CN106236752A - Drug combination antitumor drug, preparation method and application - Google Patents
Drug combination antitumor drug, preparation method and application Download PDFInfo
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- CN106236752A CN106236752A CN201610613652.1A CN201610613652A CN106236752A CN 106236752 A CN106236752 A CN 106236752A CN 201610613652 A CN201610613652 A CN 201610613652A CN 106236752 A CN106236752 A CN 106236752A
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
- A61K31/47—Quinolines; Isoquinolines
- A61K31/4738—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems
- A61K31/4745—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems condensed with ring systems having nitrogen as a ring hetero atom, e.g. phenantrolines
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/21—Esters, e.g. nitroglycerine, selenocyanates
- A61K31/215—Esters, e.g. nitroglycerine, selenocyanates of carboxylic acids
- A61K31/22—Esters, e.g. nitroglycerine, selenocyanates of carboxylic acids of acyclic acids, e.g. pravastatin
- A61K31/222—Esters, e.g. nitroglycerine, selenocyanates of carboxylic acids of acyclic acids, e.g. pravastatin with compounds having aromatic groups, e.g. dipivefrine, ibopamine
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
- A61K47/34—Macromolecular compounds obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyesters, polyamino acids, polysiloxanes, polyphosphazines, copolymers of polyalkylene glycol or poloxamers
Abstract
The invention discloses a kind of drug combination antitumor drug, including anti-angiogenic medicaments, anti-tumour cell proliferative medicine and amphipathy macromolecule material；Described anti-angiogenic medicaments is the prodrug of Combretastatin A-4 4, and described anti-tumour cell proliferative medicine is camptothecin antineoplastic agents.The invention also discloses a kind of drug combination anti-tumor medicinal preparation, preparation method and application.Cytotoxicity experiment shows that the nanoparticle in the present invention can significantly inhibit tumor cell (HT 29) and the propagation of human umbilical vein endothelial cell (HUVEC), and has dose-dependence；Scratch damage and segment dislocation experiment also confirm that in the present invention, the nanoparticle of preparation can suppress migration and the vascularization of HUVEC.Contrasting independent medication system, the drug combination nanoparticle provided in the present invention shows preferable tumor killing effect in vivo, has good clinical value and application prospect.
Description
Technical field
The invention belongs to pharmaceutical synthesis field, be specifically related to a kind of anti-angiogenic medicaments and anti-tumour cell proliferative medicine
Thing drug combination antitumor drug, preparation method and application.
Background technology
SN38 (SN38) is the activated product of clinical antitumor agents irinotecan (CPT-11).
SN38 is with topoisomerase as targeting moiety, and suppression topoisomerase I-DNA complex combines, thus stops fracture strand again
Connect, make DNA double chain structure untwist and play antitumor action.SN38 shows higher activity in vitro, is CPT-11 activity
100～1000 times.Therefore, directly utilize SN38 molecule, avoid that CPT-11 is carried out enzymolysis thus discharge the approach of SN38, can
Hope the antitumous effect being effectively improved medicine.But the water solublity of SN38 is very poor and molten insoluble in acceptable on pharmaceutics
Agent (such as tween or polyoxyethylene castor oil), therefore cannot be used directly for clinical injection.
Additionally, tumor therapy clinical research shows, the long-time chemotherapeutics using single suppression growth of tumour cell
Drug resistance can be produced.Along with deepening continuously of Anti-angiogenic therapy tumor correlational study scheme, anti-angiogenic medicaments use in conjunction
The anticancer strategy of anti-tumour cell proliferative medicine, the treatment for tumor provides new thinking and direction.
Growth and the transfer of malignant tumor depend on tumor vessel, along with neonate tumour blood vessel Mechanism Study deeply and
Medicine with tumor vessel as target spot obtain preferable curative effect when clinical treatment tumour it was confirmed by suppression tumor vessel new
Suppress the theory of tumor growth from birth.
Combretastatin A-4 4 (Combretastatin, CA4) is that the bark from Africa shrub Combretum Caffrum divides
From a kind of cis-stilbene class natural product, belong to the microtubules inhibitor of colchicine sample, it is possible to specifically target
To tumor vascular endothelial cell, destroy endotheliocyte skeleton and cause cellularity to change, thus cause tumor vascular subtracting
Few, cause center of tumor region to necrose.CA4 structural formula is shown below:
But we show in the experiment of early stage, and SN38 is due to the chemical constitution of its uniqueness, it is difficult to directly with amphipathic high score
Sub-material assembles formation nanoparticle altogether.The problem that CA4 then exists quickly release in macromolecular material nanoparticle.
