Solumaglutide soluble microneedle patch composition

Abstract

The invention discloses a semaglutide soluble microneedle patch composition. The soluble microneedle patch composition comprises a needle body material and a backing layer material, the material of the needle body can be only composed of a high-concentration semaglutide solution. According to the semaglutide micro-needle patch, the drug loading capacity of semaglutide in the micro-needle patch can be greatly improved, the hypoglycemic effect which is the same as that of subcutaneous injection administration can be achieved under the same administration amount, and a new dosage form choice is provided for the pharmaceutical application of semaglutide, such as hypoglycemic weight reduction and the like.

Description

Technical Field

[0001] The present invention relates to the technical field of pharmaceutical preparations, and in particular to a semaglutide soluble microneedle patch. Background Art

[0002] At present, most macromolecular biological drugs and vaccines are delivered by subcutaneous, intradermal or intramuscular injection. The injection route is suitable for almost any type of drug molecules to enter the human body. It is a low-cost, fast and direct way of administration, but its disadvantages are also obvious, that is, the patient is painful during injection, long-term injection leads to damage to the injection site, and it is not suitable for patients to use by themselves. The injection needle is reduced to micrometer scale, and a large number of needle-like structures are arranged on the base to form a minimally invasive transdermal drug delivery device-microneedle system. The microneedle can penetrate the stratum corneum and epidermis to the dermis. The drug carried by the microneedle is absorbed into the systemic circulation through the capillaries and lymphatic capillaries in the dermis. Because the microneedle is small enough, it can avoid stimulating nerve fibers or piercing blood vessels. It is a non-invasive and efficient transdermal drug delivery method with good patient compliance.

[0003] Microneedles can be divided into many types according to their preparation materials and drug-carrying properties, including solid microneedles, coated microneedles, hollow microneedles, swellable microneedles, and soluble microneedles. Dissolvable microneedles (DMN) are made of water-soluble and biodegradable materials containing drugs. When the microneedles penetrate the skin, the needle body can dissolve or decompose in the skin tissue fluid, and the therapeutic drug is released into the skin, thereby achieving the purpose of drug delivery. Soluble microneedles have the characteristics of high drug delivery efficiency, good biocompatibility, easy manufacturing, and low cost, and are currently a hot research topic.

[0004] Semaglutide is a GLP-1 analog developed by Novo Nordisk. It has an amino acid substitution at position 8 and a fatty acid side chain modification at position 26 of natural GLP-1, giving it a longer half-life. The injection was approved for marketing by the FDA in 2017. Its shortcomings are common problems with traditional injection routes of administration. In 2019, oral tablets were approved for marketing, avoiding the use of needles, while extremely low bioavailability and high variability have become obstacles to medication. Currently, many studies use microneedles to deliver macromolecular drugs for diabetes, such as insulin and GLP-1 analogs. The dosage of semaglutide injection for diabetes indications is 1 mg once a week, and the dosage for weight loss indications is 2.4 mg once a week. The general microneedle system has a low drug loading capacity and can only deliver sub-milligram doses of drugs, which limits the practical application of drug-loaded microneedle delivery of semaglutide. Summary of the invention

[0005] In order to overcome the problems existing in the above-mentioned prior art, one of the purposes of the present invention is to provide a semaglutide soluble microneedle patch composition, the needle body material of which can be composed only of high-concentration semaglutide, which greatly improves the drug loading of semaglutide in the microneedle patch. In addition, the effect of the soluble microneedle patch composition in the present invention in regulating blood sugar is nearly 40 times higher than that of semaglutide oral tablets, which greatly improves the bioavailability of the soluble microneedle patch composition in the present invention.

[0006] In order to achieve the above-mentioned object, the present invention provides a semaglutide soluble microneedle patch composition, comprising a needle body material and a backing layer material, wherein the needle body material comprises semaglutide.

[0007] Semaglutide of the present invention is a GLP-1 receptor agonist, which can be prepared as described in Example 4 of WO2006 / 097537 and is also referred to as N 6,26 -{18-[N-(17-carboxyheptadecanoyl)-L-γ-glutamyl]-10-oxo-3,6,12,15-tetraoxa-9,18-diazaoctadecanoyl}-[8-(2-amino-2-propionic acid), 3,4-L-arginine] human glucagon-like peptide 1 (7-37), see WHO Drug Information Vol. 24, No. 1, 2010 (SEQ ID NO: 57).

