Devices and methods for rapid formulation of nanoparticles

Abstract

Microfluidic devices for the formulation of LNPs are provided. The microfluidic device comprises a vacuum control mechanism to draw liquids through microfluidic channels. The microfluidic device controls the flow rate and mixing of liquids without the need for electronic components. This device is specifically adapted for the efficient and customizable manufacture of LNPs, enabling rapid and on-demand formulation for various gene medicine applications.

Description

TITLEDEVICES AND METHODS FOR RAPID FORMULATION OF NANOPARTICLESPRIORITY CLAIM

[0001] This application claims the benefit of and priority to U.S. Provisional App. No. 63 / 664,880, titled “DEVICES AND METHODS FOR RAPID FORMULATION OF LIPID NANOPARTICLES FOR GENE MEDICINE DELIVERY”, filed June 27, 2024, the entire contents of which are hereby incorporated by reference in their entirety.BACKGROUND

[0002] Lipid nanoparticles (“LNP”), as well as other forms of nanoparticles, have been found to improve the delivery of gene medicine, and are currently considered the industry standard to deliver these therapies in vivo. LNPs have been found to encapsulate the gene medicine and promote transfer of the gene medicine across cell barriers.

[0003] Such gene medicine deliveries are employed for such technologies as RNA interference therapies, in some cases used to fight severe acute respiratory syndrome brought on by SARS-CoV-2 and respiratory syncytial virus (commonly known as RSV). LNPs have become a highly useful vehicle in the in vivo delivery of RNA interference therapies. Other applications of gene medicine include the rapid development of vaccines (such as Pfizer and Modema vaccines against SARS-CoV-2), gene editing (such as CRISPR technology), gene replacement, and other therapeutic interventions.

[0004] However, it has been found that the efficacy of such LNPs can be affected by a number of factors (such as the flow rate of the LNP materials and the geometry of the device used to synthesize the LNPs, to name just two out of many examples), and the materials used to synthesize these LNPs may not be biocompatible. Further, the current microfluidic approaches used to manufacture LNPs are based on highly specialized electrical-powered equipment that have a high cost and require extensive expertise and development timeframes. Other methods used to synthesize theLNPs, such as sonication, may cause collateral damage in the effectiveness of the LNPs. Moreover, no current equipment enables on-demand formulation of personalized therapies in clinical settings.

[0005] As a result, there is a need for rapid formulation of LNPs using biocompatible materials in a microfluidic device that enables on-demand manufacturing of LNPs in clinical settings without requiring specialized expertise or heavy equipment.SUMMARY

[0006] The present disclosure generally relates to a microfluidic device and associated methods for the efficient formulation of LNPs.

[0007] Generally, the present disclosure solves the problem by providing a kit for rapid formulation of LNPs for gene medicine delivery, using a vertical device that enables pre-loading of validated lipid mixes. The kit employs many innovative and unique components that are described hereafter to enable manual precise operation of the microfluidic approach to formulate consistent LNPS. The device disclosed herein also can be constructed from biocompatible and chemically resistant polymers, including polydimethylsiloxane (“PDMS”) or poly(methyl methacrylate) (“PMMA”).

[0008] In light of the present disclosure, and without limiting the scope of the disclosure in any way, in an aspect of the present disclosure, which may be combined with any other aspect listed herein unless specified otherwise, microfluidic devices for the formulation of lipid nanoparticles are provided. In some embodiments, microfluidic device for the formulation of LNPs comprises a vacuum control mechanism that utilizes a manually operated or spring-loaded syringe to draw liquids through microfluidic channels, wherein said mechanism is uniquely applied to control the flow rate and mixing of liquids without requiring the use of electronic components, specifically adapted for LNP manufacturing.

[0009] In some embodiments, the microfluidic device comprises a dual compartment mixer for nano medicine.

[0010] In some embodiments, the microfluidic device comprises P-traps located at the bottom of each liquid compartment within the device, wherein each p-trap prevents unintended liquid flow without vacuum activation and contributes to a controlled flow rate through its geometry, enhancing the precision of LNP formulations.

[0011] In some embodiments, the microfluidic device comprises a Y- junction that leads to a unified mixing channel, followed by a singular or series of modular mixing geometries, wherein the mixing geometries are adaptable to modify flow rates under vacuum-induced conditions, facilitating the production of LNPs with variable sizes or characteristics.

[0012] In some embodiments, the microfluidic device comprises interchangeable mixing geometries that enable modifications of the device to produce LNPs with tailored sizes and characteristics, wherein such modifications can be made without altering the fundamental vacuum control and p-trap flow regulation mechanisms of the device.

[0013] In some embodiments, the microfluidic device comprises distinct channel geometries for organic and aqueous phases, designed using computational fluid dynamics to achieve desired flow rate ratios under a unified vacuum, thereby influencing the characteristics of the resulting LNPs, wherein the flow rate is controlled by the geometry of the microfluidic channels.

