WO2024067849A1 - Liposome pharmaceutical composition, preparation method therefor, and use thereof - Google Patents

Liposome pharmaceutical composition, preparation method therefor, and use thereof Download PDF

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Publication number
WO2024067849A1
WO2024067849A1 PCT/CN2023/122824 CN2023122824W WO2024067849A1 WO 2024067849 A1 WO2024067849 A1 WO 2024067849A1 CN 2023122824 W CN2023122824 W CN 2023122824W WO 2024067849 A1 WO2024067849 A1 WO 2024067849A1
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Prior art keywords
liposome
drug
treprostinil
aqueous phase
acid
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PCT/CN2023/122824
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French (fr)
Chinese (zh)
Inventor
姚萌
石晓晴
林一峯
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上海济煜医药科技有限公司
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Publication of WO2024067849A1 publication Critical patent/WO2024067849A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/557—Eicosanoids, e.g. leukotrienes or prostaglandins
    • A61K31/5575—Eicosanoids, e.g. leukotrienes or prostaglandins having a cyclopentane, e.g. prostaglandin E2, prostaglandin F2-alpha
    • 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
    • A61K31/557—Eicosanoids, e.g. leukotrienes or prostaglandins
    • A61K31/558—Eicosanoids, e.g. leukotrienes or prostaglandins having heterocyclic rings containing oxygen as the only ring hetero atom, e.g. thromboxanes
    • A61K31/5585—Eicosanoids, e.g. leukotrienes or prostaglandins having heterocyclic rings containing oxygen as the only ring hetero atom, e.g. thromboxanes having five-membered rings containing oxygen as the only ring hetero atom, e.g. prostacyclin
    • 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
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system

Definitions

  • patent application number 202211216387.5 entitled “A liposome, a pharmaceutical composition, a preparation method and application thereof”, filed with the State Intellectual Property Office of China on September 30, 2022;
  • the invention belongs to the field of medicines, and in particular relates to a liposome medicine composition and a preparation method and application thereof.
  • Liposomes are vesicles formed by a lipid bilayer containing an internal aqueous medium. Liposomes have been used as carriers for various therapeutic agents to provide improved delivery characteristics, such as enhancing drug circulation time in the blood, reducing cytotoxicity, sustained release of drugs, and delivery of specific drugs to selected tissues. When using liposomes for the delivery of therapeutic drugs, high drug encapsulation efficiency and drug content are required to reduce potential phospholipid excipient toxicity.
  • the present invention provides a liposome pharmaceutical composition, comprising a weakly acidic drug and a liposome; the liposome is one of the following:
  • the liposome comprises phospholipids and an internal aqueous phase
  • the liposome comprises phospholipids, an inner aqueous phase and an outer aqueous phase;
  • the internal aqueous phase comprises meglumine and a weak acid
  • the external aqueous phase suspends the liposomes.
  • the weakly acidic drug is encapsulated in the inner aqueous phase of the liposome, and the liposome encapsulating the weakly acidic drug is suspended in the outer aqueous phase.
  • the pKa of the weakly acidic drug is between 2 and 7.
  • the drug loading of the weakly acidic drug is not less than 18%, for example, 18% to 40%, and can also be 19%-30%, or 20-25%, such as 20.5%, 21%, 21.5%, 22%, 22.5%, 23%, 23.3%, 23.5% or 35%.
  • the encapsulation efficiency of the drug liposome is not less than 85%, preferably not less than 90%.
  • the encapsulation efficiency of the drug liposome is not less than 85%, preferably not less than 90%. More preferably, the drug loading of the drug liposome is not less than 18%, and the encapsulation efficiency of the drug liposome is not less than 85%, preferably not less than 90%.
  • the weakly acidic drug may be selected from: at least one drug selected from prostaglandins, antipyretics, quinoline carboxylic acid antibiotics, and interferon gene stimulator (STING) receptors;
  • the prostaglandin drugs are such as treprostinil, beraprost, iloprost, carboprost, limaprost, epoprostenol, alprostadil, unoprostone, or their derivatives (such as pharmaceutically acceptable salts, esters or prodrugs);
  • the antipyretic and analgesic drugs are such as aspirin, ibuprofen, naproxen, diclofenac sodium, or their derivatives (such as pharmaceutically acceptable salts, esters or prodrugs);
  • the quinoline carboxylic acid antibacterial drugs are such as nalidixic acid, pyrrolopyrimic acid, fluoroquinolinic acid, sitafloxacin, ciprofloxacin, enoxacin, or their derivatives (such as pharmaceutically acceptable salts, esters or prodrugs);
  • the STING drugs are such as MSA-2, STING agonist-7, Vadimezan, or their derivatives (such as pharmaceutically
  • the pH of the internal aqueous phase is 4.0 to 10.5, such as 4.5 to 10.0, 6.5 to 10.5, preferably 4.97 to 9.52;
  • the pH of the internal aqueous phase is 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, preferably 8.5.
  • the meglumine in the internal aqueous phase provides the cationic part of the internal aqueous phase
  • the weak acid provides the anionic part of the internal aqueous phase
  • the weak acid is selected from “carboxylic acids”, for example, formic acid, acetic acid, propionic acid, butyric acid, isobutyric acid, valeric acid, isovaleric acid, lactic acid or a combination thereof.
  • formic acid, acetic acid, propionic acid or a combination thereof Preferably, formic acid, acetic acid, propionic acid or a combination thereof.
  • the internal aqueous phase in the embodiments of the present invention does not contain carbonate or bicarbonate, but the internal aqueous phase system is realized by meglumine and weak acid.
  • the concentration of meglumine in the internal aqueous phase is 0.1 to 0.8 M, such as 0.2 to 0.6 M, preferably 0.3 to 0.4 M, such as 0.3 M, 0.35 M, 0.4 M;
  • the concentration of the weak acid in the inner aqueous phase is 0.1 to 0.8 M, for example 0.2 to 0.6 M, such as 0.2 M, 0.211 M, 0.268 M, 0.286 M, 0.3 M, 0.35 M, 0.40 M, 0.5 M, 0.6 M;
  • the meglumine and the weak acid are meglumine and acetic acid; preferably, the concentration of the meglumine and acetic acid in the internal aqueous phase is 0.30-0.40M.
  • the phospholipid has a bilayer lipid structure, and the bilayer lipid is composed of hydrogenated soybean lecithin (HSPC), cholesterol (CHOL) and distearoyl phosphatidylethanolamine-polyethylene glycol 2000 (DSPE-mPEG2000), and the molar ratio is 3:(1-3):(0.025-0.225), for example, 3:(2-3):(0.025-0.075), 3:(1.5-2.5):(0.025-0.075).
  • HSPC hydrogenated soybean lecithin
  • CHOL cholesterol
  • DSPE-mPEG2000 distearoyl phosphatidylethanolamine-polyethylene glycol 2000
  • the bilayer lipid is composed of hydrogenated soybean lecithin (HSPC), cholesterol (CHOL) and distearoyl phosphatidylethanolamine-polyethylene glycol 2000 (DSPE-mPEG2000), and the molar ratio is 3:2:(0.025-0.225), for example 3:2:(0.025-0.15), preferably 3:2:(0.025-0.075).
  • HSPC hydrogenated soybean lecithin
  • CHOL cholesterol
  • DSPE-mPEG2000 distearoyl phosphatidylethanolamine-polyethylene glycol 2000
  • the concentration of the phospholipid in the liposome composition is 1 to 100 mg/mL, for example, 1 to 50 mg/mL, 1 to 20 mg/mL, and in the preparation process of the liposome pharmaceutical composition of the present invention, the concentration of the phospholipid used is also, for example, 1 to 15 mg/mL, 1.2 to 5.0 mg/mL, preferably 1.3 to 4.5 mg/mL, 1.4 to 4.18 mg/mL.
  • the particle size of the liposome is 50-500 nm, for example, 100-200 nm, preferably 100-150 nm, 120-180 nm, 130-160 nm, 140-190 nm.
  • the external aqueous phase is a buffer solution with a pH of 4.5 to 6.5.
  • the buffer for achieving the corresponding pH value of the external aqueous phase is well known in the art and is not particularly limited.
  • the commonly used buffer in the art is selected from one or more of HEPES, sodium chloride, sucrose, citrate buffer, phosphate buffer, and Tris buffer.
  • the buffer can be selected from citrate buffer; the concentration of the buffer is 0.05M to 0.25M, for example, 0.08M to 0.2M, preferably 0.09 to 0.12M.
  • the liposome has a pH gradient property with a high pH in the inner aqueous phase and a low pH in the outer aqueous phase, wherein the pH value of the outer aqueous phase is higher than the pKa of the weak acid drug.
  • the present invention also provides a treprostinil liposome based on the liposome pharmaceutical composition, wherein the weakly acidic drug in the liposome pharmaceutical composition is treprostinil or a derivative thereof, the treprostinil or a derivative thereof is encapsulated in the inner aqueous phase of the liposome, and the inner aqueous phase contains meglumine;
  • the derivative of treprostinil is selected from hydrates, solvates or complexes of treprostinil; or is selected from pharmaceutically acceptable salts, esters or prodrugs thereof of treprostinil, such as treprostinil sodium salt, treprostinil potassium salt, treprostinil diethanolamine salt, treprostinil methyl ester, treprostinil ethyl ester, and treprostinil fumaryl diketopiperidine prodrug.
  • treprostinil sodium salt such as treprostinil sodium salt, treprostinil potassium salt, treprostinil diethanolamine salt, treprostinil methyl ester, treprostinil ethyl ester, and treprostinil fumaryl diketopiperidine prodrug.
  • the drug loading of treprostinil or its derivative is not less than 18%.
  • the encapsulation efficiency is not less than 85%, preferably not less than 90%.
  • the present invention also provides the use of the liposome of the treprostinil or its derivatives in the preparation of medicines.
  • the drug is a drug for treating pulmonary hypertension, pulmonary hypertension, pulmonary fibrosis, interstitial lung disease, chronic obstructive pulmonary disease, asthma, ischemic disease, heart failure, arteriosclerosis, postoperative anticoagulation, central retinal vein occlusion, thrombotic microangiopathy, peripheral vascular disease, heart and lung transplantation and the like.
  • the drug is a drug for treating peripheral arterial occlusive disease or pulmonary hypertension.
  • the present invention also provides a method for treating peripheral arterial occlusive disease or pulmonary hypertension using the liposomes of treprostinil or its derivatives, comprising administering a therapeutically effective amount of the liposomes of treprostinil or its derivatives to a patient.
  • the present invention also provides liposomes of treprostinil or its derivatives for treating peripheral arterial occlusive disease or pulmonary hypertension.
  • the present invention also provides a method for preparing a liposome pharmaceutical composition, the method comprising: loading liposomes with at least one weakly acidic drug, preferably, the loading amount of the weakly acidic drug is not less than 18%;
  • the method for preparing the liposome comprises the following steps:
  • Emulsification-extrusion The oil phase and the water phase are mixed to obtain colostrum, and the colostrum is further extruded through a polycarbonate membrane to obtain the liposome.
