CN101190334B - Thermal targeting nano ball anti-cancer medicine carrier and preparation method thereof - Google Patents
Thermal targeting nano ball anti-cancer medicine carrier and preparation method thereof Download PDFInfo
- Publication number
- CN101190334B CN101190334B CN2006101442550A CN200610144255A CN101190334B CN 101190334 B CN101190334 B CN 101190334B CN 2006101442550 A CN2006101442550 A CN 2006101442550A CN 200610144255 A CN200610144255 A CN 200610144255A CN 101190334 B CN101190334 B CN 101190334B
- Authority
- CN
- China
- Prior art keywords
- nanosphere
- water
- oil
- poly
- pharmaceutical carrier
- 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.)
- Expired - Fee Related
Links
Images
Landscapes
- Medicinal Preparation (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
The invention relates to an anticarcinogen nanometer-sphere carrier targeted for heat treatment and the preparation method thereof. The anticarcinogen nanometer-sphere carrier of the invention is madeof biodegradable material and used for enveloping anticarcinogen and achieving effects of sustained release and controlled release; thermally sensitive macromolecule with low critical solution temper ature of 37 to 43 DEG C is covalence cross-linked on the surface of the nanometer sphere, and the grafting of the thermally sensitive macromolecule leads the nanometer sphere to have the function of thermal targeting to cancerous cells. Biodegradable anticarcinogen nanometer-sphere carrier with thermally sensitive macromolecule grafted on the surface is the anticarcinogen nanometer-sphere carrier targeted for heat treatment of the invention, which can circulate inside the human body for a long time without being cleaned, speed up the release of chemotherapy medicine in cancerous tissues duringheat treatment, resolve the defect that existing chemotherapy medicine has low distribution in cancerous tissues, and reduce injury of normal cells of human body caused by chemotherapy medicines.
Description
Technical field
The present invention relates to a kind of hot targeted anticancer medicine carrier and preparation method thereof, more particularly, but the present invention relates to a kind of targeting in thermosensitive type nano ball anti-cancer medicine carrier of cancerous tissue cell and preparation method thereof.
Technical background
Cancer (being malignant tumor) is the commonly encountered diseases of a class serious harm human life Health and Living quality, at present, in the treatment for cancer method, chemotherapy is as a kind of systemic treatment means, having all has lot of advantages such as therapeutical effect to primary tumor, metastasis and subclinical metastasis, the subject matter of its existence does not lie in the kill and wound weak strength of existing cancer therapy drug to cancerous cell, but normal structure is had lethal equally, and lower at tumor tissues punishment cloth.In addition, thermotherapy also is a kind of treatment method for cancer, promptly makes tumor tissues reach 40～43 ℃ by heating, causes tumor cell tissue's growth retardation and death.At present, thermotherapy Chang Zuowei householder method can obtain better therapeutic effect with the chemotherapy combined application.Yet, in the use in conjunction of thermotherapy and chemotherapy, chemotherapeutics is still very serious to the toxic and side effects of human body, and the cancerous tissue that will make chemotherapeutics arrive thermotherapy fully is very difficult, if reduction dosage, can alleviate toxic and side effects, but can reduce again the tumor treatment effect simultaneously.
If we can make the sustained-release and controlled release dosage form with anticancer chemicals, just do not need frequent drug administration can keep drug disposition valid density in a long time yet, thereby just can reach the reduction toxic and side effects; If we can allow anticancer chemotherapeutic agent have targeting, only act on cancerous cell and do not act on normal cell, just adopt targeting drug delivery system, this just can improve curative effect and can reduce the whole body toxic and side effects, makes the patient immune function exempt from destruction.Therefore, the pharmaceutical carrier with slow release, controlled release and target function can be applicable to the chemotherapy of cancer, improves the selectivity of cancer therapy drug, reduces its toxic and side effects.
At present, in order to reduce the toxic and side effects of cancer chemotherapeutic, has slow release, the nanosphere pharmaceutical carrier of controlled release and target function has obtained extensive studies, such as, grafting the albumin nanometer ball of folic acid [International Journal ofPharmaceutics 287 (2004) 155], grafting nanoparticle [the Acta Pharmaceutica Sinica of monoclonal antibody, 30 (9) (1995) 706], magnetic albumin nanoparticle [Hepatobiliary Pancreat Dis Int., 3 (3) (2004) 365] and thermosensitive type polymer micelle [J.Control.Release 65 (2000) 93, Colloids and Surface B:Biointerfaces 16 (1999) 195-205].Wherein, the thermosensitive type polymer micelle can be in chemotherapy, combined with hyperthermia be used target cancer cell initiatively, but have shortcomings such as bad mechanical strength, stability be bad by the thermosensitive type polymer micelle that hydrophobic interaction forms, and, when the thermosensitive type polymer micelle after assemble at tumor tissue cell place, intensive shell hydrophilic polymer segment can shrink, be deposited in the surface of nano-micelle thick and fast, slow down thereby make cancer therapy drug discharge, influence chemotherapeutics the tumor treatment effect.
