Method for producing ultrasound-responsive drug carrier for delivery of hydrophobic drug, and ultrasound-responsive drug carrier using same

Abstract

Disclosed are a method for producing an ultrasonic response drug carrier for delivering a hydrophobic drug and an ultrasonic response drug carrier produced by the method, the method comprising: (a) dissolving the hydrophobic drug in a drug-loading oil to obtain a hydrophobic drug-loading oil; and (b) performing mechanical mixing at a predetermined revolutions per minute after mixing the hydrophobic drug carrier oil, an inert gas, and a phospholipid, thereby generating the ultrasound responsive drug carrier.

Description

Technical Field

[0001] The present invention relates to a method for generating a drug carrier and a drug carrier using the method, and in particular, to a method for generating an ultrasound-responsive drug carrier for effectively delivering hydrophobic drugs and an ultrasound-responsive drug carrier using the method. Background Art

[0002] A drug delivery system (DDS) can be described as a dosage formulation that effectively delivers the required dose of a drug to treat a disease by minimizing the efficacy and effect of the drug.

[0003] Such drug delivery systems include transdermal, oral, or vascular methods depending on the drug delivery route. In addition, drug delivery systems that treat the affected area by introducing microcapsules into blood vessels are attracting attention as an ideal treatment technology for the future.

[0004] In addition, it can be said that the basic technology in drug delivery system technology is the technology of accurately targeting the drug to the target affected part and the technology of controlling the release of the drug in the affected part. Therefore, targeted drug delivery systems using ultrasound and ultrasound-responsive drug carriers have received increasing attention in recent years as technologies that can solve these problems.

[0005] In particular, ultrasound-responsive drug carriers used as ultrasound contrast agents undergo cavitation due to ultrasound energy, and studies have shown that this phenomenon improves the effect of drug delivery into the skin or inside cells, so people have tried to bind ligands to desired drugs or receptors in the membrane of ultrasound-responsive drug carriers to deliver drugs to the human body.

[0006] However, this method binds drugs on the membrane surface, so there is a possibility that drugs will be lost during the process of ultrasound-responsive drug carriers moving to the target location, thus failing to fully play the role of drug carriers. In addition, there is also the limitation that a large amount of drugs cannot be loaded.

[0007] Therefore, an improved solution that can solve the above problems is needed. Summary of the invention

[0008] Technical problems to be solved

[0009] The present invention aims to solve all the above problems.

[0010] In addition, another object of the present invention is to generate an ultrasound-responsive drug carrier formed of a phospholipid monolayer.

[0011] Furthermore, another object of the present invention is to protect the drug from the external environment by loading the hydrophobic drug into the interior of the ultrasound-responsive drug carrier.

[0012] In addition, another object of the present invention is to enable a certain amount or more of hydrophobic drugs to exhibit significant drug effects by loading them into ultrasound-responsive drug carriers.

[0013] Furthermore, another object of the present invention is to effectively deliver a hydrophobic drug in a target area irradiated with ultrasonic energy by making the hydrophobic drug show high responsiveness to ultrasonic energy.

[0014] Furthermore, another object of the present invention is to centrally deliver the hydrophobic drug to the target area by preventing the natural release of the hydrophobic drug loaded in the ultrasound-responsive drug carrier.

[0015] Furthermore, another object of the present invention is to generate ultrasound-responsive drug carriers having a predetermined size distribution.

[0016] Technical solutions for technical problems

[0017] The features of the present invention are used to achieve the above-mentioned purpose of the present invention and the characteristic effects of the present invention described later, and its structure is as follows.

[0018] According to one embodiment of the present invention, a method for generating an ultrasound-responsive drug carrier for delivering hydrophobic drugs is provided, which comprises: obtaining a hydrophobic drug carrier oil by dissolving the hydrophobic drug in a drug carrier oil; and after mixing the hydrophobic drug carrier oil, an inactive gas and a phospholipid, performing mechanical mixing at a predetermined number of revolutions per minute, thereby generating the ultrasound-responsive drug carrier.

[0019] The step (b) includes: (b1) when determining the target size range of the ultrasound-responsive drug carrier, determining the target revolutions per minute corresponding to the target size range as the predetermined revolutions per minute; and (b2) after mixing the hydrophobic drug carrier oil, the inactive gas and the phospholipid, performing the mechanical mixing at the target revolutions per minute, thereby generating the ultrasound-responsive drug carrier.

[0020] The step (a) includes: (a1) when determining the target size range of the ultrasound-responsive drug carrier, referring to the viscosity range of each of a plurality of candidate drug carrier oils containing the drug carrier oil, determining a specific candidate drug carrier oil having a target viscosity range as the drug carrier oil, wherein the target viscosity range corresponds to the target size range; and (a2) obtaining the hydrophobic drug carrier oil by dissolving the hydrophobic drug in the specific candidate drug carrier oil.

[0021] The shell of the ultrasound-responsive drug carrier includes the phospholipid, the inactive gas is fixed in contact with a first part in the inner surface of the shell of the ultrasound-responsive drug carrier, and the hydrophobic drug-carrying oil is in contact with a second part as the remaining part in the inner surface of the shell of the ultrasound-responsive drug carrier.

[0022] The shell of the ultrasound-responsive drug carrier comprises the phospholipid, and the outer surface and the inner surface of the shell are hydrophilic and hydrophobic respectively.