  • the molar ratio of hydrogenated soy lecithin (HSPC), cholesterol (CHOL) and distearoyl phosphatidylethanolamine-polyethylene glycol 2000 (DSPE-mPEG2000) is 3:2:(0.025-0.225), for example 3:2:(0.025-0.15), preferably 3:2:(0.025-0.075).
  • the method for preparing liposomes further comprises an ultrafiltration step: the liposomes obtained in step (3) are ultrafiltered using a citric acid-sodium citrate buffer to replace the liposome external phase medium and remove the organic solvent to obtain the liposomes.
  • the method for preparing the liposome further comprises dispersing the liposome in the external aqueous phase to obtain a liposome containing the external aqueous phase;
  • the external aqueous phase is a buffer solution with a pH of 4.5 to 6.5;
  • the buffer for achieving the corresponding pH value of the external aqueous phase is well known in the art and is not particularly limited.
  • the commonly used buffer in the art is selected from one or more of HEPES, sodium chloride, sucrose, citrate buffer, phosphate buffer, and Tris buffer, preferably Select a citrate buffer; the concentration of the buffer is 0.05M to 0.25M, for example, 0.08M to 0.2M.
  • the method for preparing the liposome pharmaceutical composition comprises the following steps: dissolving a weakly acidic drug in an external aqueous phase, mixing with the liposome solution, to obtain the liposome pharmaceutical composition; wherein the concentration of the liposome is 1.0 to 20 mg/ml, preferably 1.0 to 5.0 mg/ml (in terms of phospholipid concentration).
  • the method for preparing the liposome pharmaceutical composition comprises the following steps:
  • the drug liposome provided by the present invention has a high drug loading capacity or a high encapsulation rate.
  • the drug liposome has a high drug loading capacity (the maximum drug loading capacity is not less than 18%); when the drug liposome is lower than the maximum drug loading capacity (18%), the encapsulation rate is not less than 85%, preferably not less than 90%.
  • the drug liposome of the present invention also has the effects of high drug loading capacity (for example, not less than 18%) and high encapsulation rate (for example, not less than 85%, preferably not less than 90%).
  • the drug loading (LE) described in the present invention is expressed as a mass percentage.
  • the definitions and explanations of encapsulation efficiency (EE) and drug loading (LE) are as follows:
  • Encapsulation efficiency EE% (1–W f /W t ) ⁇ 100% (Formula 1)
  • Wf represents the amount of free drug in the external medium
  • Wt represents the total amount of drug in the liposome suspension
  • LE represents the percentage of drug loading in the liposome
  • We represents the amount of drug encapsulated in the liposome
  • Wm represents the total weight of the drug-loaded liposome (including the amount of liposome carrier and encapsulated drug).
  • Wt is the total amount of drug in the liposome suspension
  • EE% is the encapsulation rate
  • Wl is the total lipid content of the liposome carrier.
  • treprostinil sodium (Porton); hydrogenated soybean phosphatidylcholine (HSPC) (Lipoid); distearoylphosphatidylethanolamine-polyethylene glycol 2000 (DSPE-mPEG2000) (Lipoid); cholesterol (CHOL) (Nippon Seika); sodium bicarbonate (NaHCO 3 ), sodium acetate (NaAc), meglumine (MEG) (merck); other materials are all commonly used injection-grade excipients.
  • HSPC hydrogenated soybean phosphatidylcholine
  • DSPE-mPEG2000 distearoylphosphatidylethanolamine-polyethylene glycol 2000
  • DSPE-mPEG2000 distearoylphosphatidylethanolamine-polyethylene glycol 2000
  • cholesterol CHOL
  • NaHCO 3 sodium bicarbonate
  • NaAc sodium acetate
  • MEG meglumine
  • the colostrum is extruded through a small
  • the sample is cooled to room temperature, and the liposome external phase medium is replaced with a 0.05M citric acid-sodium citrate buffer at pH 5.5 using a tangential flow TFF filtration device, and the blank liposomes after ultrafiltration are stored at 2-8°C for use.
  • Treprostinil liposomes Dissolve Treprostinil sodium in 0.05 M sodium citrate solution, then add the Treprostinil sodium solution to the blank liposomes according to a certain theoretical dosage (for example, a theoretical drug loading of 22% (w/w)), add an appropriate amount of citric acid-sodium citrate buffer to adjust the concentration of citric acid-sodium citrate in the liposome external phase to 0.09-0.12 M, and incubate at 35-45° C. for 30-60 min to obtain a Treprostinil liposome composition.
  • a certain theoretical dosage for example, a theoretical drug loading of 22% (w/w)
  • Determination of free treprostinil 1 ml of treprostinil liposomes was placed in an ultracentrifuge tube and centrifuged in an ultracentrifuge with the following parameters (temperature: 4°C, speed: 100000 rpm, time: 30 min). After 30 min, the sample was taken out and the supernatant was analyzed by HPLC.
  • Cf represents the amount of free drug in the external medium
  • Ct represents the total amount of drug in the liposome suspension
  • Ac is the peak area of the total drug content
  • Ab is the peak area of the free drug
  • a is the dilution multiple of the total drug content, 20.
  • the concentration of the liposomal drug composition was calculated using the following formula 3:
  • C is the concentration of the liposome composition, mg/ml
  • W std is the sample weight of the reference substance, mg
  • a std is the average peak area of the peak areas of the reference substance solution for five points
  • T is the content of the main drug in the reference substance, %
  • D is the dilution volume of the reference substance solution
  • a sample is the peak area of the liposome drug total content solution
  • a is the dilution volume of the liposome drug total content solution.
  • the cations and anions described in the examples represent the material concentrations that provide the corresponding ions, not the ion concentrations after the materials are ionized.
  • treprostinil liposome compositions comprising different internal aqueous phases (sodium bicarbonate (NaHCO 3 ), sodium acetate (NaAc), meglumine-acetic acid (MEG-HAc), pH about 8.5) and different drug loading amounts were prepared according to the procedure described in “1. Preparation of treprostinil liposome compositions” in the previous paragraph titled “General Experimental Procedures”, which had specific treprostinil liposomes as shown in Table 1.
  • the maximum drug loading of the treprostinil liposome of the meglumine prescription is 19.72%, while other prescriptions (such as sodium bicarbonate or sodium acetate, etc.) cannot exceed 18%; when the feed amount is low, the encapsulation rate of the meglumine prescription can also be not less than 85%, or even not less than 90%.
  • its drug loading is only 8.22%, but the encapsulation rate can reach 97.86%. Even when the drug loading of other formulations was 17.39%, the encapsulation efficiency was only 75.65%, and the encapsulation efficiency in most comparative examples was significantly lower than 60%.
  • C003 compared with Comparative Examples A001 or B001, has the same feed ratio of 21%, but the encapsulation rate of the meglumine formulation is 87.55%, which is higher than 75.65% or 79.59% of sodium bicarbonate or sodium acetate, and the drug loading is 19.72%, which is also significantly higher than 17.39% or 17.84%.
  • the meglumine formulation of the present invention is also better than the sodium bicarbonate or sodium acetate formulation.
  • treprostinil liposome compositions containing different particle sizes were prepared, and the liposome compositions were analyzed according to the above general analytical method.
  • treprostinil liposome compositions containing different concentrations of meglumine-acetic acid were prepared, and the liposome compositions were analyzed according to the above general analytical method.
  • Table 3-1 High theoretical feed ratio - Treprostinil liposome composition with different internal aqueous phase (methylglucamine) concentrations Note: The anion of the drug-carrying salt is HAc; the theoretical feed ratio is 21% (within ⁇ 5%); the particle size range of the liposome is between 100 and 150 nm;
  • Table 3-2 Low theoretical feed ratio - Treprostinil liposome composition with different internal aqueous phase (meglumine) concentrations Note: The anion of the drug-carrying salt is HAc; the theoretical feed ratio is 15% (within ⁇ 5%); the particle size range of the liposome is 100-150nm between;
  • Treprostinil liposome compositions containing different phospholipid formulations were prepared according to the above-mentioned "General Experimental Procedures", and the liposome compositions were analyzed according to the above-mentioned general analytical methods.
  • the drug-carrying salt cation is MEG; the drug-carrying salt anion is HAc; the theoretical feed ratio is 21% (within ⁇ 5%).
  • the particle size range of the above liposomes is between 100 and 150 nm.
  • the amount of acetic acid in the internal aqueous phase was adjusted (0.21-0.40 M) to prepare treprostinil liposome compositions with different internal aqueous phase pH prescriptions.
  • the liposome compositions were analyzed according to the above general analytical method.
  • Table 5 Treprostinil liposome compositions at different internal aqueous phase pH Note:
  • the drug-carrying salt cation is MEG (0.30M); the theoretical feed ratio is 22.3% (within ⁇ 5%); the particle size range is 100-200nm;
  • Treprostinil liposome compositions with different inner aqueous phase anion formulations were prepared according to the above-mentioned "general experimental procedure", and the liposome compositions were analyzed according to the above-mentioned general analytical method.
  • the prepared treprostinil liposomes have a high drug loading (not less than 18%) when the pH of the inner aqueous phase is maintained at 8.5 and acetic acid is used as the inner aqueous phase anion; valeric acid and lactic acid are used as the inner aqueous phase anions, and when the theoretical feed ratio is lower than 18%, the prepared treprostinil liposomes can obtain a high encapsulation rate (encapsulation rate not less than 85%). Gluconic acid, malic acid, citric acid, and phosphoric acid are not suitable as anions for providing gradients in this composition.
  • the drug-carrying salt cation is MEG (0.30M); the particle size range of the above liposomes is between 100 and 150 nm.
  • the liposome compositions prepared according to the above-mentioned "general experimental procedure" were loaded with different drugs, and the liposome compositions were analyzed according to the above-mentioned general analytical method.
  • Liposome drug compositions of different weakly acidic drugs Note: The cation of the drug-carrying salt is MEG (0.30M); the anion of the drug-carrying salt is HAc (0.29M); the theoretical feed ratio is 22% (error ⁇ 5%) The particle size of the liposome is between 100 and 150 nm.
  • the liposome composition prepared according to the above-mentioned "General Experimental Procedure” was loaded with treprostinil sodium, and the liposome composition was analyzed according to the above-mentioned general analytical method.
  • the liposome composition loaded with treprostinil has a high drug loading capacity and a high encapsulation efficiency.
  • the feed ratio was kept consistent and the feed was fed at different phospholipid concentrations to prepare the treprostinil liposome composition.
  • the liposome composition was analyzed according to the above general analysis method.

