Irinotecan-containing polymer micelle, preparation and use

Abstract

The present invention belongs to the technical field of pharmaceutical preparations, and specifically relates to a polymer micelle containing irinotecan, a preparation, and its use. The present invention utilizes polyethylene glycol-b-polylysine-b-polyleucine triblock micelles to encapsulate irinotecan, and optimizes the drug loading preparation process, significantly improving the drug activity closed loop rate, avoiding the ring-opening hydrolysis of the lactone ring in the irinotecan structure, effectively retaining its anti-tumor activity, and avoiding toxic side effects. At the same time, the drug loading capacity of the irinotecan polymer micelle is increased. Furthermore, the irinotecan injection of the present invention also exhibits extremely high stability and low content of related substances.

Description

Technical Field

[0001] The present invention belongs to the technical field of pharmaceutical preparations, and in particular relates to a polymer micelle containing irinotecan, a preparation and a use thereof. Background Art

[0002] Irinotecan is a semisynthetic, water-soluble camptothecin derivative. Camptothecin specifically binds to topoisomerase I, inducing reversible single-strand breaks and thereby unwinding the DNA double-strand structure. Irinotecan and its active metabolite, SN-38, bind to the topoisomerase I-DNA complex, preventing the reconnection of broken single strands. Currently available irinotecan products include irinotecan hydrochloride injection, lyophilized powder injection, and liposome injection.

[0003] Irinotecan contains a lactone ring, which readily undergoes ring-opening hydrolysis under neutral and alkaline conditions. Literature reports that camptothecin-based anticancer drugs are only effective in their closed-ring form; once the ring is opened, they lose their antitumor activity and inevitably lead to toxic side effects. Therefore, improving the active ring-closure rate of camptothecin-based drugs is a key issue for their successful application.

[0004] The Chinese patent publication number CN 102961332 A discloses a micellar preparation with a high active ring closure rate of camptothecin derivatives, which uses an amphiphilic copolymer as a carrier material, physically loads an active dose of camptothecin derivative drug molecules in micelle form, and uses an aqueous medium as the main solvent. The main structure of the carrier material is polyethylene glycol as a hydrophilic segment and polyester, polyamino acid or polyether as a hydrophobic segment. The drug molecule is one or more of camptothecin, 10-hydroxycamptothecin, 9-nitrocamptothecin, 9-aminocamptothecin, 7-ethyl-10-hydroxycamptothecin, topotecan, irinotecan, rubitecan, exitecan, berotelecan, karenitecin, gemitecan, gimatidecan, etc. This micellar preparation can significantly improve the active ring closure rate of camptothecin drugs, enhance drug efficacy, effectively reduce the systemic reaction of the drug, and can be used for tumor treatment at different stages. However, the maximum weight percentage of the drug contained therein is only 10%, the drug loading amount is low, and the drug delivery efficiency is low. Summary of the Invention

[0005] To overcome the deficiencies of the prior art, the present invention provides an irinotecan-containing polymer micelle with high drug loading and high active ring closure rate, and further provides an irinotecan-containing polymer micelle preparation with high stability.

[0006] The technical solution is as follows:

[0007] The first object of the present invention is to provide an irinotecan-containing polymer micelle composed of irinotecan and polyethylene glycol-b-polylysine-b-polyleucine triblock micelles.

[0008] In various embodiments, the irinotecan-containing polymer micelles contain 1 to 3 parts of irinotecan and 6 to 30 parts of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles by weight.

[0009] In a preferred embodiment, the irinotecan-containing polymer micelles contain 2 parts of irinotecan and 7 parts of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles by weight.

[0010] A second object of the present invention is to provide a method for preparing the above-mentioned irinotecan-containing polymer micelles, comprising: dissolving irinotecan in a buffer solution having a pH of 4.0 to 5.0, uniformly mixing the solution with polyethylene glycol-b-polylysine-b-polyleucine triblock micelles, adding an organic solvent, thoroughly shaking the solution in a water bath at -5°C to 5°C, and dialyzing the solution to obtain the irinotecan-containing polymer micelles.