[0014] In some embodiments, the microfluidic device comprises vertically positioned loading compartments for raw materials, allowing materials to be held in place by gravity and p-trap until vacuum application initiates the flow through the microfluidic channels, wherein this setup simplifies the loading process and ensures controlled initiation of liquid flow.

[0015] In some embodiments, the microfluidic device comprises a compartment containing a sponge-like material pre-filled with a precise lipid mixture, wherein the solvent is removed using freeze-drying or a similar method, wherein the lipids can be resuspended using ethanol or other solvents prior to use, ensuring accurate and consistent lipid availability for the formulation process.

[0016] To this end, in a first aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, a microfluidic device for the formulation of nanoparticles is provided, the microfluidic device comprising a vacuum control mechanism configured to draw fluids through microfluidic channels, and at least one fluid mixing junction coupled to a modular channel geometry. The vacuum control mechanism is at least one of manually, mechanically, and electronically actuated, andthe at least one fluid mixing junction is configured to enable controlled mixing of at least two fluid phases to form nanoparticles.

[0017] In a second aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, the vacuum control mechanism comprises a manually operated or spring-loaded syringe.

[0018] In a third aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, the vacuum control mechanism does not require electronic components.

[0019] In a fourth aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, the fluid mixing junction comprises a Y-shaped configuration.

[0020] In a fifth aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, the microfluidic device further comprises flow regulation geometries designed to control flow onset and rate, wherein said geometries include at least one p-trap configuration.

[0021] In a sixth aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, a flow rate of fluids through the microfluidic channels is controlled by a diameter of the microfluidic channels.

[0022] In a seventh aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, the modular channel geometry is configured to be varied, and varying the modular channel geometry includes varying the diameter of at least one microfluidic channel.

[0023] In an eighth aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, varying the diameter of at least one microfluidic channel is configured to change a ratio of flow rate between a microfluidic channel configured to transport a payload and a microfluidic channel configured to transport a nanoparticle.

[0024] In a ninth aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, the microfluidic device is a component of a kit for nanoparticle production.

[0025] In a tenth aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, the kit further comprises at least onereplaceable cartridge containing a reagent for use in nanoparticle production, a vacuumgenerating apparatus, and optional adapters for at least one of a field setting, a clinical setting, and a lab setting.

[0026] In an eleventh aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, a microfluidic device configured for vertical orientation is disclosed, wherein the microfluidic device comprises a plurality of vertically arranged input compartments and a plurality of fluid channels configured for vacuum activation and in fluid communication with each of the vertically arranged input compartments. Each of the plurality of fluid channels are configured to initiate fluid flow under gravity and vacuum pressure.

[0027] In a twelfth aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, at least one of the plurality of input compartments contains a porous or solid matrix preloaded with lipid or nanoparticle precursor materials.

[0028] In a thirteenth aspect of the present disclosure, which may be combined with any other aspect unless otherwise specified, a method for producing nanoparticles is disclosed, the method comprising introducing at least two fluid phases into a microfluidic device, initiating flow via a vacuum-generating mechanism, regulating flow using integrated passive geometries, mixing the fluid phases through modular channel configurations to form nanoparticles, and collecting the output nanoparticle suspension.

[0029] The reader will appreciate the foregoing details, as well as others, upon considering the following detailed description of certain non-limiting embodiments including microfluidic device for the formulation of LNPs.BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Fig. 1 illustrates an isometric view of a kit containing a microfluidic device according to an example embodiment of the present disclosure.

[0031] Fig. 2A illustrates an isometric view of a microfluidic device according to an example embodiment of the present disclosure.

[0032] Fig. 2B illustrates an isometric view showing the internal geometries of the microfluidic device of Fig. 2A with a second housing component removed.

[0033] Fig. 3A illustrates an isometric view of a microfluidic device according to another example embodiment of the present disclosure.

[0034] Fig. 3B illustrates an isometric view showing the internal geometries of the microfluidic device of Fig. 3A.

[0035] Fig. 3C illustrates a front view of the microfluidic device of Fig. 3A.

[0036] Fig. 3D illustrates a rear view of the microfluidic device of Fig. 3A.

[0037] Fig. 3E illustrates a right side view of the microfluidic device of Fig.3A.

[0038] Fig. 3F illustrates a left side view of the microfluidic device of Fig. 3A.

[0039] Fig. 3G illustrates a top view of the microfluidic device of Fig. 3A.

[0040] Fig. 3H illustrates a bottom view of the microfluidic device of Fig. 3A.

[0041] Fig. 4A illustrates an isometric view of a microfluidic device according to a third example embodiment of the present disclosure.

