CN102085189A - Docetaxel liposome sterile lyophilized preparation and preparation method thereof - Google Patents
Docetaxel liposome sterile lyophilized preparation and preparation method thereof Download PDFInfo
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Abstract
The invention relates to a docetaxel liposome sterile lyophilized preparation and preparation method thereof, and the docetaxel liposome sterile lyophilized preparation comprises docetaxel, natural lecithin, PEG-DSPE, cholesterol, a protective agent for lyophilization, and buffer salts. The natural lecithin, PEG-DSPE, cholesterol, and docetaxel are dissolved in a organic solvent; a buffer solution containing the protective agent for lyophilization is added into mixed solution; the solution is incubated for 10-60 min for decreasing the average particle size of the liposome to 50-200 nm; and the preparation is obtained after bacilli-eliminated filtration and lyophilization. The docetaxel liposome prepared by the invention has the advantages of less toxic and side effect and higher curative effect when compared with commercially available ordinary injections at present. The preparation method of the invention is simple and practical, and is applicable to industrial production; and the prepared docetaxel liposome has high encapsulation efficiency and good stability.
Description
Technical field
The invention belongs to field of pharmaceutical preparations, be specifically related to liposome sterile freeze-drying preparation of a kind of fat-soluble medicine docetaxel and preparation method thereof.
Background technology
The France peaceful and comfortable company of rhone-poulenc is another novel anti microtubule medicine behind paclitaxel in successful docetaxel (also the claiming Docetaxel) injection of research and development in 1986, at first go on the market in Mexico April nineteen ninety-five, subsequently in country's listings such as English, U.S., method, meaning, moral, days.
Compare with paclitaxel, the interior outflow of docetaxel cell is less and low dosage cell death inducing effect is remarkable.Many external and clinical researches confirm that all docetaxel has radiosensitizing effect to kinds of tumor cells and do not increase the weight of the radiation injury of normal structure.The affinity of docetaxel is 2 times of paclitaxel, in external active anticancer test, the docetaxel activity can reach 10 times of paclitaxel, in addition, docetaxel also has bioavailability preferably, higher IC, better water-solubility and excellent anticancer broad spectrum activity, it not only has better curative effect to breast carcinoma, pulmonary carcinoma, and the patient who suffers from head and neck cancer, gastric cancer, cancer of pancreas and soft tissue neoplasms is also had therapeutical effect preferably.In addition, docetaxel and some antitumor drug are united when using, and also have synergism.The same with paclitaxel, docetaxel is also water insoluble, the Docetaxel for Injection of clinical usefulness is the injection concentrated solution, be 2 cillin bottle packings, wherein one bottle is the tween solution of docetaxel, and another bottle is an ethanol water, earlier ethanol water is injected the tween solution of docetaxel during clinical practice, fully rock, observe just can be applied to the patient after clear and bright after 5 minutes, clinical practice is very inconvenient.And docetaxel injection during medication and uses up at the appointed time in case dilution just must be used immediately, promptly has docetaxel to separate out otherwise place a few hours, is filtered by online filter to cause the drug effect reduction.So just press for the docetaxel dosage form that exploitation makes new advances,, avoid because anaphylaxis and other toxicities that double solvent brings makes things convenient for clinical practice to increase its water solublity.
Liposome is a kind of novel form of targeting drug delivery system, it is one of focus of pharmaceutics research always, it can with drug selectivity be transported to tumor locus, the performance therapeutical effect, simultaneously do not influence the function of normal cell, tissue or organ again, thereby reach the purpose that improves curative effect, reduces toxic and side effects.Liposome has the structure that is similar to cell membrane, constitutes the film forming inner surface of the hydrophilic head shape of lipoid of bilayer, and lipophilic afterbody then is in the centre of film.Just because this membranelike structure of liposome makes it can carry various hydrophilic, hydrophobic and amphipathic materials, their are wrapped into the liposome interior water, insert the surface that class lipid bilayer or absorption are connected liposome.
Liposome had both played the protective effect to medicine as pharmaceutical carrier, had improved the targeting of medicine to the body specific part again, therefore had a lot of superior characteristic aspect the raising drug effect.Mainly show the following aspects: 1. liposome is nontoxic or toxic and side effects is little to body, and its lipid bilayer and biomembrane have bigger similarity and tissue intersolubility, thereby are easy to be organized absorption, have improved the absorption rate of medicine; 2. wrapping kmedicine by liposome is a physical process, does not change drug molecular structure, can not destroy ingredient, and the medicine of parcel then can be avoided being destroyed by hydrolytic enzyme in the body; 3. the medicine behind the parcel can reduce its toxicity to the body specific part, reduces the drug use amount, makes medicine have slow release and controlled-release function; 4. different with viral vector, liposome can biodegradation in host, non-immunogenicity; 5. in gene therapy, be easy to gene compoundly, higher targeting is arranged, realize the treatment of specific cells; 6. the material cheapness is convenient to a large amount of preparations.
In the nearly more than ten years the many pieces of bibliographical informations about docetaxel liposome have been arranged, but coming into the market of neither one product success so far.Trace it to its cause, the one, because prepared not good, the less stable of docetaxel liposome quality, can't long preservation; The 2nd, the docetaxel liposome drug loading of preparation is lower, needs a large amount of adjuvants such as phospholipid of infusion simultaneously during clinical use; The 3rd, complicated process of preparation only is suitable for laboratory and prepares on a small scale, inapplicable large-scale industrial production.At present, Chang Yong method for preparing lipidosome comprises following several method:
(1) film dispersion method: film material and fat-soluble medicine are dissolved in the organic solvent,, make it on container inner wall, form exsiccant thin film, water soluble drug is dissolved in the buffer, add in the container and constantly stir, promptly get liposome then with organic solvent evaporation.If expect the liposome that particle diameter is littler, then need to cooperate the appropriate postprocessing means, as ultrasonic, high pressure homogenize and extruding etc.
(2) injection method: lipoids such as phospholipid and fat-soluble medicine are dissolved in the organic solvent, then this medicinal liquid are slowly injected the buffer (or containing water soluble drug) of uniform temperature through syringe, the limit edged stirs, and constantly is stirred to solvent to eliminate, promptly.Preparation technology is simple for this method, easy operating, but liposome quality instability.