[0011] In many embodiments, the organic solvent is selected from at least one of methanol, ethanol, chloroform, isopropanol, n-butanol, and dimethyl sulfoxide.

[0012] In a preferred embodiment, the organic solvent is a mixed solvent of dimethyl sulfoxide and n-butanol.

[0013] In some embodiments, the water bath temperature is -2 to 2°C.

[0014] In various embodiments, the buffer solution is selected from one of disodium hydrogen phosphate-citric acid buffer solution, glycine-hydrochloric acid buffer solution, and citric acid-sodium citrate buffer solution.

[0015] In a preferred embodiment, the buffer solution is a disodium hydrogen phosphate-citric acid buffer solution.

[0016] In a preferred embodiment, the pH of the buffer solution is 4.5.

[0017] The third object of the present invention is to provide an irinotecan preparation, which is composed of the above-mentioned irinotecan-containing polymer micelles and pharmaceutically acceptable excipients.

[0018] In an embodiment, the preparation is an injection.

[0019] The fourth object of the present invention is to provide use of the irinotecan-containing polymer micelles in the preparation of drugs for treating tumors.

[0020] Compared with the prior art, the technical effects of the present invention are:

[0021] The present invention utilizes polyethylene glycol-b-polylysine-b-polyleucine triblock micelles to entrap irinotecan, optimizes the drug loading preparation process, and particularly controls the reaction temperature between -5°C and 5°C, thereby significantly improving the drug activity ring closure rate, avoiding the ring-opening hydrolysis of the lactone ring in the irinotecan structure, effectively retaining its anti-tumor activity, and avoiding toxic side effects. Simultaneously, the drug loading capacity of the irinotecan polymer micelles is increased to as high as approximately 45%, thereby greatly improving the drug delivery efficiency. Furthermore, the irinotecan-containing polymer micelles are further prepared into an irinotecan injection, and it is found that the irinotecan injection of the present invention has a low content of related substances and high stability. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 : Electron microscopic image of polymer micelles containing irinotecan in Example 1 of the present invention

[0023] Figure 2 : Schematic diagram of the polymer micelle containing irinotecan of the present invention

[0024] Figure 3 : Drug loading of irinotecan-containing polymer micelles in Examples 1 to 5 of the present invention

[0025] Figure 4 : Comparative Examples 1 to 4 Drug Loading of Irinotecan-Containing Polymer Micelle

[0026] Figure 5 : Determination of drug active ring closure rate in polymer micelles containing irinotecan in Examples 1 to 5 and Comparative Examples 1 to 4

[0027] Figure 6 : Example 6, Comparative Examples 5-6, and Results of Accelerated Tests of Related Substances of Commercially Available Preparations DETAILED DESCRIPTION

[0028] 1. Preparation of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles

[0029] (1) Synthesis of MPEG-b-PZLL: MPEG-NH and lysine anhydride were mixed in a molar ratio of 1:15 to 1:30, stirred in a water bath at 28 to 32°C for 60 to 80 hours, centrifuged, and the precipitate was collected and vacuum dried to obtain MPEG-b-PZLL.

[0030] (2) Synthesis of MPEG-b-PZLL-b-PLLeu: Mix the MPEG-b-PZLL prepared in step (1) and leucine anhydride in a molar ratio of 1:20 to 1:40, stir well in a water bath at 28 to 32°C for 60 to 80 hours, centrifuge, collect the precipitate, and vacuum dry to obtain MPEG-b-PZLL-b-PLLeu;

[0031] (3) Deprotection of the Z group of MPEG-b-PZLL-b-PLLeu: Ethanol containing 33 wt% HBr was added to the MPEG-b-PZLL-b-PLLeu obtained in step (2), and then an appropriate amount of trifluoroacetic acid was added. The mixture was reacted in a water bath at 20-25°C for 2-4 h to obtain polyethylene glycol-b-polylysine-b-polyleucine triblock micelles.