Summary of the invention
The objective of the invention is to overcome above-mentioned defective, a kind of thermal targeting nano ball anti-cancer medicine carrier is provided, but its bio-compatible and biodegradation have slow release, controlled-release function again, the energy targeting is in the cancerous tissue cell.
Above-mentioned thermal targeting nano ball anti-cancer medicine carrier of the present invention is made of two parts, promptly biodegradable nanosphere and the thermosensitive type macromolecule with lower critical solution temperature (LCST).Wherein, the material of described nanosphere is selected from following biodegradable material: protein (as bovine serum albumin and human serum albumin), polylysine, poly arginine, polyglutamic acid, poly-aspartic-acid, polypeptide, polylactic acid (PLA), polyglycolic acid (PGA), poly-(lactic acid-ethanol) (PLGA), in the derivant of chitosan, polylactic acid-poly-ethanol (PLEA), poly-(lactic acid-ethanol)-poly-ethanol, polycaprolactone, poe, poly-anhydride, poly-isobutyl group propylene hydrocyanic ester, polyphosphazene and above-mentioned each material one or more; Described thermosensitive type macromolecule is selected from the derivant of PNIPAM, polymethyl vinyl ether [poly (vinyl methyl ether)], poly-phosphorus nitrence (polyphosphazene derivatives), poly N-ethylene caprolactam [poly (N-vinylcaprolactam)], poly-siloxy ethylene glycol [poly (siloxyethylene glycol)] and above-mentioned each material one or more (as PNIPAM-acrylamide and PNIPAM-N, the N-DMAA), its LCST is 37～43 ℃.
Biodegradable nanosphere can be made carrier embedding cancer therapy drug and realize slow release, controlled-release function, and in order to prevent the generation of opsonic action in the body, the particle diameter of nanosphere is preferably less than 200nm; At a kind of LCST of surperficial covalent cross-linking of nanosphere is that 37～43 ℃ the high molecular purpose of thermosensitive type is in order to make this pharmaceutical carrier have the effect of targeting cancerous tissue cell.
The targeting principle of above-mentioned thermal targeting nano ball anti-cancer medicine carrier of the present invention is described below, if this thermal targeting nano ball anti-cancer medicine carrier of intravenous injection that (the tumor tissues position is heated to 43 ℃) during thermotherapy, because the particle diameter of thermal targeting nano ball anti-cancer medicine carrier is less than 200nm, and its surface is hydrophilic when temperature is lower than its LCST, so this thermal targeting nano ball anti-cancer medicine carrier can circulate in vivo, be not eliminated, yet, when it is recycled to when reaching the cancerous tissue place with body fluid, because the temperature at cancerous tissue place is higher than its LCST during thermotherapy, its surperficial macromolecule will experience and be transformed into hydrophobic macromolecule mutually, thereby cause this thermal targeting nano ball anti-cancer medicine carrier adsorption precipitation on cancerous cell, so just can realize the active targeting of cancer therapy drug.
The more important thing is, behind the cancerous tissue cell when thermal targeting nano ball anti-cancer medicine carrier active targeting thermotherapy, because when the temperature at cancerous tissue place is higher than the high molecular LCST of the thermosensitive type on nanosphere surface during thermotherapy, the thermosensitive type macromolecule can shrink, make the gap between the thermosensitive type macromolecular chain increase, enzyme is easier to be contacted with nanosphere, thereby causes the release of cancer therapy drug to be accelerated, and this just can overcome the existing shortcoming of micelle in the past.
The present invention also aims to provide a kind of thermosensitive type nanosphere preparation of drug carriers method, promptly make the nanosphere pharmaceutical carrier with Biodegradable material earlier, again a kind of thermosensitive type macromolecule of the surperficial covalent cross-linking of nanosphere pharmaceutical carrier (a kind of functional group of thermosensitive type macromolecule end and the functional group on nanosphere pharmaceutical carrier surface are reacted).For example, when the material of nanosphere contains carboxyl (as polyglutamic acid, poly-aspartic-acid and contain glutamic acid and the polypeptide of aspartic acid and protein etc.), can adopt segment copolymerization method to make the high molecular end of thermosensitive type have several allylamines, thereby make the high molecular end of thermosensitive type have several amino, in the phosphate buffer (100mM) of pH4.0, activate the carboxyl on nanosphere surface then with EDAC and NHS, drip the thermosensitive type macromolecular solution that has terminal amino group again, make the amino covalence reaction of activatory carboxyl and thermosensitive type macromolecule end form stable amido link; When the material of nanosphere contains amino (as polylysine, poly arginine and contain lysine and arginic polypeptide etc.), can adopt segment copolymerization method to make the high molecular end of thermosensitive type have several acrylic acid, thereby make the high molecular end of thermosensitive type have several carboxyls, take then to make it to graft on the nanosphere surface behind the carboxyl of above method activation thermosensitive type macromolecule end; For neither containing carboxyl, not containing amino Biodegradable material (as polylactic acid-based, polyglycolic acid class and polylactic acid-glycollic acid class etc.) again, can adopt copolymerization method to make it to have some amino or carboxyl, be made into nanosphere and adopt above same procedure that the thermosensitive type macromolecule is grafted on the nanosphere surface afterwards.