[0008] In the preferred technical solution, the needle solution is composed of a semaglutide solution, and the mass percentage concentration of semaglutide is 20% to 40%.

[0009] In the optional technical solution, the needle body material includes semaglutide and sodium hyaluronate, the mass percentage concentration of semaglutide in the needle body solution is 0.25% to 15%, and the mass percentage concentration of sodium hyaluronate is 4% to 15%; the needle body solution may further include mannitol, with a mass percentage concentration of 1% to 5%; the needle body material may also include at least one of polyvinyl pyrrolidone, poloxamer, and hydroxypropyl methylcellulose.

[0010] In the preferred technical solution, the backing layer material includes sodium hyaluronate or is composed of sodium hyaluronate, and the mass percentage concentration of sodium hyaluronate in the backing layer solution is 15%; the backing layer material also includes polyvinyl pyrrolidone.

[0011] The present invention also provides a method for preparing the above-mentioned soluble microneedle patch composition, comprising the following steps:

[0012] S1: preparing needle body solution and backing layer solution;

[0013] S2: adding the needle solution into the microneedle negative mold, centrifuging, and drying;

[0014] S3: adding the backing layer solution, centrifuging, and then drying and demoulding to obtain the soluble microneedle patch composition.

[0015] The centrifugation step is specifically centrifugation at 2236g centrifugal force for 5 minutes.

[0016] The present invention mainly uses sodium hyaluronate as the backing layer material and adopts a two-step mold casting method to prepare a semaglutide soluble microneedle patch. Whether the needle fluid is composed of high-concentration semaglutide, or further contains sodium hyaluronate, further contains a semaglutide protective agent such as mannitol, or further contains one or more of the molding materials polyvinyl pyrrolidone, poloxamer, and hydroxypropyl methylcellulose, the prepared microneedles all have a pyramid shape with a sharp tip, and the mechanical strength of the microneedles shows that they have good skin penetration ability.

[0017] The needle fluid is only composed of high-concentration semaglutide, which can greatly increase the drug loading of semaglutide in the microneedle drug delivery system, overcoming the disadvantages of low drug loading capacity of ordinary microneedle patches and insufficient semaglutide delivery through microneedles. The patch can effectively control blood sugar in healthy mice; in the diabetic mouse model, the semaglutide microneedle patch of the present invention can achieve a similar hypoglycemic effect as subcutaneous injection at the same dosage, providing a new dosage form option for pharmaceutical uses such as semaglutide hypoglycemic and weight loss.

[0018] Attached photos

[0019] Figure 1 Morphology of the semaglutide soluble microneedle patches prepared in Examples 1 to 6;

[0020] Figure 2 SEM images of semaglutide soluble microneedle patches prepared in Examples 1, 3, and 6;

[0021] Figure 3 Mechanical strength of the semaglutide soluble microneedle patches prepared in Examples 1 to 6;

[0022] Figure 4 In vitro pig skin puncture performance testing of the semaglutide soluble microneedle patches prepared in Examples 1 to 6;

[0023] Figure 5 Glucose tolerance test in mice with microneedle patches;

[0024] Figure 6 Test of hypoglycemic effect in diabetic mice. DETAILED DESCRIPTION

[0025] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0026] In the following examples, the purity of Semaglutide is greater than 99%, and the preparation method refers to Example 4 of patent WO2006 / 097537; HA has an average molecular weight of 8.39 kDa and was purchased from Bloomage Biotechnology Co., Ltd.; pharmaceutical grade mannitol was purchased from Shanghai Fenghong Pharmaceutical Excipients Technology Co., Ltd.; PDMS microneedle template was purchased from Micropoint Technologies Pte Ltd; unless otherwise specified, all are common commercial products and can be purchased on the market.

[0027] The present invention will be further described below by way of examples, which are not intended to further limit the present invention. It should be understood by those skilled in the art that equivalent replacements or corresponding improvements made to the present invention still fall within the scope of protection of the present invention.

