CN100471886C - Amphiphilic block copolymer and polymeric composition comprising the same for drug delivery - Google Patents

Abstract

The present invention relates to an amphiphilic block copolymer of a hydrophilic block and a hydrophobic block with a terminal hydroxyl group wherein the terminal hydroxyl group of the hydrophobic bock is substituted with a tocopherol or cholesterol group. It also relates to polymeric compositions capable of forming stable micelles in an aqueous solution, comprising the amphiphilic block copolymer and a polylactic acid derivative wherein one or more ends of the polylactic acid are covalently bound to at least one carboxyl group.

Description

Amphiphilic block copolymer and the polymeric composition that is used for administration that comprises it

Technical field

The amphiphilic block copolymer that the present invention relates to comprise hydrophilic A block and have the hydrophobic B block of terminal hydroxyl, the terminal hydroxyl of wherein said hydrophobic block is replaced by tocopherol or cholesterol group.The invention still further relates to the polymeric composition that comprises described amphiphilic block copolymer and polylactic acid derivative, wherein one or more ends of poly(lactic acid) and at least one carboxyl covalent attachment.Also relate to the polymeric composition of having fixed metal ion (metal ion-fixed) in addition, wherein the C-terminal of polylactic acid derivative with two or tervalent metal ion fix.

Background technology

In the time of in medicine is given to body, only have a spot of medicine can arrive its target position, the dosage that great majority give all is distributed on the non-target position, and can cause bad side effect.Therefore, in the past twenty years, research work just concentrates on by use and comprises that the suitable carrier development of liposome, small molecules surfactant micelle, polymer/nanometer grain and polymeric micelle (by the micelle formation polymer/nanometer grain of hardening) can effectively position the system of release administration.Use liposome to be found limitation to some extent as pharmaceutical carrier, this mainly is because its low load effectiveness, medicine unstable, medicine spills and weak storage stability fast.After being administered to the small molecules surfactant micelle in the body, when they are diluted in body fluid, just be easy to be dissociated, so they are difficult to give full play to the effect as pharmaceutical carrier.

Recently, have and report that the polymer/nanometer grain and the polymeric micelle that have utilized biodegradable polymer are the very useful technology that overcomes these problems.They have changed by the distribution in vivo of intravenously administered agents, thus therefore reduced side effect and improved its effect provide this as the cell-specific target advantage and controlled the release of medicine.They and body fluid also have good consistency, and have improved the solubleness and the bioavailability of drugs of low aqueous solubility.

Pharmaceutical carrier with nanometer size of water-wetted surface has been found identification and the picked-up that can hide reticuloendothelial system (RES), so just can circulation for a long time in blood.Another advantage of these hydrophilic nanoparticles is, because their very little, so particle can for example be drifted at the solid tumor place at pathogenic site by passive target-seeking mechanism.Yet medicine successfully is transported to specific target position need stablize the reservation medicine by carrier in circulation.Because drug targeting requires long cycling time, carrier is exposed in the blood ingredient for a long time, therefore need improve medicine-carrier-bound stability, and carrier is not removed rapidly.

Have in the pharmaceutical carrier of nanometer size of water-wetted surface at these, polymeric micelle generally includes a hundreds of segmented copolymer, and diameter is approximately 20nm to 50nm.Polymeric micelle have two spherical common-central zone, be responsible for the load dewatering medicament dense arrangement the hydrophobic substance core and can be in blood long-time round-robin be used to hide the shell of forming by hydrophilic substance of health RES.For example size is little, solubleness is high, sterilization is simple and control drug release although they have unique advantage, and owing to the medicine that may occur combining is in vivo discharged rapidly, so the physical stability of these carriers is key issues.

If the concentration of total copolymer is on micelle-forming concentration (CMC), micella is exactly thermodynamically stable so.Like this, use copolymer systems just may improve micellar body internal stability with low CMC value.Kinetic stability refers to the micellar rate of decomposition.Rate of decomposition depends on the physical condition of micella core.The micella that is formed by the multipolymer that comprises the hydrophobic block with high glass transition temperature will dissociate than the micella which has a low glass state invert point slowly.They also may be subjected to many influences that can influence the same factor of single aggressiveness rate of exchange between micella.The rate of exchange that has been found that single aggressiveness depends on for example solvent, the hydrophobic content of multipolymer and the length of hydrophilic and hydrophobic block in the core of many factors.

People have paid huge effort for biodegradable and the biocompatible core-can type pharmaceutical carrier of being devoted to develop the energy load drugs of low aqueous solubility with improved stability and effectiveness.EP 0,552,802 A2 disclose the method for preparing the chemical fixation polymeric micelle, and wherein polymkeric substance is core-can type polymkeric substance, and it contains as the hydrophilic polyethylene oxide of shell with as can be in the aqueous solution crosslinked hydrophobic biodegradability polymkeric substance of core.Yet these polymeric micelles are difficult to preparation, because must add linking agent in the hydrophobic ingredient of A-B type diblock or A-B-A type triblock copolymer, the polymkeric substance that forms core so just has a stable structure.Also have, before using never in human body used linking agent can increase worry to security.

United States Patent (USP) 6,080,396 disclose the micella that forms the block copolymer-medicine mixture.The high-molecular block copolymer-medicine mixture, wherein high-molecular weight segmented copolymer possess hydrophilic property polymer segments and hydrophobic polymer section have formed the micella that has as the hydrophilic segment of shell, and comprise anthracene nucleus class carcinostatic agent in its hydrophobic inner core.Carcinostatic agent molecule and micella core are covalently bound.Yet, when medicine and polymeric micelle covalent attachment, just be difficult to the rate of decomposition of control medicine-multipolymer bonding.

On the other hand, have report to show that the dissolving of dewatering medicament can realize by polymeric micelle, wherein for example form by two or triblock copolymer of fatty acid polyester or polyamino acid by the hydrophilic polymer that comprises polyalkylene glycol (polyalkylene glycol) derivative and hydrophobicity biodegradable polymer for micella.United States Patent (USP) 5,449,513 disclose and have contained as the polyoxyethylene glycol of hydrophilic polymer with as the polyamino acid derivative of the hydrophobic polymer Synthetic rubber, isoprene-styrene, hydrogenated, block, diblock of poly-benzyl aspartic acid etc. for example.This Synthetic rubber, isoprene-styrene, hydrogenated, block, diblock can solubilizing hydrophobic carcinostatic agent Zorubicin for example, or anti-inflammatory agent indomethacin for example.Yet polyamino acid derivative can not be hydrolyzed in vivo, will cause side effect because of the immune response that is triggered like this.

A method improving polymeric micelle stability is to improve the hydrophobicity of polymkeric substance.In order to do like this, molecular weight or concentration that just should telomerized polymer.Yet along with the increase of molecular weight, its biodegradability is also reducing, and therefore, polymkeric substance is just insufficient from the drainage of health, and they accumulate in the organ and cause toxic action at that.United States Patent (USP) 5,429,826 disclose two or the segmented copolymer that contains wetting ability polyalkylene glycol and hydrophobicity poly(lactic acid).Specifically be, this patent has been put down in writing by micellization two or segmented copolymer and has been come stable polymerization micellar method, and wherein acrylic acid derivative combines with two or the end group of segmented copolymer, and then, in the aqueous solution, polymkeric substance takes place crosslinked so that form micella.Above method can realize stable to polymeric micelle, but crosslinked polymkeric substance can not be degraded, and therefore just can not use in vivo.Above polymeric micelle can be a large amount of drugs of low aqueous solubility of dissolving in the neutral aqueous solution at pH, has just discharged at short notice but shortcoming is a medicine.In addition, at United States Patent (USP) 6,458, in 373, drugs of low aqueous solubility is dissolved in the emulsion form that has alpha-tocopherol.According to this patent,, used PEGization (PEGylated) vitamin-E as tensio-active agent for stable emulsion.The PEGization vitamin-E has the structure similar to amphiphilic block copolymer, and wherein segmented copolymer comprises hydrophilic block and hydrophobic block, and the tocopherol of high hydrophobicity has improved the avidity of multipolymer and drugs of low aqueous solubility, so just can dissolve drugs of low aqueous solubility.Yet, have limited molecular weight as the polyoxyethylene glycol of hydrophilic polymer, thus the PEGization vitamin-E separately only can solubilizing hydrophobic drug for example taxol to 2.5mg/ml.Under 2.5mg/ml or higher solubleness, formed unsettled micella, and drug crystallization is easy to form precipitation.

Clinical tumor is an inherent or posteriori to the resistance to medicine characteristic of chemotherapy.The inherent resistance is present in when diagnosis can not reply in the chemotherapeutical tumour of first line.Posteriori resistance often appears in the responsive tumour of initial therapy, but can recur, and demonstrates diverse phenotype.Resistance to medicine characteristic is produced by former used medicine with different structure and mechanism of action and novel drugs.For example, use Often failure is because cancer cells has produced posteriori resistance to the cancer chemotherapy that carries out, and this is often relevant with the variation of the overexpression of P-gp and 'beta '-tubulin.Tolerance

Cell and the other medicines that comprise dactinomycin, Zorubicin, vinealeucoblastine(VLB) and vincristine(VCR) intersect anti-medicine.Therefore, before chemotherapy can be cured the patient who suffers from metastatic carcinoma, the resistance of clinical medicine was the obstacle that mainly will overcome.

The resistance cancer cells has shown higher IC ₅₀(drug concentrations when 50% cell is suppressed) therefore for chemotherapy can effectively be carried out, just needs higher drug level to tumour cell, and normal cell then wishes to reduce drug level.Therefore, require long body circulation and the certain position of medicine in tumor tissues to improve the effectiveness of antagonism drug resistant cancer.

By foregoing, people have recognized and have thirsted for developing the polymeric micelle compositions of improvement that is used to carry dewatering medicament with biocompatibility and biodegradability.The invention provides this polymeric micelle polymkeric substance with improvement of biocompatibility and biodegradability, it can effectively be carried dewatering medicament and can not reduce its stability.

The invention summation

One aspect of the present invention relates to amphiphilic block copolymer of the hydrophobic B block that contains hydrophilic A block and have terminal hydroxyl and preparation method thereof, and the terminal hydroxyl of wherein said hydrophobic block is replaced by tocopherol or cholesterol group.Amphiphilic block copolymer of the present invention has significantly improved the hydrophobicity of hydrophobic block and has kept and natural polymer molecular weight much at one.In addition, hydrophobic functional group has significantly improved the avidity with drugs of low aqueous solubility, so just can be more stable in the aqueous solution by the polymeric micelle of this polymer formation, and can keep dissolved drugs of low aqueous solubility wherein in the time that prolongs, to keep high Plasma Concentration.In addition, amphiphilic block copolymer can with other mixed with polymers, make the polymeric composition that is used for administration.

Another aspect of the present invention relates to the amphiphilic block copolymer of the hydrophobic B block that contains hydrophilic A block and have terminal hydroxyl and the polymeric composition of polylactic acid derivative, the terminal hydroxyl of wherein said hydrophobic B block is replaced by tocopherol or cholesterol group, terminal and at least one the carboxyl covalent attachment of at least one of polylactic acid derivative.

The 3rd aspect of the present invention relates to the amphiphilic block copolymer of the hydrophobic B block that contains hydrophilic A block and have terminal hydroxyl and the polymeric composition of polylactic acid derivative, the terminal hydroxyl of wherein said hydrophobic B block is replaced by tocopherol or cholesterol group, at least one of polylactic acid derivative terminal with at least one carboxyl covalent attachment, the C-terminal group of its polylactic acid derivative with two or trivalent metal ion fix.

Polymeric composition of the present invention can form stable polymeric micelle or nanoparticle in the body fluid or the aqueous solution.Micella that is formed by the present composition or nanoparticle be poly-hydrophilic outer shell and hydrophobic inner core, and wherein a large amount of dewatering medicaments can be by the physics load.Pastille micella of the present invention and nanoparticle be in the residence time that has prolongation after the administration in blood, and can be used to form various pharmaceutical preparations.The polymeric micelle that comprises anticarcinogen by present composition preparation can be transferred to effectively and be acted on effectively in the cancer cells of tolerance anticarcinogen.Other characteristics of the present invention and advantage will occur in the following detailed description, and obtain in conjunction with the accompanying drawings, they with the mode illustrated together of embodiment characteristics of the present invention.

The accompanying drawing summary

Other characteristics of the present invention and advantage will present in the following detailed description, and obtain in conjunction with the accompanying drawings, they with the mode illustrated together of embodiment characteristics of the present invention, wherein:

Fig. 1 is the synoptic diagram of the polymeric micelle that formed in water surrounding by mono methoxy polyethylene glycol-polylactide-hydrophobic part (mPEG-PLA-hydrophobic part);

Fig. 2 is by carboxylic acid sodium deutero-D, the synoptic diagram of the polymeric micelle that the L-poly(lactic acid) forms in water surrounding;

Fig. 3 is by mono methoxy polyethylene glycol-polylactide-hydrophobic part (mPEGA-PLA-hydrophobic part) and carboxylic acid sodium deutero-D, the synoptic diagram of the polymeric micelle that the L-poly(lactic acid) forms in water surrounding;

Fig. 4 has fixed Ca ²⁺The synoptic diagram of Fig. 3 polymeric micelle;

Fig. 5 is the Ca that fixed that comprises by the dewatering medicament of load in the micella hydrophobic core ²⁺Polymeric micelle;

Fig. 6 is mPEG-PLA-cholesterol (embodiment 1) ¹The H-NMR spectrum;

Fig. 7 is mPEG-PLA-tocopherol (embodiment 7) ¹The H-NMR spectrum;

Fig. 8 has shown the polymeric micelle that comprises taxol different time plasma concentration curve at interval after administration made from different Synthetic rubber, isoprene-styrene, hydrogenated, block, diblocks;

Fig. 9 has shown the Ca that fixed that comprises taxol that makes with mPEG-PLA-tocopherol and mPEG-PLA-OH ²⁺Polymeric micelle different time plasma concentration curve at interval after administration;

Figure 10 has shown the Ca that fixed that comprises taxol ²⁺Polymeric micelle, Cremophor EL (

) and tween 80 preparation different time plasma concentration curve at interval after administration;

Figure 11 has shown the Ca that fixed that comprises taxol ²⁺Polymeric micelle and Cremophor EL (

) different time plasma concentration curve at interval after administration;

Figure 12 has shown the Ca that fixed that comprises taxol ²⁺Polymeric micelle and the tween 80 preparation (

) different time plasma concentration curve at interval after administration;

Figure 13 has shown the Ca that fixed that comprises the Japanese yew terpene ²⁺Polymeric micelle and the tween 80 preparation ( ) different time plasma concentration curve at interval after administration;

Figure 14 has shown to comprise and has fixed Ca ²⁺The medicine of polymeric micelle be anticancer effect in the MX-1 mouse having used human breast cancer cell;

Figure 15 has shown to comprise and has fixed Ca ²⁺The medicine of polymeric micelle be anticancer effect in the MDAMB435S mouse having used human breast cancer cell;

Figure 16 has shown to comprise and has fixed Ca ²⁺The medicine of polymeric micelle be anticancer effect in the SKOV-3 mouse having used Proliferation of Human Ovarian Cell;

Figure 17 has shown to comprise and has fixed Ca ²⁺The medicine of polymeric micelle be anticancer effect in the SKOV-3 mouse having used Proliferation of Human Ovarian Cell;

Figure 18 has shown to comprise and has fixed Ca ²⁺The anticancer effect (3 cycle) of medicine in having used CCL188 HT-29 mouse of polymeric micelle;

Figure 19 has shown to comprise and has fixed Ca ²⁺The anticancer effect of medicine in having used CCL188 HT-29 mouse of polymeric micelle;

Figure 20 has shown to comprise and has fixed Ca ²⁺The anticancer effect of medicine in having used PC-3 PC3 mouse of polymeric micelle;

Figure 21 has shown to comprise and has fixed Ca ²⁺The anticancer effect of medicine in having used human brain cancerous cell line U-373MG mouse of polymeric micelle;

Figure 22 has shown to comprise and has fixed Ca ²⁺Polymeric micelle medicine to taxol (

) anticancer effect in drug-fast people's carcinoma animal model; And

Figure 23 has shown to comprise and has fixed Ca ²⁺Polymeric micelle medicine to Zorubicin (

) anticancer effect in drug-fast people's carcinoma animal model.

Referring now to illustrational representational embodiment, will same content be described with specific language at this. Yet we should be understood that therefore scope of the present invention can not be restricted.

Detailed Description Of The Invention

At this polymeric compositions of disclosure and description and before using and making its method, should be understood that the present invention is not limited by this special combination disclosed herein, method step and material, this combination, method step and material can be various. We it is also to be understood that term is only for describing particular as used herein, and this does not limit the scope of the invention, and scope of the present invention only is subjected to the restriction of accessory claim and its equivalent.

Should be noted that in this specification and accessory claim, unless context stipulates that clearly otherwise singulative " ", " one " or " this " comprise plural object. Like this, for example mention the polymer that comprises " end group " and just comprise and relate to two or more this groups, mention " dewatering medicament " and just comprise and relate to two or more this medicines. In addition, mention the mixture that an amphiphilic block copolymer just comprises block copolymer, condition is that wherein block copolymer provides the composition of each A and B block, and the weight of each block ratio separately and each block and/or whole block polymerization composition or number-average molecular weight fall in the limited field defined here.

Describing and asking for protection in the process of the present invention, will use following term according to the following definition that provides.

Term " biologically active agent " or " medicine " or other similar term refer to any chemistry or biomaterial or compound as used herein, it is suitable for the method afford by existing method known in the art and/or the present invention's instruction, and biology or the pharmacotoxicological effect that can induce serious hope to have.Such effect can include but not limited to that (1) is to organic prophylactic effect with prevent that the biological action of not expecting from for example protecting from infection, (2) alleviate the situation that causes by disease and for example alleviate pain or the inflammation that causes by disease, and/or (3) are alleviated, reduced or eliminate organic disease fully.This effect can be partial, and partial anaesthetic effect for example is provided, and perhaps can be whole body.

The effect of the definition of term " biodegradable " or " biological degradation " entity (it can be enzyme or organic other product) that is material by dissolving hydrolysis or biological form is converted into less compound intermediate or end product as used herein.

Term " but physiologically acceptable " refers to material or intermediate or the end product that the effect by the entity (it can be enzyme or organic other product) of dissolving hydrolysis or biological form forms as used herein, and they can not cause the untoward reaction of human body.

" significant quantity " refers to the amount at the promoting agent that can participate in the part that can be enough under the rational danger/beneficiary proportion of any medical treatment to provide desired or whole body effect as used herein.