[0023] The step (a) comprises: (a3) ​​obtaining an intermediate hydrophobic drug carrier oil by placing the hydrophobic drug into the drug carrier oil, wherein the hydrophobic drug carrier oil is in a state in which a first portion of the hydrophobic drug is dissolved in the drug carrier oil, while a second portion of the hydrophobic drug as the remaining amount is not dissolved in the drug carrier oil; and (a4) removing the second portion of the hydrophobic drug as an insoluble portion of the hydrophobic drug from the intermediate hydrophobic drug carrier oil by centrifuging the intermediate hydrophobic drug carrier oil, thereby obtaining the hydrophobic drug carrier oil.

[0024] The inert gas includes at least a portion of perfluoromethane, perfluoroethane, perfluoropropane, perfluorobutane, perfluoropentane, perfluorohexane, perfluoroheptane, perfluorooctane, decafluoropentane, perfluoro(2-methyl-3-pentanone), perfluorotributylamine, perfluoro-15-crown-5 ether, perfluoro-1,3-dimethylcyclohexane, perfluoromethylcyclopentane, perfluorodecalin, perfluoromethyldecalin, perfluoroperhydrobenzyltetralin, PERFECTA, and sulfur hexafluoride.

[0025] According to another embodiment of the present invention, there is provided an ultrasound-responsive drug carrier for delivering hydrophobic drugs, comprising: a hydrophobic drug carrier oil obtained by dissolving the hydrophobic drug in the drug carrier oil; an inactive gas; and a shell comprising the hydrophobic drug carrier oil and the inactive gas inside.

[0026] The ultrasonic responsive drug carrier is characterized in that: when determining the target size range of the ultrasonic responsive drug carrier, the target revolutions per minute corresponding to the target size range is determined as the predetermined revolutions per minute, and after mixing the hydrophobic drug carrier oil, the inactive gas and the phospholipid, the mechanical mixing is performed at the target revolutions per minute, thereby generating the ultrasonic responsive drug carrier.

[0027] The ultrasonically responsive drug carrier is characterized in that: when determining the target size range of the ultrasonically responsive drug carrier, a specific candidate drug carrier oil having a target viscosity range is determined as the drug carrier oil with reference to the viscosity range of each of a plurality of candidate drug carrier oils containing the drug carrier oil, and the target viscosity range corresponds to the target size range, and the hydrophobic drug carrier oil is obtained by dissolving the hydrophobic drug in the specific candidate drug carrier oil.

[0028] The shell of the ultrasound-responsive drug carrier includes phospholipids, the inactive gas is fixed in contact with a first part in the inner surface of the shell of the ultrasound-responsive drug carrier, and the hydrophobic drug-carrying oil is in contact with a second part as the remaining part in the inner surface of the shell of the ultrasound-responsive drug carrier.

[0029] The shell of the ultrasound-responsive drug carrier comprises phospholipids, and the outer surface and the inner surface of the shell are hydrophilic and hydrophobic respectively.

[0030] The ultrasound-responsive drug carrier is characterized in that: the intermediate hydrophobic drug carrier oil is obtained by placing the hydrophobic drug into the drug carrier oil, wherein the hydrophobic drug carrier oil is in a state in which the first portion of the hydrophobic drug is dissolved in the drug carrier oil, while the second portion of the hydrophobic drug as the remaining portion is not dissolved in the drug carrier oil, and then the intermediate hydrophobic drug carrier oil is centrifuged to remove the second portion of the hydrophobic drug as the insoluble portion in the hydrophobic drug from the intermediate hydrophobic drug carrier oil, thereby obtaining the hydrophobic drug carrier oil.

[0031] The inert gas includes at least a portion of perfluoromethane, perfluoroethane, perfluoropropane, perfluorobutane, perfluoropentane, perfluorohexane, perfluoroheptane, perfluorooctane, decafluoropentane, perfluoro(2-methyl-3-pentanone), perfluorotributylamine, perfluoro-15-crown-5 ether, perfluoro-1,3-dimethylcyclohexane, perfluoromethylcyclopentane, perfluorodecalin, perfluoromethyldecalin, perfluoroperhydrobenzyltetralin, PERFECTA, and sulfur hexafluoride.

[0032] Beneficial Effects

[0033] The effect of the present invention is to generate an ultrasound-responsive drug carrier formed of a phospholipid monolayer.

[0034] In addition, the present invention protects the drug from the external environment by loading the hydrophobic drug into the interior of the ultrasound-responsive drug carrier.

[0035] In addition, the present invention enables a certain amount or more of hydrophobic drugs to be loaded into the ultrasound-responsive drug carrier to enable it to exhibit significant drug effects.

[0036] Furthermore, the present invention enables the hydrophobic drug to be effectively delivered in a target area irradiated with ultrasonic energy by making it show high responsiveness to ultrasonic energy.

[0037] Furthermore, the present invention enables concentrated delivery of the hydrophobic drug to the target area by preventing the natural release of the hydrophobic drug loaded in the ultrasound-responsive drug carrier.

[0038] Furthermore, another object of the present invention is to generate ultrasound-responsive drug carriers having a predetermined size distribution. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] The following drawings used to describe embodiments of the present invention are only part of the embodiments of the present invention, and technicians in the field to which the present invention belongs (hereinafter referred to as "technicians") can obtain other drawings based on these drawings without performing any creative work.

[0040] Figure 1 An ultrasound-responsive drug carrier according to an embodiment of the present invention is schematically shown.

