CN106512022A - Application of hydroxysafflor yellow A-red blood cell adhesion chondroitin sulfate A receptor protein polypeptide compound to preparing of antitumor drug - Google Patents
Application of hydroxysafflor yellow A-red blood cell adhesion chondroitin sulfate A receptor protein polypeptide compound to preparing of antitumor drug Download PDFInfo
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Abstract
The invention provides application of hydroxyl saffloryellow A-red blood cell adhesion chondroitin sulfate A receptor protein polypeptide compound (VAR2CSA-PEG-Hydroxy Saffloryellow A) to preparing of an antitumor drug. The structural formula of the hydroxysafflor yellow A-red blood cell adhesion chondroitin sulfate A receptor protein polypeptide compound (VAR2CSA-PEG-Hydroxy Saffloryellow A) is shown in the specification. In addition, a drug composition for cancer therapy and adjuvant therapy is provided. The composition is composed of the hydroxysafflor yellow A-red blood cell adhesion chondroitin sulfate A receptor protein polypeptide compound (VAR2CSA-PEG-Hydroxy Saffloryellow A) with the single dose being 1 mg-5 mg and a carrier which is acceptable in pharmacy. The composition has a very obvious inhibiting effect on malignant tumors of the epithelial tissue such as the ovarian cancer, the lung cancer, the uterine cancer and the testicular cancer.
Description
Technical field
The present invention relates to synthetic drug, and in particular to hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) acceptor egg
White polypeptide complex（VAR2CSA-PEG-Hydroxy Saffloryellow A）Preparing the application of antineoplastic.
Background technology
The provided nutrition of growth needs blood vessel of tumour and exclusion metabolite, if there is no the generation of new vessels,
Tumour can only be in the small state of 1 ~ 2mm for a long time.Once new capillary vessel progress tumor tissues, tumour are then increased rapidly
And which invades profit and transfer ability is also greatly reinforced.Therefore, how to block Tumor Angiongesis has become treating cancer in the world
Focus.
In the generating process of new blood vessel, endothelial cell activation and propagation be most basic and most important link.It is logical
Crossing suppression vascular endothelial cell proliferation can just suppress new vessels to generate.Due to tumour cell and vascular endothelial cell it is mutually interdependent
Rely and mediated neonate tumour blood vessel, therefore, study the process of Vitro Tumor angiogenesis, it is desirable to which simulation is in vivo with tumour cell altogether
Raw process, i.e.,：Vascular endothelial cell is grown under conditions of co-cultivation with tumour cell.
Research shows that the associated angiogenesis of at least 30 kinds growth factors and tumour wherein most directly promote blood vessel life
It is vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) into the factor.The blood vessel of tumour
Endothelial cell goes back the acceptor of great expression VEGF and bFGF, by vasoactive endothelium in addition to high VEGF expression and bFGF
These acceptors of cell surface and induction of vascular is newborn.
Hydroxyl radical carthamin yellow carthamus A (Hydroxy Saffloryellow A, HSYA) can by suppress tumour VEGF and
The expression of bFGF, so as to suppress tumor vascular growth, reaches antitumor action；Red blood cell sticks chondroitin sulfate A (CSA) acceptor egg
In vain（VAR2CSA）Adhesion mediation with placental receptor (Chondroitin sulfate A, CSA) is combined, and CSA is vigorous in growth
Tumour cell in altimeter reach, hydroxyl radical carthamin yellow carthamus A and red blood cell are sticked into chondroitin sulfate A (CSA) receptor protein
（VAR2CSA）After coupling, in the tumour cell that the entrance CSA altimeters that hydroxyl radical carthamin yellow carthamus A is targetted can be reached, so as to reach
To orientation antineoplastic action.
Present invention discover that hydroxyl radical carthamin yellow carthamus A (Hydroxy Saffloryellow A, HSYA) and red blood cell are sticked
Chondroitin sulfate A (CSA) receptor protein（VAR2CSA）It is coupled, forms hydroxyl radical carthamin yellow carthamus A-VAR2CSA polypeptide complexes, Neng Gouyou
Effect suppresses the growth of kinds of tumor cells, preferably further researchs and develops new drug.
The content of the invention
The technical problem to be solved is that hydroxyl radical carthamin yellow carthamus A-red blood cell sticks sulfuric acid to research and design is soft
Ossein A receptor protein polypeptide compounds（VAR2CSA-PEG-Hydroxy Saffloryellow A）Preparing antineoplastic
Application.
