Pharmaceutical composition containing Bremelantin, preparation and preparation method and application of pharmaceutical composition containing Bremelantin and preparation method and application of pharmaceutical composition containing Bremelantin

Abstract

The invention belongs to the technical field of pharmaceutical preparations, and particularly relates to a pharmaceutical composition containing Bremelinodan, a preparation as well as a preparation method and application of the pharmaceutical composition. The pharmaceutical composition provided by the invention comprises a pharmaceutical active ingredient Bramer nodan, a solvent and a polymer, and the solvent comprises one or more of N-methyl pyrrolidone, polyethylene glycol, dimethyl sulfoxide and benzyl benzoate. According to the pharmaceutical composition containing the Bramer captan, the blood concentration of the Bramer captan is stably larger than 0.1 ng / mL within 120-168 h, target slow release is achieved, direct injection can be achieved, liquid medicine does not need to be mixed before use, the preparation process is simple, the component is single, a thickening agent, a gel forming agent or other auxiliary materials do not need to be added, large-scale production can be achieved, cost is low, and the pharmaceutical composition is suitable for industrial production. Good market application prospects are realized.

Description

Technical Field

[0001] The present invention belongs to the technical field of pharmaceutical preparations, and in particular relates to a pharmaceutical composition containing bremelanotide, a preparation, and a preparation method and application thereof. Background Art

[0002] Bremelanotide, also known as Bremelanotide PT141, Bremelanotide impurity, Bremelanotide and Bremelanotide, is a cyclic heptapeptide compound with a molecular formula of C 50 H 68 N 14 O 10 , whose chemical structure is shown in Formula 1:

[0003]

[0004] Bremelanotide, as a ligand of one or more melanocortin receptors, can bind well to melanocortin receptors and produce good effects. Its pharmacological characteristics are that the drug has the potential to treat or prevent acute or chronic inflammatory diseases, autoimmune diseases, transplant rejection, metabolic diseases accompanied by weight gain, metabolic diseases accompanied by weight loss, diabetes, diabetic complications, cancer, cancerous hyperplasia, reproductive system diseases, peripheral or central nervous system diseases, cardiovascular diseases or respiratory system diseases.

[0005] Chinese invention patent application 202111481324.8 discloses a method for synthesizing bremelanotide acetate, specifically: using the Fmoc solid phase peptide synthesis method, with RinkAmide resin as a carrier, after modifying it with 4-trifluoromethylmandelic acid, according to the polypeptide sequence of bremelanotide, under the action of a coupling agent, amino acids with Fmoc-protecting groups are sequentially condensed from the C-terminus to the N-terminus to obtain a linear peptide resin; then the Fmoc protecting group and the side chain protecting agents of Lys, Asp, etc. are removed in turn, and then a ring is formed on the resin under the action of a coupling agent, the resin is cleaved with a cleavage agent, and the ester bond is disconnected under alkaline conditions to obtain a crude bremelanotide peptide, and finally purified to obtain a refined bremelanotide peptide. The method is simple, has a high yield, and has high purification efficiency.

[0006] Through the study of the original bremelanotide drug-device combination on the market, it was found that the original bremelanotide product VYLEESI (AUTOINJECTOR), as a subcutaneous drug, has a fast peak blood concentration of 0.5-1.0h, 100% bioavailability, a half-life of 1.9-4.0h, and an average clearance of 6.5±1.0L / hr, which is insufficient in potential indications requiring long-term administration. Referring to the existing sustained-release preparations on the market, these preparations often need to be mixed before use, a step that brings additional inconvenience to patients and possible medication errors.

[0007] Based on the limitations of the existing technology, there is an urgent need in the art to provide a long-acting sustained-release preparation of bremelanotide that can meet the needs of long-term administration through sustained-release technology. Summary of the invention

[0008] In view of the problems existing in the prior art, the present invention provides a pharmaceutical composition and preparation containing bremelanotide and a preparation method and application thereof.

[0009] To achieve the above purpose, the technical solution adopted by the present invention is as follows:

[0010] A pharmaceutical composition comprises bremelanotide, a solvent and a polymer, wherein the solvent comprises one or more of N-methylpyrrolidone, polyethylene glycol, dimethyl sulfoxide and benzyl benzoate.