Summary of the invention
It is an object of the invention to provide a kind of anti-angiogenic medicaments and anti-tumour cell proliferative combination therapies
Antitumor drug, it is achieved anti-angiogenic medicaments and the anticancer strategy of anti-tumour cell proliferative combination therapies, improves associating
The curative effect of medication.
Present invention also offers a kind of anti-angiogenic medicaments and the antitumor of anti-tumour cell proliferative combination therapies
Pharmaceutical preparation.
The present invention discloses a kind of anti-angiogenic medicaments and the anti-of anti-tumour cell proliferative combination therapies swells
The preparation method of tumor medicine preparation.
By using advanced nano fabrication technique in the present invention, antitumor drug and the anti-angiogenic drugs of water will be insoluble in
Thing bag is loaded in amphipathy macromolecule material, forms medicament-carried nano system, overcomes simultaneously, and SN38 is difficult to direct and amphipathic height
Molecular material assembles the problem forming nanoparticle altogether.
Asking of anti-angiogenic medicaments use in conjunction anti-tumour cell proliferative medicine existence is prepared for existing nanotechnology
Topic, the present invention will propose the anti-cancer methods of a kind of Combretastatin precursor use in conjunction camptothecins anti-tumor prodrug, and should
Drug combination method is combined with advanced nanotechnology, prepares the nanometer delivery systme of the drug combination of antineoplaston, it is achieved anti-
Angiogenesis drug and the value of anti-tumour cell proliferative Drug combination, improve the curative effect of clinical antineoplastic treatment.
A kind of drug combination antitumor drug, including anti-angiogenic medicaments, anti-tumour cell proliferative medicine and amphipathic
Macromolecular material；Described anti-angiogenic medicaments is the prodrug of Combretastatin A-4 4, and described anti-tumour cell proliferative medicine is
Camptothecin antineoplastic agents.
As preferably, the prodrug structure of described Combretastatin A-4 4 is as follows:
Described R is the alkanoyl of C1～C10.
As further preferably, the prodrug of described Combretastatin A-4 4 is Combretastatin A-4 4 (CA4) and butanoic acid or enanthic acid
Or the prodrug that the acyl chlorides of its correspondence, anhydride esterifying obtain.
As preferably, the prodrug structure of described Combretastatin A-4 4 is as follows:
The prodrug of above-mentioned Combretastatin A-4 4 can be obtained with butanoic acid or enanthic acid esterification by Combretastatin A-4 4 (CA4).
As preferably, described anti-tumour cell proliferative medicine is SN38 and unsaturated fatty acid
Connect the prodrug formed.In course of reaction, catalyst, condensing agent etc. can be added as needed on, such as can use 1-(3-bis-
Methylaminopropyl) hydrochlorate of-3-ethyl carbodiimide is as condensing agent, and DMAP, simultaneously can as catalyst
Utilize in N, N-diisopropylethylamine and hydrochloric acid in the hydrochlorate of 1-(3-dimethylamino-propyl)-3-ethyl carbodiimide.Make
For further preferably, described Combretastatin A-4 4 is 1:(1～1.5 with carboxylic acid, catalyst, the mol ratio of condensing agent): (0.5～
1.5): (1～1.5).
As preferably, described unsaturated fatty acid is selected from docosahexenoic acid, all-trans-retinoic acid, linolenic acid, oleic acid
Or linoleic acid.
As preferably, described amphipathy macromolecule material is selected from polyethylene glycol-polylactic acid (PEG-PLA), polyethylene glycol
Lactic-co-glycolic acid (PEG-PLGA), MPEG-PLA (mPEG-PLA) or methoxy poly (ethylene glycol)-poly-breast
Any one in acid-hydroxyacetic acid (mPEG-PLGA).Wherein, the molecular weight of PEG or mPEG is 2k～8k, the molecular weight of PLA
For 2k～16k, the molecular weight of PLGA is 2k～16k.It is preferably mPEG5k-PLA16k。
Present invention also offers a kind of drug combination anti-tumor medicinal preparation, described antitumor drug is any of the above-described institute
The antitumor drug stated.Described preparation can be common dosage form, can be such as powder or tablet, injection, pill etc..