[0042] Fig. 4B illustrates a front view of the microfluidic device of Fig. 4A.

[0043] Fig. 4C illustrates a rear view of the microfluidic device of Fig. 4A.

[0044] Fig. 4D illustrates a right side view of the microfluidic device of Fig.4A.

[0045] Fig. 4E illustrates a left side view of the microfluidic device of Fig. 4A.

[0046] Fig. 4F illustrates a top view of the microfluidic device of Fig. 4A.

[0047] Fig. 4G illustrates a bottom view of the microfluidic device of Fig. 4A.

[0048] Fig. 5A illustrates an isometric view of the microfluidic device of Fig. 3A with a cap coupled thereto according to an example embodiment of the present disclosure.

[0049] Fig. 5B illustrates a top view of the microfluidic device of Fig. 5A.

[0050] Fig. 5C illustrates a bottom view of the microfluidic device of Fig. 5A.

[0051] Fig. 5D illustrates a right side view of the microfluidic device of Fig.

[0052] Fig. 5E illustrates a left side view of the microfluidic device of Fig.5A.

[0053] Fig. 5F illustrates a front view of the microfluidic device of Fig. 5A.

[0054] Fig. 5G illustrates a rear view of the microfluidic device of Fig. 5A.

[0055] Fig. 6A illustrates an isometric view of the microfluidic device of Fig. 4A with a cap according to another embodiment coupled thereto.

[0056] Fig. 6B illustrates a top view of the microfluidic device of Fig. 6A.

[0057] Fig. 6C illustrates a bottom view of the microfluidic device of Fig. 6A.

[0058] Fig. 6D illustrates a right side view of the microfluidic device of Fig. 6A.

[0059] Fig. 6E illustrates a left side view of the micro fluidic device of Fig. 6A.

[0060] Fig. 6F illustrates a front view of the microfluidic device of Fig. 6A.

[0061] Fig. 6G illustrates a rear view of the microfluidic device of Fig. 6A.

[0062] Fig. 7A illustrates an isometric view of a cap for a microfluidic device according to an example embodiment of the present disclosure.

[0063] Fig. 7B illustrates a bottom isometric view of the cap of Fig. 7A.

[0064] Fig. 7C illustrates atop view of the cap of Fig. 7A.

[0065] Fig. 7D illustrates a bottom view of the cap of Fig. 7A.

[0066] Fig. 7E illustrates a right side view of the cap of Fig. 7A.

[0067] Fig. 7F illustrates a left side view of the cap of Fig. 7A.

[0068] Fig. 7G illustrates a front view of the cap of Fig. 7A.

[0069] Fig. 7H illustrates a rear view of the cap of Fig. 7A.

[0070] Fig. 8 A illustrates an isometric view of a cap for a microfluidic device according to another example embodiment of the present disclosure.

[0071] Fig. 8B illustrates atop view of the cap of Fig. 8A.

[0072] Fig. 8C illustrates a bottom view of the cap of Fig. 8A.

[0073] Fig. 8D illustrates a right side view of the cap of Fig. 8A.

[0074] Fig. 8E illustrates a left side view of the cap of Fig. 8A.

[0075] Fig. 8F illustrates a front view of the cap of Fig. 8A.

[0076] Fig. 8G illustrates a rear view of the cap of Fig. 8A.

[0077] Fig. 9 illustrates a flowchart describing an example method of forming nanoparticles using a microfluidic device according to an example embodiment of the present disclosure.DETAILED DESCRIPTION

[0078] The present disclosure generally relates to microfluidic devices for the formulation of LNPs or any other desired nanoparticle.

[0079] Specifically, the present disclosure relates to, in one embodiment, a kit for rapid formulation of lipid nanoparticles for gene medicine delivery. The kit may use a tube filled with a pre-prepared sponge-like material infused with precise doses of specific lipid mixtures, or the mixtures may be provided later by an operator (i.e. not pre-loaded). This kit utilizes microfluidic-in-a-vial platform designed for the formulation of lipid nanoparticles to be used in delivering genetic material into cells, a process pivotal in gene therapies treating a range of diseases from genetic disorders to cancers. The device utilized in this application may be constructed from biocompatible and chemically resistant polymers.

[0080] The kit in one embodiment comprises a dual compartment mixer for a nano medicine.

[0081] Vacuum Control Mechanism for Manufacturing LNPs using Microfluidic Devices

[0082] In another embodiment, the kit comprises a vacuum control mechanism that utilizes a manually operated or spring-loaded syringe to draw liquids through microfluidic channels. The mechanism may be uniquely applied to control the flow rate and mixing of liquids without the use of electronic components, specifically adapted for LNP manufacturing.

