Progesterone sustained-release delivery system

Abstract

Provided are progesterone sustained-release microspheres and a formulation thereof. Compared to progesterone injections on the market, the formulation, in the case of non-inferior bioavailability, reduces the dosing frequency from one or more injections daily to once weekly. During the administration cycle, the formulation can maintain a stable plasma concentration, avoiding peak-trough fluctuations in the drug concentration, which effectively improves the efficacy of progesterone supplementation therapy while reducing adverse reactions.

Description

A progesterone sustained-release delivery system

[0001] This application claims priority to the following prior patent applications:

[0002] The applicant filed an earlier application with the China National Intellectual Property Administration on March 26, 2025, with patent application number 202510368397.8 and title "A Progesterone Sustained-Release Delivery System";

[0003] The full text of the aforementioned prior patent applications is incorporated herein by reference. Technical Field

[0004] This invention relates to a progesterone sustained-release delivery system, specifically to an injectable microsphere and a microsphere formulation for sustained progesterone release. More specifically, this invention relates to progesterone microspheres and their formulations. Background Technology

[0005] In the field of assisted reproduction, progesterone plays a crucial role. Insufficient progesterone can lead to poor endometrial receptivity, resulting in embryo implantation failure, or even if implantation is successful, miscarriage may occur due to uterine instability. Therefore, in assisted reproduction, appropriate progesterone supplementation is an important means to improve success rates and ensure a smooth pregnancy.

[0006] Among the various existing progesterone supplementation regimens, there are still many unmet clinical needs, specifically: ① Oral route: Although convenient, progesterone has poor water solubility and low oral bioavailability (<10%). Furthermore, the metabolites produced by the first-pass effect in the liver can act on γ-aminobutyric acid (GABA) receptors in the central nervous system, enhancing the inhibitory effect of GABA and thus producing adverse reactions such as sedation and drowsiness. Clinically, oral administration is rarely used for progesterone supplementation in assisted reproduction. ② Injection route: Clinically, the main method is to administer an oil solution formed by dissolving progesterone in injectable vegetable oils such as castor oil or soybean oil via intramuscular injection. A single dose can maintain efficacy for one day, but because the oily injection solution is not easily absorbed, post-injection complications may occur. It causes local muscle inflammation, intense pain during injection, and requires frequent daily injections. There are significant peaks and troughs in blood drug concentration, resulting in numerous adverse reactions, including pain, induration, and cysts, leading to low patient compliance. Besides the oil-based solution, there is also an aqueous progesterone injection using hydroxypropyl betacyclodextrin, but this also requires daily injection, resulting in similarly poor patient compliance. Other routes include gels or suppositories, administered vaginally 1-2 times daily. Vaginal administration can cause local irritation, bleeding, and pain, leading to poor compliance. Drug leakage and other issues can also result in inaccurate dosage. Furthermore, local absorption makes it impossible to assess efficacy through systemic blood drug concentration, hindering dosage control by physicians.

[0007] To improve patient compliance, it is also necessary to reduce the frequency of administration. Currently, progesterone oil solutions or aqueous solutions for injection require once-daily administration for several weeks. Reducing the frequency to once-weekly administration can better balance compliance and the flexibility of physician dosage adjustment. The dosage of weekly formulations is significantly higher than that of daily formulations, requiring formulations to increase drug loading to reduce injection volume. Currently reported progesterone extended-release formulations generally have a drug loading of no more than 30% [①Pharm Res 34,2787–2797(2017); ②Pharm Res 35,62(2018); ③International Journal of Pharmaceutics 607(2021)121021; ④AAPS PharmSciTech(2022)23:294; ⑤J Microencapsul,2015;32(6):538], which suggests that the required intramuscular or subcutaneous injection volume exceeds 5 mL, which is difficult to achieve in clinical practice and will also cause significant adverse reactions such as injection-related pain.

[0008] In summary, to address the pain points of existing clinical treatment regimens and improve the efficacy, safety, and adherence of progesterone supplementation drugs, the developed progesterone extended-release formulations must meet the following requirements: ① Relative bioavailability calculated based on AUC is not inferior to existing injectable formulations (marketed intramuscular progesterone oil solutions or subcutaneous progesterone aqueous solutions); ② Under the premise of meeting AUC, C max It should be as low as possible; ③ The peak-valley phenomenon should be as small as possible, i.e., C max / C min The ratio should be as small as possible; ④ It is appropriate to set the dosing frequency to once a week. Summary of the Invention

[0009] To address the problems existing in the prior art, the present invention provides progesterone sustained-release microspheres and a formulation comprising the sustained-release microspheres. Specifically, the implementation scheme is shown below.

[0010] On one hand, the present invention provides progesterone sustained-release microspheres, wherein the raw materials for making the progesterone sustained-release microspheres or the components of the progesterone sustained-release microspheres include:

[0011] (1) Progesterone as an active substance; and

[0012] (2) One, two or more sustained-release polymers.

[0013] According to an embodiment of the present invention, the mass ratio of the sustained-release polymer to progesterone is 1:10-10:1, preferably 1:8-8:1, 1:5-5:1, for example 1:5, 1:4, 1:3, 1:2, 0.9:1, 0.92:1, 0.94:1, 0.96:1, 0.98:1, 1:1, 2:1, 3:1, 4:1 or 5:1.

[0014] According to an embodiment of the present invention, the progesterone sustained-release microspheres are suitable for injection administration, such as subcutaneous injection, intravenous injection or intramuscular injection; preferably intramuscular injection.

[0015] According to an embodiment of the present invention, the sustained-release polymer is selected from any one, two or more of the following: lactide-glycolic acid copolymer (PLGA), polylactide (PLA), polycaprolactone (PCL), lactide-caprolactone copolymer (PLC), lactide-glycolic acid-caprolactone copolymer (PLGC), polyorthoester (POE), and polyhydroxyalkanoate (PHA).