Present invention also offers the preparation method of a kind of above-mentioned drug combination anti-tumor medicinal preparation, including: by anti-angiogenic
Generate medicine, anti-tumour cell proliferative medicine and amphipathy macromolecule material to be dissolved in organic solvent, be then added in water,
To described anti-tumor medicinal preparation.
Specifically, including:
The first step: camptothecin derivative SN38 (SN38) is carried out molecular modification, coupling insatiable hunger
And fatty acid, selected from docosahexenoic acid (DHA), all-trans-retinoic acid (RA), linolenic acid (LNA), oleic acid or linoleic acid
(LA), drug molecule lipotropy is given.And lipotropy SN38 derivant can discharge in vivo and have the SN38 of active anticancer and divide
Son；Combretastatin A-4 4 (CA4) is carried out molecular modification, coupling satisfied fatty acid (butanoic acid or enanthic acid), slows down medicine from nanoparticle
In rate of release；
Second step: build load first step SN38 prodrug and the nanoparticle of CA4 prodrug simultaneously, this nanoparticle by CA4 prodrug,
SN38 prodrug and amphipathy macromolecule composition.
The invention provides drug combination antitumor drug application in antitumor described in a kind of any of the above-described scheme.
The load anti-angiogenic medicaments of the present invention and anti-tumour cell proliferative combination therapies nanoparticle, in vitro can
Suppression tumor cell and the propagation of vascular endothelial cell, the suppression migration of vascular endothelial cell and invasion and attack, suppression vessel lumen
Formed.The drug combination nanoparticle of the present invention also show the ability that preferably suppression tumor increases in vivo, is further characterized by
In the present invention, anti-angiogenic medicaments and anti-tumour cell proliferative combination therapies nanoparticle have extensively in antineoplaston
Wealthy using value and range of application.
And, cytotoxicity experiment shows that the nanoparticle in the present invention can significantly inhibit tumor cell (HT-29) and people
The propagation of umbilical vein vascular endothelial cells (HUVEC), and have dose-dependence；Scratch damage and segment dislocation experiment also confirm that
In the present invention, the nanoparticle of preparation can suppress migration and the vascularization of HUVEC.Contrast independent medication system, the present invention provides
Drug combination nanoparticle show preferable tumor killing effect in vivo, there is good clinical value and application prospect.
Accompanying drawing explanation
Fig. 1 is nanoparticle Electronic Speculum and grain-size graph in embodiment 9；
Fig. 2 is nanoparticle Electronic Speculum and grain-size graph in embodiment 10；
Fig. 3 is nanoparticle Electronic Speculum and grain-size graph in embodiment 11；
Fig. 4 is the influence curve of variable concentrations nanoparticles on tumor cells HT-29 survival rate in embodiment 8,9,10,11；
Fig. 5 be in embodiment 8,9,10,11 variable concentrations nanoparticle to human umbilical endothelial cell's HUVEC survival rate
Influence curve；
Fig. 6 is the nanoparticle impact on human umbilical endothelial cell's HUVEC mobility in embodiment 9,10,11；
Fig. 7 is the nanoparticle impact on human umbilical endothelial cell's HUVEC invasive ability in embodiment 9,10,11；
In figure, SN38 represents that SN38, CA4 or 1 represent Combretastatin A-4 4, butanoic acid-CA4 or 2 tables
Showing that the conjugate of butanoic acid and CA4, enanthic acid-CA4 or 3 represent the conjugate of enanthic acid and CA4,1/4-NP, 2/4-NP, 3/4-NP are respectively
Represent that CA4, butanoic acid-CA4, enanthic acid-CA4 and LNA-SN38 and amphipathy macromolecule assemble the nanoparticle of formation altogether.
Detailed description of the invention
With detailed description of the invention, the present invention is described in further detail below in conjunction with the accompanying drawings, but the present invention is not limited by it
System.
