阅读说明：本技术 一种多胺前体脂质及其应用 (Polyamine precursor lipid and application thereof ) 是由 涂家生 杜运爱 孙春萌 于 2019-08-30 设计创作，主要内容包括：本发明公开了一种多胺前体脂质及其应用,利用肿瘤组织高表达β-葡萄糖醛酸酶特点,采用β-葡萄糖醛酸修饰多胺脂质,形成多胺前体脂质,可降低阳离子多胺脂质体对人体毒副作用。本发明的β-葡萄糖醛酸修饰的多胺前体脂质在在人体生理条件下保持较强的负电荷,可使脂质体具有较好的血液相容性。而在肿瘤组织高表达的葡萄糖醛酸酶下,发生酶解反应,释放游离的多胺脂质,实现表面电荷由负电荷转变成正电荷,易于与带负电荷的肿瘤细胞结合,实现有效内吞进入肿瘤细胞内。(The invention discloses a polyamine precursor lipid and application thereof, which utilizes the characteristic of high expression of beta-glucuronidase by tumor tissues and adopts beta-glucuronic acid to modify polyamine lipid to form the polyamine precursor lipid, so that the toxic and side effects of a cationic polyamine liposome on a human body can be reduced. The beta-glucuronic acid modified polyamine precursor lipid keeps stronger negative charges under the physiological condition of a human body, so that the liposome has better blood compatibility. And under the glucuronidase highly expressed by tumor tissues, enzymolysis reaction is carried out to release free polyamine lipid, so that the surface charge is converted from negative charge to positive charge, the surface charge is easily combined with tumor cells with negative charge, and the effective endocytosis is realized to enter the tumor cells.)

1. A polyamine precursor lipid characterized by: the structural formula is shown as formula I:

wherein R is substituted tri (2-aminoethyl) amino, wherein at least one amino group of the substituted tri (2-aminoethyl) amino is substituted by a fatty acid.

2. The polyamine precursor lipid of claim 1, wherein: the fatty acid is C8-C18 saturated fatty acid or C8-C18 unsaturated fatty acid; the fatty acid is connected with the amino group by an N-alkyl chain or an amido bond-alkyl chain.

3. The polyamine precursor lipid according to claim 2, wherein: the structure of R is shown as formula II:

wherein R is₁And R₂Each and independently selected from oleic acid, linoleic acid, stearic acid, lauric acid, palmitic acid, linolenic acid, capric acid or caprylic acid.

4. A liposome, characterized by: comprises phospholipid, cholesterol and the polyamine precursor lipid; the mass ratio of the phospholipid to the cholesterol is 2-20: 1, the mass of the polyamine precursor lipid is 5-20% of the total mass of the phospholipid and the cholesterol.

5. The liposome of claim 4, wherein: the phospholipid is one or more of soybean lecithin and its derivatives, egg yolk lecithin and its derivatives, phosphatidylcholine and its derivatives, phosphatidylethanolamine and its derivatives, phosphatidylglycerol and its derivatives, phosphatidylserine and its derivatives, phosphatidic acid and its derivatives, and phosphatidylinositol and its derivatives.

6. The liposome of claim 5, wherein: the phospholipid is one or more of soybean lecithin and derivatives thereof, distearoyl phosphatidyl ethanolamine and derivatives thereof, distearoyl phosphatidyl choline and derivatives thereof or 1, 2-palmitoyl phosphatidyl glycerol and derivatives thereof.

7. The liposome of claim 6, wherein: the phospholipid is one or more of soybean lecithin, dipalmitoyl phosphatidylcholine or 1-myristoyl-2-stearoyl lecithin.

8. The liposome of claim 4, wherein: the liposome also comprises an anti-tumor drug.

9. The liposome of claim 8, wherein: the anti-tumor drug is selected from cisplatin, oxaliplatin, paclitaxel, deoxypodophyllotoxin, etoposide, SN-38, paclitaxel, docetaxel, lonidamine or adriamycin.

10. Use of a polyamine precursor lipid according to any one of claims 1 to 3 in the manufacture of a medicament for the treatment of a tumour.