The nanosphere that the present invention relates to can adopt ultrasonic emulsification-chemical crosslinking solidification method to make, and promptly adopts supersound method to make the nanometer emulsion droplet earlier, again the nanometer emulsion droplet is carried out chemical crosslinking and solidifies, and removes chemical cross-linking agent and emulsifying agent at last and can obtain nanosphere.More particularly, the making of described nanosphere pharmaceutical carrier can be taked following three kinds of methods:
1) Water-In-Oil method: with having dissolved the aqueous solution of hydrophilic anticarcinogen and hydrophilic film material as water, with the organic solvent that has dissolved oil soluble emulsifying agent as oil phase, with water mix with oil phase stir carry out rough segmentation and loose after, reuse ultrasonic cell-break machine carries out emulsifying, obtain water-in-oil type nanoemulsion liquid, under magnetic agitation, in the gained nanoemulsions, add cross-linking agent again and carry out crosslinking curing, remove excessive cross-linking agent and emulsifying agent and can obtain the nanosphere pharmaceutical carrier.
2) oil-in-water method: with having dissolved the organic solvent of hydrophobicity anticarcinogen and hydrophobic film material as oil phase, with the aqueous solution that has dissolved water soluble emulsifier as water, after oil phase stirring mixed with water carried out rough segmentation and loose, reuse ultrasonic cell-break machine carries out emulsifying, obtain the oil-in-water type nanoemulsions, under magnetic agitation, in the gained nanoemulsions, add cross-linking agent again and carry out crosslinking curing, remove excessive cross-linking agent and emulsifying agent and can obtain the nanosphere pharmaceutical carrier.
3) W/O/W multi-emulsion method: with having dissolved the aqueous solution of hydrophilic anticarcinogen as water, with the organic solvent that has dissolved hydrophobic film material and oil soluble emulsifying agent as oil phase, with water mix with oil phase stir carry out rough segmentation and loose after, reuse ultrasonic cell-break machine carries out emulsifying, obtain water-in-oil type nanoemulsion liquid, again gained water-in-oil type nanoemulsion liquid is added the aqueous phase that is dissolved with water soluble emulsifier and carry out ultrasonic emulsification, thereby obtain W/O/W type nanoemulsions, under magnetic agitation, in gained W/O/W type nanoemulsions, add cross-linking agent again and carry out crosslinking curing, remove excessive cross-linking agent and emulsifying agent and can obtain the nanosphere pharmaceutical carrier.
The thermosensitive type macromolecule that the present invention relates to is selected from one or more (as PNIPAM-acrylamide and the PNIPAM-N,N-DMAAs) in the derivant of PNIPAM, polymethyl vinyl ether [poly (vinylmethyl ether)], poly-phosphorus nitrence (polyphosphazene derivatives), poly N-ethylene caprolactam [poly (N-vinylcaprolactam)], poly-siloxy ethylene glycol [poly (siloxyethylene glycol)] and above-mentioned each material.In order to introduce functional group amino or carboxyl at the high molecular end of thermosensitive type; can be at above high molecular one section allylamine of an end block copolymerization (AA) or acrylic acid (AAc); with PNIPAM-nitrile-acrylamide-acrylic acid amine is example; can adopt radical polymerization synthetic; promptly use N-N-isopropylacrylamide (NIPAM) and third rare amide (AAm) to make comonomer; the different dibutyronitrile of azo (AIBN) is made initiator; ethanol is made solvent; under nitrogen protection, carry out radical polymerization 24h 60 ℃ the time; drip allylamine (AA) polyase 13 .0h afterwards again, at last synthetic sample is splashed into thermosensitive type macromolecule Poly (NIPAM-co-AAm-b-AA) precipitation is separated out.
Advantage of the present invention and effect are to prepare a kind of thermal targeting nano ball anti-cancer medicine carrier that reduces chemical therapy toxic side effect.The present invention adopts nanosphere to realize slow release as pharmaceutical carrier, controlled-release effect, by realizing hot targeting effect at its surface graft thermosensitive type macromolecule, because the nanosphere pharmaceutical carrier has passed through chemical crosslinking curing, therefore, have than thermosensitive type polymer micelle mechanical strength height, the advantage of good stability, and after assemble at the tumor tissues place when thermotherapy, the hydrophilic macromolecule on thermal targeting nano ball anti-cancer medicine carrier surface can shrink, make the gap between the thermosensitive type macromolecular chain increase, enzyme is easier to be contacted with nanosphere, thereby causes the release of cancer therapy drug to accelerate.Chemotherapeutics was in the not high shortcoming of cancerous tissue punishment cloth when the thermal targeting nano ball anti-cancer medicine carrier that the present invention prepares had not only solved chemotherapy combined with hyperthermia application of treatment cancer, reduce chemotherapeutics to normal histiocytic killing and wounding, also overcome shortcomings such as thermosensitive type polymer micelle mechanical strength is not high, stability drug release bad and the cancerous tissue place when thermotherapy slows down.
