Automated system for manufacturing medical nanoparticles and method for manufacturing medical nanoparticles using same

Abstract

The present invention relates to an automated system for manufacturing medical nanoparticles and a method for manufacturing medical nanoparticles using same, the system comprising: a medical nanoparticle manufacturing device; a dispensing device connected to the medical nanoparticle manufacturing device through tubing and dispensing the manufactured nanoparticles to a designated position; and an automatic washing solution injection device for washing the dispensed nanoparticles, so as to reduce quality variation and provide high-quality and safe medical nanoparticles through automatic and precise control of a manufacturing process.

Description

Automated system for manufacturing medical nanoparticles and method for manufacturing medical nanoparticles using the same

[0001] The present invention relates to an automated system for manufacturing medical nanoparticles and a method for manufacturing medical nanoparticles using the same. More specifically, it relates to a system that automates a series of processes for manufacturing nanoparticles as a carrier for delivering gene scissors into the body, and a method for manufacturing medical nanoparticles using the same.

[0002] Medical nanoparticles originated with the use of liposome-based lipid nanoparticles (LNPs) as drug delivery systems by encapsulating drugs within them to increase blood circulation time and stability. Since then, various forms of LNPs, as well as polymer or inorganic-based nanoparticles with diverse functionalities, have been actively researched as medical nanoparticles. Recently, next-generation drug delivery systems that enhance drug loading efficiency through porous structures and utilize cell-targeting characteristics via surface chemical modification are gaining attention.

[0003] Medical nanoparticles can be manufactured in large quantities using microfluidic devices, ultrasonic devices, high-pressure homogenizers, etc. However, since nanoparticles are highly sensitive to manufacturing conditions, it is not easy to mass-produce nanoparticles of consistent quality. In particular, for medical nanoparticles used in the human body, ensuring high reproducibility and yield is an important challenge.

[0004] With the recent emergence of new medical technologies such as gene therapies, mRNA vaccines, and gene editing tools, the importance of delivery vehicle technology capable of delivering these into human cells is increasing. In particular, gene editing tools are a technology capable of recognizing DNA sequences to delete or correct specific DNA; as they are known to enable the treatment of cancers caused by DNA mutations, they are regarded as a new paradigm technology, making delivery vehicle technology that can stably deliver gene editing tools into the human body extremely important.

[0005] A representative technology for delivering gene editing tools into the human body is direct injection into cells. However, this method has the disadvantages of low delivery efficiency and the requirement of external electrical energy. While there is a method utilizing viruses, its utility is limited due to the high risk of mutation and the impossibility of clinical trials.

[0006] Consequently, interest is growing in technologies that utilize porous nanoparticles as gene editing delivery vehicles. While they offer advantages such as a low risk of mutation and the elimination of external energy requirements, the lack of specialized facilities currently limits production to small-scale laboratory-based manufacturing, meaning mass production remains a long way off.

[0007] Therefore, there is a need to develop technologies related to the mass production of medical nanoparticles, and furthermore, nanoparticles capable of stably delivering gene editing tools to the human body.

[0008] The present invention aims to manufacture medical nanoparticles, specifically nanoparticles as carriers capable of carrying gene editing tools, and has one objective of solving the aforementioned technical problems and providing an automated system and a method for manufacturing medical nanoparticles capable of mass-producing medical nanoparticles of uniform and stable quality.

[0009] Another objective of the present invention is to overcome the conventional technical limitations of controlling the process simply by process time due to insufficient quality indicators in the manufacturing process of medical nanoparticles, and to provide an automated system and a method for manufacturing medical nanoparticles capable of repeatedly producing uniform, high-quality nanoparticles.

[0010] The problems of the present invention are not limited to those described above. A person skilled in the art to which the present invention pertains will have no difficulty understanding additional problems of the present invention from the overall contents of this specification.

[0011] In order to solve the technical problem of the present invention described above, according to an exemplary embodiment of the present invention, an automated system for manufacturing medical nanoparticles is provided, comprising: a medical nanoparticle manufacturing device; a dispensing device connected to the medical nanoparticle manufacturing device by tubing and dispensing nanoparticles manufactured in the nanoparticle manufacturing device to a designated location; and an automatic injection device for a washing solution for washing the dispensed nanoparticles.

[0012] The above automation system may further include a control device for controlling the above automation system.

[0013] The above medical nanoparticle manufacturing device may include: a reactor chamber; an automatic manufacturing solution injection unit connected to the reactor chamber for injecting a nanoparticle manufacturing solution at a constant rate and / or a constant volume; a temperature control unit capable of controlling the temperature inside the reactor chamber; and an optical sensor capable of analyzing the physical properties of the fluid inside the reactor chamber in real time.