" administration " and similar term refer to delivery of composition in treated individual as used herein, and composition just can be in systemic circulation like this.Preferably, composition of the present invention by subcutaneous, muscle, through skin, oral, give through mucous membrane, intravenously or intraperitoneal route of entry.The injectable thing of this purposes can be made into conventional form such as liquid solution or suspension, or is suitable for making in liquid before injection the solid form of solution or suspension, or emulsion.The proper excipient that can be used for administration comprises for example water, salt solution, glucose, glycerine, ethanol and analogue, if desired, also has auxiliary substance for example wetting or emulsifying agent, buffer reagent and analogue in a small amount.Concerning oral administration, it can be made into different forms for example solution, tablet, capsule etc.

Below provided representational embodiment, will use specific language to describe same content at this.Yet we should be understood that therefore scope of the present invention can not be restricted.In the other or another modification of this feature of the present invention that exemplifies, and, for a person skilled in the art, combine, all should be considered within the scope of the invention with these disclosed contents in other application of this principle of the invention that exemplifies.

On the one hand, originally provide the amphiphilic block copolymer that contains hydrophilic A block and have the hydrophobic B block of terminal hydroxyl, the terminal hydroxyl of wherein said hydrophobic block is replaced by tocopherol or cholesterol group.

The present invention also provides the method for preparing amphiphilic block copolymer, and for example Yi Xia method variant (variants) I is to III.

Method variant I:

The method that may further comprise the steps:

1) carboxylation has the hydrophobic compound of tocopherol or cholesterol group; With

2) exist under the situation of dicyclohexylcarbodiimide (DCC) as initiator, the amphiphilic block copolymer and the described carboxylation hydrophobic compound that will contain hydrophilic A block and have a hydrophobic B block of terminal hydroxyl react, like this hydrophobic compound of carboxylation with regard to chemical be attached on the terminal hydroxyl of hydrophobic B block.

Method variant II:

The method that may further comprise the steps:

1) carboxylation has the hydrophobic compound of tocopherol or cholesterol group and the carboxylation hydrophobic compound that obtains with the oxalyl chloride activation; With

2) amphiphilic block copolymer and the described carboxylation hydrophobic compound that has activated that will contain hydrophilic A block and have a hydrophobic B block of terminal hydroxyl reacts, like this hydrophobic compound of carboxylation with regard to chemical be attached on the terminal hydroxyl of hydrophobic B block.

3)

Method variant III:

The method that may further comprise the steps:

1) mixing has the hydrophobic compound of tocopherol or cholesterol group and the dichloride that conduct connects agent;

2) amphiphilic block copolymer that will contain hydrophilic A block and have a hydrophobic B block of terminal hydroxyl joins in the reaction mixture of step 1, like this hydrophobic compound with regard to chemical be attached on the terminal hydroxyl of hydrophobic B block; With

3) dissolving and settling step 2) segmented copolymer that obtains.

Term " hydrophobic compound of carboxylation " refers to the have hydroxyl hydrophobic compound of (carboxyl combines with it), and carboxyl can come from succinate, propanedioic acid, pentanedioic acid or hexanodioic acid.

The present invention also provides the pharmaceutical carrier that contains amphiphilic block copolymer of the present invention.The pharmaceutical composition that can form polymeric micelle in the body fluid or the aqueous solution also is provided, and it contains described amphiphilic block copolymer and drugs of low aqueous solubility.

Amphiphilic block copolymer of the present invention preferably contains the A-B type Synthetic rubber, isoprene-styrene, hydrogenated, block, diblock or the B-A-B type triblock copolymer of hydrophilic A block and hydrophobic B block, and the end group of hydrophobic block is a hydroxyl.When amphiphilic block copolymer of the present invention is placed aqueous environments, just formed the polymeric micelle of core-can type, wherein hydrophobic B block has formed core, and hydrophilic A block has formed shell.Preferably, hydrophilic A block is selected from polyalkylene glycol, polyvinyl alcohol, polyvinylpyrrolidone, polyacrylamide and derivative thereof.More preferably, hydrophilic A block is selected from mono methoxy polyethylene glycol, monovinylphenyl polyoxyethylene glycol, polyoxyethylene glycol, polyethylene-altogether-propylene glycol and polyvinylpyrrolidone.Preferably, the number-average molecular weight of hydrophilic A block is 200 to 50,000 dalton.More preferably, the number-average molecular weight of hydrophilic A block is 1,000 to 20,000 dalton.

The polymkeric substance that the hydrophobic B block of amphiphilic block copolymer of the present invention is high-biocompatibility and biodegradability, it is selected from polyester, polyanhydride, polyamino acid, poe and poly-phosphine piperazine.More preferably, hydrophobic B block is one or more materials that are selected from polylactide, polyglycolide, polycaprolactone, poly-dioxane-2-ketone, poly(lactic acid)-altogether-glycollide, poly(lactic acid)-altogether-dioxane-2-ketone, poly(lactic acid)-altogether-caprolactone and polyglycolic acid-altogether-caprolactone.Preferably, the number-average molecular weight of the hydrophobic B block of amphiphilic block copolymer is 50 to 50,000 dalton.More preferably, the number-average molecular weight of the hydrophobic B block of amphiphilic block copolymer is 200 to 20,000 dalton.

Hydrophobic B block has hydroxyl terminal, and this hydroxyl terminal is replaced by tocopherol or cholesterol, and the two all has extraordinary hydrophobicity, and it can improve the hydrophobicity of hydrophobic B block.When the hydrophobic block of amphiphilic block copolymer of the present invention was placed aqueous solution, it had been avoided and the contacting of water, and has formed kernel, to form the ball-type polymeric micelle.Like this, when being incorporated into drugs of low aqueous solubility in the amphiphilic block copolymer, this drugs of low aqueous solubility is just surrounded by hydrophobic polymer, and therefore, the kernel of polymeric micelle just can be by load in micella.Form the avidity that micellar stability depends on the hydrophobicity of hydrophobic block and it and medicine.Therefore, in the present invention, for the hydrophobicity that improves hydrophobic block keeps its molecular weight simultaneously, we have good hydrophobic functional group for example tocopherol, cholesterol etc. with connecting thinner in conjunction with last.Tocopherol and cholesterol are biocompatibility and the hydrophobic compounds with ring structure, and it can improve the avidity of segmented copolymer to drugs of low aqueous solubility.

The hydrophilic A block of amphiphilic block copolymer of the present invention and the weight ratio of hydrophobic B block are preferably in 30:70 arrives the scope of 97:3, more preferably in 4:6 arrives the scope of 7:3.If the content of hydrophilic A block is too low, polymkeric substance just may not can in the aqueous solution forms polymeric micelle so, if content is too high, the polymeric micelle of Xing Chenging is just unstable so thus.

In one embodiment, amphiphilic block copolymer of the present invention can be represented by following formula:

R _1’-O-[R _3’] _1’-[R _4’] _m’-[R _5’] _n’-C(＝O)-(CH ₂) _X’-C(＝O)-O-R _2’ (I’)

R wherein _{1 '}Be CH ₃-, H-[R _{5 '}] _{N '}-[R _{4 '}] _{M '}-or R _{2 '}-O-C (=O)-(CH ₂) _{X '}-C (=O)-[R _{5 '}] _{N '}-[R _{4 '}] _{M '}-;

R _{2 '}Be tocopherol or cholesterol;

R _{3 '}Be-CH ₂CH ₂-O-,-CH (OH)-CH ₂-,-CH (C (=O)-NH ₂)-CH ₂-or

R _{4 '}Be-C (=O)-and CHZ '-O-, wherein Z ' is hydrogen atom or methyl;

R _{5 '}Be-C (=O)-CHY "-O-, wherein Y " be hydrogen atom or methyl ,-C (=O)-CH ₂CH ₂CH ₂CH ₂CH ₂-O-or-C (=O)-CH ₂OCH ₂CH ₂-O-;

1 ' is the integer from 4-1150;

M ' is the integer from 1-300;

N ' is the integer from 0-300; With

X ' is the integer from 0-4.

Compare with existing amphiphilic block copolymer, multipolymer with hydrophobic block that is replaced by hydrophobic grouping of the present invention has the enhanced hydrophobicity, the micelle-forming concentration (CMC) that reduces and raising and avidity drugs of low aqueous solubility like this, just can be held medicine in stable environment.In addition, because functional group's combination endways, therefore the micellar size that forms in the aqueous solution can increase, and like this, just has the medicine of q.s to be comprised in the micella.Therefore, amphiphilic block copolymer can be used as pharmaceutical carrier effectively.The functional group with strong-hydrophobicity who is incorporated among the present invention has high molecular, and it just can significantly improve the avidity of hydrophobicity and segmented copolymer and medicine like this, has so just stablized the pastille micella effectively.

In addition, the polymeric micelle that is formed by amphiphilic block copolymer of the present invention has the residence time of prolongation in vivo.The haemoconcentration of polymeric micelle Chinese traditional medicine depends on the hydrophobic part that is used to replace the hydrophobic B block of amphiphilic block copolymer hydroxyl terminal.As Fig. 6 and shown in Figure 8, the polymeric micelle (composition 1-2) with amphiphilic block copolymer of the hydrophobic part (tocopherol or cholesterol) that is used for replacing hydrophobic B block hydroxyl terminal has than initial mPEG-PLA-OH polymeric micelle (composition 3) the longer residence time at blood.In addition, in all polymeric micelles, mPEG-PLA-tocopherol micella (composition 1) is the longest the cycle time in blood.This result can improve hydrophobicity by tocopherol and cholesterol moiety in the hydrophobic B block and explained.

Having hydroxyl terminal can be according to following method preparation by the segmented copolymer of the hydrophobic block of tocopherol or cholesterol replacement.In one embodiment,, be incorporated in the hydroxyl of tocopherol or cholesterol suitable linking agent for example dicarboxylic acid such as succsinic acid, propanedioic acid, pentanedioic acid or hexanodioic acid, carboxylation tocopherol or the cholesterol Chemical bond hydroxyl terminal of hydrophobic B block.

In one embodiment, according to United States Patent (USP) 6,322,805 method, weighing contains mono methoxy polyalkylene glycol (mPEG; Mn=20,000) and polylactide (PLA; Mn=1,750) amphiphilic block copolymer (mPEG-PLA) dewaters with vacuum pump at 120 ℃.Be dissolved in then in acetonitrile or the methylene dichloride.To wherein adding tocofecol succinic acid salt or cholesterol succinate, weighing dicyclohexylcarbodiimide (DCC) and 4-dimethylaminopyridine (DMAP) join wherein as initiator and catalyzer respectively, and at room temperature react.Owing in the reaction of end-OH of mPEG-PLA and hydrophobic compound-COOH, formed dicyclohexylurea (DCU), so reactant becomes opaque.After 24 hours, use glass filter to remove DCU, extract and remove DMAP with aqueous hydrochloric acid.In this purified product solution, add MgSO4 to remove any remaining moisture content; in hexane/diethylether solvent, form precipitation so that obtain to combine the amphiphilic block copolymer of tocopherol succinyl-or cholesterol succinyl-, mPEG-PLA-tocopherol or mPEG-PLA-cholesterol (wherein tocopherol or cholesterol combine with PLA by succinic diester) then.The polymerisate of filtering-depositing, dry to obtain the particulate polymkeric substance that is white in color under vacuum then.

In another embodiment, with the hydrophobic compound of oxalyl chloride activation carboxylation, need not any catalyzer, and combine with the end of mPEG-PLA.In other words, with the reaction of tocopherol (or cholesterol) succinate and oxalyl chloride, then at room temperature vacuum remove more than oxalyl chloride.Weighing mPEG-PLA also joins wherein, obtains mPEG-PLA-tocopherol (or cholesterol) in 12 hours 100 ℃ of reactions.This synthetic polymer is dissolved in acetonitrile or methylene dichloride, in hexane/diethyl ether, precipitates, filter.

In above two preparation process, tocopherol (or cholesterol) malonate, tocopherol (or cholesterol) glutarate or tocopherol (or cholesterol) adipate etc. can both be used for substituting tocopherol (or cholesterol) succinate.

In another embodiment, tocopherol or cholesterol combine with the end of mPEG-PLA by using as the dichloride that connects agent.Specifically be that weighing tocopherol or cholesterol are also dewatered with vacuum pump at 50 ℃.To wherein adding excessive connection agent, reacted 12 hours.The reaction finish after, 100 ℃ of following vacuum remove add into the connection agent.To the mPEG-PLA that wherein adds weighing, reacted 12 hours down at 100 ℃.This synthetic polymer is dissolved in the methylene dichloride, and in hexane/diethylether solvent, precipitates so that obtain amphiphilic block copolymer (wherein tocopherol or cholesterol combine with PLA by succinic diester), i.e. mPEG-PLA-tocopherol or mPEG-PLA-cholesterol.The polymerisate of filtering-depositing, vacuum-drying, the particulate polymkeric substance obtains being white in color.The connection agent that can be used for reacting can be selected from dichloride such as succinyl dichloride, oxalyl chloride, malonyl chloride, glutaryl chlorine, hexanedioyl chlorine etc.

Above synthetic segmented copolymer can mix so that obtain polymeric micelle compositions with drugs of low aqueous solubility.In other words, segmented copolymer (10-200mg) and medicine (1-50mg) are dissolved in the organic solvent for example acetonitrile, methylene dichloride etc.Mix this solution, 60 ℃ of following vacuum-dryings are to make matrix.In distilled water, freeze-drying is to obtain introducing the polymeric micelle compositions of medicine then with the matrix of drugs of low aqueous solubility and polymer dissolution.Above polymeric micelle compositions can the water solution dilution, physiological saline for example, and as injectable formulation.

Term " drugs of low aqueous solubility " or " dewatering medicament " refer to any medicine or the biologically active agent with 33.3mg/ml or lower water solubility.This comprises the Parkinsonian medicament of medicament, antihyperlipidemic, antiviral agent, treatment, dementia resisting medicine, antiemetic, immunosuppressor, anti-ulcerative drug, caccagogue and the anti-malarial agents of carcinostatic agent, microbiotic, anti-inflammatory agent, narcotic, hormone, hypotensive agent and treatment diabetes.The example of dewatering medicament comprises taxol, KETOKONAZOL, itraconazole, S-Neoral, cisapride, Paracetamol, Asprin, acetylsalicylic acid, indomethacin, naprosine, warfarin, Papaverine, Top Form Wormer, miconazole, cinarizine, Zorubicin, omeprazole, Vitamin D3 500,000 I.U/GM, melphalan, nifedipine, digoxin, the phenylformic acid tryptophane, tyrosine, phenylalanine, azthreonam, Ibuprofen BP/EP, phenoxymethylpenicillin, Thalidomide, methyltestosterone, prochlorperazine, Kendall compound, two deoxidation purine nucleoside, vitamin D2, White streptocide, sulfonylurea, Para-Aminobenzoic, melatonin, benzylpenicillin, Chlorambucil, diazepine, digoxigenin, the hydrocortisone butyric ester, the metronidazole benzoate, tolbutamide, prostaglandin(PG), the fluorine cortisone, grisovin, miconazole nitrate, leukotriene B 4 inhibitors, Proprasylyte, theophylline, flurbiprofen, Sodium Benzoate, phenylformic acid, riboflavin, benzodiazepines, phenylethyl barbituric acid, U26452, Sulphadiazine Sodium, sulfaethidole, diclofenac sodium, Phenytoin Sodium Salt (phenyroin), the hioridazine hydrochloride, bropyrimie, hydrochlorothiazide, fluconazole etc.

Above drugs of low aqueous solubility can join in the segmented copolymer with the w/w ratio of 0.1-20.0:80.0-99.9, and it suitably is included in the micellar kernel that is formed by amphiphilic block copolymer of the present invention.

In another embodiment, the invention provides the amphiphilic block copolymer of the hydrophobic B block that contains hydrophilic A block and have terminal hydroxyl and the polymeric composition of polylactic acid derivative, the terminal hydroxyl of wherein said hydrophobic B block is replaced by tocopherol or cholesterol group, terminal and at least one the carboxyl covalent attachment of at least one of polylactic acid derivative.

The amphiphilic block copolymer that contains hydrophilic A block and hydrophobic B block, wherein the hydroxyl terminal of hydrophobic block is replaced by hydrophobic tocopherol or cholesterol group, and it has excellent hydrophobic property, with described above the same.

One or more ends of polylactic acid derivative of the present invention and at least one carboxylic acid or carboxylate salt covalent attachment.The non-binding end of polylactic acid derivative of the present invention can be selected from hydroxyl, acetoxyl group, benzoyloxy group, last of the ten Heavenly stems acyloxy and functional group's covalent attachment of palm acyloxy.The function of carboxylic acid or carboxylate salt be pH be 4 or the higher aqueous solution in as hydrophilic radical, it can make poly(lactic acid) derive to be polymeric micelle wherein.When polylactic acid derivative of the present invention was dissolved in the aqueous solution, the hydrophilic and hydrophobic ingredient that exists in the polylactic acid derivative should be an equilibrated, so that form polymeric micelle.Therefore, the molecular-weight average of polylactic acid derivative of the present invention is preferably in 50 to 50,000 daltonian scopes.During preparation process, the molecular weight of polylactic acid derivative can be regulated by control reaction temperature, time and similar condition.

Polylactic acid derivative is preferably represented by following formula:

RO-CHZ-[A] _n-[B] _m-COOM (I)

Wherein A is-COO-CHZ-, B is-COO-CHY-,-COO-CH ₂CH ₂CH ₂CH ₂CH ₂-or-COO-CH ₂CH ₂OCH ₂, R is hydrogen atom or ethanoyl, benzoyl, decanoyl, palmitoyl, methyl or ethyl, and Z and Y all are hydrogen atom or methyl or phenyl, and M is H, Na, K or Li, and n is from 1 to 30 integer, m is from 0 to 20 integer.

One or more ends of polylactic acid derivative of the present invention and carboxyl or its an alkali metal salt (preferably its an alkali metal salt) covalent attachment.Metal ion in the alkali metal salt of polylactic acid derivative is a monovalent ion, for example sodium, potassium or lithium.Polylactic acid derivative in the metal ion salt form at room temperature is a solid, because its relative neutral pH, so it is highly stable.

More preferably, polylactic acid derivative is the material of following formula representative:

RO-CHZ-[COO-CHX] _p-[COO-CHY’] _q-COO-CHZ-COOM (II)

Wherein X is a methyl; Y ' is hydrogen atom or phenyl; P is from 0 to 25 integer; Q is from 0 to 25 integer, and condition is that p+q is from 5 to 25 integer; R, Z and M all are suc as formula (I) defined group.

In addition, the polylactic acid derivative of following formula (III) to (V) also is applicable to the present invention:

RO-PAD-COO-W-M’ (III)

Wherein W-M ' is Or

PAD is selected from multipolymer, the D of D.L-poly(lactic acid), D-poly(lactic acid), poly-amygdalic acid, D.L-lactic acid and oxyacetic acid, the multipolymer and the D of the multipolymer of L-lactic acid and amygdalic acid, D.L-lactic acid and caprolactone, L-lactic acid and 1, the multipolymer of 4-diox-2-ketone, R and M all are the groups suc as formula (I) definition.