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Abstract

Provided are a liposome and a composition thereof. When the liposome and the composition thereof serve as a medicament carrier, by means of specific drug-loading conditions and processes, 1) a high drug-loading capacity (with the highest drug-loading capacity not lower than 18%) is obtained; and 2) the encapsulation efficiency is not less than 85% when the drug-loading capacity is lower than the highest drug-loading capacity (18%). A medicament liposome-treprostinil liposome combination prepared from said liposome has a good therapeutic effect on the treatment of peripheral arterial occlusive disease or pulmonary arterial hypertension.

Description

一种脂质体药物组合物及其制备方法和应用A liposome drug composition and its preparation method and application
本发明要求享有：The present invention claims:
于2022年9月30日向中国国家知识产权局提交的，专利申请号为202211216387.5，名称为“一种脂质体、药物组合物及其制备方法和应用”的在先申请的优先权；Priority to the prior application, patent application number 202211216387.5, entitled “A liposome, a pharmaceutical composition, a preparation method and application thereof”, filed with the State Intellectual Property Office of China on September 30, 2022;
于2023年9月22日向中国国家知识产权局提交的，专利申请号为202311236761.2，名称为“一种脂质体药物组合物及其应用”的在先申请的优先权；Priority to the prior application, patent application number 202311236761.2, filed with the State Intellectual Property Office of China on September 22, 2023, entitled “A liposome pharmaceutical composition and its application”;
于2023年9月26日向中国国家知识产权局提交的，专利申请号为202311260280.5，名称为“一种脂质体药物组合物及其制备方法和应用”的在先申请的优先权；Priority to the prior application, patent application number 202311260280.5, filed with the State Intellectual Property Office of China on September 26, 2023, entitled “A liposome pharmaceutical composition, its preparation method and application”;
所述在先申请的全文通过引用的方式结合于本申请中。The entirety of said prior application is incorporated into the present application by reference.
技术领域Technical Field
本发明属于药物领域，具体涉及一种脂质体药物组合物及其制备方法和应用。The invention belongs to the field of medicines, and in particular relates to a liposome medicine composition and a preparation method and application thereof.
背景技术Background technique
脂质体是藉由包含内部水性介质的脂质双层形成的囊泡。脂质体已经用作各种治疗剂的载体，以提供改善的递送特性，例如增强药物在血液内的循环时间、减少细胞毒性、持续释放药物以及将特定药物递送到选定的组织。当利用脂质体进行治疗药物的递送时，需要较高的药物包封率及药含量以降低潜在的磷脂辅料毒性。Liposomes are vesicles formed by a lipid bilayer containing an internal aqueous medium. Liposomes have been used as carriers for various therapeutic agents to provide improved delivery characteristics, such as enhancing drug circulation time in the blood, reducing cytotoxicity, sustained release of drugs, and delivery of specific drugs to selected tissues. When using liposomes for the delivery of therapeutic drugs, high drug encapsulation efficiency and drug content are required to reduce potential phospholipid excipient toxicity.
US8932627B2及US20190022004A1前案技术，使用醋酸盐或碳酸氢盐将药物以分子形态负载于脂质体中，药物以离子形态贮存在内水相中，并通过调控药物的释放速率来治疗呼吸***疾病。然而，使用醋酸盐或碳酸氢盐的处方负载药物获得的药物脂质体载药量有限。有研究提出在脂质体制剂中使用碳酸氢盐会导致气体堆积，其会破坏和使脂质体不稳定，并且引起过早的药物释放，由于其对脂质体的破坏性和不稳定作用，而建议应避免以碳酸氢盐作为装载剂。The previous technologies of US8932627B2 and US20190022004A1 use acetate or bicarbonate to load the drug in liposomes in molecular form. The drug is stored in the inner aqueous phase in ionic form, and respiratory diseases are treated by regulating the release rate of the drug. However, the drug loading capacity of drug liposomes obtained by using acetate or bicarbonate prescription loading drugs is limited. Some studies have suggested that the use of bicarbonate in liposome preparations will lead to gas accumulation, which will destroy and destabilize the liposomes and cause premature drug release. Due to its destructive and destabilizing effects on liposomes, it is recommended to avoid using bicarbonate as a loading agent.
近年来，许多研究者通过远程装载技术(例如，pH梯度法或者硫酸铵梯度法)将弱碱性药物有效装载入脂质体内水相，并提高其药物的包封率。但弱酸性药物脂质体的研究相对较少。弱酸性药物脂质体制剂的开发仍然存在许多未解决的问题，例如弱酸性药物的装载量不高或包封率不佳等，本发明的技术进一步提高了药物装载量，以期在临床应用中能够提高患者顺应性和治疗效果。In recent years, many researchers have effectively loaded weakly basic drugs into the aqueous phase in liposomes by remote loading techniques (e.g., pH gradient method or ammonium sulfate gradient method), and improved the encapsulation efficiency of the drugs. However, the research on weakly acidic drug liposomes is relatively less. There are still many unresolved problems in the development of weakly acidic drug liposome preparations, such as the loading capacity of weakly acidic drugs is not high or the encapsulation efficiency is not good, etc. The technology of the present invention further improves the drug loading capacity, in the hope that patient compliance and therapeutic effect can be improved in clinical applications.
发明内容Summary of the invention
为改善上述技术问题，本发明通过如下技术方案实现：In order to improve the above technical problems, the present invention is implemented through the following technical solutions:
本发明提供一种脂质体药物组合物，包括弱酸性药物和脂质体；所述脂质体为如下中的一种： The present invention provides a liposome pharmaceutical composition, comprising a weakly acidic drug and a liposome; the liposome is one of the following:
(1)所述脂质体包括磷脂和内水相；或(1) The liposome comprises phospholipids and an internal aqueous phase; or
(2)所述脂质体包括磷脂、内水相和外水相；(2) The liposome comprises phospholipids, an inner aqueous phase and an outer aqueous phase;
其中，所述内水相包括葡甲胺与弱酸；Wherein, the internal aqueous phase comprises meglumine and a weak acid;
所述外水相使脂质体悬浮。The external aqueous phase suspends the liposomes.
根据本发明的实施方案，所述弱酸性药物包裹在脂质体的内水相中，包裹了弱酸性药物的脂质体悬浮在外水相中。According to an embodiment of the present invention, the weakly acidic drug is encapsulated in the inner aqueous phase of the liposome, and the liposome encapsulating the weakly acidic drug is suspended in the outer aqueous phase.
根据本发明的实施方案，所述弱酸性药物的pKa在2～7之间。According to an embodiment of the present invention, the pKa of the weakly acidic drug is between 2 and 7.
根据本发明的实施方案，所述弱酸性药物的载药量不低于18％，例如为18％至40％，还可以为19％-30％，或者20-25％，如20.5％，21％，21.5％，22％，22.5％，23％，23.3％、23.5％或35％。According to an embodiment of the present invention, the drug loading of the weakly acidic drug is not less than 18%, for example, 18% to 40%, and can also be 19%-30%, or 20-25%, such as 20.5%, 21%, 21.5%, 22%, 22.5%, 23%, 23.3%, 23.5% or 35%.
根据本发明的实施方案，所述药物脂质体的包封率不低于85％，优选不低于90％。根据本发明的实施方案，当所述弱酸性药物的载药量低于18％时，优选药物脂质体的包封率不低于85％，优选不低于90％。更优选地，所述药物脂质体的载药量不低于18％，且所述药物脂质体的包封率不低于85％，优选不低于90％。According to an embodiment of the present invention, the encapsulation efficiency of the drug liposome is not less than 85%, preferably not less than 90%. According to an embodiment of the present invention, when the drug loading of the weakly acidic drug is less than 18%, preferably the encapsulation efficiency of the drug liposome is not less than 85%, preferably not less than 90%. More preferably, the drug loading of the drug liposome is not less than 18%, and the encapsulation efficiency of the drug liposome is not less than 85%, preferably not less than 90%.
根据本发明的实施方案，所述弱酸性药物可以选自：***素类、解热镇痛类、喹啉羧酸抗菌类、干扰素基因刺激因子(STING)受体中的至少一种药物；According to an embodiment of the present invention, the weakly acidic drug may be selected from: at least one drug selected from prostaglandins, antipyretics, quinoline carboxylic acid antibiotics, and interferon gene stimulator (STING) receptors;
根据本发明的实施方案，所述***素类药物如曲前列尼尔、贝前列素、伊洛前列素、卡前列素、利马前列素、依前列醇、前列地尔、乌诺前列酮，或它们的衍生物(如药学上可以接受的盐、酯或前药)；所述解热镇痛类药物如阿司匹林、布洛芬、萘普生、双氯芬酸钠，或它们的衍生物(如药学上可以接受的盐、酯或前药)；所述喹啉羧酸类抗菌药物如萘啶酸、吡咯嘧酸、氟喹啉酸、西他沙星、环丙沙星、依诺沙星，或它们的衍生物(如药学上可以接受的盐、酯或前药)；所述STING药物如MSA-2、STING agonist-7、Vadimezan，或它们的衍生物(如药学上可以接受的盐、酯或前药)。According to an embodiment of the present invention, the prostaglandin drugs are such as treprostinil, beraprost, iloprost, carboprost, limaprost, epoprostenol, alprostadil, unoprostone, or their derivatives (such as pharmaceutically acceptable salts, esters or prodrugs); the antipyretic and analgesic drugs are such as aspirin, ibuprofen, naproxen, diclofenac sodium, or their derivatives (such as pharmaceutically acceptable salts, esters or prodrugs); the quinoline carboxylic acid antibacterial drugs are such as nalidixic acid, pyrrolopyrimic acid, fluoroquinolinic acid, sitafloxacin, ciprofloxacin, enoxacin, or their derivatives (such as pharmaceutically acceptable salts, esters or prodrugs); the STING drugs are such as MSA-2, STING agonist-7, Vadimezan, or their derivatives (such as pharmaceutically acceptable salts, esters or prodrugs).
根据本发明的实施方案，所述内水相的pH为4.0～10.5，例如4.5～10.0，6.5～10.5，优选4.97～9.52；According to an embodiment of the present invention, the pH of the internal aqueous phase is 4.0 to 10.5, such as 4.5 to 10.0, 6.5 to 10.5, preferably 4.97 to 9.52;
根据本发明的实施方案，所述内水相的pH为4.5、5.0、5.5、6.0、6.5、7.0、7.5、8.0、8.5、9.0、9.5，优选为8.5。According to an embodiment of the present invention, the pH of the internal aqueous phase is 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, preferably 8.5.
根据本发明的实施方案，所述内水相中葡甲胺提供了内水相的阳离子部分，而所述弱酸提供了内水相中的阴离子部分。According to an embodiment of the present invention, the meglumine in the internal aqueous phase provides the cationic part of the internal aqueous phase, and the weak acid provides the anionic part of the internal aqueous phase.
根据本发明的实施方案，所述弱酸选自“羧酸”，例如选自甲酸、乙酸、丙酸、丁酸、异丁酸、戊酸、异戊酸、乳酸或其组合。优选甲酸、乙酸、丙酸或其组合。According to an embodiment of the present invention, the weak acid is selected from "carboxylic acids", for example, formic acid, acetic acid, propionic acid, butyric acid, isobutyric acid, valeric acid, isovaleric acid, lactic acid or a combination thereof. Preferably, formic acid, acetic acid, propionic acid or a combination thereof.