(3) transmembrane gradient active loading method: the transmembrane gradient active loading method comprises pH gradient method and ammonium sulphate gradient.The pH gradient method is by regulating the pH of water inside and outside the liposome, make it to form certain pH gradient difference, utilize the difference of medicine dissociated state in different pH solution, sees through outer water with molecule-type and forms ionic drug and encapsulated at interior aqueous phase.Ammonium sulphate gradient is earlier ammonium sulfate to be wrapped in water in the liposome, removes the ammonium sulfate of outer water then by the method for dialysis, gel chromatography or ultrafiltration.It is to be diffused into outside the liposome by free ammonia, forms the pH gradient, and it is intravital to make medicine gather lipid.The transmembrane gradient active loading method is suitable for some faintly acid and weakly basic drugs, is unsuitable for insoluble drug and neutral medicine.
(4) ultrasonic dispersing method: on the basis of film dispersion method,, can obtain unilamelar liposome (SUV) again through ultrasonic Treatment.Ultrasonic technique has two kinds: a kind of is that the probe of Ultrasound Instrument is immersed in the liposome dispersion, and the method is to prepare the most widely used method of SUV on a small scale; Another is that sample is placed in test tube or the beaker, places the water-bath type Ultrasound Instrument again.Because the release of ultrasonic probe place energy can cause hot-spot, therefore cooling or cooling measure must be arranged.
(5) freeze-drying (lyophilizing rehydration method): freeze-drying is about to the lipoid high degree of dispersion in aqueous solution, lyophilization, and then be distributed in the aqueous mediator of pastille, form liposome.Within the specific limits, the envelop rate of liposome and granularity slightly increase with the increase of number of freezing and thawing; Stir after the freeze thawing or ultrasonicly all can make accumulative liposome depolymerization, the angle that never influences envelop rate is considered, stirs better than supersound process.It is to be prepared from by prefabricated vesicle that lyophilizing rehydration vesicle also is called lyophilizing rehydration liposome (FRV).This method envelop rate is very high, even also can realize higher entrapment for macromole.Drying process makes lipid bilayer and waits that wrapping medicine closely contacts, again can be bigger to the molecule sealing machine that is attached on the lipid during swelling.
(6) reverse evaporation (REV): Szoka equals the preparation technology that proposed " REV liposome " in 1978, the liposome that the water volume is big and envelop rate is high in promptly making by reverse evaporation.This method is a breakthrough of liposome technology, because it considers that first preparation has the liposome of high interior water volume-lipid ratio characteristic, and can seal the most existing water-soluble substances.The REV liposome can be made by various lipids and lipid mixture, and water volume-lipid is higher about 30 times than SUV in it, and the vesicle that makes than multilamellar liposome or hand method is high 4 times.Under low salt concn and optimum condition, there is 65% water to be encapsulated in the vesicle, in addition also very high to the envelop rate of macromolecular substances.The basis of this method is the formation of " reverse micelle ", promptly is scattered in a large amount of organic solvents by the stable little water droplet of phospholipid monolayer.This reverse micelle is that the mixture by buffering water and organic facies forms through ultrasonic, wherein cushion water and contain water soluble molecules of going into liposome to be encapsulated, and organic facies is dissolved with amphipathic phospholipid.Slowly remove organic mutual-assistance reverse micelle and change the sticky gel of formation.In the critical point of this process, gel subsides, and the part reverse micelle breaks.Reverse micelle breaks and the excessive phospholipid that produces, forms complete bilayer conversely around the micelle of remainder, and forming vesicle is the REV liposome.The REV liposome mainly is a single chamber, is to be made of several concentric bilayers but some vesicles are all arranged in each REV preparation, thereby forms few layer vesicle.The surface tension between the size of REV liposome and phospholipid type and the dissolubility in organic solvent, water buffer and the organic solvent and the factors such as relative quantity of water, organic solvent and lipid are relevant.
At present, more about the research report of docetaxel liposome, prescription composition and preparation technology are varied.Docetaxel liposome content of dispersion as reports such as MariaL.I. is lower, poor stability, can't long preservation, be not suitable for clinical practice (preparation, characterization, cytotoxicity and pharmacokinetics of liposomes containing docetaxel.Journal of controlled release 91 (2003), 417-429.)
CN200510029634.0 discloses a kind of polyene taxol liposome and preparation method thereof.But place less stable under the liquid lipidosome room temperature of the method preparation.And preparation technology is comparatively complicated, needs long-time the steaming to remove organic solvent, and this process causes phospholipid oxidation to produce lysophosphatide easily, is not suitable for suitability for industrialized production.
CN200510110973.1 discloses a kind of taxane liposome lyophilized composition and preparation method thereof, has solved the stability problem after the redissolution that improves the taxane liposome pharmaceutical composition at low cost.But added cyclodextrin in the liposome of its preparation prescription, in clinical practice, had certain risk.
CN1846692A and CN101057831A disclose Docetaxel long circulating liposomes dosage form and preparation method thereof, have significantly improved liposome circulation time in vivo, have reduced the toxicity of medicine.But the stability of the Liposomal formulation after it does not redissolve to lyophilizing is investigated the envelop rate and the change of size situation of particularly redissolving the back medicine.In addition, the present invention once according to its disclosed method with outer addition with sucrose, glucose and mannitol as freeze drying protectant, but can't obtain the preferable freeze-drying prods of outward appearance, and redissolve the back liposome and can't rebuild, drug leakage is serious.
CN101015547A discloses a kind of docetaxel liposome formulation and preparation method thereof, but its sterilization method adopts 121 degrees centigrade of autoclavings, and this can cause a large amount of oxidations of phospholipid undoubtedly, and medicine stability under this violent condition is difficult to guarantee.In addition, the stability of formulation after the sterilization is not investigated yet.
CN101317816A discloses a kind of docetaxel long-circulation formulation, and as solvent, toxicity is bigger with chloroform, needs to revolve for a long time steaming in addition to remove residual solvent, and this is difficult to carry out in suitability for industrialized production.And the liposome before and after the lyophilizing is not carried out study on the stability.
CN101322699A and CN101322689A disclose the preparation method of a kind of docetaxel long-circulating liposome and freeze-dried powder thereof.Adopted charged synthetic phospholipid as the film material, these phospholipid are imported product, cost an arm and a leg and majority is not the injection stage pharmaceutic adjuvant.It is not oxidized with protection phospholipid to add antioxidant in the prescription.In addition, complicated process of preparation is not suitable for suitability for industrialized production, particularly with chloroform methanol as solvent, toxicity is difficult to more greatly remove fully, safety can not get guaranteeing.