S-O-PAD-COO-Q (IV)

Wherein S is

, L is-NR ₁-or-O-, R ₁Be hydrogen atom or C _1-10Alkyl, Q are CH ₃, CH ₂CH ₃, CH ₂CH ₂CH ₃, CH ₂CH ₂CH ₂CH ₃Or CH ₂C ₆H ₅, a is from 0 to 4 integer, and b is from 1 to 10 integer, and M is the group suc as formula (I) definition, and PAD is the group suc as formula (III) definition.

Or

Wherein R ' is-PAD-O-C (O)-CH ₂CH ₂-C (O)-OM, M is the group suc as formula (I) definition, and PAD is that a is from 1 to 4 integer suc as formula the group of (III) definition, if a=1 for example, be exactly 3-arm (arm) PLA-COONa, if a=2 is exactly 4-arm PLA-COONa, if a=3, be exactly 5-arm PLA-COONa, if a=4 is exactly 6-arm PLA-COONa.

The initiator that is used for synthetic polymer (formula V) comprises glycerine, erythritol, threltol, tetramethylolmethane (pentaerytritol), Xylitol, ribitol, Sorbitol Powder and N.F,USP MANNITOL.

Polymeric composition of the present invention can comprise and accounts for amphiphilic block copolymer and the polylactic acid derivative gross weight is the amphiphilic block copolymer of 0.1 to 99.9 weight % and the polylactic acid derivative of 0.1 to 99.9 weight %.Preferably, polymeric composition of the present invention comprises the amphiphilic block copolymer of 20 to 95 weight % and the polylactic acid derivative of 5 to 80 weight %.More preferably, polymeric composition of the present invention comprises the amphiphilic block copolymer of 50 to 90 weight % and the polylactic acid derivative of 10 to 50 weight %.

Polylactic acid derivative of the present invention pH be 4 or the higher aqueous solution in can form micella separately, but polymeric composition can form micella in the aqueous solution, and needn't consider the pH value of solution.Since the biodegradability polymkeric substance usually pH be 10 or higher situation under hydrolysis, therefore, polymeric composition of the present invention can use in pH is 1 to 10 scope, preferred pH is in 4 to 8 scope.Can in 1 to 400nm scope, regulate by the micella of polymeric composition preparation of the present invention or the particle diameter of nanoparticle, preferred 5 to 200nm, this depends on the ratio of molecular weight and the polylactic acid derivative and the amphiphilic block copolymer of polymkeric substance.

Explanation as Fig. 1 to Fig. 3, polylactic acid derivative or amphiphilic block copolymer separately and their mixture can in the aqueous solution, form micella, the micella that is formed in the aqueous solution by the polymer composition of amphiphilic block copolymer and polylactic acid derivative demonstrates higher medicine load rate and the stability of micella that is formed separately by polylactic acid derivative or amphiphilic block copolymer than those.In the drawings, 1 represents drugs of low aqueous solubility, 10 represent mono methoxy polyethylene glycol-polylactide hydrophobic part (mPEG-PLA-hydrophobic part), 11 represent mono methoxy polyethylene glycol (mPEG), and 12 represent polylactide hydrophobic part (PLA-hydrophobic part), and 20 represent D, the sodium salt of L-poly-(lactic acid), 21 represent D, the L-poly(lactic acid), and 22 represent carboxylic acid sodium.Yet, to compare with the material that forms separately by polylactic acid derivative or amphiphilic block copolymer, polymeric composition of the present invention has significantly improved carrying drug ratio and formed micellar stability in the aqueous solution.

In one embodiment of the invention, the polymeric composition that contains amphiphilic block copolymer and polylactic acid derivative is provided, wherein amphiphilic block polymer contains hydrophilic A block and the hydrophobic B block with terminal hydroxyl, wherein said terminal hydroxyl is replaced by hydrophobic tocopherol or cholesterol group, at least one of polylactic acid derivative terminal with at least one carboxyl covalent attachment, wherein said carboxyl by two or trivalent metal ion fix.

The polymeric composition of having fixed metal ion can prepare by adding two or trivalent metal ion in the polymeric composition of amphiphilic block copolymer and polylactic acid derivative.Polymeric micelle or nanoparticle can by change be used to connect or fixedly the add-on of two or trivalent metal ion of the carboxyl terminal of polylactic acid derivative form.

Two or trivalent metal ion in preferred following: Ca ²⁺, Mg ²⁺, Ba ²⁺, Cr ³⁺, Fe ³⁺, Mn ²⁺, Ni ²⁺, Cu ²⁺, Zn ²⁺And Al ³⁺Two or trivalent metal ion can join in the polymeric composition of amphiphilic block copolymer and polylactic acid derivative preferred CaCl with the form of vitriol, muriate, carbonate, phosphoric acid salt or hydroxylate ₂, MgCl ₂, ZnCl ₂, AlCl ₃, FeCl ₃, CaCO ₃, MgCO ₃, Ca ₃(PO ₄) ₂, Mg ₃(PO ₄) ₂, AlPO ₄, MgSO ₄, Ca (OH) ₂, Mg (OH) ₂, Al (OH) ₃Or Zn (OH) ₂These forms.

As the explanation of Fig. 4 and Fig. 5, when the valent metal ion of polylactic acid derivative carboxyl terminal by two or trivalent metal ion replace when forming the metal ion key, micella of Xing Chenging or nanoparticle can have the stability that is enhanced thus.

Polymeric micelle or nanoparticle can prepare by the equivalent of metal ion that conversion adds.Specifically be that if divalent-metal ion is that 0.5 equivalent or lower amount add with the amount that is equivalent to the polylactic acid derivative carboxyl terminal, the metal ion that can form bonding with carboxyl terminal is exactly not enough so, so just can form polymeric micelle.If divalent-metal ion adds with 0.5 equivalent or higher amount, so the metal ion that can form bonding with the polylactic acid derivative carboxyl terminal just can be enough fixing micella firmly, so just formed nanoparticle.

In addition, the drug release rate of polymeric micelle or nanoparticle can be regulated by the equivalent of metal ion that change add.If metal ion is that 1 equivalent or littler amount exist to be equivalent to the polylactic acid derivative carboxyl terminal, be used for so will descending with polylactic acid derivative carboxyl terminal bonded quantity, release rate of drugs has also just improved like this.If metal ion exists with 1 equivalent or bigger amount, be used for so will increasing with polylactic acid derivative carboxyl terminal bonded quantity, therefore release rate of drugs has also just reduced like this.So, in order to improve the drug release rate in the blood, just use less normal metal ion, in order to reduce release rate of drugs, just use bigger normal metal ion.

The polymer composition of the present invention of having fixed metal ion can comprise the amphiphilic block copolymer of 5 to 95 weight %, the polylactic acid derivative of 5 to 95 weight % is 0.01 to 10 normal two or trivalent metal ion with respect to the equivalents of polylactic acid derivative carboxyl terminal.Preferably, they comprise the amphiphilic block copolymer of 20 to 80 weight %, the polylactic acid derivative of 20 to 80 weight %, 0.1 to 5 normal two or trivalent metal ion.More preferably, they comprise the amphiphilic block copolymer of 20 to 60 weight %, the polylactic acid derivative of 40 to 80 weight %, 0.2 to 2 normal two or trivalent metal ion.

This polymeric composition that contains the polymeric composition of amphiphilic block copolymer and polylactic acid derivative and fixed metal ion can form stable polymeric micelle or nanoparticle in the aqueous solution, wherein amphiphilic block copolymer contains hydrophilic block and hydroxyl terminal wherein and is had hydrophobic block, the end of polylactic acid derivative and at least one the carboxyl covalent attachment that good hydrophobic tocopherol or cholesterol group replace.Therefore, the invention still further relates to and comprise the polymeric micelle that forms by polymeric composition of the present invention with by load drugs of low aqueous solubility therein or the pharmaceutical composition of nanoparticle.After the administration, the active drug concentration of above-mentioned composition in blood has the residence time of prolongation.Pharmaceutical composition of the present invention has improved the plasma concentration of dewatering medicament, and can be used in the different pharmaceutical preparations.

Shown in Fig. 3 to 5, drugs of low aqueous solubility mixes the polymeric micelle that wherein comprises medicine to be formed on the polymeric composition of amphiphilic block copolymer and polylactic acid derivative.Can add two or trivalent metal ion come to form the metal ion key with the carboxyl terminal of polylactic acid derivative, formed stability-enhanced pastille polymeric micelle and nanoparticle thus.

The content of drugs of low aqueous solubility preferably accounts for the scope that the pharmaceutical composition gross weight is 0.1 to 30 weight %, and wherein composition contains amphiphilic block copolymer, polylactic acid derivative and hydrophobic drug.The size of pastille polymeric micelle or nanoparticle can be adjusted to 400nm from 5, and preferred 10 to 200nm, this depends on the molecular weight of polymkeric substance and the ratio of amphiphilic block copolymer and polylactic acid derivative.For instance, fixed the polymeric micelle of metal ion or the particle of nanoparticle and had the median size of 20-40nm, as shown in table 7.The micella of this magnitude range is applicable to injection formulations and sterilising filtration.

Polymeric composition or polymeric micelle or the nanoparticle of having fixed metal ion according to the present invention handled with nonmetallic ion have advantages of higher stability, and especially, the material of having fixed metal ion has higher stability in the aqueous solution.As shown in table 9, pastille polymeric micelle compositions (composition 4﹠amp; 5) dynamic stabilization has been fixed the kinetics of the polymeric micelle compositions that contains taxol of metal ion even more stable.The adding metal ion can significantly improve the residence time of polymeric micelle Chinese traditional medicine of the present invention.This is attributed to the carboxylate anion's of the polylactic acid derivative that may induce the raising of hydrophobic core rigidity crosslinked electrostatic interaction.

In addition, the amphiphilic block copolymer polymeric micelle (composition 4) of having fixed metal ion has the more stable kinetics than initial mPEG-PLA-OH (composition 7), and wherein amphiphilic block copolymer has the hydrophobic part (tocofecol succinic acid) that is used to replace hydrophobic B block hydroxyl terminal.This result has hinted that the hydrophobicity that therefore increases hydrophobic B block in the amphipathic copolymer just can form more stable micella owing to more intensive interaction between the hydrophobic part of amphipathic copolymer and the medicine.

As table 11 and shown in Figure 9, the amphiphilic Synthetic rubber, isoprene-styrene, hydrogenated, block, diblock polymeric micelle (composition 8) (wherein amphiphilic block copolymer has the hydrophobic part (tocofecol succinic acid) that is used to replace hydrophobic B block hydroxyl terminal) of having fixed metal ion had than the longer blood resides time of initial amphiphilic Synthetic rubber, isoprene-styrene, hydrogenated, block, diblock polymeric micelle (composition 9) of having fixed metal ion.This result has hinted that also as described in embodiment 36, because more intensive interaction between the hydrophobic part of amphipathic copolymer and the medicine, the hydrophobicity that therefore increases hydrophobic B block in the amphipathic copolymer just can form more stable micella.

Shown in Figure 10-13, its Chinese traditional medicine is by the composition of load in the polymeric composition of having fixed metal ion, and medicine wherein has the longer residence time in blood, therefore the Plasma Concentration that can remain valid for more time than present commercial preparation.

The present invention also provides the pharmaceutical composition as carcinostatic agent.In a preferred embodiment, pharmaceutical composition as carcinostatic agent is provided, amphiphilic block copolymer and polylactic acid derivative and anticarcinogen that it contains hydrophilic A block and has the hydrophobic B block of terminal hydroxyl, the terminal hydroxyl of wherein said hydrophobic B block is replaced by tocopherol or cholesterol group, terminal and at least one the carboxyl covalent attachment of at least one of polylactic acid derivative.The carboxyl terminal of polylactic acid derivative can be further by two or trivalent metal ion fix.

The example of anticarcinogen includes but not limited to that Taxan, taxines or taxanes are as taxol and Japanese yew terpene; Phodophyllotoxins; Camptothecin is as camptothecine, 9-nitrocamptothecin, 9-aminocamptothecin, camptothecine-11, topodecane; The anthracene nucleus class is as Zorubicin, epirubicin, aclarubicin, idarubicin, pyrarubicin; Vinca alkaloids is as vincristine(VCR), vinorelbine, vindesine, vintriptol, vinsaltine; Epostane, platinum, Etoposide, methotrexate, carmustine, 5 FU 5 fluorouracil, vitamin A acid, Vogan-Neu, tamoxifen, Mitomycin B, ametycin, amonafide, agaricol etc.

The polymeric micelle pharmaceutical composition that obtains has the much pharmacy effects of improvement.As a specific embodiment, shown in Figure 14 to 21, comprise taxol and fixed Ca ²⁺Polymeric micelle growth of cancers is had high inhibitory, and also suppress growth (the Tu22 ﹠amp of the cancer cells of tolerance anticarcinogen; 23).

(being taxol), Zorubicin etc. extensively are used in the chemotherapy of cancer.These anticarcinogens are effectively also useful in chemotherapy, but anticancer chemical sproof development always makes that these medicines are invalid in the cancer cells.Various anti-

The mechanism of medicine comprises the overexpression of P-glycoprotein (P-gp) and the distortion of 'beta '-tubulin, by characterization.At this wherein, the overexpression of P-gp has become main mechanism in order to explain multidrug resistant, comprises

The resistance phenomenon.The cancer cells of tolerance anticarcinogen has demonstrated higher IC than normal cell ₅₀(the repressed drug level of 50% cell) therefore uses the chemotherapy of anticarcinogen just to require that higher drug level is arranged in tumour cell.Therefore, the concrete location of tumor tissues Chinese traditional medicine is in order to be effective necessary.As shown in figure 10, the polymeric micelle of having fixed metal ion has longer cycling time than conventional formulation.Like this, compare with conventional formulation, it just can more optionally be assembled in tumor tissues by strengthening infiltration and being detained (EPR) effect.For the polymeric micelle of the having fixed metal ion antagonism effect to the cancer of tolerance anticarcinogen is described, we have established the opposing tolerance

The active animal model of vivo antitumor of cancer.When being inoculated into cancer cells in the mouse when being exposed in the taxol repeatedly, use

The IC of the medicine of pre-treatment cancer cells ₅₀Significantly increase than the medicine that is used for the body cancer cells.In this animal model, shown in Figure 22 and table 22, group with the polymeric micelle of having fixed metal ion (composition 10) treatment demonstrates than the higher inhibiting rate of group with polyoxyethylenated castor oil preparation (composition 11) treatment, and this may be long because be incorporated into the residence time of the concentration medicine in the polymeric micelle of having fixed metal ion.In the animal model for cancer of anti-Zorubicin, can obtain same effect (Figure 23).

Therefore, the invention provides the pharmaceutical composition that is used for the treatment of the resistance cancer, amphiphilic block copolymer and polylactic acid derivative and anticarcinogen that it contains hydrophilic A block and has the hydrophobic B block of terminal hydroxyl, the terminal hydroxyl of wherein said hydrophobic B block is replaced by tocopherol or cholesterol group, terminal and at least one the carboxyl covalent attachment of at least one of polylactic acid derivative.The carboxyl terminal of the polylactic acid derivative of above composition also can by two or trivalent metal ion fix.

In addition, the present invention includes the method for the above pharmaceutical composition of preparation.Particularly, shown in Fig. 3 and 5, the polylactic acid derivative of specified proportion, amphiphilic block copolymer and drugs of low aqueous solubility can be dissolved in one or more solvent that is selected from acetone, ethanol, methyl alcohol, ethyl acetate, acetonitrile, methylene dichloride, chloroform, acetic acid He diox.Therefrom remove organic solvent so that the uniform mixture of preparation drugs of low aqueous solubility and polymkeric substance.The uniform mixture of drugs of low aqueous solubility and polymeric composition of the present invention can join pH and in 4 to 8 the aqueous solution, obtain being mixed with the polymeric micelle aqueous solution that comprises drugs of low aqueous solubility under 0 to 80 ℃.The freeze-drying above pastille polymeric micelle aqueous solution is so that the polymeric micelle compositions of preparation solid form then.

To comprise 0.001 to 2M two or the aqueous solution of trivalent metal ion join and be mixed with in the polymeric micelle aqueous solution that comprises drugs of low aqueous solubility so that form the polymeric composition of having fixed metal ion.Slowly stirred this mixture 0.1 to 1 hour under the room temperature, freeze-drying is so that the polymeric micelle of having fixed metal ion or the nanoparticle of preparation solid form then.

Load and the polymeric micelle of the present invention or the nanoparticle that have dissolved drugs of low aqueous solubility can give by oral or parenteral route.When micella was degraded, medicine just discharged from the micellar hydrophobic core so that show pharmacological action.Especially, polymeric micelle or the nanoparticle of having fixed metal ion have kept the long period in blood, and assemble at the target injury region.

Concerning parenteral administration, polymeric micelle or nanoparticle can be by giving in intravenously, intramuscular, intraperitoneal, intranasal, internal rectum, intraocular or the lung.Concerning oral administration, medicine is mixed with polymeric micelle of the present invention, the form with tablet, capsule solution gives then.

The polymeric micelle that uses among the present invention or the dosage of nanoparticle can for example patient's symptom, age and body weight and conditions of similarity change in the scope of a broadness according to various conditions.

Following examples those skilled in the art will more clearly be understood be how to realize of the present invention.Should be appreciated that present invention is described though combine preferred specific embodiments, following content is not construed as limiting scope of the present invention.Others of the present invention will be conspicuous concerning belonging to those skilled in the art of the present invention.

Preparation embodiment 1

D, synthetic 1 of L-poly(lactic acid) (PLA-COOH)

With 100 gram D, the L-poly(lactic acid) joins in the 250ml three neck round-bottomed flasks.Flask has agitator, is heated to 80 ℃ in oil bath.Be reflected to be reduced under the pressure of 25mmHg and carry out 1 hour to remove redundant moisture by vacuum ejector.Reacting on 150 ℃ then carried out 6 hours under 25mmHg decompression pressure.The product that obtains is joined in 1 liter of distilled water with precipitation polymers, then sedimentary polymkeric substance is joined in the distilled water with remove deenergize be dissolved in pH be 4 or the lower aqueous solution in low-molecular weight polymer.Then sedimentary polymkeric substance is joined in 1 liter of distilled water, by the pH regulator to 6 of the aqueous solution being come dissolve polymer to 8 to wherein little by little adding sodium bicarbonate.By centrifugal or filtering separation and remove insoluble polymer.The 1N hydrochloric acid soln is added drop-wise to wherein, and in the aqueous solution precipitation polymers.With the sedimentary polymkeric substance twice of distilled water wash, decompression separation and drying obtain full-bodied liquid (78g D, L-poly(lactic acid), productive rate: 78%). ¹The polymkeric substance number-average molecular weight that the H-NMR spectrum records is 540 dalton.

Preparation embodiment 2 to 4

D, the Synthetic 2 of L-poly(lactic acid) (PLA-COOH)

Except that the temperature of reaction that provides according to table 1, pressure and time are controlled, all the other according to preparation among the embodiment 1 identical step obtain D, L-poly(lactic acid).By above preparation embodiment 1 to 4 synthetic D, the number-average molecular weight of L-poly(lactic acid) and productive rate are listed in the table below in 1.