Embodiment 1
Take butanoic acid (51.7 μ L, 0.57mmol), CA4 (180mg, 0.57mmol), EDC HCl (163mg,
0.855mmol), DMAP (76.6mg, 0.627mmol), DIEA (188.9 μ L, 1.14mmol) are dissolved in 4ml DCM, under room temperature
It is stirred overnight, adds ethyl acetate, utilize 5% citric acid, saturated NaHCO successively3, saturated aqueous common salt washing organic facies, organic
It is dried with anhydrous sodium sulfate, filters, collect removal of solvent under reduced pressure after filtrate；Isolated and purified (the acetic acid of solid column chromatography chromatogram
Ethyl ester: normal hexane=1:3), obtain target product butanoic acid-CA4 conjugate compound 2 (280mg, yield 93%).
Product butanoic acid-CA4's1H NMR nuclear magnetic data and mass spectrometric data are as follows:
1H NMR(400MHz,CDCl3):δ1.00-1.04(t,3H),1.73-1.78(m,2H),2.49-2.53(t,2H),
3.71 (s, 6H), 3.80 (s, 3H), 3.83 (s, 3H), 6.45-6.45 (d, 2H, J=1.2), 6.51 (s, 2H), 6.83-6.85
(d, 1H, J=8.4), 7.00-7.01 (d, 1H, J=1.6), 7.10-7.12 (q, 1H).
HR-ESI Qq-LTMS: value of calculation: [C22H26O6]+[M+H]+=387.1802；Detected value: 387.1811.
Embodiment 2
Take enanthic acid (80.8 μ L, 0.57mmol), CA4 (180mg, 0.57mmol), EDC HCl (163mg,
0.855mmol), DMAP (76.6mg, 0.627mmol), DIEA (188.9 μ L, 1.14mmol) are dissolved in 4mL DCM, under room temperature
It is stirred overnight, adds ethyl acetate, utilize 5% citric acid, saturated NaHCO successively3, saturated aqueous common salt washing organic facies, organic
It is dried with anhydrous sodium sulfate, filters, collect removal of solvent under reduced pressure after filtrate；Isolated and purified (the acetic acid of solid column chromatography chromatogram
Ethyl ester: normal hexane=1:3), obtain target product enanthic acid-CA4 conjugate compound 3 (260mg, yield 85%).
Product enanthic acid-CA4's1H NMR nuclear magnetic data and mass spectrometric data are as follows:
1H NMR(400MHz,CDCl3):δ0.88-0.91(t,3H),1.26-1.41(m,6H),1.70-1.74(t,2H),
2.51-2.54 (t, 2H), 3.71 (s, 6H), 3.80 (s, 3H), 3.84 (s, 3H), 6.45-6.45 (d, 2H, J=1.6), 6.51
(s, 2H), 6.83-6.85 (d, 1H, J=8.4), 7.00-7.00 (d, 1H, J=1.6), 7.10-7.12 (q, 1H).
HR-ESI Qq-LTMS: value of calculation [C25H32O6]+[M+H]+=429.2212；Detected value: 429.2282.
Embodiment 3
LNA-SN38 conjugate compound 4 synthetic method referenced patent document is " before SN38 medicine
Body and its preparation method and application " in the synthesis of SN38 prodrug 7, number of patent application is 201410295432.X.
Embodiment 4
LA-SN38 synthetic method referenced patent document " SN38 prodrug and preparation method thereof
And application " in the synthesis of SN38 prodrug 6, number of patent application is 201410295432.X.
Embodiment 5
OA-SN38 synthetic method referenced patent document " SN38 prodrug and preparation method thereof
And application " in the synthesis of SN38 prodrug 5, number of patent application is 201410295432.X.
Embodiment 6
RA-SN38 synthetic method referenced patent document " a kind of all-trans-retinoic acid-camptothecin cancer therapy drug conjugate and
Its preparation method and application " in the preparation of ATRA-SN38 conjugate (C10 hydroxyl), number of patent application or Patent No.
201410692682.7。
Embodiment 7
DHA-SN38 synthetic method, concrete grammar is as follows:
SN38 (119.3mg, 0.304mmol), DMAP (40.8mg, 0.334mmol), DHA (100mg, 0.304mmol),
EDC HCl (64mg, 3.334mmol) is dissolved in 4mL DMF, adds DIEA (43.1mg, 0.334mmol), and reaction overnight, removes
After reaction dissolvent, solid is dissolved in dichloromethane, washes 7 times, successively with saturated sodium bicarbonate aqueous solution, 5% citric acid and saturated
Saline solution is respectively washed 1 time, and anhydrous sodium sulfate is dried, and filters, and collects removal of solvent under reduced pressure after filtrate；Solid column chromatography chromatogram separates
Purification (ethyl acetate: normal hexane=1:1), obtains product DHA-SN38 conjugate, productivity 57.5%.
