Passive microchannel reactor for lipid nanoparticle synthesis and uses thereof

Abstract

The present application relates to the field of biotechnology and genetic engineering technology, and discloses a passive micro-channel reactor for lipid nanoparticle synthesis and application thereof, which comprises a reactor body and a reactor cover plate, the reactor body is embedded with a Y-shaped flow channel, comprising a first liquid inlet for the first liquid to enter, a first liquid inlet channel, a second liquid inlet for the second liquid to enter, a second liquid inlet channel, a mixing flow channel, an outlet channel at the end of the mixing flow channel for forming a steady-state liquid outlet flow, and an outlet; the mixing flow channel is provided with a spiral mixing flow chip for spiral disturbance; the reaction cover plate is provided with three interfaces, the first liquid inlet adapter is in communication with the first liquid inlet; the second liquid inlet adapter is in communication with the second liquid inlet; and the outlet adapter is in communication with the outlet. The flow channel structure of the present application is simple, easy to manufacture on a large scale, easy to clean, and has good product particle size distribution uniformity, and can flexibly adapt to various types of power systems.

Description

Technical Field

[0001] This invention relates to the fields of biotechnology and genetic engineering, and in particular to a passive microchannel reactor for the synthesis of lipid nanoparticles and its applications. Background Technology

[0002] Gene therapy, represented by mRNA vaccines, ASO drugs, and gene editing, repairs or replaces defective genes by introducing exogenous normal genes into target cells. It has a very broad application prospect in the treatment of diseases such as cancer, heart disease, cystic fibrosis, hemophilia, diabetes, and AIDS, and has received increasing research and development attention in recent years.

[0003] To ensure that drugs accurately target diseased cells within a given timeframe after entering the body and avoid damaging normal cells, a safe, efficient, and stable drug delivery system is crucial for gene therapy. Lipid nanoparticles (LNPs), as important non-viral carriers, encapsulate drug particles within a lipid layer, forming single-layer or multi-layer spherical particles with nanoscale dimensions. Through binding to specific ligands, they achieve specific selection of target cells, exhibiting strong tissue penetration, low cytotoxicity, and low immunogenicity, making them a popular choice for drug delivery.

[0004] Currently, commonly used methods for preparing lipid nanoparticles mainly include extrusion, ethanol injection, high-pressure homogenization, nanoprecipitation, material self-assembly, and in-situ synthesis / polymerization. These methods involve complex synthesis steps, significant batch-to-batch variations, and high synthesis losses. Furthermore, the structural uniformity and particle size distribution of nanoparticles in a single batch are poor, leading to unsatisfactory performance in large-scale industrial production. Against this backdrop, microchannel reactor technology, capable of manipulating fluid flow within micro- and nano-scale channels and achieving highly efficient fluid mixing and chemical reactions, enables the simple, rapid, and continuous production of nanoparticles, providing a new approach for the large-scale preparation of lipid nanoparticles.

[0005] To improve mixing reaction efficiency and increase reactant yield, researchers have constructed various active microchannel reactor systems by combining external conditions such as electric, magnetic, temperature, and acoustic fields. However, in the industrial preparation of LNP lipid nanoparticles, passive microchannel reactors are favored due to the stability issues of nucleic acid particles in external physical fields. By changing the topology and geometry of the flow channels, local eddies or even local turbulence can be introduced into the flow field, achieving distortion between the interfaces of different solutions, thereby increasing the contact probability and reaction efficiency between different solutions and improving the overall performance of passive microchannel reactors. Based on this idea, there are currently typical passive microchannel reactor structures such as flow focusing reactors, bifurcation recombination reactors, staggered ribbed reactors, baffle reactors, and T-junction reactors. These microchannel reactors have achieved preliminary applications in the industrial preparation of LNP lipid nanoparticles, but under current technological conditions, the production efficiency, encapsulation efficiency, particle size uniformity, and batch stability of LNP products are still relatively low. Microchannel reactors cannot be reused multiple times, and the mixing chip within them is inconvenient to replace. Summary of the Invention

[0006] To overcome or alleviate one or more of the above-mentioned technical problems, the present invention aims to provide a passive microchannel reactor for the synthesis of lipid nanoparticles and its application. It is a passive microchannel reactor based on a three-dimensional helical flow channel for the production of nanoscale LNP lipid nanoparticles. It increases the contact area of ​​nanoparticles in two solutions, increases the contact time during solution mixing, and increases the eddy current of the local flow field during the mixing time. Its flow channel structure is simple, which is conducive to large-scale manufacturing, easy to clean, reusable, and produces products with good uniformity of particle size distribution. It can be flexibly adapted to various types of power systems to achieve high-efficiency LNP preparation.