[0041] Figure 2 Schematic representation of the results of generating ultrasound-responsive drug carriers at various settings of the homogenizer's revolutions per minute.

[0042] Figure 3a to Figure 3c The separation process and results of ultrasound-responsive drug carriers of various sizes according to an embodiment of the present invention are schematically shown.

[0043] Figure 4 The cumulative amount of drugs naturally released from the ultrasound-responsive drug carrier according to an embodiment of the present invention is schematically shown.

[0044] Figure 5 The hydrophobic drug oil encapsulation rate and the hydrophobic drug content of the ultrasound-responsive drug carrier generated according to one embodiment of the present invention are schematically shown.

[0045] Figures 6a to 6c The result of irradiating ultrasound to an ultrasound-responsive drug carrier according to an embodiment of the present invention is schematically shown.

[0046] Figure 7 The result of applying the ultrasound-responsive drug carrier according to one embodiment of the present invention to breast cancer cells and then irradiating them with ultrasound is schematically shown. DETAILED DESCRIPTION

[0047] The following detailed description of the present invention refers to the accompanying drawings, which schematically illustrate specific embodiments in which the present invention can be implemented to illustrate the purpose, technical solutions and advantages of the present invention. These embodiments are described in sufficient detail to enable ordinary technicians to implement the present invention.

[0048] In addition, throughout the detailed description and claims of the present invention, the word "comprise" and its modifications are not intended to exclude other technical features, additions, constituent elements or steps. For those skilled in the art, some other purposes, advantages and characteristics of the present invention are obvious from this description, while others are obvious from the embodiments of the present invention. The following examples and drawings are exemplary and are not intended to limit the present invention.

[0049] The present invention includes any realizable combination of the embodiments in this specification. It should be understood that the various embodiments of the present invention are different but not necessarily mutually exclusive. For example, without exceeding the technical concept and scope of the present invention, the specific shapes, structures and characteristics described in this specification can be realized by other embodiments. In addition, it should be understood that without exceeding the technical concept and scope of the present invention, the position or configuration of each constituent element in each disclosed embodiment can be changed. Therefore, the detailed description described below does not have a limiting meaning, and as long as it can be properly described, the scope of the present invention is limited only by all scopes equivalent to those described in its claims and the attached claims. Similar figure marks in the drawings refer to the same or similar functions in many aspects.

[0050] In order to enable those skilled in the art to easily implement the present invention, preferred embodiments of the present invention will be described in detail below with reference to the accompanying drawings.

[0051] Figure 1 An ultrasound-responsive drug carrier 1000 for drug delivery according to an embodiment of the present invention is schematically shown.

[0052] See also Figure 1 In an ultrasound-responsive drug carrier 1000 according to an embodiment of the present invention, a shell 100 including phospholipids may be formed, and a drug-carrying oil 300 loaded with a hydrophobic drug and an inactive gas 200 may be included in the shell.

[0053] For reference, the ultrasound-responsive drug carrier 1000 according to an embodiment of the present invention may be an ultrasound-responsive microbubble having a high responsiveness to ultrasound, but is not limited thereto.

[0054] At this time, the drug may be a hydrophobic drug, which may refer to a drug that is only soluble in oil and completely insoluble in water, but is not limited thereto, and may also include a drug that is slightly soluble in water but more soluble in oil.

[0055] For example, the drug as a hydrophobic drug may include bendamustine, busulfan, carmustine, chlorambucil, cyclophosphamide, dacarbazine, ifosfamide, melphalan, procarbazine, streptozocin, temozolomide, asparaginase, capecitabine, cytarabine, 5-fluorouracil, fludarabine, gemcitabine, methotrexate, pemetrexed, raltitrexed, actinomycin-D, bleomycin, daunorubicin, epirubicin, idarubicin, mitomycin, mitoxantrone, etoposide, At least a portion of docetaxel, irinotecan, paclitaxel, topotecan, vinblastine, vincristine, vinorelbine, carboplatin, cisplatin, oxaliplatin, alemtuzumab, BCG, bevacizumab, cetuximab, denosumab, erlotinib, gefitinib, imatinib, interferon, ipilimumab, lapatinib, panitumumab, rituximab, sunitinib, sorafenib, temsirolimus, trastuzumab, clodronate, ibandronate, pamidronate and zoledronic acid.

[0056] In addition, a hydrophobic drug can be dissolved in a drug-carrying oil, thereby being able to obtain a hydrophobic drug-carrying oil.

[0057] In addition, the ultrasound-responsive drug carrier 1000 can be generated using hydrophobic drug carrier oil, inert gas, and phospholipids.

[0058] For reference, the inactive gas used to generate the ultrasound-responsive drug carrier 1000 according to an embodiment of the present invention may include perfluorocarbon gases (for example, perfluoromethane, perfluoroethane, perfluoropropane, perfluorobutane, perfluoropentane, perfluorohexane, perfluoroheptane, perfluorooctane, decafluoropentane, perfluoro(2-methyl-3-pentanone), perfluorotributylamine, perfluoro-15-crown-5 ether, perfluoro-1,3-dimethylcyclohexane, perfluoromethylcyclopentane, perfluorodecalin, perfluoromethyldecalin, perfluoroperhydrobenzyltetralin, PERFECTA, etc.), sulfur hexafluoride, and at least a portion of the air. For reference, the inactive gas included in the ultrasound-responsive drug carrier 1000 may be in liquid form, but is not limited to this, and may also be in gaseous form.