[0011] Preferably, the solvent is dimethyl sulfoxide, a mixed solution of N-methylpyrrolidone and benzyl benzoate, a mixed solution of dimethyl sulfoxide and benzyl benzoate, or a mixed solution of N-methylpyrrolidone and dimethyl sulfoxide.

[0012] Further preferably, the solvent is a mixed solution of N-methylpyrrolidone and benzyl benzoate, and N-methylpyrrolidone accounts for 5-95% of the volume of the mixed solution.

[0013] More preferably, the solvent is a mixed solution of N-methylpyrrolidone and benzyl benzoate, and N-methylpyrrolidone accounts for 95% of the volume of the mixed solution.

[0014] Further preferably, the solvent is a mixed solution of dimethyl sulfoxide and benzyl benzoate, and dimethyl sulfoxide accounts for 5-95% of the volume of the mixed solution.

[0015] More preferably, the solvent is a mixed solution of dimethyl sulfoxide and benzyl benzoate, and dimethyl sulfoxide accounts for 95% of the volume of the mixed solution.

[0016] Further preferably, the solvent is a mixed solution of N-methylpyrrolidone and dimethyl sulfoxide, and N-methylpyrrolidone accounts for 5-95% of the volume of the mixed solution.

[0017] Preferably, the polymer is a lactide-glycolide copolymer, carbomer or sodium alginate.

[0018] More preferably, the polymer is a lactide-glycolide copolymer.

[0019] Preferably, the end-capping structure of the lactide-glycolide copolymer is carboxyl-capping or ester-capping.

[0020] Preferably, the molar ratio of lactic acid monomer to glycolic acid monomer in the lactide-glycolide copolymer is 50-85:15-50.

[0021] More preferably, the molar ratio of lactic acid monomer to glycolic acid monomer in the lactide-glycolide copolymer is 50-75:20-50.

[0022] More preferably, the molar ratio of lactic acid monomer to glycolic acid monomer in the lactide-glycolide copolymer is 50:50 or 75:25.

[0023] Preferably, the average molecular weight of the lactide-glycolide copolymer is 20-50 kDa.

[0024] More preferably, the average molecular weight of the lactide-glycolide copolymer is 20-25 kDa.

[0025] More preferably, the average molecular weight of the lactide-glycolide copolymer is 23 kDa.

[0026] Preferably, the pharmaceutical composition comprises, by weight percentage, 0.1-15% of bremelanotide, 10-70% of polymer and the solvent making up to 100%.

[0027] Further preferably, the pharmaceutical composition comprises, by weight percentage, 1-5% of bremelanotide, 30-50% of polymer and the solvent making up to 100%.

[0028] More preferably, the pharmaceutical composition comprises, by weight percentage, 1-1.5% of bremelanotide, 35-40% of polymer and the solvent makes up to 100%.

[0029] The present invention also provides a method for preparing the above-mentioned pharmaceutical composition, which comprises directly mixing bremelanotide, a solvent and a polymer to obtain the pharmaceutical composition.

[0030] The present invention also provides a pharmaceutical preparation, comprising the above pharmaceutical composition or the pharmaceutical composition prepared by the above preparation method.

[0031] Preferably, the preparation is of the type of injection.

[0032] The present invention also provides the use of the above-mentioned pharmaceutical composition or the pharmaceutical composition prepared by the above-mentioned preparation method or the above-mentioned pharmaceutical preparation in the preparation of sustained-release bremelanotide drugs.

[0033] Compared with the prior art, the present invention has the following beneficial effects:

[0034] (1) The pharmaceutical composition containing bremelanotide provided by the present invention can stably maintain the blood concentration of bremelanotide above 0.1 ng / mL during 120-168 hours, thus achieving the target sustained release.

[0035] (2) The bremelanotide-containing pharmaceutical composition prepared by the present invention can be directly injected without mixing the drug solution before use.