Description of drawings
Chemical crosslinking curing, the schiff bases of Fig. 1 hydrophilic albumin nanometer ball should be reduced and be reacted with the grafting of thermosensitive type macromolecule Poly (NIPAM-co-AAm-b-AA).
The synthetic reaction of Fig. 2 thermosensitive type macromolecule Poly (NIPAM-co-AAm-b-AA).
Fig. 3 copolymer p oly (NIPAM-co-AAm-b-AA)-1 (hollow triangle), Poly (NIPAM-co-AAm-b-AA)-2 (open circles), Poly (NIPAM-co-AAm-b-AAc)-1 (black triangle) and Poly (NIPAM-co-AAm-b-AAc)-2 (filled circles) aqueous solution (c=5.00 * 10
-3G cm
-3) the temperature dependency of printing opacity value.
The stereoscan photograph of Fig. 4 albumin nanometer ball.
The hydraulic radius of PAN1, PAN2, P-PLN1 and P-PLN 2 aqueous solutions during Fig. 5 different temperatures.
Fig. 6 is embedded with the release in vitro behavior of the various pharmaceutical carriers of ADR.
The specific embodiment
Further describe the present invention below by specific embodiment, but do not limit content of the present invention.
1, thermosensitive type is high molecular synthetic
The thermosensitive type macromolecule that the present invention relates to can be a PNIPAM, polymethyl vinyl ether [poly (vinylmethyl ether)], poly-phosphorus nitrence (polyphosphazene derivatives), poly N-ethylene caprolactam [poly (N-vinylcaprolactam)], in the derivant of poly-siloxy ethylene glycol [poly (siloxyethylene glycol)] and above-mentioned each material one or more are (as PNIPAM-acrylamide and PNIPAM-N, the N-DMAA), with copolymer PNIPAM-nitrile-acrylamide-acrylic acid amine [Poly (NIPAM-co-AAm-b-AA)] of having several terminal amino groups is example, can adopt two-step method to synthesize (Fig. 2), it is the radical polymerization of initiator that synthetic method adopts with AIBN.At first, with 39.05gNIPAM, 2.95gAAm, 0.20gAIBN and 250cm
3It is 60 ℃ oil bath that the three neck round-bottomed flasks that stirrer is arranged in ethanol adds, this flask are put into temperature, stirs 24h (Fig. 2 .a) under the condition of logical nitrogen protection; Secondly, in reaction mixture, drip 2cm behind the 24h
3AA continues to stir 3.0h (Fig. 2 .b) afterwards under the same conditions.At last resulting polymers is splashed into ether make it the precipitation separate out, to remove unconverted monomer, initiator and oligomer, vacuum drying gets Poly (NIPAM-co-AAm-b-AA)-1 then.
2, nanosphere preparation of drug carriers
The material of the nanosphere that the present invention relates to can be one or more in the derivant of protein, polylysine, poly arginine, polyglutamic acid, poly-aspartic-acid, polypeptide, polylactic acid, polyglycolic acid, poly-(lactic acid-ethanol), chitosan, polylactic acid-poly-ethanol, poly-(lactic acid-ethanol)-poly-ethanol, polycaprolactone, poe, poly-anhydride, poly-isobutyl group propylene hydrocyanic ester, polyphosphazene and above-mentioned each material; The making of nanosphere pharmaceutical carrier can be taked the Water-In-Oil method, three kinds of methods such as oil-in-water method and W/O/W multi-emulsion method.Is example as material, the Water-In-Oil method of nanosphere as the manufacture method of nanosphere with the human serum albumin, is about to 0.6cm
3The 20wt.%HSA normal saline solution adds 30cm
3Contain in the cyclohexane extraction of 6.0wt.%Arlacel83, stir earlier 10min, ultrasonic emulsification 10min again, the output of ultraemulsifier (TOMY UD-200) is 165W.Add 3.0mL 0.34mol/L glutaraldehyde toluene solution (0.85mg/mg HSA) and come crosslinking curing nanometer emulsion droplet, continue during curing to stir 24h, add 0.50cm afterwards again
3The aldehyde radical that it is excessive that ethanolamine neutralizes (Fig. 1 .a), dialogue protein nano ball suspension carries out super centrifugal (20000 * g behind the reaction 1.0h, 20min), centrifugal back gained nanosphere is scattered in the 20mL cyclohexane extraction, and in magnetic agitation, dropwise add about 3.0mL acetone, separate out from cyclohexane extraction until nanosphere, superly remove supernatant after centrifugal, contain 0.2M NaBH being dissolved in behind the about 1.0h of nanosphere sample vacuum drying
420mL normal saline (schiff bases should be reduced), stir and superly behind the 1.0h can obtain hydrophilic albumin nanometer ball (albuminnanospheres) after centrifugal, can be designated as AN.