与现有技术公开的内容不同，本发明的实施方案中所述内水相不含碳酸盐或碳酸氢盐，而是通过葡甲胺和弱酸共同实现内水相体系。Different from the contents disclosed in the prior art, the internal aqueous phase in the embodiments of the present invention does not contain carbonate or bicarbonate, but the internal aqueous phase system is realized by meglumine and weak acid.
根据本发明的实施方案，所述葡甲胺在内水相中的浓度为0.1～0.8M，例如0.2～0.6M，优选0.3～0.4M，如0.3M、0.35M、0.4M；According to an embodiment of the present invention, the concentration of meglumine in the internal aqueous phase is 0.1 to 0.8 M, such as 0.2 to 0.6 M, preferably 0.3 to 0.4 M, such as 0.3 M, 0.35 M, 0.4 M;
根据本发明的实施方案，所述弱酸在内水相中的浓度为0.1～0.8M，例如0.2～0.6M，如0.2M、0.211M、0.268M、0.286M、0.3M、0.35M、0.40M、0.5M、0.6M； According to an embodiment of the present invention, the concentration of the weak acid in the inner aqueous phase is 0.1 to 0.8 M, for example 0.2 to 0.6 M, such as 0.2 M, 0.211 M, 0.268 M, 0.286 M, 0.3 M, 0.35 M, 0.40 M, 0.5 M, 0.6 M;
根据本发明的实施方案，所述葡甲胺与弱酸为葡甲胺与乙酸；优选地，所述葡甲胺与乙酸在内水相中的浓度为0.30～0.40M。According to an embodiment of the present invention, the meglumine and the weak acid are meglumine and acetic acid; preferably, the concentration of the meglumine and acetic acid in the internal aqueous phase is 0.30-0.40M.
根据本发明的实施方案，所述磷脂具有双层脂质结构，所述双层脂质由氢化大豆卵磷脂(HSPC)、胆固醇(CHOL)和二硬脂酰基磷脂酰乙醇胺-聚乙二醇2000(DSPE-mPEG2000)构成，摩尔比为3:(1-3):(0.025-0.225)，例如3:(2-3):(0.025-0.075)、3:(1.5-2.5):(0.025-0.075)。According to an embodiment of the present invention, the phospholipid has a bilayer lipid structure, and the bilayer lipid is composed of hydrogenated soybean lecithin (HSPC), cholesterol (CHOL) and distearoyl phosphatidylethanolamine-polyethylene glycol 2000 (DSPE-mPEG2000), and the molar ratio is 3:(1-3):(0.025-0.225), for example, 3:(2-3):(0.025-0.075), 3:(1.5-2.5):(0.025-0.075).
根据本发明的实施方案，所述双层脂质由氢化大豆卵磷脂(HSPC)、胆固醇(CHOL)和二硬脂酰基磷脂酰乙醇胺-聚乙二醇2000(DSPE-mPEG2000)构成，摩尔比为3:2:(0.025-0.225)，例如3:2:(0.025-0.15)，优选3:2:(0.025-0.075)。According to an embodiment of the present invention, the bilayer lipid is composed of hydrogenated soybean lecithin (HSPC), cholesterol (CHOL) and distearoyl phosphatidylethanolamine-polyethylene glycol 2000 (DSPE-mPEG2000), and the molar ratio is 3:2:(0.025-0.225), for example 3:2:(0.025-0.15), preferably 3:2:(0.025-0.075).
根据本发明的实施方案，所述磷脂在脂质体组合物中的浓度为1～100mg/mL，例如1～50mg/mL、1～20mg/mL，而在本发明的脂质体药物组合物的制备过程中，磷脂的使用浓度还例如1～15mg/mL，1.2～5.0mg/mL，优选为1.3～4.5mg/mL、1.4～4.18mg/mL。According to an embodiment of the present invention, the concentration of the phospholipid in the liposome composition is 1 to 100 mg/mL, for example, 1 to 50 mg/mL, 1 to 20 mg/mL, and in the preparation process of the liposome pharmaceutical composition of the present invention, the concentration of the phospholipid used is also, for example, 1 to 15 mg/mL, 1.2 to 5.0 mg/mL, preferably 1.3 to 4.5 mg/mL, 1.4 to 4.18 mg/mL.
根据本发明的实施方案，所述脂质体的粒径为50～500nm，例如100～200nm，优选为100～150nm、120～180nm、130～160nm、140～190nm。According to an embodiment of the present invention, the particle size of the liposome is 50-500 nm, for example, 100-200 nm, preferably 100-150 nm, 120-180 nm, 130-160 nm, 140-190 nm.
根据本发明的实施方案，所述外水相为pH为4.5～6.5的缓冲液。According to an embodiment of the present invention, the external aqueous phase is a buffer solution with a pH of 4.5 to 6.5.
根据本发明的实施方案，实现外水相相应pH值的缓冲液为本领域公知的，并无特别限定。本领域常用的缓冲液选自HEPES、氯化钠、蔗糖、枸橼酸盐缓冲液、磷酸盐缓冲液、Tris缓冲液中的一种或几种，根据本发明的优选实施方案，所述缓冲液可以选自枸橼酸盐缓冲液；所述缓冲液的浓度为0.05M～0.25M，例如为0.08M～0.2M，优选为0.09～0.12M。According to an embodiment of the present invention, the buffer for achieving the corresponding pH value of the external aqueous phase is well known in the art and is not particularly limited. The commonly used buffer in the art is selected from one or more of HEPES, sodium chloride, sucrose, citrate buffer, phosphate buffer, and Tris buffer. According to a preferred embodiment of the present invention, the buffer can be selected from citrate buffer; the concentration of the buffer is 0.05M to 0.25M, for example, 0.08M to 0.2M, preferably 0.09 to 0.12M.
根据本发明的实施方案，所述脂质体具有内水相pH高，外水相pH低的pH梯度特性，其中外水相的pH值高于弱酸药物的pKa。According to an embodiment of the present invention, the liposome has a pH gradient property with a high pH in the inner aqueous phase and a low pH in the outer aqueous phase, wherein the pH value of the outer aqueous phase is higher than the pKa of the weak acid drug.
本发明还提供一种基于所述脂质体药物组合物的曲前列尼尔脂质体，其中，所述脂质体药物组合物中的弱酸性药物为曲前列尼尔或其衍生物，所述曲前列尼尔或其衍生物包裹于所述脂质体的内水相，所述内水相中包含葡甲胺；The present invention also provides a treprostinil liposome based on the liposome pharmaceutical composition, wherein the weakly acidic drug in the liposome pharmaceutical composition is treprostinil or a derivative thereof, the treprostinil or a derivative thereof is encapsulated in the inner aqueous phase of the liposome, and the inner aqueous phase contains meglumine;
根据本发明的实施方案，所述曲前列尼尔的衍生物选自曲前列尼尔的水合物、溶剂合物或络合物；或者选自曲前列尼尔的药学上可接受的盐、酯或其前药等，如曲前列尼尔钠盐、曲前列尼尔钾盐、曲前列尼尔二乙醇胺盐、曲前列尼尔甲酯、曲前列尼尔乙酯、曲前列尼尔的富马酰基二酮哌啶前药。According to an embodiment of the present invention, the derivative of treprostinil is selected from hydrates, solvates or complexes of treprostinil; or is selected from pharmaceutically acceptable salts, esters or prodrugs thereof of treprostinil, such as treprostinil sodium salt, treprostinil potassium salt, treprostinil diethanolamine salt, treprostinil methyl ester, treprostinil ethyl ester, and treprostinil fumaryl diketopiperidine prodrug.
根据本发明的实施方案，所述曲前列尼尔或其衍生物的载药量不低于18％，当低于最高载药量(18％)时，包封率不低于85％，优选不低于90％。According to an embodiment of the present invention, the drug loading of treprostinil or its derivative is not less than 18%. When it is less than the maximum drug loading (18%), the encapsulation efficiency is not less than 85%, preferably not less than 90%.
所述的曲前列尼尔分子结构如下：
The molecular structure of treprostinil is as follows:
本发明还提供所述曲前列尼尔或其衍生物的脂质体在制备药物中的应用。The present invention also provides the use of the liposome of the treprostinil or its derivatives in the preparation of medicines.
根据本发明的实施方案，所述药物为治疗肺动脉高压、肺高血压、肺纤维化、间质性肺疾病、慢性阻塞性肺病、哮喘、缺血性疾病、心脏衰竭、动脉硬化、术后抗凝血、视网膜中央静脉阻塞、血栓性微血管病、周围性血管疾病、心肺移植等疾病的药物。According to an embodiment of the present invention, the drug is a drug for treating pulmonary hypertension, pulmonary hypertension, pulmonary fibrosis, interstitial lung disease, chronic obstructive pulmonary disease, asthma, ischemic disease, heart failure, arteriosclerosis, postoperative anticoagulation, central retinal vein occlusion, thrombotic microangiopathy, peripheral vascular disease, heart and lung transplantation and the like.
根据本发明的实施方案，所述药物为治疗外周动脉闭塞性疾病或肺动脉高压药物。According to an embodiment of the present invention, the drug is a drug for treating peripheral arterial occlusive disease or pulmonary hypertension.
本发明还提供所述曲前列尼尔或其衍生物的脂质体在治疗外周动脉闭塞性疾病或肺动脉高压的方法，包括给与患者治疗有效量的曲前列尼尔或其衍生物的脂质体。本发明还提供用于治疗外周动脉闭塞性疾病或肺动脉高压的曲前列尼尔或其衍生物的脂质体。The present invention also provides a method for treating peripheral arterial occlusive disease or pulmonary hypertension using the liposomes of treprostinil or its derivatives, comprising administering a therapeutically effective amount of the liposomes of treprostinil or its derivatives to a patient. The present invention also provides liposomes of treprostinil or its derivatives for treating peripheral arterial occlusive disease or pulmonary hypertension.
本发明还提供了一种脂质体药物组合物的制备方法，该方法包括，将脂质体负载至少一种弱酸性药物，优选地，所述弱酸性药物的载药量不低于18％；The present invention also provides a method for preparing a liposome pharmaceutical composition, the method comprising: loading liposomes with at least one weakly acidic drug, preferably, the loading amount of the weakly acidic drug is not less than 18%;
根据本发明的实施方案，所述脂质体的制备方法，包括以下步骤：According to an embodiment of the present invention, the method for preparing the liposome comprises the following steps:
(1)油相制备：按重量份，称取氢化大豆卵磷脂、二硬脂酰基磷脂酰乙醇胺-聚乙二醇2000和胆固醇溶于乙醇得到油相；(1) Preparation of oil phase: Weigh hydrogenated soybean lecithin, distearoyl phosphatidylethanolamine-polyethylene glycol 2000 and cholesterol in parts by weight and dissolve them in ethanol to obtain an oil phase;
(2)水相制备：将葡甲胺溶于水后，加入乙酸调节pH，得到水相，即内水相；(2) Preparation of aqueous phase: After dissolving meglumine in water, acetic acid is added to adjust the pH to obtain an aqueous phase, i.e., an inner aqueous phase;
(3)乳化-挤出：将所述油相和水相混合后得到初乳，进一步将初乳经过聚碳酸酯膜挤出，得到所述脂质体。(3) Emulsification-extrusion: The oil phase and the water phase are mixed to obtain colostrum, and the colostrum is further extruded through a polycarbonate membrane to obtain the liposome.
根据本发明的实施方案，所述氢化大豆卵磷脂(HSPC)、胆固醇(CHOL)和二硬脂酰基磷脂酰乙醇胺-聚乙二醇2000(DSPE-mPEG2000)的摩尔比为3:2:(0.025-0.225)，例如3:2:(0.025-0.15)，优选3:2:(0.025-0.075)。According to an embodiment of the present invention, the molar ratio of hydrogenated soy lecithin (HSPC), cholesterol (CHOL) and distearoyl phosphatidylethanolamine-polyethylene glycol 2000 (DSPE-mPEG2000) is 3:2:(0.025-0.225), for example 3:2:(0.025-0.15), preferably 3:2:(0.025-0.075).
根据本发明的实施方案，所述脂质体的制备方法还包括超滤的步骤：将步骤(3)得到的脂质体用枸橼酸-枸橼酸钠缓冲液进行超滤，置换脂质体外相介质并去除有机溶剂，得到所述脂质体。According to an embodiment of the present invention, the method for preparing liposomes further comprises an ultrafiltration step: the liposomes obtained in step (3) are ultrafiltered using a citric acid-sodium citrate buffer to replace the liposome external phase medium and remove the organic solvent to obtain the liposomes.
根据本发明的实施方案，当所述脂质体包括外水相时，所述脂质体的制备方法还包括将所述脂质体分散于外水相，得到含外水相的脂质体；According to an embodiment of the present invention, when the liposome includes an external aqueous phase, the method for preparing the liposome further comprises dispersing the liposome in the external aqueous phase to obtain a liposome containing the external aqueous phase;
根据本发明的实施方案，所述外水相为pH为4.5～6.5的缓冲液；According to an embodiment of the present invention, the external aqueous phase is a buffer solution with a pH of 4.5 to 6.5;