[0016] According to embodiments of the present invention, the L / G (molar ratio of lactide to glycolide) of the lactide-glycol copolymer is 85:15-40:60, 85:15-30:70; preferably 80:20-40:60, 60:40-40:60, for example 75:25, 70:30, 65:35, 60:40, 55:45, 50:50, 45:55, 40:60, 35:65, 30:70.

[0017] According to an embodiment of the present invention, the molecular weight of the sustained-release polymer is in the range of about 4 kDa to about 17 kDa, for example, about 7 kDa to about 17 kDa or about 4 kDa to about 15 kDa; preferably, the molecular weight is about 10 kDa to 17 kDa; for example, the molecular weight is about 5 kDa, 6 kDa, 7 kDa, 8 kDa, 9 kDa, 10 kDa, 11 kDa, 12 kDa, 13 kDa, 14 kDa, 15 kDa, 16 kDa or 17 kDa.

[0018] According to an embodiment of the present invention, the chain end of the sustained-release polymer is capped with a carboxyl group or an ester group; preferably, it is capped with a carboxyl group.

[0019] According to embodiments of the present invention, the raw materials for making the progesterone sustained-release microspheres or the components of the progesterone sustained-release microspheres optionally further include one, two or more amphiphilic polymers.

[0020] According to embodiments of the present invention, the amphiphilic polymer is selected from block polymers containing polyester and / or polyether, and / or other amphiphilic polymers. Preferably, the amphiphilic polymer is selected from polyoxyethylene-polyoxypropylene block copolymer (PEO-PPO-PEO, commonly known as poloxamer), polyoxyethylene-polysiloxane block copolymer, polyethylene glycol-long-chain fatty acid, polyethylene glycol-polyester block copolymer (wherein the polyester can be a polymer formed by polymerizing / copolymerizing one or more monomers such as lactide, glycolide, and caprolactone). More preferably, the amphiphilic polymer is selected from one, two, or more of poloxamer, polyethylene glycol-polylactide (PEG-PLA), and polyethylene glycol-lactide-glycolic acid copolymer (PEG-PLGA).

[0021] According to an exemplary embodiment of the present invention, the amphiphilic polymer is selected from poloxamer.

[0022] According to an exemplary embodiment of the present invention, the amphiphilic polymer is selected from poloxamer F68 and poloxamer F127.

[0023] According to an exemplary embodiment of the present invention, the amphiphilic polymer is selected from PEG-PLA, such as PEG(2k~10k)-PLA(1k~30k) and PEG(5k~6k)-PLA(1k~15k), with PEG(5k)-PLA(15k) and PEG(6k)-PLA(1k) being exemplary examples.

[0024] According to embodiments of the present invention, the weight ratio of the sustained-release polymer (e.g., PLGA) to the amphiphilic polymer is 100:0-80:20. Preferably, the weight ratio is 99.8:0.2-90:10. For example, 99.5:0.5, 99:1, 98.5:1.5, 98:2, 97.5:2.5, 97:3, 96.5:3.5, 96:4, 95.5:4.5, 95:5, 94.5:5.5, 94:6, 93.5:6.5, 93:7, 92.5:7.5, 92:8, 91.5:8.5, 91:9, 90.5:9.5, 90:10.

[0025] According to an embodiment of the present invention, the progesterone sustained-release microspheres have the following particle size distribution:

[0026] D10 is approximately 5μm-20μm, preferably approximately 5μm-15μm; for example, approximately 6.0μm, 7.0μm, 8.0μm, 9.0μm, 10.0μm, 10.1μm, 10.3μm, 10.6μm, 10.9μm, 11.0μm, 11.4μm, 11.6μm, 12.0μm, 13.0μm, and 14.0μm.

[0027] And / or, D50 is about 15μm-35μm, preferably about 20μm-30μm; for example, about 21.0μm, 21.5μm, 22.0μm, 23.0μm, 23.4μm, 23.7μm, 24.0μm, 25.0μm, 25.1μm, 25.3μm, 26.0μm, 26.8μm, 27.0μm, 28.0μm, 29.0μm;

[0028] And / or, D90 is about 30μm-70μm, preferably about 35μm-65μm; for example, 36.0μm, 37.0μm, 37.4μm, 37.7μm, 38.0μm, 38.1μm, 39.0μm, 40.0μm, 41.0μm, 42.0μm, 43.0μm, 43.4μm, 44.0μm, 45.0μm, 46.0μm, 47.0μm, 48.0μm, 48.9μm, 49.0μm, 49.2μm, 50.0μm, 50.5μm, 51.0μm, 52.0μm, 52.2μm, 53.0μm, 54.0μm, 55.0μm, 58.0μm, 60.0μm.

[0029] According to an embodiment of the present invention, the progesterone sustained-release microspheres, with the particle size distribution described above, exhibit significantly improved in vivo bioavailability.

[0030] According to an embodiment of the present invention, the progesterone sustained-release microspheres can stably release most of the encapsulated progesterone within 7 days, consistent with the target dosing cycle (7 days). According to an embodiment of the present invention, the progesterone sustained-release microspheres are administered once a week.

[0031] According to an embodiment of the present invention, the drug loading of the progesterone sustained-release microspheres is about 30%-60%; preferably, the drug loading is 40%-55%; for example, 41.0%, 42.0%, 43.0%, 44.0%, 45.0%, 46.0%, 46.5%, 46.9%, 47.0%, 47.7%, 48.0%, 48.1%, 48.6%, 49.0%, 49.1%, 50.0%, 50.5%, 51.0%, 51.5%, 52.0%, and 53.0%. The drug loading refers to the mass ratio of the active molecule, i.e., progesterone, to the microspheres.

[0032] According to embodiments of the present invention, the encapsulation efficiency of the progesterone sustained-release microspheres is about 80.0%-99.9%; preferably, the encapsulation efficiency is 92.0%-99.8%; for example, about 81.0%, 82.0%, 83.0%, 84.0%, 85.0%, 86.0%, 87.0%, 88.0%, 89.0%, 90.0%, 91.0%, 92.0%, 93.0%, 93.8%, 94.0%, 95.0%, 95.4%, 96.0%, 96.2%, 97.0%, 97.2%, 98.0%, 98.2%, 99.0%, and 99.5%.