3, the high molecular grafting of thermosensitive type
Remove supernatant behind the nanosphere after the centrifugal above-mentioned normal saline washing, make it to be scattered in 10mL phosphate buffer (pH4.0,100mM), weighing 0.25g EDAC and 0.30g NHS make it to be dissolved in the 10mL phosphate buffer (ice bath), and then the EDAC/NHS mixed liquor is added dropwise in the albumin nanometer ball suspension, mixed solution continues to stir 2.0h under room temperature; 2.0h the super centrifugal supernatant of removing in back can be removed unreacted EDAC and NHS, and nanosphere is scattered in the 10mL phosphate buffer again, adds Poly (NIPAM-co-AAm-b-AA)-1 solution of 5.0mL 0.10g/mL, reacts 3.0h (Fig. 1 .b) under the room temperature; Superly the nanosphere sample is carried out lyophilization, ℃ preservation then-20 after centrifugal.Grafting has the albumin nanometer ball of Poly (NIPAM-co-AAm-b-AA)-1 can be designated as PAN1.
Change the 20wt.%HSA normal saline solution into be dissolved with 2.5 μ gADR/mg HSA 20wt.%HSA normal saline solution, can make ADR-AN and ADR-PAN1 with above-mentioned identical experimental technique, ADR is an anticancer drugs, doxorubicin.
Embodiment 2
With among the embodiment 1 during the thermosensitive type Polymer Synthesizing consumption of AIBN change 0.10g into, other synthesis conditions are all constant, synthetic product be Poly (NIPAM-co-AAm-b-AA)-2.Adopt above-mentioned with quadrat method with Poly (NIPAM-co-AAm-b-AA)-2 grafting to blank albumin nanometer ball (AN) be embedded with on the albumin nanometer ball ADR-AN surface of anticarcinogen amycin ADR, obtain sample P AN2 and ADR-PAN2 respectively.
Embodiment 3
Change the material of nanosphere among the embodiment 1 into polylysine (MW:3-7 ten thousand), other conditions are identical, make polylysine nanosphere (Polylysine nanospheres), can be designated as PLN, at last gained PLN is scattered in the 10mL phosphate buffer (pH4.0,100mM) in.With in embodiment 1 and 2 with the 2cm of N-isopropylacrylamide and acrylamide copolymerization
3Allylamine all changes 1.45g acrylic acid (AAc) into, synthesizes Poly (NIPAM-co-AAm-b-AAc)-1 and Poly (NIPAM-co-AAm-b-AAc)-2.Weighing 0.25g EDAC and 0.30g NHS make it to be dissolved in the 10mL phosphate buffer (ice bath), prepare two parts, and then two portions of EDAC/NHS mixed liquors are added dropwise to respectively in Poly (NIPAM-co-AAm-b-AAc)-2 solution of Poly (NIPAM-co-AAm-b-AAc)-1 solution of 10.0mL 0.10g/mL and 10.0mL 0.10g/mL, mixed solution continues to stir 2.0h under room temperature, again two kinds of mixed liquors are splashed in the ether respectively and precipitate, remove supernatant, vacuum drying 24h again can obtain Poly (NIPAM-co-AAm-b-AAc)-1 and Poly (NIPAM-co-AAm-b-AAc)-2 that terminal carboxyl group has activated; The Poly (NIPAM-co-AAm-b-AAc)-1 and the Poly (NIPAM-co-AAm-b-AAc)-2 that have activated of weighing 0.5g terminal carboxyl group respectively makes it to be dissolved in respectively the phosphate buffer of 10mL PLN, reacts 3.0h under the room temperature, continues magnetic agitation in the course of reaction; Superly the nanosphere sample is carried out lyophilization, ℃ preservation then-20 after centrifugal.Grafting has the polylysine nanosphere of Poly (NIPAM-co-AAm-b-AAc)-1 and Poly (NIPAM-co-AAm-b-AAc)-2 can be designated as P-PLN1 and P-PLN2 respectively.
The measurement of thermosensitive type macromolecule temperature-sensitive behavior
Poly (NIPAM-co-AAm-b-AA)-1, Poly (NIPAM-co-AAm-b-AA)-2, Poly (NIPAM-co-AAm-b-AAc)-1 and Poly (NIPAM-co-AAm-b-AAc)-2 are made into 5.00 * 10 respectively
-3G cm
-3Solution, with the printing opacity value T of each solution of spectrophotometer measurement and the relation and the LCST value of temperature, temperature when getting printing opacity value T=50% is LCST, the results are shown in Figure 3, the LCST of Poly (NIPAM-co-AAm-b-AA)-1, Poly (NIPAM-co-AAm-b-AA)-2, Poly (NIPAM-co-AAm-b-AAc)-1 and Poly (NIPAM-co-AAm-b-AAc)-2 is 42.6 ℃, 42.4 ℃, 41.5 ℃ and 41.1 ℃.