[0014] The above optical sensor may analyze at least one of turbidity and absorbance.

[0015] The above medical nanoparticle manufacturing device may further include a stirring unit for stirring a solution inside a reactor chamber.

[0016] The above medical nanoparticle manufacturing device may further include a transfer device that connects a reactor chamber and a dispensing device to move the manufactured nanoparticles in a fixed amount and / or at a constant speed.

[0017] The reactor chamber includes an upper inlet and a lower outlet, the inlet is connected to a first line of an optical sensor, and the outlet is connected to a second line of an optical sensor, so that physical property monitoring can be performed through the movement of a fluid containing nanoparticles within the reactor chamber.

[0018] The above medical nanoparticle manufacturing device may further include a circulation pump that moves the manufactured nanoparticle solution to an optical sensor.

[0019] The above optical sensor can be placed inside the reactor chamber.

[0020] When the data measured by the above optical sensor reaches a reference value, the valve connected to the lower outlet of the reactor chamber is automatically opened and closed through the control device, allowing the manufactured nanoparticles to move to the dispensing device through the transfer device.

[0021] If the data measured by the optical sensor above does not reach a reference value, a solution containing nanoparticles can be moved to the upper inlet of the reactor through the first line according to the control device.

[0022] The dispensing device may include: an automatic nanoparticle solution injection unit; a dispensing nozzle; a three-dimensional moving unit for moving the dispensing nozzle to a predetermined position; and a two-dimensional moving unit for moving a container for dispensing the nanoparticle solution to a predetermined position.

[0023] The above automatic cleaning solution injection device may include an automatic cleaning solution injection unit; a dispensing nozzle; a three-dimensional moving unit for moving the dispensing nozzle to a predetermined position; and a two-dimensional moving unit for moving a container containing a nanoparticle solution to a predetermined position.

[0024] The above medical nanoparticles may be nanoparticles for gene editing delivery.

[0025] According to another exemplary embodiment of the present invention, a method for manufacturing medical nanoparticles using an automated system is provided, comprising: a step of injecting a nanoparticle manufacturing solution into a reactor chamber at a fixed quantity and / or constant rate through an automatic manufacturing solution injection unit; a step of manufacturing nanoparticles within the reactor chamber; a step of measuring the physical properties of a solution containing nanoparticles within the reactor chamber using an optical sensor; a step of controlling the movement of the solution containing the manufactured nanoparticles to a dispensing device through a transfer device connected to a lower discharge port of the reactor chamber when the physical properties of the solution containing the nanoparticles satisfy a reference value; a step of dispensing the solution containing nanoparticles moved to the dispensing device at a fixed quantity and / or constant rate to a predetermined location; and a step of washing the dispensed solution containing nanoparticles using an automatic washing solution injection unit.

[0026] The above-mentioned centrifugation step may be further included prior to the washing step of the dispersed nanoparticles.

[0027] The above nanoparticles may be silica-based porous nanoparticles.

[0028] The present invention provides an automated system for manufacturing medical nanoparticles and a method for manufacturing medical nanoparticles capable of repeatedly producing uniform, high-quality nanoparticles.

[0029] The automated system for mass-producing medical nanoparticles and the method for manufacturing medical nanoparticles according to the present invention overcome the conventional technical limitations of controlling the process simply by process time due to insufficient quality indicators in the manufacturing process of medical nanoparticles, and provide the advantage of being able to repeatedly produce uniform, high-quality nanoparticles.

[0030] FIG. 1 illustrates the configuration of an automated system (1000) for manufacturing medical nanoparticles, which is an exemplary embodiment of the present invention.

[0031] FIG. 2 shows a front view of a medical nanoparticle manufacturing device (100) among an automated system for manufacturing medical nanoparticles according to one embodiment of the present invention.

[0032] FIG. 3 illustrates a top view of a medical nanoparticle manufacturing device (100) among an automated system for manufacturing medical nanoparticles according to one embodiment of the present invention.

[0033] FIG. 4 illustrates a left view of a medical nanoparticle manufacturing device (100) among an automated system for manufacturing medical nanoparticles according to one embodiment of the present invention.

[0034] FIG. 5 illustrates a right view of a medical nanoparticle manufacturing device (100) among an automated system for manufacturing medical nanoparticles according to one embodiment of the present invention.

[0035] FIG. 6 shows a front view of a dispensing device and an automatic washing solution injection device (200) among an automated system for manufacturing medical nanoparticles according to one embodiment of the present invention.

[0036] FIG. 7 illustrates a top view of a dispensing device and an automatic washing solution injection device (200) among an automated system for manufacturing medical nanoparticles according to one embodiment of the present invention.

