WO2019218857A1 - 一种多西他赛棕榈酸酯脂质体及其制备方法 - Google Patents

一种多西他赛棕榈酸酯脂质体及其制备方法 Download PDF

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WO2019218857A1
WO2019218857A1 PCT/CN2019/084543 CN2019084543W WO2019218857A1 WO 2019218857 A1 WO2019218857 A1 WO 2019218857A1 CN 2019084543 W CN2019084543 W CN 2019084543W WO 2019218857 A1 WO2019218857 A1 WO 2019218857A1
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liposome
docetaxel
palmitate
docetaxel palmitate
injection
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PCT/CN2019/084543
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English (en)
French (fr)
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陈建明
高保安
许幼发
武鑫
史亚敏
严朗
傅志勤
郦小平
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上海维洱生物医药科技有限公司
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Priority to US17/056,139 priority Critical patent/US20210212947A1/en
Publication of WO2019218857A1 publication Critical patent/WO2019218857A1/zh

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/127—Liposomes
    • A61K9/1271—Non-conventional liposomes, e.g. PEGylated liposomes, liposomes coated with polymers
    • A61K9/1273—Polymersomes; Liposomes with polymerisable or polymerised bilayer-forming substances
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/127—Liposomes
    • A61K9/1271—Non-conventional liposomes, e.g. PEGylated liposomes, liposomes coated with polymers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/337—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having four-membered rings, e.g. taxol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/08—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing oxygen, e.g. ethers, acetals, ketones, quinones, aldehydes, peroxides
    • A61K47/12—Carboxylic acids; Salts or anhydrides thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/16—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing nitrogen, e.g. nitro-, nitroso-, azo-compounds, nitriles, cyanates
    • A61K47/18—Amines; Amides; Ureas; Quaternary ammonium compounds; Amino acids; Oligopeptides having up to five amino acids
    • A61K47/183—Amino acids, e.g. glycine, EDTA or aspartame
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/24—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing atoms other than carbon, hydrogen, oxygen, halogen, nitrogen or sulfur, e.g. cyclomethicone or phospholipids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/26—Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/28—Steroids, e.g. cholesterol, bile acids or glycyrrhetinic acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/127—Liposomes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/127—Liposomes
    • A61K9/1277—Processes for preparing; Proliposomes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/19—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles lyophilised, i.e. freeze-dried, solutions or dispersions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • the invention relates to the technical field of medicine, in particular to a docetaxel palmitate liposome and a preparation method thereof.
  • Docetaxel also known as docetaxel, is a taxane antitumor drug modified with 10-deacetylbaccatin III as the core skeleton. Its chemical structure is shown in Figure 1. The chemical name is: [2aR-(2a ⁇ , 4 ⁇ , 4a ⁇ , 6 ⁇ , 9 ⁇ , (aR*, ⁇ S*), 11a, 12a, 12a ⁇ , 12b ⁇ )]- ⁇ -[[1,1 dimethyl Oxy)carbonyl]amino]- ⁇ -hydroxyphenylpropionic acid [12b-acetoxy-12-benzoyloxy-2a,3,4,4a,5,6,9,10,11,12,12a,12b - dodecahydro-4,6,11-trihydroxy-4a,8,13,13-tetramethyl-5-oxo-7,11-methylene-1H-cyclopentapenta[3,4 Benzo[1,2-b]oxabutan-9-yl]ester, molecular formula C 43 H 53 NO 14
  • the anti-tumor activity of docetaxel is 1.3-12 times that of paclitaxel.
  • the curative effect is exact.
  • the FDA has approved its use for the treatment of cancers such as breast cancer, ovarian cancer, non-small cell lung cancer and pancreatic cancer. It is the most valuable discovery to date.
  • One of the anticancer drugs One of the anticancer drugs.
  • the docetaxel preparation for clinical use is its injection, docetaxel injection, which is the only clinical preparation of docetaxel.
  • the injection consists of docetaxel solution of docetaxel and 13% ethanol solution. When used, 13% ethanol solution is injected into the Tween 80 solution of docetaxel and shaken. Dilute in % glucose solution or normal saline before intravenous drip. It can be seen that the clinical application process of the injection is cumbersome, inconvenient to use, and easy to secondary pollution.
  • the effect of docetaxel injection is remarkable, the adverse reactions are particularly prominent, and its bone marrow suppression toxicity is the primary adverse reaction, which is clearly described in the specification.
  • Tumor stromal disrupting agent enhances the anti-ceremon efficacy of docetaxel loaded PEGylated liposomes in lung cancer [J] .Nanomedicine, 2016, 11 (11): 1377-1392.); Chinese patent (patent number: CN101584663A) provides a novel injection of docetaxel liposome and its preparation method, prepared by emulsion volatilization method Docetaxel liposome, the preparation process is complex and uncontrollable, and the formulation contains sodium cholesteryl sulfate, sodium dodecyl benzene sulfonate, etc., solubilizing Has a strong hemolytic property (Cui Fu. Pharmacy [M].
  • Chinese patent discloses a docetaxel freeze-dried liposome and The preparation method is to increase the water solubility of docetaxel, increase the encapsulation efficiency, and improve the stability of the liposome by adding a cyclodextrin having a hemolytic property, even if the drug loading of the liposome is only 0.5. Mg/mL, can not meet the requirements of clinical drugs;
  • Chinese patent (patent number: CN102379849A) provides a pH-sensitive docetaxel liposome and its preparation method, the drug loading is still low, and the liposome The path is too large.
  • docetaxel liposomes are poorly implementable, mainly because docetaxel is poorly soluble and does not match the compatibility of the lipid material.
  • the plastid has a series of problems such as low drug loading, low encapsulation efficiency and poor stability.
  • docetaxel liposomes are poorly prepared, so it is especially important to develop a docetaxel liposome that is truly efficient, low-toxic, stable in quality, and simple in preparation. This will be docetaxel. It lays a solid foundation for research and application in the field of anti-tumor.
  • the present invention provides a docetaxel palmitate liposome.
  • docetaxel palmitate liposome In order to solve the problem of poor fat solubility and poor drug-forming properties of docetaxel, we intend to improve the fat solubility of docetaxel through structural modification, that is, the esterification reaction of docetaxel with palmitic acid to obtain docetaxel.
  • the fat-soluble prodrug of the race namely docetaxel palmitate.
  • Tests have shown that the preparation of docetaxel palmitate by a specific prescription process is excellent, and the drug loading is as high as 10 mg/ml (see Example 10).
  • the anti-tumor effect in mice is better than that in the market.
  • the docetaxel injection see Example 19
  • the present invention transforms the docetaxel structure into docetaxel palmitate, which is one of the key technical features of the present invention that has received significant results.
  • docetaxel palmitate liposome can be fully inspired by the above patents to achieve the intended purpose, and in fact, as mentioned, The anti-tumor effect is indeed superior to the commercially available docetaxel injection (see Example 19).
  • the chelating agent was introduced into the prescription, the in vivo action time of the drug was prolonged (see Example 20), and the anti-tumor effect was also better; in addition, after the addition of the chelating agent, the liposome-related Good results have also been obtained in terms of quality indicators, such as narrower particle size distribution, smoother sterilization filtration, and industrial batch production, which greatly improves the applicability of the present invention (see Example 21). Therefore, the inclusion of a chelating agent in a docetaxel palmitate liposome provided by the present invention is the core technical feature of the present invention.
  • a metal atom or ion acts with a ligand containing two or more than two coordinating atoms to form a chelate compound having a cyclic structure, and such a ligand substance capable of forming a chelate compound is called a chelating agent.
  • chelating agents are widely used, but are basically added to improve the chemical stability of the active ingredient, especially as an antioxidant, but can be used in the efficacy of the preparation and prolonging the action time in the body, etc. It is rare to show better results. Through the literature search at home and abroad, there is no relevant report.
  • a first object of the present invention is to provide a docetaxel palmitate liposome.
  • the present invention provides a docetaxel palmitate liposome comprising docetaxel palmitate as a main drug.
  • Docetaxel palmitate is used in an amount of 0.1 to 2% by weight.
  • the present invention provides a docetaxel palmitate liposome, mainly docetaxel palmitate, and a chelating agent.
  • Docetaxel palmitate is used in an amount of from 0.1 to 2%; and the chelating agent is used in an amount of from 0.001 to 1% by weight.
  • the present invention provides a docetaxel palmitate liposome, mainly docetaxel palmitate, and further comprises a chelating agent, lecithin, and DSPE-PEG2000.
  • the dosage of docetaxel palmitate is 0.1-2%; the amount of chelating agent is 0.001-1%; the amount of lecithin is 1-10%, and the amount of DSPE-PEG2000 is 0.05-1% (weight by volume).