Summary of the invention
At the deficiencies in the prior art, main purpose of the present invention provides and a kind ofly can obviously reduce the docetaxel liposome sterile freeze-drying preparation that existing docetaxel injection toxic and side effects, preparation technology are simple, be easy to industrial amplification production.Said preparation entrapment efficiency height, simple to operate is a kind of nanometer colloid system that can stablize placement in solution, and this preparation can steady in a long-termly be placed after lyophilization, and the main character in lyophilized formulations redissolution back remains unchanged.
The phospholipid of selecting for use in the prepared docetaxel liposome preparation of the present invention is natural phospholipid, and is cheap, be easy to obtain.Content that the more important thing is phosphatidylcholine in the selected natural phospholipid is moderate, and this stability and sealing of medicine for final Liposomal formulation all is very favorable.
Cholesterol is significant for Liposomal formulation, and how many one side of its content have determined liposome stability in vivo on the other hand medicine release behavior in vivo to be had material impact.The present invention has suitably reduced content of cholesterol on the basis of existing bibliographical information, this is significant for the present invention.Improved the drug loading of fat-soluble medicine in lipid bilayer by reducing content of cholesterol on the one hand, guaranteed that on the other hand Liposomal formulation is in external and intravital stability.
Prepared its mean diameter of Liposomal formulation of the present invention is about 100 nanometers, and this has guaranteed that on the one hand Liposomal formulation can not removed rapidly by reticuloendothelial system in vivo, helps gathering, the release of antitumor drug at tumor locus on the other hand.
Another object of the present invention provides the preparation method of above-mentioned docetaxel liposome sterile freeze-drying preparation.This preparation method biggest advantage is to need not other step after various film materials are dissolved in the organic solvents such as the adequate amount of ethanol or the tert-butyl alcohol at a certain temperature to remove organic solvent and can enter next step operation.Why select the ethanol or the tert-butyl alcohol to be because these solvent toxicity are less, need not painstakingly to steam to remove, only need step of freeze drying can reach the requirement of dissolvent residual.The selected film material of the present invention is easy to dissolving in the ethanol or the tert-butyl alcohol in addition.This preparation method is simple and be suitable for large-scale production.
The term explanation:
The DSPE of PEGization (PEG-DSPE): the DSPE of Pegylation is a kind of long circulation film material;
MPEG
2000-DSPE: methoxy poly (ethylene glycol) 2000 DSPE;
But above raw material market is buied, and also can prepare voluntarily by prior art.
Being described in detail as follows of technical solution of the present invention:
Technical scheme of the present invention is at first to select one or more phospholipid that are suitable for the character of docetaxel medicine own and medicine and phospholipid are dissolved in the suitable quantity of water solubleness organic solvent fully dissolving under suitable temperature and stirring condition.In above-mentioned solution, add an amount of buffer salt solution that contains excipient and freeze drying protectant, after fully mixing under suitable temperature and the stirring condition, can obtain rough liposome turbid liquor.Lipid granule in this suspension has bigger particle size distribution, and the particle diameter of particle is between between tens nanometers to tens micron.Select suitable technology the particle diameter of particle can be reduced to below the 1um afterwards, preferably, mean diameter is controlled at below the 200nm.By above-mentioned steps, can obtain the docetaxel liposome preparation, this preparation, can be stablized under the 2-8 ℃ of condition and place more than 12 months more than 90 days and do not assemble, precipitate through placement that can be at room temperature stable after the aseptic filtration.For further improving stability of formulation, being convenient to preserve and clinical practice, can select cryodesiccated mode that above-mentioned preparation is handled.Docetaxel liposome preparation after lyophilization can steady in a long-termly be placed, and character does not have obvious change after redissolving.
Used main adjuvant is a phospholipid in the preparation process of liposome, comprises natural phospholipid such as soybean phospholipid, egg yolk lecithin, cuorin and sphingomyelins etc.; Also comprise various synthetic or semi-synthetic phospholipid such as hydrogenated soya phosphatide, hydrogenation egg yolk lecithin, dioleoyl phospholipid phatidylcholine (DOPC), dimyristoyl phosphatidyl choline (DMPC), dipalmitoyl phosphatidyl choline (DPPC), distearoyl phosphatidylcholine (DSPC), dilinoleoylphosphatidylcholine (DLPC) etc.; Can also be to have the phospholipid of electric charge such as DOPG (DOPG), dioleoyl phospholipid acyl serine (DOPS), GLYCEROL,DIMYRISTOYL PHOSPHATIDYL (DMPG) etc.Phospholipid used in the present invention can be one or more the compositions in the above-mentioned phospholipid, wherein preferably uses natural phospholipid, and the content of these its phosphatidylcholines of natural phospholipid is preferably more than more than 70% greater than more than 50%.Phospholipid that the present invention is selected and antitumor drug docetaxel have certain ratio, and this ratio is between 1: 100 to 1: 10 (weight ratio), preferably between 1: 50 to 1: 10.
Also added cholesterol in the liposome of the present invention, it is more stable that the adding of cholesterol can make lipid bilayer in the liposome, be higher than the flowability that can reduce liposome membrane under the situation of phase transition temperature, and be lower than the flowability that can increase film under the prerequisite of phase transition temperature.In the environment, the existence of cholesterol can reduce the removing speed of liposome on the one hand in vivo, also helps the slow release of medicine on the other hand.Moreover, cholesterol can also reduce the generation of free radical in liposome, thereby has reduced oxidation level, prevents the generation of hemolytic phospholipid.But for fat-soluble medicine, the affiliation that adds of cholesterol reduces the ability that medicine embeds the liposome bilayer, thereby reduces the drug loading of liposome.So the addition of cholesterol needs well-designed.The present invention is by repeatedly experiment discovery, and when the addition of cholesterol surpassed 5% (weight ratio) of phospholipid weight, precipitation will appear in the Liposomal formulation of preparation at short notice.Therefore the addition of cholesterol is between the 0-5% (weight ratio) of phospholipid weight among the present invention.
In order to improve the stability of liposome, circulation time in vivo particularly, increase the half-life of medicine, in liposome, add an amount of long recycled material such as the phospholipid of sphingomyelins, ganglioside (GM1) and Polyethylene Glycol (PEG) derivatization also suits.The present invention has adopted in liposome the phospholipid that adds PEGization to prolong the half-life in the body of liposome as the mode of (PEG-DSPE, the DSPE of Pegylation), raising oncotherapy effect.Wherein, the addition of PEG-DSPE need keep certain ratio with the amount of phospholipid, and this ratio can be 0-50% (weight ratio), preferably selects 0-30%.If select 0, mean that then this liposome is a conventional liposome, does not have long circulatory function.