[0059] In addition, the drug-carrying oil included in the ultrasound-responsive drug carrier 1000 according to an embodiment of the present invention may include at least a portion of almond oil, apricot oil, avocado oil, rapeseed oil, castor oil, coconut oil, cocoa butter, corn oil, cottonseed oil, linseed oil, medium-chain triglyceride (MCT) oil, palm oil, soybean oil and sunflower oil.

[0060] In addition, the phospholipids contained in the shell formed on the outer surface of the ultrasound-responsive drug carrier 1000 according to an embodiment of the present invention may include at least a portion of DPPC, DPPA, DSPE-mPEG-2000 and cholesterol. However, the present invention is not limited thereto, and the phospholipids contained in the shell formed on the outer surface of the ultrasound-responsive drug carrier 1000 according to an embodiment of the present invention may include at least a portion of phosphatidylcholine substances (HSPC, DEPC, DOPC and DMPC, etc.), DMPA-NA, DPPA-Na, DOPA-Na, DSPE, DSPE-mPEG, DSPE-mPEG-2000-Na, DSPE-mPEG-5000-Na, DSPE-Maleimide PEG-2000-Na.

[0061] For example, a hydrophobic drug can be placed in a drug carrier oil to obtain an intermediate hydrophobic drug carrier oil, wherein the hydrophobic drug carrier oil is in a state in which a first portion (a first amount) of the hydrophobic drug is dissolved in the drug carrier oil, while a second portion, which is the remaining amount of the hydrophobic drug, is not dissolved in the drug carrier oil. The intermediate hydrophobic drug carrier oil is then centrifuged to remove the insoluble hydrophobic drug (i.e., the second portion) in the hydrophobic drug, thereby obtaining the hydrophobic drug carrier oil.

[0062] For reference, the insoluble hydrophobic drug may refer to a hydrophobic drug that is no longer dissolved in the saturated state of the drug carrier oil when the hydrophobic drug is excessively put into the drug carrier oil in an amount greater than the solubility of the hydrophobic drug in the drug carrier oil.

[0063] Furthermore, after mixing the hydrophobic drug carrier oil, the inactive gas, and the phospholipid, mechanical mixing is performed at a predetermined number of revolutions per minute to generate the ultrasound-responsive drug carrier 1000 .

[0064] The diameter of the ultrasound-responsive drug carrier 1000 according to one embodiment of the present invention generated by the above process can be 500nm to 3um. However, the present invention is not limited thereto, and the size of the ultrasound-responsive drug carrier 1000 can be determined by changing the number of revolutions per minute of mechanical mixing or selecting oils of different viscosities.

[0065] For example, a target size range of an ultrasound-responsive drug carrier can be determined, and a specific candidate drug carrier oil having a target viscosity range (which corresponds to the target size range) can be determined as the drug carrier oil by referring to the viscosity range of each of a plurality of candidate drug carrier oils containing drug carrier oils, and then a hydrophobic drug can be dissolved in the specific candidate drug carrier oil, thereby obtaining a hydrophobic drug carrier oil. In addition, by mixing such a hydrophobic drug carrier oil with an inactive gas and a phospholipid, an ultrasound-responsive drug carrier corresponding to the target size range can be generated.

[0066] For another example, by determining the target size range of the ultrasound-responsive drug carrier, and determining the target revolutions per minute corresponding to the target size range as a predetermined revolutions per minute, after mixing the hydrophobic drug carrier oil, inactive gas and phospholipids, mechanical mixing is performed at the target revolutions per minute, thereby generating an ultrasound-responsive drug carrier corresponding to the target size range.

[0067] In addition, see Figure 1 It can be confirmed that the shell 100 formed outside the ultrasound-responsive drug carrier 1000 has a phospholipid monolayer shape.

[0068] For reference, the head of a phospholipid is hydrophilic and the tail is hydrophobic, so Figure 1 The interior of the ultrasound-responsive drug carrier shown to include a shell formed as a single layer is hydrophobic.

[0069] In the prior art, both sides (ie, the inner surface and the outer surface) of the shell of the ultrasound-responsive drug carrier whose shell is formed by a phospholipid bilayer are hydrophilic, and thus hydrophobic substances cannot be loaded.

[0070] On the contrary, according to an embodiment of the present invention, Figure 1 As shown, by forming the outer shell of the ultrasound-responsive drug carrier with a phospholipid monolayer, a hydrophobic substance (eg, a hydrophobic drug-carrying oil) can be loaded in large quantities into the inner space of the ultrasound-responsive drug carrier.

[0071] Meanwhile, according to one embodiment of the present invention, instead of directly loading the hydrophobic drug into the ultrasound-sensitive drug carrier, the hydrophobic drug is dissolved in a drug carrier oil to obtain a hydrophobic drug carrier oil, thereby the hydrophobic drug carrier oil can be loaded into the ultrasound-responsive drug carrier.

[0072] If the hydrophobic drug is not dissolved in oil but loaded into the ultrasound-responsive drug carrier as it is, the hydrophobic drug loaded in the ultrasound-responsive drug carrier may crystallize. When the ultrasound-responsive drug carrier loaded with such crystallized hydrophobic drug is irradiated with ultrasound, the problem of uneven release of the hydrophobic drug may occur.

[0073] Therefore, according to one embodiment of the present invention, after dissolving the hydrophobic drug in the drug carrier oil, the vial mixed with the hydrophobic drug carrier oil, the inactive gas and the phospholipid aqueous solution is mechanically mixed, thereby generating an ultrasound-responsive drug carrier 1000.