[0032] 2. Irinotecan-containing polymer micelles

[0033] Example 1

[0034] formula:

[0035] 1 part by weight of irinotecan

[0036] 3 parts by weight of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles

[0037] Preparation method:

[0038] Irinotecan was dissolved in a sodium hydrogen phosphate-citric acid buffer solution at pH 4.5, and then mixed evenly with polyethylene glycol-b-polylysine-b-polyleucine triblock micelles. A mixed solvent of dimethyl sulfoxide and n-butanol (volume ratio 1:1) was added, and the mixture was fully shaken in a 0°C water bath and dialyzed to obtain polymer micelles containing irinotecan.

[0039] Example 2

[0040] formula:

[0041] 2 parts by weight of irinotecan

[0042] 7 parts by weight of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles

[0043] Preparation method:

[0044] Irinotecan was dissolved in a sodium hydrogen phosphate-citric acid buffer solution at pH 4.0, and then mixed evenly with polyethylene glycol-b-polylysine-b-polyleucine triblock micelles. A mixed solvent of dimethyl sulfoxide and n-butanol (volume ratio 2:1) was added, and the mixture was fully shaken in a -2°C water bath and dialyzed to obtain polymer micelles containing irinotecan.

[0045] Example 3

[0046] formula:

[0047] 3 parts by weight of irinotecan

[0048] 10 parts by weight of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles

[0049] Preparation method:

[0050] Irinotecan was dissolved in a sodium hydrogen phosphate-citric acid buffer solution at pH 5.0, and then mixed evenly with polyethylene glycol-b-polylysine-b-polyleucine triblock micelles. A mixed solvent of dimethyl sulfoxide and n-butanol (volume ratio 3:1) was added, and the mixture was fully shaken in a 2°C water bath and dialyzed to obtain polymer micelles containing irinotecan.

[0051] Example 4

[0052] formula:

[0053] 2 parts by weight of irinotecan

[0054] 20 parts by weight of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles

[0055] Preparation method:

[0056] Irinotecan was dissolved in a glycine-hydrochloric acid buffer solution at pH 4.0, and then mixed evenly with polyethylene glycol-b-polylysine-b-polyleucine triblock micelles. Dimethyl sulfoxide was added, and the mixture was fully shaken in a -5°C water bath and dialyzed to obtain polymer micelles containing irinotecan.

[0057] Example 5

[0058] formula:

[0059] 2 parts by weight of irinotecan

[0060] 30 parts by weight of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles

[0061] Preparation method:

[0062] Irinotecan was dissolved in a citric acid-sodium citrate buffer solution with a pH of 4.0 to 5.0, and then mixed evenly with polyethylene glycol-b-polylysine-b-polyleucine triblock micelles. N-butanol was added, and the mixture was fully shaken in a 5° C. water bath and dialyzed to obtain polymer micelles containing irinotecan.

[0063] Comparative Example 1

[0064] formula:

[0065] 1 part by weight of irinotecan

[0066] 3 parts by weight of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles

[0067] Preparation method:

[0068] Irinotecan and polyethylene glycol-b-polylysine-b-polyleucine triblock micelles were mixed evenly, and a mixed solvent of dimethyl sulfoxide and n-butanol (volume ratio 1:1) was added. The mixture was fully shaken in a 0°C water bath and dialyzed to obtain polymer micelles containing irinotecan.

[0069] Comparative Example 2

[0070] formula:

[0071] 1 part by weight of irinotecan

[0072] 3 parts by weight of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles

[0073] Preparation method:

[0074] Irinotecan was dissolved in water and then mixed evenly with polyethylene glycol-b-polylysine-b-polyleucine triblock micelles. A mixed solvent of dimethyl sulfoxide and n-butanol (volume ratio 1:1) was added, and the mixture was fully shaken in a 0°C water bath and dialyzed to obtain polymer micelles containing irinotecan.