The preparation method of liposome also is diversified, comprises that the various preparation methoies of modes such as film evaporation method, injection method, ultrasonic dispersion, freeze-drying, reverse evaporation all are suitable for the present invention.But because the prepared liposome of these methods differs greatly at aspects such as stability, envelop rate, drug loading, therefore the liposome mass discrepancy for preparing is bigger, sometimes even second-rate situation occurs.In order to prepare the higher liposome of quality, simplify preparation technology simultaneously and make it be applicable to industrial-scale production, the present invention has adopted a kind of preparation technology of improvement.Specifically, at first an amount of phospholipid, cholesterol, docetaxel are dissolved in the organic solvent that dissolves each other with water jointly, this organic solvent can be one or more the compositions in ethanol, the tert-butyl alcohol, methanol, ether, dichloromethane, the chloroform etc.Because the present invention does not need to remove organic solvent, therefore selecting the less ethanol of toxicity and the tert-butyl alcohol is preferable selection.For better dissolving, can pass through elevated temperature and stirring or ultransonic mode.In addition, the addition of organic solvent also is to need to select meticulously, the addition of organic solvent of the present invention and final preparation volume have dependency, the addition of organic solvent should not surpass 20% of final preparation volume specifically, wherein again to be no more than 10% for good, if for example prepare volume 1000ml, then the addition of organic solvent is good to be not more than 100ml.
After the above-mentioned steps, can resemble prior art report select rotary evaporation or other suitable technology that organic solvent is removed.But need not this step among the present invention, this has simplified preparation technology greatly, has shortened manufacturing cycle, is particularly suitable for mass preparation.Then, can directly the buffer salt solution for preparing be added in the solvent that is dissolved with medicine and phospholipid and hatch.This moment incubation temperature be between 30-70 ℃, with 40-60 ℃ be the best, incubation time is 10-60 minute.Above-mentioned buffer salt solution has multiple choices, can be phosphate buffer, acetate buffer, glycine buffer night or the like, and pH is between 3.0-8.0, between the preferred 4.0-7.0.In addition, carry out for the ease of cryodesiccated, it also is essential adding an amount of freeze drying protectant in the buffer salt solution.These freeze drying protectants can be one or more glucides, are good with monosaccharide and disaccharidase wherein, as glucose, sucrose, maltose, trehalose, lactose, galactose etc.; Also can be other non-glucides such as mannitol, albumin, glycine etc.One or more materials in the above-mentioned freeze drying protectant have been selected among the present invention.The w/v of freeze drying protectant in buffer is good with 2-20%, preferably between 5-15%.
The suspension that forms after above-mentioned incubation step is the thick suspension of liposome, and the liposome in this suspension mostly is multilamelar liposome or multilamellar liposome, and particle diameter is also bigger, generally between between tens nanometers to tens micron.This suspension is not suitable as the intravenous fluid liposome, and is also unstable.Therefore need further processing, can extrude by high pressure specifically, mode such as ultrasonic, high-speed stirred further reduces particle diameter with this suspension and improve stability.The present invention has adopted microjet equipment that thick suspension is processed, and pressure is selected between 1000-20000psi, and cycle-index is between 2-20.Preferably technology is to carry out 2-10 microjet circulation through low pressure (2000-6000psi) earlier, passes through high pressure (7000-20000psi) again and carries out 2-10 microjet circulation.So after the technology, the mean diameter of the liposome of preparation can be reduced to below the micron, if process choice is suitable, mean diameter can be reduced between the 50-500nm.
The placement that the liposome of preparation can be stable under room temperature after the filtration sterilization step by step after above-mentioned steps is more than 90 days and tangible quality can not take place change.
Above-mentioned liposome solutions can be directly used in intravenous injection after aseptic filtration.In order further to improve stability, can select lyophilization or spray-dired mode to remove moisture.The moisture of final preparation is controlled at below 3%.
Preferably, in the docetaxel liposome sterile freeze-drying preparation of the present invention, contain following component in per 100 gram docetaxel liposome sterile freeze-drying preparations:
This docetaxel liposome sterile freeze-drying preparation makes as follows:
Take by weighing natural phosphatidyl choline, mPEG
2000-DSPE and cholesterol are dissolved in an amount of organic solvent, and fully the dissolving back adds the docetaxel of recipe quantity, adds buffer, hatches under suitable temperature and heat-retaining condition 10-60 minute; By the known method of those skilled in the art the mean diameter of liposome is reduced to 50-200nm; Through filtering with microporous membrane degerming postlyophilization promptly.
Preferably, the detailed preparation process of Liposomal formulation of the present invention is summarized as follows:
(1) take by weighing the natural phosphatidyl choline of recipe quantity, the DSPE of PEGization (PEG-DSPE) and cholesterol and be dissolved in the organic solvent, stirring and dissolving gets phospholipid solution under 40-60 ℃ of condition;
(2) docetaxel that takes by weighing recipe quantity is dissolved in the above-mentioned phospholipid solution, fully mixing;
(3) adopt the water for injection preparation to contain the buffer salt solution of freeze drying protectant, regulate between the pH to 4.0-7.0; And it is added in the solution that above-mentioned steps (2) makes, keep 40-60 ℃ of temperature, stir or ultrasound condition under hatched 10-60 minute suspension;
(4) suspension that step (3) is obtained is added in the microjet instrument, and adjusting pressure is 2000-6000psi, and after circulation 2-10 time, adjusting pressure is 7000-20000psi, and circulation 2-10 time obtains liposome solutions;
(5) liposome solutions that step (4) is obtained filters, gets docetaxel liposome step by step through 0.8um, 0.45um, 0.22um filter membrane successively; Packing under aseptic condition, lyophilization promptly get the docetaxel liposome sterile freeze-drying preparation.
Above-mentioned organic solvent is selected from one or both the compositions in ethanol, the tert-butyl alcohol, the consumption of organic solvent be no more than final preparation the liquid preparation volume 20%, be preferably and be no more than 10%.
Above-mentioned natural phosphatidyl choline is natural soybean lecithin or Ovum Gallus domesticus Flavus lecithin.Preferably, in described soybean lecithin or Ovum Gallus domesticus Flavus lecithin, the content of phosphatidylcholine is greater than 70wt%, and the content of hemolytic phospholipid is less than 3wt%.
