Applicator for drug patch

Abstract

A patch applicator device is provided. The device can include a reusable system or a disposable system for applying a push-through patch or a breakaway patch to a patient's skin. The device can improve the application and effectiveness of a microneedle-based patch.

Description

Technical Field

[0001] This invention was made with government support under R44AI142948 and SB1AI164584 awarded by the National Institutes of Health. The government has certain rights in this invention.

[0002] This application claims priority under 35 USC §119 to U.S. Provisional Application No. 63 / 355,301, filed on June 24, 2022. The entire contents of the provisional application are incorporated herein by reference.

[0003] This disclosure generally relates to applicator devices and systems for drug patches. The devices and systems can be used to securely apply a patch to a patient's skin.

Background Art

[0004] Currently, numerous devices are available or under development for administering drugs (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccines) through the skin to patients. The simplest of these devices may include systems that release or apply a substance to the skin for absorption. More complex systems can facilitate administration through various techniques such as the use of small needles that facilitate deeper administration into the skin, or other systems such as iontophoresis that promote the movement of substances into the skin.

[0005] Recently, patch-type devices that can place biodegradable needle-shaped devices into the skin have been developed. These devices carry the substance to be administered and slowly decompose over time, thereby optionally protecting the substance to be administered or, together with other agents that provide other therapeutic advantages (e.g., control of release rate, improvement of biological function), exposing the body to the substance to be administered for a desired period. Such devices can be used for the administration of vaccines, small molecule drugs, biologics, combination products, or other therapeutic or prophylactic substances.

[0006] The effectiveness of such biodegradable needle patches can be improved by ensuring that the patch is securely applied with sufficient force to deposit the biodegradable needles at the desired depth within the skin. However, while such patches can be effectively applied by manual application by a patient or healthcare provider, it would be beneficial to provide an improved system for reliably applying the biodegradable patch with minimal or minimal training and with a high level of reproducibility.

[0007] Accordingly, the present disclosure provides an improved device for applying a medical patch, including a biodegradable needle patch. SUMMARY OF THE INVENTION MEANS FOR SOLVING THE PROBLEM

[0008] The present disclosure relates to an applicator device for applying a patch-type device to a patient's skin. The device can enable secure application of the patch, including uniform and secure application, thereby achieving sufficient force and / or depth of skin penetration such that the needle-like portion of the patch is disposed at a desired depth within or under a portion of the skin. The applicator can be configured to provide a predetermined force to quickly and securely apply the patch to a desired location, thereby helping to improve the administration of an active agent (e.g., a drug, vaccine, biologic, or other material) using the selected patch.

[0009] The applicator can include an outer body portion that is substantially cylindrical and has a hollow interior, a piston slidably coupled within the hollow interior of the outer body portion, a compressible member disposed within the outer body and configured to apply a downward pressure to the piston, and an actuator disposed beneath the piston and slidably engaged within the hollow interior, the actuator extending from the bottom of the outer body such that an upward pressure on the actuator pushes up the piston and compresses the compressible member. The hollow interiors of the piston and the outer body portion are slidably coupled via a protrusion and a cam path, and the cam path forms a continuous loop such that an upward pressure on the piston compresses the compressible member and moves the piston upward within the hollow interior of the outer body portion until the protrusion reaches the top of the cam path and engages the downward portion of the cam path, whereupon the piston is released to the downward portion of the continuous loop and the piston is pushed down. The drug patch is held in a ring holder at the bottom of the actuator, pushed through the ring holder, and pressed against a target that abuts the bottom surface of the ring holder.

[0010] The accompanying drawings, which are incorporated herein and constitute a part of this specification, illustrate exemplary embodiments of the present disclosure and, together with the specification, serve to explain the principles of the present disclosure.

Brief Description of the Drawings

[0011]

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Modes for Carrying Out the Invention

[0012] Next, specific exemplary embodiments according to the present disclosure will be referred to in detail. Specific examples of these embodiments are shown in the accompanying drawings. As much as possible, the same reference numerals are used throughout the drawings to refer to the same or similar parts.

[0013] In this application, unless otherwise specified, the use of the singular form includes the plural form. In this application, the use of "or" means "and / or" unless otherwise specified. Further, the use of the term "including", as well as other forms such as "includes" and "included", is not limiting. The ranges described in this specification are to be understood to include all values between the endpoints.

[0014] The headings of the sections used in this specification are for structuring purposes only and should not be construed as limiting the subject matter being described. All documents or parts of documents (including but not limited to patents, patent applications, articles, books, and papers) cited in this application are hereby expressly incorporated by reference in their entirety for any purpose.

[0015] It should be understood that the principles of the present disclosure are described herein with reference to exemplary embodiments for specific applications, but the present disclosure is not limited thereto. Those skilled in the art, who have access to the teachings provided herein, will recognize that all additional changes, applications, embodiments, and substitutions of equivalents are within the scope of the embodiments described herein. Therefore, the present invention should not be regarded as limited by the foregoing description.

[0016] As described above, various patch devices are available for administering drugs or other substances into or through the skin. As used herein, "drug" or "drug patch" shall be understood to refer to any substance or patch that transports a substance that produces a biological effect in a patient. "Drug" shall be understood to refer to any pharmaceutical, small molecule drug, vaccine (including any vaccine such as mRNA vaccine, protein, glycoprotein, live virus, live attenuated virus, inactivated virus, recombinant vaccine, or other vaccines), peptide, biologic, antibody, vitamin, mineral, hormone, or other substance that can be administered into or through the skin.

[0017] Drug patches can include micro-needle-based devices, which will be described in more detail below. Such micro-needle devices can include one or more (preferably a group or array) of micro-needles. The micro-needles are disposed on the skin-facing surface of a flexible or semi-rigid patch, and by applying the patch to the patient's skin, the needles can penetrate the skin to a desired depth. In some cases, the micro-needles may include biodegradable components that deposit at a desired distance in the skin, decompose at a desired rate, and release or present the drug to the patient, thereby inducing a desired response such as an immunological response to a vaccine. Further details regarding exemplary patches including micro-needle patches will be further described below.