The invention provides hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) receptor protein polypeptide compound
（VAR2CSA-PEG-Hydroxy Saffloryellow A）Preparing the application of antineoplastic.
Hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) receptor protein polypeptide compound（VAR2CSA-PEG-
Hydroxy Saffloryellow A）Structural formula it is as follows：
。
Hydroxyl radical carthamin yellow carthamus A-red blood cell of the present invention sticks chondroitin sulfate A (CSA) receptor protein polypeptide compound
（VAR2CSA-PEG-Hydroxy Saffloryellow A）Prepared by following method：
By hydroxyl radical carthamin yellow carthamus A (Hydroxy Saffloryellow A, HSYA),N,N'-succinimidyl carbonate
(N, N '-Disuccinimidyl Carbonate, DSC) and amino polyethylene glycol（NPEG）In molar ratio 1：20：20 third
Stirring reaction more than 6 hours in ketone, by gained hydroxyl radical carthamin yellow carthamus A-polyethylene glycol complex（HSYA-NPEG）, reduce pressure dry
It is dry, after removing acetone, with purifying water dissolves, extracted by ethyl acetate repeatedly, by acetic acid ethyl ester extract 200-300 purposes
Thin layer silica gel is separated, and carries out gradient elution with methylene chloride-methanol, and thin-layer chromatography control test collects Sydroxy carthamin
A- polyethylene glycol complexes（HSYA-NPEG）It is dissolved in dimethyl sulfoxide（Dimethyl sulfoxide, DMSO）In；Will be red thin
Born of the same parents stick chondroitin sulfate A (CSA) receptor protein（VAR2CSA）With 1- (3- dimethylamino-propyls) -3- ethyl-carbodiimide hydrochlorides
（1- (3-Dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride, EDC）,N- hydroxysuccinimidyl
Acid imide（N- Hydroxysuccinimide, NHS）Activation, in molar ratio 1：30 ratio is by hydroxyl radical carthamin yellow carthamus A-poly- second
Glycol compound（HSYA-NPEG）Stick chondroitin sulfate A (CSA) receptor protein with the red blood cell for having activated（VAR2CSA）Mixing, 4 DEG C
Stirring 24 hours, takes out dialysis desalination, and over-molecular sieve, collection end-product obtain hydroxyl radical carthamin yellow carthamus A-red blood cell and stick sulfuric acid
Chondroitin A receptor protein polypeptide compound（VAR2CSA-PEG-Hydroxy Saffloryellow A）.
Hydroxyl radical carthamin yellow carthamus A-red blood cell is sticked chondroitin sulfate A (CSA) receptor protein polypeptide compound by the present invention
（VAR2CSA-PEG-Hydroxy Saffloryellow A）The animal test of pesticide effectiveness is carried out, Sydroxy carthamin is as a result shown
A- red blood cells stick chondroitin sulfate A (CSA) receptor protein polypeptide compound（VAR2CSA-PEG-Hydroxy Saffloryellow
A）The growth of the malignant cell of the epithelial tissues such as oophoroma, lung cancer, the cancer of the uterus and carcinoma of testis can be suppressed, cancer cell is turned
Shifting has obvious inhibitory action；Hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) receptor protein polypeptide compound
（VAR2CSA-PEG-Hydroxy Saffloryellow A）Nest cancer, lung will be slower than to the inhibition of other tumour cells
Cancer, the cancer of the uterus and carcinoma of testis etc., show that hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) receptor protein polypeptide and is combined
Thing（VAR2CSA-PEG-Hydroxy Saffloryellow A）The tumour cell of CSA acceptors overexpression is enriched in easily, it is right
The effect of other cells is much smaller.Therefore, hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) receptor protein polypeptide
Compound（VAR2CSA-PEG-Hydroxy Saffloryellow A）Can be used for prepare treatment oophoroma, lung cancer, the cancer of the uterus and
The medicine of the malignant tumours such as carcinoma of testis.