  • a second object of the present invention is to provide a docetaxel palmitate liposome which is a lyophilized powder injection or a liposome solution for injection.
  • a third object of the present invention is to provide a docetaxel palmitate prodrug, wherein the docetaxel palmitate prodrug is docetaxel as a parent drug, and an ester bond is linked to a molecule of palm. It forms a prodrug with acid and is a fat-soluble prodrug with good stability and functionality.
  • the structure of the docetaxel palmitate prodrug is as follows:
  • the docetaxel palmitate prodrug characterized in that palmitic acid is linked to the 2' position of the docetaxel side chain, and the preparation process is as follows: docetaxel 10.00 g, palmitic acid 3.81 g, 1- (3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC) 2.43 g and 4-dimethylaminopyridine (DMAP) 1.82 g, placed in a reaction vessel, made with 50 mL of anhydrous dichloromethane Dissolve, under nitrogen protection conditions, stir the reaction at room temperature for 4-24h to obtain the reaction solution; wash the reaction solution twice with 5% aqueous citric acid solution, then wash it once with saturated sodium chloride solution, remove the anhydrous water by rotary evaporation Methyl chloride; isolated and purified to give docetaxel palmitate.
  • the reaction synthesis route map is as follows:
  • the present invention provides a docetaxel palmitate liposome, the liposome being an injection containing a chelating agent; the injection containing the chelating agent may also be an injection solution, or a kind Lyophilized powder needle; containing a chelating agent is a core technical feature of the present invention.
  • the docetaxel palmitate liposome is prepared by the following formula:
  • the lyoprotectant is 0;
  • the lyoprotectant is 0.1-40% g/ml.
  • the docetaxel palmitate liposome is prepared by the following formula:
  • the docetaxel palmitate liposome is prepared by the following formula:
  • the lecithin described in the above formula is selected from the group consisting of high purity egg yolk lecithin (EPCS), hydrogenated soybean lecithin (HSPC), dipalmitoylphosphatidylcholine (DPPC), phosphatidylcholine, egg yolk lecithin, One or a combination of two or more of soybean lecithin, phosphatidylserine, dimyristoyl phosphatidylcholine, distearoylphosphatidylcholine, phosphatidylethanolamine, and sphingomyelin; preferably high-purity egg yolk lecithin (EPCS) And one or a combination of two or more of hydrogenated soybean lecithin (HSPC).
  • EPCS high purity egg yolk lecithin
  • HSPC hydrogenated soybean lecithin
  • DPPC dipalmitoylphosphatidylcholine
  • phosphatidylcholine egg yolk lecithin
  • the chelating agent described in the appeal formulation is selected from the group consisting of citric acid, disodium citrate, trisodium citrate, lactic acid, sodium lactate, malic acid, sodium malate, ethylenediaminetetraacetic acid, disodium edetate, and ethylene.
  • One or a mixture of two or more kinds of trisodium aminotetraacetate is preferably used in combination of one or more of citric acid, disodium citrate, trisodium citrate, lactic acid, and sodium lactate.
  • the lyoprotectant described in the above formula is one or more selected from the group consisting of trehalose, sucrose, maltose, lactose, mannitol, glucose, sorbitol, xylitol, erythritol, and threonine; One or a combination of mannitol, trehalose, and sucrose is used in combination.
  • the pH adjuster described in the above formula is one or more selected from the group consisting of sodium hydroxide and hydrochloric acid.
  • a fourth object of the present invention is to provide a process for the preparation of said docetaxel palmitate liposome.
  • the preparation method of the docetaxel palmitate liposome is an injection method.
  • the docetaxel palmitate liposome is prepared by the following steps:
  • a liposome solution Homogeneous emulsification in a homogenizer, or placing it in an extruder through successive extrusion membranes of different pore diameters, or high-pressure homogenization and then extruding to obtain a liposome solution;
  • the dry protective agent is placed in the above liposome solution, stirred to dissolve, and made up to the full amount with water for injection; pH is adjusted with a pH adjuster; the filter is sterilized, dispensed, and sealed by a 0.22 ⁇ m filter, ie, He is a palmitate liposome; it can also be lyophilized to prepare a docetaxel palmitate liposome lyophilized powder.
  • the organic solvent for injection is selected from one or more selected from the group consisting of propylene glycol, absolute ethanol and t-butanol, and the amount is 1-8% g/ml; preferably anhydrous ethanol, preferably used in an amount of 2 - 5% g/ml.
  • the organic solvent for injection may be retained in the liposome, or may be removed by ultrafiltration after emulsification of the crude liposome, or may be removed by freeze drying.
  • the liposome crude product is emulsified, preferably by extrusion emulsification, and the obtained liposome particle size distribution will be more uniform; the extruded membrane pore diameter is selected from 0.8 ⁇ m, 0.6 ⁇ m, 0.4 ⁇ m, 0.2 ⁇ m, 0.1 ⁇ m, 0.05 ⁇ m, one or two or more extrusions of large pore size to small pore diameter are used, preferably 0.4 ⁇ m, 0.2 ⁇ m, 0.1 ⁇ m, 0.05 ⁇ m.
  • the chelating agent may be dissolved in the oil phase, or may be dissolved in the aqueous phase or dissolved in the liposome solution.
  • the lyoprotectant is dissolved in the liposome solution and may also be dissolved in the aqueous phase.
  • a docetaxel palmitate liposome of the invention having a particle size of 50-150 nm.
  • the docetaxel palmitate liposome of the invention containing a chelating agent is a core technical feature of the invention, and the addition of the chelating agent makes the docetaxel act longer in time, has better antitumor effect, and is related to the preparation. The characteristics are better, which is the fundamental manifestation of the substantial effects of the present invention.
  • the average particle diameter was determined to be 92.4 nm.
  • the volume was adjusted to 100 ml; the pH was adjusted to 5.5 with sodium hydroxide; the filter was sterilized by 0.22 ⁇ m filter, divided, lyophilized, and sealed to obtain paclitaxel palmitate liposome lyophilized powder.
  • the average particle diameter was determined to be 86.6 nm.
  • the average particle diameter was determined to be 120.7 nm.
  • the average particle diameter was determined to be 50.36 nm.
  • the high-pressure homogenizer is homogeneously emulsified to obtain a liposome solution; 10 g of sucrose and 5 g of mannitol are weighed, placed in the above liposome solution, stirred to dissolve, and made up to 100 ml with water for injection; pH is adjusted with sodium hydroxide. The value is 7.0; after 0.22 ⁇ m filter membrane sterilization, dispensing, lyophilization, and sealing, the paclitaxel palmitate liposome freeze-dried powder is obtained
  • the average particle diameter was determined to be 60.7 nm.
  • docetaxel palmitate 0.7g, high-purity egg yolk lecithin (EPCS) 6g, 0.7g DSPE-PEG2000, cholesterol 0.5g, citric acid 0.5g, placed in 6g absolute ethanol, in Heating at 55 ° C to dissolve, to obtain an organic phase; weigh 80g of water for injection, heated to 55 ° C to obtain an aqueous phase; the organic phase is injected into the aqueous phase under stirring, and mixed to obtain a crude liposome; The crude body is homogenized and emulsified by a high-pressure homogenizer to obtain a liposome solution; the volume is adjusted to 100 ml with water for injection; the pH value is adjusted to 4.8 with sodium hydroxide; the membrane is sterilized, dispensed and sealed by 0.22 ⁇ m. A paclitaxel palmitate liposome solution was obtained.
  • EPCS high-purity egg yolk lecithin
  • the average particle diameter was determined to be 80.4 nm.
  • the average particle diameter was determined to be 150.0 nm.
  • docetaxel palmitate 0.8g, dipalmitoylphosphatidylcholine (DPPC) 3g, phosphatidylcholine 3g, egg yolk lecithin 1g, 0.8g DSPE-PEG2000, placed in 8g propylene glycol
  • the organic phase is obtained by heating at 70 ° C to obtain an organic phase; 80 g of water for injection is weighed and heated to 55 ° C to obtain an aqueous phase; the organic phase is poured into the aqueous phase under stirring, and mixed to obtain a crude liposome;
  • the crude liposome was extruded through an extrusion membrane having a pore diameter of 0.8 ⁇ m, 0.4 ⁇ m, 0.2 ⁇ m, and 0.1 ⁇ m to obtain a liposome solution; ultra-filtration was used to remove propylene glycol by ultrafiltration; and trisodium citrate 0.8 g was weighed.
  • the average particle diameter was determined to be 145.2 nm.
  • docetaxel palmitate 0.1g, soy lecithin 2g, 0.1g DSPE-PEG2000, placed in 6g of absolute ethanol, stir at 25 ° C to dissolve, to obtain the organic phase; 0.5 g of disodium diamine tetraacetate and 0.5 g of disodium citrate, placed in 80 g of water for injection, and stirred at 25 ° C to dissolve to obtain an aqueous phase; the organic phase is poured into the aqueous phase under stirring, and mixed.