[0075] Comparative Example 3

[0076] formula:

[0077] 1 part by weight of irinotecan

[0078] 3 parts by weight of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles

[0079] Preparation method:

[0080] Irinotecan was dissolved in a sodium hydrogen phosphate-citric acid buffer solution at pH 4.5, and then mixed evenly with polyethylene glycol-b-polylysine-b-polyleucine triblock micelles. A mixed solvent of dimethyl sulfoxide and n-butanol (volume ratio 1:1) was added, and the mixture was fully shaken in a 20°C water bath and dialyzed to obtain polymer micelles containing irinotecan.

[0081] Comparative Example 4

[0082] formula:

[0083] 1 part by weight of irinotecan

[0084] 3 parts by weight of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles

[0085] Preparation method:

[0086] Irinotecan was dissolved in a sodium hydrogen phosphate-citric acid buffer solution at pH 3.0, and then mixed evenly with polyethylene glycol-b-polylysine-b-polyleucine triblock micelles. A mixed solvent of dimethyl sulfoxide and n-butanol (volume ratio 1:1) was added, and the mixture was fully shaken in a 0°C water bath and dialyzed to obtain polymer micelles containing irinotecan.

[0087] 3. Quality evaluation of irinotecan-containing polymer micelles

[0088] 1. Average particle size of irinotecan-containing polymer micelles

[0089] Table 1 Average particle size of irinotecan-containing polymer micelles (nm)

[0090] Average particle size (nm) Example 1 79.64±4.32 Example 2 85.34±6.74 Example 3 88.21±5.92 Example 4 82.45±5.18 Example 5 84.03±6.25 Comparative Example 1 91.82±9.68 Comparative Example 2 95.71±7.50 Comparative Example 3 124.18±12.72 Comparative Example 4 94.25±8.04

[0091] The experimental data shown in Table 1 show that the particle size of the irinotecan-containing polymer micelles prepared in Examples 1 to 5 of the present invention is uniform. Figure 1 This is a scanning electron microscopy morphology display of the irinotecan-containing polymer micelles in Example 1, showing that the morphology is complete and the micelle outline is clear.

[0092] 2. Drug loading of irinotecan-containing polymer micelles

[0093] Irinotecan in the irinotecan-containing polymer micelles of Examples 1 to 5 and Comparative Examples 1 to 4 was separated from the micelle carrier, and the content of irinotecan in the irinotecan-containing polymer micelles was determined.

[0094] like Figure 3 、 4 As shown, the irinotecan-containing polymer micelles obtained in Examples 1 to 5 of the present invention have a large drug loading capacity, which can reach up to about 45%.

[0095] 3. Active ring closure rate of irinotecan in polymer micelles containing irinotecan

[0096] The irinotecan-containing polymer micelles of Examples 1 to 5 and Comparative Examples 1 to 4 were dissolved in an equal volume of phosphate buffer, stirred at room temperature until completely dissolved, and the pH was adjusted to 7.4. The drug active ring closure rate was determined by HPLC.

[0097] Figure 5 The results of the determination of the drug active ring-closure rate in the irinotecan-containing polymer micelles of Examples 1 to 5 and Comparative Examples 1 to 4 show that the drug active ring-closure rate of the irinotecan-containing polymer micelles obtained in the embodiments of the present invention can effectively avoid the ring-opening hydrolysis of the lactone ring in the irinotecan structure, effectively retain its anti-tumor activity, and avoid toxic side effects.

[0098] 4. Irinotecan preparations

[0099] Example 6 Irinotecan injection

[0100] The irinotecan-containing polymer micelles prepared in Example 1 and sorbitol are dissolved in an appropriate amount of water for injection, the pH is adjusted to 4.0-5.0 with a pH regulator, and water for injection is added to make up the volume. The finished product is obtained by filtering, filling, stoppering, capping, sterilizing and packaging.

