Patent Application
20210085606
The technical field relates to a powder form, and in particular, it relates to a powder form having a therapeutic agent for the use of slowing joint degeneration and repairing tendon rupture.
Degeneration is an unavoidable process that everyone will experience during their lifetime. Degeneration is often accompanied by pain and inflammation, thereby reducing the quality of life of patients. Common degenerative diseases caused by aging or excessive use are including degenerative arthritis and tendonopathy. Degenerative arthritis of the knee joint can cause cartilage wear, inflammation, and deformation of the joints, thereby causing significant pain and inconvenience to the patient.
Methods used clinically for treating degenerative diseases include hyaluronic acid injection, which injects a mucous-like macromolecular-hyaluronic-acid-containing agent, serving as a lubricant, into the knee joint to prevent joints or tendons from wearing, to inhibit inflammation, to stimulate secretion of synovial fluid, and to reduce joint pain. The declared effects, however, have not yet been supported by strong scientific evidence. It should be noted that the injected hyaluronic-acid can be spread over the non-affected region due to the fluidity of hyaluronic acid, resulting in a reduced therapeutic efficacy.
Therefore, there is a need to develop a novel slow-releasing therapeutic agent, which can be attached to defects or lesions of the joints or tendons, for use in treatment of joints and tendons.
According to embodiments of the disclosure, the disclosure provides a powder form, wherein the powder form consists essentially of 120-380 parts by weight of biodegradable copolymer, 15-75 parts by weight of urea, and 100 parts by weight of platelet-rich plasma (PRP). The biodegradable copolymer has a structure of Formula (I) or Formula (II):
wherein A is a hydrophilic polyethylene glycol polymer; B is a hydrophobic polyester polymer; BOX is a bifunctional group monomer of 2, 2′-bis(2-oxazoline) used for coupling the blocks A-B or B-A-B; and n is 0 or an integer greater than 0.
According to embodiments of the disclosure, the disclosure further provides a use of hydrogel composition for alleviating degenerative joint and tendon tear, wherein the hydrogel composition includes 100 parts by weight of therapeutic agent and 120-380 parts by weight of biodegradable copolymer. The therapeutic agent includes platelet-rich plasma (PRP), doxorubicin, transforming growth factor, or a combination thereof, wherein the biodegradable copolymer has a structure of Formula (I) or Formula (II):
wherein A is a hydrophilic polyethylene glycol polymer; B is a hydrophobic polyester polymer; BOX is a bifunctional group monomer of 2, 2′-bis(2-oxazoline) used for coupling the blocks A-B or B-A-B ; and a is 0 or an integer greater than 0.
A detailed description is given in the following embodiments with reference to the accompanying drawings.
The disclosure provides a powder form, wherein the powder form consists essentially of 120-380 parts by weight of biodegradable copolymer, 15-75 parts by weight of urea, and 100 parts by weight of platelet-rich plasma (PRP). The biodegradable copolymer has a structure of Formula (I) or Formula (II):
wherein A is a hydrophilic polyethylene glycol polymer; B is a hydrophobic polyester polymer; BOX is a bifunctional group monomer of 2, 2′-bis(2-oxazoline) used for coupling the blocks A-B or B-A-B; and n is 0 or an integer greater than 0.
According to embodiments of the disclosure, A can be polyethylene glycol (PEG), or methoxy-polyethylene glycol (mPEG).
According to embodiments of the disclosure, B can be poly(lactide-co-glycolide) (PLGA), poly(propionic-co-lactic (PPLA), poly(valeric-co -lactic) (PVLA), or poly(caproic-co-lactic (PCLA). The hydrophobic polyester polymer can have a molecular weight of 500-5000 g/mole. The term molecular weight disclosed in the disclosure means weight average molecular weight.