Above-mentioned freeze drying protectant is one or more the compositions in maltose, mannitol, galactose, lactose, glucose, sucrose, the trehalose.
Above-mentioned buffer salt is selected from phosphate, acetate or glycinate.
The all employing prior aries that do not specify in the above-mentioned preparation method.
The prepared docetaxel liposome sterile freeze-drying preparation of the present invention adds normal saline, glucose injection or water for injection and redissolves the back mean diameter between 50-200nm, and entrapment efficiency is greater than 95%, and pH is between 4.0-7.0.
The lyophilized formulations of method for preparing can be stablized under 2-8 ℃ of condition and places more than 2 years and obvious change do not take place.After this lyophilized formulations redissolved, mean diameter was compared no significant change before with lyophilizing, but the distribution of particle diameter is more concentrated, and main quality index such as the envelop rate of medicine, stability, drug loading, zeta current potential is compared also not obviously change before with lyophilizing.
Compared with prior art, major advantage of the present invention comprises:
1. aseptic freeze-dried Liposomal formulation provided by the present invention with the dilution of normal saline or glucose injection after direct intravenously administrable, do not contain toxic organic solvent and other solubilizing agents, can effectively reduce the untoward reaction that normal injection is brought.The liposome mean diameter helps the release of entrapped drug at tumor locus less than 200nm, therefore has certain tumor-targeting;
2. good stability.The present invention has selected the moderate natural phospholipid of phosphatidylcholine content for use, adds cholesterol and fat-soluble medicine with specific proportioning, and with about these docetaxel liposome mean diameter 100 nanometers that prepare, the medicine that surpasses more than 95% is encapsulated in the liposome.Can stablize placement under the liquid condition 90 days and phenomenons such as drug leakage, liposome gathering, precipitation do not take place.The liposome particle size distribution is more concentrated after lyophilizing, and can stablize and place 2 years;
3. preparation is simple, be suitable for amplifying a difficult problem that has overcome the prior art processes complexity, has been difficult to extensive amplification.Particularly need not in the liposome preparation process to use the bigger organic solvent of toxicity also to need not to remove this step of organic solvent, so the restriction that the manufacturing cycle of sample is short, scale is not subjected to appointed condition;
4. both can prepare the conventional liposome preparation, and also can prepare long circulating liposomes (hidden liposome) by the mode of adding long recycled material in prescription, both are no significant difference aspect main quality index such as envelop rate, drug loading, percolation ratio, particle diameter;
5. animal test results shows, the docetaxel liposome of the present invention's preparation is compared with present commercially available normal injection, has the advantage that toxic and side effects is little, curative effect is high.
The specific embodiment
Further specify the present invention below in conjunction with embodiment, but be not limited thereto.
The preparation of embodiment 1 docetaxel long-circulating liposome
Take by weighing soybean lecithin 6 grams, mPEG
2000-DSPE 1 gram, cholesterol 0.15 gram are dissolved in the 10ml tert-butyl alcohol, and stirring and dissolving under 55 ℃ of conditions obtains phospholipid solution.Taking by weighing docetaxel 0.2 gram is dissolved in the phospholipid solution.Compound concentration is the phosphate buffer of 10mM, transfers pH to 4.5, adds mannitol 4 grams, maltose 8 grams, fully stirring and dissolving.Measure the 100ml buffer and be added in the phospholipid solution, stir under 50 ℃ of conditions and hatched 30 minutes, obtain the thick suspension of liposome.This thick suspension is added in the microjet instrument 5000psi circulation 5 times, 15000psi circulation 8 times.Behind the membrane filtration of 0.8um, 0.45um and 0.22um, promptly obtain docetaxel long-circulating liposome respectively.
After measured, according to the liposome mean diameter 85nm that above-mentioned steps obtains, drug loading is 2mg/ml, and entrapment efficiency is 99%.Place after 90 days under the room temperature and detect particle diameter, drug loading, envelop rate and related substance, all do not have obviously change.
To redissolve after the above-mentioned liposome solutions lyophilizing, mean diameter is 95nm after testing, and drug loading is 2mg/ml, and entrapment efficiency is 99%.Liposome after the lyophilizing is placed after 18 months and is detected, and main quality index does not all have obvious change.
The preparation of embodiment 2 common docetaxel liposomes
Take by weighing soybean phospholipid 5 gram, cholesterol 0.1 gram is dissolved in the 10ml ethanol, stirring and dissolving under 55 ℃ of conditions obtains phospholipid solution.Taking by weighing docetaxel 0.3 gram is dissolved in the phospholipid solution.Compound concentration is the phosphate buffer of 10mM, transfers pH to 4.5, adds mannitol 3 grams, maltose 6 grams, fully stirring and dissolving.Measure the 100ml buffer and be added in the phospholipid solution, stir under 50 ℃ of conditions and hatched 30 minutes, obtain the thick suspension of liposome.This thick suspension is added in the microjet instrument 5000psi circulation 3 times, 15000psi circulation 5 times.Behind the membrane filtration of 0.8um, 0.45um and 0.22um, promptly obtain docetaxel liposome respectively.
After measured, according to the liposome mean diameter 80nm that above-mentioned steps obtains, drug loading is 3mg/ml, and entrapment efficiency is 98.9%.Place after 90 days under the room temperature and detect particle diameter, drug loading, envelop rate and related substance, all do not have obviously change.
To redissolve after the above-mentioned liposome solutions lyophilizing, mean diameter is 90nm after testing, and drug loading is 3mg/ml, and entrapment efficiency is 99.2%, and ethanol content is less than 0.1%, moisture 1.5%.Liposome after the lyophilizing is placed after 24 months and is detected, and main quality index does not all have obvious change.
Embodiment 3 does not contain the preparation of the docetaxel liposome of cholesterol
Take by weighing soybean lecithin 60 grams and be dissolved in the 100ml tert-butyl alcohol, stirring and dissolving under 55 ℃ of conditions obtains phospholipid solution.Taking by weighing docetaxel 1.5 grams is dissolved in the phospholipid solution.Compound concentration is the phosphate buffer of 10mM, transfers pH to 4.5, adds mannitol 50 grams, maltose 100 grams, fully stirring and dissolving.Measure the 1000ml buffer and be added in the phospholipid solution, stir under 50 ℃ of conditions and hatched 30 minutes, obtain the thick suspension of liposome.This thick suspension is added in the microjet instrument 5000psi circulation 5 times, 15000psi circulation 8 times.Behind the membrane filtration of 0.8um, 0.45um and 0.22um, promptly obtain docetaxel liposome respectively.