The configuration of surface of albumin nanometer ball is observed
Testing sample is bonded on the sample platform by two-sided conducting resinl, with the gold-plated device of decompression (JFC-1600 fine coater, JEOL, Japan) when pressure is lower than 5Pa, sample is carried out gold-plated (120A, 20s), reuse scanning electron microscope (JSM-6700F, JEOL, Japan) configuration of surface of observation albumin nanometer ball, the result as shown in Figure 4.
The hydraulics character and the vitro drug release thereof of thermal targeting nano ball anti-cancer medicine carrier
(1) the hydraulics character of thermal targeting nano ball anti-cancer medicine carrier
PAN1, PAN2, P-PLN1 and the P-PLN2 hydraulic radius R in pure water when measuring different temperatures with DLS
hSample solution all to balance 10min under the fixed temperature, is measured post-heating and is measured to next temperature next time before each the measurement.Hydraulic radius R
hBe to estimate, the results are shown in Figure 5 by the diffusion coefficient that the utilization of Stokes-Einstein equation records.
The result shows, PAN1, PAN2, P-PLN1 and the P-PLN2 hydraulic radius R in pure water
hCan reduce along with the rising of temperature, this is because the thermosensitive type macromolecule on albumin nanometer ball surface is changed for contraction state by extended configuration along with the rising of temperature.By this experiment is that provable this thermosensitive type protein nano ball pharmaceutical carrier has temperature-sensitive effect preferably, and because the contraction of macromolecular chain, the gap between the macromolecular chain can increase.
(2) measurement of the making of standard curve and embedding rate: with the light absorption value (ABS value) of anticancer drugs, doxorubicin (ADR) solution (all containing 4.0mg/mL HSA and 20mg/mL trypsin) of spectrophotometer measurement variable concentrations.Reference solution is for containing the tryptic aqueous solution of 4.0mg/mL HSA and 20mg/mL.The concentration of ADR is respectively 0.5,1.0,2.5,5.0,15.0 and 30.0ug/mL.Can obtain standard curve by ADR concentration and the mapping of ABS value.Weighing 24mg embedding and the not pharmaceutical carrier of embedding ADR again, make it to be dissolved in the trypsin solution of 6mL 20mg/mL, at 37 ℃ of degraded 4.0h, the ABS value of the pharmaceutical carrier solution that is embedded with ADR of having degraded with spectrophotometer measurement (with the pharmaceutical carrier solution of the not embedding ADR that has degraded as reference solution), can obtain the concentration of corresponding ADR by above-mentioned standard curve, medicine charging ratio and embedding rate can calculate by following two formula:
(3) release in vitro of thermal targeting nano ball anti-cancer medicine carrier
Twice weighing 30mgADR-AN makes it to be scattered in respectively 37 ℃ phosphate buffer (PBS respectively, pH7.4,10mM) and contain the tryptic phosphate buffer of 0.20mg/mL, more respectively twice weighing 30mg ADR-PAN1 and the ADR-PAN2 20mL that makes it to be scattered in respectively 37 ℃ and 43 ℃ contain the tryptic phosphate buffer of 0.20mg/mL.Place water-bath slowly to stir each suspension, each sample is carried out that super centrifugal (44900 * g 20min), and takes out 1.0mL and carries out spectrophotometric analysis (concentration of the ADR that measurement discharges) from supernatant through predetermined time interval.The amount of the ADR that discharges accounts for the ratio of total embedding amount (amount of the initial ADR that adds of=embedding rate *) for discharging percentage ratio.The ADR that contains ADR-AN in the tryptic phosphate buffer of 0.20mg/mL in the neutralization of 37 ℃ phosphate buffers discharges, discharge and the results are shown in Figure 6 at the ADR of ADR-PAN1 and ADR-PAN2 that contains in the tryptic phosphate buffer of 0.20mg/mL of 37 ℃ and 43 ℃.
The result shows: (1) ADR of ADR-AN in not containing tryptic phosphate buffer discharges slowly, yet the ADR of ADR-AN discharges faster in containing tryptic phosphate buffer.Reason is the quickening that trypsin degradation albumin nanometer ball causes rate of release; (2) in the time of 37 ℃, the ADR of ADR-AN discharges the fastest, the ADR of ADR-PAN1 discharges and takes second place, the ADR of ADR-PAN2 discharges the slowest, in addition, in the time of 43 ℃, it is fast that the ADR of ADR-PAN1 discharges the also ADR release than ADR-PAN2, the high molecular molecule quantitative change of thermosensitive type conference of this explanation surface graft causes the drug release of this carrier slack-off, and it is faster than the drug releasing rate that is connected to the high molecular pharmaceutical carrier of thermosensitive type not to be connected to the high molecular pharmaceutical carrier of thermosensitive type; (3) ADR-PAN1 and ADR-PAN2 are fast than the drug release medicine under 37 ℃ at the drug release under 43 ℃, this is because 43 ℃ of high molecular LCST of thermosensitive type greater than ADR-PAN1 and ADR-PAN2 surface graft, therefore, contraction has taken place in the hydrophilic polymer on ADR-PAN1 and ADR-PAN2 surface, make the gap between the macromolecular chain increase, thereby cause enzyme easier, so the release of cancer therapy drug can be accelerated with the contact of protein nano ball.