[0018] To improve the efficacy of any drug patch, it is desirable to ensure that the patch is applied properly, which includes uniform application to ensure that as many micro-needles as possible are placed at the appropriate depth. Further, it is desirable to ensure that the patch is applied such that the micro-needles are pushed into the skin at the desired depth without inadvertently shearing the micro-needles on the skin. By applying the patch properly, the overall efficacy can be improved.

[0019] An automated applicator device may be desirable to improve the consistency and effectiveness of patch application. Accordingly, the present disclosure provides embodiments of an applicator device having one or more features or advantages over existing devices or simple manual application. The disclosed device can be configured for repeated use or single use, and the device may have a patch pre-loaded thereon (i.e., as a kit or patch product including the applicator and the patch). Alternatively, the applicator may be separate from the patch, and the patch may be selected and attached to the device (e.g., based on the desired agent, patient characteristics, or the need for additional application to multiple patients or a patient with multiple patches).

[0020] The applicator can improve patch application by one or more of: (1) properly holding and / or stretching / pre-tensioning the skin to receive the patch; (2) applying a reliable degree of force and / or depth of force to the skin to ensure proper placement of the microneedles; and / or (3) controlling the distribution of the application force across the patch. The structure and function of the applicator in various embodiments will be described in more detail below.

[0021] FIG. 1 is a perspective view of a drug patch 10 applied to a patient's arm. As shown, the patch 10 is substantially square or square with rounded corners, although other shapes and configurations are contemplated and described below. The patch typically has an adhesive for securing the patch to the patient's skin. At least the backing layer is semi-flexible and can be deployed using the applicator described herein.

[0022] Figures 2A and 2B are, respectively, a side perspective view and a bottom perspective view of an exemplary applicator 100 for use with a medicament patch 10 similar to that shown in FIG. 1. As shown, the applicator 100 has an upper body portion 110 and a patient contact lower portion 105 (e.g., the bottom surface 120 of an actuator 122 or a ring holder 50 described below). The upper body portion 110 can be held by a user, and the patient contact lower portion 105 can be pressed against the skin of a patient. When the device is pressed against the skin of the patient, potential energy is generated until sufficient pressure is applied, at which point the internal piston system pushes down on the upper portion of the patch 10, causing the patch to be properly deployed onto the skin of the patient. Specific details of the internal components of the applicator and their functions will be described with the following figures.

[0023] FIGS. 3 and 4A-4D show the detailed assembled components and operations of the applicator 100 according to various embodiments. Specifically, FIG. 3 is a perspective view of an exemplary applicator 100 showing the internal components. FIG. 3 shows a patch applicator 100 including a patch 10 in the form of a generally flexible sheet.

[0024] The applicator 100 includes an outer body portion 110 that is substantially cylindrical and has a hollow interior 112. A piston 114 is slidably coupled to the hollow interior 112 of the outer body portion. A compressible member 118 (shown in FIG. 10A) is disposed within the outer body 110 and is configured to apply a downward pressure on the piston 114. An actuator 122 is disposed below the piston 114 and is slidably engaged with the hollow interior 112. The actuator extends from the outer body 100 such that an upward pressure on the bottom region 120 of the actuator 122 pushes up the piston 114 and compresses the compressible member 118.

[0025] The compressible member 118 can be any suitable spring or other compressible structure. For example, the spring can be a common spring, a compression spring, a wave spring, a dome spring, or a leaf spring. Alternatively, a compressible member such as a balloon, a compressible bladder, or a similar structure can also be used.

[0026] As described with reference to FIGS. 4A-4D, the piston 114 and the hollow interior 112 of the outer body portion are slidably connected by the protrusion 130 and the cam path 140, and until the protrusion reaches the top of the cam path and engages the downward portion of the cam path, the upward pressure on the piston compresses the compressible member 118 by the piston 114 and moves the piston upward within the hollow interior of the outer body portion. The cam path forms a continuous loop such that the piston is released to the downward portion of the continuous loop to release the piston and the piston is pushed down. Further, the drug patch 10 is held in a ring holder near the bottom of the actuator 122, and when the applicator is actuated, the drug patch 10 is pushed down through the ring holder and pressed against the target in contact with the bottom surface of the ring holder.

[0027] Next, with reference to FIGS. 4A-4D, the operation of the applicator component will be described. FIG. 4A is a side view of the embodiment of FIG. 3 with the compression member 118 and the piston 114 in the downward position. FIGS. 4B and 4C are additional views of the exemplary applicator of FIG. 4A in a partially compressed position prior to deployment. FIG. 4D is an additional view of the exemplary applicator of FIGS. 3, 4A, 4B, and 4C after releasing the piston to deploy the device and apply the patch.

[0028] As shown, piston 114 has, on at least one side, a protrusion 130 that engages a cam path 140 on the inner surface of the outer body 110. Protrusion 130 is also shown in FIG. 13A, which provides additional separate details regarding the piston embodiment. As shown, protrusion 130 is located near the upper portion 132 of piston 114, although the position of the protrusion is changeable. In some cases, and generally, piston 114 includes a plurality of protrusions, each of which engages a separate cam path 140 on the inner surface of the outer body 110. For example, in one embodiment, the piston has two protrusions as shown in FIG. 13A, and the two protrusions are located on opposite sides of the piston. Accordingly, each protrusion engages an individual cam path 140 on the opposite side surface of the inner surface of the outer body 110. It is also conceivable that more than two protrusions and cam paths are used (for example, three or more protrusions are evenly spaced around the piston along with the same number of cam paths). However, generally two protrusions are used because such an embodiment properly controls the movement of piston 114.

[0029] Next, the specific configuration of cam path 140 will be described. The movement of protrusion 114 in cam path 140 is shown in FIGS. 4A-4D. As shown, the cam path forms a continuous loop. For example, as shown, the loop includes a first upward section 141, a second upwardly inclined section 142, a third downward section 143, and a fourth downwardly inclined section 144. Sections 141-144 are ordered based on the order of movement during normal operation of applicator 100. The movement paths of protrusion 130 and cam path 140 are shown in enlarged views in FIGS. 10A and 10B.