Hydroxyl radical carthamin yellow carthamus A-red blood cell of the present invention sticks chondroitin sulfate A (CSA) receptor protein polypeptide compound
（VAR2CSA-PEG-Hydroxy Saffloryellow A）Preparing treatment treatment oophoroma, lung cancer, the cancer of the uterus and carcinoma of testis
It is that chondroitin sulfate A (CSA) receptor protein polypeptide compound is sticked by hydroxyl radical carthamin yellow carthamus A-red blood cell etc. the medicine of malignant tumour
（VAR2CSA-PEG-Hydroxy Saffloryellow A）As active component and drug regimen made by conventional pharmaceutical carrier
Thing.Described pharmaceutical composition can be tablet, dispersible tablet, lozenge, oral disintegrating tablet, sustained release tablets, capsule, soft capsule, dripping pill,
Granula, injection, powder-injection or aerosol etc..There is larger clinical value.
Description of the drawings
Fig. 1 red blood cells stick chondroitin sulfate A (CSA) receptor protein（VAR2CSA）Scheme with tumour cell A549 specific bindings
1 in figure：rContr、2：RVAR2CSA groups, 3：CSA groups
Fig. 2 red blood cells stick chondroitin sulfate A (CSA) receptor protein（VAR2CSA）Scheme with tumour cell skov-3 specific bindings
1 in figure：rContr、2：RVAR2CSA groups, 3：CSA groups
Fig. 3 red blood cells stick chondroitin sulfate A (CSA) receptor protein（VAR2CSA）Scheme with tumour cell ishikawa specific bindings
1 in figure：rContr、2：RVAR2CSA groups, 3：CSA groups
Fig. 4 red blood cells stick chondroitin sulfate A (CSA) receptor protein（VAR2CSA）Scheme with tumour cell HUVEC specific bindings
1 in figure：rContr、2：RVAR2CSA groups, 3：CSA groups
Fig. 5 difference group people's venous endothelial cell proliferative conditions；
1 in figure：Normal group, 2：Model group, 3：Administration group；*P<0.01, and normally compare； ** P<0.05, with model group ratio
Compared with
Effect of Fig. 6 hydroxyl radical carthamin yellow carthamus As to abnormality proliferation HUVEC apoptosis
1 in figure：Normal group, 2：Model group, 3：Administration group；*P<0.01, and normally compare； ** P<0.05, with model group ratio
Compared with
Fig. 7 hydroxyl radical carthamin yellow carthamus As-red blood cell sticks the external suppression tumour of chondroitin sulfate A (CSA) receptor protein polypeptide compound
Experiment
1 in figure：Control group, 2：It is high that hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) receptor protein polypeptide compound
Dosage group, 3：Hydroxyl radical carthamin yellow carthamus A-red blood cell stick chondroitin sulfate A (CSA) receptor protein polypeptide compound middle dose group, 4：Hydroxyl
Base carthamus tinctorius yellow colour A-red blood cell stick chondroitin sulfate A (CSA) receptor protein polypeptide compound it is low, 5：Endoxan group.
Specific embodiment
The invention discloses a kind of hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) receptor protein polypeptide being combined
Thing（VAR2CSA-PEG-Hydroxy Saffloryellow A）The application of antineoplastic is being prepared, with reference to concrete reality
Apply example to be further described the present invention, but protection scope of the present invention is not limited thereto, it is any to be familiar with this technology neck
The technical staff in domain the invention discloses technical scope in, technology according to the present invention scheme and its inventive concept are equal to
Replace or change, should all be included within the scope of the present invention.
In following embodiments, material used, reagent etc., if no special instructions, commercially obtain.