[0083] P-Trap Design in Microfluidic Channels

[0084] In another embodiment, the kit may comprise a plurality of p-traps located at the bottom of each liquid compartment within the device. The p-traps may prevent unintended liquid flow without vacuum activation and contribute to a controlled flow rate through their geometry, enhancing the precision of LNP formulations. It should be noted that any flow regulation element can be used in the microfluidic channels, and that a p-trap is used here as an illustrative example embodiment of the present disclosure . For instance, a valve may be used to regulate flow instead of a p-trap.

[0085] Modular Mixing Geometry for Microfluidic LNP Production

[0086] In another embodiment, the kit may comprise a Y -junction that leads to a unified mixing channel, followed by at least one modular mixing geometry. The at least one mixing geometry may be adaptable to modify flow rates under vacuum- induced conditions, facilitating the production of LNPs with variable sizes or characteristics.

[0087] Scalable and Modular Design Features for Tailored LNP Characteristics

[0088] In another embodiment, the kit is designed for customizable lipid nanoparticle production. The kit may comprise interchangeable mixing geometries that enable modifications of the device to produce LNPs with tailored sizes and characteristics. The modifications may be made without altering the fundamental vacuum control and p-trap flow regulation mechanisms of the device.

[0089] Controlling Flow Rate by Geometry

[0090] In another embodiment, the kit may be configured such that the flow rate is controlled by the geometry of the microfluidic channels. The kit may comprise distinct channel geometries for organic and aqueous phases, designed using computational fluid dynamics to achieve desired flow rate ratios under a unified vacuum, thereby influencing the characteristics of the resulting LNPs. For instance, the total flow rate of the microfluidic device and the ratio of flow between the channels for the payload and the lipid may be controlled by varying the diameter of any of the channels or a portion thereof. The flow may also be manipulated by the addition or subtraction of various features within the channels (for example, flow regulators).

[0091] Vertical Setup for Material Loading and Retention

[0092] In another embodiment, the kit may be configured for vertical setup. The kit may comprise vertically positioned loading compartments for raw materials, allowing materials to be held in place by gravity and, optionally, at least one p-trap until vacuum application initiates the flow through the microfluidic channels. The kit may simplify the loading process and ensure controlled initiation of liquid flow.

[0093] Sponge Pre-Filled with Lipids

[0094] In another embodiment, the kit may comprise a compartment containing a sponge-like material pre-filled with a precise lipid mixture. The solventmay be removed using freeze-drying or a similar method. The lipids may be resuspended using ethanol or other solvent(s) prior to use, ensuring accurate and consistent lipid availability for the formulation process.

[0095] Description of the Drawings

[0096] Turning now to Fig. 1, a kit 10 for LNP formulation is provided, including a syringe 12 and a LNP formulation device 100, 200, coupled to a device seat 14. As described in further detail below, the LNP formulation device 100, 200 is provided with a lipid in one compartment and a payload in another compartment. The syringe 12 is used to draw the lipid formulation and the payload formulation into mixing compartments of the device and into the syringe using vacuum pressure profdes, such that a fully mixed LNP formulation is provided in the syringe barrel as a result. The syringe 12 may be a manually operated syringe or a spring loaded syringe which helps ensure a steady draw rate. In some alternative embodiments, a syringe is not employed,. Rather, any vacuum-generating mechanism may be used to form a vacuum in the LNP formulation device 100, 200. In this sense, the kit 10 comprises a system for the development of LNPs by vacuum pressure profiles (i.e. without the use of electronics or electronic components).

[0097] Figs. 2A-2B illustrate isometric views of an exterior and an interior of one embodiment of the LNP formulation device 100, respectively, according to one embodiment of the present disclosure. The LNP formulation device 100 includes a housing 102 which may be formed as two or more housing components 104, 106 resiliently coupled. As may be appreciated from the following description of the interior, one of the housing components 104 may house the entirety of the microfluidic channels of the LNP formulation device 100, while the other housing component 106 may act as a sort of seat for the first housing component 104. In this way, the housing components 104, 106 may be separable from each other and a different first housing component 104 with a different interior microfluidic channel geometry may be coupled to the second housing component 106 if different flow conditions are desired, for example. This design allows for the LNP formulation device 100 to be modular. In some alternative embodiments, the housing 102 is formed as a single integrated unit or the second housing component 106 is not provided altogether (similar to the illustration of Fig. 2B).