[0028] Example 1

[0029] Dissolve 2.5 mg of semaglutide in 797.5 mg of purified water, add 50 mg of mannitol and 150 mg of sodium hyaluronate, stir and dissolve to obtain the needle fluid. Add 150 mg of sodium hyaluronate to 850 mg of purified water, stir and dissolve to obtain the backing layer solution. The needle fluid and backing layer solution are degassed by centrifugation at 5000 rpm for 10 min. Pipette 30 μl of needle fluid and spread it on the surface of the PDMS microneedle mold, centrifuge at 2236 × g for 5 min, cycle twice, absorb excess needle fluid, and dry in an oven at 30 ° C for 3 h. Pipette 70 μl of backing layer solution and spread it on the surface of the mold, centrifuge at 2236 × g for 5 min, cycle twice, add 30 μl of backing layer solution, place it in an oven at 30 ° C for 24 h, and then demold it to obtain a soluble semaglutide layered microneedle patch.

[0030] Example 2

[0031] Dissolve 50 mg of semaglutide in 797.5 mg of purified water, add 38.125 mg of mannitol and 114.375 mg of sodium hyaluronate, stir and dissolve to obtain the needle fluid. Add 150 mg of sodium hyaluronate to 850 mg of purified water, stir and dissolve to obtain the backing layer solution. The needle fluid and backing layer solution are degassed by centrifugation at 5000 rpm for 10 min. Absorb 30 μl of needle fluid and spread it on the surface of the PDMS microneedle mold, centrifuge at 2236 × g for 5 min, cycle twice, absorb excess needle fluid, and dry in an oven at 30 ° C for 3 h. Absorb 70 μl of backing layer solution and spread it on the surface of the mold, centrifuge at 2236 × g for 5 min, cycle twice, add 30 μl of backing layer solution, place it in an oven at 30 ° C for 24 h, and then demold it to obtain a soluble semaglutide layered microneedle patch.

[0032] Example 3

[0033] Dissolve 100 mg of semaglutide in 797.5 mg of purified water, add 25.625 mg of mannitol and 76.875 mg of sodium hyaluronate, stir and dissolve to obtain needle fluid. Add 150 mg of sodium hyaluronate to 850 mg of purified water, stir and dissolve to obtain the backing layer solution. The needle fluid and backing layer solution are degassed by centrifugation at 5000 rpm for 10 min. Pipette 30 μl of needle fluid and spread it on the surface of the PDMS microneedle mold, centrifuge at 2236 × g for 5 min, cycle twice, absorb excess needle fluid, and dry in an oven at 30 ° C for 3 h. Pipette 70 μl of backing layer solution and spread it on the surface of the mold, centrifuge at 2236 × g for 5 min, cycle twice, add 30 μl of backing layer solution, place it in an oven at 30 ° C for 24 h, and then demold it to obtain a soluble semaglutide layered microneedle patch.

[0034] Example 4

[0035] Dissolve 150 mg of semaglutide in 797.5 mg of purified water, add 13.125 mg of mannitol and 39.375 mg of sodium hyaluronate, stir and dissolve to obtain needle fluid. Add 150 mg of sodium hyaluronate to 850 mg of purified water, stir and dissolve to obtain the backing layer solution. The needle fluid and backing layer solution are degassed by centrifugation at 5000 rpm for 10 min. Absorb 30 μl of needle fluid and spread it on the surface of the PDMS microneedle mold, centrifuge at 2236 × g for 5 min, cycle twice, absorb excess needle fluid, and dry in an oven at 30 ° C for 3 h. Absorb 70 μl of backing layer solution and spread it on the surface of the mold, centrifuge at 2236 × g for 5 min, cycle twice, add 30 μl of backing layer solution, place it in an oven at 30 ° C for 24 h, and then demold it to obtain a soluble semaglutide layered microneedle patch.