Technical Field

The invention belongs to the technical field of high molecular materials, and particularly relates to polyamine precursor lipid based on tumor microenvironment extracellular enzyme activation and application thereof in a pharmaceutical preparation.

Background

The enzyme is an important stimulating factor in organisms and plays an important role in maintaining normal physiological functions in the organisms. In certain pathological environments, the expression of enzymes changes, for example, certain proteases, glycosidases or phospholipases can be used as biomarkers for the diagnosis and treatment of inflammation, tumors and neurodegenerative diseases. The tumor tissue over-expressed enzyme is combined with a nano delivery carrier such as a polymer and a liposome, and a tumor extracellular enzyme response type delivery system is designed by utilizing the characteristics of catalytic specificity and high efficiency of the enzyme to a substrate, so that the functions of charge reversal, cell penetrating, active targeting, triggered drug release and the like are exerted, and the aim of diagnosis and treatment integrating accurate diagnosis and treatment of tumors or integration of the two is fulfilled.

The liposome as new antitumor medicine carrier has the advantages of improving the pharmacokinetics property of medicine in vivo, increasing antitumor effect, reducing the toxic and side effect of medicine, etc. The clinically used liposome medicaments such as adriamycin liposome/paclitaxel liposome show good anti-tumor effect. However, the liposome preparation has the problems of low tumor targeting, slow tumor cell uptake, slow drug release and the like, and the clinical application of the liposome anti-tumor preparation is limited. In recent years, researchers develop pH sensitive liposome, extracellular enzyme sensitive liposome, ROS sensitive liposome and the like based on a tumor microenvironment to overcome the defects of poor targeting property, low tumor cell uptake efficiency and slow drug release of liposome preparations.

Disclosure of Invention

The invention aims to provide a polyamine precursor lipid based on activation of tumor microenvironment ectoenzyme, which utilizes the characteristic of high expression of beta-glucuronidase in tumor tissues and adopts beta-glucuronic acid to modify polyamine lipid to form polyamine precursor lipid, so that the toxic and side effects of a cationic polyamine liposome on a human body can be reduced.

A polyamine proliposome having a structural formula shown in formula I:

I

wherein R is substituted tri (2-aminoethyl) amino, wherein at least one amino group of the substituted tri (2-aminoethyl) amino is substituted by a fatty acid.

Further, the fatty acid is a saturated fatty acid of C8-C18 or an unsaturated fatty acid of C8-C18; the fatty acid is connected with the amino group by an N-alkyl chain or an amido bond-alkyl chain.

Further, the structure of R is shown as formula II:

wherein R is₁And R₂Each and independently selected from oleic acid, linoleic acid, stearic acid, lauric acid, palmitic acid, linolenic acid, capric acid or caprylic acid.

A liposome comprises phospholipid, cholesterol and the polyamine precursor lipid; the mass ratio of the phospholipid to the cholesterol is 2-20: 1, the mass of the polyamine precursor lipid is 5-20% of the total mass of the phospholipid and the cholesterol.

Further, the phospholipid is one or a mixture of more of soybean lecithin and derivatives thereof, egg yolk lecithin and derivatives thereof, phosphatidylcholine and derivatives thereof, phosphatidylethanolamine and derivatives thereof, phosphatidylglycerol and derivatives thereof, phosphatidylserine and derivatives thereof, phosphatidic acid and derivatives thereof, and phosphatidylinositol and derivatives thereof.

Further, the phospholipid is one or a mixture of more of soybean lecithin and derivatives thereof, distearoylphosphatidylethanolamine and derivatives thereof, distearoylphosphatidylcholine and derivatives thereof or 1, 2-palmitoylphosphatidylglycerol and derivatives thereof.

Further, the phospholipid is one or a mixture of more of soybean lecithin, dipalmitoyl phosphatidylcholine and 1-myristoyl-2-stearoyl lecithin.

Furthermore, the liposome also comprises an anti-tumor drug.

Further, the anti-tumor drug is selected from cisplatin, oxaliplatin, paclitaxel, deoxypodophyllotoxin, etoposide, SN-38, paclitaxel, docetaxel, lonidamine or adriamycin.

The application of the polyamine precursor lipid in preparing the medicine for treating tumor.