1 hydroxyl radical carthamin yellow carthamus A of embodiment-red blood cell sticks chondroitin sulfate A (CSA) receptor protein polypeptide compound
（VAR2CSA-PEG-Hydroxy Saffloryellow A）It is coupled
The round-bottomed flask of a cleaning is taken, hydroxyl radical carthamin yellow carthamus A (Hydroxy Saffloryellow A, HSYA) is weighed respectively
(purity >=95%, purchased from middle inspection institute) 0.612g,N,N'-succinimidyl carbonate (N, N '-Disuccinimidyl
Carbonate, DSC) (purity >=95%, purchased from Sigma-Aldrich) 5.2g and amino polyethylene glycol（NPEG）It is (senior
It is pure, purchased from the U.S.) 6mL, 250mL acetone (chromatographically pure, purchased from Sigma-Aldrich) is added, stirring reaction more than 6 hours；
By gained hydroxyl radical carthamin yellow carthamus A-polyethylene glycol complex（HSYA-NPEG）, remove acetone under reduced pressure, add 100mL purified waters
Dissolving, adds 100mL ethyl acetate and extracts repeatedly, extracts three times；Acetic acid ethyl ester extract is collected, drying under reduced pressure concentrates second
Acetoacetic ester extract is to about 50mL；Acetic acid ethyl ester extract is separated with the thin layer silica gel of 200-300 mesh, with dichloromethane-first
Alcohol carries out gradient elution, and thin-layer chromatography control test collects hydroxyl radical carthamin yellow carthamus A-polyethylene glycol complex（HSYA-NPEG）
Component, drying under reduced pressure；By solid crude product with 20 mL ethyl acetate/methanols (volume ratios 3:1) mixed solvent dissolving, using dry
Method dress post carries out pillar layer separation with methyl alcohol as eluant, eluent, obtains brown powder target product；Gained object is dissolved in
20mL dimethyl sulfoxide (DMSO)s（Dimethyl sulfoxide, DMSO）In (chromatographically pure, purchased from Sigma-Aldrich)；Accurately claim
Take 0.10g red blood cells and stick chondroitin sulfate A (CSA) receptor protein（VAR2CSA）It is dissolved in 20mL MES buffer solutions（pH6.0）In, plus
Enter 0.286g 1- (3- dimethylamino-propyls) -3- ethyl-carbodiimide hydrochlorides（1-(3-Dimethylaminopropyl)-3-
Ethylcarbodiimide hydrochloride, EDC）(chromatographically pure, purchased from Sigma-Aldrich) and 0.365g N- hydroxyl
Base succinimide（N- Hydroxysuccinimide, NHS）(chromatographically pure, purchased from Sigma-Aldrich) is stirred at room temperature reaction
30min, will be dissolved in the hydroxyl radical carthamin yellow carthamus A-polyethylene glycol complex of DMSO（HSYA-NPEG）With the red blood cell for having activated
Stick chondroitin sulfate A (CSA) receptor protein（VAR2CSA）Mixing, is stirred at room temperature 30min；Reaction terminates, with 4 DEG C of phosphate buffer
Dialysis desalination, changes a not good liquor per 6h, and collection end-product obtains hydroxyl radical carthamin yellow carthamus A-red blood cell and sticks chondroitin sulfate A (CSA) acceptor
Polypeptide compound（VAR2CSA-PEG-Hydroxy Saffloryellow A）.
2 red blood cell of embodiment sticks chondroitin sulfate A (CSA) receptor protein（VAR2CSA）With the specific binding of tumour
1. Preparatory work of experiment
Complete medium：The hyclone of incomplete culture medium+10%（GIBCO）, standby after mixing, 4 DEG C of preservations.It is wherein not complete
Full culture medium selects RPMI1640 culture mediums（GIBCO）
The preparation of variable concentrations CSA：Electronic balance weighs a small amount of tested material（1-3mg）, dissolved with a small amount of PBS, with corresponding culture
Basigamy makes the solution of 7 concentration gradients, and after 0.22 μm of first concentration is filtered, concentration doubling dilution below is prepared.
2. experiment packet and experimental program
4 cells are had, each cell is respectively divided into 3 groups, per group of 3 holes.Respectively rContr, rVAR2CSA group, CSA groups.Point
A549, skov-3, ishikawa, HUVEC exponential phase cell is not taken（It is purchased from Sheng Ke institutes of Chinese Academy of Sciences cell bank）, it is inoculated in
96 orifice plates（Purchased from Corning companies）In, per 100 μ L of hole, after 24h, it is separately added into rContr, rVAR2CSA and CSA, control group
Add 100 μ L of culture medium, control group and detection group to be all provided with 3 multiple holes per hole, separately set blank control group, be not added with cell and only add culture medium
Conduct zeroing group.96 orifice plates are placed in into 37 DEG C, 5%CO2Incubator culture, when cell growth is to 70%-80% density, goes
Supernatant, PBS of the incubation containing 2% hyclone, 4 DEG C of incubation 30min, being subsequently adding V5-FITC antibody carries out secondary incubating
Educate, after incubation, flow cytometer detection is analyzed.