[0074] In addition, the following describes the generation process of the ultrasound-responsive drug carrier 1000 according to an embodiment of the present invention using a specific hydrophobic drug (e.g., paclitaxel), a specific drug carrier oil (e.g., MCT oil), a specific inactive gas (e.g., perfluorohexane), and a specific phospholipid (e.g., DPPC, DPPA, and cholesterol, etc.).

[0075] However, the generation process of the ultrasound responsive drug carrier 1000 described below is only an example to help understand the present invention, and the present invention is not limited to this. Those skilled in the art can easily understand without undue effort the method of generating the ultrasound responsive drug carrier 1000 according to the present invention by combining other hydrophobic drugs, other drug carrier oils, other inactive gases, and other phospholipids as described above.

[0076] For reference, the ultrasound-responsive drug carrier described below may be ultrasound-responsive microbubbles having a high responsiveness to ultrasound waves, but is not limited thereto.

[0077] For example, 15 mg of paclitaxel can be placed in a 20 mL microtube, and then 10 mL of MCT oil can be added to the microtube.

[0078] Alternatively, paclitaxel in the microtubes can be dissolved in MCT oil using an ultrasonic water bath at 40°C for about 2 hours.

[0079] Furthermore, the mixture in the microtube was centrifuged at 14,000 rpm for 20 minutes using a centrifuge to remove paclitaxel (powder) that was not dissolved in the MCT oil.

[0080] In addition, 200 mg of 1,2-Dipalmitoyl-sn-glycerol-3-phosphocholine (DPPC), (ii) 10 mg of diphenylphosphoryl azide (DPPA), (iii) 35 mg of cholesterol, and (iv) 10 mg of DSPE-PEG can be dispersed in 30 mL of propylene glycol, ultrasonically treated at 40° C. for about 1 hour using an ultrasonic water bath, and 70 mL of tri-distilled water is added, and ultrasonically treated at 40° C. for about 1 hour using an ultrasonic water bath, thereby obtaining a phospholipid aqueous solution for preparing an ultrasound-responsive drug carrier 1000. For reference, the refrigeration temperature of the phospholipid aqueous solution can be 4° C.

[0081] Alternatively, (i) 1 mL of MCT oil in which paclitaxel is dissolved at a concentration of 1.5 mg / mL, (ii) 18 mL of a phospholipid aqueous solution, and (iii) 1 mL of perfluorohexane may be placed in a 30 mL glass vial.

[0082] For reference, the dosages of the above-mentioned MCT oil, phospholipid aqueous solution and perfluorohexane are only examples, and the present invention is not limited thereto.

[0083] For example, (i) 2 mL of MCT oil dissolved with paclitaxel (paclitaxel / MCT oil concentration: 1.5 mg / mL), 16 mL of phospholipid aqueous solution, and 2 mL of perfluorohexane can be placed in a 30 mL glass vial, or (ii) 4 mL of MCT oil dissolved with paclitaxel (paclitaxel / MCT oil concentration: 1.5 mg / mL), 12 mL of phospholipid aqueous solution, and 4 mL of perfluorohexane can be placed in a 30 mL glass vial.

[0084] In addition, an emulsion (i.e., an ultrasound-responsive drug carrier) can be generated by mechanically mixing (i) a phospholipid aqueous solution, (ii) MCT oil dissolved with paclitaxel, and (iii) perfluorohexane placed in a glass vial. For reference, a solution having an ultrasound-responsive drug carrier 1000 can be obtained by (i) mechanically mixing at 4500 rpm for 45 seconds when using a vial mixer, and (ii) mechanically mixing at 35000 rpm for 5 minutes when using a homogenizer.

[0085] For reference, Figure 2 Shown are ultrasound-responsive drug carriers generated at different revolutions per minute.

[0086] See also Figure 2 It can be confirmed that when the rpm of the homogenizer is set to 18,000, almost no ultrasonic responsive drug carrier is generated, while when the rpm of the homogenizer is set to 25,000, a small amount of ultrasonic responsive drug carrier is generated. In addition, it can be confirmed that when the rpm of the homogenizer is set to 30,000, more ultrasonic responsive drug carriers are generated, and when the rpm of the homogenizer is set to 35,000, more ultrasonic responsive drug carriers with a target size are generated compared to other examples.

[0087] In addition, according to an embodiment of the present invention, in order to further improve the drug delivery effect, ultrasound responsive drug carriers other than specific ultrasound responsive drug carriers with a predetermined size (e.g., 500nm to 3μm) can be removed from ultrasound responsive drug carriers generated in multiple sizes (e.g., 100nm to 10μm).

[0088] For example, after setting the rpm of the homogenizer to 35,000 and performing mechanical mixing, 40 mL of distilled water can be added to 20 mL of a solution containing ultrasound-responsive drug carriers of various sizes, and then the entire solution can be placed in a 100 mL syringe.

[0089] In addition, after the syringe is refrigerated at 4°C and normal pressure (e.g., 1 standard atmosphere) for 1 hour, when a large ultrasonic responsive drug carrier is precipitated at the bottom of the syringe due to gravity, the solution or precipitate located at the bottom of the syringe can be removed. For reference, the volume of the removed solution or precipitate can be less than 10% of the volume of the entire solution in the syringe.

[0090] See also Figure 3a , it can be seen that the oversized ultrasound-responsive drug carriers (precipitates) and the rest of the supernatant were separated due to gravity.