[0030] Figure 4A shows the applicator 100 before use with the actuator 122 in the lower position. As shown, the protrusion 130 is at the lowest point of the cam path 140, and thus the piston 122 is in the lower position. As shown in Figure 13A, when an upward pressure is applied to the actuator 122, i.e., by pressing the device against the patient's skin, the actuator moves upward to a point where the extension 126 of the actuator (Figure 12) engages the flange or extended region 138 of the piston 114. By continuing to apply pressure to the actuator 122, the piston 114 begins to move upward along the first section of the cam path 141, thereby compressing the compressible member 118. By continuing to apply upward pressure, the protrusion of the piston 130 moves along the second section 142 of the cam path 140, and accordingly the piston 114 continues to move further upward, compressing the compressible member.

[0031] Finally, as shown in Figure 4C, the protrusion approaches the third downward section 143 of the cam path. When it reaches the third downward section 143 of the cam path, since the cam path is directed downward, the protrusion 130 can move downward, and when the protrusion 140 reaches the third downward section 143 of the cam path 140, the actuator extension 126 is disengaged from the flange or extended region 138 of the piston. The specific structure and function of the piston 114 and the actuator 122 will be described in more detail below. In some cases, the device 100 can also be reset or configured for additional use. In some cases, the holder is removed and the actuator is pulled to change the position of the device and reset it. In some cases, the device is configured to allow only single use.

[0032] As shown in FIG. 4D, when the piston moves downward, the piston 114 engages the patch 10, applying a quick and sufficient pressure to the patch such that the patch 10 is pressed against the patient's skin through the applicator's ring holder. The force actually applied is variable based on the particular patch configuration and the patient's target area. Generally, the piston applies the patch to the patient's skin target area at about 90 N to 130 N.

[0033] The cam path 140 is shown in four sections forming a loop that closely resembles a parallelogram, but the cam path 140 can be envisioned to have other shapes. For example, any shape can be used that forms a loop allowing for repeated movement of the protrusion through the cam path 140 (i.e., allowing the applicator 100 to be used repeatedly). For example, a suitable shape could be more rounded and near an oval shape, but generally, the cam path should have a downward section similar to the fourth section 143, as this allows the piston 114 to move quickly downward along the fourth section 143, enabling the patch to be pushed out of the applicator.

[0034] Generally, to enable a configuration that accurately orients the patch with respect to the patient's skin target area and allows the piston 114 to push the patch out of the applicator 100, the patch 10 is held in a ring-type holder. Specifically, the patch is held by a holder having a generally open area and a rigid rim or perimeter. The patch is secured to the inner rim of the ring holder at one or more points along the edge of the patch such that the patch spans the open area. The piston, when actuated, passes through the open area and pushes the patch downward.

[0035] Specific embodiments of the ring holder will be described below. For example, FIGS. 5A-5C are perspective, top, and bottom views of a ring holder 50 for a drug patch 10 according to an exemplary embodiment. The holder has a rigid periphery 58 and a central opening 56. The patch is fixed on the opening 56 and on the inner portion of the rim 58, as shown in FIGS. 5B and 5C. In some cases, the patch 10 is held at one or more points along its edge by a retaining ring 52.

[0036] As shown, the holder 50 is circular, but the use of the term ring does not mean that the holder must be circular. The holder may be of other shapes, such as square, elliptical, triangular, or other shapes, depending on factors such as the patch shape and configuration.

[0037] The ring holder 50 may be fixed to the bottom 120 of the actuator 122 using several connection mechanisms. For example, the holder 50 and the actuator 122 can be attached via an extension 54 having a clip, as shown in FIG. 7. Other connection mechanisms such as screwing, press-fitting, friction fitting, or adhesive connection may also be used.

[0038] Variations in the configuration of the ring holder and the method of holding the patch are conceivable. For example, FIGS. 5A-5C show a circular retaining ring 52, but the retaining ring may be of other shapes, including a ring 52'(FIGS. 9A-9C) having an extension 53 to assist in removal from and placement on the holder. Further, the retaining ring 52' may form a sandwich connection 57 (FIG. 9C) with flanges and grooves or similar structures to stabilize the connection between the retaining ring 52' and the patch.

[0039] Furthermore, the patch can be held in a holder attachable to the bottom of the actuator 122, although other configurations are conceivable. For example, as shown in FIG. 8, a small ring holder 60 with a retaining ring 62 may be used, and such a holder can be placed on the upper part of the actuator, thereby avoiding a larger ring holder attached to the bottom of the actuator. Further, instead of using a retaining ring, the patch 10 may be held by a separable connector. For example, FIG. 15 shows an alternative separating mechanism 1400 for holding a drug patch in a patch applicator. In the separating mechanism, a thin or relatively weak connector 1410 holds the patch to the applicator, and the connector 1410 is broken or torn by the pressure of the piston.

[0040] A suitable patch needs to be designed to be released from the actuator by the application of pressure from the piston 114. FIGS. 6A and 6B are exemplary patch configurations for use in the disclosed applicator and patch system. As shown, the patches 10, 10' can include backing layers 12, 12', and adhesive regions 16, and drug-containing regions 14. The adhesive region 16 can extend within the drug-containing region 14 (e.g., a microneedle array) as long as the adhesive is not disposed in such a way as to adversely affect the microneedles or the detachment of the microneedles from the skin.

[0041] As shown in FIG. 6A, the backing layer can extend from around the adhesion region 16 along the entire periphery, or as shown in FIG. 6B, for example, it can extend from a part of the corner adhesive. As described above, at least a part of the patch, for example, the backing layer, can have sufficient flexibility to allow the patch to be pushed through the ring holder of the applicator device 100. To provide sufficient flexibility, the backing layer may be formed of a material having mechanical properties and / or dimensions that provide the desired degree of flexibility. For example, materials suitable for the backing layer may be polyester (e.g., polyethylene terephthalate or polyethylene terephthalate glycol between about 0.002 inches or 0.005 inches thick), paper, aluminum, or other flexible materials.