The biodegradable copolymer can be PEG-PLGA, PEG-PLGA-PEG, PLGA-PEG-PLGA, or combinations thereof. The term PEG means a hydrophilic polyethylene glycol (PEG) polymer, such as polyethylene glycol (PEG), or methoxy-poly(ethylene glycol) (mPEG)(methoxy-poly(ethylene glycol), mPEG). The hydrophilic polyethylene glycol (PEG) polymer can have a molecular weight of between 350-2000 g/mole. The term PLGA means hydrophobic poly(lactic-co-glycolic acid), and can be derived from D,L-Lactide, D-Lactide, L-Lactide, D,L-Lactic acid, D-Lactic acid, L-Lactic acid, glycolide, β-propiolactone, δ-valerolactone, or ε-caprolactone, such as poly(lactide-co-glycolide) (PLGA), poly(propionic-co-lactic (PPLA), poly(valeric-co -lactic) (PVLA), or poly(caproic-co-lactic) (PCLA). The hydrophobic poly(lactic-co-glycolic acid) can have a molecular weight of between 1000-3500 g/mole.
According to embodiments of the disclosure, the disclosure provides a use of hydrogel composition for alleviating degenerative joint and tendon tear, wherein the hydrogel composition includes 100 parts by weight of therapeutic agent and 120-380 parts by weight of biodegradable copolymer, wherein the therapeutic agent includes platelet-rich plasma (PRP), doxorubicin, transforming growth factor (TGF-b1), bovine serum albumin, or a combination thereof. The biodegradable copolymer has a structure of Formula (I) or Formula (II):
wherein A is a hydrophilic polyethylene glycol polymer; B is a hydrophobic polyester polymer; BOX is a bifunctional group monomer of 2, 2′-bis(2-oxazoline) used for coupling the blocks A-B or B-A-B; and n is 0 or an integer greater than 0.
According to embodiments of the disclosure, A can be polyethylene glycol (PEG), or methoxy-poly(ethylene glycol (mPEG).
According to embodiments of the disclosure, B can be poly(lactide-co-glycolide) (PLGA), poly(propionic-co-lactic (PPLA), poly(valeric-co -lactic) (PVLA), or poly(caproic-co-lactic (PCLA).
According to embodiments of the disclosure, the therapeutic agent of the disclosure includes platelet-rich plasma (PRP) and doxorubicin, wherein the weight ratio of the platelet-rich plasma (PRP) to the doxorubicin is 1:2.
According to embodiments of the disclosure, the therapeutic agent of the disclosure includes platelet-rich plasma (PRP) and transforming growth factor, wherein the weight ratio of the platelet-rich plasma (PRP) to the transforming growth factor is 1:2.
According to embodiments of the disclosure, the therapeutic agent of the disclosure includes platelet-rich plasma (PRP) and bovine serum albumin, wherein the weight ratio of the platelet-rich plasma (PRP) to the bovine serum albumin is 1:2.
According to embodiments of the disclosure, the use of hydrogel composition for alleviating degenerative joint and tendon tear of the disclosure further includes an ultrasonic treatment, which forces the hydrogel composition to release the therapeutic agent. In addition, the output intensity of the ultrasonic treatment can be 100-10000 W/cm2. The ultrasonic treatment has a treatment period from 1 min to 5 min. Under the ultrasound stimulation, the release amount of the therapeutic agent can be increased.
According to embodiments of the disclosure, the use of hydrogel composition for alleviating degenerative joint and tendon tear of the disclosure, the hydrogel composition further includes 50-400 parts by weight of water.
Below, exemplary embodiments will be described in detail so as to be easily realized by a person having ordinary knowledge in the art. The inventive concept may be embodied in various forms without being limited to the exemplary embodiments set forth herein. Descriptions of well-known parts are omitted for clarity.