DHA-SN38 conjugate nuclear-magnetism and mass spectrometric data are as follows:
1H NMR(400MHz,CDCl3): δ 0.88-0.91 (t, 3H), 0.91-0.99 (t, 3H), 1.31-1.35 (t, 3H),
1.77-1.88(m,2H),1.96-2.02(m,2H),2.50-2.55(m,2H),2.65-2.85(m,12H),3.06-3.12(m,
2H), 3.75 (br, 1H), 5.20-5.35 (m, 13H), 5.43-5.46 (m, 2H), 5.66-5.70 (d, 1H, J=16.4),
7.47-7.50 (m, 1H), 7.61 (s, 1H), 7.75-7.76 (d, 1H, J=2), 8.17-8.20 (d, 1H, J=8.8).
HR-ESI Qq-LTMS: value of calculation [C44H50N2O6]+[M+H]+=703.3742；Detected value: 703.3741.
Embodiment 8
The LNA-of butanoic acid-CA4 conjugate compound 2 (CA4 content 0.5mg) prepared by embodiment 1, embodiment 3 preparation
SN38 (SN38 content 0.5mg) is respectively and mPEG5k-PLA16k(20 times of drug quality) are dissolved in 1mL acetone, are uniformly dissolved
After be added drop-wise in 10mL water, after dropping, decompression remove acetone, i.e. can get butanoic acid-CA4 and LNA-SN38 Nano medication
(being respectively labeled as 2-NP, 4-NP).
Embodiment 9
By LNA-SN38 compound 4 (1.67mg) and the mPEG of preparation in CA4 compound 1 (0.5mg), embodiment 35k-
PLA16k(43.4mg) it is dissolved in 3mL acetone, is added drop-wise to after being uniformly dissolved in 10mL water, after dropping, removal third of reducing pressure
Ketone, i.e. can obtain loading anti-angiogenic medicaments and the Nano medication (being designated as 1/4-NP) of anti-tumour cell proliferative medicine.Electronic Speculum
Lower nanometer appearance is as it is shown in figure 1, DLS records size 31 ± 4nm.
Embodiment 10
The LNA-SN38ization of preparation in butanoic acid-CA4 compound 2 (0.6mg) that purification in embodiment 1 is obtained, embodiment 3
Compound 4 (1.67mg) and mPEG5k-PLA16(45.4mg) it is dissolved in 3mL acetone, is added drop-wise to after being uniformly dissolved in 10mL water, drip
After adding, acetone is removed in decompression, i.e. can obtain loading anti-angiogenic medicaments and the nanometer medicine of anti-tumour cell proliferative medicine
Thing (is designated as 2/4-NP).Nanometer appearance under Electronic Speculum is as in figure 2 it is shown, DLS records size 38 ± 17nm.
Embodiment 11
The LNA-SN38 of preparation in enanthic acid-CA4 compound 3 (0.63mg) that purification in embodiment 2 is obtained, embodiment 3
Compound 4 (1.67mg) and mPEG5k-PLA8k(46mg) it is dissolved in 3mL acetone, is added drop-wise to after being uniformly dissolved in 10mL water, drip
After adding, acetone is removed in decompression, i.e. can obtain loading anti-angiogenic medicaments and the nanometer medicine of anti-tumour cell proliferative medicine
Thing (is designated as 3/4-NP).Nanometer appearance under Electronic Speculum is as it is shown on figure 3, DLS records size 38 ± 14nm.