[0098] The housing 102 has a lipid compartment 110 separated from a payload compartment 120 by a separator 130. The lipid compartment 110 and payload compartment 120 may be formed as cavities in the housing 102 configured to hold liquid formulations of a lipid mixture and a nucleic acid mixture, respectively, in some embodiments. In some embodiments, the lipid (or payload, if a payload able to be reconstituted) formulations may be provided as a sponge-like material pre-filled with a precise lipid mixture. The solvent may be removed using freeze-drying or a similar method. The lipids may be resuspended using ethanol or other solvent(s) prior to use, ensuring accurate and consistent lipid availability for the formulation process. The sponge is then provided in the lipid compartment 110 (or payload compartment 120 if applicable) . The separator 130 aids in prevent premature mixture of the lipid and payload formulations whether when provided in liquid form or in a dehydrated form. Further, in some embodiments, the LNP formulation device 100 is configured to have vertically positioned loading compartments for raw materials, allowing materials to be held in place by gravity and, optionally, at least one p-trap 142, 144 (described in more detail below) until vacuum application initiates the flow through the microfluidic channels. This may ensure controlled initiation of liquid flow.

[0099] Indicators 160a, b are provided in some embodiments in order to show a user which part of the housing 102 pertains to the lipid compartment 110 and the payload compartment 120, or for use in preparing the kit 10. The lipid compartment 110 and the payload compartment 120 each form a respective funnel 112, 122 which begins a fluid channel for each fluid, which in some embodiments are, at least in part, microfluidic channels. The use of microfluidic channels helps enhance the controllability of the volume extracted for mixing and for delivery to the syringe 12 and may also prevent the introduction of unwanted gases into the fluid stream. The fluid channels meet at a Y -junction 145 which is where the lipid agent mixes with the payload to form the LNP. Although the flow regulation element is described as a Y -junction, any geometry may be employed in alternative embodiments to form a region where the fluid channels are merged into a single fluid channel.

[0100] Liquid is drawn into the fluid channels by introducing a vacuum at the outlet 150 by the syringe 12 of the kit 10. In this way a flow is created from an upstream direction (closer to the lipid compartment 110 and the payload compartment120) towards a downstream direction (closer to the outlet 150). Each fluid channel has its own flow regulation element 142, 144 that may prevent unintended liquid flow without vacuum activation and contribute to a controlled flow rate through their geometry, enhancing the precision of LNP formulations. Such flow regulation element 142, 144 may be a p-trap in some embodiments.

[0101] A mixing region 146 is included in some embodiments downstream of the Y-junction 145. The mixing region 146 is configured to alter the flow to promote mixing of the lipid and the payload, such as by causing an acceleration in at least one direction. In some embodiments the mixing region 146 comprises a serpentine or sinusoidal pattern. In alternative embodiments, the mixing region 146 comprises a cylindrical or ring banded pattern, a serial hourglass pattern, a corkscrew pattern, a funnel pattern, a herringbone pattern, or any other patter designed to promote accelerations of the fluid therein to enhance mixing. In some embodiments, the LNP formulation device 100 is configured to be modular, for example by having interchangeable portions of the housing 102 that change the internal geometries of the fluid channels. In some embodiments, the mixing region 146 is modular and may be exchanged for a mixing region 146 having a different geometry. The LNP formulation device 100 may include at least one modular mixing geometry. The at least one mixing geometry may be adaptable to modify flow rates under vacuum-induced conditions, facilitating the production of LNPs with variable sizes or characteristics. The mixing region 146 terminates at an outlet channel 148 which is sized and shaped to deliver the LNP formulation to the outlet 150 at a desired velocity and volumetric flow rate.

[0102] The total flow rate of the microfluidic device and the ratio of flow between the channels for the payload and the lipid may be controlled by varying the diameter of any of the channels or a portion thereof. Lor example, the diameter of each channel may be increased by a certain percentage or absolute number in order to increase a volumetric flowrate of the fluid in each of the channels, or the diameter of the payload channels may be increased while the other channel diameters are held constant in order to deliver a higher ratio of payload material to the resulting suspension. The modular design of the system may allow for a simple replacement of channel geometries in the microfluidic device in order to accommodate a wide range of desired flow conditions.The flow may also be manipulated by the addition or subtraction of various features within the channels (for example, flow regulators).

[0103] In some embodiments, the payload of the payload compartment 120 is any formulation configured to be suspended in a lipid mixture. For example, the payload may be a nucleic acid (such as RNA, mRNA, etc.), chemotherapeutics, antibiotics, gene editing and protein replacement therapies, small molecules, supplements, vitamins, proteins, and any other agent for in vivo placement by nanoparticle vehicle.

[0104] Turning now to Figs. 3A-3H, an alternative embodiment of the LNP formulation device 200 is illustrated in isometric exterior and interior views, respectively. The housing 202 of this embodiment of the LNP formulation device 200 takes on a more elliptical profile than the embodiment of Figs. 2A-2B. The separator 230 in this embodiment is offset towards the lipid compartment 210, such that the lipid compartment 210 holds a smaller volume than the payload compartment 220. In alternative embodiments, the separator 230 is offset such that the lipid compartment 210 is larger than the payload compartment 220. In some alternative embodiments, the housing 202 comprises a rectangular or square profile, an octagonal profile, a hexagonal profile, or any other desired geometrical profile. The housing 202 may include indicators 260a, b which delineate the agent which each respective compartment 210, 220 is configured to have disposed within.