[0036] Example 5

[0037] Dissolve 202.5 mg of semaglutide in 797.5 mg of purified water, stir and dissolve to obtain needle fluid. Add 150 mg of sodium hyaluronate to 850 mg of purified water, stir and dissolve to obtain the backing layer solution. Centrifuge the needle fluid and backing layer solution at 5000 rpm for 10 min to degas. Pipette 30 μl of needle fluid and spread it on the surface of the PDMS microneedle mold, centrifuge at 2236 × g for 5 min, cycle twice, absorb excess needle fluid, and dry in an oven at 30°C for 3 h. Pipette 70 μl of backing layer solution and spread it on the surface of the mold, centrifuge at 2236 × g for 5 min, cycle twice, add 30 μl of backing layer solution, place in an oven at 30°C to dry for 24 h, and then demold to obtain a soluble semaglutide layered microneedle patch.

[0038] Example 6

[0039] Dissolve 400 mg of semaglutide in 600 mg of purified water, stir and dissolve to obtain the needle body fluid. Add 150 mg of sodium hyaluronate to 850 mg of purified water, stir and dissolve to obtain the backing layer solution. Centrifuge the needle body fluid and backing layer solution at 5000 rpm for 10 min to degas. Pipette 30 μl of needle body fluid and spread it on the surface of the PDMS microneedle mold, centrifuge at 2236 × g for 5 min, cycle twice, absorb excess needle body fluid, and dry in an oven at 30°C for 3 h. Pipette 70 μl of backing layer solution and spread it on the surface of the mold, centrifuge at 2236 × g for 5 min, cycle twice, add 30 μl of backing layer solution, place it in an oven at 30°C to dry for 24 h, and then demold it to obtain a soluble semaglutide layered microneedle patch.

[0040] Table 1 Summary of the mass percentage of the needle fluid and the backing layer solution in Examples 1-6

[0041]

[0042]

[0043] Example 7

[0044] The morphology of the microneedle patch was observed using a stereo microscope (DVM6A, Leica, Germany) and a scanning electron microscope (SEM, JSM-IT200, JEOL, Japan). In the preparation of the semaglutide microneedle patch in Examples 1-4, sodium hyaluronate (HA) was used as the only skeleton material for the needle body and the backing layer, and mannitol was the protective agent for semaglutide; in the preparation of the semaglutide microneedle patch in Examples 5-6, sodium hyaluronate (HA) was used as the only skeleton material for the backing layer, and the needle body fluid contained only semaglutide. Microscope images of the prepared semaglutide microneedle patch are shown in the attached figure. Figure 1 As shown, it can be seen that the microneedle patches prepared with different ratios of main and auxiliary materials all have good morphology, complete needle bodies and sharp needle tips.

[0045] The SEM images of the semaglutide soluble microneedle patches prepared in Examples 1, 3, and 6 are shown in the attached figure. Figure 2 As shown, it can be seen that the microneedles prepared with different ratios of semaglutide to skeleton materials all have a pyramid shape with a sharp tip. It is worth mentioning that the sharpness of the microneedle is not affected by the reduction of the skeleton material content. Even if the skeleton material is removed, the good shape of the microneedle can be maintained by relying on the moldability of semaglutide itself, which is of breakthrough significance for improving drug loading capacity.

[0046] Example 8

[0047] The mechanical strength of the semaglutide soluble microneedles prepared in Examples 1 to 6 was analyzed using a geoanalyzer (TA-XT, Stable Micro Systems, Britain): the microneedle patch was placed on the stage of the geoanalyzer with the needle tip facing vertically upward. A P / 5 probe was used to press the needle tip and record the force-distance curve when the probe moved. The moving speed of the probe was set to 0.01 mm / s. The results are shown in the attached figure. Figure 3 shown.

[0048] Characterization of the skin puncture performance of the semaglutide soluble microneedle patch prepared in Example 1-6 using ex vivo pig skin: The pig skin of Bama Xiang pig was cut into squares, washed with saline to remove surface fat, and dried with absorbent paper for further use. The microneedle patch containing trypan blue dye in the needle body (prepared in the same way as in Example 1-6) was pressed vertically into the skin with fingers. After 5 minutes, the patch was removed and the skin was observed using a stereomicroscope. The results are shown in the attached figure. Figure 4 shown.