First, 10.04 g of mPEG (methoxy poly(ethylene glycol) (with a molecular weight of 550 g/mole), 20 g of lactide, and 5.64 g of glycolide were subsequently added into a reactor, and the reaction bottle was heated slowly to force that the components were completely dissolved in the solvent. After heating to 160° C., the stannous 2-ethyl-hexanoate (14.0 μl) as catalyst was added into the reaction bottle. After heating for 8 hr, poly(lactide-co-glycolide) (PLGA) was obtained via polymerization of lactide and glycolide. After the reaction was completed, 1.84 g of succinic anhydride (SA) (with a molecular weight of 100.07 g/mole) was added into the reaction bottle. Next, after reacting for 4 hr, 1.28 g of 2, 2′-bis(2-oxazoline) (BOX) (with a molecular weight of 140.14 g/mole) was added into the reaction bottle. After the components were completely dissolved in the solvent, stannous octoate as catalyst was added into the reaction bottle. After reacting for 4 hr, the result (translucent gel) was reprecipitated with a solution (including diethyl ether and n-hexane, and the volume ratio of diethyl ether to n-hexane is 1:9). The result was washed three times to remove the residual monomers and dried in a vacuum for 24 hr at a temperature of 40° C., thus obtaining mPEG-PLGA biblock polymer.
100 mL, of pig blood was treated with a platelet rich plasma extraction kit (manufactured by Biosafe) and a cell separation system (manufactured by Sepax) (with a centrifugation time of 15 min and a centrifugation speed of 3400 rpm), obtaining a platelet-rich plasma (PRP).
A solution including 0.5 mL of mPEG-PLGA biblock polymer of Preparation Example 1 and urea was cooled to −20° C. to form a powder form, wherein the urea concentration of the powder form was 2%.
The platelet-rich plasma prepared by Preparation Example 1 served as 100% PRP sample solution.
The powder form of Preparation Example was dispersed in a microcentrifuge tube, and 0.5 mL of 100% PRP sample solution of Preparation Example 3 was added into the tube to mix with the powder from via a tube oscillator. After standing for 20 min and then resolving, 100% PRP sample solution (including the mPEG-PLGA biblock polymer) was obtained.
1 mL of 100% PRP sample solution (without the mPEG-PLGA biblock polymer) of preparation Example 4 was diluted with 1 mL of phosphate buffered saline (PBS), obtaining 50% PRP sample solution (without the mPEG-PLGA biblock polymer).
100 mg of powder form of Preparation Example 3 was dispersed in a microcentrifuge tube, and 0.5 mL of 50% PRP sample solution of Preparation Example 5 was added into the tube to mix with the powder from via a tube oscillator. After standing for 20 min and then resolving, 50% PRP sample solution (including the mPEG-PLGA biblock polymer) was obtained.
First, 0.1 mL of 100% PRP sample solution 120 (without the mPEG-PLGA biblock polymer) of Preparation Example 4 was added into a multiple well plate 110. After the sample solution 140 was transferred into a gel, human dermal fibroblasts cells (HDF) 160 were implanted in the cell culture plate 170. 0.6 mL of medium (DMEM) 150 was added. Next, the multiple well plate 110 was hung on the cell culture plate 170, and the co-culture was performed at 37° C. (as shown in
First, 0.1 mL of 100% PRP sample solution (including the mPEG-PLGA biblock polymer) 120 of Preparation Example 5 was added into a multiple well plate 110. After the sample solution 140 was transferred into a gel, human dermal fibroblasts cells (HDF) 160 were implanted in the cell culture plate 170. 0.6 mL of medium (DMEM) 150 was added. Next, the multiple well plate 110 was hung on the cell culture plate 170, and the co-culture was performed at 37° C. (as shown in
First, 0.1 mL of 50% PRP sample solution 120 (without the mPEG-PLGA biblock polymer) of Preparation Example 6 was added into a multiple well plate 110. After the sample solution 140 was transferred into a gel, human dermal fibroblasts cells (HDF) 160 were implanted in the cell culture plate 170. 0.6 mL of medium (DMEM) 150 was added. Next, the multiple well plate 110 was hung on the cell culture plate 170, and the co-culture was performed at 37° C. (as shown in
First, 0.1 mL of 50% PRP sample solution 120 (including the mPEG-PLGA biblock polymer) of Preparation Example 7 was added into a multiple well plate 110. After the sample solution 140 was transferred into a gel, human dermal fibroblasts cells (HDF) 160 were implanted in the cell culture plate 170. 0.6 mL of medium (DMEM) 150 was added. Next, the multiple well plate 110 was hung on the cell culture plate 170, and the co-culture was performed at 37° C. (as shown in
First, 0.1 mL of 100% PRP sample solution (without the mPEG-PLGA biblock polymer) of Preparation Example 4 was added into a multiple well plate. After the sample solution was transferred into a gel, 0.6 mL of 0.6 mL of medium (DMEM) was implanted in the cell culture plate. Next, the multiple well plate 110 was hung on the cell culture plate 170. The transforming growth factor (TGF-b1) released from PRP was slowly penetrated into the cell culture plate via the hole (with a diameter of 0.4 μm) in the bottom of the multiple well plate. After the reaction was completed, 0.6 mL of culture fluid was sampled and analyzed.