[0033] According to an embodiment of the present invention, the raw materials for making the progesterone sustained-release microspheres or the components of the progesterone sustained-release microspheres include: progesterone as an active substance, a sustained-release polymer, and an amphiphilic polymer.

[0034] According to embodiments of the present invention, the amount of progesterone used, based on the total weight of the raw materials, is about 30%-60%; preferably 45%-55%; for example, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, or 60%.

[0035] According to embodiments of the present invention, the amount of the slow-release polymer, based on the total weight of the raw materials, is about 30%-60%; preferably 45%-55%; for example, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, or 60%.

[0036] According to embodiments of the present invention, the amount of the amphiphilic polymer, based on the total weight of the raw materials, is about 0.2%-10%; preferably 0.5%-5%; for example, 0.5%, 0.8%, 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5%, 5.5%, 6%, 6.5%, 7%, 7.5%, 8%, 8.5%, 9%, 9.5%, and 10%. In this application, the percentage of usage is a weight percentage or a mass percentage.

[0037] According to an embodiment of the present invention, the weight ratio of the sustained-release polymer to the amphiphilic polymer is 100:0-80:20; preferably 99.8:0.2-90:10.

[0038] According to an embodiment of the present invention, the sustained-release polymer is lactide-glycolic acid copolymer (PLGA).

[0039] According to an embodiment of the present invention, the PLGA has a carboxyl end cap or the PLGA chain is end capped with a carboxyl group.

[0040] According to an embodiment of the present invention, the L / G (molar ratio of lactide to glycolide) of the lactide-glycol copolymer is 75:25-25:75 (e.g., 50:50).

[0041] According to an embodiment of the present invention, the molecular weight of the PLGA is 7kDa-17kDa.

[0042] According to an embodiment of the present invention, the amphiphilic polymer is selected from poloxamer; for example, poloxamer F68 and poloxamer F127.

[0043] According to an embodiment of the present invention, the amphiphilic polymer is selected from PEG-PLA, such as PEG(2k~10k)-PLA(1k~30k), exemplarily PEG(5k)-PLA(15k) and PEG(6k)-PLA(1k).

[0044] According to an embodiment of the present invention, the raw materials for preparing the progesterone sustained-release microspheres include, or the components of the progesterone sustained-release microspheres are:

[0045] The total weight of the raw materials includes 45%-55% (e.g., 47%-53%, 50%) of progesterone, 45%-55% (e.g., 45%-49%, 47%-49%) of PLGA and 0.5%-5% (e.g., 1%-3%) of amphiphilic polymers.

[0046] Preferably, the amphiphilic polymer is selected from poloxamer or PEG-PLA.

[0047] According to an embodiment of the present invention, the raw materials for preparing the progesterone sustained-release microspheres include, or the components of the progesterone sustained-release microspheres are:

[0048] Based on the total weight of the raw materials, it contains 45%-55% progesterone, 45%-55% PLGA, and 0.5%-5% poloxamer.

[0049] Alternatively, by total weight of raw materials, 45%-55% progesterone, 45%-55% PLGA and 0.5%-5% PEG-PLA.

[0050] According to an embodiment of the present invention, the raw materials for preparing the progesterone sustained-release microspheres include, or the components of the progesterone sustained-release microspheres are:

[0051] The total weight of the raw materials is 47%-53% (e.g., 50%) of progesterone, 45%-49% (e.g., 47%-49%) of PLGA and 0.5%-5% (e.g., 1%-3%) of poloxamer.

[0052] Alternatively, by total weight of raw materials, 47%-53% (e.g., 50%) of progesterone, 45%-49% (e.g., 47%-49%) of PLGA and 0.5%-5% (e.g., 1%-3%) of PEG-PLA.

[0053] On the other hand, the present invention also provides a method for preparing the above-mentioned progesterone sustained-release microspheres, wherein the progesterone sustained-release microspheres are prepared by an oil-in-water (O / W) emulsification-solvent removal method.

[0054] According to an embodiment of the present invention, the preparation method includes:

[0055] (1) Prepare an organic dispersion containing progesterone, a sustained-release polymer (and an amphiphilic polymer optionally added or not added) and an organic solvent;

[0056] (2) The organic dispersed phase is mixed with an aqueous continuous phase containing water and a stabilizer (such as polyvinyl alcohol (PVA)) to form an O / W emulsion;

[0057] (3) Remove organic solvents from O / W emulsions by extraction and / or evaporation to form solid microspheres;

[0058] (4) The solid microspheres are dried to obtain progesterone sustained-release microspheres loaded with progesterone.

[0059] In some embodiments of the present invention, the organic solvent is selected from any one, two or more of the following: dichloromethane, ethyl acetate, acetone, benzyl alcohol, methanol, ethanol, isopropanol, 1-butanol, isobutanol, 1-pentanol, isoamyl alcohol, 1-hexanol (a single organic solvent, or a mixture of two or more organic solvents).

[0060] According to an embodiment of the present invention, the mass-volume ratio (g:mL) of progesterone to organic solvent is 3:1-3:100, for example 3:5, 3:8, 3:10, 3:20, 3:30, 3:40, 3:50, 3:60, 3:70, 3:80, 3:90.

[0061] According to an embodiment of the present invention, the aqueous continuous phase (including aqueous continuous phase-1 and aqueous continuous phase-2) is a 1% aqueous solution of polyvinyl alcohol.

[0062] According to an embodiment of the present invention, step (2) specifically involves: shearing and emulsifying the organic dispersed phase and the aqueous continuous phase-1, and then introducing the sheared emulsion into the aqueous continuous phase-2 to form solid microspheres.