[0036] (3) The pharmaceutical composition containing bremelanotide provided by the present invention has a simple preparation process and a single component. It can be mass-produced without adding a thickener, a gel-forming agent or other excipients, has low cost and has good market application prospects. DETAILED DESCRIPTION

[0037] The present invention is further described below in conjunction with specific examples, and the advantages and features of the present invention will become clearer as the description proceeds. However, these examples are merely exemplary and do not constitute any limitation to the scope of the present invention. The methods, ingredients and amounts involved in the following examples are all conventional methods known to those skilled in the art unless otherwise specified.

[0038] It is worth noting that the raw materials used in the present invention are all common commercially available products, and the sources of some of the raw materials are shown in Table 1.

[0039] Table 1 Raw material source information

[0040]

[0041]

[0042] 1. The abbreviations used in the present invention have the following meanings:

[0043] API: essential pharmaceutical ingredient;

[0044] NMP: N-methyl-2-pyrrolidone;

[0045] DMSO: dimethyl sulfoxide;

[0046] PLGA: poly(lactide-co-glycolide);

[0047] PBS: phosphate buffered saline;

[0048] BA: benzyl alcohol;

[0049] BB: benzyl benzoate.

[0050] 2. The specific method of the in vitro release experiment of the present invention is as follows:

[0051] Dissolution method: Add an appropriate amount of prepared drug gel, weighing about 20 mg, into the medium and place it in a 37°C water bath;

[0052] Dissolution medium: PBS at pH 7.4 and a concentration of 20 mg / mL;

[0053] Medium volume: 10 mL;

[0054] Sampling method: Take 1 mL each time and add 1 mL of fresh medium;

[0055] Sampling points: Samples were taken at 0.5h and 24h for determination of API bremelanotide content.

[0056] 3. The content of API in the present invention is detected by HPLC, and the specific method is as follows:

[0057] 3.1 Solution preparation

[0058] 3.1.1 Diluent: Weigh about 1g of bovine serum albumin into a blue-capped bottle, add 800mL of purified water to dissolve, add 200mL of acetonitrile, add 1mL of trifluoroacetic acid, shake well, and you have it.

[0059] 3.1.2 Preparation of mobile phase: Measure 700 mL of purified water, put it into a blue-capped bottle, add 300 mL of acetonitrile, mix well, add 1 mL of trifluoroacetic acid, mix well, filter with a 0.45 μm filter membrane, and degas by ultrasonication.

[0060] 3.1.3 Preparation of control solution: Take about 10 mg of API bremelanotide, accurately weigh it, put it into a 20 mL volumetric flask, add diluent to dissolve and dilute to the scale, shake well; measure 1.0 mL of the above solution into a 20 mL volumetric flask, add diluent to dilute to the scale, shake well.

[0061] 3.1.4 Preparation of test sample solution: Take about 50 mg of the test sample, accurately weigh it, put it into a 20 mL volumetric flask, add 4 mL of DMSO and dissolve it by ultrasonic, add diluent and dilute to the scale, shake well, take an appropriate amount into a centrifuge tube, centrifuge it at 8000 rpm for 10 min, and take the supernatant.

[0062] Chromatographic conditions

[0063] Instrument: High performance liquid chromatography;

[0064] Chromatographic column: Waters Xbridge C18, 150 mm × 4.6 mm × 3.5 μm;

[0065] Mobile phase: acetonitrile: water = 30:70 (v:v, containing 0.1% trifluoroacetic acid);

[0066] Column temperature: 35°C;

[0067] Flow rate: 1.0 mL / min;

[0068] Detection wavelength: 215nm;

[0069] Injection volume: 20 μL;

[0070] Running time: 10 minutes;

[0071] Elution mode: isocratic elution.

[0072] Example 1

[0073] The composition of a pharmaceutical composition containing bremelanotide is shown in Table 2 below.

[0074] Table 2 Prescription

[0075]

[0076] A method for preparing a pharmaceutical composition containing bremelanotide: accurately weighing a prescribed amount of bremelanotide, PLGA and DMSO and placing them in a sample bottle, and then rolling the sample bottle in a roller mixer until bremelanotide and PLGA are completely dissolved to obtain the pharmaceutical composition.