It is further elucidated with the nanoparticle of the present invention therapeutic effect to tumor below by way of test case:
Test case 1 vitro cytotoxicity is tested
Take the logarithm trophophase cell (HT-29, HUVEC), after trypsinization, plant plate (96 orifice plates, 5 × 103Individual cell/
Hole), hatch 24h for 37 DEG C and make cell attachment, with SN38 as matched group, add in the embodiment 8,9,10,11 of variable concentrations and prepare
Nanoparticle (100 μ L/ hole), cultivate after 48h, every hole adds 30 μ L tetrazolium bromide (MTT, 5mg/mL), removes after continuing to cultivate 4h
Supernatant, every hole adds 100 μ L dimethyl sulfoxide (DMSO), vibration, makes crystallized product fully dissolve, and 492nm is in microplate reader
Detect each hole absorbance, draw out cell survival curve, and calculate each group of nanoparticle 503nhibiting concentration (IC to cell50) value.
The in vitro toxicity result of various tumor cells is shown in Fig. 4 and Fig. 5 and Biao 1 by antitumor medicinal liposome.
Table 1: the IC of nanoparticles on tumor cells in embodiment 8,9,10,1150(μM)
Can be seen that from Fig. 4 and Fig. 5 and Biao 1, the nanoparticle 4-NP being made up of SN38 prodrug has similar to SN38 freely
Cytotoxicity, but with CA4 prodrug 1,2,3 constitute combination nanoparticle 1/4-NP, 2/4-NP, 3/4-NP cytotoxicity substantially increase
By force, it was demonstrated that effectiveness associated with two medicines.
Test case 2: migrate experiment
Static HUVEC cell is incubated in 6 orifice plates, with rifle head, cell does in centre position, hole straight drawing
Trace, removes culture medium, add free CA4 medicine group and embodiment 9,10,11 are prepared respectively 1/4-NP, 2/4-NP, 3/
4-NP (in terms of CA4, its concentration is 5nM), does matched group with blank non-dosing group.Each group cell is placed in incubator cultivation 24h
After, take pictures, and the width granularity of cut is processed.Result shows (Fig. 6), compared with blank group, free CA4 medicine group with
And the transfer ability of the nanoparticle of preparation all energy significance suppression HUVEC cells in embodiment 9,10,11.
Test case 3: Matrigel
Take the logarithm trophophase HUVEC cell, by 4 × 104The density of individual cells/well is seeded to be covered with 30 μ L Matrigel
The upper room of Transwell cell, lower room adds 700 μ L culture medium, uniformly rocks and makes cell distribution uniform, cell is placed in 37
DEG C, 5%CO2Incubator is cultivated, 24h, after cell attachment, carry out next step operation.Room culture medium in taking-up, adds 200 μ L and contains
There is medicine (nanoparticle prepared in free CA4 medicine group and embodiment 9,10,11, drug level calculates) with CA4 concentration 5nM
Culture medium, lower room adds 700 μ L containing the culture medium of 20% hyclone, after cultivating 24h, abandons room culture medium, PBS two
Secondary, methanol is fixed, violet staining, wipes upper room glue and cell with cotton swab, and basis of microscopic observation penetrates the cell of room, claps
According to, counting.
Result is as it is shown in fig. 7, the invasion and attack of each dosing group HUVEC cell are significantly suppressed, and blank group is invaded cell number and is
834, and in free CA4 medicine group and embodiment 9,10,11, in the nanoparticle group of preparation, HUVEC cell invasion number is respectively
252,330,325,308, each group all has significant differences (p < 0.01) with blank group.
Test case 4: vessel lumen Forming ability is tested
Matrigel adds in 96 orifice plates, 30 μ L/ holes, 37 DEG C of solidification 1h, adds 100 μ L containing 2 × 104The training of individual HUVEC
Support base, add 1/4-NP, 2/4-NP, the 3/4-NP prepared respectively in free CA4 medicine group and embodiment 9,10,11 (with CA4
Meter, its concentration is 5nM), with DMSO for blank group.After cultivating 4h, basis of microscopic observation plastidogenetic tube chamber pattern, point
The analysis medicine impact on segment dislocation.It is circular that blank group cell forms obvious tube chamber, and dosing group CA4 and 1/4-NP, 2/4-
NP, 3/4-NP cell is at random, less formation tube chamber pattern.Confirm that in the present invention, the nanoparticle of preparation can suppress HUVEC to be formed
The ability of vessel lumen.