First, 0.1 mL of 100% PRP sample solution (including the mPEG-PLGA biblock polymer) of Preparation Example 5 was added into a multiple well plate. After the sample solution was transferred into a gel, 0.6 mL, of medium (DMEM) was implanted in the cell culture plate. Next, the multiple well plate 110 was hung on the cell culture plate 170. The transforming growth factor (TGF-b1) released from PRP was slowly penetrated into the cell culture plate via the hole (with a diameter of 0.4 μm) in the bottom of the multiple well plate. After the reaction was completed, 0.6 mL of culture fluid was sampled and analyzed.
First, 0.1 mL of 50% PRP sample solution (without the mPEG-PLGA biblock polymer) of Preparation Example 6 was added into a multiple well plate. After the sample solution was transferred into a gel, 0.6 mL of medium (DMEM) was implanted in the cell culture plate. Next, the multiple well plate 110 was hung on the cell culture plate 170. The transforming growth factor (TGF-b1) released from PRP was slowly penetrated into the cell culture plate via the hole (with a diameter of 0.4 μm) in the bottom of the multiple well plate. After the reaction was completed, 0.6 mL of culture fluid was sampled and analyzed.
First, 0.1 mL of 50% PRP sample solution (including the mPEG-PLGA biblock polymer) of Preparation Example 7 was added into a multiple well plate. After the sample solution was transferred into a gel, 0.6 mL of medium (DMEM) was implanted in the cell culture plate. Next, the multiple well plate 110 was hung on the cell culture plate 170. The transforming growth factor (TGF-b1) released from PRP was slowly penetrated into the cell culture plate via the hole (with a diameter of 0.4 μm) in the bottom of the multiple well plate. After the reaction was completed, 0.6 mL of culture fluid was sampled and analyzed.
0.6 mL of culture fluid was extracted from the cell culture plates of Preparation Examples 12-15 individually after reacting 24 hr, and then stored at −20° C. Next, 0.6 mL of medium (DMEM) was implanted in the cell culture plate of Preparation Examples 12-15, and then 0.6 mL of culture fluid was extracted from the cell culture plates of Preparation Examples 12-15 individually after reacting 24 hr, and then stored at −20° C. The aforementioned steps were repeated until 20 days. Finally, the release amount of the TGF-b1 of the culture fluids were determined by enzyme-linked immunosorbent assay (ELISA), and the results are shown in
As shown in
0.2 mL of culture fluid was extracted from the cell culture plates of Preparation Examples 8-11 individually after reacting 24 hr. Next, 0.2 mL of medium (DMEM) was implanted in the cell culture plate of Preparation Examples 8-11. After reacting another day, the agent (alamar blue assay, sold by Bio-Rad) was added into the cell culture plate. After reacting, the human dermal fibroblasts cell (HDF) proliferation rate of the result was determined by measuring absorbance at 570 nm by a spectrophotometer.
0.2 mL of culture fluid was extracted from the cell culture plates of Preparation Examples 8-11 individually after reacting 24 hr. Next, 0.2 mL of medium (DMEM) was implanted in the cell culture plate of Preparation Examples 8-11. The aforementioned steps were repeated until 5 days. The agent (alamar blue assay, sold by Bio-Rad) was added into the cell culture plate. After reacting, the human dermal fibroblasts cell (HDF) proliferation rate of the result was determined by measuring absorbance at 570 nm by a spectrophotometer.