[0037] FIG. 8 illustrates a left view of a dispensing device and an automatic washing solution injection device (200) among an automated system for manufacturing medical nanoparticles according to one embodiment of the present invention.

[0038] FIG. 9 illustrates a right view of a dispensing device and an automatic washing solution injection device (200) among an automated system for manufacturing medical nanoparticles according to one embodiment of the present invention.

[0039] FIG. 10 is data showing the absorbance of nanoparticles measured during the operation of an automated system for manufacturing medical nanoparticles according to one embodiment of the present invention.

[0040] Figure 11 shows SEM images according to the absorbance of nanoparticles during the implementation of an automated system for manufacturing medical nanoparticles according to one embodiment of the present invention.

[0041] FIG. 12 shows an SEM image of medical nanoparticles produced according to an automated system for manufacturing medical nanoparticles according to one embodiment of the present invention.

[0042] Preferred embodiments of the present invention are described herein with reference to the accompanying drawings. However, embodiments of the present invention may be modified in various other forms, and the scope of the present invention is not limited to the embodiments described below.

[0043] In addition, embodiments of the present invention are provided to more fully explain the present invention to those with average knowledge in the relevant technical field.

[0044] In drawings, the shapes and sizes of elements may be exaggerated for clearer explanation.

[0045] In describing the embodiments of the present invention, if it is determined that a detailed description of known technology related to the present invention may unnecessarily obscure the essence of the present invention, such detailed description will be omitted. Furthermore, the terms described below are defined considering their functions in the present invention, and these may vary depending on the intentions or conventions of the user or operator. Therefore, such definitions should be based on the content throughout this specification. The terms used in the detailed description are merely for describing the embodiments of the present invention and should not be limited in any way. Unless explicitly stated otherwise, expressions in the singular form include the meaning of the plural form.

[0046] In this description, expressions such as “include” or “equipped” are intended to refer to certain characteristics, numbers, steps, actions, elements, parts or combinations thereof, and should not be interpreted to exclude the existence or possibility of one or more other characteristics, numbers, steps, actions, elements, parts or combinations thereof other than those described.

[0047] In this specification, terms such as 'top', 'upper', 'upper surface', 'lower', 'lower surface', 'lower surface', and 'side surface' are based on the drawings and may actually vary depending on the direction in which the elements or components are arranged.

[0048] Additionally, throughout the specification, when it is said that one part is 'connected' to another part, this includes not only cases where they are 'directly connected,' but also cases where they are 'indirectly connected' with other elements in between.

[0049] Hereinafter, an automated system for manufacturing medical nanoparticles according to exemplary embodiments of the present invention and a method for manufacturing medical nanoparticles using the same will be described in detail.

[0050] FIG. 1 illustrates the configuration of an automated system (1000) for manufacturing medical nanoparticles according to an exemplary invention. Referring to FIG. 1, the automated system (1000) for manufacturing medical nanoparticles includes a medical nanoparticle manufacturing device (100), a dispensing device connected to the medical nanoparticle manufacturing device by tubing for dispensing nanoparticles manufactured by the nanoparticle manufacturing device to a designated location, and an automatic washing solution injection device (200) for washing the dispensed nanoparticles.

[0051] In another example, the automation system (1000) for manufacturing the medical nanoparticles may further include a transfer device (170) that connects a reactor chamber and a dispensing device to transfer the manufactured nanoparticles at a constant quantity and / or constant speed. In yet another example, the automation system (1000) for manufacturing the medical nanoparticles may further include a control device (300) that controls the automation system.

[0052] Refer to FIGS. 2 to 5 for various exemplary embodiments of a medical nanoparticle manufacturing apparatus (100). FIG. 2 shows the front view of the manufacturing apparatus (100), FIG. 3 shows the view from above, FIG. 4 shows the view from the left, and FIG. 5 shows the view from the right.

[0053] Looking specifically at FIGS. 2 to 5, the medical nanoparticle manufacturing device (100) may include a reactor chamber (110), an automatic manufacturing solution injection unit (120) connected to the reactor chamber for injecting a nanoparticle manufacturing solution in a fixed amount and / or at a fixed rate, a temperature control unit (140) capable of controlling the temperature inside the reactor chamber, and an optical sensor (150) capable of analyzing the physical properties of the fluid inside the reactor chamber in real time.

[0054] In addition, the above-described medical nanoparticle manufacturing device may include a stirring unit (130) capable of stirring a solution at a desired speed inside a reactor chamber. Specifically, the stirring unit (130) may be equipped with a magnetic impeller or a magnetic bar. For example, if the stirring unit is a magnetic impeller, the stirring unit may be installed in the upper central part of the reactor chamber. At this time, the stirring speed can be controlled to a level of 0.1 to 1000 rpm, and the stirring direction can be unidirectional, such as forward or reverse, or bidirectional, and if necessary, stirring can be performed including an intermediate stopping time.