[0105] As with the embodiment of Figs. 2A-2B, the housing 202 may comprise at least two housing components 204, 206. Also as similar to the embodiment of Figs. 2A-2B, one of the housing components 204 may house the entirety of the microfluidic channels of the LNP formulation device 200, while the other housing component 206 may act as a sort of seat for the first housing component 204. In this way, the housing components 204, 206 may be separable from each other and a different first housing component 204 with a different interior microfluidic channel geometry may be coupled to the second housing component 206 if different flow conditions are desired, for example. This design allows for the LNP formulation device 200 to be modular. As perhaps best illustrated in Fig. 3B, one or more key features 207, 208 may be provided to help place the first housing component 204 in correct relation to the second housing component 206 for proper coupling. In other embodiments, the keyfeatures 207, 208 may aid in the coupling of the housing components 204, 206, such as by comprising magnets or clasps, for example. It should be understood that “key” as used herein does not mean “essential”. In some alternative embodiments, the housing 202 is formed as a single integrated unit or the second housing component 206 is not provided. The housing 202 (and especially the second housing component 206, but also optionally the first housing component 204) may also include one or more grips 203, 205 which aid an operator in gripping the LNP formulation device 200 and provide further aesthetic and ergonomic enhancements to the LNP formulation device 200.

[0106] In much a similar way to the embodiment of Figs. 2A-2B, with modifications as necessary to the embodiment illustrated here, the LNP formulation device 200 comprises funnels 212, 222 at a terminus of each of the lipid compartment 210 and the payload compartment 220, respectively, which begin fluid channels as described above. In this embodiment, flow regulation elements 242, 244 such as p-traps are employed to ensure fluid and gases do not escape the fluid channels inadvertently. The fluid channels meet at a Y -junction 245 at which point the lipid formulation and the payload formation begin to mix in a single channel. A mixing region 246 promotes the efficient mixture of the lipid formulation and the payload by changing the acceleration of the fluid in the channel. Akin to the configurations described above in the embodiment of Figs. 2A-2B, the mixing region 246 comprises a serpentine or sinusoidal pattern, and in alternative embodiments, the mixing region 246 comprises a cylindrical or ring banded pattern, a serial hourglass pattern, a corkscrew pattern, a funnel pattern, a herringbone pattern, or any other pattern designed to promote accelerations of the fluid therein to enhance mixing. In some embodiments, the LNP formulation device 200 is configured to be modular, for example by having interchangeable portions of the housing 202 that change the internal geometries of the fluid channels. In some embodiments, the mixing region 246 is modular and may be exchanged for a mixing region 246 having a different geometry. The LNP formulation device 100 may include at least one modular mixing geometry. The at least one mixing geometry may be adaptable to modify flow rates under vacuum-induced conditions, facilitating the production of LNPs with variable sizes or characteristics. The mixing region 246 terminates at an outlet channel 148 which is sized and shaped to deliver the LNP formulation to the outlet 250 at a desired velocity and volumetric flow rate.

[0107] In some alternative embodiments ofthe configurations of any of Figs. 1-3B, the LNP formulation device 100, 200 includes a cap or cover which may form a hermetic seal over the lipid compartment 110, 210 and / or the payload compartment 120, 220. This may allow for embodiments in which the LNP formulation device 100, 200 is disposed in a generally horizontal configuration.

[0108] As with the embodiment of Figs. 2A-2B, the total flow rate of the microfluidic device and the ratio of flow between the channels for the payload and the lipid may be controlled by varying the diameter of any of the channels or a portion thereof. The flow may also be manipulated by the addition or subtraction of various features within the channels (for example, flow regulators). It should further be understood that any features which would be apparent as common between the illustrated embodiments are intended to apply to each embodiment, as well as to any other embodiment which may be appreciated by those of skill in the art, even if not explicitly stated.

[0109] Figs. 4A-4G illustrate an alternative embodiment of the LNP formulation device of Figs. 3A-3H where certain features are made optional. For example, the LNP formulation device 200a may contain no indicators on the housing, may have a housing that is one integrated piece, may not include any grips, and may have an outlet disposed in any location of the housing, or any of these features may have a different configuration and / or geometric profile.

[0110] Turning now to Figs. 7A-7H, an optional cap 300 for an LNP formulation device 200, 200a is illustrated. Fig. 5A shows, for example, how the cap 300 may be coupled to the LNP formulation device 200. The cap 300 includes a top surface 310 and a second surface 314 opposite the top surface 310 and configured to interface with a top portion of the LNP formulation device 200. The cap 300 also includes a side surface 311 disposed about a circumference ofthe top surface 310 and circumscribing a circumference of the second surface 314. The side surface 311 may culminate in a lip 312 which may provide coupling structure to resiliently couple the cap 300 to the LNP formulation device 200 in some embodiments.