Claims (6)
1. thermal targeting nano ball anti-cancer medicine carrier, it is characterized in that, the surface that has the Biodegradable nanometer ball of amino or carboxyl on the surface has lower critical solution temperature 37-43 ℃ and terminal PNIPAM class thermosensitive type macromolecule with carboxyl or amino by carboxyl and amino reaction covalent cross-linking, and its structure is
Wherein, described thermosensitive type macromolecule is selected from one or more in one section allylamine of an end block copolymerization or the acrylic acid PNIPAM class material, and its lower critical solution temperature is 37-43 ℃;
Wherein, the material of described nanosphere is selected from protein, polylysine, poly arginine, polyglutamic acid, poly-aspartic-acid, polypeptide, chitosan.
2. a kind of thermal targeting nano ball anti-cancer medicine carrier as claimed in claim 1, the particle diameter that it is characterized in that described nanosphere is less than 200nm.
3. the preparation method of the described thermal targeting nano ball anti-cancer medicine carrier of claim 1 is characterized in that comprising the steps:
1) makes the nanosphere pharmaceutical carrier with surperficial Biodegradable material with amino or carboxyl;
2) the surperficial covalent cross-linking of nanosphere pharmaceutical carrier a kind of have lower critical solution temperature 37-43 ℃ and terminal have carboxyl or an amino PNIPAM class thermosensitive type macromolecule.
4. the preparation method of a kind of thermal targeting nano ball anti-cancer medicine carrier as claimed in claim 3 is characterized in that, the making of described nanosphere pharmaceutical carrier is selected from following three kinds of methods:
1) Water-In-Oil method: with having dissolved the aqueous solution of hydrophilic anticarcinogen and hydrophilic film material as water, with the organic solvent that has dissolved oil soluble emulsifying agent as oil phase, with water mix with oil phase stir carry out rough segmentation and loose after, reuse ultrasonic cell-break machine carries out emulsifying, obtain water-in-oil type nanoemulsion liquid, under magnetic agitation, in the gained nanoemulsions, add cross-linking agent again and carry out crosslinking curing, remove excessive cross-linking agent and emulsifying agent and can obtain the nanosphere pharmaceutical carrier;
2) oil-in-water method: with having dissolved the organic solvent of hydrophobicity anticarcinogen and hydrophobic film material as oil phase, with the aqueous solution that has dissolved water soluble emulsifier as water, after oil phase stirring mixed with water carried out rough segmentation and loose, reuse ultrasonic cell-break machine carries out emulsifying, obtain the oil-in-water type nanoemulsions, under magnetic agitation, in the gained nanoemulsions, add cross-linking agent again and carry out crosslinking curing, remove excessive cross-linking agent and emulsifying agent and can obtain the nanosphere pharmaceutical carrier;
3) W/O/W multi-emulsion method: with having dissolved the aqueous solution of hydrophilic anticarcinogen as water, with the organic solvent that has dissolved hydrophobic film material and oil soluble emulsifying agent as oil phase, with water mix with oil phase stir carry out rough segmentation and loose after, reuse ultrasonic cell-break machine carries out emulsifying, obtain water-in-oil type nanoemulsion liquid, again gained water-in-oil type nanoemulsion liquid is added the aqueous phase that is dissolved with water soluble emulsifier and carry out ultrasonic emulsification, thereby obtain W/O/W type nanoemulsions, under magnetic agitation, in gained W/O/W type nanoemulsions, add cross-linking agent again and carry out crosslinking curing, remove excessive cross-linking agent and emulsifying agent and can obtain the nanosphere pharmaceutical carrier.
5. the manufacture method of a kind of nanosphere pharmaceutical carrier as claimed in claim 4 is characterized in that, the concentration range of described film material is 1.0%-30.0% (w/v).
6. the manufacture method of a kind of nanosphere pharmaceutical carrier as claimed in claim 4 is characterized in that, described emulsifier concentration scope is 0.1%-8.0% (w/v).