  • the crude liposome was obtained by extruding the crude liposome with an extrusion membrane having a pore size of 0.6 ⁇ m, 0.2 ⁇ m, 0.1 ⁇ m, and 0.05 ⁇ m to obtain a liposome solution; and diluting to 100 ml with water for injection; Hydrochloric acid adjusts the pH value to 3.5; after 0.22 ⁇ m filter membrane sterilization, dispensing, sealing, the paclitaxel palmitate liposome solution is obtained.
  • the average particle diameter was determined to be 118.8 nm.
  • the machine is homogenized and emulsified to obtain a liposome solution; ultra-filter is used to remove anhydrous ethanol by ultrafiltration; sodium lactate 0.5g and sodium malate 0.5g are weighed, placed in ultra-filtered liposome solution, stirred to dissolve The volume was adjusted to 100 ml with water for injection; the pH value was adjusted to 5.0 with hydrochloric acid; the solution was sterilized, dispensed, and sealed through a 0.22 ⁇ m filter to obtain a paclitaxel palmitate liposome solution.
  • the average particle diameter was determined to be 130.2 nm.
  • the average particle diameter was determined to be 90.7 nm.
  • the crude liposome was obtained; the crude liposome was extruded through an extrusion membrane having a pore diameter of 0.8 ⁇ m, 0.6 ⁇ m, 0.4 ⁇ m, and 0.1 ⁇ m to obtain a liposome solution; the volume was adjusted to 100 ml with water for injection; The pH value was adjusted to 7.5 with sodium hydroxide; the 0.22 ⁇ m filter was sterilized, dispensed, lyophilized, and sealed to obtain paclitaxel palmitate liposome lyophilized powder.
  • the average particle diameter was determined to be 104.3 nm.
  • the organic phase is obtained by heating at 70 ° C to obtain an organic phase; 50 g of water for injection is weighed and heated to 70 ° C to obtain an aqueous phase; the organic phase is poured into the aqueous phase under stirring, and mixed to obtain a crude liposome; The crude liposome was homogenized by high-pressure homogenizer to obtain a liposome solution; 15 g of sucrose, 15 g of mannitol, 5 g of erythritol and 5 g of threonine were weighed and placed in the above liposome solution, and stirred.
  • the average particle diameter was determined to be 78.4 nm.
  • the average particle diameter was determined to be 140.7 nm.
  • the average particle diameter was determined to be 105.3 nm.
  • the average particle diameter was determined to be 89.4 nm.
  • docetaxel palmitate 0.3g, high-purity egg yolk lecithin (EPCS) 3g, 0.7g DSPE-PEG2000, citric acid 0.1g, placed in 4g propylene glycol, heated at 70 ° C to dissolve
  • the organic phase is obtained; 10 g of sucrose and 5 g of trehalose are weighed, placed in 70 g of water for injection, and heated to 70 ° C to obtain an aqueous phase; the organic phase is poured into the aqueous phase under stirring, and mixed to obtain a liposome.
  • EPCS high-purity egg yolk lecithin
  • the crude liposome was extruded through an extrusion membrane having a pore size of 0.8 ⁇ m, 0.6 ⁇ m, 0.4 ⁇ m, and 0.1 ⁇ m to obtain a liposome solution; the volume was adjusted to 100 ml with water for injection; and the pH was adjusted with sodium hydroxide. It is 6.0; the 0.22 ⁇ m filter membrane is sterilized, divided, lyophilized and sealed to obtain paclitaxel palmitate liposome lyophilized powder.
  • the average particle diameter was determined to be 113.6 nm.
  • docetaxel palmitate 0.3g, high-purity egg yolk lecithin (EPCS) 3g, 0.5g DSPE-PEG2000, 0.01g citric acid, placed in 4g propylene glycol, heated at 60 ° C to dissolve
  • the organic phase is obtained; 17 g of sucrose and 5 g of mannitol are weighed, placed in 65 g of water for injection, and heated to 70 ° C to obtain an aqueous phase; the organic phase is poured into the aqueous phase under stirring, and mixed to obtain a liposome.
  • EPCS high-purity egg yolk lecithin
  • the crude liposome was extruded with an extrusion membrane having a pore size of 0.6 ⁇ m, 0.4 ⁇ m, 0.1 ⁇ m, and 0.05 ⁇ m to obtain a liposome solution; the volume was adjusted to 100 ml with water for injection; and the pH was adjusted with sodium hydroxide. It is 6.7; the 0.22 ⁇ m filter membrane is sterilized, divided, lyophilized and sealed to obtain paclitaxel palmitate liposome lyophilized powder.
  • the average particle diameter was determined to be 106.3 nm.
  • Example 19 Effect of chelating agents on antitumor effects of docetaxel palmitate liposomes
  • a chelating agent in the formulation is the key to the substantial effect of the docetaxel palmitate lipid of the present invention.
  • a chelating agent and a chelating agent-free liposome were separately prepared under the same prescription process by using multiple parallel comparison methods, and the mouse S180 sarcoma was used as a tumor model.
  • the anti-tumor effects of the chelating agent and the docetaxel palmitate liposome containing no chelating agent were compared. The experimental design and results are shown below.
  • the commercially available docetaxel injection was used as a positive control drug; the chelating agent docetaxel palmitate liposome prepared in Example 1 was used as a chelating agent-containing test preparation; the prescription according to Example 1 was completely obtained.
  • the chelating agent-free liposomes were prepared in parallel as a chelating agent-free control formulation.
  • Mouse ascites S180 cells were cultured in DMEM medium, routinely cultured at 37 ° C, 5% CO 2 , subcultured every two days, and adjusted to a cell concentration of 5 ⁇ 10 7 cells in the logarithmic growth phase. mL, under sterile conditions, was injected into the abdominal cavity of mice. After a week of obvious ascites, the ascites of tumor-bearing mice were taken under aseptic conditions, diluted with normal saline at a ratio of 1:5, and inoculated with 0.2 mL of mice. In the abdominal cavity.
  • the second-generation ascites of the tumor-bearing mice was taken under aseptic conditions, diluted with a 1:5 amount of normal saline, and S180 cell suspension was subcutaneously injected into the left axilla of the mice, each 0.2 mL.
  • mice after 24 hours of inoculation were weighed and randomly divided into 4 groups of 8 animals each.
  • a blank group a commercially available docetaxel injection group, a chelating agent-free control group, and a chelating agent-containing control group were separately set.
  • the mice were administered by tail vein injection at a dose of 10 mg/kg each (according to docetaxel), and the blank group was given 0.2 ml of physiological saline, once a day for a total of 4 doses.
  • the mice were sacrificed on the third day after withdrawal, the body weight of the mice was weighed, the tumors were weighed, and the tumor inhibition rate was calculated.
  • Tumor inhibition rate (normal weight of saline group - tumor weight of administration group) / tumor weight of physiological saline group ⁇ 100%
  • the prescription contains a chelating agent, and the anti-tumor effect of docetaxel palmitate liposomes will be better, which is a manifestation of the substantial effects of the present invention, so the inclusion of a chelating agent is a core technical feature of the present invention. .
  • Example 20 Effect of chelating agents on pharmacokinetics of docetaxel palmitate liposomes in vivo
  • Example 1 The commercially available docetaxel injection was used as a reference preparation; the docetaxel palmitate liposome prepared in Example 1 was used as a chelating agent docetaxel palmitate liposome sample; The formulation procedure of Example 1 produced a chelating agent-free docetaxel palmitate liposome as a chelating agent-free docetaxel palmitate liposome sample.
  • Rat tail vein injection was administered at a dose of 10 mg/kg (according to docetaxel) at 0.033 h, 0.083 h, 0.167 h, 0.25 h, 0.5 h, 0.75 h, 1 h after administration.
  • 0.5 mL of blood was taken from the orbital venous plexus, placed in a centrifuge tube containing heparin sodium and shaken, centrifuged at 4500 r/min for 10 min, and 150 ⁇ l of plasma was taken. Cryopreservation at -20 °C. Plasma samples were processed in a conventional manner and the concentration of docetaxel in plasma was determined by high performance liquid chromatography.
  • Model fitting was performed using DAS 2.0 software, and pharmacokinetic parameters were calculated.
  • the in vivo pharmacokinetic results of the commercially available docetaxel injection group, the chelating agent docetaxel palmitate liposome group, and the chelating agent docetaxel palmitate liposome are shown in Table 2. .
  • Docetaxel palmitate liposomes, AUC ⁇ and T 1/2 are significantly larger than commercially available docetaxel palmitate injections.