[0042] Furthermore, the patch 10 (including the entire patch or the backing layer) can have various shapes. For example, FIGS. 16A-16C are alternative exemplary embodiments of the backing portion of a drug patch. The patch can include polygons (e.g., hexagon (FIG. 16A) or octagon), square (FIG. 16B), or circle (FIG. 16C). Furthermore, as shown in FIG. 6A or FIG. 6B, other shapes such as triangles, ovals, squares with rounded edges (scround or squircle) are also conceivable. Additionally, the flexibility of a specific region can also be enhanced by modifying the backing layer. For example, as shown in FIG. 16C, cuts or depressions 1500 can be provided in specific regions along the periphery of the backing layer.

[0043] Figures 11A, 11B, 12, 13, and 14 provide more details regarding the structure and interaction of the actuator 122 and the piston 114 according to various embodiments. Figures 11A and 11B are a perspective view and a side view showing the connection and movement of the actuator 122 within an exemplary patch applicator device. Figure 12 is a perspective view of the actuator shown in Figures 11A and 11B. As shown, the actuator includes a bottom 128 and an extension 126. Generally, the actuator and the outer body 110 are engaged to allow only a substantially linear sliding motion therebetween. Thus, the extension 126 can engage with the outer body via a groove, a tube, a runway, or a connection of the protrusion and cam path type. For example, the actuator 122 can have a protrusion 124 that engages with the linear cam path 111 of the body 110. It is contemplated that the configuration can be changed, for example, by placing the protrusion on the body and the path on the actuator 122. Further, although two extensions, protrusions, and cam paths are shown, three or more can be used for each.

[0044] As described above, the piston 114 can have several configurations. Generally, the piston is substantially cylindrical to allow rotation within the body 110 due to its circular cross-section. Figure 13 is a perspective view of the piston 114 of an exemplary patch applicator device. As shown, the piston 114 has a protrusion 130, an upper section 132, and a flange or expansion region 138 (all as described above).

[0045] The piston 114 has a bottom surface 136 configured to push down the patch. As shown, the surface 136 is convex, but it is contemplated that the surface can be flat, or other modifications such as a smooth surface or a textured surface can be added.

[0046] As described above with respect to the movement of the piston and actuator, the actuator pushes up the piston to compress the spring or compressible member, but when the piston reaches a particular point in the cam path, the piston is released, thereby releasing the energy stored in the now-compressed spring and pushing the piston down to apply the patch. The piston and actuator may be engaged in different configurations that allow the piston to be released from the actuator. In one embodiment, the piston can rotate relative to the actuator within the outer body.

[0047] Figures 14A-14C show the interaction between the actuator 122 and the piston 114 of the applicator according to various embodiments. As shown, the actuator extension 126 pushes on the flange or extended section 138 of the piston 114. However, the flange or extended section 138 does not extend entirely around the piston 114, and the extended section 138 has a gap 139 (Figs. 13B and 14C). When the projection 130 of the piston reaches the third downward section 143 of the cam path 140, the extension 126 of the actuator reaches the gap 139 of the piston extended section 138. This effect is due to the rotation of the piston 114 as the projection 130 passes through the loop-shaped cam path 140 along with the linear movement of the actuator. Thus, when the engagement between the piston projection 130 and the cam path 140 and the engagement between the piston extended section 138 and the actuator extension 126 are simultaneously released, the piston 114 is released downward.

[0048] The components including the piston 114, the actuator 122, and the outer body have been described with the interacting projections 130 and 124 and the cam paths 140 and 111, but it should be noted that the positions of the projections and cam paths can be changed. For example, the cam path may be disposed on the actuator and / or the piston having projections on the inner surface of the outer body 110, or a combination of such configurations.

[0049] Furthermore, although the ring holder is attached to the bottom surface of the actuator, the bottom surface of the actuator may be the skin contact surface of the device, and the path may be fixed to the upper surface of the bottom portion 128 of the actuator (for example, using the holder 60).

[0050] Detailed example of an exemplary patch As discussed above, the applicator can be used to apply a number of types of drug patches, but may be particularly desirable for the application of microneedle devices. Thus, suitable patches containing microneedles are described in more detail below. The applicator and / or the ring holder or sub-component can be provided as a kit or system including an applicator with one or more patches used with the applicator and patch, the patch and ring holder, or a reusable applicator. Suitable microneedle devices are further described in PCT Patent Application PCT / US2011 / 056856, entitled "Silk fibroin-based microneedles and methods of making the same", filed on October 19, 2011; PCT Patent Application PCT / US2019 / 025467, entitled "Microneedle comprising silk fibroin applied to a dissolvable base", filed on April 2, 2019; PCT Patent Application PCT / US2020 / 055139, entitled "Silk Fibroin-Based Microneedles and Uses Thereof", filed on October 9, 2020; PCT Patent Application PCT / US2021 / 033776, entitled "Compositions and devices for vaccine release and uses thereof", filed on May 21, 2021; and PCT Patent Application PCT / US2022 / 030177, entitled "Microneedle Vaccine Against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2)", filed on May 20, 2022, each of which is incorporated herein by reference in its entirety.

[0051] Suitable patches may preferably include silk fibroin-based microneedles and microneedle devices (e.g., microneedle arrays and patches) for the administration, transport, and release (e.g., controlled release or sustained release) of therapeutic agents such as vaccines, antigens, and / or immunogens (e.g., influenza vaccines and / or coronavirus vaccines, e.g., mRNA-based vaccines) through biological barriers such as the skin, mucosa, oral cavity, tissue, or cell membrane.

[0052] The term "administer" or "administering" includes the routes by which therapeutic agents are introduced into a subject to perform their intended functions. In certain embodiments, the administration of a therapeutic agent by a microneedle or microneedle device as described herein may be repeated, and this administration may be at intervals of at least about 1 day, 2 days, 3 days, 5 days, 10 days, 15 days, 30 days, 45 days, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 12 weeks, 2 months, 75 days, 3 months, or at least 6 months. In other embodiments, the administration of a therapeutic agent by a microneedle or microneedle device as described herein may be repeated annually. In other embodiments, the administration of a therapeutic agent by a microneedle or microneedle device as described herein may be repeated the number of times necessary to achieve a therapeutic or prophylactic effect. Administration "in combination" with one or more additional therapeutic agents includes simultaneous (parallel) administration and sequential administration in any order.