[0055] In addition, the above-mentioned medical nanoparticle manufacturing device may further include a manufacturing solution tank (121) that supplies conventional manufacturing solutions required for nanoparticle manufacturing.

[0056] First, a manufacturing solution required for manufacturing nanoparticles is supplied from a manufacturing solution tank (121). For example, if silica-based porous nanoparticles are being manufactured, the manufacturing solution tank may be a storage container made of a material capable of accommodating ultrapure water, an organic solvent, a surfactant, etc. The organic solvent is not particularly limited, but, for example, a solvent capable of dissolving the surfactant for manufacturing silica-based porous nanoparticles may be used. In this case, the solubility stability of the surfactant in the manufacturing solution is high, which may be more advantageous for manufacturing silica-based porous nanoparticles.

[0057] There may be multiple manufacturing solution tanks (121) that supply conventional manufacturing solutions required for manufacturing nanoparticles. For example, it is preferable to have at least two, three, four, or five tanks. The upper limit is determined by the type of manufacturing solution required and is not specifically limited, but for example, to simplify the device, it may be 20 or fewer.

[0058] Each manufacturing solution tank is connected to an automatic manufacturing solution injection unit (120), and the manufacturing solution can be injected from the automatic manufacturing solution injection unit (120) into the reactor chamber at a constant volume and / or constant speed through a control device (300). The automatic manufacturing solution injection unit (120) may specifically be a syringe pump, but is not particularly limited as long as it can be controlled at a constant volume and / or constant speed. In this case, there may also be multiple syringe pumps as needed. In this case, the nanoparticle manufacturing reaction can be controlled more precisely.

[0059] When injecting fluid into a reactor chamber connected by tubing, a dropwise method of slowly injecting in the form of small droplets is also possible, making it easy to control the precise volume and reaction rate. The reactor chamber (110) is capable of inducing the synthesis of porous nanoparticles and, specifically, may be composed of one or more chambers.

[0060] The reactor chamber (110) can be a continuous flow reactor, a microfluidic reactor, a millifluidic reactor, a jacketed reactor, etc.

[0061] The reactor chamber (110) may include a temperature control unit (140) capable of precisely controlling the temperature. In a non-limiting example, the reactor chamber (110) may be configured as a double-jacketed type or a water bath type reactor, and may be operated in a circulation or water bath manner by connecting it to an oil bath.

[0062] The reactor chamber includes an upper inlet and a lower outlet, the inlet is connected to a first line of an optical sensor, and the outlet is connected to a second line of an optical sensor, so that physical property monitoring can be performed through the movement of a fluid containing nanoparticles within the reactor chamber.

[0063] The nanoparticle manufacturing reaction proceeds within the reactor, and even during the process of manufacturing nanoparticles, the physical properties of the fluid within the reactor can be measured through the optical sensor (150). A solution containing nanoparticles can be moved to the optical sensor (150) through a second line connected to the lower outlet of the reactor chamber (110).

[0064] To facilitate the movement of the above nanoparticles, the medical nanoparticle manufacturing device (100) may further include a circulation pump (160) that moves the manufactured nanoparticle solution to an optical sensor (150). The circulation pump (160) is connected to a first line (151) connecting the optical sensor (150) to the upper inlet of the reactor chamber (110) and a second line (152) connecting the optical sensor (150) to the lower outlet of the reactor chamber, thereby enabling the solution containing nanoparticles to circulate well.

[0065] The optical sensor (150) is intended to measure the physical properties of the fluid in the reactor and may analyze at least one of turbidity and absorbance. In such a case, for example, the optical sensor (150) may be an absorbance sensor. The absorbance sensor may be installed outside or inside the reactor chamber.

[0066] The optical sensor (150) measures at least one physical property selected from turbidity and absorbance, and if the measured value does not meet the reference value, the solution containing nanoparticles can be moved back into the reactor via the first line through the control device (300) so that the reaction can be completed. Specifically, depending on the control device, the solution containing nanoparticles can be moved to the upper inlet of the reactor via the first line.

[0067] In a non-limiting embodiment, the optical sensor (150) may be placed inside the reactor chamber (110). This allows monitoring of the process of nanoparticle synthesis proceeding inside the reactor.

[0068] The inside of the reactor chamber can be controlled to a temperature above room temperature through the temperature control unit (140), and can be raised to a temperature above 200°C or up to 250°C. A temperature sensor may be provided to check the internal temperature. The inside of the reactor chamber may be coated with Teflon to protect the surface and facilitate cleaning.