  • the experimental results show that after the preparation of docetaxel as a prodrug liposome, the metabolism of the drug in the body is obviously delayed, and the action time is longer.
  • a chelating agent-containing docetaxel palmitate liposome with an increase in AUC ⁇ and a prolonged T 1/2 compared to a docetaxel palmitate liposome without a chelating agent. From the perspective of drug efficacy, liposomes containing chelating agents have better anti-tumor effects, perhaps because they have a relatively long duration of action in vivo, so the anti-tumor effect is better.
  • a chelating agent to the formulation of docetaxel palmitate liposomes prolongs the in vivo action time of the drug and enhances the anti-tumor effect.
  • the inclusion of a chelating agent in the formulation is particularly important for the superiority of the docetaxel palmitate liposomes of the present invention and is a key technical feature.
  • Example 21 Effect of chelating agents on the properties of docetaxel palmitate liposomal formulations
  • the study is a sterile liposome, and the liposome cannot be autoclaved during the production process. Therefore, the 0.22 ⁇ m filter is usually used for sterilization and filtration to achieve a sterile effect.
  • the sterilization filtration is not smooth, which seriously affects the production efficiency.
  • docetaxel palmitate liposomes we paid particular attention to the smoothness of the sterilization filtration and found that the addition of a chelating agent can make the filtration more smooth.
  • the liposome added with a chelating agent has a slightly smaller particle size and a smaller particle size distribution PDI, so the filtration is smoother.
  • the experimental design and results are shown below.
  • Example 1 1000 ml of a docetaxel palmitate liquid liposome containing a chelating agent and a chelating agent was prepared in accordance with the formulation process described in Example 1, respectively, using a plate filter having a diameter of 11 mm, 0.22 ⁇ m.
  • the polyethersulfone filter was filtered to record the filtration volume; the particle size and particle size distribution PDI of the chelating agent and the chelating agent-free docetaxel palmitate liquid liposome were determined, and the test results are shown in Table 3. :
  • Chelating agent docetaxel palmitate liposomes Chelating agent docetaxel palmitate liposome The average particle size 102.1nm 92.4nm PDI 0.237 0.124
  • the docetaxel palmitate liposome of the invention has smaller particle size, narrower distribution and smoother sterilization filtration. It can be seen that after the addition of the chelating agent, the basic properties of the preparation are obviously improved, and the production implementation will be smoother. It further embodies the superiority of the chelating agent in the prescription.

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Abstract

一种多西他赛棕榈酸酯脂质体及其制备方法，以多西他赛棕榈酸酯为主药，还包括螯合剂、卵磷脂以及DSPE-PEG2000。所述脂质体延长了药物在体内的作用时间，提高了抗肿瘤效果，不含增溶剂，制备工艺简单。

Description

一种多西他赛棕榈酸酯脂质体及其制备方法 技术领域
本发明涉及医药技术领域，具体涉及一种多西他赛棕榈酸酯脂质体及其制备方法。
背景技术
多西他赛(Docetaxel，DTX)又名多西紫杉醇，是以10-脱乙酰基浆果赤霉素Ⅲ为母核骨架经修饰得到的紫杉烷类抗肿瘤药，其化学结构式如图1所示，化学名称为：[2aR-(2aα,4β,4aβ,6β,9α,(aR*,βS*),11a,12a,12aα,12bα)]-β-[[(1,1二甲基乙氧基)羰基]氨基]-α-羟基苯丙酸[12b-乙酰氧-12-苯甲酰氧-2a,3,4,4a,5,6,9,10,11,12,12a,12b-十二氢-4,6,11-三羟基-4a,8,13,13-四甲基-5-氧代-7,11-亚甲基-1H-环癸五烯并[3,4]苯并[1,2-b]氧杂丁环-9-基]酯，分子式C 43H 53NO 14，分子量807.88，不溶于水，易溶于乙醇、丙酮、乙醚以及苯等有机溶剂。多西他赛的抗肿瘤活性是紫杉醇的1.3-12倍，疗效确切，FDA已批准其用于乳腺癌、卵巢癌、非小细胞肺癌以及胰腺癌等癌症的治疗，是迄今发现最有价值的抗癌药物之一。
Figure PCTCN2019084543-appb-000001
多西他赛化学结构式
目前，应用于临床的多西他赛制剂为其注射液，即多西他赛注射液，系多西他赛的临床唯一制剂。该注射液是由多西他赛的吐温80溶液与13％的乙醇溶液两部组成，临用时将13％乙醇溶液注入在多西他赛的吐温80溶液中摇匀，然后再用5％葡萄糖溶液或生理盐水稀释，方可静脉滴注。由此可见，该注射液临床应用过程繁琐、使用不便、易二次污染。多西他赛注射液虽然疗效显著，但不良反应尤为突出，其骨髓抑制毒性是首要不良反应，在其说明书中已明确记载。多西他赛单独治疗时，骨髓抑制发生率高达76.4％，当与其他化疗药物联合使用时骨髓抑制毒性更加严重，发生率更高，严重影响化疗进程，削弱患者的治疗效果(多西他赛注射液产品说明书；朱琨,周灿,闫融,等.多西他赛常规剂量化疗后Ⅳ度骨髓抑制患者白细胞计数波动规律及治疗策略[J].现代肿瘤医学,2012,20(1):000159-161.；曹建伟,耿明飞,朱东山,等.多西他赛化疗所致食管癌患者骨髓抑制的特点及对策[J].中国民康医学,2016,28(11):10-11.；沙红玉,郑文文,郭晨煜,等.多西他赛所致不良反应报告分析[J].中国医院药学杂志,2015,35(6):536-539.)，再加上其处方中含具有溶血性与过敏性的吐温80，给临床用药带来了严重的安全隐患，严重限制了多西他赛抗肿瘤药效的发挥，故迫切需要开发一种更加高效的、低骨髓抑制毒性的、不含增溶剂的多西他赛注射液应用于临床，为多西他赛的深入研究与应用奠定基础。