[0053] As used herein, "subject" refers to a human or an animal. Usually, the animal is a vertebrate, such as a primate, rodent, livestock, or game animal. Examples of primates include chimpanzees, cynomolgus monkeys, rhesus monkeys, and macaques (e.g., rhesus monkeys). Examples of rodents include mice, rats, woodchucks, ferrets, rabbits, and hamsters. Examples of livestock and game animals include cows, horses, pigs, deer, bison, buffalo, felines (e.g., domestic cats), canines (e.g., dogs, foxes, wolves), birds (e.g., chickens, emus, ostriches), and fish (e.g., trout, catfish, and salmon). In certain embodiments of the aspects described herein, the subject is a mammal (e.g., a primate, e.g., a human). The subject may be male or female. In certain embodiments, the subject is a mammal. The mammal may be a human, non-human primate, mouse, rat, dog, cat, horse, or cow, but is not limited to these examples. Further, the methods and formulations described herein may be used to treat livestock and / or pets. In some embodiments, the term "subject" is intended to include an organism capable of eliciting an immune response (e.g., a mammal, e.g., a human).

[0054] In certain embodiments, the subject is a human. The subject can be of any age. In one embodiment, the subject is an elderly human subject, e.g., 65 years of age or older. In one embodiment, the subject is a human subject who is not an elderly person, e.g., under 65 years of age. In one embodiment, the subject is a pediatric human subject, e.g., 18 years of age or younger. In one embodiment, the subject is an adult subject, e.g., over 18 years of age.

[0055] As used herein, the term "antigen" refers to a molecule (e.g., a gene product (e.g., a protein or peptide), a pathogen fragment, a whole pathogen, a viral vector, or a virus particle) that can induce a humoral and / or cellular immune response, for example, leading to the activation of B lymphocytes and / or T lymphocytes and / or innate immune cells and / or antigen-presenting cells. Any macromolecule containing a protein or peptide can also be an antigen. An antigen can also be derived from a genome and / or recombinant DNA. For example, any DNA containing a nucleotide sequence or a partial nucleotide sequence encoding a protein that can elicit an immune response encodes an "antigen". In some embodiments, an antigen need not be encoded only by the full-length nucleotide sequence of a gene, nor need it be encoded by a gene at all. In some embodiments, an antigen may be synthesized or derived from a biological sample, such as a tissue sample, a tumor sample, a cell, or a fluid containing other biological components. In some embodiments, an antigen can be derived from a virus, such as an inactivated virus, a virus-like particle, or a viral vector. The antigens used herein may also be a mixture of several individual antigens.

[0056] As used herein, the term "immunogen" refers to any substance (e.g., antigen, combination of antigens, pathogen fragment, whole pathogen) that can induce an immune response in an organism. An "immunogen" can induce an immunological response against itself after administration to a mammalian subject. As used herein, the term "immunological" with respect to an immune response refers to the occurrence of a humoral (antibody-mediated) and / or cellular (mediated by antigen-specific T cells or their secreted products) response in a recipient subject against an immunogen. Such responses can be an active response induced by administration of an immunogen or immunogenic peptide to a subject, or a passive response induced by administration of antibodies or sensitized T cells directed against an immunogen. In some embodiments, the immunogen is a coronavirus antigen. In some embodiments, the immunogen is a coronavirus. In some embodiments, the immunogen is an influenza virus. In some embodiments, the immunogen is a viral vaccine (e.g., a monovalent (also referred to as single-valent) or multivalent (also referred to as polyvalent) vaccine such as for coronavirus and / or influenza). In some embodiments, the vaccine (e.g., coronavirus vaccine and / or influenza vaccine) can be monovalent, bivalent, trivalent, tetravalent (also referred to as 4-valent), or pentavalent. In some embodiments, the immunogen is a replicating or non-replicating vaccine vector (e.g., including an adenovirus vector, adeno-associated virus vector, alphavirus vector, herpesvirus vector, measles virus vector, poxvirus vector, or vesicular stomatitis virus vector).

[0057] As used herein, the terms "therapeutic agent" and "active agent" are terms recognized in the art and refer to any chemical moiety that is a biologically, physiologically, or pharmacologically active substance that acts locally or systemically in a subject. Upon administration to a subject, various forms of therapeutic agents that can be released from the microneedles described herein into adjacent tissue or fluid may be used. Examples of therapeutic agents, also referred to as "drugs," are described in well-known references such as the Merck Index, Physicians Desk Reference, and The Pharmacological Basis of Therapeutics, which include, but are not limited to, pharmaceuticals, vitamins, mineral supplements, substances used in the treatment, prevention, diagnosis, cure, or alleviation of diseases or disorders such as viral infections, substances that affect the structure or function of the body, or prodrugs that become biologically active or more active after being placed in a physiological environment.

[0058] In certain embodiments, the therapeutic agent includes, but is not limited to, vaccines, antigens, and / or immunogens. In certain embodiments, the therapeutic agent includes a coronavirus vaccine, antigen, and / or immunogen. In certain embodiments, the therapeutic agent includes an influenza vaccine, antigen, and / or immunogen.

[0059] In certain embodiments, the therapeutic agent includes amino acid molecules such as, but not limited to, peptides and / or proteins. In certain embodiments, the therapeutic agent includes a recombinant protein vaccine.

[0060] In certain embodiments, the therapeutic agent includes nucleic acid molecules such as, but not limited to, deoxyribonucleic acid (DNA) molecules and / or ribonucleic acid (RNA) molecules. In certain embodiments, the therapeutic agent includes mRNA. In some embodiments, the therapeutic agent includes nucleic acid-based vaccines such as DNA-based vaccines and / or RNA-based vaccines. In some embodiments, the therapeutic agent includes mRNA-based vaccines.