[0049] The mechanical strength of microneedles is one of the most important properties and plays a decisive role in effective skin puncture and transdermal drug delivery. Figure 3 The results of mechanical strength analysis show that the microneedle patches prepared in Examples 1-6 have similar mechanical strengths. When the displacement is less than 300 μm, which is half of the theoretical needle height, no inflection point is observed in the force / displacement curve, which indicates that the needle is not easy to break. As the displacement further increases to 400 μm and 500 μm, slower force changes appear in the force / displacement curves of the microneedle patches of Examples 5 and 6, indicating that slight fractures occur at large deformations of the needles. The mechanical strength of the microneedle patch without sodium hyaluronate under large deformation is lower than that of other formulations, and the force at 50% deformation (300 μm) is still greater than 21N / 85 needles. Therefore, the microneedle patches prepared in Examples 1 to 6 all have good skin penetration capabilities.

[0050] The skin penetration ability of the microneedle patches was further evaluated using Bama Xiang pig skin, which has a similar structure to the human body. The results showed that all the microneedle patches had good skin penetration ability. The microneedle patches prepared in Examples 1 to 3 had a penetration efficiency of almost 100%, while the microneedle patches prepared in Examples 4 to 6 had a slightly lower penetration ability, but still had a penetration efficiency of more than 95%.

[0051] Example 9

[0052] The drug loading of semaglutide in the microneedle patch was determined by high performance liquid chromatography (HPLC, 1260VWD, Agilent, USA) and external standard method: the accurately weighed semaglutide standard was dissolved in 10mmol / L phosphate buffer (PBS) at pH 7.5 as a reference solution, and the microneedle patches prepared in Example 1 and Example 6 were dissolved in an appropriate volume of 10mmol / L PBS at pH 7.5 to prepare a sample solution with a concentration of not less than 5μg / mL. A C4 chromatographic column (150mm×4.6mm) with a filler diameter of 3μm was used, and a gradient elution method was adopted. The mobile phase A was a mixture of 0.1mol / L diammonium phosphate solution and acetonitrile in a volume ratio of 90:10, and the mobile phase B was a mixture of acetonitrile and ultrapure water in a volume ratio of 80:20. The column temperature was set to 30°C, the mobile phase flow rate was 1.0mL / min, the injection volume was 20μL, and the ultraviolet detection wavelength was set to 215nm. The test results are shown in the following table:

[0053] sample Example 1 Example 6 Drug loading (7.85±1.01)μg / tablet (1158.86±33.92)μg / tablet

[0054] In Example 1, when the mass ratio of semaglutide in the needle fluid was 0.25%, the drug loading was only (7.85±1.01) μg / tablet. In Example 6, when the mass ratio of semaglutide in the needle fluid was increased to 40% without adding HA and mannitol, the drug loading increased significantly to (1158.86±33.92) μg / tablet, which can meet the dosage requirements of semaglutide in clinical use.

[0055] Example 10

[0056] Healthy mice aged 6-8 weeks were used to evaluate glucose tolerance after using the microneedle patch: After the healthy mice fasted for 12 hours, the microneedle patch prepared in Example 6 (the needle body contained 5.5 μg of semaglutide) was applied to the back skin of the mice, removed after 4 hours, and injected with glucose intraperitoneally at a dose of 3 g / kg. Blood glucose was tested with a blood glucose meter at 15 minutes, 30 minutes, 1 hour and 2 hours after the injection of glucose. A blank microneedle patch was used as a control. The preparation method of the blank microneedle patch was referred to Example 4, except that the needle body did not contain semaglutide. The changes in blood glucose concentration are shown in the attached figure. Figure 5 shown.

[0057] 0.25h after intraperitoneal injection of glucose in the control mice, the blood glucose concentration increased from 5.45±0.64mmol / L to 20.17±3.30mmol / L, and the rate of change of blood glucose concentration from baseline was (278.57±109.45)%. In contrast, 0.25 hours after intraperitoneal injection of glucose in mice treated with microneedle patches, the blood glucose concentration increased from 3.30±0.93mmol / L to 8.13±2.21mmol / L, and the rate of change from baseline was only (169.87±125.02)%, which was significantly lower than that of the control group. In addition, when the glucose solution was injected intraperitoneally again, the blood glucose control effect was still observed 24 hours after microneedle patch treatment. Therefore, it can be inferred that semaglutide in the microneedle patch was successfully delivered and maintained its biological activity after being prepared into a microneedle patch.