Has the advantages that: the polyamine precursor lipid of the invention has beta-glucuronic acid modified group, keeps stronger negative charge under the physiological condition of human body, and can make the liposome have better blood compatibility after being used for preparing the liposome. And under the glucuronidase highly expressed by tumor tissues, enzymolysis reaction is carried out to release free polyamine lipid, so that the surface charge is converted from negative charge to positive charge, the surface charge is easily combined with tumor cells with negative charge, and the effective endocytosis is realized to enter the tumor cells. After entering lysosome by endocytosis, the lysosome has high activity beta-glucuronidase, which can hydrolyze glucuronic acid and release free primary amine. Polyamine lipids capture protons, exert a proton sponge effect or act with the lysosome membrane, lyse the lysosome membrane, allow the polyamine liposomes to enter the cytosol, and aggregate within the liposomes through electrostatic interactions.

Drawings

FIG. 1 is a graph of the potential versus time curve for the GluAcNA liposomes of example 1.

FIG. 2 is a graph of buffering capacity of the liposomes of each type in example 1.

Figure 3 is a cytotoxicity curve of liposomes from example 2.

Detailed Description

The invention discloses a polyamine precursor lipid based on tumor microenvironment extracellular enzyme activation and application thereof in liposome preparation, and can be realized by appropriately improving process parameters by referring to the content. It is expressly intended that all such similar substitutes and modifications which would be obvious to one skilled in the art are deemed to be included in the invention. While the products and applications of this invention have been described in terms of preferred embodiments, it will be apparent to those of ordinary skill in the art that variations and modifications in the methods and applications described herein, as well as other suitable variations and combinations, may be made to implement and use the techniques of this invention without departing from the spirit and scope of the invention.

In a specific embodiment, the design idea of the invention is as follows:

aiming at the technical problem that the cationic polyamine liposome has toxic and side effects on human bodies, the invention designs a class of beta-glucuronidase sensitive polyamine precursor lipid, and the lipid material takes tri (2-aminoethyl) amine as a connecting arm to modify hydrophilic beta-glucuronic acid on an oleic acid derivative.

In the present invention, the oleic acid derivative is preferably one selected from the following:

、、

、。

because the beta-glucuronidase is highly expressed in the tumor tissue, especially in a tumor necrosis area, the invention utilizes the characteristic of the beta-glucuronidase highly expressed in the tumor tissue, adopts the beta-glucuronic acid to modify the polyamine lipid to form the polyamine precursor lipid, and the polyamine precursor lipid is doped into the liposome, thereby reducing the toxic and side effects of the cationic polyamine liposome on the human body. Meanwhile, the polyamine precursor lipid can be hydrolyzed by beta-glucuronidase in a tumor region to expose free polyamine lipid and increase the uptake of tumor cells to the liposome. Because the tri (2-aminoethyl) amine contains a tertiary amine structure, the liposome can escape from the inside of a lysosome to enter the cell through the proton sponge effect, and can be gathered in the inner part of the mitochondria under the drive of positive charges, so that the drug can be delivered to the mitochondria of tumor cells, and the aim of gradually delivering and targeting the mitochondria under the drive of the charges can be realized.

The polyamine precursor lipid material is light yellow solid powder which is easily dissolved in dichloromethane and chloroform. Under the action of beta-glucuronidase, glucuronide chain can be broken, and polyamine lipid is released through 1,6 self-elimination reaction. The polyamine precursor polyamine lipid can be prepared into a liposome responding to beta-glucuronidase together with phospholipid, cholesterol and the like with different amounts. Under the action of tumor acidity and high expression beta-glucuronidase, electronegative beta-glucuronic acid is removed to form polyamine cationic liposome with positive charge, as shown in figure 1. The polyamine cationic liposome has strong pH buffering capacity, and can ensure that the liposome generates proton sponge effect under the acidic condition of lysosome and escapes from the lysosome, as shown in figure 2.

The polyamine precursor lipid based on the activation of tumor microenvironment extracellular enzyme and the application thereof provided by the invention are explained in detail below, and the experimental method without specific conditions and the reagent without formula are all according to the conventional conditions in the field