[0061] As used herein, the term "vaccine" refers to any composition that induces a protective immune response in a subject exposed to the composition. The immune response can include induction of antibodies and / or induction of a T cell response. Typically, an "immune response" includes, but is not limited to, one or more of the following effects: production or activation of antibodies, B cells, helper T cells, suppressor T cells, and / or cytotoxic T cells, particularly with respect to an antigen (s) included in or derived from the composition or vaccine of interest. Preferably, the subject exhibits either a therapeutic or protective immunological (memory) response, resulting in enhanced resistance to new infections and / or reduced clinical severity of the disease. Such protection is evidenced by a decrease in the number or severity of clinical signs associated with pathogen infection, or the absence of one or more clinical signs, a delay in the onset of viremia, a decrease in virus persistence, a decrease in overall virus load, and / or a decrease in virus excretion. In some embodiments, a "vaccine" is an antigen or immunogen (including subunit antigens, toxoid antigens, conjugate antigens, or other types of antigen molecules, or nucleic acid molecules encoding them) or any preparation of killed or live attenuated microorganisms that, when introduced into a subject's body, affects the immune response to a specific antigen or microorganism by causing activation of the immune system (including, for example, induction of antibody formation, T cell response, and / or B cell response) against the specific antigen or microorganism. Generally, vaccines against microorganisms are directed at at least a portion of a virus, bacterium, parasite, mycoplasma, or other infectious agent.

[0062] The term "therapeutically effective amount" refers to the amount of a composition as defined herein that is effective to prevent, ameliorate, and / or treat a condition resulting from a disease described herein, such as a viral infection.

[0063] The term "treatment" refers to therapeutic measures and prophylactic or preventive means for curing or halting or at least retarding the progression of a disease. Those in need of treatment include those already suffering from a condition resulting from a viral infection as described herein, as well as those in whom viral infection should be prevented. Subjects who have partially or fully recovered from a viral infection as described herein may also be in need of treatment. Prevention includes steps for inhibiting or reducing the spread of a virus, or for inhibiting or reducing the onset, occurrence, or progression of one or more symptoms associated with a viral infection as described herein.

[0064] As used herein, the term "virus" refers to an infectious agent composed of a nucleic acid encapsulated in a protein. Such infectious agents are unable to replicate autonomously (i.e., replication requires the use of host cell machinery). The viral genome may be single-stranded (ss) or double-stranded (ds), RNA or DNA, and may or may not be able to use reverse transcriptase (RT). Further, ssRNA viruses may be either sense (+) or antisense (-). Exemplary viruses include, but are not limited to, dsDNA viruses (e.g., adenovirus, herpesvirus, poxvirus), ssDNA viruses (e.g., parvovirus), dsRNA viruses (e.g., reovirus), (+)ssRNA viruses (e.g., picornavirus, togavirus, coronavirus), (-)ssRNA viruses (e.g., orthomyxovirus, rhabdovirus), ssRNA-RT viruses, i.e., (+) sense RNA having DNA in the middle of its life cycle (e.g., retrovirus), and dsDNA-RT viruses (e.g., hepadnavirus). In some embodiments, the virus may also include wild-type (natural) virus, inactivated virus, live attenuated virus, modified virus, recombinant virus, or any combination thereof. An exemplary retrovirus is human immunodeficiency virus (HIV). Other examples of viruses include, but are not limited to, enveloped viruses, respiratory syncytial virus, non-enveloped viruses (e.g., human papillomavirus (HPV)), bacteriophage, recombinant viruses, and viral vectors. The term "bacteriophage" as used herein refers to a virus that infects bacteria.

[0065] As used herein, the term "coronavirus" refers to a plus-sense ssRNA virus of the genus Coronaviridae. Coronaviruses can be alpha coronaviruses, beta coronaviruses, gamma coronaviruses, or delta coronaviruses. Coronaviruses can be live wild-type viruses, live attenuated viruses, inactivated viruses (e.g., UV-inactivated viruses), chimeric viruses, or recombinant viruses. Coronaviruses are known to infect humans and other animals (e.g., birds and mammals). Examples of coronaviruses include severe acute respiratory syndrome coronavirus (SARS-CoV), severe acute respiratory syndrome virus 2 (SARS-CoV-2), Middle East respiratory syndrome coronavirus (MERS-CoV), human coronavirus 229E (HCoV-229E), human coronavirus NL63 (HCoV-NL63), human coronavirus OC43 (HCoV-OC43), and human coronavirus HKU1 (HCoV-HKU1).

[0066] As used herein, the term "influenza virus" refers to a negative-sense ssRNA virus of the family Orthomyxoviridae. Influenza viruses can be live wild-type viruses, live attenuated viruses, inactivated viruses, chimeric viruses, or recombinant viruses. Examples of influenza viruses include influenza A, influenza B, influenza C, and influenza D.

[0067] The microneedles described herein can be of any shape and / or geometry suitable for piercing a biological barrier (e.g., a layer of skin) to enable the release (e.g., controlled release or sustained release) of a vaccine within a subject. Non-limiting examples of the shape and / or geometry of the microneedles include cylindrical, wedge-shaped, conical, pyramidal, and / or irregular shapes, or any combination thereof.

[0068] As used herein, the terms “release” and “controlled release or sustained release” refer to the release of a vaccine, antigen, and / or immunogen (e.g., from a microneedle, microneedle device, formulation, composition, article, device, and preparation described herein, e.g., from a silk fibroin-based microneedle tip described herein), e.g., a coronavirus vaccine, an influenza vaccine, or a combination thereof, over a period of time, e.g., for at least about 1 to about 28 days (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, or 28 days or more, e.g., between about 4 days to about 25 days, between about 10 days to about 20 days, between about 10 days to about 15 days, between about 12 days to about 16 days, e.g., for about 1 to 2 weeks, about 1 to 3 weeks, or about 1 to 4 weeks, e.g., for about 1 month to about 3 months). In some embodiments, the controlled release or sustained release of a vaccine such as a coronavirus vaccine and / or an influenza vaccine over a period of about 1 day to about 14 days, e.g., for a period of about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14 days, can result in broad-spectrum immunity in a subject by a microneedle, microneedle device, formulation, composition, article, device, or preparation described herein. In some embodiments, vaccine formulations and preparations containing silk fibroin have controlled release or sustained release properties (e.g., the vaccine is formulated and / or configured to release over a period of time, or for at least 1, 5, 10, 15, 30, 45 minutes; over a period of time, or for at least 1, 2, 3, 4, 5, 10, 24 hours; over a period of time, or for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 days; over a period of time, or for at least 1, 2, 3, 4, 5, 6, 7, 8 weeks; over a period of time, or for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 months; over a period of time, or for at least 1, 2, 3, 4, 5 years, or more, e.g., into the subject's skin).