[0091] Thereafter, the solution remaining in the syringe may be centrifuged at 2000 rpm for 20 minutes to remove the unprecipitated ultrasound-responsive drug carrier in the upper layer (ie, the ultrasound-responsive drug carrier with too small a size).

[0092] See also Figure 3b , it can be confirmed that the ultrasound-responsive drug carriers of a predetermined size (precipitate) and the remaining supernatant (ultrasound-responsive drug carriers of too small a size) are separated due to centrifugation.

[0093] In addition, see Figure 3a and Figure 3b It can be confirmed that the interior of the ultrasound-responsive drug carrier 1000 according to an embodiment of the present invention is divided into a crescent shape.

[0094] Specifically, it can be confirmed that the interior of the ultrasound-responsive drug carrier 1000 is divided into (i) a meniscus-shaped space occupied by the hydrophobic drug-carrying oil, and (ii) the remaining space occupied by the inactive gas.

[0095] In addition, through Figure 3a and Figure 3b It can be confirmed that the inert gas is fixed at a specific position inside the ultrasound-responsive drug carrier 1000 .

[0096] For example, in the inner surface of the shell of the ultrasound responsive drug carrier 1000 according to an embodiment of the present invention, the inactive gas is fixed by contacting the first portion, and the hydrophobic drug carrier oil can be in contact with the rest of the inner surface of the shell of the ultrasound responsive drug carrier (i.e., the second portion).

[0097] If the inactive gas is not fixed at a specific position inside the ultrasound-responsive drug carrier, but moves freely inside the ultrasound-responsive drug carrier, it may not show a crescent shape (e.g. Figure 3aand the partition form shown in Figure 3D).

[0098] Specifically, when the inactive gas is fixed at a specific position inside the ultrasonic responsive drug carrier, the arrangement direction of the ultrasonic responsive drug carrier can be determined according to the weight of the inactive gas. Therefore, irradiating the ultrasonic wave according to the arrangement direction of the ultrasonic responsive drug carrier can exert a more effective cavitation effect and effectively deliver the hydrophobic drug to the target area.

[0099] In addition, see Figure 3c It can be confirmed that (i) when the process described in the above example is not performed, ultrasound-responsive drug carriers with different sizes ranging from 50 nm to 6 μm are obtained, and on the contrary, (ii) when the process described in the above example is performed, ultrasound-responsive drug carriers with relatively uniform sizes ranging from 500 nm to 3 μm are obtained.

[0100] Furthermore, as described above, after dissolving a hydrophobic drug in a drug-carrying oil to obtain a hydrophobic drug-carrying oil, a stable ultrasound-responsive drug carrier can be generated by loading the hydrophobic drug-carrying oil into an ultrasound-responsive drug carrier.

[0101] Relatedly, Figure 4 The cumulative amount of drug naturally released from the ultrasound-responsive drug carrier 1000 generated according to an embodiment of the present invention is shown.

[0102] See also Figure 4 It can be confirmed that there is no naturally released drug 1 hour, 3 hours, and 6 hours after the generation of the ultrasound-responsive drug carrier 1000 loaded with hydrophobic drugs. In addition, it can be confirmed that only about 30% of all the drugs loaded in the ultrasound-responsive drug carrier 1000 are released even after 1 day from the generation of the ultrasound-responsive drug carrier 1000, and only about 60% of the drugs are released after 4 days and 7 days from the generation of the ultrasound-responsive drug carrier 1000.

[0103] Therefore, it can be confirmed that the ultrasound-responsive drug carrier 1000 generated according to an embodiment of the present invention can load the hydrophobic drug very stably.

[0104] also, Figure 5 1 and 2 show (i) the hydrophobic drug oil encapsulation rate and (ii) the hydrophobic drug content of the ultrasound-responsive drug carrier 1000 generated according to an embodiment of the present invention.

[0105] For reference, Figure 5 The horizontal axis represents the concentration of the hydrophobic drug carrier oil in which the hydrophobic drug is dissolved. Figure 5 The left vertical axis represents the hydrophobic drug oil encapsulation rate of the ultrasound-responsive drug carrier 1000, Figure 5The right vertical axis represents the hydrophobic drug content of the ultrasound-responsive drug carrier 1000 .

[0106] See also Figure 5 It can be confirmed that the hydrophobic drug oil encapsulation rate of the ultrasound-responsive drug carrier 1000 generated according to one embodiment of the present invention is independent of the concentration of the hydrophobic drug dissolved in the drug oil (e.g., 25ug / mL, 50ug / mL, 75ug / mL), and has a constant encapsulation rate of about 30% (i.e., uniform quality).

[0107] In addition, see Figure 5 When the concentration of the hydrophobic drug dissolved in the drug carrier oil increases to 25ug / mL, 50ug / mL, and 75ug / mL as described above, the content of the hydrophobic drug loaded in the ultrasound-responsive drug carrier 1000 generated according to an embodiment of the present invention also increases proportionally (for example, 10ug / mL, 15ug / mL, 20ug / mL).

[0108] It can be seen that by controlling the amount of the hydrophobic drug dissolved in the hydrophobic drug carrier oil, the content of the hydrophobic drug loaded in the ultrasound-responsive drug carrier 1000 generated according to an embodiment of the present invention can be controlled.