0.2 mL of culture fluid was extracted from the cell culture plates of Preparation Examples 8-11 individually after reacting 24 hr. Next, 0.2 mL of medium (DMEM) was implanted in the cell culture plate of Preparation Examples 8-11. The aforementioned steps were repeated until 5 days. The agent (alamar blue assay, sold by Bio-Rad) was added into the cell culture plate. After reacting, the human dermal fibroblasts cell (HDF) proliferation rate of the result was determined by measuring absorbance at 570 nm by a spectrophotometer. In the meantime, the clear supernatant liquid of culture fluid was extracted, and the concentration of TGF-b1 of the clear supernatant liquid was determined by enzyme-linked immunosorbent assay (ELISA), and the result is shown in
As shown in
0.6 mL of culture fluid was extracted from the cell culture plates of Preparation Examples 12-15 individually after reacting 24 hr, and then stored at −20° C. Next, 0.6 mL of medium (DMEM) was implanted in the cell culture plate of Preparation Examples 12-15, and then 0.6 mL of culture fluid was extracted from the cell culture plates of Preparation Examples 12-15 individually after reacting 24 hr, and then stored at −20° C. The aforementioned steps were repeated until 13 days (analyzed at the second, fourth, sixth, eighth, ninth, twelfth and thirteenth days). After implanting, human dermal fibroblasts cell (HDF) into a 96 well plate, the above sample solutions (0.1 mL) were added into the 96 well plate individually. After standing at a cell incubator for 72 hr, the agent (alamar blue assay) was added into the 96 well plate. The human dermal fibroblasts cell (HDF) proliferation rate of the results were determined by measuring absorbance at 570 nm by a spectrophotometer, and the results are shown in
As shown in
50 mg of mPEG-PLGA-BOX-PLGA-mPEG polymer, 6.6 mg of urea, and 34.4 mg of 100% PRP sample solution (without the mPEG-PLGA biblock polymer) were dissolved in water and stirred at 4° C., obtaining a solution. Next, the aforementioned solution was cooled to −20° C. to form a solid mixture. Finally, the solid mixture was subjected to a freeze-drying process at −20° C. under 8 mTorr for 2 days, obtaining a powder form. The powder form was stored at room temperature and −20° C. individually. After two weeks, the powder forms (stored at room temperature and stored −20° C.) were dissolved in 409 mg of water individually, and then the release amount of the TGF-b1 of the results was determined by enzyme-linked immunosorbent assay (ELISA, and the results are shown in
50 mg of mPEG-PLGA-BOX-PLGA-mPEG polymer, 8.3 mg of urea, and 34.4 mg of 100% PRP sample solution (without the mPEG-PLGA biblock polymer) were dissolved in water and stirred at 4° C., obtaining a solution. Next, the aforementioned solution was cooled to −20° C. to form a solid mixture. Finally, the solid mixture was subjected to a freeze-drying process at −20° C. under 8 mTorr for 2 days, obtaining a powder form. The powder form was stored at room temperature and −20° C. individually. After two weeks, the powder forms (stored at room temperature and stored at −20° C.) were dissolved in 407.3 mg of water individually, and then the release amount of the TGF-b1 of the results was determined by enzyme-linked immunosorbent assay (ELISA), and the results are shown in
50 mg of mPEG-PLGA-BOX-PLGA-mPEG polymer, 10 mg of urea, and 34.4 mg of 100% PRP sample solution (without the mPEG-PLGA biblock polymer) were dissolved in water and stirred at 4° C., obtaining a solution. Next, the aforementioned solution was cooled to −20° C. to form a solid mixture. Finally, the solid mixture was subjected to a freeze-drying process at −20°C. under 8 mTorr for 2 days, obtaining a powder form. The powder form was stored at room temperature and −20° C. individually. After two weeks, the powder forms (stored at room temperature and stored at −20° C.) were dissolved in 405.6 mg of water individually, and then the release amount of the TGF-b1 of the results was determined by enzyme-linked immunosorbent assay (ELISA), and the results are shown in