[0077] Example 2

[0078] The composition of a pharmaceutical composition containing bremelanotide is shown in Table 3 below.

[0079] Table 3 Prescription

[0080]

[0081] A method for preparing a pharmaceutical composition containing bremelanotide: accurately weighing a prescribed amount of bremelanotide, PLGA, DMSO and BB and placing them in a sample bottle, and then rolling the sample bottle in a roller mixer until bremelanotide and PLGA are completely dissolved to obtain the pharmaceutical composition.

[0082] Experimental Example 1 In vitro release experiment

[0083] 1. Solubility of API bremelanotide

[0084] In order to select a suitable solvent and minimize the amount of API used while meeting the drug concentration requirements in the preparation, the concentration of the drug bremelanotide was set to 40 mg / mL, and the saturated solubility (mg / mL) of bremelanotide in 5 mL of different aqueous or organic solvents was investigated, wherein the aqueous solvent was water or PBS with a pH of 7.4 and a concentration of 20 mg / mL, and the organic solvent was NMP, DMSO, a mixed solution of NMP and BB with a volume ratio of 1:0.05, or a mixed solution of NMP and BA with a volume ratio of 1:0.05. The test results are shown in Table 4.

[0085] As shown in Table 4, the saturated solubility of bremelanotide in pH=7.4 PBS is relatively low.

[0086] Table 4 Saturated solubility results of API in different solvents

[0087]

[0088]

[0089] 2. Prescription screening

[0090] Since the API bremelanotide has good water solubility, it is quite difficult to develop a sustained-release preparation. Therefore, during the development process, priority is given to trying the feasibility of the extreme prescription, that is, taking the in vitro release rate as the evaluation index, focusing on screening the average molecular weight of PLGA, the end-capping structure of PLGA, the molar ratio of lactic acid monomers to glycolic acid monomers in PLGA, and the solvent system.

[0091] 2.1 Average molecular weight of PLGA

[0092] PLGA with carboxyl end-capping, a molar ratio of lactic acid monomer to glycolic acid monomer of 50:50 and different average molecular weights (20, 23, 25 and 40 kDa) were selected, and the solvents were NMP or DMSO, respectively.

[0093] By weight percentage, 1.1% of bremelanotide, 36% of PLGA and 62.9% of solvent were mixed in a roller mixer to prepare a pharmaceutical composition containing bremelanotide. The in vitro release rate was used as an investigation index to screen the molecular weight of PLGA. The results are shown in Table 5.

[0094] As can be seen from Table 5, the in vitro release rates of the drug compositions prepared from PLGA with average molecular weights of 20, 23 and 25 kDa are relatively low, and the combinations of solvents and PLGA average molecular weights shown can all be used for the formulation of bremelanotide compositions.

[0095] Table 5 In vitro release test results of drug compositions prepared with different PLGA average molecular weights

[0096]

[0097] 2.2 PLGA capping structure

[0098] PLGA with a molar ratio of 50:50 between lactic acid monomer and glycolic acid monomer, an average molecular weight of 40 kDa and different end-capping structures (carboxyl end-capping and ester end-capping) was selected, and the solvent was NMP.

[0099] By weight percentage, 1.1% of bremelanotide, 36% of PLGA and 62.9% of solvent were mixed in a roller mixer to prepare a pharmaceutical composition containing bremelanotide. The end-capping structure of PLGA was screened with the in vitro release rate as the evaluation index. The results are shown in Table 6.

[0100] As shown in Table 6, both carboxyl-terminated and ester-terminated PLGA can be used in the formulation of bremelanotide composition, among which the in vitro release rate of the drug composition prepared by ester-terminated PLGA is lower.

[0101] Table 6 In vitro release test results of drug compositions prepared with different PLGA end-capping structures

[0102]

[0103] 2.3 Molar ratio of lactic acid monomer to glycolic acid monomer in PLGA

[0104] PLGA with carboxyl end-capping, average molecular weight of 23 kDa and different molar ratios of lactic acid monomer and glycolic acid monomer was selected, and the solvent was NMP.