After measured, according to the liposome mean diameter 90nm that above-mentioned steps obtains, drug loading is 1.5mg/ml, and entrapment efficiency is 97.8%.Place after 90 days under the room temperature and detect particle diameter, drug loading, envelop rate and related substance, all do not have obviously change.
To redissolve after the above-mentioned liposome solutions lyophilizing, mean diameter is 95nm after testing, and drug loading is 1.5mg/ml, and entrapment efficiency is 97.0%, and t butanol content is less than 0.15%.Liposome after the lyophilizing is placed after 18 months and is detected, and main quality index does not all have obvious change.
Embodiment 4 cholesterol levels are to the influence of docetaxel liposome
Take by weighing 5 gram soybean phospholipids, 1.5 gram cholesterol and 0.2 gram docetaxel and be dissolved in the 10ml dehydrated alcohol, stirring and dissolving under 50 ℃ of conditions gets phospholipid solution.Compound concentration is the acetate buffer solution of 10mM, transfers pH to 4.5, adds trehalose 10 grams, fully stirring and dissolving.Measure the 100ml buffer and be added in the phospholipid solution, stir under 50 ℃ of conditions and hatched 60 minutes, obtain the thick suspension of liposome.This thick suspension is added in the microjet instrument 5000psi circulation 2 times, 10000psi circulation 5 times.Behind the membrane filtration of 0.8um, 0.45um and 0.22um, promptly obtain docetaxel liposome respectively.
After measured, according to the liposome mean diameter 90nm that above-mentioned steps obtains, drug loading is 2mg/ml, and entrapment efficiency is 96.8%.Place after 48 hours under 4 ℃ of conditions and detect, particle diameter 120nm, drug loading 1.5mg/ml, envelop rate 75%, and have precipitation to produce.
Embodiment 5 contains the preparation of the docetaxel liposome of Ovum Gallus domesticus Flavus lecithin
Take by weighing 50 gram Ovum Gallus domesticus Flavus lecithins, 1.5 gram cholesterol and 2 gram docetaxels and be dissolved in the 100ml dehydrated alcohol, stirring and dissolving under 50 ℃ of conditions gets phospholipid solution.Compound concentration is the acetate buffer solution of 10mM, transfers pH to 4.5, adds lactose 100 grams, mannitol 50 grams, fully stirring and dissolving.Measure the 1000ml buffer and be added in the phospholipid solution, stir under 50 ℃ of conditions and hatched 20 minutes, obtain the thick suspension of liposome.This thick suspension is added in the microjet instrument 5000psi circulation 2 times, 10000psi circulation 10 times.Behind the membrane filtration of 0.8um, 0.45um and 0.22um, promptly obtain docetaxel liposome respectively.
After measured, according to the liposome mean diameter 100nm that above-mentioned steps obtains, drug loading is 2mg/ml, and entrapment efficiency is 98.8%.Place after 30 days under 4 ℃ of conditions and detect particle diameter, drug loading, envelop rate and related substance, all do not have obviously change.
The preparation of embodiment 6 extensive common docetaxel liposomes
Take by weighing soybean lecithin 6kg, cholesterol 0.1kg and be dissolved in the 10L dehydrated alcohol, stirring and dissolving under 55 ℃ of conditions obtains phospholipid solution.Taking by weighing docetaxel 0.15kg is dissolved in the phospholipid solution.Compound concentration is the phosphate buffer of 10mM, transfers pH to 6.0, adds mannitol 4kg, maltose 8kg, and fully stirring and dissolving is removed the insoluble granule material through the 0.45um membrane filtration.Measure the 100L buffer and be added in the phospholipid solution, in retort, stir under 50 ℃ of conditions and hatched 40 minutes, obtain the thick suspension of liposome.This thick suspension is added in the microjet instrument 5000psi circulation 2 times, 10000psi circulation 5 times.Behind the filter element filtering of 0.8um, 0.45um and 0.22um, promptly obtain docetaxel liposome respectively.
Above-mentioned liposome carries out packing, lyophilizing under aseptic condition.
After measured, according to the liposome mean diameter 80nm that above-mentioned steps obtains, drug loading is 1.5mg/ml, and entrapment efficiency is 99.1%.Place after 90 days under the room temperature and detect particle diameter, drug loading, envelop rate and related substance, all do not have obviously change.
To redissolve after the above-mentioned liposome solutions lyophilizing, mean diameter is 95nm after testing, and drug loading is 1.5mg/ml, and entrapment efficiency is 99.2%, and ethanol content is less than 0.1%.Liposome after the lyophilizing is placed after 24 months and is detected, and main quality index does not all have obvious change.
The lyophilizing of embodiment 7 docetaxel liposomes
The liposome solutions of embodiment 1 preparation is carried out aseptic subpackaged, every bottled amount is 10ml, highly about 15mm.Add two fork plugs after the packing, place freeze dryer to carry out lyophilizing.Carry out lyophilizing according to following freeze-drying curve: sample temperature is brought down below-45 ℃ with the speed of 0.1 degrees celsius/minute, keeps after 4-6 hour and finishes pre-freeze; Freeze dryer flaggy temperature is set is-40 ℃, open vacuum pump, begin sublimation process when vacuum is reduced to 20Pa, this process was kept about 60 hours.Begin slow intensification afterwards, heat up, reach 25 degrees centigrade until temperature with 2 degrees centigrade/hour temperature.25 ℃ keep 12 hours after, finish lyophilizing and also charge into the nitrogen outlet.
After testing, be lower than 1.5% by the freeze dried liposome water content of above-mentioned technology, outward appearance is good, and the main quality index of liposome does not have obvious change before and after the lyophilizing.
Embodiment 8 film dispersion methods prepare docetaxel liposome
Take by weighing 6 gram natural phosphatidyl cholines, 0.15 gram cholesterol and 0.2 gram docetaxel and be dissolved in the 30ml dehydrated alcohol, stirring and dissolving under 50 ℃ of conditions gets phospholipid solution.Phospholipid solution is transferred in the rotary evaporation bottle revolves steaming, wave to remove most ethanol and make phospholipid be film like and be dispersed on the rotary evaporation bottle wall, place the vacuum drying oven dried overnight to remove residual ethanol in the rotary evaporation bottle.Compound concentration is the phosphate buffer of 10mM, transfers pH to 4.5, adds lactose 10 grams, mannitol 5 grams, fully stirring and dissolving.Measure the 100ml buffer and be added in the rotary evaporation bottle, stir under 50 ℃ of conditions and hatched 40 minutes, obtain the thick suspension of liposome.This thick suspension is added in the microjet instrument 5000psi circulation 2 times, 10000psi circulation 10 times.Behind the membrane filtration of 0.8um, 0.45um and 0.22um, promptly obtain docetaxel liposome respectively.