根据本发明的实施方案，实现外水相相应pH值的缓冲液为本领域公知的，并无特别限定。本领域常用的缓冲液选自HEPES、氯化钠、蔗糖、枸橼酸盐缓冲液、磷酸盐缓冲液、Tris缓冲液中的一种或几种，优 选枸橼酸盐缓冲液；所述缓冲液的浓度为0.05M～0.25M，例如为0.08M～0.2M。According to the embodiment of the present invention, the buffer for achieving the corresponding pH value of the external aqueous phase is well known in the art and is not particularly limited. The commonly used buffer in the art is selected from one or more of HEPES, sodium chloride, sucrose, citrate buffer, phosphate buffer, and Tris buffer, preferably Select a citrate buffer; the concentration of the buffer is 0.05M to 0.25M, for example, 0.08M to 0.2M.
根据本发明的实施方案，所述脂质体药物组合物的制备方法，包括以下步骤：将弱酸性药物溶解在外水相中，与所述脂质体的溶液混合，得到所述脂质体药物组合物；其中所述脂质体的浓度为1.0～20mg/ml，优选1.0～5.0mg/ml(以磷脂浓度计)。According to an embodiment of the present invention, the method for preparing the liposome pharmaceutical composition comprises the following steps: dissolving a weakly acidic drug in an external aqueous phase, mixing with the liposome solution, to obtain the liposome pharmaceutical composition; wherein the concentration of the liposome is 1.0 to 20 mg/ml, preferably 1.0 to 5.0 mg/ml (in terms of phospholipid concentration).
根据本发明的实施方案，所述脂质体药物组合物的制备方法，包括以下步骤：According to an embodiment of the present invention, the method for preparing the liposome pharmaceutical composition comprises the following steps:
(1)将所述弱酸性药物溶于枸橼酸钠水溶液中，得到药物溶液；(1) dissolving the weakly acidic drug in a sodium citrate aqueous solution to obtain a drug solution;
(2)将所述脂质体的溶液与枸橼酸盐缓冲液混合，得到载体溶液；(2) mixing the liposome solution with a citrate buffer to obtain a carrier solution;
(3)将所述药物溶液加入到所述载体溶液中进行负载，得到所述脂质体药物组合物。(3) adding the drug solution into the carrier solution for loading to obtain the liposome drug composition.
有益效果Beneficial Effects
本发明提供的药物脂质体的载药量高或者包封率高。其中，该药物脂质体的载药量高(最高载药量不低于18％)；所述药物脂质体当低于最高载药量(18％)时，包封率不低于85％，优选不低于90％。优选地，本发明的药物脂质体还同时具有载药量高(例如不低于18％)和包封率高(例如不低于85％，优选不低于90％)的效果。The drug liposome provided by the present invention has a high drug loading capacity or a high encapsulation rate. The drug liposome has a high drug loading capacity (the maximum drug loading capacity is not less than 18%); when the drug liposome is lower than the maximum drug loading capacity (18%), the encapsulation rate is not less than 85%, preferably not less than 90%. Preferably, the drug liposome of the present invention also has the effects of high drug loading capacity (for example, not less than 18%) and high encapsulation rate (for example, not less than 85%, preferably not less than 90%).
具体实施方式Detailed ways
下文将结合具体实施例对本发明的技术方案做更进一步的详细说明。应当理解，下列实施例仅为示例性地说明和解释本发明，而不应被解释为对本发明保护范围的限制。凡基于本发明上述内容所实现的技术均涵盖在本发明旨在保护的范围内。The technical scheme of the present invention will be further described in detail below in conjunction with specific embodiments. It should be understood that the following embodiments are only exemplary descriptions and explanations of the present invention, and should not be construed as limiting the scope of protection of the present invention. All technologies implemented based on the above content of the present invention are included in the scope that the present invention is intended to protect.
除非另有说明，以下实施例中使用的原料和试剂均为市售商品，或者可以通过已知方法制备。Unless otherwise specified, the raw materials and reagents used in the following examples are commercially available or can be prepared by known methods.
本发明所述的载药量(LE)用质量百分数来表示。包封率(EE)和载药量(LE)的定义及解释如下：The drug loading (LE) described in the present invention is expressed as a mass percentage. The definitions and explanations of encapsulation efficiency (EE) and drug loading (LE) are as follows:
包封率
EE％＝(1–Wf/Wt)×100％    (公式1)Encapsulation efficiency
EE%＝(1–W f /W t )×100% (Formula 1)
式中Wf表示在外部介质中的游离药物的量；Wt表示脂质体悬液中药物的总量。Where Wf represents the amount of free drug in the external medium; Wt represents the total amount of drug in the liposome suspension.
使用以下公式2计算脂质体药物组合物的载药量：
LE％＝[We/Wm]×100％＝[(Wt×EE％)/(Wl+Wt×EE％)]×100％     (公式2)The drug loading of the liposome drug composition was calculated using the following formula 2:
LE% = [ We / Wm ] × 100% = [( Wt × EE%) / ( Wl + Wt × EE%)] × 100% (Formula 2)
式中LE表示脂质体中药物的载药量百分数；We表示包封于脂质体内的药物量；Wm表示载药脂质体的总重量(包含脂质体载体和包载药物的量)。其中，Wt为脂质体悬液中总药物量；EE％为包封率；Wl为脂质体载体的总脂质含量。Where LE represents the percentage of drug loading in the liposome; We represents the amount of drug encapsulated in the liposome; Wm represents the total weight of the drug-loaded liposome (including the amount of liposome carrier and encapsulated drug). Where Wt is the total amount of drug in the liposome suspension; EE% is the encapsulation rate; Wl is the total lipid content of the liposome carrier.
实验设备：pH计(梅特勒，S220K)；集热式恒温加热磁力搅拌器(上海豫康科教仪器设备有限公司，DF-101S)；小试脂质体挤出器(ATS EX200，氮气源动力)；超速离心机(Backman，MAX-XP)； HPLC(Agilent1260，USA)；纳米激光粒度仪(malvern，ZS90)；切向流超滤设备(Millipore Pellicon)；Experimental equipment: pH meter (Mettler, S220K); heat-collecting constant temperature heating magnetic stirrer (Shanghai Yukang Science and Education Instrument Equipment Co., Ltd., DF-101S); small-scale liposome extruder (ATS EX200, nitrogen source power); ultracentrifuge (Backman, MAX-XP); HPLC (Agilent 1260, USA); nanometer laser particle size analyzer (malvern, ZS90); tangential flow ultrafiltration equipment (Millipore Pellicon);
实验物料：曲前列尼尔钠(Porton)；氢化大豆磷脂酰胆碱(HSPC)(Lipoid)；二硬脂酰基磷脂酰乙醇胺-聚乙二醇2000(DSPE-mPEG2000)(Lipoid)；胆固醇(CHOL)(日本精化)；碳酸氢钠(NaHCO3)，醋酸钠(NaAc)，葡甲胺(MEG)(merck)；其他物料均为常用注射级辅料。Experimental materials: treprostinil sodium (Porton); hydrogenated soybean phosphatidylcholine (HSPC) (Lipoid); distearoylphosphatidylethanolamine-polyethylene glycol 2000 (DSPE-mPEG2000) (Lipoid); cholesterol (CHOL) (Nippon Seika); sodium bicarbonate (NaHCO 3 ), sodium acetate (NaAc), meglumine (MEG) (merck); other materials are all commonly used injection-grade excipients.
一般实验程序General Experimental Procedure
1.曲前列尼尔脂质体组合物的制备1. Preparation of Treprostinil Liposome Composition
实施例1：曲前列尼尔脂质体的制备Example 1: Preparation of Treprostinil Liposomes
(1)空白脂质体的制备：首先，称取HSPC 2.367g，CHOL 0.78g和DSPE-mPEG2000 0.21g(HSPC/CHOL/DSPE-mPEG2000＝3/2/0.075(摩尔比))，加入到洁净干燥的容器中，加入11.3g无水乙醇，密封，于60～65℃下溶解，得到脂质溶液。然后，将上述脂质溶液注入至内水相(如葡甲胺溶液或碳酸氢钠溶液，以0.3M，pH＝8.5的碳酸氢钠溶液作为内水相为例，为注入至86.7g碳酸氢钠溶液)，在60～65℃下搅拌水合20min；乳化结束后，于60～65℃下，将初乳经过小试脂质体挤出器进行挤出，0.6～1.0MPa压力下，经200nm和/或100nm的聚碳酸酯膜多次挤出，使样品粒径为100～200nm，PDI小于0.2。然后，将样品冷却至常温，使用切向流TFF过滤设备将脂质体外相介质置换成pH5.5的0.05M枸橼酸-枸橼酸钠缓冲液，超滤后的空白脂质体于2～8℃保存备用。(1) Preparation of blank liposomes: First, weigh 2.367 g of HSPC, 0.78 g of CHOL, and 0.21 g of DSPE-mPEG2000 (HSPC/CHOL/DSPE-mPEG2000=3/2/0.075 (molar ratio)), add them into a clean and dry container, add 11.3 g of anhydrous ethanol, seal the container, and dissolve at 60-65°C to obtain a lipid solution. Then, the lipid solution is injected into the internal aqueous phase (such as meglumine solution or sodium bicarbonate solution, taking 0.3M sodium bicarbonate solution with pH=8.5 as the internal aqueous phase as an example, 86.7g sodium bicarbonate solution is injected), and stirred and hydrated at 60-65°C for 20min; after the emulsification is completed, the colostrum is extruded through a small-scale liposome extruder at 60-65°C, and extruded through a 200nm and/or 100nm polycarbonate membrane multiple times at a pressure of 0.6-1.0MPa, so that the sample particle size is 100-200nm, and the PDI is less than 0.2. Then, the sample is cooled to room temperature, and the liposome external phase medium is replaced with a 0.05M citric acid-sodium citrate buffer at pH 5.5 using a tangential flow TFF filtration device, and the blank liposomes after ultrafiltration are stored at 2-8°C for use.
(2)曲前列尼尔脂质体的制备：将曲前列尼尔钠溶解在0.05M枸橼酸钠溶液中，然后按照一定的理论投药量(例如，理论载药量为22％(w/w))，将曲前列尼尔钠溶液投入到上述空白脂质体中，加入适量枸橼酸-枸橼酸钠缓冲液使脂质体外相中枸橼酸-枸橼酸钠浓度调整至0.09～0.12M，并在35～45℃下孵育30～60min，得到曲前列尼尔脂质体组合物。(2) Preparation of Treprostinil liposomes: Dissolve Treprostinil sodium in 0.05 M sodium citrate solution, then add the Treprostinil sodium solution to the blank liposomes according to a certain theoretical dosage (for example, a theoretical drug loading of 22% (w/w)), add an appropriate amount of citric acid-sodium citrate buffer to adjust the concentration of citric acid-sodium citrate in the liposome external phase to 0.09-0.12 M, and incubate at 35-45° C. for 30-60 min to obtain a Treprostinil liposome composition.
一般分析方法General analytical methods
1.曲前列尼尔脂质体组合物的定量表征1. Quantitative Characterization of Treprostinil Liposome Composition
a.对照品溶液配制：a. Preparation of reference solution:
精密称取约10mg曲前列尼尔钠原料药置于50ml量瓶中，加2.5ml纯化水溶解，并用甲醇稀释至刻度，摇匀，作为曲前列尼尔纳对照品溶液。Accurately weigh about 10 mg of treprostinil sodium raw material and place it in a 50 ml volumetric flask, add 2.5 ml of purified water to dissolve it, and dilute it to the scale with methanol. Shake well and use it as the treprostinil sodium reference solution.
b.游离和总曲前列尼尔浓度：b. Free and total treprostinil concentrations:
精密量取脂质体药物组合物溶液1ml置20ml量瓶中，加入甲醇约3ml轻轻摇晃，除去产生的气泡后，用甲醇稀释至刻度，摇匀，即得。取样进HPLC分析，测定组合物中曲前列尼尔总含量。Accurately measure 1 ml of the liposome drug composition solution and place it in a 20 ml volumetric bottle, add about 3 ml of methanol and shake gently, remove the bubbles, dilute to the mark with methanol, shake well, and obtain the product. Take a sample and conduct HPLC analysis to determine the total content of treprostinil in the composition.
游离曲前列尼尔测定：取曲前列尼尔脂质体1ml至超速离心管中，在参数为(温度：4℃，转速：100000rpm，时间：30min)超速离心机中离心，30min后取出样品，取上清液进HPLC分析；Determination of free treprostinil: 1 ml of treprostinil liposomes was placed in an ultracentrifuge tube and centrifuged in an ultracentrifuge with the following parameters (temperature: 4°C, speed: 100000 rpm, time: 30 min). After 30 min, the sample was taken out and the supernatant was analyzed by HPLC.
HPLC具体色谱条件如下：