Table 1

Preparation embodiment	Temperature (℃)	Time (hour)	Pressure (mmHg)	Mn	Productive rate (%)
						1	150	6	25	540	78
2	160	12	10	1140	83
						3	160	24	10	1550	84
4	160	24	5	2100	87

* productive rate=(monomer of resulting polymers/use) * 100

Preparation embodiment 5

D, synthetic 1 of L-lactic acid and ethanol copolymer (PLGA-COOH)

With 55 (55) gram D, L-lactic acid (0.6 mole) and 45 gram oxyacetic acids (0.6 mole) together join in the 250ml three neck round-bottomed flasks.Under being reflected at 150 ℃ temperature and under the decompression pressure of 10mmHg, carry out 12 hours, according to preparation embodiment 1 in identical step carry out.

Preparation embodiment 6

D, the Synthetic 2 of L-lactic acid and ethanol copolymer (PLGA-COOH)

With 73 (73) gram D, L-lactic acid (0.8 mole) and 27 gram oxyacetic acids (0.35 mole) together join in the 250ml three neck round-bottomed flasks.Under being reflected at 160 ℃ temperature and under the decompression pressure of 10mmHg, carry out 12 hours, according to preparation embodiment 1 in identical step carry out.

Preparation embodiment 7

D, synthetic 3 of L-lactic acid and ethanol copolymer (PLGA-COOH)

With 91 (91) gram D, L-lactic acid (1.0 moles) and 9 gram oxyacetic acids (0.12 mole) together join in the 250ml three neck round-bottomed flasks.Under being reflected at 160 ℃ temperature and under the decompression pressure of 10mmHg, carry out 12 hours, according to preparation embodiment 1 in identical step carry out.

Preparation embodiment 8

D, synthetic 4 of L-lactic acid and ethanol copolymer (PLGA-COOH)

With 73 (73) gram D, L-lactic acid (0.8 mole) and 27 gram oxyacetic acids (0.35 mole) together join in the 250ml three neck round-bottomed flasks.Under being reflected at 180 ℃ temperature and under the decompression pressure of 5mmHg, carry out 24 hours, according to preparation embodiment 1 in identical step carry out.

Above preparation embodiment 5 to 8 synthetic multipolymers have been shown in the table 2.

Table 2

Preparation embodiment 9

D, L-lactic acid and amygdalic acid multipolymer (PLMA-COOH) synthetic

With 75 (75) gram D, L-lactic acid (0.83 mole) and 25 gram D, L-amygdalic acid (0.16 mole) together joins in the 250ml three neck round-bottomed flasks.Under being reflected at 180 ℃ temperature and under 10 to 20mmHg decompression pressure, carry out 5 hours, according to preparation embodiment 1 in identical step carry out.Obtain 54 (54) gram (productive rates: D 54%), the multipolymer of L-lactic acid and amygdalic acid.D, the mol ratio of L-lactic acid and amygdalic acid is 85/15.Warp ¹The polymkeric substance number-average molecular weight that the H-NMR spectrum records is 1,096 dalton.

Preparation embodiment 10

Acetyl oxygen D, L-polylactic acid derivative (AcO-PLA-COOH) synthetic

By preparation embodiment 2 synthetic D, L-poly(lactic acid) (Mn:1,140 dalton) and 20ml Mono Chloro Acetic Acid together join in the 250ml three neck round-bottomed flasks with 50 (50) grams.Flask has condenser, and reaction mixture refluxed in nitrogen gas stream 4 hours.Unnecessary Mono Chloro Acetic Acid is removed in distillation, and reaction product is joined in the mixture of ice and water.Slowly stir whole mixture so that precipitation polymers.The polymkeric substance of precipitation separation with distilled water wash twice, is dissolved in the anhydrous propanone then.To wherein adding anhydrous magnesium sulfate to remove unnecessary moisture content.The product that filtration obtains is to remove sal epsom.Utilize vacuum aspirator to remove acetone, thereby obtain liquid acetyl oxygen D, L-poly(lactic acid) (46g, productive rate: 92%).Warp ¹H-NMR determines the unimodal at the 2.02ppm place of acetoxyl group group.

Preparation embodiment 11

Palm acyl-oxygen D, L-polylactic acid derivative (PalmO-PLA-COOH) synthetic

By preparation embodiment 2 synthetic D, the L-poly(lactic acid) (Mn:1,140 dalton) join in the 250ml three neck round-bottomed flasks with 20 (20) grams.Reactant is dehydration fully under vacuum in 120 ℃ oil bath.Oil bath is cooled to 50 ℃, to wherein adding 50ml acetone with complete dissolve polymer.To wherein adding five (5) ml chlorine palmitinic acids, under 50 ℃ temperature, in nitrogen, reacted 10 hours.With excessive hexane wash reaction product to remove remaining reactants.Then product is dissolved in the acetone, this solution is joined in the mixture of ice and water.Slowly stir whole mixture and obtain the oligomer precipitation.Separate this oligomer and use distilled water wash twice, be dissolved in the anhydrous propanone then.In this solution, add anhydrous magnesium sulfate to remove unnecessary moisture content.Filter products therefrom to remove sal epsom.Remove acetone with vacuum aspirator, thereby obtain palm acyl-oxygen D, L-polylactic acid derivative (19.1g, productive rate: 96%).Warp ¹The peak that H-NMR determines palmitoyl is 0.88,1.3 and the 2.38ppm place.

Preparation embodiment 12

Synthesizing of 3 arm poly(lactic acid) (3 arm PLA-COOH)

One (1) gram glycerine (0.011mol) is joined in the 100ml three neck round-bottomed flasks.Flask has agitator, is heated to 80 ℃ in oil bath.Be reflected to be reduced under the pressure of 25mmHg and carry out 30 minutes to remove unnecessary moisture content by vacuum aspirator.The catalysts stannous octoate (Tin (Oct) 2) that is dissolved in the toluene is joined in the glycerine.Stirred reaction mixture 30 minutes, at 110 ℃ pressure is reduced to 1mmHg and carry out 1 hour solvent (toluene) to remove catalyst-solvent, to the rac-Lactide (35.8g that wherein adds purifying, 0.249mol, 10 weight %), mixture heating up to 130 ℃ was carried out 6 hours and under the decompression pressure of 25mmHg.With the polymer dissolution that forms in acetone, dropwise to wherein adding 0.2N NaHCO ₃The aqueous solution is with precipitation polymers.With the sedimentary polymkeric substance three of distilled water wash or four times, decompression separates down and drying obtains powder (3 arm PLA-OH).

100 (100) grams, 3 arm PLA-OH (0.033mol) are joined in the single neck round-bottomed flask of 100ml.Be reflected to be reduced under the pressure of 25mmHg and carry out 30 minutes to remove unnecessary moisture content by vacuum aspirator.To wherein adding 19.8 gram succinyl oxides (0.198mol), and mixture heating up to 125 ℃ was carried out 6 hours.With the polymer dissolution that forms in acetone, to wherein dropwise adding distilled water with precipitation polymers.Should sedimentary polymer dissolution under 60 ℃ at 0.2N NaHCO ₃In the aqueous solution.Remove by filter insoluble polymkeric substance.To wherein dropwise adding the 2N HCL aqueous solution with precipitation polymers.With the sedimentary polymkeric substance five of distilled water wash or six times, decompression separates down and drying obtains powder (3 arm PLA-COOH).Warp ¹The H-NMR spectrum determines that the number-average molecular weight of polymkeric substance is 3,000 dalton.

Preparation embodiment 13

Synthesizing of 5 arm poly(lactic acid) (5 arm PLA-COOH)

One (1) gram Xylitol (0.0066mol) is joined in the 100ml three neck round-bottomed flasks.Flask has agitator, is heated to 80 ℃ in oil bath.Be reflected to be reduced under the pressure of 25mmHg and carry out 30 minutes to remove unnecessary moisture content by vacuum aspirator.The catalysts stannous octoate (Tin (Oct) 2) that is dissolved in the toluene is joined in the glycerine.Stirred reaction mixture 30 minutes is reduced to 1mmHg at 110 ℃ with pressure and is carried out 1 hour to remove the solvent (toluene) that has dissolved catalyzer.To the rac-Lactide that wherein adds purifying (31.7g, 0.151mol, 10 weight %), and under the decompression pressure of 25mmHg, mixture heating up to 130 ℃ was carried out 6 hours.With the polymer dissolution that forms in acetone, dropwise to wherein adding 0.2N NaHCO ₃The aqueous solution is with precipitation polymers.With the sedimentary polymkeric substance three of distilled water wash or four times, decompression separates down and drying obtains powder (5 arm PLA-OH).

100 (100) grams, 5 arm PLA-OH (0.033mol) are joined in the single neck round-bottomed flask of 100ml.Be reflected to be reduced under the pressure of 25mmHg and carry out 30 minutes to remove unnecessary moisture content by vacuum aspirator.To wherein adding 33 (33.0) gram succinyl oxides (0.33mol), and mixture heating up to 125 ℃ was carried out 6 hours.With the polymer dissolution that forms in acetone, to wherein dropwise adding distilled water with precipitation polymers.Should sedimentary polymer dissolution under 60 ℃ at 0.2N NaHCO ₃In the aqueous solution.Remove by filter insoluble polymkeric substance.To wherein dropwise adding the 2N HCL aqueous solution with precipitation polymers.With the sedimentary polymkeric substance five of distilled water wash or six times, decompression separates down and drying obtains powder (3 arm PLA-COOH).Warp ¹The H-NMR spectrum determines that the number-average molecular weight of polymkeric substance is 3,000 dalton.

Preparation embodiment 14

Synthetic 1 of poly(lactic acid) sodium salt (PLA-COONa)

Will be by preparation embodiment 1 synthetic D, the L-poly(lactic acid) (Mn:540 dalton) be dissolved in the acetone.This solution is joined in the round-bottomed flask, and flask has agitator.Slowly stir this solution under the room temperature, make pH reach 7 to wherein slowly adding sodium hydrogen carbonate solution (1N).To wherein adding anhydrous sodium sulphate, therefrom remove unnecessary moisture content.Filter the gained mixture, and evaporate acetone with the solvent evaporation device.Therefrom obtain white solid.This solid is dissolved in the anhydrous propanone, and filtering solution is to remove insoluble part.Evaporation acetone has been left white solid D, L-poly(lactic acid) sodium salt (productive rate: 96%).As shown in Figure 2, warp ¹H-NMR observes the hydrogen peak of contiguous carboxyl at the 4.88ppm place, and when polymer dissolution was in water, its pH was 6.5 to 7.5.

Preparation embodiment 15

The Synthetic 2 of poly(lactic acid) sodium salt (PLA-COONa)

Remove to use 2 synthetic D, outside L-poly(lactic acid) (Mn:1,140 dalton) and the aqueous sodium carbonate, according to the step synthesizing polylactic acid sodium salt (productive rate 95%) identical with preparing embodiment 14 by preparation embodiment.

Preparation embodiment 16

Acetyl oxygen D, L-poly(lactic acid) sodium salt (AcO-PLA-COONa) synthetic

Remove to use 10 synthetic acetyl oxygen-D, outside L-poly(lactic acid) (Mn:1,140 dalton) and the aqueous sodium carbonate, according to the step synthesis of acetyl oxygen-D identical, L-poly(lactic acid) sodium salt (productive rate 95%) with preparing embodiment 14 by preparation embodiment.

Preparation embodiment 17

Palm acyl-oxygen D, synthetic 1 of L-poly(lactic acid) sodium salt (PalmO-PLA-COONa)

Will be by preparation embodiment 11 synthetic palm acyl-oxygen D, L-poly(lactic acid) (Mn:1,140 dalton) is dissolved in (28.6v/v%) in the aqueous acetone solution fully.This solution is joined in the round-bottomed flask, and flask has agitator.Slowly stir this solution under the room temperature, then to wherein adding sodium bicarbonate aqueous solution (1N) in order to neutralization.Slowly stir this solution under the room temperature, and make pH reach 7 to wherein slowly adding sodium bicarbonate aqueous solution (1N).To wherein adding anhydrous magnesium sulfate to remove unnecessary moisture content.Filter gained solution, with solvent evaporation device evaporation acetone soln.Therefrom obtain white solid.This solid is dissolved in the acetone, and filtering solution is to remove any insoluble particle.Evaporation acetone obtains white solid palm acyl-oxygen D, L-poly(lactic acid) sodium salt (productive rate: 96%).

Preparation embodiment 18

Synthesizing of poly(lactic acid) sylvite (PLA-COOK)

Remove to use 3 synthetic D, outside L-lactic acid (Mn:1,550 dalton) and the potassium bicarbonate aqueous solution, according to the step synthesizing polylactic acid sylvite (productive rate 98%) identical with preparing embodiment 14 by preparation embodiment.

Preparation embodiment 19

Synthetic 3 of poly(lactic acid) sodium salt (PLA-COONa)

Remove to use 4 synthetic D, outside the L-lactic acid (Mn:2,100 dalton), according to the step synthesizing polylactic acid sodium salt (productive rate 95%) identical with preparation embodiment 14 by preparation embodiment.

Preparation embodiment 20

D, synthetic 1 of L-lactic acid and ethanol copolymer sodium salt (PLGA-COONa)

Remove to use 5 synthetic D, outside L-lactic acid and ethanol copolymer (Mn:920 dalton) and the aqueous sodium carbonate, synthesize D, the sodium salt of L-lactic acid and ethanol copolymer (productive rate 98%) according to the step identical with preparation embodiment 14 by preparation embodiment.

Preparation embodiment 21

D, the Synthetic 2 of L-lactic acid and ethanol copolymer sodium salt (PLGA-COONa)

Remove to use 6 synthetic D, outside L-lactic acid and the ethanol copolymer (Mn:1,040 dalton), according to the synthetic D of step identical, the sodium salt of L-lactic acid and ethanol copolymer (productive rate 93%) with preparation embodiment 14 by preparation embodiment.

Preparation embodiment 22

D, L-lactic acid and ethanol copolymer sylvite (PLGA-COOK) synthetic

Remove to use 7 synthetic D, outside L-lactic acid and ethanol copolymer (Mn:1,180 dalton) and the wet chemical, synthesize D, the sylvite of L-lactic acid and ethanol copolymer (productive rate 92%) according to the step identical with preparation embodiment 14 by preparation embodiment.

Preparation embodiment 23

D, synthetic 3 of L-lactic acid and ethanol copolymer sodium salt (PLGA-COONa)

Remove to use 8 synthetic D, outside L-lactic acid and the ethanol copolymer (Mn:1,650 dalton), according to the synthetic D of step identical, the sodium salt of L-lactic acid and ethanol copolymer (productive rate 98%) with preparation embodiment 14 by preparation embodiment.

Preparation embodiment 24

D, L-lactic acid and amygdalic acid multipolymer sodium salt (PLMA-COONa) synthetic

Remove to use 9 synthetic D, outside L-lactic acid and the amygdalic acid multipolymer (Mn:1,096 dalton), according to the synthetic solid D that is white in color of step identical, the sodium salt of L-lactic acid and amygdalic acid multipolymer (productive rate 96%) with preparation embodiment 14 by preparation embodiment.

Preparation embodiment 25

Synthesizing of 3 arm poly(lactic acid) sodium salts (3 arm PLA-COONa)

Remove to use by preparation embodiment 12 synthetic 3 arm D, outside the L-lactic acid copolymer (Mn:3,000 dalton), according to the synthetic solid 3 arm poly(lactic acid) sodium salts that are white in color of step identical with preparation embodiment 14.

Preparation embodiment 26

Synthesizing of 5 arm poly(lactic acid) sodium salts (5 arm PLA-COONa)

Remove to use 13 synthetic 5-arm D, outside the L-lactic acid copolymer (Mn:3,000 dalton), according to the synthetic solid 5 arm poly(lactic acid) sodium salts that are white in color of step identical with preparation embodiment 14 by preparation embodiment.

Table 3 has shown the carboxylate salt by above preparation embodiment 14 to 26 synthetic polylactic acid derivatives.

Table 3

Preparation embodiment	Reactant (Mn)	Alkali	Product	Mn (dalton)	Productive rate (%)
						14	PLA-COOH(540)	NaHCO 3	PLA-COONa	540	96
15	PLA-COOH (1,140)	Na 2CO 3	PLA-COONa	1,140	95
						16	AcO-PLA-COOH(1,140)	Na 2CO 3	AcO-PLA-COONa	1,140	95
17	Palmityl O-PLA-COOH (1,140)	NaHCO 3	Palmityl O-PLA-COONa	1,140	96
						18	PLA-COOH (1,550)	KHCO 3	PLA-COOK	1,550	98
19	PLA-COOH (2,100)	NaHCO 3	PLA-COONa	2,100	95
						20	PLGA-COOH(920)	Na 2CO 3	PLGA-COONa	920	98
21	PLGA-COOH(1,140)	NaHCO 3	PLGA-COONa	1,040	93
						22	PLGA-COOH(1,180)	K 2CO 3	PLGA-COOK	1,180	92
23	PLGA-COOH(1,650)	NaHCO 3	PLGA-COONa	1,650	98
						24	PLMA-COOH(1,096)	NaHCO 3	PLMA-COONa	1,096	96
25	3 arm PLA-COOH (3,000)	NaHCO 3	3 arm PLA-COONa	3,000	98
						26	5 arm PLA-COOH (3,000)	NaHCO 3	5 arm PLA-COONa	3,000	98

Preparation embodiment 27

The polymerization of mPEG-PLGA (mPEG-PLA) segmented copolymer (AB type)

Five (5) gram mono methoxy polyethylene glycols (Mn:2,000 dalton) are joined in the 100ml two neck round-bottomed flasks, and decompression following (1mmHg) is heated to 100 ℃ and carries out making in 2 to 3 hours the mixture dehydration.In reaction flask, fill drying nitrogen, use syringe catalysts stannous octoate (Sn (Oct) ₂) be injected in the rac-Lactide of 0.1 weight % (5mg).Stirred reaction mixture 30 minutes carries out 1 hour to remove the solvent (toluene) that has dissolved catalyzer at 110 ℃ of decline low pressures to 1mmHg.To the rac-Lactide that wherein adds purifying (5g), and mixture heating up to 130 ℃ carried out 12 hours.With the polymer dissolution that forms in ethanol, to wherein adding ether with precipitation polymers.The dry mixture that obtains is 48 hours in vacuum oven.The number-average molecular weight of gained mPEG-PLA is 2,000-1,765 dalton, warp ¹H-NMR is defined as the AB type.

Preparation embodiment 28

Mono methoxy polyethylene glycol-poly-(lactic acid-altogether-glycollide) (mPEG-PLGA) polymerization of segmented copolymer (AB type)

For synthetic mPEG-PLGA segmented copolymer, the step according to identical with preparation embodiment 27 exists under the stannous situation of octoate catalyst, in 120 ℃ with mono methoxy polyethylene glycol (Mn:5,000 dalton) and rac-Lactide and glycollide reaction 12 hours.The number-average molecular weight of gained mPEG-PLGA is 5,000-4,000 dalton, warp ¹H-NMR is defined as the AB type.