After measured, according to the liposome mean diameter 120nm that above-mentioned steps obtains, drug loading is 2mg/ml, and entrapment efficiency is 97.6%.Place after 30 days under 4 ℃ of conditions and detect particle diameter, drug loading, envelop rate and related substance, all do not have obviously change.
Embodiment 9 alcohol injections prepare docetaxel liposome
Take by weighing 50 gram natural phosphatidyl cholines, 1.5 gram cholesterol and 2 gram docetaxels and be dissolved in the 100ml dehydrated alcohol, stirring and dissolving under 50 ℃ of conditions gets phospholipid solution.Compound concentration is the acetate buffer solution of 10mM, transfers pH to 4.5, adds lactose 100 grams, mannitol 50 grams, fully stirring and dissolving.Measure the 1000ml buffer in beaker and place 50 ℃ of water-baths, simultaneously high-speed stirred.Be added in the syringe phospholipid solution and the slow buffer that injects, constantly stir, until whole phospholipid solutions are injected buffer, continuation was stirred 30 minutes under 50 ℃ of conditions, obtained the thick suspension of liposome.This thick suspension is added in the microjet instrument 5000psi circulation 2 times, 10000psi circulation 10 times.Behind the membrane filtration of 0.8um, 0.45um and 0.22um, promptly obtain docetaxel liposome respectively.
After measured, according to the liposome mean diameter 100nm that above-mentioned steps obtains, drug loading is 2mg/ml, and entrapment efficiency is 54.3%.Place under 4 ℃ of conditions and find have precipitation to produce after 7 days, envelop rate is reduced to about 10%.
Embodiment 10 organic solvent volumes are to the influence of docetaxel liposome
Respectively take by weighing 6 gram natural phosphatidyl cholines, 0.15 gram cholesterol and 0.2 gram docetaxel and be dissolved in respectively in (a) 6ml, (b) 10ml, (c) 15ml, (d) 20ml dehydrated alcohol, stirring and dissolving under 50 ℃ of conditions gets phospholipid solution.Compound concentration is the phosphate buffer of 10mM, transfers pH to 4.5, adds lactose 10 grams, mannitol 5 grams, fully stirring and dissolving.Measure respectively the 100ml buffer be added to above-mentioned (a) (b) (c) stir under 50 ℃ of conditions and hatched 40 minutes (d) in four groups of phospholipid solutions, obtain the thick suspension of liposome.Each is organized thick suspension be added to respectively in the microjet instrument, 5000psi circulation 2 times, 10000psi circulation 10 times.Behind the membrane filtration of 0.8um, 0.45um and 0.22um, promptly obtain docetaxel liposome respectively successively.
After measured, the result shows that the liposome of (a) group prepared and the main quality index zero difference of the liposome entrapment efficiency of (b) organizing prepared all reach more than 95%, particle diameter all about 90nm, place after 30 days and detect by room temperature, the main equal no change of quality index.Entrapment efficiency of liposome according to (c) group prepared is 85%, and particle diameter is 96nm, and the room temperature placement small amount of precipitate occurs after 2 days and envelop rate reduces to 50%.Entrapment efficiency of liposome according to (d) group prepared is 60%, particle diameter 92nm, and room temperature is placed and is occurred precipitation after 24 hours, and envelop rate reduces to 10%.
The preparation of the docetaxel liposome of embodiment 11 different medicine fat ratios
Medicine fat according to 1: 10,1: 20,1: 30,1: 50,1: 100 ratio prepares docetaxel liposome than (weight ratio) according to the preparation technology that embodiment 2 provides respectively.
Described medicine fat ratio is meant: the weight ratio of docetaxel medicine and phospholipid.
Each liposome to preparation detects.The result shows that when medicine fat ratio was 1: 10, entrapment efficiency was 75%, and the liposome room temperature is placed and occurred precipitation after 24 hours.When medicine fat ratio was 1: 100, entrapment efficiency was 99.2%, and drug loading is 0.6mg/ml, and room temperature is placed 30 days no changes, but outward appearance is relatively poor after the lyophilizing.The liposome quality of preparation is better when medicine fat ratio is respectively 1: 20,1: 30,1: 50.
The toxicity assessment of embodiment 12 docetaxel liposomes
Terminological interpretation:
The KM mice: i.e. kunming mice, the outbreeding group of mean people Mus of the amount of being to use maximum is widely used in the research in fields such as pharmacology, toxicology, and the production and the calibrating of medicine, biological product.But market is bought.
Iv: intravenous administration.
Q4d * 3: be administered once administration 3 times in per four days.
The docetaxel liposome of docetaxel liposome group selection embodiment 1,2,3,6 preparations carries out the toxicity assessment experiment, and docetaxel injection group selection Qilu Pharmaceutical Co., Ltd. produces the docetaxel injection of selling.
Select 50 of healthy KM mices, male and female half and half, age in Mus 6-8 in age week, body weight 18～22g.Mice is divided into 5 groups at random, and 10 every group, the initial body weight of KM mice is no more than or is lower than 20% of average weight, between group average weight difference less than 1g, every group of male and female half and half.
By the administration of tail intravenous injection, q4d * 3, administration volume 0.2ml/10g.After the administration at once to the administration 7～14d observe the reaction of animals situation, record body weight, poisoning symptom, outward appearance sign and death condition.Dosage regimen and the weight of animals situation of change are as shown in table 1.
Table 1 dosage regimen and the weight of animals situation of change table
The result shows, all there is dose-effect relationship preferably Docetaxel for Injection liposome mice state, body weight, mortality rate aspect after the administration, and the same dose liposome reduces than docetaxel injection toxicity.
During single dose 15mg/kg, liposome treated animal survival quantity all is higher than the docetaxel injection group.During single dose 33mg/kg, liposome group survival 5-6 only illustrates that this dosage is near median lethal dose(LD 50); And the docetaxel injection group is survived 2, illustrates that this dosage is higher than median lethal dose(LD 50).This experiment shows that the same dose docetaxel liposome is littler than the toxicity of docetaxel injection.