[0069] In some embodiments, the microneedles of the present invention can include the following layers: (1) a backing material; (2) a solubility base; and (3) a transplantable controlled-release or sustained-release tip. For example, the microneedles described herein can include a backing material applied to a solubility base layer that supports a distal controlled-release or sustained-release transplantable tip containing silk fibroin and a vaccine (e.g., an influenza vaccine and / or a coronavirus vaccine, e.g., an mRNA-based vaccine, an antigen, and / or an immunogen).

[0070] As used herein, the term "backing" refers to a material suitable for binding and / or adhering to components of the microneedle. In some embodiments, the backing material is suitable for binding and / or adhering to the base (e.g., the solubility base) of the microneedles described herein.

[0071] As used herein, the term "base" or "solubility base" refers to a layer that forms the base of the microneedle (e.g., functions as a support for the distal microneedle tip (e.g., a silk fibroin tip) loaded with a vaccine, an antigen, and / or an immunogen (e.g., a coronavirus vaccine, an influenza vaccine, or a combination thereof)) and / or also functions as a layer connecting adjacent microneedles to form a continuous microneedle array or microneedle patch. In some embodiments, at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more of the base is dissolved after application to a biological barrier, such as the skin, mucosal surface, or oral cavity.

[0072] As used interchangeably herein, the terms "sustained release tip," "implantable sustained release tip," "implantable microneedle tip," or "releasable tip" refer to the distal end, e.g., the tip, of a microneedle that can penetrate a biological barrier, such as the skin, mucosal surface, or oral cavity of a subject, and can deposit within a biological barrier, such as a skin layer (e.g., the dermis). In embodiments, the tip contains an amount of silk fibroin protein sufficient to maintain the release of a vaccine, e.g., a coronavirus vaccine (e.g., a SARS-CoV-2 vaccine) and / or an influenza vaccine, for an extended period of time, e.g., at least about 1 day (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14 or more days, e.g., between about 4 days to about 30 days, between 5 days to about 25 days, between about 10 days to about 20 days, between about 10 days to about 15 days, between about 4 days to about 14 days, between about 14 days to about 15 days, e.g., about 1 to 2 weeks, about 1 to 3 weeks, or about 1 to 4 weeks, e.g., about 2 to 12 months). In some embodiments, the implantable sustained release tip contains a coronavirus vaccine, an antigen, and / or an immunogen. In some embodiments, the implantable sustained release tip contains an influenza vaccine, an antigen, and / or an immunogen.

[0073] As used herein, the term "microneedle" refers to a structure having at least two, more typically three components, such as layers, for transporting or administering a vaccine, antigen, and / or immunogen across a biological barrier, such as skin, tissue, or cell membrane. In some embodiments, the microneedle includes a base (e.g., the solubility base described herein), a tip (e.g., the implantable tip described herein), and optionally a backing material. In embodiments, the microneedle has dimensions with a height between about 350 μm and about 1500 μm (e.g., about 350 μm to about 1500 μm, e.g., about 350 μm, about 400 μm, about 450 μm, about 500 μm, about 550 μm, about 600 μm, about 650 μm, about 700 μm, about 750 μm, about 800 μm, about 850 μm, about 900 μm, about 950 μm, about 1000 μm, about 1050 μm, about 1100 μm, about 1150 μm, about 1200 μm, about 1250 μm, about 1300 μm, about 1350 μm, about 1400 μm, about 1450 μm, about 1500 μm). In some embodiments, the microneedle is fabricated to have any dimensions and / or geometries that enable deployment of a microneedle tip (e.g., a silk fibroin tip), such as an implantable sustained-release tip, at a depth between about 100 μm and about 900 μm (e.g., at a depth of about 800 μm) into the dermal layer of the skin for controlled or sustained release of a vaccine (e.g., a coronavirus vaccine and / or an influenza vaccine).

[0074] As used herein, the terms "microneedle patch" and "microneedle array" refer to devices that include a plurality of microneedles, such as silk fibroin-based microneedles, arranged in a random or predefined pattern, such as an array.

[0075] In some embodiments, the length of the microneedle can be between about 350 μm and about 1500 μm (e.g., about 350 μm, about 400 μm, about 450 μm, about 500 μm, about 550 μm, about 600 μm, about 650 μm, about 700 μm, about 750 μm, about 800 μm, about 850 μm, about 900 μm, about 950 μm, about 1000 μm, about 1050 μm, about 1100 μm, about 1150 μm, about 1200 μm, about 1250 μm, about 1300 μm, about 1350 μm, about 1400 μm, about 1450 μm, about 1500 μm). In embodiments, the length of the microneedle can be made, as described herein, of a length sufficient to administer a transplantable tip containing a vaccine, antigen, and / or immunogen for controlled or sustained release to the epidermis (e.g., from about 10 μm to about 120 μm below the skin surface), for example, to induce an immune response. In some embodiments, the length of the microneedle can be made, as described herein, of a length sufficient to enable administration of a transplantable tip containing a vaccine, antigen, and / or immunogen for controlled or sustained release to the dermis (e.g., from about 60 μm to about 2.1 mm below the skin surface). One of ordinary skill in the art can adjust the microneedle length with respect to a number of factors including, but not limited to, the thickness of the tissue, such as the thickness of the skin (e.g., age, gender, body location, subject species (e.g., animal), drug administration profile, diffusion characteristics of the vaccine, antigen, and / or immunogen for controlled or sustained release (e.g., as a function of the ionic charge and / or molecular weight and / or shape of the vaccine, antigen, and / or immunogen for controlled or sustained release), or any combination thereof. However, without wishing to be bound by theory, a sustained release tip transplantable at a depth between about 100 μm and about 600 μm within the dermal layer of the subject's skin can be positioned to achieve controlled or sustained release of the vaccine from the tip using a microneedle having a height of about 650 μm. In some embodiments, the height of the microneedle can be about 800 μm (e.g., between about 500 μm and 1200 μm in height).