[0105] By weight percentage, 1.2% of bremelanotide, 39.5% of PLGA and 59.3% of solvent were placed in a roller mixer and mixed to prepare a pharmaceutical composition containing bremelanotide. The molar ratio of lactic acid monomer to glycolic acid monomer in PLGA was screened with the in vitro release rate as the evaluation index. The results are shown in Table 7.

[0106] As shown in Table 7, the molar ratio of lactic acid monomer to glycolic acid monomer in PLGA is 75:25 or 50:50, which can be used for the formulation of bremelanotide composition. Among them, the in vitro release rate of the drug composition prepared by PLGA with a molar ratio of lactic acid monomer to glycolic acid monomer of 50:50 is lower.

[0107] Table 7 In vitro release test results of the pharmaceutical compositions prepared by PLGA with different molar ratios of lactic acid monomer and glycolic acid monomer

[0108]

[0109] 2.4 Solvent system

[0110] PLGA with carboxyl end-capping, average molecular weight of 23 kDa and molar ratio of lactic acid monomer to glycolic acid monomer of 50:50 was selected, and the solvents were NMP, DMSO, a mixed solution of NMP and BB with a volume ratio of 0.95:0.05, and a mixed solution of DMSO and BB with a volume ratio of 0.95:0.05.

[0111] By weight percentage, in the NMP system, 1.2% of bremelanotide, 39.5% of PLGA and 59.3% of solvent; in the DMSO system, 1.4% of bremelanotide, 39.4% of PLGA and 59.2% of solvent, the above materials are respectively placed in a roller mixer and mixed to prepare a pharmaceutical composition containing bremelanotide;

[0112] The solvents were screened using the in vitro release rate as the evaluation index. The results are shown in Table 8.

[0113] As can be seen from Table 8, all the solvents in the table can be used for the formulation of the bremelanotide composition, among which the in vitro release rate of the pharmaceutical composition prepared by the solvent DMSO, a mixed solution of NMP and BB in a volume ratio of 0.95:0.05, or a mixed solution of DMSO and BB in a volume ratio of 0.95:0.05 is low.

[0114] Table 8 In vitro release test results of pharmaceutical compositions prepared with different solvents

[0115]

[0116] 3. Prescription design

[0117] Combining the above PLGA and solvent screening, four formulations with lower in vitro release rates were obtained. The specific information is shown in Table 9. Two of the formulations were selected as Examples 1 and 2.

[0118] Table 9 Prescription of the pharmaceutical composition containing bremelanotide

[0119]

[0120] Experimental Example 2 Pharmacokinetic (PK) experiment in rats

[0121] Experimental animals: 10 SD rats, SPF grade male, weighing 210-240 g, purchased from Guangzhou Ruige Biotechnology Co., Ltd., production license number SYXK (Guangdong) 2022-0296, adapted to the environment for 3-5 days, free access to food and water, feed Co 60 The irradiated experimental mice were maintained on feed. The animal room environment was maintained at a temperature of 20-26°C, a humidity of 40-70%, 12h light and dark alternation, 10-20 times / h ventilation, and no more than 3 mice were kept in each cage.

[0122] Experimental method: 10 SD rats were randomly divided into 3 groups. Group 1 (n=4) was subcutaneously injected with 3 mg / kg of the pharmaceutical composition containing bremelanotide prepared in Example 1; Group 2 (n=3) was subcutaneously injected with 6 mg / kg of the pharmaceutical composition containing bremelanotide prepared in Example 1; Group 3 (n=3) was subcutaneously injected with 6 mg / kg of the pharmaceutical composition containing bremelanotide prepared in Example 2. Pharmacokinetic blood samples were collected before administration (0 h) and at 0.5, 1, 1.5, 2, 3, 4, 6, 12, 24, 48, 72, 96, 120 and 168 h after administration, and serum was separated to detect plasma drug concentration.