[0007] This invention provides the following technical solution:

[0008] A passive microchannel reactor for lipid nanoparticle synthesis includes a reactor body (1) and a reactor cover (2). The reactor body (1) is embedded with a recessed Y-shaped flow channel, which includes a first inlet (11) for a first liquid, a first inlet channel (12), a second inlet (13) for a second liquid, a second inlet channel (14), a mixing channel (15) formed by the confluence of the first and second inlet channels, and a flow channel (15) at the end of the mixing channel (15) for forming a steady-state effluent flow. The liquid outlet channel (16) and the liquid outlet (17) are provided; the mixing channel (15) is provided with a spiral mixing chip (3) for spiral turbulence; the reaction cover plate (2) is provided with three interfaces, namely a first liquid inlet adapter (4), a second liquid inlet adapter (5) and a liquid outlet adapter (6), the first liquid inlet adapter (4) is connected to the first liquid inlet (11); the second liquid inlet adapter (5) is connected to the second liquid inlet (13); the liquid outlet adapter (6) is connected to the liquid outlet (17).

[0009] According to some possible implementations, the spiral mixing chip (3) is formed by scanning along a spiral line with a specific geometry, the central axis of the spiral mixing chip (3) coincides with the central axis of the mixing channel (15), and the specific geometry is selected from line segments, rectangles, arbitrary planar shapes or hollow planar shapes.

[0010] According to some possible implementations, the characteristic diameter of the spiral mixing chip (3) accounts for 20% to 80% of the characteristic diameter of the mixing channel (15).

[0011] According to some possible implementations, when the specific geometry is a line segment, the length of the spiral mixing chip (3) is 0.5~40 mm.

[0012] According to some possible implementations, when the specific geometry is rectangular, the diameter of the spiral mixing chip (3) is 0.1~10 mm and the length is 4~40 mm.

[0013] According to some possible implementations, the included angle between the mixing channel (15) and the outlet channel (16) is an obtuse angle.

[0014] According to some possible implementations, the spiral mixing chip (3) has 2 to 10 spiral turns.

[0015] According to some possible implementations, the spiral mixing chip (3) is made of polypropylene, stainless steel or PEEK.

[0016] An application of the passive microchannel reactor described above for the synthesis of lipid nanoparticles, which is used to produce nanoscale LNP lipid nanoparticles.

[0017] Compared with the prior art, the present invention has the following beneficial effects:

[0018] (1) The passive microchannel reactor for lipid nanoparticle synthesis provided by the present invention is provided with a spiral mixing chip in the mixing channel, which can introduce spiral turbulence during liquid mixing and promote the mixing between different liquids.

[0019] (2) The passive microchannel reactor provided by the present invention can control the mixing of particles with uniform particle size, which is beneficial to drug delivery and stable storage.

[0020] (3) The system components of the passive microchannel reactor of the present invention are made of stainless steel and PEEK or polypropylene, which can withstand acids, alkalis and common cleaning agents. The connection is chamfered to leave no dead corners for cleaning, and the material in contact with the liquid can withstand ethanol and acid and alkali solvents.

[0021] (4) The passive microchannel reactor of the present invention only requires the preparation of 2 portions of solution for the formation of nanoparticles. It is simple to operate in production, uses fewer containers, filters and other materials, and has low cost.

[0022] (5) The mixing chip involved in the passive microchannel reactor of the present invention has rich three-dimensional geometric features, which makes the mixing channel present a richer equivalent three-dimensional structure, the material is more fully mixed in the mixing channel, and it can be scaled up.

[0023] (6) The mixing chip and mixing channel involved in the passive microchannel reactor of the present invention are detachable and can be quickly replaced to meet the mixing requirements under different material properties. Attached Figure Description

[0024] Figure 1 This is a three-dimensional structural schematic diagram of a passive microchannel reactor for the synthesis of lipid nanoparticles provided in an embodiment of the present invention.

[0025] Figure 2 for Figure 1 Exploded view.

[0026] Figure 3 This is a schematic diagram of the internal flow channel of a passive microchannel reactor for the synthesis of lipid nanoparticles, provided in an embodiment of the present invention.

[0027] Figure 4 This is a schematic diagram of the structure of a spiral mixing chip in a first embodiment of a passive microchannel reactor for lipid nanoparticle synthesis provided in this invention.

[0028] Figure 5 This is a schematic diagram of the structure of a spiral mixing chip in a second embodiment of a passive microchannel reactor for lipid nanoparticle synthesis provided in this invention.