[0063] According to an embodiment of the present invention, the volume of the aqueous continuous phase-1 is 5 to 30 times the volume of the organic solvent, for example, 8, 10, 12, 15, 16, 18, 20, 23, 25, or 28 times.

[0064] According to an embodiment of the present invention, the volume of aqueous continuous phase-2 is 1 to 30 times the volume of aqueous continuous phase-1, for example, 3 times, 5 times, 8 times, 10 times, 12 times, 15 times, 16 times, 18 times, 20 times, 22 times, 25 times, and 28 times.

[0065] According to an embodiment of the present invention, the shear emulsification rate is 3000rpm-6000rpm, for example 3500rpm, 3800rpm, 4000rpm, 4300rpm, 4500rpm, 4800rpm, 5000rpm, 5300rpm, 5500rpm, and 5800rpm.

[0066] According to an embodiment of the present invention, in step (3), after removing the organic solvent, the solid microsphere suspension is subjected to sieving and dehydration treatment.

[0067] According to an embodiment of the present invention, the material is sieved using a 100-800 mesh sieve (e.g., 100 mesh, 200 mesh, 300 mesh, 400 mesh, 500 mesh, 600 mesh, 700 mesh, 800 mesh).

[0068] According to an embodiment of the present invention, dehydration is performed by vacuum filtration; for example, dehydration is performed by vacuum filtration on a filter membrane (e.g., 5 μm).

[0069] According to an embodiment of the present invention, after step (3) is completed, the solid microspheres are optionally washed with water and then concentrated.

[0070] According to an embodiment of the present invention, in step (4), the drying method is vacuum freeze drying.

[0071] In another aspect, the present invention also provides a formulation comprising the above-mentioned progesterone sustained-release microspheres.

[0072] According to an embodiment of the present invention, the formulation is an injectable sustained-release formulation.

[0073] According to an embodiment of the present invention, the formulation comprises the above-mentioned progesterone sustained-release microspheres and (appropriate amount) diluent.

[0074] According to an embodiment of the present invention, the above-mentioned progesterone sustained-release microspheres are suspended in a diluent to obtain the injectable sustained-release formulation.

[0075] According to an embodiment of the invention, the diluent comprises water (preferably water for injection) and one, two or more pharmaceutically acceptable excipients.

[0076] According to embodiments of the present invention, the pharmaceutically acceptable excipient comprises any one or any combination thereof of surfactants (e.g., Tween 80), thickeners (e.g., sodium carboxymethyl cellulose), osmotic pressure regulators (e.g., mannitol), and pH buffers (e.g., citric acid, disodium hydrogen phosphate).

[0077] According to embodiments of the present invention, the diluent comprises water, an osmotic pressure regulator (e.g., mannitol), and a thickener (e.g., sodium carboxymethyl cellulose). Preferably, the concentration of the osmotic pressure regulator (e.g., mannitol) is 1%-10%, for example, 3%, 4%, 5%, 6%, 7%, or 8%. Preferably, the concentration of the thickener (e.g., sodium carboxymethyl cellulose) is 0.1%-5%, for example, 0.2%, 0.3%, 0.4%, 0.5%, 1%, 1.5%, 2%, 2.5%, or 3%.

[0078] According to embodiments of the invention, the diluent optionally comprises a surfactant (e.g., Tween 80). Preferably, the concentration of the surfactant (e.g., Tween 80) is 0.005%-0.8%, for example 0.01%, 0.03%, 0.05%, 0.08%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, or 0.7%.

[0079] According to an embodiment of the invention, the diluent comprises water, an osmotic pressure regulator (e.g., mannitol), a thickener (e.g., sodium carboxymethyl cellulose), and a surfactant (e.g., Tween 80).

[0080] According to embodiments of the present invention, the relative bioavailability of a formulation comprising the progesterone sustained-release microspheres, after a single dose, can reach 80%-125% (F%) of the corresponding dose of a marketed progesterone injection administered over 7 consecutive days, for example, 81%, 81.1%, 83%, 85%, 85.6%, 88%, 88.6%, 90%, 93%, 93.3%, 95%, 98%, 98.6%, 100%, 100.6%, 103%, 105%, 108%, 108.3%, 110%, 113%, 115%, 118%, 120%, 120.1%, and 123%. The corresponding dose refers to the total dose of a marketed progesterone injection administered over 7 consecutive days, with the same dose injected each day, which is equivalent to the dose of a single dose of the progesterone sustained-release microspheres.

[0081] According to an embodiment of the present invention, the dosage form comprising the progesterone sustained-release microspheres is administered once a week.

[0082] According to an embodiment of the present invention, a formulation comprising the progesterone sustained-release microspheres, administered once, has a C max C compared to a single dose of a marketed progesterone injection max The ratio is between 1:1 and 1:10. More preferably, the ratio is between 1:1.5 and 1:8, for example, 1:1.5, 1:1.8, 1:2.0, 1:2.8, 1:3.0, 1:3.1, 1:3.5, 1:4.0, 1:4.5, 1:5.0, 1:5.3, 1:5.5, 1:6.0, 1:6.5, 1:7.0, 1:7.5, and 1:8.0.

[0083] According to an embodiment of the present invention, a formulation comprising the progesterone sustained-release microspheres, administered once, has a C max / C min The ratio ranges from 2:1 to 15:1, for example, 2.5:1, 2.8:1, 3.0:1, 3.5:1, 4.0:1, 4.5:1, 5.0:1, 5.5:1, 5.7:1, 6.0:1, 6.5:1, 6.6:1, 6.7:1, 7.0:1, 7.4:1, 7.5:1, 8.0:1, 9.0:1, 10.0:1, 11.0:1, 11.2:1, 12.0:1, 13.0:1, and 14.0:1.