[0123] Experimental results and analysis: The experimental results are shown in Table 10. Seven days after administration (i.e., 168 hours), no obvious abnormalities were found in the administration sites of the three groups of animals. From the results of the PK test, it can be seen that the drug was released 24 hours after administration. The plasma drug concentrations of Group 1 and Group 2, which were administered with doses of 3 mg / kg and 6 mg / kg, were higher than 0.1 ng / mL at 120 hours after administration; the plasma drug concentrations of Group 2 and Group 3, which were administered with a dose of 6 mg / kg, were higher than 0.1 ng / mL at 168 hours after administration.

[0124] Table 10 Rat PK experimental results

[0125]

[0126] Note: BQL means below limit of quantitation.

[0127] The pharmacokinetic properties of the pharmaceutical composition containing bremelanotide prepared in Examples 1-2 of the present invention were further studied.

[0128] The PK experimental data of Example 1-2 were compared with the PK data of the original product VYLEESI (AUTOINJECTOR) in the FDA review report, and the pharmacokinetic data are shown in Table 11.

[0129] VYLEESI: Each 0.3 mL prefilled syringe contains 1.75 mg of bremelalotide (equivalent to 1.89 mg of bremelalotide acetate) and inactive ingredients include 2.5% glycerol and sterile water for injection. Hydrochloric acid or sodium hydroxide is used to adjust the pH.

[0130] As shown in Table 11, the pharmaceutical compositions containing bremelanotide prepared in Examples 1-2 all showed good sustained-release effects and effectively reduced C max , T max Moved back and greatly extended T 1 / 2 , effectively achieving the sustained-release target.

[0131] Table 11 Pharmacokinetic data

[0132]

[0133] Note: * Indicates that the VYLEESI data comes from the FDA review report of the original product.

[0134] Finally, it should be noted that the above content is only used to illustrate the technical solution of the present invention, rather than to limit the scope of protection of the present invention. Simple modifications or equivalent substitutions of the technical solution of the present invention by ordinary technicians in this field do not deviate from the essence and scope of the technical solution of the present invention.

Claims

1. A pharmaceutical composition, characterized in that The invention comprises a pharmaceutically active ingredient, bremelanotide, a solvent and a polymer, wherein the solvent comprises one or more of N-methylpyrrolidone, polyethylene glycol, dimethyl sulfoxide and benzyl benzoate.

2. The pharmaceutical composition according to claim 1, characterized in that The solvent is a mixed solution of dimethyl sulfoxide, N-methyl pyrrolidone and benzyl benzoate, a mixed solution of dimethyl sulfoxide and benzyl benzoate, or a mixed solution of N-methyl pyrrolidone and dimethyl sulfoxide.

3. The pharmaceutical composition according to claim 1, characterized in that The polymer is a lactide-glycolide copolymer, carbomer or sodium alginate.

4. The pharmaceutical composition according to claim 3, characterized in that The polymer is a lactide-glycolide copolymer, and the end-capping structure of the lactide-glycolide copolymer is a carboxyl end-capping structure or an ester end-capping structure.

5. The pharmaceutical composition according to claim 4, characterized in that The molar ratio of the lactic acid monomer to the glycolic acid monomer in the lactide-glycolide copolymer is 50-85:15-50.

6. The pharmaceutical composition according to claim 4, characterized in that The average molecular weight of the lactide-glycolide copolymer is 20-50 kDa.

7. The pharmaceutical composition according to any one of claims 1 to 6, characterized in that In terms of weight percentage, the pharmaceutical composition comprises 0.1-15% of bremelanotide, 10-70% of a polymer and a solvent to make up to 100%.

8. A method for preparing the pharmaceutical composition according to any one of claims 1 to 7, characterized in that: The method comprises directly mixing the active pharmaceutical ingredient bremelanotide, a solvent and a polymer to obtain the product.

9. A pharmaceutical preparation, characterized in that A pharmaceutical composition comprising any one of claims 1 to 7 or a pharmaceutical composition prepared by the preparation method of claim 8, wherein the pharmaceutical preparation is an injection.

10. Use of the pharmaceutical composition according to any one of claims 1 to 7, the pharmaceutical composition prepared by the preparation method according to claim 8, or the pharmaceutical preparation according to claim 9 in the preparation of sustained-release bremelanotide drugs.

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