Preparation embodiment 29

Mono methoxy polyethylene glycol-poly-(lactic acid-altogether-p-diox-2-ketone) (mPEG-PLDO) polymerization of segmented copolymer (AB type)

For synthetic mPEG-PLDO segmented copolymer, according to the identical step of preparation embodiment 27, exist under the stannous situation of octoate catalyst, in 120 ℃ with mono methoxy polyethylene glycol (Mn:12,000 dalton) and rac-Lactide and p-diox-2-reactive ketone 12 hours.The number-average molecular weight of gained mPEG-PLDO is 12,000-10,000 dalton, warp ¹H-NMR is defined as the AB type.

Preparation embodiment 30

The polymerization of mono methoxy polyethylene glycol-polycaprolactone (mPEG-PCL) segmented copolymer (AB type)

For synthetic mPEG-PCL segmented copolymer, the step according to identical with preparation embodiment 27 exists under the stannous situation of octoate catalyst, in 130 ℃ with mono methoxy polyethylene glycol (Mn:12,000 dalton) and caprolactone reaction 12 hours.The number-average molecular weight of gained mPEG-PCL is 12,000-5,000 dalton, warp ¹H-NMR is defined as the AB type.

Following table 4 has shown by above preparation embodiment 27 to 30 synthetic block polymers.

Table 4

Preparation embodiment	Amphiphilic block copolymer	Mn (dalton)	Productive rate (%)
				27	mPEG-PLA	2,000-1,765	86
28	mPEG-PLGA	5,000-4,000	90
				29	mPEG-PLDO	12,000-10,000	78
30	mPEG-PCL	12,000-5,000	93

Preparation embodiment 31

The polymerization of mono methoxy polyethylene glycol-mono methoxy polyethylene glycol (PLA-mPEG-PLA) segmented copolymer (BAB type)

Remove to use 25g methoxy poly (ethylene glycol) (MW=2,000) and 50g D, outside the L-rac-Lactide, obtain PLA-mPEG-PLA according to the step identical with preparing embodiment 27.The number-average molecular weight of gained PLA-mPEG-PLA is 1,765-2,000-1,765 dalton, warp ¹H-NMR is defined as the BAB type.

Embodiment 1

The polymerization of mPEG-PLA-cholesterol

A) The synthetic cholesterol succinate

In round-bottomed flask, 7.6 gram cholesterol and 2.36 gram succinyl oxides are dissolved in 1 of 100ml, the 4-diox.To wherein adding 2.9 gram catalysts 4-(diformazan ammonia) pyridines (DMAP), stirred the mixture under the room temperature 24 hours.Reaction mixture is joined in the HCl solution with precipitation cholesterol succinate (9.1g; Productive rate=95%).

B) connect mPEG-PLA and cholesterol succinate

In round-bottomed flask, ten (10) grams are dissolved in the 50ml acetonitrile by preparation embodiment 27 synthetic mPEG-PLA and 1.55 gram (1.2 times of moles of polymkeric substance) cholesterol succinates.To wherein adding catalyzer 0.76 gram dicyclohexylcarbodiimide (DCC) and 0.045 gram 4-(diformazan ammonia) pyridine (DMAP), stirred this mixture 24 hours under the room temperature.When reaction is finished, use the glass filter filtering mixt to remove dicyclohexyl carbon urea (dicyclohexylcarbourea) by product.Extract to remove remaining catalyzer with aqueous hydrochloric acid.In the solution of this purified product, add sal epsom removing any residual moisture content, and this mixture joined contain n-hexane/ether (v/v=7/3) and be used for the cosolvent of recrystallization to obtain the mPEG-PLA-cholesterol (10g of purifying; Productive rate=88.6%).Fig. 6 has shown its NMR spectrum.

Embodiment 2

The polymerization of mPEG-PLA-cholesterol

A) The synthetic cholesterol succinate

7.6 gram cholesterol and succinyl dichlorides (cholesterol two moles) are joined in the flask, and be dissolved in the 50ml acetonitrile.Be reflected at and carry out 12 hours under 50 ℃, in the HCL aqueous solution, form precipitation then and obtain cholesterol succinate (8.2g the succinate group is connected on the hydroxyl of cholesterol; Productive rate=92%).

B) connect mPEG-PLA and cholesterol succinate

Except that use 10 gram mPEG-PLA and by embodiment 2a) the synthetic cholesterol succinate (1.2 times of moles of polymkeric substance), according to embodiment 1b) identical step obtains mPEG-PLA-cholesterol (9.52g: productive rate 85%).

Embodiment 3 to 5

The polyase 13 to 5 of mPEG-PLA-cholesterol

Except that the malonyl chloride (embodiment 3), glutaryl chlorine (embodiment 4) and the hexanedioyl chlorine (embodiment 5) that use respectively to two times of molar weights of polymkeric substance, obtain the mPEG-PLA-cholesterol according to the step identical with embodiment 2.

Embodiment 6 to 9

The polymerization 1 to 4 of mPEG-PLA-tocopherol

Except that using the 8.5g tocopherol and be respectively malonyl chloride (embodiment 6), succinyl dichloride (embodiment 7), glutaryl chlorine (embodiment 8) and the hexanedioyl chlorine (embodiment 9) of two times of molar weights of polymkeric substance, obtain the mPEG-PLA-tocopherol according to the step identical with embodiment 2.

Embodiment 10

The polymerization of mono methoxy polyethylene glycol-poly-(lactic acid-be total to-glycollide) tocopherol (mPEG-PLGA-tocopherol) segmented copolymer (AB type)

Except that using 10g by preparation embodiment 28 synthetic mPEG-PLGA and the 1.767 gram tocofecol succinic acid salt, according to embodiment 1b) identical step obtains the mPEG-PLGA-tocopherol (10g of purifying; Productive rate=87.5%).

Embodiment 11

The polymerization of mono methoxy polyethylene glycol-poly-(lactic acid-be total to-glycollide) cholesterol (mPEG-PLGA-cholesterol) segmented copolymer (AB type)

Except that using 10g by preparation embodiment 28 synthetic mPEG-PLGA and the 0.70 gram cholesterol succinate, according to embodiment 1b) identical step obtains the mPEG-PLGA-cholesterol (10g of purifying; Productive rate=88.6%).

Embodiment 12

The polymerization of mono methoxy polyethylene glycol-poly-(lactic acid-be total to-p-diox-2-ketone) tocopherol (mPEG-PLDO-tocopherol) segmented copolymer (AB type)

Except that using 10g by preparation embodiment 29 synthetic mPEG-PLDO and the 0.314 gram tocofecol succinic acid salt, according to embodiment 1b) identical step obtains the mPEG-PLDO-tocopherol (10g of purifying; Productive rate=87.5%).

Embodiment 13

The polymerization of mono methoxy polyethylene glycol-poly-(lactic acid-totally-dioxs-2-ketone) cholesterol (mPEG-PLDO-cholesterol) segmented copolymer (AB type)

Except that using 10g by preparation embodiment 29 synthetic mPEG-PLDO and the 0.288 gram cholesterol succinate, according to embodiment 1b) identical step obtains the mPEG-PLDO-cholesterol (10g of purifying; Productive rate=88.6%).

Embodiment 14

Mono methoxy polyethylene glycol-polycaprolactone tocopherol (mPEG-PCL-tocopherol) block

The polymerization of multipolymer (AB type)

Except that using 10g by preparation embodiment 30 synthetic mPEG-PCL and the 0.406g tocofecol succinic acid salt, according to embodiment 1b) identical step obtains the mPEG-PCL-tocopherol (10g of purifying; Productive rate=87.5%).

Embodiment 15

The polymerization of mono methoxy polyethylene glycol-polycaprolactone cholesterol (mPEG-PCL-cholesterol) segmented copolymer (AB type)

Except that using 10g by preparation embodiment 30 synthetic mPEG-PCL and the 0.372 gram cholesterol succinate, according to embodiment 1b) identical step obtains the mPEG-PCL-cholesterol (10g of purifying; Productive rate=88.6%).

Embodiment 16

The polymerization 6 of mPEG-PLA-cholesterol

Weighing four (4) gram cholesterol use vacuum pump 50 ℃ of dehydrations down.To wherein adding succinyl dichloride (3.0g; 2.0 times of moles of cholesterol), reacted 12 hours.After reaction is finished, remove unnecessary succinyl dichloride in 100 ℃ under the vacuum.To wherein adding mPEG-PLA (36g; 0.95 times of mole of cholesterol), reacted 12 hours.The synthetic polymer dissolution in methylene dichloride, is precipitated in hexane/ether solvent to obtain to have the amphiphilic block copolymer of cholesterol group, mPEG-PLA-cholesterol then.The polymerisate of filtering-depositing, the vacuum-drying particulate polymkeric substance (35g that obtains being white in color then; Productive rate 88%).

Embodiment 17 to 20

The polymerase 17 to 10 of mPEG-PLA-cholesterol

Be respectively oxalyl chloride (embodiment 17), malonyl chloride (embodiment 18), glutaryl chlorine (embodiment 19) and the hexanedioyl chlorine (embodiment 20) of 2 times of molar weights of cholesterol except that using, obtain the mPEG-PLA-cholesterol according to the step identical with embodiment 16.

Embodiment 21 to 25

The polymerization 5 to 9 of mPEG-PLA-tocopherol

Except that using the 4.3g tocopherol and be respectively oxalyl chloride (embodiment 21), malonyl chloride (embodiment 22), succinyl dichloride (embodiment 23), glutaryl chlorine (embodiment 24) and the hexanedioyl chlorine (embodiment 25) of 2 times of molar weights of tocopherol, obtain the mPEG-PLA-tocopherol according to the step identical with embodiment 16.

Embodiment 26

The polymerization 11 of mPEG-PLA-cholesterol

Weighing cholesterol succinate (4.9g) and oxalyl chloride (2.53g; 2.0 times of moles of cholesterol succinate), reacted 6 hours down at 50 ℃.After reaction is finished, remove unnecessary oxalyl chloride under the vacuum.Weighing and to wherein adding mPEG-PLA (36g; 0.95 times of mole of cholesterol succinate).Temperature of reaction is 100 ℃, and reaction was carried out 12 hours.The synthetic polymer dissolution in methylene dichloride, is precipitated in hexane/ether solvent and filtration then.The vacuum-drying product is to obtain mPEG-PLA-cholesterol (34.6g; Productive rate 91%).

Embodiment 27 to 29

The polymerization 12 to 14 of mPEG-PLA-cholesterol

Except that using cholesterol malonate (embodiment 27), cholesterol glutarate (embodiment 28) and cholesterol adipate (embodiment 29), obtain the mPEG-PLA-cholesterol according to the step identical with embodiment 26.

Embodiment 30 to 33

The polymerization 10 to 13 of mPEG-PLA-tocopherol

Except that using tocopherol malonate (embodiment 30), tocofecol succinic acid salt (embodiment 31), tocopherol glutarate (embodiment 32) and tocopherol adipate (embodiment 33), obtain the mPEG-PLA-tocopherol according to the step identical with embodiment 26.

Embodiment 34

The preparation of tocopherol-PLA-mPEG-PLA-tocopherol

Except that using 10g by preparation embodiment 31 synthetic PLA-mPEG-PLA and the tocofecol succinic acid salt (2.4 times of moles of polymkeric substance), according to embodiment 1b) identical step obtains tocopherol-PLA-mPEG-PLA-tocopherol (productive rate=92.4%).

Embodiment 35

The preparation of cholesterol-PLA-mPEG-PLA-cholesterol

Except that using 10g by the preparation embodiment 31 synthetic PLA-mPEG-PLA, according to embodiment 1b) identical step obtains cholesterol-PLA-mPEG-PLA-cholesterol (productive rate=94.2%).

Embodiment 36

The pharmacokinetics of the polymeric micelle of the amphiphilic block copolymer of puting together with hydrophobic part that contains taxol

For estimate be used to replace amphiphilic block copolymer (mPEG-PLA, Mn2001-1765) hydrophobic part of hydrophobic B block hydroxyl terminal is to the influence of the blood resides time of the polymeric micelle that contains taxol, we have prepared following composition.With the amphiphilic block copolymer of taxol and embodiment 1,7 or preparation embodiment 27 mixed, then mixture is dissolved in the 5ml dehydrated alcohol with the preparation clear soln with 1:99.Utilize vacuum aspirator to remove wherein ethanol contains taxol with preparation polymeric composition.To wherein adding distilled water (4ml), stir the mixture in 60 ℃ and to contain the polymeric micelle aqueous solution of taxol in 10 minutes with preparation.With the strainer of mixture by the 200nm aperture, freeze-drying then.

Table 5 has been summarized above composition and medicament contg.

Table 5

Concerning experimentation on animals, be the femoral vein and the femoral artery place intubate of the male Sprague-Dawley rat of 250-300g in weight.The dosage of composition 1 to 3 with 5mg/kg was injected in the femoral vein through 15 seconds.After the injection, when 1,5,15 and 30 minute and 1,2,3,4 and 6 hour, from femoral artery, get the 0.3ml whole blood, centrifugal then to obtain clarifying supernatant blood plasma.

In order to analyze the plasma concentration of medicine, we pour 0.1ml blood plasma in the Glass tubing that is capped, and contain the acetonitrile solution of internal standard substance to wherein adding 0.1ml.Ten (10) ml ethyl acetate are joined in the above solution, powerful stirred this mixture 30 seconds, then 2, under the 500rpm centrifugal 10 minutes.Round ethyl acetate layer and be transferred in the test tube, in nitrogen gas stream, under 40 ℃, evaporate organic solvent then fully.To the acetonitrile solution that wherein adds 0.1ml 40% (v/v), powerful this mixture of stirring 30 seconds carries out HPLC then.The condition of HPLC is as follows:

Inject volume: 0.075ml

Flow velocity: 1.0ml/ minute

Wavelength: 227nm

Moving phase: 24% acetonitrile solution carried out 5 minutes, brought up to 58% and carried out 16 minutes, brought up to 70% and carried out 2 minutes, was reduced to 34% and carried out 4 minutes, kept 5 minutes.

Post: 4.6 * 50nm (C18, Vydac, USA).

Following table 6 and Fig. 8 have shown the micellar size and to drug plasma concentration result's analysis.

Table 6

As table 6 and shown in Figure 8, the polymeric micelle (composition 1 and 2) of amphiphilic block copolymer of hydrophobic part (tocofecol succinic acid or cholesterol succsinic acid) with the C-terminal that is used to replace hydrophobic B block is than the blood resides time significant prolongation of natural mPEG-PLA-OH polymeric micelle (composition 3).This result has hinted that owing to more powerful interaction between the hydrophobic part of amphiphilic polymer and medicine the hydrophobic performance that therefore increases hydrophobic B block in the amphiphilic block copolymer forms more stable micella.

In addition, confirmed that also mPEG-PLA-tocopherol micella (composition 1) is longer the cycle time in blood than mPEG-PLA-cholesterol micella (composition 2).

Embodiment 37

The preparation of ionization fixed polymeric micelle

Step 1:D, the polymeric gel of L-PLA-COONa and mPEG-PLA-tocopherol segmented copolymer The preparation of bundle

With the D of 248.1mg (0.218 mole) from preparation embodiment 15, L-PLA-COONa (Mn:1,140) and 744.3mg be dissolved in fully in the 5ml ethanol to obtain clear soln from the mPEG-PLA-tocopherol (Mn:2,000-1,800 dalton) of embodiment 7.Therefrom remove ethanol to make polymeric composition.To wherein adding distilled water (6.2ml), stir the mixture 30 minutes to make the polymeric micelle aqueous solution at 60 ℃

Step 2: fix with divalent-metal ion

The aqueous solution of 0.121ml (0.109mmol) 0.9M Calcium Chloride Powder Anhydrous is joined in the polymeric micelle aqueous solution of step 1 preparation, stirred the mixture under the room temperature 20 minutes.With mixture by having the strainer in 200nm aperture, freeze-drying then.The particle diameter of measuring according to dynamic light scattering (DLS) method is 25nm.

Embodiment 38

Fixed Ca ²⁺The D that contains taxol, the preparation of the micellar of L-PLA-COONa and mPEG-PLA-tocopherol segmented copolymer

Step 1: contain the D of taxol, L-PLA-COONa and mPEG-PLA-tocopherol block are altogether The preparation of the polymeric micelle of polymers

With the D of 248.1mg (0.218 mole) from preparation embodiment 15, L-PLA-COONa (Mn:1,140), 7.5mg taxol and 744.3mg are dissolved in the 5ml ethanol to obtain clear soln fully from the mPEG-PLA-tocopherol (Mn:2,000-1,800 dalton) of embodiment 7.Therefrom remove ethanol to make the polymeric composition that contains taxol.To wherein adding distilled water (6.2ml), stir the mixture 30 minutes to make the polymeric micelle aqueous solution that contains taxol at 60 ℃.

Step 2: fix with divalent-metal ion

The aqueous solution of 0.121ml (0.109mmol) 0.9M Calcium Chloride Powder Anhydrous is joined in the polymeric micelle aqueous solution of step 1 preparation, stirred the mixture under the room temperature 20 minutes.With mixture by having the strainer in 200nm aperture, freeze-drying then.Measure the content of taxol and solubleness and measure particle diameter through HPLC according to dynamic light scattering (DLS) method.

D, L-PLA-COONa/mPEG-PLA-tocopherol (weight ratio): 1/3

Content of taxol: 0.75 weight %

Particle diameter: 29nm

Embodiment 39

Fixed Mg ²⁺The D that contains taxol, the preparation of the polymeric micelle of L-PLMA-COONa and mPEG-PLA-tocopherol segmented copolymer

Remove and use the D of 248.1mg (0.226mmol) from preparation embodiment 24, L-PLMA-COONa (Mn:1,096), 7.5mg taxol and 744.3mg are from the mPEG-PLA-tocopherol (Mn:2 of embodiment 7,000-1,800 dalton), and outside the aqueous solution (Mw:203.31) of 0.230 (0.113mmol) 0.5M magnesium chloride, 6 hydrates, fixed Mg according to the step preparation identical with embodiment 38 ²⁺The polymeric micelle compositions that contains taxol.

D, L-PLMA-COONa/mPEG-PLA-tocopherol (weight ratio): 1/3

Content of taxol: 0.75 weight %

Particle diameter: 30nm

Embodiment 40

Fixed Ca ²⁺The D that contains taxol, the preparation of the polymeric micelle of L-PLMA-COONa and mPEG-PLA-tocopherol segmented copolymer

Remove and use the D of 248.1mg (0.226mmol) from preparation embodiment 24, L-PLMA-COONa (Mn:1,096), 7.5mg taxol and 744.4mg are from the mPEG-PLA-tocopherol (Mn:2 of embodiment 7,000-1,800 dalton), and outside the aqueous solution of the Calcium Chloride Powder Anhydrous of 0.126 (0.113mmol) 0.9M, fixed Ca according to the step preparation identical with embodiment 38 ²⁺The polymeric micelle compositions that contains taxol.