[0084] According to an embodiment of the present invention, the AUC of a formulation comprising the progesterone sustained-release microspheres 0-168h It is 1100h·ng / mL-2500h·ng / mL, such as 1200h·ng / mL, 1300h·ng / mL, 1400h·ng / mL, 1433.4h·ng / mL, 1500h·ng / mL, 1513.7h·ng / mL, 1566.5h·ng / mL, 1600h·ng / mL, 1650.0h·ng / mL, 1700 h·ng / mL, 1743.0h·ng / mL, 1778.0h·ng / mL, 1800h·ng / mL, 1900h·ng / mL, 1914.2h·ng / mL, 2000h·ng / mL, 2100h·ng / mL, 2123.6h·ng / mL, 2200h·ng / mL, 2300h·ng / mL, 2400h·ng / mL.

[0085] According to an embodiment of the present invention, the formulation comprises the above-mentioned progesterone sustained-release microspheres and a diluent;

[0086] The diluent contains water, 1%-10% mannitol, and 0.1%-3% sodium carboxymethyl cellulose;

[0087] Preferably, the diluent further includes 0.01%-0.5% Tween 80;

[0088] Preferably, the formulation containing progesterone sustained-release microspheres has a single-dose C max / C min The ratio is between 2:1 and 10:1.

[0089] The present invention also provides the use of the above-mentioned progesterone sustained-release microspheres and formulations containing the above-mentioned progesterone sustained-release microspheres in the preparation of medicaments for treating gynecological conditions, or in the preparation of medicaments for supporting pregnancy, assisted reproduction and / or improving fertility.

[0090] The present invention also provides a method for treating gynecological conditions, supporting pregnancy, assisted reproduction and / or improving fertility, the method comprising administering (e.g., intramuscular injection) a therapeutically effective amount of the above-mentioned progesterone sustained-release microspheres or a formulation containing the above-mentioned progesterone sustained-release microspheres to a subject in need.

[0091] According to an embodiment of the present invention, the gynecological condition is selected from amenorrhea, endometriosis, abnormal uterine bleeding due to hormonal imbalance, menstrual disorders, and dysmenorrhea. Beneficial effects

[0092] This invention addresses clinical pain points in existing treatment regimens by using microsphere encapsulation technology to prepare progesterone sustained-release microspheres and formulations containing these microspheres. These formulations, while maintaining bioavailability comparable to marketed progesterone injections, can reduce the dosing frequency from once or multiple daily injections to once weekly. More importantly, these formulations maintain stable blood drug concentrations throughout the dosing cycle, avoiding fluctuations in drug concentration peaks and troughs, effectively improving the efficacy of progesterone replacement therapy and reducing adverse reactions. Attached Figure Description

[0093] Figure 1 is a time-in-time curve of animal testing for comparative formulation 1 and formulations 4, 5, and 6 of the present invention. The horizontal axis represents time (hours), and the vertical axis represents plasma concentration (ng / mL).

[0094] Terminology Explanation

[0095] "More than three" means three or more types.

[0096] In this invention, if a numerical variable is selected from ab, it means that the variable can be the point value of any specific number within the range of ab (including the endpoints) or the range value of any number within the range of ab (including the endpoints). For example, if a variable is selected from any integer from 1 to 20, it means that the variable can be 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20, or it can be 1-20, 1-16, 1-10, 1-6, 1-4, 1-3, 1-2, 2-20, 2-16, 2-10, 2-6, 2-4, 2-3, 3-20, 3-16, 3-10, 3-6, 3-4, 4-20, 4-16, 4-10, 4-6, 6-20, 6-16, 6-10, 10-20, 10-16, or 16-20, etc. Similar descriptions in this invention can be understood with reference to the foregoing definitions.

[0097] Unless otherwise specified, the term "L / G" indicates the molar ratio of lactide to glycolide in PLGA.

[0098] Unless otherwise specified, all figures representing the amount, properties (e.g., molecular weight, experimental data), etc., of components as used in this specification and claims should be understood to be modified in all cases by the term "about". Therefore, unless otherwise stated, the numerical characteristics set forth herein are approximate values ​​that can vary depending on the desired properties sought to be obtained in embodiments of the invention or common knowledge in the art. In some embodiments, the term "about" means within ±5% of a specific value or range, preferably within 2%; even within 1%.

[0099] Unless otherwise stated, the amount of each raw material used when calculating "by total weight of raw materials" does not include raw materials used in the microsphere preparation process but not contained in the final product, such as solvents used. In other words, substances not contained in the final microsphere product are not included in the category of raw materials mentioned herein. Detailed Implementation

[0100] Before further describing the invention, it should be understood that the invention is not limited to the specific embodiments described. It should also be understood that the terminology used herein is for the purpose of describing specific embodiments only and is not intended to be limiting.

[0101] It should be noted that, as used herein, unless the context clearly specifies otherwise, the singular forms “a,” “an,” and “the / said” include a plural of indicators.

[0102] Example 1. Preparation of comparative formulations 1-2

[0103] The preparation of progesterone sustained-release microspheres is described in patent CN115605187B. Referring to Example 1 in the specification, two types of progesterone microspheres—comparative formulations 1 and 2—were prepared using an emulsification-solvent evaporation method with a copolymer of progesterone and lactide (PLGA).

[0104] The preparation of contrast formulations 1-2 was as follows: An organic phase solution was prepared by mixing 2.4 g of one of the PLGAs listed in Table 1, 2.4 g of progesterone, and 16 mL of dichloromethane (DCM). The mixture was stirred at room temperature to form a clear solution. Subsequently, the organic phase solution was mixed with 240 mL of a 1% PVA (polyvinyl alcohol 17-88, Jiangxi Alpha High-Tech, ChP) aqueous solution and fed into a wire shear press (Kinematica) at a flow rate ratio of 1:15 using a peristaltic pump. The microspheres were sheared and emulsified at 3000 rpm using MT3100S2, and the resulting emulsion was passed into 2.4 L of 0.1% PVA aqueous solution. The emulsion was stirred at medium speed for about 4 hours to remove the solvent. The solidified microsphere suspension was sieved through a 150 μm sieve to separate any large agglomerates, and the product was dehydrated and collected through a 25 μm sieve. The microspheres were further washed with purified water and the suspension was concentrated. The microsphere suspension was freeze-dried under vacuum for 48 h. The drug loading, encapsulation efficiency, and particle size of the control formulations 1-2 were then determined, and the results are shown in Table 2.