鉴于多西他赛注射液存在的不足，其新制剂的研究报道屡见不鲜，尝试将多西他赛开 发成脂质体、胶束、纳米粒等纳米包裹制剂(程树仓,庞鑫,翟光喜.多烯紫杉醇纳米制剂的研究进展[J].药学研究,2013,32(1):45-48.)，遗憾的是迄今为止还没有一种新的多西他赛纳米制剂上市。从降低毒性提高疗效的角度来看，脂质体作为抗肿瘤药物的载体研究与应用较为广泛，特别是已应用于临床的阿霉素脂质体以及伊立替康脂质体，在降低毒性与提高疗效方面效果尤为突出，故广大医药工作者拟将多西他赛开发成脂质体，拟达到安全高效的目的。现有文献研究了一种多西他赛脂质体，其载药量仅为0.75mg/mL，且稳定性差，无法长期保存，无法应用于临床(Immordino M L,Brusa P,Arpicco S,et al.Preparation,characterization,cytotoxicity and pharmacokinetics of liposomes containing docetaxel[J].Journal of Controlled Release,2003,91(3):417.)；现有文献研究的多西他赛脂质体，载药量最多不超过1mg/mL，且稳定性差，制备工艺复杂，不利于产业化(Patel K,Doddapaneni R,Chowdhury N,et al.Tumor stromal disrupting agent enhances the anticancer efficacy of docetaxel loaded PEGylated liposomes in lung cancer[J].Nanomedicine,2016,11(11):1377-1392.)；中国专利(专利号：CN101584663A)提供了一种新型的注射用多西他赛脂质体及其制备方法，利用乳化挥散法制备多西他赛脂质体，制备工艺复杂不可控，且处方中含有胆固醇硫酸酯钠、十二烷基苯磺酸钠等离子型的增溶剂，该类增溶剂具有较强的溶血性(崔福德.药剂学[M].北京:人民卫生出版社,2011:42)；中国专利(专利号：CN103830181A)公开了一种多西他赛冻干脂质体及其制备方法，是通过添加具有溶血性的环糊精来达到提高多西他赛水溶性、提高包封率、提高脂质体稳定性的目的，即便如此脂质体的载药量也仅为0.5mg/mL，无法满足临床用药的要求；中国专利(专利号：CN102379849A)提供一种具有pH敏感的多西他赛脂质体及其制备方法，载药量依然较低，且脂质体粒径偏大。由此可见，直接将多西他赛制备成脂质体的可实施性较差，主要原因是多西他赛脂溶性很差，与脂质材料的相容性不匹配所致，制备的脂质体存在载药量低、包封率低、稳定性差等一系列问题。总而言之，多西他赛脂质体的成药性很差，故研发一种真正意义上高效低毒、质量稳定、制备工艺简单的多西他赛脂质体尤为重要，这将为多西他赛在抗肿瘤领域的研究与应用奠定坚实的基础。
发明内容
本发明提供一种多西他赛棕榈酸酯脂质体。为了解决多西他赛脂溶性差以及脂质体成药性差的问题，我们拟通过结构改造来提高多西他赛的脂溶性，即将多西他赛与棕榈酸进行成酯反应，得到多西他赛的脂溶性前药，即多西他赛棕榈酸酯。试验证明，将多西他赛棕榈酸酯通过特定的处方工艺制备的脂质体成药极佳，载药量居然高达10mg/ml(见实施例10)，小鼠体内抗肿瘤效果好于市售的多西他赛注射液(见实施例19)，故本发明将多西他赛结构改造成多西他赛棕榈酸酯，是本发明收到显著效果的关键技术特征之一。
在中国专利CN201610301096.4(公开号CN105853403A)中明确记载了一种紫杉醇的脂溶性前药紫杉醇棕榈酸酯，且成功开发成了脂质体，体内抗肿瘤效果以及安全性亦显著提高，故通过合成脂溶性前药来解决原药脂质体成药性差的方案，以及收到高效低毒的动物体内效果已不足为奇。另外，多西他赛与紫杉醇同属紫杉烷类化合物，理论上讲多西他 赛棕榈酸酯脂质体可完全受上述专利的启发即可达到预期的目的，而事实上也正如所述，抗肿瘤效果的确优于市售的多西他赛注射液(见实施例19)。但在研究过程中，意外发现当处方中引入螯合剂后，会延长药物的体内作用时间(见实施例20)，同时抗肿瘤效果也更好；另外，添加螯合剂后，在脂质体相关质量指标等方面也收到了更好的效果，如粒径分布将更窄、除菌过滤更加顺畅，有利于产业化批量生产，大大提高了本发明的适用性(见实施例21)。故本发明提供的一种多西他赛棕榈酸酯脂质体中含有螯合剂是本发明最为核心的技术特征。
金属原子或离子与含有两个或连两个以上配位原子的配位体作用，生成具有环状结构的螯合物，能生成螯合物的这种配体物质叫螯合剂。在药物制剂中，螯合剂广泛使用，但基本上均是为了提高活性成分的化学稳定性而添加的，特别是作为抗氧化剂使用效果较好，但能够在制剂的药效以及延长体内作用时间等方面表现出更好的效果则少见。通过对国内外文献检索，亦未见有相关报道，本脂质体中的螯合剂到底是对药物产生作用还是对脂质体微粒本身有某种结合，机理暂时不明，后续我们将进一步深入予以探讨研究。出于好奇，我们开展了系列对比试验，发现在多西他赛棕榈酸酯脂质体中添加螯合剂后，药物在体内的循环明显延长，曲线下面积AUC∞也大于不加螯合剂的脂质体(见实施例20)。我们知道，药物在体内缓慢代谢，不易失活，故药效自然提高。故本发明要想达到更好的实质效果，仅凭中国专利CN105853403A的启发是远远不够的，因为本发明提供的一种多西他赛棕榈酸酯脂质体，其创新技术特征就是含有螯合剂，同时也收到了实质性的效果。
本发明的第一目的，是提供一种多西他赛棕榈酸酯脂质体。
本发明提供一种多西他赛棕榈酸酯脂质体，以多西他赛棕榈酸酯为主药。多西他赛棕榈酸酯用量为0.1-2％(重量体积百分比)。
本发明提供一种多西他赛棕榈酸酯脂质体，以多西他赛棕榈酸酯为主药，还包括螯合剂。多西他赛棕榈酸酯用量为0.1-2％；螯合剂用量0.001-1％(重量体积百分比)。
本发明提供一种多西他赛棕榈酸酯脂质体，以多西他赛棕榈酸酯为主药，还包括螯合剂、卵磷脂以及DSPE-PEG2000。多西他赛棕榈酸酯用量为0.1-2％；螯合剂用量0.001-1％；卵磷脂用量1-10％，DSPE-PEG2000用量0.05-1％(重量体积百分比)。
本发明的第二个目的，是提供一种多西他赛棕榈酸酯脂质体，是冻干粉针剂，也可以是注射用脂质体溶液。
本发明的第三个目的，是提供一种多西他赛棕榈酸酯前药，所述的多西他赛棕榈酸酯前药以多西他赛为母药，以酯键链接一分子棕榈酸而形成前药，是一种稳定性好、功能性强的脂溶性前药。
所述的多西他赛棕榈酸酯前药的结构如下：
Figure PCTCN2019084543-appb-000002
多西他赛棕榈酸酯化学结构式
所述的多西他赛棕榈酸酯前药，其特征在于，棕榈酸与多西他赛侧链2’位相连，其制备过程如下：多西他赛10.00g、棕榈酸3.81g、1-(3-二甲氨基丙基)-3-乙基碳二亚胺(EDC)2.43g和4-二甲氨基吡啶(DMAP)1.82g，置于反应容器中，用50mL无水二氯甲烷使溶解，在氮气保护条件下，室温搅拌反应4-24h，得反应溶液；将反应溶液用5％柠檬酸水溶液洗涤两遍，再用饱和氯化钠溶液洗涤一遍，旋转蒸发减压去除无水二氯甲烷；分离纯化，即得到多西他赛棕榈酸酯。反应合成路线图如下所示：
Figure PCTCN2019084543-appb-000003
多西他赛棕榈酸酯合成线路图
本发明提供一种多西他赛棕榈酸酯脂质体，所述的脂质体是一种含有螯合剂的注射剂；所述含有螯合剂的注射剂也可以是注射用溶液，也可以是一种冻干粉针；含有螯合剂是本发明的核心技术特征。
所述的一种多西他赛棕榈酸酯脂质体，具体由下列配方配制而成：
Figure PCTCN2019084543-appb-000004
制备注射用溶液时，冻干保护剂为0；
制备注射用冻干粉针时，冻干保护剂为0.1-40％克/毫升
较优的，所述的一种多西他赛棕榈酸酯脂质体，具体由下列配方配制而成：
Figure PCTCN2019084543-appb-000005
较优的，所述的一种多西他赛棕榈酸酯脂质体，具体由下列配方配制而成：
Figure PCTCN2019084543-appb-000006
其中，上述配方中所述的卵磷脂，选自高纯蛋黄卵磷脂(EPCS)、氢化大豆卵磷脂(HSPC)、二棕榈酰磷脂酰胆碱(DPPC)、磷脂酰胆碱、蛋黄卵磷脂、大豆卵磷脂、磷脂酰丝氨酸、二肉豆蔻酰磷脂酰胆碱、二硬脂酰磷脂酰胆碱、磷脂酰乙醇胺、鞘磷脂中的一种或两种以上合用；优选高纯蛋黄卵磷脂(EPCS)、氢化大豆卵磷脂(HSPC)中的一种或两种以上合用。
上诉配方中所述的螯合剂选自柠檬酸、柠檬酸二钠、柠檬酸三钠、乳酸、乳酸钠、苹果酸、苹果酸钠、乙二胺四乙酸、乙二胺四乙酸二钠、乙二胺四乙酸三钠中的一种或者两种以上的混合物，优选柠檬酸、柠檬酸二钠、柠檬酸三钠、乳酸、乳酸钠中的一种或两种以上合用。
上述配方中所述的冻干保护剂为海藻糖、蔗糖、麦芽糖、乳糖、甘露醇、葡萄糖、山梨醇、木糖醇、赤藓糖醇、苏氨酸中的一种或两种以上；优选甘露醇、海藻糖、蔗糖中的一种或两种以上合用。
上述配方中所述的pH调节剂为氢氧化钠、盐酸中的一种或两种以上。
本发明的第四个目的，是提供所述的一种多西他赛棕榈酸酯脂质体的制备方法。