D, L-PLMA-COONa/mPEG-PLA-tocopherol (weight ratio): 1/3

Content of taxol: 0.75 weight %

Particle diameter: 34nm

Embodiment 41

Fixed Ca ²⁺The D that contains taxol, the preparation of the polymeric micelle of L-PLA-COOK and mPEG-PLA-cholesterol segmented copolymer

Remove and use the D of 248.1mg (0.160mmol) from preparation embodiment 18, L-PLA-COOK (Mn:1,550), 7.5mg taxol and 744.4mg are from the mPEG-PLA-cholesterol (Mn:2 of embodiment 1,000-1,800 dalton), and outside the aqueous solution of the Calcium Chloride Powder Anhydrous of 0.089 (0.080mmol) 0.9M, fixed Ca according to the step preparation identical with embodiment 38 ^{2 +}The polymeric micelle compositions that contains taxol.

D, L-PLMA-COONa/mPEG-PLA-cholesterol (weight ratio): 1/3

Content of taxol: 0.75 weight %

Particle diameter: 34nm

Embodiment 42

Fixed Ca ²⁺The D that contains taxol, the preparation of the polymeric micelle of L-PLMA-COONa and mPEG-PLA-cholesterol segmented copolymer

Remove and use the D of 248.1mg (0.226mmol) from preparation embodiment 24, L-PLMA-COONa (Mn:1,096), 7.5mg taxol and 744.4mg are from the mPEG-PLA-cholesterol (Mn:2 of embodiment 1,000-1,800 dalton), and outside the aqueous solution of the Calcium Chloride Powder Anhydrous of 0.126 (0.113mmol) 0.9M, fixed Ca according to the step preparation identical with embodiment 38 ²⁺The polymeric micelle compositions that contains taxol.

D, L-PLMA-COONa/mPEG-PLA-cholesterol (weight ratio): 1/3

Content of taxol: 0.75 weight %

Particle diameter: 34nm

Embodiment 43

Fixed Ca ²⁺The 3 arm PLA-COONa that contain taxol and the preparation of the polymeric micelle of mPEG-PLA-tocopherol segmented copolymer

Remove and use the 3 arm PLA-COONa (Mn:3s of 248.1mg (0.0827mmol) from preparation embodiment 25,000), 7.5mg taxol and 744.4mg are from the mPEG-PLA-tocopherol (Mn:2 of embodiment 7,000-1,800 dalton), and outside the aqueous solution of the Calcium Chloride Powder Anhydrous of 0.1377 (0.124mmol) 0.9M, fixed Ca according to the step preparation identical with embodiment 38 ²⁺The polymeric micelle compositions that contains taxol.

3 arm PLACOONa/mPEG-PLA-tocopherols (weight ratio): 1/3

Content of taxol: 0.75 weight %

Particle diameter: 29nm

Embodiment 44

Fixed Ca ²⁺The 5 arm PLA-COONA that contain taxol and the preparation of the polymeric micelle of mPEG-PLA-tocopherol segmented copolymer

Remove and use the 5 arm PLA-COONa (Mn:3s of 248.1mg (0.0827mmol) from preparation embodiment 26,000), 7.5mg taxol and 744.4mg are from the mPEG-PLA-tocopherol (Mn:2 of embodiment 7,000-1,800 dalton), and outside the aqueous solution of the Calcium Chloride Powder Anhydrous of 0.2295 (0.207mmol) 0.9M, fixed Ca according to the step preparation identical with embodiment 38 ²⁺The polymeric micelle compositions that contains taxol.

5 arm PLACOONa/mPEG-PLA-tocopherols (weight ratio): 1/3

Content of taxol: 0.75 weight %

Particle diameter: 29nm

Embodiment 45

The D that contains Zorubicin, the preparation of the polymeric micelle of L-PLMA-COONa and mPEG-PLA-tocopherol segmented copolymer

With mPEG-PLA-tocopherol (Mn:2,000-1,800), D, L-PLMA-COONa (Mn:969) and Zorubicin HCl mix with the weight ratio of 78.62:17.24:1.00, then mixture are dissolved in the 5ml anhydrous methanol with the preparation settled solution.Use vacuum aspirator therefrom to remove methyl alcohol contains Zorubicin with preparation polymeric composition.To wherein adding distilled water (4ml), stir the mixture in 60 ℃ and to contain the polymeric micelle aqueous solution of Zorubicin in 10 minutes with preparation.Is the strainer of 200nm, freeze-drying then with mixture by the aperture.

D, L-PLMA-COONa/mPEG-PLA-tocopherol (weight ratio): 1/4.56

Zorubicin content: 1.03 weight %

Particle diameter: 35nm

Embodiment 46

The D that contains epirubicin, the preparation of the polymeric micelle of L-PLMA-COONa and mPEG-PLA-tocopherol segmented copolymer

With mPEG-PLA-tocopherol (Mn:2,000-1,800), D, L-PLMA-COONa (Mn:969) and epirubicin HCl mix with the weight ratio of 78.26:17.24:1.00, then mixture are dissolved in the 5ml anhydrous methanol with the preparation clear soln.Use vacuum aspirator therefrom to remove methyl alcohol contains epirubicin with preparation polymeric composition.To wherein adding distilled water (4ml), stir the mixture in 60 ℃ and to contain the polymeric micelle aqueous solution of epirubicin in 10 minutes with preparation.Is the strainer of 200nm, freeze-drying then with mixture by the aperture.

D, L-PLMA-COONa/mPEG-PLA-tocopherol (weight ratio): 1/4.56

Epirubicin content: 1.03 weight %

Particle diameter: 30nm

Embodiment 47

Fixed Ca ²⁺The particle diameter of polymeric micelle

In order to determine to have fixed Ca ²⁺The particle diameter of polymeric micelle, we have prepared following polymeric micelle compositions.

With mPEG-PLA (Mn:2,000-1,800) and D, L-PLMA-COONa (Mn:866,994,1,156,1,536) mixes with the equivalence ratio of 1:1, then mixture is dissolved in the 5ml dehydrated alcohol with the preparation clear soln.Use vacuum aspirator therefrom to remove ethanol with the preparation polymeric composition.To wherein adding distilled water, stir the mixture in 60 ℃ and to contain the polymeric micelle aqueous solution of taxol in 10 minutes with preparation.Add and D in the above polymeric micelle aqueous solution, L-PLMA-COONa solution has the CaCl of identical equivalents ₂The aqueous solution (concentration: 100mg/ml), and under room temperature, stirred the mixture 20 minutes.Is the strainer of 200nm with mixture by the aperture, is 7.4 PBS damping fluid obtains 40mg/ml concentration with the diluted mixture thing polymkeric substance to wherein adding pH then.After using the 0.22um film filter to filter, measure particle diameter with the related particle size analysis device of photon.

Table 7

As shown in table 7, fixed Ca ²⁺The median size of polymeric micelle be 20-40nm.The micella of this particle size range is suitable for injection formulations and sterile filtration.As for being elevated to 1536 D from 866, the molecular weight of L-PLMA-COONa is used Ca ²⁺Processing and untreated micellar particle diameter all have increase slightly.Fixed Ca ²⁺The particle diameter ratio Ca of polymeric micelle ²⁺Handle larger about 10nm.

Embodiment 48

Fixed Ca ²⁺The kinetic stability of the polymeric micelle that contains taxol

In order to detect the stability of nano-composition, we have prepared following polymeric micelle compositions.

With taxol, mPEG-PLA-tocopherol (Mn:2,000-1,800) and D, L-PLMA-COONa (Mn:1,096) mixes with the equivalence ratio of 1:3:3 (composition 4), then mixture is dissolved in the 5ml dehydrated alcohol with the preparation clear soln.Use vacuum aspirator therefrom to remove ethanol contains taxol with preparation polymeric composition.To wherein adding distilled water (4ml), stir the mixture in 60 ℃ and to contain the polymeric micelle aqueous solution of taxol in 10 minutes with preparation.Add and D in the above polymeric micelle aqueous solution, L-PLMA-COONa solution has the CaCl of identical equivalents ₂The aqueous solution (concentration: 100mg/ml), and under room temperature, stirred the mixture 20 minutes.Is the strainer of 200nm, freeze-drying then with mixture by the aperture.

With taxol, mPEG-PLA-tocopherol (Mn:2,000-1,800) and D, L-PLMA-COONa (Mn:1,096) mixes with the equivalence ratio of 1:3:3 (composition 5), then mixture is dissolved in the 5ml dehydrated alcohol with the preparation clear soln.Use vacuum aspirator therefrom to remove ethanol contains taxol with preparation polymeric composition.To wherein adding distilled water (4ml), stir the mixture in 60 ℃ and to contain the polymeric micelle aqueous solution of taxol in 10 minutes with preparation.Is the strainer of 200nm, freeze-drying then with mixture by the aperture.

(composition 6) mixes taxol, mPEG-PLA-tocopherol (Mn:2,000-1,800) with the equivalence ratio of 1:3, then mixture is dissolved in the 5ml dehydrated alcohol with the preparation clear soln.Use vacuum aspirator therefrom to remove ethanol contains taxol with preparation polymeric composition.To wherein adding distilled water (5ml), stir the mixture in 60 ℃ and to contain the polymeric micelle aqueous solution of taxol in 10 minutes with preparation.Is the strainer of 200nm, freeze-drying then with mixture by the aperture.

With taxol, mPEG-PLA (Mn:2,000-1,765) and D, L-PLMA-COONa (Mn:1,096) mixes with the equivalence ratio of 1:3:3 (composition 7), then mixture is dissolved in the 5ml dehydrated alcohol with the preparation clear soln.Use vacuum aspirator therefrom to remove ethanol contains taxol with preparation polymeric composition.To wherein adding distilled water (4ml), stir the mixture in 6O ℃ and to contain the polymeric micelle aqueous solution of taxol in 10 minutes with preparation.Add and D in the above polymeric micelle aqueous solution, L-PLMA-COONa solution has the CaCl of identical equivalents ₂The aqueous solution (concentration: 100mg/ml), and under room temperature, stirred the mixture 20 minutes.Is the strainer of 200nm, freeze-drying then with mixture by the aperture.

Table 8

	MPEG-PLA-tocopherol (mg)	mPEG-PLA(mg)	D，L-PLMA-COONa(mg)	Taxol (mg)	CaCl 2(mg)	Content of taxol (mg/ml)
							Composition 4	267.0	-	77.0	20.0	3.9	1.0
Composition 5	267.0	-	77.0	20.0	-	1.0
							Composition 6	267.0	-	-	20.0	-	1.0
Composition 7	-	267.0	77.0	20.0	3.9	1.0

With pH is that 7.4 PBS damping fluid joins in the lyophilised compositions to make the taxol of 1.0mg/ml concentration.Mixture leaves standstill at 37 ℃, measures the concentration of passing taxol in time by HPLC.The result is presented in the table 9.

Table 9

As shown in table 9, fixed Ca ²⁺The polymeric micelle compositions that contains taxol (composition 4) than not using Ca ²⁺The kinetics of the composition of handling (composition 5) is more stable.Add Ca ²⁺Can significantly improve the delay of taxol in the polymeric micelle of the present invention.This is owing to D, L-PLA-COO ^-And Ca ²⁺Crosslinked electrostatic interaction, it may cause the rigidity of hydrophobic core to increase.Fixed Ca ²⁺The polymeric micelle (composition 4) of the amphiphilic block copolymer that has the hydrophobic part (tocofecol succinic acid) that is used to replace hydrophobic B block hydroxyl terminal than having fixed Ca ²⁺Residence time of polymeric micelle (composition 7) of natural mPEG-PLA-OH long.This result has hinted also that owing to the hydrophobic part of amphiphilic block copolymer and the more powerful effect between the medicine hydrophobic performance that therefore increases hydrophobic B block in the amphiphilic block copolymer forms more stable micella.

Embodiment 49

Fixed Ca ²⁺The pharmacokinetics of the polymeric micelle that contains taxol

Be used to replace the hydrophobic part of the hydrophobic B block of amphiphilic Synthetic rubber, isoprene-styrene, hydrogenated, block, diblock hydroxyl terminal to having fixed Ca in order to estimate ²⁺The influence of the polymeric micelle that contains taxol residence time in blood, we have prepared following composition.

With taxol, mPEG-PLA-tocopherol (Mn:2,000-1,800) or mPEG-PLA-OH, and D, L-PLMA-COONa (Mn:1,004) mixes with the weight ratio of 74.25:24.75:1.00, then mixture is dissolved in the 5ml dehydrated alcohol with the preparation clear soln.Use vacuum aspirator therefrom to remove ethanol contains taxol with preparation polymeric composition.To wherein adding distilled water (4ml), stir the mixture in 60 ℃ and to contain the polymeric micelle aqueous solution of taxol in 10 minutes with preparation.Add and D in the above polymeric micelle aqueous solution, L-PLMA-COONa solution has the CaCl of identical equivalents ₂The aqueous solution (concentration: 100mg/ml), and under room temperature, stirred the mixture 20 minutes.Is the strainer of 200nm, freeze-drying then with mixture by the aperture.

Table 10 has been summarized above composition and content of medicines.

Table 10

Concerning experimentation on animals, be the femoral vein and the femoral artery place intubate of the male Sprague-Dawley rat of 220-270g in weight.The composition of embodiment 8 and 9 dosage with 5mg/kg was injected in the femoral vein through 15 seconds.After the injection, when 1,5,15 and 30 minute and 1,2,3,4 and 6 hour, from femoral artery, get the 0.3ml whole blood, centrifugal then to obtain clarifying supernatant blood plasma.

Process analysis Plasma Concentration according to identical with embodiment 36 has shown the analytical results to drug plasma concentration among following table 11 and Fig. 9.

Table 11

As table 11 and shown in Figure 9, fixed Ca ²⁺The polymeric micelle (composition 8) of amphiphilic Synthetic rubber, isoprene-styrene, hydrogenated, block, diblock of hydrophobic part (tocofecol succinic acid) with the terminal hydroxyl that is used to replace hydrophobic B block than having fixed Ca ²⁺Blood resides time of natural mPEG-PLA-OH polymeric micelle (composition 9) obviously prolong.This result hinted, such as embodiment 36 record because more powerful interaction between the hydrophobic part of amphiphilic polymer and medicine, the hydrophobic performance that therefore increases hydrophobic B block in the amphipathic copolymer forms more stable micella.

Embodiment 50

Fixed Ca ²⁺The pharmacokinetics of the polymeric micelle that contains taxol

In order relatively to have fixed Ca ²⁺The polymeric micelle that contains taxol with contain the residence time of preparation in blood of other carrier, we have prepared following composition.

(composition 10) fixed Ca ²⁺The polymeric micelle that contains taxol

With taxol, mPEG-PLA-tocopherol (Mn:2,000-1,800) and D, L-PLMA-COONa (Mn:1,004) mixes with the weight ratio of 99.25:33.08:1.00, then mixture is dissolved in the 5ml dehydrated alcohol with the preparation clear soln.Use vacuum aspirator therefrom to remove ethanol contains taxol with preparation polymeric composition.To wherein adding distilled water (4ml), stir the mixture in 60 ℃ and to contain the polymeric micelle aqueous solution of taxol in 10 minutes with preparation.Add and D in the above polymeric micelle aqueous solution, L-PLMA-COONa solution has the CaCl of identical equivalents ₂The aqueous solution (concentration: 100mg/ml), and under room temperature, stirred the mixture 20 minutes.Is the strainer of 200nm, freeze-drying then with mixture by the aperture.The hydrokinetics particle diameter of polymeric micelle is 34nm.

(composition 11) contains the composition of taxol, polyoxyethylenated castor oil and dehydrated alcohol.

Taxol (30mg) is dissolved in the mixing solutions of 5ml polyoxyethylenated castor oil and dehydrated alcohol (50:50v/v) to obtain clear soln.With solution is the strainer of 200nm by the aperture.

(composition 12) contains the composition of taxol, Polysorbate 80 (tween 80) and dehydrated alcohol.

Taxol (30mg) is dissolved in the mixing solutions of 5ml Polysorbate 80 and dehydrated alcohol (50:50v/v) to obtain clear soln.With solution is the strainer of 200nm by the aperture.

Table 12 has been summarized above composition and content of medicines

Table 12

Concerning experimentation on animals, be the femoral vein and the femoral artery place intubate of the male Sprague-Dawley rat of 230-250g in weight.Composition 10,11 and 12 dosage with 5mg/kg were injected in the femoral vein through 15 seconds.After the injection, when 1,5,15 and 30 minute and 1,2,3,4 and 6 hour, from femoral artery, get the 0.3ml whole blood, centrifugal then to obtain clarifying supernatant blood plasma.

Process analysis Plasma Concentration according to identical with embodiment 36 has shown the analytical results to drug plasma concentration among following table 13 and Figure 10.

Table 13

As table 13 and shown in Figure 10, fixed Ca ²⁺Polymeric micelle (composition 10) longer than the blood resides time of the injection liquid that contains other tensio-active agent ( composition 11 and 12).Because the Ca that fixed of the present invention ²⁺Polymeric micelle (composition 10) than the listing preparation,

The blood resides time of (composition 11) is long, therefore, and biodegradable of the application of the invention and biocompatible polymer, with

Can improve the residence time of medicine in blood than the present invention.

Embodiment 51

Fixed Ca ²⁺The pharmacokinetics of the polymeric micelle that contains taxol

In order relatively to have fixed Ca ²⁺The polymeric micelle that contains taxol with contain the residence time of preparation in blood of other carrier, we have prepared following composition.

(composition 13) fixed Ca ²⁺The polymeric micelle that contains taxol

Taxol, mPEG-PLA-tocopherol (Mn:2,000-1,800) and 5 arm PLA-COONa (Mn:3,000) are mixed with the weight ratio of 99.25:33.08:1.00, then mixture is dissolved in the 5ml dehydrated alcohol with the preparation clear soln.Use vacuum aspirator therefrom to remove ethanol contains taxol with preparation polymeric composition.To wherein adding distilled water (4ml), stir the mixture in 60 ℃ and to contain the polymeric micelle aqueous solution of taxol in 10 minutes with preparation.In the above polymeric micelle aqueous solution, add and have identical normal CaCl with 5 arm PLA-COONa ₂The aqueous solution (concentration: 100mg/ml), and under room temperature, stirred the mixture 20 minutes.Is the strainer of 200nm, freeze-drying then with mixture by the aperture.The hydrokinetics particle diameter of polymeric micelle is 32nm.

(composition 11) contains the composition of taxol, polyoxyethylenated castor oil and dehydrated alcohol.

Taxol (30mg) is dissolved in the mixing solutions of 5ml polyoxyethylenated castor oil and dehydrated alcohol (50:50v/v) to obtain clear soln.With solution is the strainer of 200nm by the aperture.