[0029] In the picture:

[0030] 1. Reactor body; 2. Reactor cover plate; 3. Spiral mixing chip; 4. First inlet adapter; 5. Second inlet adapter; 6. Outlet adapter; 11. First inlet; 12. First inlet channel; 13. Second inlet; 14. Second inlet channel; 15. Mixing channel; 16. Outlet channel; 17. Outlet. Detailed Implementation

[0031] The present invention will now be described in detail with reference to embodiments and accompanying drawings. However, it should be understood that the embodiments and drawings are for illustrative purposes only and do not constitute any limitation on the scope of protection of the present invention. All reasonable modifications and combinations included within the inventive spirit of the present invention fall within the scope of protection of the present invention.

[0032] The present invention will be further described below with reference to the accompanying drawings. Example

[0033] like Figure 1 and Figure 2 The passive microchannel reactor for lipid nanoparticle synthesis provided in this embodiment includes a reactor body 1 and a reactor cover 2 covering it. The reactor cover 2 has three interfaces, namely a first inlet adapter 4, a second inlet adapter 5, and an outlet adapter 6. The reactor body 1 has recessed flow channels in a Y-shape, such as... Figure 3 The flow channel layout includes a first inlet 11, a first inlet channel 12, a second inlet 13, a second inlet channel 14, a mixing channel 15, an outlet channel 16, and an outlet 17. The first inlet channel 12 and the second inlet channel 14 respectively receive two liquids to be mixed, and ultimately converge in the mixing channel 15, where the mixing process is completed under the turbulence effect of the spiral mixing chip 3. The spiral mixing chip 3 and the mixing channel 15 are mechanically integrated to change the equivalent flow cross-section and flow path of the mixing channel 15, thus promoting liquid mixing. The mechanical integration is detachable, referring to snap-fit ​​methods that do not require irreversible assembly such as bonding.

[0034] The first inlet 11 and the second inlet 13 are respectively connected to the first inlet adapter 4 and the second inlet adapter 5 to form a complete inlet channel, allowing the liquid to be mixed to seamlessly enter the mixing channel 15. The outlet 17 and the outlet adapter 6 are connected to form a complete outlet channel, allowing the mixed liquid to be output to the subsequent process in a standard manner. The first inlet adapter 4 is connected to the first inlet 11 and is perpendicular to the first inlet channel 12; the second inlet adapter 5 is connected to the second inlet 13 and is perpendicular to the second inlet channel 14; the end of the mixing channel 15 is directly connected to the outlet channel 16 for forming a steady-state outlet flow; the outlet adapter 6 is connected to the outlet 17 and is perpendicular to the outlet channel 17. The vertical arrangement is for the convenience of installing the inlet and outlet adapters. In actual operation, the reactors are very thin, and to ensure that the solution smoothly enters the flow channel to complete mixing and flows out of the reactor, the flow channel and outlet positions need to be vertically arranged.

[0035] The first inlet 11, the second inlet 13, and the outlet 17 have the same geometry and can be matched with standard Luer interfaces, through interfaces, or other interfaces. Correspondingly, the first inlet adapter 4, the second inlet adapter 5, and the outlet adapter 6 adopt corresponding interface forms.

[0036] Figure 3 In this process, there is a certain angle between the mixing channel 15 and the outlet channel 16. This angle is obtuse, so as to reduce the influence of the eddy current in the mixing channel 15 and form a steady-state outlet flow in the outlet channel 16, without affecting the flowability of the mixed liquid.

[0037] The spiral mixing chip 3 and the mixing channel 15 are designed to be detachable, so as to facilitate disassembly, cleaning and replacement, and can be adjusted according to different mixing requirements.

[0038] The spiral mixing chip 3 is formed by scanning along a spiral line with a specific geometry. This embodiment illustrates two implementations of the spiral mixing chip 3, as follows: Figure 4 and Figure 5 As shown.

[0039] The central axis of the spiral mixing chip 3 coincides with the central axis of the mixing channel 15. On the cross-section of the mixing channel 15, the characteristic diameter corresponding to the spiral mixing chip 3 accounts for 20% to 80% of the characteristic dimension of the mixing channel 15. Considering that the cross-sectional shapes of the spiral mixing chip 3 and the mixing channel 15 can be chosen in various ways, the characteristic dimension here refers to the dimension most relevant to the interface size. For example, when the cross-section is a line segment, the characteristic dimension is the length of the line segment; when the cross-section is a rectangle, the characteristic dimension is the dimension of the rectangle along the radial direction; when the cross-section is other irregular polygons, the characteristic dimension is the difference between its maximum and minimum dimensions along the radial direction.

[0040] The length of the spiral mixing chip 3 is generally less than the effective length of the mixing channel 15, and the appropriate length can be selected according to the liquid mixing requirements. The spiral mixing chip 3 is made of materials resistant to organic solvents and acids and alkalis, such as stainless steel, polypropylene (PP), or PEEK, and is processed by methods such as 3D printing, blow molding, and photolithography. This embodiment breaks through the 2D structural design of existing microchannel reactors by incorporating the spiral mixing chip 3, which allows for more uniform and thorough mixing of active ingredients and excipients in three-dimensional space, such as nucleic acids in lipid nanoparticles and the mixing of SM-102:DSPC:cholesterol:DMG-PEG-2k in a specified ratio.