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2006101442550A CN101190334B (en) | 2006-11-30 | 2006-11-30 | Thermal targeting nano ball anti-cancer medicine carrier and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2006101442550A CN101190334B (en) | 2006-11-30 | 2006-11-30 | Thermal targeting nano ball anti-cancer medicine carrier and preparation method thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN101190334A CN101190334A (en) | 2008-06-04 |
| CN101190334B true CN101190334B (en) | 2010-09-15 |
Family
ID=39485628
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2006101442550A Expired - Fee Related CN101190334B (en) | 2006-11-30 | 2006-11-30 | Thermal targeting nano ball anti-cancer medicine carrier and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN101190334B (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102389572A (en) * | 2011-11-02 | 2012-03-28 | 中国科学院过程工程研究所 | Chitosan-carboxylated chitosan nanosphere loading insoluble antitumor drug and preparation method and application thereof |
| CN103239410B (en) * | 2013-05-02 | 2015-01-28 | 上海交通大学 | Preparation method of fluorescence and heat/pH sensitive organic-inorganic composite microsphere carrier |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1720996A (en) * | 2005-06-17 | 2006-01-18 | 南京大学医学院附属鼓楼医院 | Antineoplastic nanometer heat sensitive target medicine carrier in conjunction with the application of chemotherapy thermotherapy |
-
2006
- 2006-11-30 CN CN2006101442550A patent/CN101190334B/en not_active Expired - Fee Related
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1720996A (en) * | 2005-06-17 | 2006-01-18 | 南京大学医学院附属鼓楼医院 | Antineoplastic nanometer heat sensitive target medicine carrier in conjunction with the application of chemotherapy thermotherapy |
Non-Patent Citations (6)
| Title |
|---|
| J. E. Chung et. al.."Thermo-responsive drug delicery from polyneric micellesconstructed using block copolymers ofpoly(N-isoptopylacrylamide) and poly(butylmethacrylate)".J. controlled Release62 1999.1999,62(1999),115-124. |
| J.E.Chung et.al.."Thermo-responsive drug delicery from polyneric micellesconstructed using block copolymers ofpoly(N-isoptopylacrylamide) and poly(butylmethacrylate)".J. controlled Release62 1999.1999,62(1999),115-124. * |
| Zhe-Yu Shen et. al.."Preparation and characterization of thermo-responsivealbumin nanospheres".International Journal of Pharmaceutics346 2008.2007,346(2008),133-142. |
| Zhe-Yu Shen et. al.."Preparation and characterization of thermo-responsivealbumin nanospheres".International Journal of Pharmaceutics346 2008.2007,346(2008),133-142. * |
| 王恺."白蛋白微球的制备、改性和应用的研究进展".国外医学药学分册30 6.2003,30(6),366-369. |
| 王恺."白蛋白微球的制备、改性和应用的研究进展".国外医学药学分册30 6.2003,30(6),366-369. * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN101190334A (en) | 2008-06-04 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Preman et al. | Recent developments in stimuli-responsive polymer nanogels for drug delivery and diagnostics: A review | |
| Xu et al. | Multifunctional silica nanoparticles as a promising theranostic platform for biomedical applications | |
| Crucho | Stimuli‐responsive polymeric nanoparticles for nanomedicine | |
| Maya et al. | Smart stimuli sensitive nanogels in cancer drug delivery and imaging: a review | |
| Hamidi et al. | Hydrogel nanoparticles in drug delivery | |
| Sahil et al. | Microsphere: A review | |
| Zhang et al. | Advanced materials and processing for drug delivery: the past and the future | |
| Sun et al. | Temperature-sensitive gold nanoparticle-coated pluronic-PLL nanoparticles for drug delivery and chemo-photothermal therapy | |
| Miao et al. | Charge reversible and biodegradable nanocarriers showing dual pH-/reduction-sensitive disintegration for rapid site-specific drug delivery | |
| Aggarwal et al. | Gemcitabine-loaded PLGA-PEG immunonanoparticles for targeted chemotherapy of pancreatic cancer | |
| Adamo et al. | Functionalization of nanoparticles in specific targeting and mechanism release | |
| Nakache et al. | Biopolymer and polymer nanoparticles and their biomedical applications | |
| JP2014518862A (en) | Polymer nanoparticles for drug delivery | |
| US20050084456A1 (en) | Functionalized particles | |
| Meng et al. | Uptake and metabolism of novel biodegradable poly (glycerol-adipate) nanoparticles in DAOY monolayer | |
| CN107617108A (en) | A kind of core crosslinking nano grain of double targetings and pH/ redox sensitives and its preparation method and application | |
| WO2015027342A1 (en) | Microgel compositions for delivery of substances to the brain | |
| CN101190334B (en) | Thermal targeting nano ball anti-cancer medicine carrier and preparation method thereof | |
| Du et al. | Temperature/pH-responsive carmofur-loaded nanogels rapidly prepared via one-pot laser-induced emulsion polymerization | |
| Asem et al. | Polymeric nanoparticles explored for drug-delivery applications | |
| Van Herk et al. | Controlled release systems: advances in nanobottles and active nanoparticles | |
| Daramola et al. | Polymer nanoparticles (nanomedicine) for therapeutic applications | |
| Malikmammadov et al. | Dual-and multistimuli-responsive polymers for biomedical applications | |
| Chouhan et al. | An updated review on nanoparticle based approach for nanogel drug delivery system | |
| Patel et al. | Targeting aspects of hydrogels in drug delivery |
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 | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20100915 Termination date: 20151130 |
|
| EXPY | Termination of patent right or utility model |