The specific HPLC chromatographic conditions are as follows:

c.包封率(EE)和载药量(LE)：c. Encapsulation efficiency (EE) and drug loading (LE):
本发明所述的包封率(EE)和载药量(LE)的定义及解释已在上文描述，结合具体的分析方法，公式补充如下：The definitions and explanations of the encapsulation efficiency (EE) and drug loading (LE) described in the present invention have been described above. Combined with the specific analysis method, the formula is supplemented as follows:
包封率
Encapsulation efficiency
式中Cf表示外部介质中的游离药物的量；Ct表示脂质体悬液中药物的总量；Ac为药物总含量的峰面积；Ab为游离药物的峰面积；a为总药物含量的稀释倍数，20。Wherein Cf represents the amount of free drug in the external medium; Ct represents the total amount of drug in the liposome suspension; Ac is the peak area of the total drug content; Ab is the peak area of the free drug; and a is the dilution multiple of the total drug content, 20.
使用以下公式3计算脂质体药物组合物的浓度：
The concentration of the liposomal drug composition was calculated using the following formula 3:
C为脂质体组合物的浓度，mg/ml；Wstd为对照品的称样量，mg；Astd五针对照品溶液的峰面积平均峰面积；T为对照品中主药的含量，％；D为对照品溶液的稀释体积；Asample为脂质体药物总含量溶液的峰面积；a为脂质体药物总含量溶液的稀释体积。C is the concentration of the liposome composition, mg/ml; W std is the sample weight of the reference substance, mg; A std is the average peak area of the peak areas of the reference substance solution for five points; T is the content of the main drug in the reference substance, %; D is the dilution volume of the reference substance solution; A sample is the peak area of the liposome drug total content solution; a is the dilution volume of the liposome drug total content solution.
d.平均粒径和多分散性指数(PdI)：d. Average particle size and polydispersity index (PdI):
取30μl曲前列尼尔脂质体组合物，用纯化水稀释至2ml，混匀，通过动态光散射原理，利用马尔文纳米激光粒度仪，进样检测(参数：温度25℃，折射系数1.33，平衡时间120s，循环3次)组合物的粒径分布和多分散系数。Take 30 μl of the treprostinil liposome composition, dilute it to 2 ml with purified water, mix it, and use the dynamic light scattering principle to detect the particle size distribution and polydispersity coefficient of the composition using a Malvern nano-laser particle size analyzer (parameters: temperature 25°C, refractive index 1.33, equilibrium time 120s, and 3 cycles).
实施例中所述阳离子和阴离子表示提供相应离子的物料浓度并非物料电离后的离子浓度。The cations and anions described in the examples represent the material concentrations that provide the corresponding ions, not the ion concentrations after the materials are ionized.
实施例1.不同载药盐种类的不同投料比的曲前列尼尔脂质体组合物Example 1. Treprostinil liposome compositions with different drug-carrying salt types and different feed ratios
为了研究不同载药盐种类对曲前列尼尔脂质体载药的影响，根据如前段标题“一般实验程序”的“1.曲前列尼尔脂质体组合物的制备”中所述的程序制备了包含不同内水相(碳酸氢钠(NaHCO3)、醋酸钠(NaAc)、葡甲胺-乙酸(MEG-HAc)，pH约8.5)和不同载药量的曲前列尼尔脂质体组合物，其具有如表1中所示的特定曲前列尼尔脂质体。 In order to study the effect of different types of drug-loading salts on the drug loading of treprostinil liposomes, treprostinil liposome compositions comprising different internal aqueous phases (sodium bicarbonate (NaHCO 3 ), sodium acetate (NaAc), meglumine-acetic acid (MEG-HAc), pH about 8.5) and different drug loading amounts were prepared according to the procedure described in “1. Preparation of treprostinil liposome compositions” in the previous paragraph titled “General Experimental Procedures”, which had specific treprostinil liposomes as shown in Table 1.
曲前列尼尔脂质体组合物的定量表征，包括曲前列尼尔脂质体的理论投料比(即理论载药量)、包封率和载药量，其根据如前段标题为“一般分析方法”的“1.曲前列尼尔脂质体组合物的定量表征”所述的程序进行。The quantitative characterization of the treprostinil liposome composition, including the theoretical feed ratio (i.e., theoretical drug loading), encapsulation efficiency and drug loading of the treprostinil liposome, was performed according to the procedure described in "1. Quantitative Characterization of Treprostinil Liposome Composition" in the previous paragraph entitled "General Analytical Methods".
由下表的结果可见：由于脂质体内水相容积有限，随着投料量的增加，药物的载药量呈现先增大(从8.22％增加至19.72％)，后基本维持不变的趋势，故投料量为21％时，包封率为87.55％，载药量为19.72％，但后续投料量再增加，而载药量基本保持不变，故会导致包封率显著下降。其二，高投料量时，葡甲胺处方的曲前列尼尔脂质体的最大载药量为19.72％，而其他处方(如碳酸氢钠或者醋酸钠等)均无法突破18％；低投料量时，葡甲胺处方的包封率也能够不低于85％，甚至不低于90％，以实施例C001为例，其载药量仅为8.22％，但包封率可以达到97.86％。而其他处方即便在载药量为17.39％时，包封率也仅为75.65％，且大部分比较例中的包封率都显著低于60％。As can be seen from the results in the following table: Due to the limited volume of the aqueous phase in the liposome, as the feed amount increases, the drug loading of the drug first increases (from 8.22% to 19.72%), and then basically remains unchanged. Therefore, when the feed amount is 21%, the encapsulation rate is 87.55% and the drug loading is 19.72%, but the subsequent feed amount increases again, and the drug loading remains basically unchanged, so the encapsulation rate will be significantly reduced. Second, when the feed amount is high, the maximum drug loading of the treprostinil liposome of the meglumine prescription is 19.72%, while other prescriptions (such as sodium bicarbonate or sodium acetate, etc.) cannot exceed 18%; when the feed amount is low, the encapsulation rate of the meglumine prescription can also be not less than 85%, or even not less than 90%. Taking Example C001 as an example, its drug loading is only 8.22%, but the encapsulation rate can reach 97.86%. Even when the drug loading of other formulations was 17.39%, the encapsulation efficiency was only 75.65%, and the encapsulation efficiency in most comparative examples was significantly lower than 60%.
从下表还可以看出，C003相较于比较例A001或者B001，均为相同的投料比21％，但葡甲胺处方的包封率87.55％，高于碳酸氢钠或者醋酸钠的75.65％或79.59％，而载药量更是19.72％，也是明显高于17.39％或者17.84％。同样的，其他相同投料比的条件，也均是本发明的葡甲胺处方优于碳酸氢钠或者醋酸钠处方。It can also be seen from the table below that C003, compared with Comparative Examples A001 or B001, has the same feed ratio of 21%, but the encapsulation rate of the meglumine formulation is 87.55%, which is higher than 75.65% or 79.59% of sodium bicarbonate or sodium acetate, and the drug loading is 19.72%, which is also significantly higher than 17.39% or 17.84%. Similarly, under other conditions with the same feed ratio, the meglumine formulation of the present invention is also better than the sodium bicarbonate or sodium acetate formulation.
表1不同载药盐制备的曲前列尼尔脂质体组合物

备注：*理论投料比表示理论载药量(误差±5％以内)。脂质体的粒径范围是100～150nm之间；Table 1 Treprostinil liposome compositions prepared with different drug-loaded salts

Note: *Theoretical feed ratio indicates the theoretical drug loading (within ±5%). The particle size range of liposomes is between 100 and 150 nm;
实施例2.不同粒径的葡甲胺-乙酸载药盐的曲前列尼尔脂质体组合物Example 2. Treprostinil liposome compositions containing meglumine-acetic acid drug-loaded salts of different particle sizes
按照上述“一般实验程序”制备包含不同粒径(100～150nm，150～200nm)的曲前列尼尔脂质体组合物。并按照上述的一般分析方法对脂质体组合物进行分析。According to the above "general experimental procedure", treprostinil liposome compositions containing different particle sizes (100-150 nm, 150-200 nm) were prepared, and the liposome compositions were analyzed according to the above general analytical method.
结果：具有不同粒径的曲前列尼尔脂质体组合物的包封率和载药量见下表2。从表中可以看出，在140～190nm的粒径范围内，曲前列尼尔脂质体的包封率和载药量相当。 Results: The encapsulation efficiency and drug loading of the treprostinil liposome compositions with different particle sizes are shown in Table 2. As can be seen from the table, within the particle size range of 140 to 190 nm, the encapsulation efficiency and drug loading of the treprostinil liposomes are comparable.
表2不同粒径的曲前列尼尔组合物

备注：载药盐阳离子为MEG(0.30M)；载药盐阴离子为HAc(0.29M)；理论投料比为21％(误差±5％
以内)；Table 2 Treprostinil compositions of different particle sizes

Note: The cation of the drug-carrying salt is MEG (0.30M); the anion of the drug-carrying salt is HAc (0.29M); the theoretical feed ratio is 21% (error ± 5%)
within );
实施例3不同内水相浓度的曲前列尼尔脂质体组合物Example 3 Treprostinil liposome compositions with different internal aqueous phase concentrations
按照上述“一般实验程序”制备包含不同浓度的葡甲胺-乙酸(葡甲胺浓度为0.20～0.60M，pH为8.5)的曲前列尼尔脂质体组合物。并按照上述的一般分析方法对脂质体组合物进行分析。According to the above "general experimental procedure", treprostinil liposome compositions containing different concentrations of meglumine-acetic acid (meglumine concentration is 0.20-0.60 M, pH is 8.5) were prepared, and the liposome compositions were analyzed according to the above general analytical method.
结果：从表中可以看出，当理论投料比较高为21％时，水相浓度为0.30～0.40M时制备的曲前列尼尔脂质体具有高载药量(不低于18％)；当理论投料比低于18％时，水相浓度在0.30～0.40M之间制得的曲前列尼尔脂质体可获得高包封率(包封率不低于90％)。Results: It can be seen from the table that when the theoretical feed ratio is as high as 21%, the Treprostinil liposomes prepared when the aqueous phase concentration is 0.30-0.40M have a high drug loading (not less than 18%); when the theoretical feed ratio is lower than 18%, the Treprostinil liposomes prepared when the aqueous phase concentration is between 0.30 and 0.40M can obtain a high encapsulation rate (encapsulation rate is not less than 90%).
表3-1高理论投料比-不同内水相(葡甲胺)浓度的曲前列尼尔脂质体组合物

备注：载药盐阴离子为HAc；理论投料比为21％(误差±5％以内)；脂质体的粒径范围是100～150nm之
间；Table 3-1 High theoretical feed ratio - Treprostinil liposome composition with different internal aqueous phase (methylglucamine) concentrations

Note: The anion of the drug-carrying salt is HAc; the theoretical feed ratio is 21% (within ±5%); the particle size range of the liposome is between 100 and 150 nm;
表3-2低理论投料比-不同内水相(葡甲胺)浓度的曲前列尼尔脂质体组合物

备注：载药盐阴离子为HAc，；理论投料比为15％(误差±5％以内)；脂质体的粒径范围是100～150nm
之间；Table 3-2 Low theoretical feed ratio - Treprostinil liposome composition with different internal aqueous phase (meglumine) concentrations

Note: The anion of the drug-carrying salt is HAc; the theoretical feed ratio is 15% (within ±5%); the particle size range of the liposome is 100-150nm
between;
实施例4不同磷脂处方的曲前列尼尔脂质体组合物Example 4 Treprostinil liposome compositions with different phospholipid prescriptions
按照上述“一般实验程序”制备包含不同磷脂处方的曲前列尼尔脂质体组合物。并按照上述的一般分析方法对脂质体组合物进行分析。 Treprostinil liposome compositions containing different phospholipid formulations were prepared according to the above-mentioned "General Experimental Procedures", and the liposome compositions were analyzed according to the above-mentioned general analytical methods.
结果：从表中可以看出，在高理论投料比时，HSPC、CHOL和DSPE-mPEG2000的摩尔比为3/2/(0.025-0.150)时，制备的曲前列尼尔脂质体具有高载药量(不低于18％)；HSPC、CHOL和DSPE-mPEG2000的摩尔比为3/(2～3)/0.075时，制备的曲前列尼尔脂质体具有高载药量(不低于18％)和高包封率(不低于85％)。Results: It can be seen from the table that at high theoretical feed ratios, when the molar ratio of HSPC, CHOL and DSPE-mPEG2000 is 3/2/(0.025-0.150), the prepared treprostinil liposomes have a high drug loading (not less than 18%); when the molar ratio of HSPC, CHOL and DSPE-mPEG2000 is 3/(2-3)/0.075, the prepared treprostinil liposomes have a high drug loading (not less than 18%) and a high encapsulation rate (not less than 85%).
表4-1不同磷脂处方的曲前列尼尔脂质体组合物