Table 14 has been summarized above composition and content of medicines

Table 14

Concerning experimentation on animals, be the femoral vein and the femoral artery place intubate of the male Sprague-Dawley rat of 230-250g in weight.Composition 13 and 11 dosage with 5mg/kg were injected in the femoral vein through 15 seconds.After the injection, when 1,5,15 and 30 minute and 1,2,3,4 and 6 hour, from femoral artery, get the 0.3ml whole blood, centrifugal then to obtain clarifying supernatant blood plasma.

Process analysis Plasma Concentration according to identical with embodiment 36 has shown the analytical results to drug plasma concentration among following table 15 and Figure 11.

Table 15

As table 15 and shown in Figure 11, fixed Ca ²⁺Polymeric micelle (composition 13) longer than the blood resides time of the injection liquid that contains other tensio-active agent (composition 11).Because the Ca that fixed of the present invention ²⁺Polymeric micelle (composition 13) than the listing preparation,

The blood resides time of (composition 11) is long, therefore, and biodegradable of the application of the invention and biocompatible polymer, with

Compare the present invention and can improve the residence time of medicine in blood.

Embodiment 52

Fixed Ca ²⁺The pharmacokinetics of the polymeric micelle that contains many Xi Taqi

In order relatively to have fixed Ca ²⁺The polymeric micelle that contains many Xi Taqi with contain the residence time of preparation in blood of other carrier, we have prepared following composition.

(composition 14) fixed Ca ²⁺The polymeric micelle that contains many Xi Taqi

Many Xi Taqi, mPEG-PLA-tocopherol (Mn:2,000-1,800) and 3 arm PLA-COONa (Mn:3,000) are mixed with the weight ratio of 99.25:33.08:1.00, then mixture is dissolved in the 5ml dehydrated alcohol with the preparation clear soln.Use vacuum aspirator therefrom to remove ethanol contains many Xi Taqi with preparation polymeric composition.To wherein adding distilled water (4ml), stir the mixture in 60 ℃ and to contain the polymeric micelle aqueous solution of many Xi Taqi in 10 minutes with preparation.In the above polymeric micelle aqueous solution, add and have identical normal CaCl with 3 arm PLA-COONa ₂The aqueous solution (concentration: 100mg/ml), and under room temperature, stirred the mixture 20 minutes.Is the strainer of 200nm, freeze-drying then with mixture by the aperture.The hydrokinetics particle diameter of polymeric micelle is 30nm.

(composition 15) contains the composition of many Xi Taqi, Polysorbate 80 (tween 80) and dehydrated alcohol.

Many Xi Taqi (20mg) and tween 80 (520mg) are dissolved in the aqueous ethanolic solution of 1.5ml 13% (v/v) to obtain clear soln.With solution is the strainer of 200nm by the aperture.

Table 16 has been summarized above composition and content of medicines

Table 16

Concerning experimentation on animals, be the femoral vein and the femoral artery place intubate of the male Sprague-Dawley rat of 210-240g in weight.Composition 14 and 15 dosage with 10mg/kg were injected in the femoral vein through 15 seconds.After the injection, when 5,15 and 30 minutes and 1,2,3,6 and 8 hour, from femoral artery, get the 0.3ml whole blood, centrifugal then to obtain clarifying supernatant blood plasma.

Process analysis Plasma Concentration according to identical with embodiment 36 has shown the analytical results to drug plasma concentration among following table 17 and Figure 12.

Table 17

As table 17 and shown in Figure 12, fixed Ca ²⁺Polymeric micelle (composition 14) longer than the blood resides time of the injection liquid that contains tween 80 (composition 15).Because the Ca that fixed of the present invention ²⁺Polymeric micelle (composition 14) than the listing preparation,

The blood resides time of (composition 15) is long, therefore, and biodegradable of the application of the invention and biocompatible polymer, with

Compare the present invention and can improve the residence time of medicine in blood.

Embodiment 53

Fixed Ca ²⁺The pharmacokinetics of the polymeric micelle that contains many Xi Taqi

In order relatively to have fixed Ca ²⁺The polymeric micelle that contains many Xi Taqi and its residence time of preparation in blood of containing other carrier, we have prepared following composition.

(composition 16) fixed Ca ²⁺The polymeric micelle that contains many Xi Taqi

With many Xi Taqi, mPEG-PLA-tocopherol (Mn:2,000-1,800) and D, L-PLA-COONa (Mn:1,700) mixes with the weight ratio of 75.0:25.0:1.00, then mixture is dissolved in the 5ml dehydrated alcohol with the preparation clear soln.Use vacuum aspirator therefrom to remove ethanol contains many Xi Taqi with preparation polymeric composition.To wherein adding distilled water (4ml), stir the mixture in 60 ℃ and to contain the polymeric micelle aqueous solution of many Xi Taqi in 10 minutes with preparation.Add and D in the above polymeric micelle aqueous solution, L-PLA-COONa has identical normal CaCl ₂The aqueous solution (concentration: 100mg/ml), and under room temperature, stirred the mixture 20 minutes.Is the strainer of 200nm, freeze-drying then with mixture by the aperture.The hydrokinetics particle diameter of polymeric micelle is 32nm.

(composition 15) contains many Xi Taqi, tween 80 and 13% alcoholic acid composition.

Many Xi Taqi (20mg) and tween 80 (520mg) are dissolved in the aqueous ethanolic solution of 1.5ml 13% (v/v) to obtain clear soln.With solution is the strainer of 200nm by the aperture.

Table 18 has been summarized above composition and content of medicines

Table 18

Concerning experimentation on animals, be the femoral vein and the femoral artery place intubate of the male Sprague-Dawley rat of 230-250g in weight.Composition 16 and 15 dosage with 5mg/kg were injected in the femoral vein through 15 seconds.After the injection, when 1,5,15 and 30 minute and 1,2,3,4 and 6 hour, from femoral artery, get the 0.3ml whole blood, centrifugal then to obtain clarifying supernatant blood plasma.

Process analysis Plasma Concentration according to identical with embodiment 36 has shown the analytical results to drug plasma concentration among following table 19 and Figure 13.

Table 19

As table 19 and shown in Figure 13, fixed Ca ²⁺Polymeric micelle (composition 16) longer than the blood resides time of the injection that contains tween 80 (composition 15).Because the Ca that fixed of the present invention ²⁺Polymeric micelle (composition 16) than the listing preparation,

The blood resides time of (composition 15) is long, therefore, and biodegradable of the application of the invention and biocompatible polymer, with

Compare the present invention and can improve the residence time of medicine in blood.

Embodiment 54

Fixed Ca ²⁺The maximum tolerated dose of the polymeric micelle that contains taxol

With the timetable of 0-, 1-and 2-days, give ten (10) group Tac:Cr:(Ncr through the tail vein)-nude mouse (female, 8 weeks, 20.5 ± 0.50g; Male, 8 weeks, 21.3 ± 1.6) intravenous injection dosage be 16,20,25 and 30mg/kg fixed Ca ²⁺The polymeric micelle solution that contains taxol (composition 10).In 30 days, observe the mouse of survival in all groups and the variation of body weight every day.

With the timetable of 0-, 2-and 4-days, give five (5) group Tac:Cr:(Ncr through the tail vein)-nude mouse (female, 8 weeks, 24.7 ± 1.2; Male, 8 weeks, 24.2 ± 1.3) intravenous injection dosage be 20,25,30 and 35mg/kg fixed Ca ²⁺The polymeric micelle solution that contains taxol (composition 10).In 30 days, observe the mouse of survival in all groups and the variation of body weight every day.

With 0-, 2-, 4-and 6-days timetable, give four (4) group Tac:Cr:(Ncr through the tail vein)-nude mouse (female, 8 weeks, 22.5 ± 0.8; Male, 8 weeks, 24.3 ± 1.6) intravenous injection dosage be 20,25 and 30mg/kg fixed Ca ²⁺The polymeric micelle solution that contains taxol (composition 10).In 30 days, observe the mouse of survival in all groups and the variation of body weight every day.

With the timetable of 0-, 4-and 8-days, give ten (10) group Tac:Cr:(Ncr through the tail vein)-nude mouse (female, 8 weeks, 19.3 ± 0.71g; Male, 8 weeks, 23.3 ± 1.1) intravenous injection dosage be 25,28,30,35 and 39mg/kg fixed Ca ²⁺The polymeric micelle solution that contains taxol (composition 10).In 30 days, observe the mouse of survival in all groups and the variation of body weight every day.

The definition of MTD is in 2 weeks, both not cause that death (because toxic action) when also not having sign of life significantly to change, is about the tolerance of the intermediate value that loses weight that contrasts 10-20% after administration.Shown in table 20, the MTD in each administration time table is all in the scope of 20-30mg/kg.

We have also carried out the toxicity research of vehicle.The Ca that accepted not contained pharmaceutical fixed ²⁺The fast growth of animal of polymeric micelle, its weight ratio has been accepted salt solution or has not been had the animal of injection high slightly.This is attributed to the calorie content of preparation.

Table 20

Embodiment 55

Fixed Ca ²⁺The antitumour activity of the polymeric micelle that contains taxol

Get the cell that is stored in the liquid nitrogen, set up cell in vitro and cultivate.After the results, washed cell in sterile phosphate buffered saline (PBS) is determined viable count.With cell with about 7 * 10 ⁷The concentration of cell/ml is resuspended among the aseptic PBS.Right side rib place subcutaneous injection 0.1ml at health naked (nu/nu) athymic mouse (20-25g, 8 ages in week) contains 7 * 10 ⁶The cell suspension of human cancer cell (MX-1, SKOV-3, MDAMB435S, HT29, PC-3, U373MG).After the cancer knurl reaches a certain size, they are carried out xenotransplantation for 3 times to form the heterograft fragment of 3-4mm.Trocar with No. 12 (gauge) is subcutaneously injected into healthy athymia (nu/nu) the nude mice right side rib in (20-25g, 8 ages in week) with this heterograft fragment.When gross tumor volume reaches 100-300mm ³The time, give medicine, this time point is designated as the 0th day.In the time of the 0th day, mouse is divided into 5 groups, at the 0th, 1 and 2 day, the 0th, 2 and 4 day or the 0th, 4 and 8 day, by tail vein had fixing of various dose taxol polymeric micelle of metal ion (composition 10) and polyoxyethylenated castor oil preparation (composition 11), and measure the volume of cancer at different timed interval places.The volume of cancer is by formula (W ²* L)/2 calculate, wherein W is a minor axis, L is a major axis.

For treatment being estimated the following gross tumor volume that calculated:

Gross tumor volume (TV)=0.5 * L * W ²(L: major axis, W: minor axis)

Relative tumour volume (RTV)=(V _t/ V ₀) * 100% (Vt: t days TV, V ₀: the 0th day TV)

3 standards of parallel use are determined result of treatment: average tumor growth curve, optimum growh suppress (T/C%) and particular growth postpones (SGD).

By the treatment group optimum growh that the average RTV of control group on duty 100% calculates 4 all inherent certain day after in the end injecting is suppressed (T/C%).

One or twice calculate SGD in the doubling time, as follows:

Particular growth postpones (SGD): SGD=(T _{The D treatment}-T _{The D contrast})/T _{The D contrast}

T _D: tumour-doubling time

Activity level is determined as follows:

	T/C％ SGD
		(+) + ++ +++ ++++	＜50 or 1.0＜50 Hes 1.0＜40 Hes 1.5＜25 Hes 2.0＜10 Hes 3.0

According to the NCI standard, T/C≤42%th, minimum activity-level.T/C＜10% has been considered to prove the maximum anti-tumor activity level that further develops.

Concerning the experiment of estimating that is considered, each treatment group has 4 mouse at least to control group, every group of at least 4 tumours.When the treatment beginning, the smallest tumors diameter is 4mm or 30mm ³Volume.Dead animal is considered to toxicity and causes death in 2 weeks after giving last medicine, and eliminating is outside any evaluation.There is treatment group to be considered to be used to estimate antitumous effect greater than 1/3 toxicity causes death or the body weight loss intermediate value is not recovered fully greater than 15%.

Shown in Figure 14 to 21 and table 21, with according to the group compare, the treatment group and the polyoxyethylenated castor oil preparation for treating group of having fixed the polymeric micelle of metal ion have shown significant inhibition to tumor growth, particularly, fixed the treatment group of polymeric micelle (composition 10) of metal ion than the inhibiting rate height of polyoxyethylenated castor oil preparation (composition 11) treatment group.

Table 21

(3 cycles of *:, in 21,25,29 days (2 cycle), during 42,46 and 50 days (3 cycle), give the medicine of the single intravenous injection dosage in the salt solution through intravenously the 0th, 4,8 day (1 cycle).

Embodiment 56

Fixed Ca ²⁺The polymeric micelle that contains taxol to the tolerance

The antitumour activity of carcinoma animal model

Get the cell that is stored in the liquid nitrogen, set up cell in vitro and cultivate.After the results, washed cell in sterile phosphate buffered saline (PBS) is determined viable count.With cell with about 7 * 10 ⁷The concentration of cell/ml is resuspended among the aseptic PBS.Right side rib place subcutaneous injection 0.1ml healthy athymia (nu/nu) nude mice (20-25g, 8 ages in week) contains 7 * 10 ⁶The cell suspension of human cancer cell (HT29).After cancer reaches a certain size, they are carried out xenotransplantation for 3 times to form the heterograft fragment of 3-4mm.With No. 12 trocars this heterograft fragment is subcutaneously injected into healthy athymia (nu/nu) the nude mice right side rib in (20-25g, 8 ages in week).When gross tumor volume reaches specific when size, under the administration time table of q1dX5, with 20mg/kg/ days dosage through the tail vein give taxol (the polyoxyethylenated castor oil preparation,

).After 3 weeks, under the administration time table of q1dX5, once more with 20mg/kg/ days dosed administration to obtain tolerance

The heterograft fragment of cancer.When cancer reaches specific when big or small, this heterograft fragment (3-4mm) is subcutaneously injected into healthy athymia (nu/nu) the nude mice right side rib in (20-25g, 8 ages in week) with No. 12 trocars.Reach 100-300mm when corpus carcinosus is long-pending ³The time, give medicine, this time point is designated as the 0th day.In the time of the 0th day, mouse is divided into 5 groups, at the 0th, 2 and 4 day,, and measure the volume of cancer at different timed interval places by tail vein had fixing of various dose taxol polymeric micelle of metal ion (composition 10) and polyoxyethylenated castor oil preparation (composition 11).

As described in above experiment, in order to illustrate that the polymeric micelle of having fixed metal ion is to tolerance

The effect of cancer, we have set up tolerance

The active animal model of vivo antitumor of cancer.Cancer cells repeated exposure in being inoculated into mouse exists The time, taxol is to using

The IC of Zhi Liao cancer cells in advance ₅₀IC than the taxol that is used for natural cancer cells ₅₀(not having display data) is significantly improved.In this animal model, polymeric micelle (composition 10) the treatment group of having fixed metal ion has shown than polyoxyethylenated castor oil preparation (composition 11) treatment group high inhibitory, this may be owing to the effective concentration residence time of medicine in blood that is incorporated in the polymeric micelle of having fixed metal ion is long, shown in Figure 22 and table 22.

Table 22

Embodiment 57

Fixed Ca ²⁺The polymeric micelle that contains taxol to the antitumour activity of carcinoma animal model of tolerance Zorubicin

From American Type Culture Colection (ATCC) purchaser sarcoma of uterus, Zorubicin (

) tolerance subbreed (MES-SA/Dx5; The MDR varient), cultivate and in being supplemented with the PRMI-1640 medium of 10%FBS and separate.After the results, washed cell in sterile phosphate buffered saline (PBS) is determined viable count.With cell with about 7 * 10 ⁷The concentration of cell/ml is resuspended among the aseptic PBS.Right side rib place subcutaneous injection 0.1ml healthy athymia (nu/nu) nude mice (20-25g, 8 ages in week) contains 7 * 10 ⁶The cell suspension of human cancer cell (MES-SA/Dx5).After cancer reaches a certain size (500-700mg), the section that the cancer graft is cut into 3 * 3 * 3mm, and transplant with trocar, go down to posterity 3 times then to form the heterograft fragment of 3-4mm.With No. 12 trocars this heterograft fragment is subcutaneously injected into healthy athymia (nu/nu) the nude mice right side rib in (20-25g, 8 ages in week).When gross tumor volume reaches 100-300mm ³The time, give medicine, this time point is designated as the 0th day.In the time of the 0th day, mouse is divided into 5 groups, at the 0th, 2 and 4 day, with the dosage of 20mg/kg taxol by tail vein administration of fixed polymeric micelle of metal ion (composition 10) and polyoxyethylenated castor oil preparation (composition 11).Measure the volume of cancer at different timed interval places.

As described in above experiment, for the polymeric micelle of the having fixed metal ion effect to the cancer of tolerance Zorubicin is described, we have set up the active animal model of vivo antitumor of the cancer of tolerance Zorubicin.In this animal model, polymeric micelle (composition 10) the treatment group of having fixed metal ion has shown than polyoxyethylenated castor oil preparation (composition 11) treatment group high inhibitory, this may be owing to the effective concentration residence time of medicine in blood that is incorporated in the polymeric micelle of having fixed metal ion is long, shown in Figure 23 and table 23.

Table 23

Polymeric micelle according to the present invention by the amphiphilic block copolymer preparation is harmless, and it has higher medicine load rate, and can keep medicine for a long time in the aqueous solution, therefore, in the time of in being expelled to body, can improve Plasma Concentration.

In addition, polymeric composition of the present invention can form stable polymeric micelle or nanoparticle in the body fluid or the aqueous solution.The micella or the nanoparticle that are formed by the present composition have hydrophilic outer shell and hydrophobic inner core, and wherein a large amount of dewatering medicaments can be by the physics load.Pastille micella of the present invention and nanoparticle are giving the back has prolongation in blood residence time, and can be used to make various pharmaceutical preparations.

Be understandable that above-mentioned embodiment only is the explanation that the principle of the invention is used.Multiple variant and other embodiment can both be derived, and do not deviate from the spirit and scope of the present invention.Therefore, though the present invention has shown in the accompanying drawings, and in conjunction with coming special and at length carried out abovely comprehensively describing with thinking the most practical and embodiment preferred of the present invention at present, but obviously, those skilled in the art can form various variants and not deviate from principle of the present invention and notion, as proposing in the claims.

Claims (38)

1. one kind contains hydrophilic A block and the amphiphilic block copolymer with hydrophobic B block of terminal hydroxyl, and the terminal hydroxyl of wherein said hydrophobic block is replaced by tocopherol or cholesterol group; Wherein hydrophilic A block is selected from polyalkylene glycol, polyvinyl alcohol, polyvinylpyrrolidone and polyacrylamide; Wherein hydrophobic B block is selected from polylactide, polyglycolide, polycaprolactone, poly-dioxane-2-ketone, poly(lactic acid)-altogether-glycollide, poly(lactic acid)-altogether-dioxane-2-ketone, poly(lactic acid)-altogether-caprolactone and polyglycolic acid-altogether-caprolactone.