[0105] Table 1: List of PLGA polymers used to prepare comparative formulations 1 and 2

[0106] Table 2: Detailed information on comparative formulations 1 and 2

[0107] The following HPLC methods were used to determine the drug loading (%) and encapsulation efficiency (%):

[0108] Chromatographic parameters:

[0109] Chromatographic column: YMC Triart C18, 3μm, 4.6mm × 150mm;

[0110] Column temperature: 30℃;

[0111] Mobile phase: 80:20 (v / v) acetonitrile / water, isocratic;

[0112] Flow rate: 1.0 mL / min;

[0113] Injection volume: 10 μL;

[0114] Wavelength: 241nm.

[0115] The standard solution was prepared by accurately weighing approximately 10 mg of progesterone reference standard into a 100 mL volumetric flask, adding approximately 80 mL of methanol, mixing and sonicating to dissolve, and then diluting to volume with methanol to prepare a working standard solution of approximately 100 μg / mL progesterone.

[0116] The sample solution was prepared as follows: Approximately 20 mg of PLGA microspheres loaded with progesterone was accurately weighed into a 100 mL volumetric flask, and approximately 50 mL of acetonitrile was added. The mixture was then stirred and sonicated to dissolve the microspheres, and the volume was brought to a final volume with water. After centrifugation, the supernatant was collected for HPLC analysis.

[0117] The percentage of drug loading is calculated using the following formula:

[0118] Drug loading (%) = weight of progesterone in the microsphere sample / total weight of microspheres × 100%.

[0119] Encapsulation efficiency is calculated using the following formula:

[0120] Encapsulation efficiency (%) = Actual drug loading percentage / Theoretical drug loading percentage × 100%.

[0121] The following particle size distribution tests were performed on the progesterone-loaded microspheres:

[0122] Instrument parameters:

[0123] Dispersant: 0.1% Tween 20 aqueous solution;

[0124] Refractive index: 1.33;

[0125] Lower limit of light blocking: 3%;

[0126] Upper limit of opacity: 15%;

[0127] Stirring speed: 2100 rpm;

[0128] Result type: Volume distribution;

[0129] Number of measurements: 3;

[0130] Data output: Average value is created after SOP is completed;

[0131] Add approximately 200 mL of 0.1% Tween 20 solution to the liquid sample dispersion device of the Malvern 3000 laser particle size analyzer, set the instrument parameters as described above, add an appropriate amount of progesterone microsphere sample, and ensure the detector occlusion is within the range of 3%-15% before performing particle size distribution determination.

[0132] Example 2. Preparation of modified microsphere formulation

[0133] The polymer (PLGA RG502H, or a combination of PLGA RG502H and an amphiphilic polymer), progesterone, and dichloromethane (DCM) were mixed according to the ratios shown in Table 3 (g:g:mL). The mixture was stirred at room temperature to form a clear solution. Subsequently, the organic phase solution was fed into a wire shear mill at a flow rate ratio of 1:15 with a 1% PVA aqueous solution (W1, the volume of which is 15 times that of the dichloromethane) via a peristaltic pump and sheared at a rate of 4500 rpm. The resulting emulsion was then passed into 10 times the volume of W1 in a 0.1% PVA aqueous solution. The emulsion was stirred at a medium speed for approximately 4 hours to remove the solvent. The solidified microsphere suspension was sieved through a 200-mesh sieve to separate any large agglomerates, and the product was dehydrated and collected by vacuum filtration through a 5 μm filter membrane. The microspheres were further washed with purified water, and the suspension was concentrated. The microsphere suspension was then freeze-dried under vacuum for 48 hours. The drug loading, encapsulation efficiency, and particle size of the modified microsphere formulation were then determined using the method described in Example 1, and the results are shown in Table 3.

[0134] Table 3: Detailed information on the microsphere formulation of the present invention

[0135] Example 3. Preparation of modified microsphere formulation

[0136] The polymer (PLGA RG502H, or a combination of PLGA RG502H and an amphiphilic polymer), progesterone, and DCM were mixed according to the ratios in Table 4 (g:g:mL). The mixture was stirred at room temperature to form a clear solution. Subsequently, the organic phase solution was fed into a wire shear mill at a flow rate ratio of 1:15 with a 1% PVA aqueous solution (W1, the volume of which is 15 times that of dichloromethane and DCM) via a peristaltic pump and sheared at a rate of 4500 rpm. The resulting emulsion was then passed into 10 times the volume of W1 in a 0.1% PVA aqueous solution. The emulsion was stirred at a medium speed for approximately 4 hours to remove the solvent. The solidified microsphere suspension was sieved through a 600-mesh sieve to separate any large agglomerates, and the product was dehydrated and collected by vacuum filtration through a 5 μm filter membrane. The microspheres were further washed with purified water, and the suspension was concentrated. The microsphere suspension was then freeze-dried under vacuum for 48 hours. The drug loading, encapsulation efficiency, and particle size of the modified microsphere formulation were then determined using the method described in Example 1, and the results are shown in Table 4.

[0137] Table 4: Detailed information on the microsphere formulation of the present invention

[0138] Example 4. Diluent

[0139] The diluents were prepared according to the formulations shown in Table 5, and their viscosity was measured. The results are shown in Table 5; the viscosities of diluents 1-4 all meet the requirements for injectable formulations.

[0140] Table 5: Diluent Formulation and Viscosity

[0141] The sustained-release microspheres prepared in this invention can be suspended in a diluent to obtain an injectable sustained-release formulation.