75 mg of mPEG-PLGA-BOX-PLGA-mPEG polymer, 10 mg of urea, and 34.4 mg of 100% PRP sample solution (without the mPEG-PLGA biblock polymer) were dissolved in water and stirred at 4° C., obtaining a solution. Next, the aforementioned solution was cooled to −20° C. to form a solid mixture. Finally, the solid mixture was subjected to a freeze-drying process at −20° C. under 8 mTorr for 2 days, obtaining a powder form. The powder form was stored at room temperature and −20° C. individually. After two weeks, the powder forms (stored at room temperature and stored at −20° C.) were dissolved in 380.6 mg of water individually, and then the release amount of the TGF-b1 of the results was determined by enzyme-linked immunosorbent assay (ELISA), and the results are shown in
75 mg of mPEG-PLGA-BOX-PLGA-mPEG polymer, 12.5 mg of urea, and 34.4 mg of 100% PRP sample solution (without the mPEG-PLGA biblock polymer) were dissolved in water and stirred at 4° C., obtaining a solution. Next, the aforementioned solution was cooled to −20° C. to form a solid mixture. Finally, the solid mixture was subjected to a freeze-drying process at −20° C. under 8 mTorr for 2 days, obtaining a powder form. The powder form was stored at room temperature and −20° C. individually. After two weeks, the powder forms (stored at room temperature and stored at −20° C.) were dissolved in 378.1 mg of water individually, and then the release amount of the TGF-b1 of the results was determined by enzyme-linked immunosorbent assay (ELISA), and the results are shown in
75 mg of mPEG-PLGA-BOX-PLGA-mPEG polymer, 15 mg of urea, and 34.4 mg of 100% PRP sample solution (without the mPEG-PLGA biblock polymer) were dissolved in water and stirred at 4° C., obtaining a solution. Next, the aforementioned solution was cooled to −20° C. to form a solid mixture. Finally, the solid mixture was subjected to a freeze-drying process at −20° C. under 8 mTorr for 2 days, obtaining a powder form. The powder form was stored at room temperature and −20° C. individually. After two weeks, the powder forms (stored at room temperature and stored at −20° C.) were dissolved in 375.6 mg of water individually, and then the release amount of the TGF-b1 of the results was determined by enzyme-linked immunosorbent assay (ELISA), and the results are shown in
125 mg of mPEG-PLGA-BOX-PLGA-mPEG polymer, 16.6 mg of urea, and 34.4 mg of 100% PRP sample solution (without the mPEG-PLGA biblock polymer) were dissolved in water and stirred at 4° C., obtaining a solution. Next, the aforementioned solution was cooled to −20° C. to form a solid mixture. Finally, the solid mixture was subjected to a freeze-drying process at −20° C. under 8 mTorr for 2 days, obtaining a powder form. The powder form was stored at room temperature and −20° C. individually. After two weeks, the powder forms (stored at room temperature and stored at −20° C.) were dissolved in 324 mg of water individually, and then the release amount of the TGF-b1 of the results was determined by enzyme-linked immunosorbent assay (ELISA), and the results are shown in
125 mg of mPEG-PLGA-BOX-PLGA-mPEG polymer, 20.8 mg of urea, and 34.4 mg of 100% PRP sample solution (without the mPEG-PLGA biblock polymer) were dissolved in water and stirred at 4° C., obtaining a solution. Next, the aforementioned solution was cooled to −20° C. to form a solid mixture. Finally, the solid mixture was subjected to a freeze-drying process at −20° C. under 8 mTorr for 2 days, obtaining a powder form. The powder form was stored at room temperature and −20° C. individually. After two weeks, the powder forms (stored at room temperature and stored at −20° C.) were dissolved in 319.8 mg of water individually, and then the release amount of the TGF-b1 of the results was determined by enzyme-linked immunosorbent assay (ELISA), and the results are shown in