[0069] After medical nanoparticles are manufactured in the reactor chamber (110), when the data measured by the optical sensor (150) reaches a reference value, the valve connected to the lower outlet of the reactor chamber is automatically opened and closed through the control device (300), and the manufactured nanoparticles can be transferred to the dispensing device through the transfer device (170).

[0070] The automated system (1000) for manufacturing medical nanoparticles according to the present invention can derive indicators such as automatic quantitative injection of manufacturing solutions, the completion time of nanoparticle manufacturing through measurement of turbidity or absorbance of reactants, and automatic transfer to a washing process through a control device (300).

[0071] FIGS. 6 to 9 illustrate a dispensing device for manufactured medical nanoparticles and an automatic washing solution injection device (200). With reference to FIGS. 6 to 9, the operation of the dispensing device for medical nanoparticles and the automatic washing solution injection device (200) will be described in detail.

[0072] When a solution containing synthesized medical nanoparticles (hereinafter referred to as "nanoparticle solution") is transferred via a transfer device (170) to a medical nanoparticle dispensing device and a washing solution automatic injection device (200), it can be dispensed at a constant volume and / or speed at a desired location via the dispensing device. The volume and speed of the nanoparticle solution being transferred can also be efficiently controlled according to the size of the container to be dispensed.

[0073] Specifically, the dispensing device may include an automatic nanoparticle solution injection unit (210), a dispensing nozzle (231), a three-dimensional moving unit (220) for moving the dispensing nozzle to a predetermined position, and a two-dimensional moving unit (230) for moving a container for dispensing the nanoparticle solution to a predetermined position.

[0074] The nanoparticle solution is injected at a constant volume and / or rate through the nanoparticle solution automatic injection unit (210) and dispensed to a predetermined position according to the movement of the dispensing nozzle (231). At this time, the dispensing nozzle can be moved to a pre-set position in the control device (300) by the 3D moving unit (220) that moves the dispensing nozzle to prepare for dispensing. Similarly, the 2D moving unit (230) can move the container to be dispensed with the nanoparticle solution to a pre-set position in the control device (300) to prepare for dispensing. The container to be dispensed with the nanoparticle solution shown in FIG. 7 is an example and may be a conical tube. As an example, 1 L of nanoparticle solution can be prepared and automatically dispensed into 20 50 mL conical tubes.

[0075] According to one embodiment of the present invention, the dispensed nanoparticle solution may undergo a washing process. That is, a washing solution is additionally dispensed into the dispensed nanoparticle solution.

[0076] The above washing process can be performed through an automatic washing solution injection device. Specifically, the automatic washing solution injection device may include an automatic washing solution injection unit, a dispensing nozzle, a three-dimensional moving unit that moves the dispensing nozzle to a predetermined position, and a two-dimensional moving unit that moves a container containing a nanoparticle solution to a predetermined position.

[0077] A cleaning solution can be supplied from a cleaning solution tank (241), and since various cleaning solutions can be used, there may be multiple cleaning solution tanks (241). Each cleaning solution tank (241) is connected to a cleaning solution automatic injection unit (240), and the cleaning solution automatic injection unit (240) can dispense the cleaning solution through a dispensing nozzle (231) at a predetermined quantity and / or constant speed in the control device (300). The above process can be repeated several times, and by automatically processing this repeated process, the cleaning process of medical nanoparticles can be performed efficiently.

[0078] Through the above washing process, impurities, unreacted substances, and surfactants generated during the synthesis process can be removed, the size of the nanoparticles can be made uniform, and stability can be improved by preventing aggregation. Since medical nanoparticles are applied to the human body, the purity and safety of the nanoparticles are extremely important, as low-purity nanoparticles can cause potential toxicity.

[0079] The present invention may provide a method for manufacturing medical nanoparticles using an automated system for manufacturing medical nanoparticles as described above. Specifically, the method may include: (S10) injecting a nanoparticle manufacturing solution into a reactor chamber at a fixed quantity and / or constant rate through an automatic manufacturing solution injection unit; (S20) manufacturing nanoparticles within the reactor chamber; (S30) measuring the physical properties of a solution containing nanoparticles within the reactor chamber using an optical sensor; (S40) controlling the movement of the solution containing the manufactured nanoparticles to a dispensing device through a transfer device connected to a lower discharge port of the reactor chamber when the physical properties of the solution containing the nanoparticles satisfy a reference value; (S50) dispensing the solution containing nanoparticles moved to the dispensing device at a fixed quantity and / or constant rate to a predetermined location; and (S60) washing the dispensed solution containing nanoparticles using an automatic washing solution injection unit.