[0142] Example 5. Zoological Experiment

[0143] Test sample: Take comparative formulations 1 and 2, and formulations 1 to 8 of the present invention. Weigh 150 mg of each sample, suspend it in 1 mL of diluent 4 (see Table 5 of Example 4) to prepare the test sample with a strength of 65 mg / mL.

[0144] Reference standard: Commercially available progesterone aqueous solution injection, specification 1.112mL:25mg (Changchun Jinsai Pharmaceutical Co., Ltd.).

[0145] Experimental animals: Male beagles, weighing 8-12 kg, 4 per group.

[0146] Dosage: The test sample was administered as a single dose of 6.2 mg / kg, and the control sample was administered as a single dose of 0.9 mg / kg.

[0147] Administration route: All test samples were administered via intramuscular injection, while the reference samples were administered via subcutaneous injection.

[0148] Sample collection: Plasma was collected for the test sample at the following time points: 0h (before drug administration), 0.5h, 1h, 2h, 4h, 8h, 12h, 24h, 48h, 72h, 96h, 120h, 144h, and 168h after drug administration. Plasma was collected for the control sample at the following time points: 0h (before drug administration), 0.25h, 0.5h, 0.75h, 1h, 1.5h, 2h, 3h, 4h, 6h, 9h, 12h, and 24h after drug administration.

[0149] The PK results of each group of zoological experiments are shown in Table 6.

[0150] Table 6: PK parameters of test sample and reference sample

[0151] The bioavailability (F%) of the formulation is calculated using the formula: F% = AUC of the formulation 0-168h / (AUC of reference standard) 0-24h ×7)×100%

[0152] Comparison of PK data shows that progesterone sustained-release microspheres significantly reduced peak-trough phenomena compared to the control, i.e., C max / C min The ratio of [specific component] to [specific component] is significantly reduced, which is beneficial for improving drug efficacy. The C [specific component] of microsphere formulations... maxAll concentrations were significantly reduced, and the situation of instantaneous high drug concentrations was improved, which is beneficial to reducing the irritation caused by progesterone itself. This indicates that microsphere encapsulation of progesterone can significantly alter the release behavior of progesterone, giving it sustained-release properties. More specific comparisons:

[0153] ① The bioavailability of the control formulations 1 and 2 is insufficient, failing to reach the equivalence level of ≥80%, and cannot be used as weekly formulations to achieve clinical efficacy; while the bioavailability of the formulations 1-8 of this invention is not inferior to that of the control.

[0154] ②Compared with control formulation 1, formulation 1 has the same formulation composition, but formulation 1 has a smaller particle size, which can improve bioavailability to a certain extent; indicating that smaller particle size can increase the specific surface area of ​​microspheres and increase the exposure of progesterone during the expected release cycle.

[0155] ③ Compared with formulation 1, formulation 2 has the addition of poloxamer F68 in its formulation, which further improves its bioavailability;

[0156] ④ Compared with formulation 2, formulation 4 has a further reduced particle size, especially a reduced D90, which can achieve higher bioavailability and further prove the correlation between particle size and progesterone release in microspheres;

[0157] ⑤ Compared with formulation 4, formulation 5 changed the amphiphilic polymer from poloxamer F68 to poloxamer F127, further improving bioavailability; formulation 6, based on formulation 5, further increased the content of poloxamer F127, resulting in improved bioavailability, C max It reduces blood drug concentration and makes blood drug concentrations more stable;

[0158] ⑥ Compared with formulation 4, formulations 7 and 8 show that although PEG-PLA can also control burst release, its effect on improving release rate is not as good as poloxamer.

[0159] ⑦ Compared with formulation 5, formulations 7 and 8 have smaller microspheres, but their effect on improving release is significantly lower than that of formulation 5 microspheres. This indicates that the selection of excipients is more important for improving the release characteristics of microspheres than the improvement of particle size.

[0160] The above research results demonstrate that the formulation provided by this invention can better meet clinical needs.

[0161] The embodiments of the present invention have been described above. However, the present invention is not limited to the above embodiments. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A progesterone sustained-release microsphere, characterized in that, The raw materials for preparing the progesterone sustained-release microspheres or the components of the progesterone sustained-release microspheres include: (1) Progesterone as an active substance; and (2) One, two or more sustained-release polymers; Preferably, the progesterone sustained-release microspheres have the following particle size distribution: D10 is approximately 5μm-20μm, preferably approximately 5μm-15μm; And / or, D50 is about 15μm-35μm, preferably about 20μm-30μm; And / or, D90 is about 30μm-70μm, preferably about 35μm-65μm.

2. The progesterone sustained-release microspheres according to claim 1, characterized in that, The mass ratio of the sustained-release polymer to progesterone is 1:10-10:1, preferably 1:5-5:1; Preferably, the sustained-release polymer is selected from any one, two or more of the following: lactide-glycolic acid copolymer (PLGA), polylactide (PLA), polycaprolactone (PCL), lactide-caprolactone copolymer (PLC), lactide-glycolic acid-caprolactone copolymer (PLGC), polyorthoester (POE), and polyhydroxyalkanoate (PHA). Preferably, the molar ratio (L / G) of lactide to glycolide in the lactide-glycol copolymer is 85:15-40:60; Preferably, the molecular weight of the sustained-release polymer is in the range of about 4 kDa to about 17 kDa; Preferably, the chain ends of the sustained-release polymer are capped with carboxyl groups or ester groups.