备注：载药盐阳离子为MEG(0.30M)；载药盐阴离子为HAc(0.29M)；理论投料比为21％(误差±5％
以内)；脂质体的粒径范围是100～150nm之间。Table 4-1 Treprostinil liposome compositions with different phospholipid prescriptions

Note: The cation of the drug-carrying salt is MEG (0.30M); the anion of the drug-carrying salt is HAc (0.29M); the theoretical feed ratio is 21% (error ± 5%)
The particle size of the liposome is between 100 and 150 nm.
表4-2不同磷脂处方的曲前列尼尔脂质体组合物

备注：载药盐阳离子为MEG；载药盐阴离子为HAc；理论投料比为21％(误差±5％以内)。上述脂质体
的粒径范围是100～150nm之间。Table 4-2 Treprostinil liposome compositions with different phospholipid prescriptions

Note: The drug-carrying salt cation is MEG; the drug-carrying salt anion is HAc; the theoretical feed ratio is 21% (within ±5%). The particle size range of the above liposomes is between 100 and 150 nm.
实施例5不同内水相pH的曲前列尼尔脂质体组合物Example 5 Treprostinil liposome compositions with different internal aqueous phase pH
按照上述“一般实验程序”，调整内水相乙酸的用量(0.21～0.40M)，制备不同内水相pH处方的曲前列尼尔脂质体组合物。并按照上述一般分析方法对脂质体组合物进行分析。According to the above "general experimental procedure", the amount of acetic acid in the internal aqueous phase was adjusted (0.21-0.40 M) to prepare treprostinil liposome compositions with different internal aqueous phase pH prescriptions. The liposome compositions were analyzed according to the above general analytical method.
结果：从表中可以看出，内水相溶液中的乙酸浓度在0.21～0.40M，对应的内水相溶液pH为9.52～4.97时，制备的曲前列尼尔脂质体具有高载药量(不低于18％)。Results: As can be seen from the table, when the acetic acid concentration in the inner aqueous phase solution is 0.21-0.40 M and the corresponding inner aqueous phase solution pH is 9.52-4.97, the prepared treprostinil liposomes have a high drug loading capacity (not less than 18%).
表5不同内水相pH的曲前列尼尔脂质体组合物


备注：载药盐阳离子为MEG(0.30M)；理论投料比为22.3％(误差±5％以内)；粒径范围为100～200nm；Table 5 Treprostinil liposome compositions at different internal aqueous phase pH


Note: The drug-carrying salt cation is MEG (0.30M); the theoretical feed ratio is 22.3% (within ±5%); the particle size range is 100-200nm;
实施例6不同内水相阴离子的曲前列尼尔脂质体组合物Example 6 Treprostinil liposome compositions with different internal aqueous phase anions
按照上述“一般实验程序”制备不同内水相阴离子处方的曲前列尼尔脂质体组合物。并按照上述的一般分析方法对脂质体组合物进行分析。Treprostinil liposome compositions with different inner aqueous phase anion formulations were prepared according to the above-mentioned "general experimental procedure", and the liposome compositions were analyzed according to the above-mentioned general analytical method.
结果：从表中可以看出，保持内水相pH为8.5，乙酸作为内水相阴离子，制备的曲前列尼尔脂质体具有高载药量(不低于18％)；戊酸和乳酸作为内水相阴离子，在理论投料比低于18％时，制得的曲前列尼尔脂质体可获得高包封率(包封率不低于85％)。葡萄糖酸、苹果酸、柠檬酸、磷酸在本组合物中不适宜作为提供梯度的阴离子。Results: As can be seen from the table, the prepared treprostinil liposomes have a high drug loading (not less than 18%) when the pH of the inner aqueous phase is maintained at 8.5 and acetic acid is used as the inner aqueous phase anion; valeric acid and lactic acid are used as the inner aqueous phase anions, and when the theoretical feed ratio is lower than 18%, the prepared treprostinil liposomes can obtain a high encapsulation rate (encapsulation rate not less than 85%). Gluconic acid, malic acid, citric acid, and phosphoric acid are not suitable as anions for providing gradients in this composition.
表6不同内水相酸的种类的曲前列尼尔脂质体组合物

备注：载药盐阳离子为MEG(0.30M)；上述脂质体的粒径范围是100～150nm之间。Table 6 Treprostinil liposome compositions with different types of internal aqueous phase acid

Note: The drug-carrying salt cation is MEG (0.30M); the particle size range of the above liposomes is between 100 and 150 nm.
实施例7不同弱酸性药物种类的脂质体药物组合物Example 7 Liposome pharmaceutical compositions of different weakly acidic drug types
按照上述“一般实验程序”制备的脂质体组合物，分别负载不同药物，并按照上述的一般分析方法对脂质体组合物进行分析。The liposome compositions prepared according to the above-mentioned "general experimental procedure" were loaded with different drugs, and the liposome compositions were analyzed according to the above-mentioned general analytical method.
结果：从表中可以看出，负载不同药物的脂质体组合物均具有高载药量(不低于18％)。Results: As can be seen from the table, the liposome compositions loaded with different drugs all have high drug loading (not less than 18%).
表7不同弱酸性药物的脂质体药物组合物

备注：载药盐阳离子为MEG(0.30M)；载药盐阴离子为HAc(0.29M)；理论投料比为22％(误差±5％
以内)；脂质体的粒径范围是100～150nm之间。Table 7 Liposome drug compositions of different weakly acidic drugs

Note: The cation of the drug-carrying salt is MEG (0.30M); the anion of the drug-carrying salt is HAc (0.29M); the theoretical feed ratio is 22% (error ± 5%)
The particle size of the liposome is between 100 and 150 nm.
实施例8曲前列尼尔脂质体药物组合物Example 8 Treprostinil liposome pharmaceutical composition
按照上述“一般实验程序”制备的脂质体组合物，负载曲前列尼尔钠，并按照上述的一般分析方法对脂质体组合物进行分析。The liposome composition prepared according to the above-mentioned "General Experimental Procedure" was loaded with treprostinil sodium, and the liposome composition was analyzed according to the above-mentioned general analytical method.
结果：从表中可以看出，负载曲前列尼尔的脂质体组合物具有高载药量和高包封率。 Results: As can be seen from the table, the liposome composition loaded with treprostinil has a high drug loading capacity and a high encapsulation efficiency.
表8曲前列尼尔脂质体药物组合物

备注：载药盐阳离子为MEG(0.30M)；载药盐阴离子为HAc(0.29M)；Table 8 Treprostinil liposome pharmaceutical composition

Note: The cation of the drug-loaded salt is MEG (0.30M); the anion of the drug-loaded salt is HAc (0.29M);
实施例9.不同磷脂浓度的曲前列尼尔脂质体组合物Example 9. Treprostinil liposome compositions with different phospholipid concentrations
按照上述“一般实验程序”，保持投料比一致，按照不同磷脂浓度投料制备曲前列尼尔脂质体组合物。并按照上述的一般分析方法对脂质体组合物进行分析。According to the above "general experimental procedure", the feed ratio was kept consistent and the feed was fed at different phospholipid concentrations to prepare the treprostinil liposome composition. The liposome composition was analyzed according to the above general analysis method.
结果：从表中可以看出，当理论投料比为22％时，载药时的组合物，其磷脂浓度1.4～4.18mg/ml范围内制备的曲前列尼尔脂质体具有高载药量(不低于18％)。Results: It can be seen from the table that when the theoretical feed ratio is 22%, the drug-loaded composition, whose phospholipid concentration is in the range of 1.4-4.18 mg/ml, has a high drug loading capacity (not less than 18%).
表9不同磷脂浓度的曲前列尼尔脂质体组合物

备注：载药盐阳离子为MEG(0.30M)；载药盐阴离子为HAc(0.29M)；理论投料比为22％(误差±5％
以内)；脂质体的粒径范围是100～150nm之间。Table 9 Treprostinil liposome compositions with different phospholipid concentrations

Note: The cation of the drug-carrying salt is MEG (0.30M); the anion of the drug-carrying salt is HAc (0.29M); the theoretical feed ratio is 22% (error ± 5%)
The particle size of the liposome is between 100 and 150 nm.
以上对本发明技术方案的实施方式进行了示例性的说明。应当理解，本发明的保护范围不拘囿于上述实施方式。凡在本发明的精神和原则之内，本领域技术人员所做的任何修改、等同替换、改进等，均应包含在本申请权利要求书的保护范围之内。 The above is an exemplary description of the implementation of the technical solution of the present invention. It should be understood that the protection scope of the present invention is not limited to the above implementation. Any modification, equivalent substitution, improvement, etc. made by those skilled in the art within the spirit and principle of the present invention should be included in the protection scope of the claims of this application.

Claims (10)