3rd, result
1st, flow cytomery MFI values analysis
In here experiment, human cancer cell line A549, skov-3, ishikawa and the mankind that we test patient source is normal
The Percentage bound of the HUVEC and rVAR2CSA of cell derived, each cell are respectively divided into 3 groups, per group of 3 multiple holes.RContr groups（Control
Group）RContr, rVAR2CSA groups is only added only to add rVAR2CSA, CSA groups to add rVAR2CSA and CSA simultaneously.Add medicine
After take out when cell grows to 70%-80% density, after adding V5-FITC antibody incubations, carry out MFI with flow cytometer (average
Fluorescence intensity) detection.
According to obtaining a result after MFI value analysis and assessment, rVAR2CSA and human cancer cell line show high-bonding-ratio, while
After adding inhibitor C SA, Percentage bound is substantially reduced, and illustrates that rVAR2CSA has the function of identification human cancer cell.
Effect of 3 hydroxyl radical carthamin yellow carthamus A of embodiment to HUVEC
The HUVEC models of abnormality proliferation are set up, HSYA is added（Purchased from Nat'l Pharmaceutical ＆ Biological Products Control Institute's standard items）Afterwards, MTT
Detection cell proliferative conditions, Flow cytometry cell cycle and apoptosis rate.
Experiment packet and experimental program：
Normal group：Normal HUVEC
Model group：Normal HUVEC+SGC7901 supernatants（50%）
Administration group：Normal HUVEC+SGC7901 supernatants（50%）+HSYA(0.075g/L)
Normal HUVEC 80% is passed on after merging, by 1 × 105It is inoculated in 25cm2Blake bottle in, add 2.5mL/ bottles training
Nutrient solution, it is after cell attachment 6h, random to be grouped, nutrient solution is suctioned out, normal group adds the RPMI-1640 of 5mL/ bottles（It is purchased from
GIBCO）, model group RPMI-1640 5ml/ bottle of the addition containing 50% A549 tumor supernatants, administration group are added containing 50% A549
The common 5mL/ bottles of 250 μ L of RPMI-1640 4.75mL and 0.075g/L HSYA of tumour supernatant, after continuing culture 20h, collect thin
Born of the same parents, for FCM analysis.
HUVEC cells 80% are passed on after merging, and cell concentration is adjusted to 0.5 × 10 with culture medium5Individual/ml, is inoculated in 96
Orifice plate, puts in incubator after adherent 6 hours, suctions out supernatant, and normal group adds 200 μ L/ holes of RPMI-1640.37℃,5%CO2Point
5mg/mL MTT Pei Yang not be added after 48 hours（Purchased from Sigma-Aldrich）10 μ L/ holes, continue culture 4h, remove supernatant, then
Add 100 μ L/ holes of DMSO, ELIASA（Purchased from match Mo Feishier companies）Survey 560nm OD values.
Cell concentration is adjusted to 106Individual/mL, adds ice ethanol（75%）It is resuspended, it is fixed, add the cold PBS of 1mL（It is purchased from
Hyclone, GE）, gently shaking makes cell suspend.1000rpm, 4 DEG C of centrifugations, 10 minutes, abandons supernatant, stays precipitation, and the process is entered
Row is twice.Cell is resuspended in into 200 μ L Binding Buffer.Add 10 μ L propidium iodides（PI）Dyeing, gently mixes, keeps away
Light or room temperature reaction react 30 minutes for 15 minutes or 4 DEG C, add 300 μ L Binding Buffer, flow cytometer（Purchased from U.S.
BD companies of state）Detection.
Experimental result：
1）MTT is detected
Administration group cell absorbance is significantly lower than model group.Compare with model group, administration group cell OD values are significantly lower than model group,
Illustrate that administration group cell growth receives suppression.
2）Flow cytometer detection
Administration group Apoptosis peak is significantly raised；Administration group apoptosis rate is apparently higher than model group.Compared with model group, administration
Ratio shared by the group cell S phases is substantially reduced, and the change of other each groups is not obvious.Illustrate that administration group HSYA suppresses cell life
It is long, and apoptosis may occur.
Embodiment 4：Hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) receptor protein polypeptide compound
（VAR2CSA-PEG-Hydroxy Saffloryellow A）External suppression tumor experiment
The HSYA of low concentration is a kind of stronger AI, after HSYA chains VAS2CSA, specific recognition tumour
Tissue, is finally reached effect of Growth of Tumors Transplanted.It is thin that hydroxyl radical carthamin yellow carthamus A-red is studied in this experiment by animal model
Born of the same parents stick chondroitin sulfate A (CSA) receptor protein polypeptide compound（VAR2CSA-PEG-Hydroxy Saffloryellow A）To stomach
The inhibition of cancer transplantable tumor.