[0080] In another example, the step of measuring the physical properties of a solution containing nanoparticles in the reactor chamber using the optical sensor (S30) may involve measuring the turbidity or absorbance appearing in the physical property value of the target nanoparticles. Specifically, for example, the particle distribution, porosity, size, etc. of the target nanoparticles, and the turbidity or absorbance appearing in the physical property value may be calculated in advance, and (S40) the physical properties of the solution containing the nanoparticles may be confirmed through whether the turbidity or absorbance satisfies the reference value.

[0081] The method for manufacturing medical nanoparticles according to the present invention may further include a step of centrifuging the dispensed nanoparticles prior to step (S60). Through centrifugation, the solvent used in the synthesis process is removed to increase the purity of the nanoparticles, and it is possible to selectively separate only nanoparticles of a desired size. Additionally, if necessary, a surface treatment step may be further included.

[0082] The automated system for manufacturing medical nanoparticles and the method for manufacturing medical nanoparticles according to the present invention can repeatedly produce uniform, high-quality nanoparticles. This overcomes the conventional technical limitations of controlling the process simply by process time due to insufficient quality indicators during the manufacturing process of medical nanoparticles, thereby enabling the repeated production of uniform, high-quality nanoparticles. More specifically, the automated system for manufacturing medical nanoparticles and the method for manufacturing medical nanoparticles according to the present invention may have a quality deviation of nanoparticles produced therefrom that is, for example, 38% or less, 35% or less, 30% or less, 28% or less, or 25% or less. In such cases, it provides a more advantageous effect for use as medical nanoparticles, more specifically as nanoparticles for delivering gene editing tools.

[0083] Quality deviations may be deviations in measurable physical properties, such as deviations in particle size, particle distribution, porosity, and surface roughness.

[0084] In an exemplary embodiment of the present invention, the medical nanoparticle may be a gene editing delivery nanoparticle. The gene editing delivery nanoparticle may be loaded with a gene editing component and injected into the body via injection or other administration methods. The gene editing delivery nanoparticle loaded with the gene editing component may pass through the cell membrane and enter the cytoplasm, and release the gene editing component in response to the intracellular environment. The gene editing delivery nanoparticle manufactured by the automated system according to the present invention has the advantage of being able to repeatedly reproduce high-quality nanoparticles in large quantities according to quality indicators, thereby increasing the therapeutic efficiency using gene editing.

[0085] The following shows the performance evaluation of nanoparticles for gene editing delivery manufactured according to the automated system of the present invention.

[0086] Examples

[0087] 1. Manufacturing of Gene Editing Delivery Vehicles

[0088] Using automated equipment for manufacturing medical nanoparticles according to an exemplary embodiment of the present invention, 1 liter of nanoparticles for the in vivo delivery of a gene editing tool (hereinafter referred to as "gene editing tool delivery vehicle") was prepared. The internal temperature of the reactor chamber was set to 80°C, and ultrapure water and triethanolamine (TEA) were quantitatively injected into the reactor chamber at a rate of 200 ml / min using a syringe pump, followed by stirring for 30 minutes. When the temperature of the solution inside the reactor reached 80°C, sodium salicylate (NaSal) and cetyltrimethylammonium chloride solution (CTAC) were injected into the reactor at the same rate, and stirring was maintained for about 1 hour. Subsequently, the rate was set so that a mixed solution of tetraethylorthosilicate (TEOS) and ethanol could be injected into the reactor chamber at a rate of 1 drop per second. Subsequently, monitoring of turbidity or absorbance was started, and for this purpose, a circulation pump was operated at a rate of 200 ml / min.

[0089] When the desired absorbance value was reached, the manufacturing process was automatically terminated, and the manufactured gene editing delivery solution was transferred to a dispensing device and an automatic washing solution injection device (200) and quantitatively dispensed 50 ml into 20 50 ml conical tubes. The dispensed delivery solution underwent a centrifugation process, was then placed back into the dispensing device and the automatic washing solution injection device (200), and after repeating the washing process, the process was terminated.

[0090] FIG. 10 is a graph showing the monitoring of turbidity or absorbance data during the manufacturing process of a gene editing delivery vehicle. Even in the case of a reproduction experiment under the same conditions, the same turbidity or absorbance results were obtained, confirming that the same quality can be secured when manufacturing a gene editing delivery vehicle according to the present invention.

[0091] Figure 12 shows an SEM image of the final gene editing delivery system, which can be seen as porous nanoparticles of uniform size and shape.