From the weight of animals situation of change, when the accumulation dosage is 45mg/kg, the docetaxel liposome group is all bigger always than docetaxel injection group body weight, and the average weight of the average weight of liposome treated animal and injection treated animal has significant difference during off-test; When accumulated dose is 99mg/kg, docetaxel liposome and docetaxel injection group average weight basically identical during off-test, but because injection group surviving animals negligible amounts, so not statistically significant.
In addition, the symptom of having loose bowels in various degree appears in the low dose group of docetaxel injection group animal when testing the 7th day, and the low dose group of docetaxel liposome group above-mentioned symptom also do not occur until off-test.The animal symptom of having loose bowels all appears in docetaxel liposome group and injection group during high dose, but the time of liposome group outlet symptom will be later than the injection group.
Present embodiment proves absolutely and adopts the docetaxel liposome preparation of the present invention's preparation can reduce the toxicity that common docetaxel injection brings.
Claims (8)
1. docetaxel liposome sterile freeze-drying preparation is characterized in that containing in per 100 gram docetaxel liposome sterile freeze-drying preparations following component:
This docetaxel liposome sterile freeze-drying preparation makes as follows:
The natural phosphatidyl choline, mPEG2000-DSPE and the cholesterol that take by weighing above-mentioned amount are dissolved in an amount of organic solvent, and fully the dissolving back adds docetaxel, adds buffer, hatches under suitable temperature and heat-retaining condition 10-60 minute; Again the mean diameter of liposome is reduced to 50-200nm; Through filtering with microporous membrane degerming postlyophilization promptly.
2. docetaxel liposome sterile freeze-drying preparation as claimed in claim 1 is characterized in that preparation method comprises the steps:
(1) DSPE (PEG-DSPE) and the cholesterol that takes by weighing natural phosphatidyl choline, PEGization is dissolved in the organic solvent, and stirring and dissolving gets phospholipid solution under 40-60 ℃ of condition;
(2) take by weighing docetaxel and adding in the above-mentioned phospholipid solution;
(3) adopt the water for injection preparation to contain the buffer salt solution of freeze drying protectant, regulate between the pH to 4.0-7.0; And it is added in the phospholipid solution that above-mentioned steps (2) makes, keep 40-60 ℃ of temperature, stir or ultrasound condition under hatched 10-60 minute suspension;
(4) suspension that step (3) is obtained is added in the microjet instrument, and adjusting pressure is 2000-6000psi, and after circulation 2-10 time, adjusting pressure is 7000-20000psi, circulation 2-10 time; Obtain liposome solutions;
(5) liposome solutions that step (4) is obtained filters, gets docetaxel liposome step by step through 0.8um, 0.45um, 0.22um filter membrane successively; Packing under aseptic condition, lyophilization promptly get the docetaxel liposome sterile freeze-drying preparation.
3. docetaxel liposome sterile freeze-drying preparation as claimed in claim 1 or 2 is characterized in that described natural phospholipid is natural soybean lecithin or Ovum Gallus domesticus Flavus lecithin.
4. docetaxel liposome sterile freeze-drying preparation as claimed in claim 3 is characterized in that in described soybean lecithin or Ovum Gallus domesticus Flavus lecithin the content of phosphatidylcholine is greater than 70wt%, and the content of hemolytic phospholipid is less than 3wt%.
5. docetaxel liposome sterile freeze-drying preparation as claimed in claim 1 or 2 is characterized in that described freeze drying protectant is one or more the compositions in maltose, mannitol, galactose, lactose, glucose, sucrose, the trehalose.
6. docetaxel liposome sterile freeze-drying preparation as claimed in claim 1 or 2 is characterized in that described buffer salt is selected from phosphate, acetate or glycinate.
7. docetaxel liposome sterile freeze-drying preparation as claimed in claim 1 or 2, it is characterized in that organic solvent used in the preparation process is selected from one or both the compositions in ethanol, the tert-butyl alcohol, the consumption of organic solvent be no more than final preparation the liquid preparation volume 20%, be preferably and be no more than 10%.
8. docetaxel liposome sterile freeze-drying preparation as claimed in claim 1 or 2, it is characterized in that the docetaxel liposome sterile freeze-drying preparation adds normal saline, glucose injection or water for injection and redissolves the back mean diameter between 50-200nm, entrapment efficiency is greater than 95%, and pH is between 4.0-7.0.
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Cited By (4)
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| CN103768018A (en) * | 2012-10-17 | 2014-05-07 | 南京绿叶思科药业有限公司 | Cabazitaxel liposome injection and preparation method thereof |
| CN104208030A (en) * | 2014-08-01 | 2014-12-17 | 中国人民解放军军事医学科学院放射与辐射医学研究所 | Albumin-combined taxol long-circulation nano-particle freeze-dried preparation |
| CN105853403A (en) * | 2016-05-09 | 2016-08-17 | 上海天氏利医药科技有限公司 | Paclitaxel palmitate liposome and preparation method thereof |
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| CN101015547A (en) * | 2006-05-15 | 2007-08-15 | 沈阳药科大学 | Docetaxel liposome formulation and preparation method thereof |
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| CN101317816B (en) * | 2007-06-08 | 2011-03-30 | 中国科学院上海药物研究所 | Docetaxel long-circulation formulation |
| CN101244039B (en) * | 2008-03-20 | 2012-09-12 | 江苏先声药物研究有限公司 | Novel method for preparing indissoluble medicaments liposome preparations |
| CN101584663B (en) * | 2008-05-22 | 2013-02-20 | 广州瑞济生物技术有限公司 | Novel delivery system of Duoxitasai lipidosome for injection and preparation method thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN103768018A (en) * | 2012-10-17 | 2014-05-07 | 南京绿叶思科药业有限公司 | Cabazitaxel liposome injection and preparation method thereof |
| CN104208030A (en) * | 2014-08-01 | 2014-12-17 | 中国人民解放军军事医学科学院放射与辐射医学研究所 | Albumin-combined taxol long-circulation nano-particle freeze-dried preparation |
| CN105853403A (en) * | 2016-05-09 | 2016-08-17 | 上海天氏利医药科技有限公司 | Paclitaxel palmitate liposome and preparation method thereof |
| CN105853403B (en) * | 2016-05-09 | 2019-03-29 | 上海天氏利医药科技有限公司 | A kind of paclitaxel palmitate liposome and preparation method thereof |
| CN111760022A (en) * | 2020-08-07 | 2020-10-13 | 山东泰邦生物制品有限公司 | Human immunoglobulin liposome pharmaceutical composition and preparation method thereof |
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