所述的一种多西他赛棕榈酸酯脂质体的制备方法为注入法。
所述的一种多西他赛棕榈酸酯脂质体由如下步骤制备而成：
称取处方量的多西他赛棕榈酸脂酯、胆固醇、磷脂、DSPE-PEG2000、螯合剂，置于注入用有机溶媒中，在25～70℃下加热使溶解，得有机相；称取注射用水适量，加热至25～70℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品进行乳化，可将其置于高压均质机中进行均质乳化，或将其置于挤出器中依次通过不同孔径的挤出膜挤出，或高压均质后再进行挤出，得脂质体溶液；称取配方量冻干保护剂，置于上述脂质体溶液中，搅拌使溶解，并用注射用水定容至全量；用pH调节剂调节pH值；过0.22μm滤膜除菌、分装、封口，即得多西他赛棕榈酸酯脂质体；也可冻干制备成多西他赛棕榈酸酯脂质体冻干粉针。
其中，所述的注入用有机溶媒，选自丙二醇、无水乙醇、叔丁醇中的一种或两种以上合用，用量为1-8％克/毫升；优选无水乙醇，用量优选为2-5％克/毫升。
所述的注入用有机溶媒，可保留在脂质体中，也可在脂质体粗品乳化后再通过超滤去除，也可通过冷冻干燥去除。
所述的将脂质体粗品进行乳化，优选挤出乳化法，得到的脂质体粒径分布将更均一； 挤出膜孔径选自0.8μm、0.6μm、0.4μm、0.2μm、0.1μm、0.05μm，选用一种或两种以上依次通过大孔径到小孔径的挤出，优选0.4μm、0.2μm、0.1μm、0.05μm。
所述的螯合剂可以溶解在油相中，也可以溶解在水相中，也可以溶解在脂质体溶液中。
所述的冻干保护剂溶解在脂质体溶液中，也可以溶解在水相中。
发明的一种多西他赛棕榈酸酯脂质体，粒径为50-150nm。
本发明的一种多西他赛棕榈酸酯脂质体，含有螯合剂是本发明的核心技术特征，螯合剂的添加使得多西他赛体内作用时间更长、抗肿瘤效果更佳、制剂相关特性更好，这是本发明实质性效果的根本体现。
具体实施方式
以下结合具体实施例，对本发明作进一步说明。应理解，以下实施例仅用于说明本发明而非用于限定本发明的范围。
实施例1：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.5g、高纯蛋黄卵磷脂(EPCS)3g、0.3g DSPE-PEG2000，置于4g无水乙醇中，在50℃下加热使溶解，得有机相；称取乙二胺四乙酸二钠0.05g，置于90g注射用水中，在50℃下加热搅拌使溶解，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品分别用孔径为0.4μm、0.1μm、0.05μm的挤出膜挤出，得脂质体溶液；用注射用水定容至100ml；用盐酸调节剂调节pH值4.5；过0.22μm滤膜除菌、分装、封口，即得多西他赛棕榈酸酯脂质体溶液。
经测定，平均粒径为92.4nm。
实施例2：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.3g、高纯蛋黄卵磷脂(EPCS)3g、0.2g DSPE-PEG2000、柠檬酸0.1g，置于4g丙二醇中，在60℃下加热使溶解，得有机相；称取注射用水70g，加热至60℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品分别用孔径为0.4μm、0.2μm、0.1μm、0.05μm的挤出膜挤出，得脂质体溶液；称取蔗糖15g、甘露醇5g，置于上述脂质体溶液中，搅拌使溶解，并用注射用水定容至100ml；用氢氧化钠调节pH值为5.5；过0.22μm滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉。
经测定，平均粒径为86.6nm。
实施例3：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.7g、高纯蛋黄卵磷脂(EPCS)6g、0.5g DSPE-PEG2000、乳酸0.3g，置于6g无水乙醇中，在45℃下加热使溶解，得有机相；称取注射用水65g，加热至45℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品分别用孔径为0.6μm、0.4μm、0.1μm的挤出膜挤出，得脂质体溶液；称取海藻糖20g，置于上述脂质体溶液中，搅拌使溶解，并用注射用水定容至100ml；用氢氧化钠调节pH值为6.2；过0.22μm滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉。
经测定，平均粒径为120.7nm。
实施例4：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.3g、高纯蛋黄卵磷脂(EPCS)5g、0.3g DSPE-PEG2000、苹果酸0.1g、柠檬酸0.2g，置于5g无水乙醇中，在65℃下加热使溶解，得有机相；称取注射用水70g，加热至65℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品用高压均质机均质乳化，再分别用孔径为0.1μm、0.05μm的挤出膜挤出，得脂质体溶液；称取蔗糖10g、海藻糖5g，置于上述脂质体溶液中，搅拌使溶解，并用注射用水定容至100ml；用氢氧化钠调节pH值为6.0；过0.22μm滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉。
经测定，平均粒径为50.36nm。
实施例5：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.2g、高纯蛋黄卵磷脂(EPCS)3g、0.2g DSPE-PEG2000、柠檬酸0.01g，置于3g无水乙醇中，在50℃下加热使溶解，得有机相；称取注射用水75g，加热至50℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品用高压均质机均质乳化，得脂质体溶液；称取蔗糖10g、甘露醇5g，置于上述脂质体溶液中，搅拌使溶解，并用注射用水定容至100ml；用氢氧化钠调节pH值为7.0；过0.22μm滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉。
经测定，平均粒径为60.7nm。
实施例6：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.7g、高纯蛋黄卵磷脂(EPCS)6g、0.7g DSPE-PEG2000、胆固醇0.5g、柠檬酸0.5g，置于6g无水乙醇中，在55℃下加热使溶解，得有机相；称取注射用水80g，加热至55℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品用高压均质机均质乳化，得脂质体溶液；用注射用水定容至100ml；用氢氧化钠调节pH值为4.8；过0.22μm滤膜除菌、分装、封口，即得紫杉醇棕榈酸酯脂质体溶液。
经测定，平均粒径为80.4nm。
实施例7：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.7g、蛋黄卵磷脂2g、氢化大豆卵磷脂(HSPC)1g、0.5g DSPE-PEG2000、胆固醇0.1g，置于4g无水乙醇中，在55℃下加热使溶解，得有机相；称取柠檬酸三钠0.2g、海藻糖10g、甘露醇12g、葡萄糖8g，置于60g注射用水中，加热至55℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品分别用孔径为0.8μm、0.4μm、0.2μm的挤出膜挤出，得脂质体溶液；用注射用水定容至100ml；用盐酸调节pH值为4.5；过0.22μm滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉。
经测定，平均粒径为150.0nm。
实施例8：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.8g、二棕榈酰磷脂酰胆碱(DPPC)3g、磷脂酰胆碱3g、蛋黄卵磷脂1g、0.8g DSPE-PEG2000，置于8g丙二醇中，在70℃下加热使溶解，得有机相；称取注射用水80g，加热至55℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品分别用孔径为0.8μm、0.4μm、0.2μm、0.1μm的挤出膜挤出，得脂质体溶液；用超滤仪超滤去除丙二醇；称取柠檬酸三钠0.8g，置于超滤后的脂质体溶液中，搅拌溶解；用注射用水定容至100ml；用盐酸调节pH值为9.0；过0.22μm滤膜除菌、分装、封口，即得紫杉醇棕榈酸酯脂质体溶液。
经测定，平均粒径为145.2nm。
实施例9：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.1g、大豆卵磷脂2g，0.1g DSPE-PEG2000，置于6g无水乙醇中，在25℃下搅拌使溶解，得有机相；称取乙二胺四乙酸三钠0.5g、柠檬酸二钠0.5g，置于注射用水80g中，在25℃下加热搅拌使溶解，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品分别用孔径为0.6μm、0.2μm、0.1μm、0.05μm的挤出膜挤出，得脂质体溶液；用注射用水定容至100ml；用盐酸调节pH值为3.5；过0.22μm滤膜除菌、分装、封口，即得紫杉醇棕榈酸酯脂质体溶液。