[0111] A plurality of cap apertures 320 may be provided radially on the side surface 311. The cap apertures 320 may also extend into channels 322 in the second surface 314 of the cap 300. The cap apertures 320 and channels 322 may provide an airpathway so that when vacuum pressure is applied to the LNP formulation device 200 to draw fluids therethrough while a cap 300 is coupled thereto, a constant fluid pressure may be maintained throughout the LNP formulation device assembly 400. In some embodiments, as few as one cap aperture 320 and associated channel 322 is provided, and air is able to flow to both the lipid compartment 210 and the payload compartment 220. In other embodiments, as few as two cap apertures 320 and associated channels 322 are provided, one for each of the lipid compartment 210 and the payload compartment 220. Any desired number of cap apertures 320 and / or associated channels 322 are able to be provided to meet desired flow conditions. Further, the cap aperture 320 may be provided without an associated channel 322 and may be positioned such that air is still able to flow from an exterior of the cap 300 to an interior of the cap 300 and LNP formulation device 200 while the cap 300 and LNP formulation device 200 are coupled.

[0112] Figs. 8A-8G illustrate an alternative embodiment of the cap of Figs. 7A-7H where certain features are made optional or variable. For example, the cap 300a may contain no cap apertures or channels, or these features may have a different configuration and / or geometric profile.

[0113] Figs. 5A-5G illustrate a LNP formulation device assembly 400 where a LNP formulation device 200 is coupled to a cap 300 such that the top surface 310 of the cap faces away from the LNP formulation device 200 and the lip 3120 of the cap 300 surrounds an exterior portion of the LNP formulation device 200. The cap apertures 320 allow air flow to reach an interior portion of the assembly 400 between the cap 300 and the LNP formulation device 200.

[0114] Figs. 6A-6G illustrate an alternative embodiment of the LNP formulation device assembly 400a where some of the features of the LNP formulation device 200a and cap 300a are optional or variable as described above in the embodiments of Figs. 4A-4G and 8A-8G.

[0115] Turning now to Fig. 9, a method M500 for the production of nanoparticles is illustrated as a flowchart. At a first step S501, at least two fluid phases are introduced into a microfluidic device, such as the LNP formulation devices 100, 200 described above. At a second step S502, flow is initiated in the microfluidic device for each of the fluid phases via a vacuum-generating mechanism, such as the syringe 12 ofthe kit 10 described above. At a third step S503, integrated passive geometries are used to regulate the flow of the fluid phases through the microfluidic device. These passive geometries may include the Y-junctions 145, 245, the p-traps 142, 144, 242, 244, the mixing regions 146, 246, and the outlet channels 148, 248 described above, among other passive geometrical manipulations. At a fourth step, S504, the fluid phases are mixed through channel configurations of the microfluidic device, which in some embodiments includes modular channel configurations, to form nanoparticles. At a fifth step S505, the output nanoparticle suspension is collected.

[0116] Conclusion

[0117] It should be understood that various changes and modifications to the presently preferred embodiments described herein will be apparent to those skilled in the art. Such changes and modifications can be made without departing from the spirit and scope of the present subject matter and without diminishing its intended advantages. It is therefore intended that such changes and modifications be covered by the appended claims.

[0118] For example, in the foregoing, reference is made to lipid nanoparticles. It should be understood that any nanoparticle suitable to a particular use may be employed. Such nanoparticles include lipid nanoparticles, metallic nanoparticles (such as gold nanoparticles), polymeric nanoparticles, or any other nanoparticles as appreciated by those skilled in the art.

[0119] The preceding disclosures are illustrative embodiments. It should be appreciated by those of skill in the art that the devices, techniques and methods disclosed herein elucidate representative embodiments that function well in the practice of the present disclosure. However, those of skill in the art should, in light of the present disclosure, appreciate that many changes can be made in the specific embodiments that are disclosed and still obtain a like or similar result without departing from the spirit and scope of the invention.

[0120] Unless otherwise indicated, all numbers expressing quantities of ingredients, properties such as molecular weight, reaction conditions, and so forth used in the specification and claims are to be understood as being modified in all instances by the term “about.” Accordingly, unless indicated to the contrary, the numerical parameters set forth in the following specification and attached claims areapproximations that may vary depending upon the desired properties sought to be obtained by the present invention. At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of the claims, each numerical parameter should at least be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. Notwithstanding that the numerical ranges and parameters setting forth the broad scope of the invention are approximations, the numerical values set forth in the specific examples are reported as precisely as possible. Any numerical value, however, inherently contains certain errors necessarily resulting from the standard deviation found in their respective testing measurements.