Experiment packet and experimental program：
A549 cell lines are dressed to sufficient amount, when exponential phase is in adherent cell, are disappeared with 0.25% pancreatin
Change, count under inverted microscope, and observation of cell vigor is more than 95%, adjust concentration, be configured to 1 × 107Individual/mL is unicellular
Suspension, is only inoculated in C57BL/6 mouse by 0.2mL/（It is purchased from Guangdong Province's animal experimental center）Left rib abdomen is subcutaneous, replicates Mouse Stomach
Cancer Transplanted tumor model.Five groups are randomly divided into after being inoculated with 1 week, 6 per group.Inoculation next day, hydroxyl radical carthamin yellow carthamus A-red thin
Born of the same parents stick chondroitin sulfate A (CSA) receptor protein polypeptide compound（VAR2CSA-PEG-Hydroxy Saffloryellow A）Greatly,
In, small dose group difference lumbar injection respective concentration medicine（1.4th, 0.7,0.35mg/kg/ only, 2 times/d）；Model group gives
Amount sterile saline（0.2mL/, 2 times/d）；Endoxan inoculation group starts intraperitoneal injection of cyclophosphamide on the 2nd day（25mg/
Kg/, per 1 time on the 2nd）.22nd day mouse weights, put to death.
It is inoculated with the 22nd day, after whole sacrifices, routine disinfection, completely strips the left rib abdomen hypodermic tumour of mouse, use electronics
Balance weighs knurl weight, calculates the inhibition rate of tumor growth of each group.
Experimental result：
After concentration when the 7th day whole C57BL/6 mouse naked eyes be it is visible have microspike under the stomach wall of left side, size about 0.25 ×
0.22cm or so subcutaneous nodules, gradually substantially, have gone out knurl, tumor formation rate 100%；From tumor growth curve it can be seen that tumour body
It is physiological saline group that product growth is most fast, and it is endoxan group that growth is most slow, medication the 14th day, and each group gross tumor volume is set up out
Existing difference, physiological saline group are substantially accelerated with the heavy dose of group mice-transplanted tumor system growths of HSYA, and HSYA small dose groups mouse moves
Plant knurl volume growth relatively slow, the growth of CTX groups is most slow.When 16d, each group mice-transplanted tumor volume increases all obvious
Accelerate, wherein HSYA each groups tumor volume change is most obvious.
Claims (5)
1. hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) receptor protein polypeptide compound（VAR2CSA-PEG-
Hydroxy Saffloryellow A）Preparing the application of antineoplastic, it is characterised in that the Sydroxy carthamin
A- red blood cells stick chondroitin sulfate A (CSA) receptor protein polypeptide compound（VAR2CSA-PEG-Hydroxy Saffloryellow
A）Structural formula it is as follows：
。
2. according to claim 1, it is characterised in that the hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) to be received
Body protein polypeptide complex（VAR2CSA-PEG-Hydroxy Saffloryellow A）Prepared by following method：
By hydroxyl radical carthamin yellow carthamus A (Hydroxy Saffloryellow A, HSYA),N,N'-succinimidyl carbonate
(N, N '-Disuccinimidyl Carbonate, DSC) and amino polyethylene glycol（NPEG）In molar ratio 1：20：20 third
Stirring reaction more than 6 hours in ketone, by gained hydroxyl radical carthamin yellow carthamus A-polyethylene glycol complex（HSYA-NPEG）, reduce pressure dry
It is dry, after removing acetone, with purifying water dissolves, extracted by ethyl acetate repeatedly, by acetic acid ethyl ester extract 200-300 purposes
Thin layer silica gel is separated, and carries out gradient elution with methylene chloride-methanol, and thin-layer chromatography control test collects Sydroxy carthamin
A- polyethylene glycol complexes（HSYA-NPEG）It is dissolved in dimethyl sulfoxide（Dimethyl sulfoxide, DMSO）In, by red blood cell
Stick chondroitin sulfate A (CSA) receptor protein（VAR2CSA）With 1- (3- dimethylamino-propyls) -3- ethyl-carbodiimide hydrochlorides（1-
(3-Dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride, EDC）,N- hydroxysuccinimidyl acyl is sub-
Amine（N- Hydroxysuccinimide, NHS）Activation, in molar ratio 1：30 ratio is by hydroxyl radical carthamin yellow carthamus A-polyethylene glycol
Compound（HSYA-NPEG）Stick chondroitin sulfate A (CSA) receptor protein with the red blood cell for having activated（VAR2CSA）Mixing, 4 DEG C of stirrings
24 hours, dialysis desalination is taken out, over-molecular sieve, collection end-product obtain hydroxyl radical carthamin yellow carthamus A-red blood cell and stick chondroitin sulfate
Plain A receptor protein polypeptides compound（VAR2CSA-PEG-Hydroxy Saffloryellow A）.