[0092] 2. Performance Evaluation of Gene Editing Delivery Vehicles

[0093] CRISPR RNP was delivered to HCT116 cell lines (negative control) and MT50 cell lines (anti-cancer) using a gene editing delivery vehicle prepared according to the above example, and the cell viability was measured to evaluate whether an effective gene editing effect occurred. The values ​​compared with the cell viability of a gene editing delivery vehicle prepared by a conventional laboratory method, such as the comparative example, rather than the automated system of the present invention, are shown in Table 1 below.

[0094] (Comparative Example)

[0095] A solution of triethanolamine dissolved in ultrapure water was placed in a beaker, and the temperature was controlled using a magnetic hot plate and a water bath. The mixture was stirred at 1,000 rpm for 30 minutes using a magnetic bar until it reached 80 ℃. Subsequently, a mixed solution of NaSal and CTAC was added to the beaker, and the mixture was stirred at 1,000 rpm for 1 hour while maintaining the temperature at 80 ℃. Afterward, a mixed solution of TEOS and ethanol was added, and the mixture was stirred at 1,000 rpm for 2 hours. Finally, porous silica nanoparticles were obtained through centrifugation and a washing process. The entire process was carried out while controlling the reaction time based on visual observation of turbidity.

[0096] (Evaluation Method)

[0097] (a) Evaluation Item 1: EMX1 Toxicity Analysis

[0098] A delivery vehicle was prepared by loading a gene scissors containing the Guide RNA of EMX1 onto porous silica nanoparticles prepared in the previous examples and comparative examples, respectively. Cancer cell lines were incubated and cultured in a Petri dish at 37°C and 25% CO2, and then the cell viability was measured by counting the cells after 3 days of culture following the injection of the delivery vehicle.

[0099] (b) Evaluation Item 2: MT50 Cancer Cell Apoptosis Effect

[0100] A delivery vehicle was prepared by loading the MT50 gene scissors, which have a cancer-killing effect, onto porous silica nanoparticles prepared in the previous examples and comparative examples, respectively. Cancer cell lines were incubated and cultured in a Petri dish under conditions of 37°C and 25% CO2, and then the cell viability was measured by counting the cells after 3 days of culture following the injection of the delivery vehicle.

[0101] Example Comparative Example EMX1 Toxicity Analysis 100% 91% MT50 Cancer Cell Killing Effect 25% 39%

[0102] Looking at Table 1 above, through EMX1 toxicity analysis, it can be confirmed that the gene editing delivery vehicle manufactured according to the automated system of the present invention was completely washed and free of residual toxic substances, showing a cell viability of 100%, whereas in the case of the comparative example, trace amounts of residual toxic substances remained. In addition, through the analysis of the MT50 cancer cell death effect, it can be seen that the gene editing delivery vehicle manufactured according to the automated system of the present invention contributed to the death of a much larger number of cancer cells compared to the comparative example.

[0103] Based on the above results, it can be determined that the product homogeneity of the manufactured gene editing delivery vehicle has been secured, as the product evaluation results appear very similar when the process indicators are operated identically through the automation system according to the present invention.

[0104] In other words, it is presumed that in the case according to the present invention, the gene editing efficiency was significantly higher because the gene scissors were accurately delivered to target cells or tissues. On the other hand, in the case of the comparative example, although it was carried out with the same mass balance as the example, it is judged that the quality of the gene scissors delivery vehicle was poor and it was difficult to form homogeneous particles because the reference value for quality control during the manufacturing process was not clear and the volume control of the reactants was not precisely performed.

[0105] It should be noted that the embodiments described above are intended merely to illustrate and embody the present invention and are not intended to limit the scope of the present invention. This is because the scope of the present invention is determined by the matters described in the patent claims and matters reasonably inferred therefrom.

[0106] [Explanation of the symbol]

[0107] 1000: Automated system for manufacturing medical nanoparticles

[0108] 100: Medical nanoparticle manufacturing device 110: Reactor chamber

[0109] 112: Reactor chamber outlet

[0110] 120: Automatic manufacturing solution injection unit 121: Manufacturing solution tank

[0111] 130: Stirring unit 140: Temperature control unit

[0112] 150: Optical sensor 151: 1st line

[0113] 152: Line 2 160: Circulation pump

[0114] 170: Transfer device 200: Nanoparticle dispensing and automatic washing solution injection device

[0115] 210: Automatic nanoparticle solution injection unit

[0116] 220: 3D Movement Unit (xyz Stage System)

[0117] 230: 2D Movement Unit (xy Stage System)

[0118] 231: Dispensing nozzle 240: Automatic cleaning solution injection unit

[0119] 241: Cleaning solution tank 300: Control device

Claims

1. Medical nanoparticle manufacturing device; A dispensing device connected to the above-mentioned medical nanoparticle manufacturing device via tubing and dispensing nanoparticles manufactured by the above-mentioned nanoparticle manufacturing device to a designated location; and An automated system for manufacturing medical nanoparticles comprising: an automatic injection device for a washing solution for washing the above-mentioned nanoparticles.