125 mg of mPEG-PLGA-BOX-PLGA-mPEG polymer, 25 mg of urea, and 34.4 mg of 100% PRP sample solution (without the mPEG-PLGA biblock polymer) were dissolved in water and stirred at 4° C., obtaining a solution. Next, the aforementioned solution was cooled to −20° C. to form a solid mixture. Finally, the solid mixture was subjected to a freeze-drying process at −20° C. under 8 mTorr for 2 days, obtaining a powder form. The powder form was stored at room temperature and −20° C. individually. After two weeks, the powder forms (stored at room temperature and stored at −20° C.) were dissolved in 315.6 mg of water individually, and then the release amount of the TGF-b1 of the results was determined by enzyme-linked immunosorbent assay (ELISA), and the results are shown in
As shown in
Platelet-rich plasma (PRP) of Preparation Example 1 was cooled to −20° C. to form a powder form. The powder form was stored at room temperature and −20° C. individually. After one month, the powder forms (stored at room temperature and stored at −20° C.) were dissolved in water individually, and then the release amount of the TGF-b1 of the results was determined by enzyme-linked immunosorbent assay (ELISA), and the results are shown in
Platelet-rich plasma (PRP) of Preparation Example 1 was cooled to −20° C. to form a powder form. The powder form was stored at room temperature and −20° C. individually. After one month, the powder forms (stored at room temperature and stored at −20° C.) were dissolved in water individually. After mixing with the agent (alamar blue assay) and then reacting, the human dermal fibroblasts cell (HDF) proliferation rate of the result was determined by measuring absorbance at 570 nm by a spectrophotometer, and the result is shown in
mPEG-PLGA biblock polymer (dissolved in water) was mixed with platelet-rich plasma (PRP) to prepare PRP solution with various concentration. mPEG-PLGA and platelet-rich plasma were mixed uniformly at 25° C. Next, the mixture was disposed into a specific release element, and 500 μL of the mixture was released by the release element at predetermined time instants for measuring the cumulative release amount and release rate of TGF-b1. The result is shown in
First, 1 mg/mL of doxorubicin was added into 15% mPEG-PLGA aqueous solution, and the mixture was stirred at 25° C. for uniformly mixing mPEG-PLGA and doxorubicin. Next, 1 mL of the result was disposed on the bottom of the release element (10 mL), and then heated at 37° C. for 5 min to form a gel. Next, 9 mL of phosphate buffered saline (PBS) was added into the release element, and the release element was disposed on a thermostat (37° C.) and shaken at 50 rpm, obtaining a PBS-miscible solution. A small amount of PBS-miscible solution was analyzed by HPLC to determine the release ratio of doxorubicin each day, and the result is shown in
First, TGF-b1 was added into 15% mPEG-PLGA aqueous solution, and the mixture was stirred at 25° C. for uniformly mixing mPEG-PLGA and TGF-b1. Net 1 mL of the result was disposed on the bottom of the release element (10 mL), and then heated at 37° C. for 5 min to form a gel. Next, 9 mL of phosphate buffered saline (PBS) was added into the release element, and the release element was disposed on a thermostat (37° C.) and shaken at 50 rpm, obtaining a PBS-miscible solution. A small amount of PBS-miscible solution was analyzed by HPLC to determine the release ratio of TGF-b1 each day, and the result is shown in
First, doxorubicin was added into a 15% mPEG-PLGA aqueous solution (the concentration of doxorubicin was 1 mg/mL). The mixture was stirred at 25° C. for uniformly mixing mPEG-PLGA 230 and doxorubicin. Next, 1 mL of the result was disposed on the bottom of the release element 240 (10 mL), and then heated at 37° C. for 5 min to form a gel 230. Next, 9 mL of phosphate buffered saline (PBS) was added into the release element, and the release element was disposed on a thermostat 210 (37° C.) and shaken at 50 rpm, obtaining a PBS-miscible solution. At predetermined time instants, PBS-miscible solution was subjected to an ultrasound stimulation (as indicated by the arrow) via an ultrasound stimulation element 220 (as shown in
The PBS-miscible solution of Example 18 was provided. The conditions with ultrasound stimulation or without ultrasound stimulation were compared. As shown in