[0109] also, Figures 6a to 6c The result of irradiating ultrasound on the ultrasound-responsive drug carrier 1000 is schematically shown to confirm the responsiveness of the ultrasound-responsive drug carrier 1000 to ultrasound energy according to an embodiment of the present invention.

[0110] First, see Figure 6a When the ultrasound responsive drug carrier 1000 according to an embodiment of the present invention is applied inside a silicone tube 6002, the result of applying ultrasound to the ultrasound responsive drug carrier 1000 using an ultrasound irradiation device (transducer) 6001 shows that the brightness of the area where the ultrasound responsive drug carrier exists is higher than that of other areas. Therefore, it can be seen that the ultrasound responsive drug carrier 1000 according to an embodiment of the present invention has a high responsiveness to ultrasonic energy.

[0111] also, Figure 6b Shown, in use Figure 6a The ultrasound irradiation device shown takes a picture of the silicone tube to which the ultrasound responsive drug carrier is applied before irradiating the ultrasound responsive drug carrier 1000 with ultrasound, and takes a picture of the silicone tube 10 minutes after irradiating the ultrasound.

[0112] See also Figure 6b In the above figure, before the ultrasound is applied, the inside of the silicone tube is bright, but see Figure 6bAs can be seen from the figure below, the inside of the silicone tube becomes darker after 10 minutes of ultrasonic irradiation. The reason is that when ultrasonic waves are irradiated on the ultrasonic responsive drug carrier 1000, which has a high response to ultrasonic energy, the ultrasonic responsive drug carrier 1000 is destroyed due to the cavitation effect, and the destroyed particles aggregate, and then the aggregated particles precipitate, thereby the inside of the silicone tube becomes darker.

[0113] also, Figure 6c The image brightness of (i) the ultrasound-responsive drug carrier 1000 irradiated with ultrasound and (ii) the image brightness of the ultrasound-responsive drug carrier 1000 not irradiated with ultrasound are shown. Figure 6c The image brightness when the ultrasound-responsive drug carrier 1000 according to an embodiment of the present invention is irradiated with ultrasound is much lower than the image brightness when it is not irradiated with ultrasound. This confirms that the ultrasound-responsive drug carrier 1000 according to an embodiment of the present invention exhibits high responsiveness to ultrasonic energy.

[0114] also, Figure 7 The schematic diagram shows the results of observation through a confocal microscope while changing the wavelength of light after applying the ultrasound-responsive drug carrier 1000 according to an embodiment of the present invention to breast cancer and irradiating ultrasound.

[0115] For reference, in order to easily confirm whether the ultrasound-responsive drug carrier or the drug loaded therein is effectively introduced into the cell interior, Nile red having fluorescence is loaded into the ultrasound-responsive drug carrier instead of the hydrophobic drug.

[0116] Figure 7 (a) shows the results of observing the drug delivery effect in the visible light region, see Figure 7 (a) It can be confirmed that Nile red and / or ultrasound responsive drug carrier 1000 is introduced into breast cancer cells 7001. Figure 7 (b) to 7(e) are further confirmed.

[0117] Figure 7 (b) shows the observation results in the wavelength region of 300 nm to 600 nm (corresponding to the excitation wavelength (350 nm) and emission wavelength (461 nm) of Hoechst 3342) after the nuclei of breast cancer cells were stained with Hoechst 3342. The position of the nuclei of breast cancer cells can be confirmed by the blue part.

[0118] also, Figure 7 (c) shows the observation results in the wavelength region of 400 nm to 700 nm (corresponding to the excitation wavelength (540 nm) and emission wavelength (660 nm) of Nile Red), and the region where Nile Red exists can be confirmed by the red-colored portion.

[0119] also, Figure 7 (d) Yes Figure 7 (a) to Figure 7 (c) shows the merged graph of the observations, Figure 7 (e) Yes Figure 7 (b) and Figure 7 (c) shows the combined results of the observations shown in (d) and Figure 7 (e) It can be confirmed that the ultrasound-responsive drug carrier 1000 and / or Nile red loaded therein are effectively introduced into the interior (particularly, the cytoplasm) of breast cancer cells.

[0120] The present invention is described above through specific matters such as specific constituent elements and limited embodiments and drawings, but this is only to help a comprehensive understanding of the present invention. The present invention is not limited thereto, and technicians in the field of the present invention can make various modifications and changes based on the above description.

[0121] Therefore, the technical concept of the present invention should not be limited to the embodiments described above, and the claims and modifications equivalent or equivalent to the claims all belong to the technical concept scope of the present invention.

Claims

1. A method for producing an ultrasound-responsive drug carrier for delivering a hydrophobic drug, comprising: (a) dissolving the hydrophobic drug in a drug-carrying oil to obtain a hydrophobic drug-carrying oil; as well as (b) after mixing the hydrophobic drug carrier oil, the inert gas and the phospholipid, performing mechanical mixing at a predetermined number of revolutions per minute, thereby generating the ultrasound-responsive drug carrier.

2. The method according to claim 1, characterized in that: The step (b) comprises: (b1) when determining a target size range of the ultrasound-responsive drug carrier, determining the revolutions per minute of the target corresponding to the target size range as the predetermined revolutions per minute; as well as (b2) After mixing the hydrophobic drug carrier oil, the inert gas, and the phospholipid, performing the mechanical mixing at the target revolutions per minute, thereby generating the ultrasound-responsive drug carrier.