2. In Paragraph 1, An automation system for manufacturing medical nanoparticles, further comprising a control device for controlling the above automation system.

3. In Paragraph 1, The above medical nanoparticle manufacturing device is, Reactor chamber; An automatic solution injection unit connected to the above reactor chamber for injecting a nanoparticle manufacturing solution at a constant rate and / or a constant volume; A temperature control unit capable of controlling the temperature within the reactor chamber; and An automated system for manufacturing medical nanoparticles, comprising an optical sensor capable of analyzing the physical properties of the fluid in the reactor chamber in real time.

4. In Paragraph 3, The above optical sensor is an automated system for manufacturing medical nanoparticles, which analyzes at least one of turbidity and absorbance.

5. In Paragraph 3, The above-described medical nanoparticle manufacturing apparatus is an automated system for manufacturing medical nanoparticles, further comprising a stirring unit for stirring a solution within a reactor chamber.

6. In Paragraph 3, The above-mentioned medical nanoparticle manufacturing apparatus is an automated system for manufacturing medical nanoparticles, further comprising a transfer device that connects a reactor chamber and a dispensing device to move the manufactured nanoparticles at a constant volume and / or constant speed.

7. In Paragraph 3, An automated system for manufacturing medical nanoparticles, wherein the reactor chamber includes an upper inlet and a lower outlet, the inlet is connected to a first line of an optical sensor, and the outlet is connected to a second line of an optical sensor, and physical property monitoring is performed through the movement of a fluid containing nanoparticles within the reactor chamber.

8. In Paragraph 3, The above-described medical nanoparticle manufacturing device is an automated system for manufacturing medical nanoparticles, further comprising a circulation pump that moves the manufactured nanoparticle solution to an optical sensor.

9. In Paragraph 3, The above optical sensor is an automated system for manufacturing medical nanoparticles, placed inside a reactor chamber.

10. In Paragraph 7, An automated system for manufacturing medical nanoparticles, wherein when data measured by the optical sensor reaches a reference value, a valve connected to the lower outlet of the reactor chamber is automatically opened and closed via a control device, and the manufactured nanoparticles are transferred to a dispensing device via a transfer device.

11. In Paragraph 7, An automated system for manufacturing medical nanoparticles, wherein if the data measured by the optical sensor does not reach a reference value, a solution containing nanoparticles moves to the upper inlet of the reactor through the first line according to the control device.

12. In Paragraph 1, The above dispensing device is, Automatic nanoparticle solution injection unit; Dispensing nozzle; A three-dimensional moving part that moves the above dispensing nozzle to a predetermined position; and An automated system for manufacturing medical nanoparticles, comprising: a two-dimensional moving unit for moving a container for dispensing the nanoparticle solution to a predetermined position.

13. In Paragraph 1, The above automatic cleaning solution injection device is, Automatic cleaning solution injection unit; Dispensing nozzle; A three-dimensional moving part that moves the above dispensing nozzle to a predetermined position; and An automated system for manufacturing medical nanoparticles, comprising: a two-dimensional moving unit for moving a container containing a nanoparticle solution to a predetermined position.

14. In Paragraph 1, The above medical nanoparticles are nanoparticles for gene editing delivery, and the automated system for manufacturing medical nanoparticles.

15. A step of injecting a nanoparticle manufacturing solution into a reactor chamber at a fixed quantity and / or constant rate through an automatic manufacturing solution injection unit; A step of manufacturing nanoparticles within the reactor chamber above; A step of measuring the physical properties of a solution containing nanoparticles in the reactor chamber using an optical sensor; A step of controlling the transfer of the solution containing the nanoparticles to a dispensing device through a transfer device connected to the lower outlet of the reactor chamber when the physical properties of the solution containing the nanoparticles satisfy a reference value; A step of dispensing a solution containing nanoparticles transferred to the dispensing device at a fixed rate and / or a fixed rate to a predetermined location; and A method for manufacturing medical nanoparticles using an automated system, comprising the step of washing the solution containing the above-mentioned dispensed nanoparticles using an automatic washing solution injection device.

16. In Paragraph 15, A method for manufacturing medical nanoparticles using an automated system, further comprising a step of centrifuging prior to the step of washing the above-mentioned nanoparticles.

17. In Paragraph 15, A method for manufacturing medical nanoparticles using an automated system, wherein the above nanoparticles are silica-based porous nanoparticles.

Images