Test case 5: anti-tumor in vivo effect assessment
5 week old Balb/c mouse bare subcutaneous injections contain 5 × 106HT-29 cell suspension 200 μ L, treats that tumor body length is to 70mm3Size
After, it is divided into 4 groups (often groups 7): the 2-NP (CA4 equivalent dose is 15mg/kg) and 4-NP of preparation in normal saline, embodiment 8
The 2/4-NP (CA4 and SN38 equivalent dose is 15mg/kg) of (SN38 equivalent dose is 15mg/kg), embodiment 10 preparation.Often
Every three days one time tail intravenously administrable, 3 times altogether.Being 0 day to be administered for the first time, separated in time measures tumor volume, and right
Result carries out statistical analysis.Result shows, 4-NP, 2/4-NP have highly significant relative to normal saline group to suppression tumor growth
Effect, and anti-angiogenic medicaments and anti-tumour cell proliferative medication combined group (2/4-NP) be better than by the 2-NP of independent medication
With 4-NP group.In vivo test is further characterized by, the anti-angiogenic medicaments prepared in the present invention and anti-tumour cell proliferative medicine
Drug combination nanoparticle has the ability that stronger suppression tumor increases.
Claims (10)
1. a drug combination antitumor drug, it is characterised in that include anti-angiogenic medicaments, anti-tumour cell proliferative medicine
With amphipathy macromolecule material；Described anti-angiogenic medicaments is the prodrug of Combretastatin A-4 4, and described antitumor cell increases
Growing medicine is camptothecin antineoplastic agents.
Drug combination antitumor drug the most according to claim 1, it is characterised in that before the medicine of described Combretastatin A-4 4
Body structure is as follows:
Described R is the alkanoyl of C1～C10.
Drug combination antitumor drug the most according to claim 1, it is characterised in that before the medicine of described Combretastatin A-4 4
The prodrug that body is Combretastatin A-4 4 to be obtained with butanoic acid or enanthic acid or its corresponding acyl chlorides, anhydride esterifying.
Drug combination antitumor drug the most according to claim 1, it is characterised in that before the medicine of described Combretastatin A-4 4
Body structure is as follows:
Drug combination antitumor drug the most according to claim 1, it is characterised in that described anti-tumour cell proliferative medicine
Thing is that SN38 is connected the prodrug formed with unsaturated fatty acid.
Drug combination antitumor drug the most according to claim 5, it is characterised in that described unsaturated fatty acid is selected from two
Dodecahexaene acid, all-trans-retinoic acid, linolenic acid, oleic acid or linoleic acid.
Drug combination antitumor drug the most according to claim 1, it is characterised in that described amphipathy macromolecule material selects
Any one in PEG-PLA, PEG-PLGA, mPEG-PLA and mPEG-PLGA.
8. a drug combination anti-tumor medicinal preparation, it is characterised in that described antitumor drug is that claim 1～7 is arbitrary
Antitumor drug described in Xiang.
9. the preparation method of the drug combination anti-tumor medicinal preparation described in a claim 8, it is characterised in that including: will
Anti-angiogenic medicaments, anti-tumour cell proliferative medicine and amphipathy macromolecule material are dissolved in organic solvent, are then added to
In water, obtain described anti-tumor medicinal preparation.
10. the application in antitumor of the drug combination antitumor drug described in an any one of claim 1～7.
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| CN109675039A (en) * | 2018-12-21 | 2019-04-26 | 中国科学院长春应用化学研究所 | Pharmaceutical composition, anti-tumor drug and application |
| CN109771657A (en) * | 2019-03-27 | 2019-05-21 | 湘潭大学 | A kind of bonding vascular disrupting agents and the high molecular antineoplastic drug of immunomodulator and preparation method thereof |
| CN111253411A (en) * | 2020-03-26 | 2020-06-09 | 山东省科学院生物研究所 | Berberine linoleic acid conjugate and preparation method and application thereof |
| CN111362837A (en) * | 2020-03-23 | 2020-07-03 | 新乡医学院 | NQO1 activated Combretastatin A4 prodrug and synthesis method and application thereof |
| CN114848644A (en) * | 2022-04-20 | 2022-08-05 | 深圳市龙华区人民医院 | Nano-targeting sustained-release drug, and preparation method, device and application thereof |
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| CN114848644A (en) * | 2022-04-20 | 2022-08-05 | 深圳市龙华区人民医院 | Nano-targeting sustained-release drug, and preparation method, device and application thereof |
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Application publication date: 20161221 |