3. The method according to claim 1, characterized in that: The step (a) comprises: (a1) when determining a target size range of the ultrasound-responsive drug carrier, referring to the viscosity range of each of a plurality of candidate drug carrier oils containing the drug carrier oil, determining a specific candidate drug carrier oil having a target viscosity range as the drug carrier oil, the target viscosity range corresponding to the target size range; and (a2) obtaining the hydrophobic drug carrier oil by dissolving the hydrophobic drug in the specific candidate drug carrier oil.

4. The method according to claim 1, characterized in that: The shell of the ultrasound-responsive drug carrier comprises the phospholipid, In the inner surface of the shell of the ultrasound-responsive drug carrier, the inactive gas contacts with a first portion to be fixed, and in the inner surface of the shell of the ultrasound-responsive drug carrier, the hydrophobic drug-carrying oil contacts with a second portion as the remaining portion.

5. The method according to claim 1, characterized in that: The shell of the ultrasound-responsive drug carrier comprises the phospholipid, The outer surface and the inner surface of the shell are hydrophilic and hydrophobic, respectively.

6. The method according to claim 1, characterized in that: The step (a) comprises: (a3) placing the hydrophobic drug in the drug-carrying oil to obtain an intermediate hydrophobic drug-carrying oil, wherein the hydrophobic drug-carrying oil is in a state where a first portion of the hydrophobic drug is dissolved in the drug-carrying oil, and a second portion of the hydrophobic drug as the remaining amount is not dissolved in the drug-carrying oil; and (a4) removing the second portion of the hydrophobic drug, which is an insoluble portion of the hydrophobic drug, from the intermediate hydrophobic drug carrier oil by centrifuging the intermediate hydrophobic drug carrier oil, thereby obtaining the hydrophobic drug carrier oil.

7. The method according to claim 1, characterized in that: The inert gas includes at least a portion of perfluoromethane, perfluoroethane, perfluoropropane, perfluorobutane, perfluoropentane, perfluorohexane, perfluoroheptane, perfluorooctane, decafluoropentane, perfluoro(2-methyl-3-pentanone), perfluorotributylamine, perfluoro-15-crown-5 ether, perfluoro-1,3-dimethylcyclohexane, perfluoromethylcyclopentane, perfluorodecalin, perfluoromethyldecalin, perfluoroperhydrobenzyltetralin, PERFECTA, and sulfur hexafluoride.

8. An ultrasound-responsive drug carrier for delivering a hydrophobic drug, comprising: A hydrophobic drug-carrying oil, which is obtained by dissolving the hydrophobic drug in the drug-carrying oil; Inert gases; as well as A housing includes the hydrophobic drug carrier oil and the inert gas in an inner space.

9. The ultrasound-responsive drug carrier according to claim 8, characterized in that: When the target size range of the ultrasound-responsive drug carrier is determined, the target revolutions per minute corresponding to the target size range is determined as the predetermined revolutions per minute, and after mixing the hydrophobic drug carrier oil, the inactive gas and the phospholipid, the mechanical mixing is performed at the target revolutions per minute, thereby generating the ultrasound-responsive drug carrier.

10. The ultrasound-responsive drug carrier according to claim 8, characterized in that: When determining the target size range of the ultrasound-responsive drug carrier, a specific candidate drug carrier oil having a target viscosity range is determined as the drug carrier oil with reference to the viscosity range of each of a plurality of candidate drug carrier oils containing the drug carrier oil, wherein the target viscosity range corresponds to the target size range, and the hydrophobic drug carrier oil is obtained by dissolving the hydrophobic drug in the specific candidate drug carrier oil.

11. The ultrasound-responsive drug carrier according to claim 8, characterized in that: The shell of the ultrasound-responsive drug carrier includes phospholipids, In the inner surface of the shell of the ultrasound-responsive drug carrier, the inactive gas contacts with a first portion to be fixed, and in the inner surface of the shell of the ultrasound-responsive drug carrier, the hydrophobic drug-carrying oil contacts with a second portion as the remaining portion.

12. The ultrasound-responsive drug carrier according to claim 8, characterized in that: The shell of the ultrasound-responsive drug carrier includes phospholipids, The outer surface and the inner surface of the shell are hydrophilic and hydrophobic, respectively.

13. The ultrasound-responsive drug carrier according to claim 8, characterized in that: The intermediate hydrophobic drug carrier oil is obtained by placing the hydrophobic drug into the drug carrier oil, wherein the hydrophobic drug carrier oil is in a state in which a first portion of the hydrophobic drug is dissolved in the drug carrier oil, while a second portion of the hydrophobic drug as the remaining amount is not dissolved in the drug carrier oil, and then the intermediate hydrophobic drug carrier oil is centrifuged to remove the second portion of the hydrophobic drug as an insoluble portion in the hydrophobic drug from the intermediate hydrophobic drug carrier oil, thereby obtaining the hydrophobic drug carrier oil.

14. The ultrasound-responsive drug carrier according to claim 8, characterized in that: The inert gas includes at least a portion of perfluoromethane, perfluoroethane, perfluoropropane, perfluorobutane, perfluoropentane, perfluorohexane, perfluoroheptane, perfluorooctane, decafluoropentane, perfluoro(2-methyl-3-pentanone), perfluorotributylamine, perfluoro-15-crown-5 ether, perfluoro-1,3-dimethylcyclohexane, perfluoromethylcyclopentane, perfluorodecalin, perfluoromethyldecalin, perfluoroperhydrobenzyltetralin, PERFECTA, and sulfur hexafluoride.

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