[0041] In the first implementation, such as Figure 4 As shown, the spiral mixing chip 3 has a spiral surface structure, formed by a line segment spirally scanning along the axis. In the second embodiment, as... Figure 5 As shown, the cross-section of the spiral mixing chip 3 is rectangular, and the specific dimensions of the rectangle can be adjusted according to actual needs. In some other embodiments, the cross-section can adopt any feasible shape, including triangles, polygons, circles, semicircles, rhombuses, D-shapes, or even hollow or perforated planar structures. When the spiral mixing chip 3 is used in the first or second embodiment, the number of spiral scans of the spiral mixing chip 3 can be selected, such as 2-10 turns, to adapt to the effective mixing length required for different liquids to be mixed. Both of these flow channel control chips facilitate the mixing and encapsulation of two-phase solutions during nanoparticle synthesis. The flow channel control chip provided by this invention controls the formation of uniform particles.

[0042] In use, the two liquids to be mixed enter the first inlet channel 12 and the second inlet channel 14 through the first inlet 11 and the second inlet 13 respectively, and then converge at the ends of the first inlet channel 12 and the second inlet channel 14 into the mixing channel 15. In the mixing channel 15, due to the presence of the spiral mixing chip 3, the flow channel is transformed into a spiral shape, so that the two liquids are continuously affected by spiral turbulence during the flow process, thereby improving the mixing effect. After flowing out of the mixing channel, the mixed liquid will further flow through the outlet channel 16 to form a gently flowing mixed liquid for use in subsequent stages.

[0043] This embodiment is applicable to the production of nanoscale LNP lipid nanoparticles.

[0044] The above embodiments are merely preferred embodiments of the present invention, and the scope of protection of the present invention is not limited to the above embodiments. All technical solutions falling within the scope of the present invention's concept are within the scope of protection of the present invention. It should be noted that for those skilled in the art, improvements and modifications made without departing from the principles of the present invention should also be considered within the scope of protection of the present invention.

Claims

1. A passive microchannel reactor for the synthesis of lipid nanoparticles, characterized in that: The reactor includes a reactor body (1) and a reactor cover (2). The reactor body (1) is embedded with a recessed Y-shaped flow channel. The flow channel includes a first inlet (11) for a first type of liquid to enter, a first inlet channel (12), a second inlet (13) for a second type of liquid to enter, a second inlet channel (14), a mixing channel (15) formed by the intersection of the first and second inlet channels, an outlet channel (16) and an outlet (17) at the end of the mixing channel (15) for forming a steady-state liquid flow. The mixing channel (15) is provided with a spiral mixing chip (3) for spiral turbulence. The reactor cover (2) is provided with three interfaces, namely the first... The system includes an inlet adapter (4), a second inlet adapter (5), and an outlet adapter (6). The first inlet adapter (4) is connected to the first inlet port (11); the second inlet adapter (5) is connected to the second inlet port (13); and the outlet adapter (6) is connected to the outlet port (17). The spiral mixing chip (3) is formed by scanning along a spiral line with a specific geometric shape. The central axis of the spiral mixing chip (3) coincides with the central axis of the mixing channel (15). The specific geometric shape is selected from any planar shape. The characteristic diameter of the spiral mixing chip (3) accounts for 20% to 80% of the characteristic diameter of the mixing channel (15).

2. The passive microchannel reactor for lipid nanoparticle synthesis according to claim 1, characterized in that: When the specific geometry is a line segment, the length of the spiral mixing chip (3) is 0.5~40 mm.

3. The passive microchannel reactor for lipid nanoparticle synthesis according to claim 1, characterized in that: When the specific geometry is rectangular, the diameter of the spiral mixing chip (3) is 0.1~10 mm and the length is 4~40 mm.

4. The passive microchannel reactor for lipid nanoparticle synthesis according to claim 1, characterized in that: The angle between the mixing channel (15) and the outlet channel (16) is an obtuse angle.

5. The passive microchannel reactor for lipid nanoparticle synthesis according to claim 1, characterized in that: The spiral mixing chip (3) has 2 to 10 spiral turns.

6. The passive microchannel reactor for lipid nanoparticle synthesis according to claim 1, characterized in that: The spiral mixing chip (3) is made of polypropylene, stainless steel or PEEK.

7. An application of the passive microchannel reactor for the synthesis of lipid nanoparticles according to any one of claims 1 to 6, characterized in that: It is used to produce nanoscale LNP lipid nanoparticles.

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