经测定，平均粒径为118.8nm。
实施例10：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂1.0g、高纯蛋黄卵磷脂(EPCS)10g、1.0g DSPE-PEG2000、胆固醇1g，置于10g无水乙醇中，在60℃下加热使溶解，得有机相；称取注射用水74g，加热至60℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品用高压均质机均质乳化，得脂质体溶液；用超滤仪超滤去除无水乙醇；称取乳酸钠0.5g、苹果酸钠0.5g，置于超滤后的脂质体溶液中，搅拌使溶解；用注射用水定容至100ml；用盐酸调节pH值为5.0；过0.22μm滤膜除菌、分装、封口，即得紫杉醇棕榈酸酯脂质体溶液。
经测定，平均粒径为130.2nm。
实施例11：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.1g、二硬脂酰磷脂酰胆碱1g、0.05g DSPE-PEG2000，置于1g无水乙醇中，在55℃下加热使溶解，得有机相；称取注射用水95g，加热至55℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品用高压均质机均质乳化，得脂质体溶液；称取乳酸钠0.001g，置于上述脂质体溶液中，搅拌使溶解；用注射用水定容至100ml；用氢氧化钠调节pH值为8.0；过0.22μm滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉针剂。
经测定，平均粒径为90.7nm。
实施例12：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.1g、磷脂酰乙醇胺1g、二肉豆蔻酰磷脂酰胆碱1g、0.5g DSPE-PEG2000、乙二胺四乙酸0.005g，置于4g无水乙醇中，在55℃下加热使溶解，得有机相；称取海藻糖5g，置于70g注射用水中，加热至55℃，得水相；将有机相在搅拌 条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品分别用孔径为0.8μm、0.6μm、0.4μm、0.1μm的挤出膜挤出，得脂质体溶液；用注射用水定容至100ml；用氢氧化钠调节pH值为7.5；过0.22μm滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉。
经测定，平均粒径为104.3nm。
实施例13：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.2g、磷脂酰丝氨酸1g、鞘磷脂1g、0.2g DSPE-PEG2000、柠檬酸0.01g，置于2g无水乙醇与4g丙二醇的混合溶剂中，在70℃下加热使溶解，得有机相；称取注射用水50g，加热至70℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品用高压均质机均质乳化，得脂质体溶液；称取蔗糖15g、甘露醇15g、赤藓糖醇5g、苏氨酸5g，置于上述脂质体溶液中，搅拌使溶解，并用注射用水定容至100ml；用氢氧化钠调节pH值为6.0；过0.22μm滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉。
经测定，平均粒径为78.4nm。
实施例14：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.4g、高纯蛋黄卵磷脂(EPCS)5g、0.5g DSPE-PEG2000、柠檬酸0.01g，置于5g无水乙醇中，在50℃下加热使溶解，得有机相；称取注射用水50g，加热至50℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品用高压均质机均质乳化，再分别用孔径为0.2μm、0.1μm、0.05μm的挤出膜挤出的，得脂质体溶液；称取木糖醇10g、山梨醇15g、甘露醇10g，置于上述脂质体溶液中，搅拌使溶解，并用注射用水定容至100ml；用氢氧化钠调节pH值为6.0；过0.22μm滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉。
经测定，平均粒径为140.7nm。
实施例15：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.3g、高纯蛋黄卵磷脂(EPCS)3g、0.1g DSPE-PEG2000，置于5g叔丁醇中，在45℃下加热使溶解，得有机相；称取注射用水80g，加热至45℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品用高压均质机均质乳化，得脂质体溶液；称取柠檬酸三钠0.1g、蔗糖13g、甘露醇5g，置于上述脂质体溶液中，搅拌使溶解；用注射用水定容至100ml；用盐酸调节pH值为5.5；过0.22μm滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉针剂。
经测定，平均粒径为105.3nm。
实施例16：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.3g、高纯蛋黄卵磷脂(EPCS)1.5g、氢化大豆卵磷脂(HSPC)0.5g、0.1g DSPE-PEG2000，置于6g无水乙醇中，在55℃下加热使溶解，得有机相；称取注射用水60g，加热至55℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品用高压均质机均质乳化，得脂质体溶液；用超滤仪超滤去除无水乙醇；称取乙二胺四乙酸二钠0.3g、海藻糖19g、乳糖5g，置于超滤后的脂质体溶液中，搅拌使溶解；用注射用水定容至100ml；用氢氧化钠调节pH值为7.0；过0.22μm 滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉针剂。
经测定，平均粒径为89.4nm。
实施例17：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.3g、高纯蛋黄卵磷脂(EPCS)3g、0.7g DSPE-PEG2000、柠檬酸0.1g，置于4g丙二醇中，在70℃下加热使溶解，得有机相；称取蔗糖10g、海藻糖5g，置于70g注射用水中，加热至70℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品分别用孔径为0.8μm、0.6μm、0.4μm、0.1μm的挤出膜挤出，得脂质体溶液；用注射用水定容至100ml；用氢氧化钠调节pH值为6.0；过0.22μm滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉。
经测定，平均粒径为113.6nm。
实施例18：多西他赛棕榈酸酯脂质体的制备
称取处方量的多西他赛棕榈酸脂脂0.3g、高纯蛋黄卵磷脂(EPCS)3g、0.5g DSPE-PEG2000、柠檬酸0.01g，置于4g丙二醇中，在60℃下加热使溶解，得有机相；称取蔗糖17g、甘露醇5g，置于65g注射用水中，加热至70℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品分别用孔径为0.6μm、0.4μm、0.1μm、0.05μm的挤出膜挤出，得脂质体溶液；用注射用水定容至100ml；用氢氧化钠调节pH值为6.7；过0.22μm滤膜除菌、分装、冻干、封口，即得紫杉醇棕榈酸酯脂质体冻干粉。
经测定，平均粒径为106.3nm。
实施例19：螯合剂对多西他赛棕榈酸酯脂质体抗肿瘤效果的影响
处方中含有螯合剂，是本发明的多西他赛棕榈酸酯脂质体现实质性效果的关键所在。为了进一步验证螯合剂在本发明中的优越性，采取多次平行对比的方式，在相同处方工艺下分别制备含螯合剂与不含螯合剂的脂质体，以小鼠S180肉瘤作为肿瘤模型，分别对含螯合剂与不含螯合剂的多西他赛棕榈酸酯脂质体进行抗肿瘤效果比较，试验设计与结果如下所示。
1.样品来源
取市售多西他赛注射液，作为阳性对照药；取实施例1制备的含螯合剂多西他赛棕榈酸酯脂质体，作为含螯合剂供试制剂；完全按照实施例1的处方工艺，平行制备不含螯合剂的脂质体，作为不含螯合剂对照制剂。
2.小鼠S180肿瘤模型的建立与给药方案的设计
小鼠腹水瘤S180细胞，培养于DMEM培养液中，37℃，5％CO 2下常规培养，平均每两天传代一次，至对数生长期时用培养基调整细胞浓度5×10 7个/mL，在无菌条件下注射于小鼠腹腔内，待一周左右后可见明显腹水时，无菌条件下取荷瘤小鼠腹水，用生理盐水以1:5适量稀释，接种0.2mL于小鼠腹腔内。待一周左右可见明显二代腹水时，无菌条件下取荷瘤小鼠二代腹水，用生理盐水1:5适量稀释，于小鼠左侧腋窝皮下注射S180细胞悬液，每只0.2mL。
将接种24h后的ICR小鼠称重，随机分为4组，每组8只。分别设定空白组、市售多西他赛注射液组、不含螯合剂对照组、含螯合剂对照组。采用小鼠尾静脉注射方式给药， 剂量为每次10mg/kg(按多西他赛计)，空白组给于0.2ml生理盐水，隔一天给一次药，总共给药4次。停药第三日处死小鼠，称取小鼠体重，剥离瘤体称重，计算抑瘤率。