[0121] The terms "a" and "an" and "the" and similar referents used in the context of describing the invention (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. Recitation of ranges of values herein is merely intended to serve as a shorthand method of referring individually to each separate value falling within the range. Unless otherwise indicated herein, each individual value is incorporated into the specification as if it were individually recited herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g. "such as") provided herein is intended merely to better illuminate the invention and does not pose a limitation on the scope of the invention otherwise claimed. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the invention.

[0122] The use of the term "or" in the claims is used to mean "and / or" unless explicitly indicated to refer to alternatives only or the alternatives are mutually exclusive, although the disclosure supports a definition that refers to only alternatives and "and / or."

[0123] Groupings of alternative elements or embodiments of the invention disclosed herein are not to be construed as limitations. Each group member may be referred to and claimed individually or in any combination with other members of the group or other elements found herein. It is anticipated that one or more members of a group may be included in, or deleted from, a group for reasons of convenience and / orpatentability. When any such inclusion or deletion occurs, the specification is herein deemed to contain the group as modified thus fulfilling the written description of any and all Markush groups used in the appended claims.

[0124] Preferred embodiments of this invention are described herein, including the best mode known to the inventors for carrying out the invention. Of course, variations on those preferred embodiments will become apparent to those of ordinary skill in the art upon reading the foregoing description. The inventor expects those of ordinary skill in the art to employ such variations as appropriate, and the inventors intend for the invention to be practiced otherwise than specifically described herein. Accordingly, this invention includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the above-described elements in all possible variations thereof is encompassed by the invention unless otherwise indicated herein or otherwise clearly contradicted by context.

[0125] Specific embodiments disclosed herein may be further limited in the claims using consisting of or consisting essentially of language. When used in the claims, whether as filed or added per amendment, the transition term “consisting of’ excludes any element, step, or ingredient not specified in the claims. The transition term “consisting essentially of’ limits the scope of a claim to the specified materials or steps and those that do not materially affect the basic and novel characteristic(s). Embodiments of the invention so claimed are inherently or expressly described and enabled herein.

[0126] Further, it is to be understood that the embodiments of the invention disclosed herein are illustrative of the principles of the present invention. Other modifications that may be employed are within the scope of the invention. Thus, by way of example, but not of limitation, alternative configurations of the present invention may be utilized in accordance with the teachings herein. Accordingly, the present invention is not limited to that precisely as shown and described.

Claims

CLAIMS1. A microfluidic device for the formulation of nanoparticles, comprising: a vacuum control mechanism configured to draw fluids through microfluidic channels, wherein said vacuum control mechanism is at least one of manually, mechanically, and electronically actuated; and at least one fluid mixing junction coupled to a modular channel geometry, configured to enable controlled mixing of at least two fluid phases to form nanoparticles.

2. The device of claim 1, wherein the vacuum control mechanism comprises a manually operated or spring-loaded syringe.

3. The device of claim 1, wherein the vacuum control mechanism does not require electronic components.

4. The device of claim 1, wherein the fluid mixing junction comprises a Y -shaped configuration.

5. The device of claim 1, further comprising flow regulation geometries designed to control flow onset and rate, wherein said geometries include at least one p-trap configuration.

6. The device of claim 1, wherein a flow rate of fluids through the microfluidic channels is controlled by a diameter of the microfluidic channels.

7. The device of claim 6, wherein the modular channel geometry is configured to be varied, and wherein varying the modular channel geometry includes varying the diameter of at least one microfluidic channel.

8. The device of claim 7, wherein varying the diameter of at least one microfluidic channel is configured to change a ratio of flow rate between a microfluidic channel configured to transport a payload and a microfluidic channel configured to transport a nanoparticle.

9. The device of claim 1, wherein the micro fluidic device is a component of a kit for nanoparticle production, wherein the kit further comprises: a vacuum-generating apparatus; and optional adapters for at least one of a field setting, a clinical setting, and a lab setting.

10. The device of claim 9, wherein the kit further comprises: at least one replaceable cartridge containing a reagent for use in nanoparticle production.

11. A microfluidic device configured for vertical orientation, comprising a plurality of vertically arranged input compartments and a plurality of fluid channels configured for vacuum activation and in fluid communication with each of the vertically arranged input compartments, wherein each of the plurality of fluid channels are configured to initiate fluid flow under gravity and vacuum pressure.

12. The microfluidic device of claim 11 , wherein at least one of the plurality of input compartments contains a porous or solid matrix preloaded with lipid or nanoparticle precursor materials.

13. A method for producing nanoparticles, comprising: introducing at least two fluid phases into a microfluidic device; initiating flow via a vacuum-generating mechanism; regulating flow using integrated passive geometries; mixing the fluid phases through modular channel configurations to form nanoparticles; and collecting the output nanoparticle suspension.

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