[0058] Embodiment 11

[0059] The db / db mice and STZ-induced diabetic mice were used as models to evaluate the blood sugar control ability of the semaglutide microneedle patch. After the mice fasted for 12 hours, the microneedle patch prepared in Example 6 (the needle body contained 5.5 μg of semaglutide) was applied to the back skin of the mice, and mice subcutaneously injected with semaglutide at a dose of 5.5 μg / mouse were used as positive controls. Untreated mice were used as negative controls. Blood sugar was measured 15 minutes, 30 minutes, 1 hour, 2 hours, 4 hours and 8 hours after the application of the microneedle patch, and the changes in blood sugar concentration are shown in the attached figure. Figure 6 shown.

[0060] Four hours after subcutaneous injection of semaglutide in db / db mice, blood glucose concentration continued to decrease from 29.5±3.76mmol / L to 3.95±0.68mmol / L. In mice treated with semaglutide microneedle patch, blood glucose concentration hardly changed before 0.5 hours, and began to decrease after 0.5 hours. There was no significant difference compared with the control group before 4 hours. However, 4 hours after administration, blood glucose concentration in the semaglutide microneedle patch treatment group was significantly lower than that in the control group. Compared with the baseline, the percentage change of blood glucose concentration in the semaglutide microneedle patch treatment group reached -(62.14±3.37)%.

[0061] After subcutaneous injection of semaglutide solution in STZ-induced diabetic mice, the blood glucose concentration dropped from 24.14±2.80mmol / L to 12.74±4.91mmol / L after 8 hours, with a change rate of -(45.16±26.77)%. The fasting blood glucose of mice with microneedle patches was 28.77±3.49mmol / L. After 8 hours of microneedle patch application, blood glucose dropped to 12.40±3.33mmol / L, with a change rate of -(55.87±15.15)%. The blood glucose concentration change trend of the microneedle patch treatment group and the subcutaneous injection group was similar, and the blood glucose concentrations of the microneedle patch treatment group and the subcutaneous injection group were significantly lower than those of the control group. In summary, compared with subcutaneous injection, the semaglutide microneedle patch has a similar blood glucose control effect, which indicates that semaglutide can be effectively delivered, and the delivery dosage is equivalent to subcutaneous injection, which can produce corresponding physiological functions.

Claims

1. A semaglutide soluble microneedle patch composition, characterized in that: The soluble microneedle patch composition comprises a needle body material and a backing layer material; the needle body material comprises semaglutide.

2. The soluble microneedle patch composition according to claim 1, characterized in that: The needle solution is composed of a semaglutide solution, wherein the mass percentage concentration of the semaglutide is 20% to 40%.

3. The soluble microneedle patch composition according to claim 1, characterized in that: The needle body material comprises semaglutide and sodium hyaluronate, wherein the mass percentage concentration of semaglutide in the needle body solution is 0.25% to 15%, and the mass percentage concentration of sodium hyaluronate is 4% to 15%.

4. The soluble microneedle patch composition according to claim 3, characterized in that: The needle body material also includes mannitol, and the mass percentage concentration in the needle body solution is 1% to 5%.

5. The soluble microneedle patch composition according to claim 4, characterized in that: The needle body material also includes at least one of polyvinyl pyrrolidone, poloxamer, and hydroxypropyl methylcellulose.

6. The soluble microneedle patch composition according to any one of claims 1 to 5, characterized in that: The backing layer material includes sodium hyaluronate.

7. The soluble microneedle patch composition according to claim 6, characterized in that: The backing layer material is composed of sodium hyaluronate, and the mass percentage concentration of sodium hyaluronate in the backing layer solution is 15%.

8. The soluble microneedle patch composition according to claim 6, characterized in that: The backing layer material also includes polyvinyl pyrrolidone.

9. The method for preparing the soluble microneedle patch composition according to any one of claims 1 to 8, characterized in that: The following steps are involved: S1: preparing needle body solution and backing layer solution; S2: adding the needle solution into the microneedle negative mold, centrifuging, and drying; S3: adding the backing layer solution, centrifuging, and then drying and demoulding to obtain the soluble microneedle patch composition.

10. The preparation method according to claim 9, characterized in that: The centrifugation step is specifically centrifugation at a centrifugal force of 2236 g for 5 minutes.

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