抑瘤率＝(生理盐水组瘤重-给药组瘤重)/生理盐水组瘤重×100％
3.抗肿瘤效果
以小鼠S180肉瘤为模型，考察含螯合剂与不含螯合剂的多西他赛棕榈酸酯脂质体以及市售多西他赛注射液的抗肿瘤效果，结果如表1所示。
表1含螯合剂与不含螯合剂的多西他赛棕榈酸酯脂质体以及市售多西他赛注射液抗肿瘤效果对比结果
组别 平均瘤重(g) 抑瘤率
空白组 1.17±0.49 /
市售多西他赛注射液组 0.44±0.21 62.39％
不含螯合剂多西他赛棕榈酸酯脂质体对照组 0.30±0.11 74.36％
含螯合剂多西他赛棕榈酸酯脂质体供试组 0.21±0.06 82.05％
结果分析：
①含螯合剂与不含螯合剂的多西他赛棕榈酸酯脂质体抗肿瘤效果，均显著好于市售多西他赛注射液，说明将多西他赛开发成前药多西他赛棕榈酸酯脂质体后，抗肿瘤效果明显提高，是本发明实质性效果的重要方面。
②平行对比含螯合剂与不含螯合剂的多西他赛棕榈酸酯脂质体抗肿瘤效果。结果显示，含螯合剂脂质体抗肿瘤效果要好于不含螯合剂组。
终上所述，处方中含有螯合剂，多西他赛棕榈酸酯脂质体的抗肿瘤效果将更好，这是本发明实质性效果的体现，故含有螯合剂是本发明的核心技术特征。
实施例20：螯合剂对多西他赛棕榈酸酯脂质体体内药代动力学的影响
1.样品来源
取市售的多西他赛注射液，作为参比制剂；取实施例1制备的多西他赛棕榈酸脂质体，作为含螯合剂多西他赛棕榈酸脂质体样品；完全按照实施例1的处方工艺制备得不含螯合剂的多西他赛棕榈酸脂质体，作为不含螯合剂的多西他赛棕榈酸脂质体样品。
2.药代动力学试验设计
取SD雄性大鼠18只，随机分为三组，每组6只，实验前禁食过夜，自由饮水。分别设定市售多西他赛注射液组、不含螯合剂多西他赛棕榈酸脂质体、含螯合剂多西他赛棕榈酸脂质体组。采用大鼠尾静脉注射方式给药，剂量为每次10mg/kg(按多西他赛计)，分别于给药后0.033h、0.083h、0.167h、0.25h、0.5h、0.75h、1h、1.5h、2h、3h、4h、6h、8h、10h、12h、24h经眼眶静脉丛取血0.5mL，置于含有肝素钠的离心管并摇匀，4500r/min离心10min，取血浆150μl，于-20℃下冻存。按常规方法进行血浆样品的处理，用高效液相色谱法测定血浆中多西他赛的浓度。
3.结果与分析
采用DAS 2.0软件进行模型拟合，计算得药动学参数。市售多西他赛注射液组、含螯合剂多西他赛棕榈酸脂质体组、不含螯合剂多西他赛棕榈酸酯脂质体的体内药代动力学结 果如表2所示。
表2含螯合剂与不含螯合剂多西他赛棕榈酸酯脂质体以及市售多西他赛注射液多西他赛大鼠药代动力学主要参数的比较
Figure PCTCN2019084543-appb-000007
结果分析：
1.多西他赛棕榈酸酯脂质体，AUC∞与T 1/2明显比市售多西他赛棕榈酸酯注射液大。该实验结果，说明将多西他赛制备成前药脂质体后，明显延缓药物在体内的代谢，使作用时间更长。
2.含螯合剂的多西他赛棕榈酸酯脂质体，与不含螯合剂的多西他赛棕榈酸酯脂质体比，AUC∞增大，T 1/2延长。从药效来看，含螯合剂的脂质体抗肿瘤效果更好，也许就是因为其具有体内相对较长的作用时间，故抗肿瘤效果更好。
终上所述，在多西他赛棕榈酸酯脂质体的处方中添加螯合剂后，延长了药物的体内作用时间，提高了抗肿瘤作用。由此可见，处方中含有螯合剂，对本发明的多西他赛棕榈酸酯脂质体优越性的体现尤为重要，是关键技术特征。
实施例21：螯合剂对多西他赛棕榈酸酯脂质体制剂特性的影响
研究的是一种无菌脂质体，而脂质体在生产过程中无法高温灭菌，故人们通常用0.22μm的滤膜进行除菌过滤，达到无菌的效果。而在实际生产过程中，往往由于脂质体粒径较大或者是粒径分布不均匀导致除菌过滤不顺畅，严重影响生产效率。就多西他赛棕榈酸酯脂质体而言在研究过程中，我们特别关注除菌过滤的顺畅程度，发现螯合剂的添加能够使过滤性更顺畅。通过检测，加螯合剂的脂质体，粒径略小，粒径分布PDI更小，故过滤更加顺畅。试验设计与结果如下所示。
以实施例1为例，完全按照实施例1记载的处方工艺分别制备含螯合剂与不含螯合剂的多西他赛棕榈酸酯液体脂质体1000ml，分别用直径11mm的板式滤器，0.22μm的聚醚砜滤膜过滤，记录过滤体积；分别测定含螯合剂与不含螯合剂的多西他赛棕榈酸酯液体脂质体的粒径与粒径分布PDI，试验结果见表3所示：
表3本发明的多西他赛棕榈酸酯脂质体制剂相关特性的比较
指标 不含螯合剂多西他赛棕榈酸酯脂质体 含螯合剂多西他赛棕榈酸酯脂质体
平均粒径 102.1nm 92.4nm
PDI 0.237 0.124
过滤量 625ml 867ml
结果分析：
本发明的多西他赛棕榈酸酯脂质体粒径更小、分布更窄、除菌过滤更顺畅。由此可见，添加螯合剂后，制剂基本特性明显改善，生产实施将更加顺畅。进一步体现了处方中含螯合剂的优越性。
以上已对本发明创造的较佳实施例进行了具体说明，但本发明创造并不限于所述实施例，熟悉本领域的技术人员在不违背本发明创造精神的前提下还可做出种种的等同的变型或替换，这些等同的变型或替换均包含在本申请权利要求所限定的范围内。

Claims (17)

  1. 一种多西他赛棕榈酸酯脂质体，以多西他赛棕榈酸酯为主药，还包括螯合剂、卵磷脂以及DSPE-PEG2000；且多西他赛棕榈酸酯用量为0.1-2％，螯合剂用量为0.001-1％，卵磷脂用量为1-10％，DSPE-PEG2000用量为0.05-1％。
  2. 根据权利要求1所述的多西他赛棕榈酸酯脂质体，其特征在于，该脂质体为一种冻干粉针剂。
  3. 根据权利要求1所述的多西他赛棕榈酸酯脂质体，其特征在于，该脂质体是一种注射用脂质体溶液。
  4. 根据权利要求1、2或3所述的多西他赛棕榈酸酯脂质体，其特征在于，具体由下列配方配制而成：
    Figure PCTCN2019084543-appb-100001
  5. 根据权利要求1、2或3所述的多西他赛棕榈酸酯脂质体，其特征在于，具体由下列配方配制而成：
    Figure PCTCN2019084543-appb-100002
  6. 根据权利要求1或2或3所述的多西他赛棕榈酸酯脂质体，其特征在于，具体由下列配方配制而成：
    Figure PCTCN2019084543-appb-100003
  7. 根据权利要求1-6任一项所述的多西他赛棕榈酸酯脂质体，其特征在于，所述的卵磷脂选自高纯蛋黄卵磷脂(EPCS)、氢化大豆卵磷脂(HSPC)、二棕榈酰磷脂酰胆碱(DPPC)、磷脂酰胆碱、蛋黄卵磷脂、大豆卵磷脂、磷脂酰丝氨酸、二肉豆蔻酰磷脂酰胆碱、二硬脂酰磷脂酰胆碱、磷脂酰乙醇胺、鞘磷脂中的一种或两种以上。
  8. 据权利要求1-6任一项所述的多西他赛棕榈酸酯脂质体，其特征在于，所述的螯合剂选自柠檬酸、柠檬酸二钠、柠檬酸三钠、乳酸、乳酸钠、苹果酸、苹果酸钠、乙二胺四乙酸、乙二胺四乙酸二钠、乙二胺四乙酸三钠中的一种或者两种以上。
  9. 根据权利要求4-6任一项所述的多西他赛棕榈酸酯脂质体，其特征在于，所述的冻干保护剂选自海藻糖、蔗糖、麦芽糖、乳糖、甘露醇、葡萄糖、山梨醇、木糖醇、赤藓糖醇、苏氨酸中的一种或两种以上。
  10. 根据权利要求4-6任一项所述的多西他赛棕榈酸酯脂质体，其特征在于，所述的pH调节剂为氢氧化钠、盐酸中的一种或两种以上。
  11. 根据权利要求1-10任一项所述的多西他赛棕榈酸酯脂质体，其特征在于，脂质体的粒径为50-150nm。
  12. 如权利要求1所述的多西他赛棕榈酸酯脂质体的制备方法，其特征在于，所述的制备方法如下：
    称取处方量的多西他赛棕榈酸脂酯、胆固醇、磷脂、DSPE-PEG2000、螯合剂，置于注入用有机溶媒中，在25～70℃下加热使溶解，得有机相；称取注射用水适量，加热至25～70℃，得水相；将有机相在搅拌条件下注入水相中，混匀，即得脂质体粗品；将脂质体粗品进行乳化，可将其置于高压均质机中进行均质乳化，或将其置于挤出器中依次通过不同孔径的挤出膜挤出，或高压均质后再进行挤出，得脂质体溶液；称取配方量冻干保护剂，置于上述脂质体溶液中，搅拌使溶解，并用注射用水定容至全量；用pH调节剂调节pH值；过0.22μm滤膜除菌、分装、封口，即得多西他赛棕榈酸酯脂质体；也可冻干制备成多西他赛棕榈酸酯脂质体冻干粉针。
  13. 根据权利要求12所述的多西他赛棕榈酸酯脂质体的制备方法，其特征在于，所述的注入用有机溶媒，选自丙二醇、无水乙醇、叔丁醇中的一种或两种以上，用量为1-10％克/毫升。
  14. 根据权利要求12所述的多西他赛棕榈酸酯脂质体的制备方法，其特征在于，所述的注入用有机溶媒，可保留在脂质体中，也可在脂质体粗品乳化后再通过超滤去除，也可通过冷冻干燥去除。
  15. 根据权利要求12所述的多西他赛棕榈酸酯脂质体的制备方法，其特征在于，所述的将脂质体粗品进行乳化，挤出膜孔径选自0.8μm、0.6μm、0.4μm、0.2μm、0.1μm、0.05μm，选用一种，或两种以上依次通过大孔径到小孔径的挤出。
  16. 根据权利要求12所述的多西他赛棕榈酸酯脂质体的制备方法，其特征在于，所述的螯合剂溶解在油相中、水相中或者脂质体溶液中。
  17. 根据权利要求12所述的多西他赛棕榈酸酯脂质体的制备方法，其特征在于，所述的冻干保护剂溶解在脂质体溶液中，或者溶解在水相中。
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