3. The progesterone sustained-release microspheres according to claim 1 or 2, characterized in that, The raw materials used to make the progesterone sustained-release microspheres or the components of the progesterone sustained-release microspheres may optionally include one, two or more amphiphilic polymers; Preferably, the amphiphilic polymer is selected from block polymers containing polyester and / or polyether, and / or other amphiphilic polymers; more preferably, the amphiphilic polymer is selected from polyoxyethylene-polyoxypropylene block copolymers, polyoxyethylene-polysiloxane block copolymers, polyethylene glycol-long-chain fatty acids, and polyethylene glycol-polyester block copolymers; more preferably, the amphiphilic polymer is selected from poloxamer, polyethylene glycol-polylactide (PEG-PLA), and polyethylene glycol-lactide-glycolic acid copolymer (PEG-PLGA); Preferably, the amphiphilic polymer is selected from poloxamer; preferably, the amphiphilic polymer is selected from poloxamer F68 and poloxamer F127. Preferably, the amphiphilic polymer is selected from PEG (2k~10k)-PLA (1k~30k); more preferably PEG (5k)-PLA (15k) or PEG (6k)-PLA (1k); Preferably, the weight ratio of the sustained-release polymer to the amphiphilic polymer is 100:0-80:20; more preferably, the weight ratio is 99.8:0.2-90:

10.

4. A progesterone sustained-release microsphere, characterized in that, The raw materials for making the progesterone sustained-release microspheres include, or the components of the progesterone sustained-release microspheres include: progesterone as the active substance, a sustained-release polymer, and an amphiphilic polymer; Preferably, the amount of progesterone used is about 30%-60% based on the total weight of the raw materials; more preferably, it is 45%-55%. Preferably, the amount of the slow-release polymer is about 30%-60% based on the total weight of the raw materials; more preferably, it is 45%-55%. Preferably, the amount of amphiphilic polymer used is about 0.2%-10% based on the total weight of the raw materials; more preferably, it is 0.5%-5%. Preferably, the weight ratio of the sustained-release polymer to the amphiphilic polymer is 100:0-80:20; more preferably, it is 99.8:0.2-90:

10. Preferably, the sustained-release polymer is lactide-glycolic acid copolymer (PLGA); Preferably, the PLGA has carboxyl-terminated ends; Preferably, the molar ratio (L / G) of lactide to glycolide in the lactide-glycol copolymer is 75:25-25:75; Preferably, the molecular weight of the PLGA is in the range of about 7 kDa to about 17 kDa; Preferably, the amphiphilic polymer is selected from poloxamer; for example, poloxamer F68 and poloxamer F127. Preferably, the amphiphilic polymer is selected from PEG (2k~10k)-PLA (1k~30k); more preferably PEG (5k)-PLA (15k) or PEG (6k)-PLA (1k); Preferably, the raw materials for preparing the progesterone sustained-release microspheres include, or are components of, the progesterone sustained-release microspheres: 45%-55% progesterone, 45%-55% PLGA and 0.5%-5% poloxamer, by weight of the total raw materials; or, 45%-55% progesterone, 45%-55% PLGA and 0.5%-5% PEG-PLA, by weight of the total raw materials. Preferably, the raw materials for preparing the progesterone sustained-release microspheres include, or are components of, the progesterone sustained-release microspheres: 47%-53% (e.g., 50%) of progesterone, 45%-49% (e.g., 47%-49%) of PLGA and 0.5%-5% (e.g., 1%-3%) of poloxamer, by weight of the raw materials; or, 47%-53% (e.g., 50%) of progesterone, 45%-49% (e.g., 47%-49%) of PLGA and 0.5%-5% (e.g., 1%-3%) of PEG-PLA, by weight of the raw materials.

5. The method for preparing progesterone sustained-release microspheres according to any one of claims 1-4, wherein the progesterone sustained-release microspheres are prepared by an oil-in-water (O / W) emulsification-solvent removal method; Preferably, the preparation method includes: (1) Prepare an organic dispersion containing progesterone, a sustained-release polymer, an amphiphilic polymer (optionally added or not added), and an organic solvent; (2) The organic dispersed phase is mixed with an aqueous continuous phase containing water and a stabilizer to form an O / W emulsion; (3) Remove organic solvents from O / W emulsions by extraction and / or evaporation to form solid microspheres; (4) The solid microspheres are dried to obtain progesterone sustained-release microspheres loaded with progesterone.

6. The preparation method according to claim 5, characterized in that, Step (2) specifically involves shear emulsifying the organic dispersed phase with the aqueous continuous phase-1, and then introducing the sheared emulsion into the aqueous continuous phase-2 to form solid microspheres; preferably, the shear emulsification rate is 3000 rpm-6000 rpm. Preferably, in step (3), after removing the organic solvent, the solid microsphere suspension is subjected to sieving and dehydration treatment; preferably, it is sieved through a 100-800 mesh sieve.

7. A formulation comprising the progesterone sustained-release microspheres according to any one of claims 1-4; Preferably, the formulation is an injectable sustained-release formulation; Preferably, the formulation containing the progesterone sustained-release microspheres is administered once a week.

8. The formulation according to claim 7, characterized in that, The formulation comprises the progesterone sustained-release microspheres and diluent as described in any one of claims 1-4; Preferably, the diluent comprises water and one, two or more pharmaceutically acceptable excipients; preferably, the pharmaceutically acceptable excipients comprise surfactants, thickeners, osmotic pressure regulators, pH buffers or combinations thereof; preferably, the diluent comprises water, osmotic pressure regulators, and thickeners.

9. The use of the progesterone sustained-release microspheres according to any one of claims 1-4, and the formulation comprising the progesterone sustained-release microspheres according to any one of claims 1-4, in the preparation of a medicament for treating gynecological conditions, or in the preparation of a medicament for supporting pregnancy, assisted reproduction, and / or improving fertility; Preferably, the gynecological conditions are selected from amenorrhea, endometriosis, abnormal uterine bleeding due to hormonal imbalance, menstrual disorders, and dysmenorrhea.

10. A method for treating gynecological conditions, supporting pregnancy, assisting reproduction and / or improving fertility, the method comprising administering to a subject in need a therapeutically effective amount of the progesterone sustained-release microspheres of any one of claims 1-4 or a formulation comprising the progesterone sustained-release microspheres of any one of claims 1-4; Preferably, the gynecological conditions are selected from amenorrhea, endometriosis, abnormal uterine bleeding due to hormonal imbalance, menstrual disorders, and dysmenorrhea.

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