  1. 一种脂质体药物组合物，包括弱酸性药物和脂质体；A liposome pharmaceutical composition comprising a weakly acidic drug and liposomes;
    所述脂质体为如下中的一种：The liposome is one of the following:
    (1)所述脂质体包括磷脂和内水相；(1) The liposome comprises phospholipids and an internal aqueous phase;
    (2)所述脂质体包括磷脂、内水相和外水相；(2) The liposome comprises phospholipids, an inner aqueous phase and an outer aqueous phase;
    其中，所述内水相包括葡甲胺与弱酸，所述内水相的pH为4.0～10.5，例如4.5～10.0，6.5～10.5，优选4.97～9.52；Wherein, the inner aqueous phase comprises meglumine and a weak acid, and the pH of the inner aqueous phase is 4.0 to 10.5, for example, 4.5 to 10.0, 6.5 to 10.5, preferably 4.97 to 9.52;
    所述外水相使脂质体悬浮；The external aqueous phase suspends the liposomes;
    所述弱酸性药物包裹在脂质体的内水相中，包裹了弱酸性药物的脂质体悬浮在外水相中。The weakly acidic drug is encapsulated in the inner water phase of the liposome, and the liposome encapsulating the weakly acidic drug is suspended in the outer water phase.
  2. 根据权利要求1所述的脂质体药物组合物，其特征在于，所述弱酸性药物的pKa在2～7之间；The liposome pharmaceutical composition according to claim 1, characterized in that the pKa of the weakly acidic drug is between 2 and 7;
    优选地，所述弱酸性药物选自：***素类、解热镇痛类、喹啉羧酸抗菌类、干扰素基因刺激因子(STING)受体，或它们的衍生物(如药学上可以接受的盐、酯或前药)中的至少一种药物；Preferably, the weakly acidic drug is selected from: at least one drug selected from: prostaglandins, antipyretics, quinoline carboxylic acid antibiotics, interferon gene stimulator (STING) receptors, or their derivatives (such as pharmaceutically acceptable salts, esters or prodrugs);
    优选地，所述***素类药物选自曲前列尼尔、贝前列素、伊洛前列素、卡前列素、利马前列素、依前列醇、前列地尔、乌诺前列酮，或它们的衍生物；所述解热镇痛类药物选自阿司匹林、布洛芬、萘普生、双氯芬酸钠，或它们的衍生物；所述喹啉羧酸类抗菌药物选自萘啶酸、吡咯嘧酸、氟喹啉酸、西他沙星、环丙沙星、依诺沙星，或它们的衍生物；所述STING药物选自MSA-2、STING agonist-7、Vadimezan，或它们的衍生物；Preferably, the prostaglandin drugs are selected from treprostinil, beraprost, iloprost, carboprost, limaprost, epoprostenol, alprostadil, unoprostone, or their derivatives; the antipyretic and analgesic drugs are selected from aspirin, ibuprofen, naproxen, diclofenac sodium, or their derivatives; the quinoline carboxylic acid antibacterial drugs are selected from nalidixic acid, pyroximate, fluoroquinolinic acid, sitafloxacin, ciprofloxacin, enoxacin, or their derivatives; the STING drug is selected from MSA-2, STING agonist-7, Vadimezan, or their derivatives;
    优选地，所述药物脂质体的载药量低于18％，其包封率不低于85％，优选不低于90％；Preferably, the drug loading of the drug liposome is less than 18%, and the encapsulation efficiency is not less than 85%, preferably not less than 90%;
    优选地，当所述弱酸性药物的载药量不低于18％，不高于40％。Preferably, the drug loading of the weakly acidic drug is not less than 18% and not more than 40%.
  3. 根据权利要求1或2所述的脂质体药物组合物，其特征在于，所述弱酸选自羧酸，例如选自甲酸、乙酸、丙酸、丁酸、异丁酸、戊酸、异戊酸、乳酸或其组合；优选甲酸、乙酸、丙酸或其组合；The liposome pharmaceutical composition according to claim 1 or 2, characterized in that the weak acid is selected from carboxylic acids, such as formic acid, acetic acid, propionic acid, butyric acid, isobutyric acid, valeric acid, isovaleric acid, lactic acid or a combination thereof; preferably formic acid, acetic acid, propionic acid or a combination thereof;
    优选地，所述葡甲胺在内水相中的浓度为0.1～0.8M，例如0.2～0.6M，优选0.3～0.4M；Preferably, the concentration of meglumine in the inner aqueous phase is 0.1 to 0.8 M, such as 0.2 to 0.6 M, preferably 0.3 to 0.4 M;
    优选地，所述弱酸在内水相中的浓度为0.1～0.8M，例如0.2～0.6M。Preferably, the concentration of the weak acid in the inner aqueous phase is 0.1-0.8M, such as 0.2-0.6M.
  4. 根据权利要求1-3任一项所述的脂质体药物组合物，其特征在于，所述葡甲胺与弱酸为葡甲胺与乙酸；优选地，所述葡甲胺与乙酸在内水相中的浓度为0.30～0.40M。The liposome pharmaceutical composition according to any one of claims 1 to 3, characterized in that the meglumine and the weak acid are meglumine and acetic acid; preferably, the concentration of the meglumine and acetic acid in the inner aqueous phase is 0.30 to 0.40 M.
  5. 根据权利要求1-4任一项所述的脂质体药物组合物，其特征在于，所述磷脂具有双层脂质结构，所述双层脂质由氢化大豆卵磷脂(HSPC)、胆固醇(CHOL)和二硬脂酰基磷脂酰乙醇胺-聚乙二醇2000(DSPE-mPEG2000)构成，摩尔比为3:(1-3):(0.025-0.225)，优选3:2:(0.025-0.150)；The liposome pharmaceutical composition according to any one of claims 1 to 4, characterized in that the phospholipid has a bilayer lipid structure, and the bilayer lipid is composed of hydrogenated soybean phosphatidylcholine (HSPC), cholesterol (CHOL) and distearoyl phosphatidylethanolamine-polyethylene glycol 2000 (DSPE-mPEG2000), and the molar ratio is 3: (1-3): (0.025-0.225), preferably 3: 2: (0.025-0.150);
    优选地，所述磷脂在脂质体组合物中的浓度为1～100mg/mL，例如1～50mg/mL、1～20mg/mL。Preferably, the concentration of the phospholipid in the liposome composition is 1-100 mg/mL, such as 1-50 mg/mL, 1-20 mg/mL.
  6. 根据权利要求1-5任一项所述的脂质体药物组合物，其特征在于，所述脂质体的粒径为50～500nm，例如100～200nm； The liposome pharmaceutical composition according to any one of claims 1 to 5, characterized in that the particle size of the liposome is 50 to 500 nm, for example 100 to 200 nm;
    优选地，所述脂质体具有内水相pH高，外水相pH低的pH梯度特性。Preferably, the liposome has a pH gradient property with a high pH in the inner aqueous phase and a low pH in the outer aqueous phase.
  7. 根据权利要求1-6任一项所述的脂质体药物组合物，其特征在于，所述外水相为pH为4.5～6.5的缓冲液；The liposome pharmaceutical composition according to any one of claims 1 to 6, characterized in that the external aqueous phase is a buffer solution with a pH of 4.5 to 6.5;
    优选地，所述缓冲液选自HEPES、氯化钠、蔗糖、枸橼酸盐缓冲液、磷酸盐缓冲液、Tris缓冲液中的一种或几种，优选枸橼酸盐缓冲液；所述缓冲液的浓度为0.05M～0.25M，例如为0.08M～0.2M，优选为0.09～0.12M。Preferably, the buffer is selected from one or more of HEPES, sodium chloride, sucrose, citrate buffer, phosphate buffer, and Tris buffer, preferably citrate buffer; the concentration of the buffer is 0.05M to 0.25M, for example, 0.08M to 0.2M, preferably 0.09 to 0.12M.
  8. 一种基于权利要求1-7任一项所述脂质体药物组合物的曲前列尼尔脂质体，其中，所述脂质体药物组合物中的弱酸性药物为曲前列尼尔或其衍生物，所述曲前列尼尔或其衍生物包裹于所述脂质体的内水相；A treprostinil liposome based on the liposome pharmaceutical composition according to any one of claims 1 to 7, wherein the weakly acidic drug in the liposome pharmaceutical composition is treprostinil or a derivative thereof, and the treprostinil or a derivative thereof is encapsulated in the inner aqueous phase of the liposome;
    优选地，所述曲前列尼尔的衍生物选自曲前列尼尔的水合物、溶剂合物或络合物，或者选自曲前列尼尔的药学上可接受的盐、酯或其前药等，如曲前列尼尔钠盐、曲前列尼尔钾盐、曲前列尼尔二乙醇胺盐、曲前列尼尔甲酯、曲前列尼尔乙酯、曲前列尼尔的富马酰基二酮哌啶前药。Preferably, the derivative of treprostinil is selected from hydrates, solvates or complexes of treprostinil, or pharmaceutically acceptable salts, esters or prodrugs thereof of treprostinil, such as treprostinil sodium salt, treprostinil potassium salt, treprostinil diethanolamine salt, treprostinil methyl ester, treprostinil ethyl ester, and treprostinil fumaryl diketopiperidine prodrug.
  9. 权利要求8所述曲前列尼尔脂质体或其衍生物在制备药物中的应用；Use of the treprostinil liposome or its derivatives as claimed in claim 8 in the preparation of medicines;
    优选地，所述药物为治疗肺动脉高压、肺高血压、肺纤维化、间质性肺疾病、慢性阻塞性肺病、哮喘、缺血性疾病、心脏衰竭、动脉硬化、术后抗凝血、视网膜中央静脉阻塞、血栓性微血管病、周围性血管疾病、心肺移植等疾病的药物；Preferably, the drug is a drug for treating pulmonary hypertension, pulmonary hypertension, pulmonary fibrosis, interstitial lung disease, chronic obstructive pulmonary disease, asthma, ischemic disease, heart failure, arteriosclerosis, postoperative anticoagulation, central retinal vein occlusion, thrombotic microangiopathy, peripheral vascular disease, heart and lung transplantation, and the like;
    优选地，所述药物为治疗外周动脉闭塞性疾病或肺动脉高压药物。Preferably, the drug is a drug for treating peripheral arterial occlusive disease or pulmonary hypertension.
  10. 权利要求1-7任一项脂质体药物组合物的制备方法，该方法包括，将脂质体负载至少一种弱酸性药物；A method for preparing a liposome pharmaceutical composition according to any one of claims 1 to 7, the method comprising loading the liposomes with at least one weakly acidic drug;
    优选地，所述弱酸性药物的载药量不低于18％；Preferably, the drug loading of the weakly acidic drug is not less than 18%;
    优选地，所述脂质体药物组合物的制备方法，包括以下步骤：将弱酸性药物溶解在外水相中，与所述脂质体的溶液混合，得到所述脂质体药物组合物；其中所述脂质体的浓度为1.0～20mg/ml，优选1.0～5.0mg/ml(以磷脂浓度计)；Preferably, the method for preparing the liposome pharmaceutical composition comprises the following steps: dissolving a weakly acidic drug in an external aqueous phase and mixing the drug with the liposome solution to obtain the liposome pharmaceutical composition; wherein the concentration of the liposome is 1.0 to 20 mg/ml, preferably 1.0 to 5.0 mg/ml (based on phospholipid concentration);
    优选地，所述脂质体药物组合物的制备方法，包括以下步骤：Preferably, the method for preparing the liposome pharmaceutical composition comprises the following steps:
    (1)将所述弱酸性药物溶于枸橼酸钠水溶液中，得到药物溶液；(1) dissolving the weakly acidic drug in a sodium citrate aqueous solution to obtain a drug solution;
    (2)将所述脂质体的溶液与枸橼酸盐缓冲液混合，得到载体溶液；(2) mixing the liposome solution with a citrate buffer to obtain a carrier solution;
    (3)将所述药物溶液加入到所述载体溶液中进行负载，得到所述脂质体药物组合物。 (3) adding the drug solution into the carrier solution for loading to obtain the liposome drug composition.
PCT/CN2023/122824 2022-09-30 2023-09-28 Liposome pharmaceutical composition, preparation method therefor, and use thereof WO2024067849A1 (en)

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Citations (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5939096A (en) * 1995-04-18 1999-08-17 Yissum Research Development Company Of The Hebrew University Of Jerusalem Liposome drug-loading method and composition
CN1717221A (en) * 2002-11-26 2006-01-04 吉里德科学公司 Method of drug loading in liposomes by gradient
CN101795671A (en) * 2007-08-21 2010-08-04 阿尔扎公司 Liposome compositions for in vivo administration of boronic acid compounds
CN106999419A (en) * 2014-08-04 2017-08-01 佐尼奥尼制药股份有限公司 Remote loading sparingly water-soluble medicine is to lipid film bubble
CN111372580A (en) * 2017-07-24 2020-07-03 国邑药品科技股份有限公司 Liposome composition comprising weak acid drug and use thereof
CN112004527A (en) * 2018-04-23 2020-11-27 美商Tlc生物医药公司 Inhalable liposome sustained-release composition for treating pulmonary diseases
CN112384224A (en) * 2018-05-07 2021-02-19 国邑药品科技股份有限公司 Pharmaceutical compositions for controlling the release of treprostinil
CN113811307A (en) * 2019-05-14 2021-12-17 国邑药品科技股份有限公司 Pharmaceutical composition of weak acid drug and administration method

Patent Citations (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5939096A (en) * 1995-04-18 1999-08-17 Yissum Research Development Company Of The Hebrew University Of Jerusalem Liposome drug-loading method and composition
CN1717221A (en) * 2002-11-26 2006-01-04 吉里德科学公司 Method of drug loading in liposomes by gradient
CN101795671A (en) * 2007-08-21 2010-08-04 阿尔扎公司 Liposome compositions for in vivo administration of boronic acid compounds
CN106999419A (en) * 2014-08-04 2017-08-01 佐尼奥尼制药股份有限公司 Remote loading sparingly water-soluble medicine is to lipid film bubble
CN111372580A (en) * 2017-07-24 2020-07-03 国邑药品科技股份有限公司 Liposome composition comprising weak acid drug and use thereof
CN112004527A (en) * 2018-04-23 2020-11-27 美商Tlc生物医药公司 Inhalable liposome sustained-release composition for treating pulmonary diseases
CN112384224A (en) * 2018-05-07 2021-02-19 国邑药品科技股份有限公司 Pharmaceutical compositions for controlling the release of treprostinil
CN113811307A (en) * 2019-05-14 2021-12-17 国邑药品科技股份有限公司 Pharmaceutical composition of weak acid drug and administration method

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