[0101] It should be noted that the applicant also prepared irinotecan injection using the irinotecan-containing polymer micelles of Examples 2 to 5, and the stability and other effects thereof were similar to those of Example 6.

[0102] Comparative Example 5 Irinotecan Injection

[0103] The irinotecan-containing polymer micelles prepared in Comparative Example 1 and sorbitol were dissolved in an appropriate amount of water for injection, and the pH was adjusted to 4.0-5.0 with a pH regulator. Water for injection was added to make up the volume, and the finished product was obtained after filtration, filling, stoppering, capping, sterilization and packaging.

[0104] Comparative Example 6 Irinotecan Injection

[0105] The irinotecan-containing polymer micelles prepared in Comparative Example 3 and sorbitol were dissolved in an appropriate amount of water for injection, and the pH was adjusted to 4.0-5.0 with a pH regulator. Water for injection was added to make up the volume, and the finished product was obtained after filtration, filling, stoppering, capping, sterilization and packaging.

[0106] Commercially available irinotecan injection (H20030263)

[0107] 5. Stability Verification of Irinotecan Preparations

[0108] Accelerated tests were conducted on Example 6, Comparative Examples 5-6, and commercially available formulations. The formulations were packaged according to the commercially available formulations and placed at a temperature of 40±2°C and a relative humidity of 75%±5% for 6 months. Samples were taken at the end of the first, second, third, and sixth months of the test, and the content of related substances was determined by high performance liquid chromatography.

[0109] Figure 6 The results of the accelerated test on related substances of Example 6, Comparative Examples 5-6, and commercially available preparations show that the irinotecan injection provided by the present invention has low content of related substances and high stability.

Claims

1. A polymer micelle containing irinotecan, characterized in that The irinotecan-containing polymer micelles are composed of 1 to 3 parts of irinotecan and 6 to 30 parts of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles by weight. The preparation method of the irinotecan-containing polymer micelles comprises: dissolving irinotecan in a buffer solution with a pH value of 4.0 to 5.0, uniformly mixing the irinotecan with the polyethylene glycol-b-polylysine-b-polyleucine triblock micelles, adding an organic solvent, fully shaking the mixture in a water bath at -5°C to 5°C, and dialyzing the mixture to obtain the irinotecan-containing polymer micelles; the organic solvent is selected from at least one of methanol, ethanol, chloroform, isopropanol, n-butanol, and dimethyl sulfoxide.

2. The irinotecan-containing polymer micelle according to claim 1, characterized in that Calculated by weight, the irinotecan-containing polymer micelles consist of 2 parts of irinotecan and 7 parts of polyethylene glycol-b-polylysine-b-polyleucine triblock micelles.

3. The irinotecan-containing polymer micelle according to claim 1, wherein The organic solvent is a mixed solvent of dimethyl sulfoxide and n-butanol.

4. The irinotecan-containing polymer micelle according to claim 1, characterized in that The temperature of the water bath is -2~2°C.

5. The irinotecan-containing polymer micelle according to claim 1, characterized in that The buffer solution is selected from one of disodium hydrogen phosphate-citric acid buffer solution, glycine-hydrochloric acid buffer solution, and citric acid-sodium citrate buffer solution.

6. The irinotecan-containing polymer micelle according to claim 5, characterized in that The buffer solution is a disodium hydrogen phosphate-citric acid buffer solution.

7. The irinotecan-containing polymer micelle according to claim 1, characterized in that The pH of the buffer solution is 4.

5.

8. An irinotecan preparation, characterized in that The invention comprises the irinotecan-containing polymer micelles according to claim 1 and pharmaceutically acceptable excipients.

9. The preparation according to claim 8, characterized in that The preparation is an injection.

10. Use of the irinotecan-containing polymer micelle according to claim 1 in preparing a drug for treating tumors.

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