The PBS-miscible solution of Example 19 was provided. At predetermined time instants, PBS-miscible solution was subjected to an ultrasound stimulation (as indicated by the arrow) via an ultrasound stimulation element 220 (as shown in
First, bovine serum albumin (BSA) was added into 15% mPEG-PLGA aqueous solution. The mixture was stirred at 25° C. for uniformly mixing mPEG-PLGA 230 and bovine serum albumin. Next, 1 mL of the result was disposed on the bottom of the release element (10 mL), and then heated at 37° C. for 5 min to form a gel. Next, 9 mL of phosphate buffered saline (PBS) was added into the release element, and the release element was disposed on a thermostat 210 (37° C.) and shaken at 50 rpm, obtaining a PBS-miscible solution. At predetermined time instants, PBS-miscible solution was subjected to an ultrasound stimulation (as indicated by the arrow) via an ultrasound stimulation element 220 (as shown in
10% mPEG-PLGA was mixed with doxorubicin to form a drug-containing hydrogel (the doxorubicin concentration was 1 mg/mL). 2 mL of hyaluronic acid (sold by Hyalgan) and the drug-containing hydrogel were subjected to stagnation tests. The stagnation test included following steps. The hyaluronic acid and the drug-containing hydrogel were injected onto an inclined plate with a slop of 15 degree, and the flowing situations of the hyaluronic acid and the drug-containing hydrogel were observed. As a result, the hyaluronic acid was flowable at 37° C., and the drug-containing hydrogel was gel and not flowable. Therefore, the drug-containing hydrogel could be trapped in the affected area, thereby extending the drug effect.
150 μL of the drug-containing hydrogel (A) (10% mPEG-PLGA/Doxorubicin, wherein the doxorubicin concentration was 1 mg/mL), the hyaluronic acid (B) (sold by Hyalgan), and the saline solution (C) were injected into the subcutaneous tissue of mice individually. The visual appearances of mice was observed via visual inspection, and the stagnation condition in the subcutaneous tissue was imaged by ultrasonic detection. Finally, the fluorescence intensity was detected by IVIS spectrum. After seven days, the subcutaneous mass (as indicated by the arrow), which the drug-containing hydrogel (A) was injected thereinto, had a relatively high drug retention, as shown in
The drug-containing hydrogel (A) (10% mPEG-PLGA/Doxorubicin, wherein the doxorubicin concentration was 1 mg/mL), the hyaluronic acid (B) (sold by Hyalgan), and the saline solution (C) were injected into the subcutaneous tissue of mice individually.
The mPEG-PLGA biblock polymer of the disclosure has high thermal-sensitivity. The convenience of using the mPEG-PLGA biblock polymer can be improved by modifying the components and amounts. Drugs can be fixed in specific regions by means of the mPEG-PLGA biblock polymer to avoid drug flow. The mPEG-PLGA biblock polymer can be turned into a gel by body temperature by means of the advantage of phase change in the human body (37° C.) gel. Furthermore, drugs or transforming growth factor can be affixed to the wound tissue by means of the mPEG-PLGA biblock polymer, thereby effectively promoting tissue healing and restoring tissue integrity.
In addition, according to the Examples of the disclosure, the mPEG-PLGA biblock polymer can effectively cover doxorubicin, bovine serum albumin, or transforming growth factor. Furthermore, the release amount of the aforementioned substances can be increased 1.45-3 times via ultrasound stimulation. This means that the mPEG-PLGA biblock polymer exhibits a controllable release ability.
It will be clear that various modifications and variations can be made to the disclosed methods and materials. It is intended that the specification and examples be considered as exemplary only, with the true scope of the disclosure being indicated by the following claims and their equivalents.
This application is a Divisional of U.S. patent application Ser. No. 15/855,260, filed Dec. 27, 2017, the entire contents of which are herein incorporated by reference.
| Number | Date | Country | |
|---|---|---|---|
| Parent | 15855260 | Dec 2017 | US |
| Child | 17113785 | US |