3. according to claim 1, it is characterised in that a kind of medicine is to stick sulfuric acid by hydroxyl radical carthamin yellow carthamus A-red blood cell
Chondroitin A receptor protein polypeptide compound（VAR2CSA-PEG-Hydroxy Saffloryellow A）As active component with
Pharmaceutical composition made by pharmaceutical carrier.
4. according to claim 3, a kind of pharmaceutical composition it is characterized in that described pharmaceutical composition be tablet, dispersible tablet,
Lozenge, oral disintegrating tablet, sustained release tablets, capsule, soft capsule, dripping pill, granule, injection, powder-injection or aerosol.
5. according to claim 3, it is characterised in that the single dose hydroxyl carthamin yellow A-containing of described pharmaceutical composition-red
Cell adhesion chondroitin sulfate A (CSA) receptor protein polypeptide compound（VAR2CSA-PEG-Hydroxy Saffloryellow A）For
1mg-5 mg。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610889714.1A CN106512022B (en) | 2016-10-12 | 2016-10-12 | Hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) receptor protein polypeptide compound in the application for preparing anti-tumor drug |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610889714.1A CN106512022B (en) | 2016-10-12 | 2016-10-12 | Hydroxyl radical carthamin yellow carthamus A-red blood cell sticks chondroitin sulfate A (CSA) receptor protein polypeptide compound in the application for preparing anti-tumor drug |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN106512022A true CN106512022A (en) | 2017-03-22 |
| CN106512022B CN106512022B (en) | 2019-09-27 |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109387627A (en) * | 2018-10-16 | 2019-02-26 | 中国科学院深圳先进技术研究院 | A kind of Reagent Protocol of screening for cancer and early diagnosis based on placenta sample chondroitin sulfate A (CSA) |
| CN112789504A (en) * | 2018-07-13 | 2021-05-11 | 瓦克特诊断有限责任公司 | Isolation of fetal-derived circulating cells using the recombinant malaria protein VAR2CSA |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101744802A (en) * | 2008-11-28 | 2010-06-23 | 张前 | Novel application of hydroxysafflor yellow A to preparation of anti-tumour drugs |
| CN104136041A (en) * | 2012-02-09 | 2014-11-05 | Var2制药有限公司 | Targeting of chondroitin sulfate glycans |
| WO2015095952A1 (en) * | 2013-12-27 | 2015-07-02 | The Centre For Drug Research And Development | Var2csa-drug conjugates |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101744802A (en) * | 2008-11-28 | 2010-06-23 | 张前 | Novel application of hydroxysafflor yellow A to preparation of anti-tumour drugs |
| CN104136041A (en) * | 2012-02-09 | 2014-11-05 | Var2制药有限公司 | Targeting of chondroitin sulfate glycans |
| WO2015095952A1 (en) * | 2013-12-27 | 2015-07-02 | The Centre For Drug Research And Development | Var2csa-drug conjugates |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112789504A (en) * | 2018-07-13 | 2021-05-11 | 瓦克特诊断有限责任公司 | Isolation of fetal-derived circulating cells using the recombinant malaria protein VAR2CSA |
| CN109387627A (en) * | 2018-10-16 | 2019-02-26 | 中国科学院深圳先进技术研究院 | A kind of Reagent Protocol of screening for cancer and early diagnosis based on placenta sample chondroitin sulfate A (CSA) |
| CN109387627B (en) * | 2018-10-16 | 2021-09-24 | 中国科学院深圳先进技术研究院 | Reagent method for screening and early diagnosis of cancer based on placenta-like chondroitin sulfate A |
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