacy
CN110041375A (en) Compound, preparation method and its application in preparation of anti-tumor drugs with asymmetric monosubstituted naphthalimide tetravalence platinum structure
CN101317835B (en) Application of cantharidin and its derivant in preparing sensitization medicament for tumour chemotherapy
CN104557909B (en) 3- acyloxy replaces dextrorotation deoxidation tylophorinine derivative, its preparation method and pharmaceutical composition and purposes
CN103893168A (en) Application of isopimpinellin in preparation of antitumor drugs and anticancer reversal agents
CN104138369A (en) Anticancer drug
CN108030777B (en) Chloroguanide application in preparation of anti-tumor drugs
CN105213366B (en) The medical usage and its pharmaceutical composition of gamboge ketone compound
CN107108693A (en) New peptide compounds, Its Preparation Method And Use
CN107056739B (en) Bola type quercetin derivative and its preparation method and application
CN108484637A (en) Target anticancer new drug X-76 salt-forming compounds and application thereof, preparation method
CN106822129A (en) Application of the aloperine derivative in the medicine for preparing treatment tumour
CN103012394B (en) Rhodanine derivative and preparation method thereof
CN107880060A (en) Polyether compound purposes
CN102499927B (en) Medicine composition for preventing and curing tumors
CN103417536A (en) Applications of harmol in preparation of antitumor drugs

Legal Events

Date Code Title Description
C06 Publication
PB01 Publication
C10 Entry into substantive examination
SE01 Entry into force of request for substantive examination
C14 Grant of patent or utility model
GR01 Patent grant
ASS Succession or assignment of patent right

Owner name: NORTHEAST NORMAL UNIVERSITY

Free format text: FORMER OWNER: INST. OF GENETICS AND CYTOLOGY, NORTHEAST NORMAL UNIV.

Effective date: 20131230

C41 Transfer of patent application or patent right or utility model
COR Change of bibliographic data

Free format text: CORRECT: ADDRESS; FROM: 130021 CHANGCHUN, JILIN PROVINCE TO: 130022 CHANGCHUN, JILIN PROVINCE

TR01 Transfer of patent right

Effective date of registration: 20131230

Address after: 130022 Jilin City, Changchun province people's street, No. 5268

Patentee after: Northeast Normal University

Address before: 130021 No. 5268 Renmin Street, Jilin, Changchun

Patentee before: Inst. of Genetics and Cytology, Northeast Normal Univ.