2. according to the amphiphilic block copolymer of claim 1, it is A-B type Synthetic rubber, isoprene-styrene, hydrogenated, block, diblock or B-A-B type triblock copolymer.

3. according to the amphiphilic block copolymer of claim 1, the weight ratio of wherein hydrophilic A block and hydrophobic B block is in 30:70 arrives the scope of 97:3.

4. according to the amphiphilic block copolymer of claim 1, wherein the number-average molecular weight of hydrophilic block is 200 to 50,000 dalton.

5. according to the amphiphilic block copolymer of claim 1, wherein the number-average molecular weight of hydrophobic block is 50 to 50,000 dalton.

6. the amphiphilic block copolymer of following formula representative:

R _1’-O-[R _3’] _1’-[R _4’] _m’-[R _5’] _n’-C(＝O)-(CH ₂) _X，-C(＝O)-O-R _2’　　　(I’)

R wherein _{1 '}Be CH ₃-, H-[R _{5 '}] _{N '}-[R _{4 '}] _{M '}-or R _{2 '}-O-C (=O)-(CH ₂) _{X '}-C (=O)-[R _{5 '}] _{N '}-[R _{4 '}] _{M '}-;

R _{2 '}Be tocopherol or cholesterol;

R _{3 '}Be-CH ₂CH ₂-O-,-CH (OH)-CH ₂-,-CH (C (=O)-NH ₂)-CH ₂-or

R _{4 '}Be-C (=O)-and CHZ '-O-, wherein Z ' is hydrogen atom or methyl;

R _{5 '}Be-C (=O)-CHY "-O-, wherein Y " be hydrogen atom or methyl ,-C (=O)-CH ₂CH ₂CH ₂CH ₂CH ₂-O-or-C (=O)-CH ₂OCH ₂CH ₂-O-;

L ' is the integer of 4-1150;

M ' is the integer of 1-300;

N ' is the integer of O-300; With

X ' is the integer of 0-4.

7. preparation contains the method for the hydrophilic A block and the amphiphilic block copolymer of the hydrophobic B block with terminal hydroxyl, and the terminal hydroxyl of wherein said hydrophobic block is replaced by tocopherol or cholesterol group, and it comprises step:

1) carboxylation has the hydrophobic compound of tocopherol or cholesterol group; With

2) under the situation that exists dicyclohexylcarbodiimide as initiator, to contain hydrophilic A block and react, the hydrophobic compound chemical of carboxylation is attached on the terminal hydroxyl of hydrophobic B block with amphiphilic block copolymer and described carboxylation hydrophobic compound with hydrophobic B block of terminal hydroxyl; Wherein hydrophilic A block is selected from polyalkylene glycol, polyvinyl alcohol, polyvinylpyrrolidone and polyacrylamide; Wherein hydrophobic B block is selected from polylactide, polyglycolide, polycaprolactone, poly-dioxane-2-ketone, poly(lactic acid)-altogether-glycollide, poly(lactic acid)-altogether-dioxane-2-ketone, poly(lactic acid)-altogether-caprolactone and polyglycolic acid-altogether-caprolactone.

8. preparation contains the method for the hydrophilic A block and the amphiphilic block copolymer of the hydrophobic B block with terminal hydroxyl, and the terminal hydroxyl of wherein said hydrophobic block is replaced by tocopherol or cholesterol group, and it comprises step:

1) carboxylation has the hydrophobic compound of tocopherol or cholesterol group and the carboxylation hydrophobic compound that obtains with the oxalyl chloride activation; With

2) amphiphilic block copolymer and the described carboxylation hydrophobic compound that has activated that will contain hydrophilic A block and have a hydrophobic B block of terminal hydroxyl reacts, and the hydrophobic compound chemical of carboxylation is attached on the terminal hydroxyl of hydrophobic B block; Wherein hydrophilic A block is selected from polyalkylene glycol, polyvinyl alcohol, polyvinylpyrrolidone and polyacrylamide; Wherein hydrophobic B block is selected from polylactide, polyglycolide, polycaprolactone, poly-dioxane-2-ketone, poly(lactic acid)-altogether-glycollide, poly(lactic acid)-altogether-dioxane-2-ketone, poly(lactic acid)-altogether-caprolactone and polyglycolic acid-altogether-caprolactone.

9. preparation contains the method for the hydrophilic A block and the amphiphilic block copolymer of the hydrophobic B block with terminal hydroxyl, and the terminal hydroxyl of wherein said hydrophobic block is replaced by tocopherol or cholesterol group, and it comprises step:

1) mixing has the hydrophobic compound of tocopherol or cholesterol group and the dichloride that conduct connects agent;

2) will contain hydrophilic A block and join in the reaction mixture of step 1, the hydrophobic compound chemical is attached on the terminal hydroxyl of hydrophobic B block with amphiphilic block copolymer with hydrophobic B block of terminal hydroxyl; With

3) dissolving and settling step 2) segmented copolymer that obtains;

Wherein hydrophilic A block is selected from polyalkylene glycol, polyvinyl alcohol, polyvinylpyrrolidone and polyacrylamide; Wherein hydrophobic B block is selected from polylactide, polyglycolide, polycaprolactone, poly-dioxane-2-ketone, poly(lactic acid)-altogether-glycollide, poly(lactic acid)-altogether-dioxane-2-ketone, poly(lactic acid)-altogether-caprolactone and polyglycolic acid-altogether-caprolactone.

10. the pharmaceutical carrier that contains the amphiphilic block copolymer of with good grounds claim 1.

11. can form the pharmaceutical composition of polymeric micelle in the body fluid or the aqueous solution, it contains the amphiphilic block copolymer and the drugs of low aqueous solubility of with good grounds claim 1.

12. according to the composition of claim 11, wherein drugs of low aqueous solubility has 33.3mg/ml or lower water solubility.

13. be used for the polymeric composition of administration, described composition contains hydrophilic A block and has the amphiphilic block copolymer and the polylactic acid derivative of the hydrophobic B block of terminal hydroxyl, the terminal hydroxyl of wherein said hydrophobic B block is replaced by tocopherol or cholesterol group, and at least one of polylactic acid derivative is terminal covalently bound with at least one carboxyl; Wherein hydrophilic A block is selected from polyalkylene glycol, polyvinyl alcohol, polyvinylpyrrolidone and polyacrylamide; Wherein hydrophobic B block is selected from polylactide, polyglycolide, polycaprolactone, poly-dioxane-2-ketone, poly(lactic acid)-altogether-glycollide, poly(lactic acid)-altogether-dioxane-2-ketone, poly(lactic acid)-altogether-caprolactone and polyglycolic acid-altogether-caprolactone; Wherein polylactic acid derivative is selected from D, L-poly(lactic acid), D-poly(lactic acid), poly-amygdalic acid, D, the multipolymer of L-lactic acid and oxyacetic acid, D, the multipolymer of L-lactic acid and amygdalic acid, D, the multipolymer of L-lactic acid and caprolactone and D, L-lactic acid and 1, the multipolymer of 4-diox-2-ketone.

14. according to the polymeric composition of claim 13, wherein polylactic acid derivative is represented with following formula:

RO-CHZ-[A] _n-[B] _m-COOM (I)

Wherein A is-COO-CHZ-, B is-COO-CHY-,-COO-CH ₂CH ₂CH ₂CH ₂CH ₂-or-COO-CH ₂CH ₂OCH ₂, R is hydrogen atom or ethanoyl, benzoyl, decanoyl, palmitoyl, methyl or ethyl, and Z and Y all are hydrogen atom or methyl or phenyl, and M is H, Na, K or Li, and n is 1 to 30 integer, m is 0 to 20 integer.

15. according to the polymeric composition of claim 13, wherein polylactic acid derivative is represented with following formula:

RO-CHZ-[COO-CHX] _p-[COO-CHY’] _q-COO-CHZ-COOM (II)

Wherein X is a methyl; Y ' is hydrogen atom or phenyl; p is 0 to 25 integer; q is 0 to 25 integer; condition is that p+q is 5 to 25 integer; R is hydrogen atom or ethanoyl, benzoyl, decanoyl, palmitoyl, methyl or ethyl, and Z is hydrogen atom or methyl or phenyl, and M is H, Na, K or Li.

16. according to the polymeric composition of claim 13, wherein polylactic acid derivative is represented with following formula:

RO-PAD-COO-W-M’ (III)

Wherein W-M ' is

Or

PAD is selected from D; L-poly(lactic acid), D-poly(lactic acid), poly-amygdalic acid, D; the multipolymer of L-lactic acid and oxyacetic acid, D; the multipolymer of L-lactic acid and amygdalic acid, D; the multipolymer of L-lactic acid and caprolactone and D, L-lactic acid and 1, the multipolymer of 4-diox-2-ketone; R is hydrogen atom or ethanoyl, benzoyl, decanoyl, palmitoyl, methyl or ethyl, and M is H, Na, K or Li.

17. according to the polymeric composition of claim 13, wherein polylactic acid derivative is represented with following formula:

S-O-PAD-COO-Q (IV)

Wherein S is

L is-NR ₁-or-O-, R ₁Be hydrogen atom or C _1-10Alkyl, Q are CH ₃, CH ₂CH ₃, CH ₂CH ₂CH ₃, CH ₂CH ₂CH ₂CH ₃Or CH ₂C ₆H ₅, a is 0 to 4 integer, and b is 1 to 10 integer, and M is H, Na, K or Li; PAD is selected from D, L-poly(lactic acid), D-poly(lactic acid), poly-amygdalic acid, D, the multipolymer of L-lactic acid and oxyacetic acid, D, the multipolymer of L-lactic acid and amygdalic acid, D, the multipolymer of L-lactic acid and caprolactone and D, L-lactic acid and 1, the multipolymer of 4-diox-2-ketone.

18. according to the polymeric composition of claim 13, wherein polylactic acid derivative is represented with following formula:

Or

Wherein R ' is-PAD-O-C (O)-CH ₂CH ₂-C (O)-OM, PAD is selected from D, L-poly(lactic acid), D-poly(lactic acid), poly-amygdalic acid, D, the multipolymer of L-lactic acid and oxyacetic acid, D, the multipolymer of L-lactic acid and amygdalic acid, D, the multipolymer of L-lactic acid and caprolactone and D, L-lactic acid and 1, the multipolymer of 4-diox-2-ketone; M is that a is 1 to 4 integer suc as formula the group of (I) definition.

19. according to the polymeric composition of claim 13, wherein hydrophilic A block is selected from polyalkylene glycol, polyvinylpyrrolidone, polyvinyl alcohol and polyacrylamide; Hydrophobic B block is selected from polylactide, polyglycolide, poly-dioxane-2-ketone, polycaprolactone, poly(lactic acid)-be total to-glycollide, poly(lactic acid)-be total to-caprolactone, poly(lactic acid)-be total to-dioxane-2-ketone.

20. according to the polymeric composition of claim 13, wherein amphiphilic block copolymer is represented with following formula:

R _1’-O-[R _3’] _1’-[R _4’] _m’-[R _5’] _n’-C(＝O)-(CH ₂) _X’-C(＝O)-O-R _2’ (I’)

R wherein _{1 '}Be CH ₃-, H-[R _{5 '}] _{N '}-[R _{4 '}] _{M '}-or R _{2 '}-O-C (=O)-(CH ₂) _{X '}-C (=O)-[R _{5 '}] _{N '}-[R _{4 '}] _{M '}-;

R _{2 '}Be tocopherol or cholesterol;

R _{3 '}Be-CH ₂CH ₂-O-,-CH (OH)-CH ₂-,-CH (C (=O)-NH ₂)-CH ₂-or

R _{4 '}Be-C (=O)-and CHZ '-O-, wherein Z ' is hydrogen atom or methyl;

R _{5 '}Be-C (=O)-CHY "-O-, wherein Y " be hydrogen atom or methyl ,-C (=O)-CH ₂CH ₂CH ₂CH ₂CH ₂-O-or-C (=O)-CH ₂OCH ₂CH ₂-O-;

L ' is the integer of 4-1150;

M ' is the integer of 1-300;

N ' is the integer of 0-300; With

X ' is the integer of 0-4.

21. according to the polymeric composition of claim 13, the number-average molecular weight of wherein hydrophilic A block and hydrophobic B block is respectively in 200 to 50,000 dalton and 50 to 50,000 daltonian scopes.

22. according to the polymeric composition of claim 13, wherein hydrophilic A block and the weight ratio of hydrophobic B block in amphiphilic block copolymer are 30:70 to 97:3.

23., wherein contain and account for amphiphilic block copolymer that the composition gross weight is 0.1 to 99.9 weight % and the polylactic acid derivative of 0.1 to 99.9 weight % according to the polymeric composition of claim 13.

24. according to the polymeric composition of claim 13, wherein the number-average molecular weight of polylactic acid derivative is 50 to 50,000 dalton.

25. according to the polymeric composition of claim 13, wherein polylactic acid derivative is by condensation reaction and then with in yellow soda ash, sodium bicarbonate, saleratus or the salt of wormwood and the form of resulting sodium or sylvite under the situation that does not have catalyzer.

26. according to the polymeric composition of claim 13, the carboxyl terminal of wherein relative 1 molar equivalent polylactic acid derivative also contains two or trivalent metal ion of 0.01 to 10 molar equivalent.

27. according to the polymeric composition of claim 26, wherein two or trivalent metal ion be selected from Ca ²⁺, Mg ²⁺, Ba ²⁺, Cr ³⁺, Fe ³⁺, Mn ²⁺, Ni ²⁺, Cu ²⁺, Zn ²⁺And Al ³⁺

28. by micella or nanoparticle according to the preparation of the polymeric composition of claim 13.

29. according to the micella or the nanoparticle of claim 28, wherein the particle diameter of micella or nanoparticle arrives in the scope of 400nm 1.

30. pharmaceutical composition that contains 70 to 99.9 weight % according to the drugs of low aqueous solubility of the polymeric composition of claim 13 and 0.1 to 30 weight %.

31. pharmaceutical composition that contains 70 to 99.9 weight % according to the drugs of low aqueous solubility of the polymeric composition of claim 26 and 0.1 to 30 weight %.

32. a method for preparing the polymeric composition that contains drugs of low aqueous solubility, it may further comprise the steps:

1) with hydrophilic A block with have the amphiphilic block copolymer of hydrophobic B block of terminal hydroxyl and polylactic acid derivative and drugs of low aqueous solubility are dissolved in the organic solvent, the terminal hydroxyl of wherein said hydrophobic B block is replaced by tocopherol or cholesterol group, terminal and at least one the carboxyl covalent attachment of at least one of polylactic acid derivative; With

2) evaporation organic solvent wherein to wherein adding the aqueous solution, contains the polymeric micelle of drugs of low aqueous solubility with formation;

Wherein hydrophilic A block is selected from polyalkylene glycol, polyvinyl alcohol, polyvinylpyrrolidone and polyacrylamide; Wherein hydrophobic B block is selected from polylactide, polyglycolide, polycaprolactone, poly-dioxane-2-ketone, poly(lactic acid)-altogether-glycollide, poly(lactic acid)-altogether-dioxane-2-ketone, poly(lactic acid)-altogether-caprolactone and polyglycolic acid-altogether-caprolactone; Wherein polylactic acid derivative is selected from D, L-poly(lactic acid), D-poly(lactic acid), poly-amygdalic acid, D, the multipolymer of L-lactic acid and oxyacetic acid, D, the multipolymer of L-lactic acid and amygdalic acid, D, the multipolymer of L-lactic acid and caprolactone and D, L-lactic acid and 1, the multipolymer of 4-diox-2-ketone.

33. according to the method for claim 32, wherein also comprise in the polymeric micelle of drugs of low aqueous solubility add two or trivalent metal ion with the fixing step of polylactic acid derivative C-terminal.

34. according to the method for claim 32, wherein organic solvent is selected from acetone, ethanol, methyl alcohol, ethyl acetate, acetonitrile, methylene dichloride, chloroform, acetic acid and diox.

35. pharmaceutical composition as carcinostatic agent, amphiphilic block copolymer and polylactic acid derivative and cancer therapy drug that it contains hydrophilic A block and has the hydrophobic B block of terminal hydroxyl, the terminal hydroxyl of wherein said hydrophobic B block is replaced by tocopherol or cholesterol group, terminal and at least one the carboxyl covalent attachment of at least one of polylactic acid derivative;

Wherein hydrophilic A block is selected from polyalkylene glycol, polyvinyl alcohol, polyvinylpyrrolidone and polyacrylamide; Wherein hydrophobic B block is selected from polylactide, polyglycolide, polycaprolactone, poly-dioxane-2-ketone, poly(lactic acid)-altogether-glycollide, poly(lactic acid)-altogether-dioxane-2-ketone, poly(lactic acid)-altogether-caprolactone and polyglycolic acid-altogether-caprolactone; Wherein polylactic acid derivative is selected from D, L-poly(lactic acid), D-poly(lactic acid), poly-amygdalic acid, D, the multipolymer of L-lactic acid and oxyacetic acid, D, the multipolymer of L-lactic acid and amygdalic acid, D, the multipolymer of L-lactic acid and caprolactone and D, L-lactic acid and 1, the multipolymer of 4-diox-2-ketone.

36. according to the pharmaceutical composition of claim 35, the carboxyl terminal of wherein relative 1 molar equivalent polylactic acid derivative also contains two or trivalent metal ion of 0.01 to 10 molar equivalent.

37. pharmaceutical composition that is used for the treatment of the resistance cancer, amphiphilic block copolymer and polylactic acid derivative and cancer therapy drug that it contains hydrophilic A block and has the hydrophobic B block of terminal hydroxyl, the terminal hydroxyl of wherein said hydrophobic B block is replaced by tocopherol or cholesterol group, terminal and at least one the carboxyl covalent attachment of at least one of polylactic acid derivative;

Wherein hydrophilic A block is selected from polyalkylene glycol, polyvinyl alcohol, polyvinylpyrrolidone and polyacrylamide; Wherein hydrophobic B block is selected from polylactide, polyglycolide, polycaprolactone, poly-dioxane-2-ketone, poly(lactic acid)-altogether-glycollide, poly(lactic acid)-altogether-dioxane-2-ketone, poly(lactic acid)-altogether-caprolactone and polyglycolic acid-altogether-caprolactone; Wherein polylactic acid derivative is selected from D, L-poly(lactic acid), D-poly(lactic acid), poly-amygdalic acid, D, the multipolymer of L-lactic acid and oxyacetic acid, D, the multipolymer of L-lactic acid and amygdalic acid, D, the multipolymer of L-lactic acid and caprolactone and D, L-lactic acid and 1, the multipolymer of 4-diox-2-ketone.

38. according to the pharmaceutical composition of claim 37, the carboxyl terminal of wherein relative 1 molar equivalent polylactic acid derivative also contains two or trivalent metal ion of 0.01 to 10 molar equivalent.

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