[0076] Exemplary microneedles of the present invention are shown in FIGS. 5A - 5B.

[0077] In some embodiments, as described herein, multiple microneedles can be arranged randomly or in a predetermined pattern to form a microneedle array and / or patch. The patch can include a carrier, backing, or "handle" layer adhered to the back of the base (see, e.g., FIG. 4). This layer can provide structural support and an area where the patch can be processed and manipulated without disturbing the array of needles.

[0078] Microneedle array The microneedle array can include about 121 needles in an 11×11 square grid with a pitch of about 0.75 mm. The individual needles are about 0.65 mm in length and are cones with a base diameter of about 0.35 mm and an included angle of about 30°. The tip of the needle must be sharp enough to penetrate the skin. The radius of curvature of the tip should ideally be 0.01 mm or less.

[0079] Backing Exemplary backing materials that can be used in the fabrication of the microneedles of the present invention include, but are not limited to, solid supports such as paper - based materials, plastic materials, polymer materials, or polyester - based materials (e.g., Whatman 903 paper, polymer tape, plastic tape, adhesive - coated polyester tape, or other medical tapes). In some embodiments, the backing includes Whatman 903 paper. In some embodiments, the backing includes a polyester tape. In some embodiments, the polyester tape includes an adhesive - coated polyester tape. In some embodiments, the backing material may be coated (e.g., on at least one side) with an adhesive suitable for binding and / or adhering to the solubility - based microneedles described herein.

[0080] The backing material used in the microneedles of the present invention may have various characteristics including, but not limited to, the ability to bind and / or adhere to the solubility-based layer to enable release. The backing material, for example, if the solubility-based layer has cracks or discontinuities, must be strong enough for the backing to maintain the integrity of the patch. The backing material may, for example, be sufficiently flexible to conform to a non-flat surface such as the skin surface. In particular, the backing may be sufficiently flexible during the wearing time, such as after the patch has been applied to the skin (e.g., pressed against the skin). The backing may include and / or consist of a non-soluble material such that the backing maintains its integrity after application of the patch to the skin surface and during removal of the patch from the skin surface.

[0081] The backing can have any dimensions suitable for application to the target skin surface. In some embodiments, the dimensions of the backing can be a circle with a diameter of 12 mm. In some embodiments, the dimensions of the backing can be a 12 mm wide strip with a "handle" section up to 12 mm in length extending beyond the edge of a 12 mm × 12 mm patch.

[0082] Solubility-based The solubility-based layer forms the base of the conical needles (e.g., functions as a support for the distal silk fibroin tip loaded with vaccine, antigen, and / or immunogen). The solubility-based layer can also function as a layer that connects adjacent needles to form a microneedle array or patch. In some embodiments, the solubility-based layer constitutes less than 98% (e.g., less than about 98%, less than about 90%, less than about 80%, less than about 70%, less than about 60%, less than about 50%, less than about 40%, less than about 30%, less than about 20%, less than about 10%, less than about 9%, less than about 8%, less than about 7%, less than about 6%, less than about 5%, less than about 4%, less than about 3%, less than about 2%, less than about 1%) of the total amount (e.g., dose) of vaccine, antigen, and / or immunogen loaded into the microneedles and / or microneedle device. In some embodiments, the solubility-based layer does not contain, for example, a detectable amount of vaccine, antigen, and / or immunogen. In some embodiments, the solubility-based layer is formulated to limit and / or reduce the amount of leakage (e.g., diffusion) of vaccine, antigen, and / or immunogen from the silk fibroin tip to the solubility-based layer as compared to, for example, a base layer formulation known in the art, such as a base layer formulation containing PAA. In some embodiments, the limited amount and / or decreased amount of leakage (e.g., diffusion) of vaccine, antigen, and / or immunogen from the silk fibroin tip can be determined, for example, after fabrication and storage (e.g., storage at about 4°C (e.g., refrigeration), about 25°C (e.g., room temperature), about 37°C (e.g., body temperature), about 45°C, and / or about 50°C) for about 1 day, about 2 days, about 3 days, about 4 days, about 5 days, or about 6 days; about 1 week, about 2 weeks, or about 3 weeks; about 1 month, about 2 months, about 3 months, about 4 months, about 5 months, about 6 months, about 7 months, about 8 months, about 9 months, about 10 months, or about 11 months; or about 1 year or more as compared to, for example, a base layer formulation containing PAA.

[0083] The solubility-based layer includes, for example, a material that can dissolve in the skin within an intended wearing time (e.g., about 5 minutes). In some embodiments, at least about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% of the solubility-based layer dissolves within the intended wearing time (e.g., within about 1 minute, about 2 minutes, about 3 minutes, about 4 minutes, about 5 minutes, about 6 minutes, about 7 minutes, about 8 minutes, about 9 minutes, or about 10 minutes or more) after application to the skin, for example.

[0084] The materials used to fabricate the solubility-based layer must be strong enough for the microneedles to penetrate the skin and also robust enough to allow for release (e.g., not overly fragile). The solubility-based material must be suitable for everyday handling without catastrophic failure and retain its mechanical properties between release and application (e.g., not hygroscopic enough to melt due to ambient humidity). The solubility-based layer material must be non-toxic and non-reactive at the dosages used in the patch. In some embodiments, the solubility-based layer includes water-soluble components. In some embodiments, the solubility-based layer described herein has improved biocompatibility compared to, for example, a solubility-based layer that includes poly(acrylic acid) (PAA). In some embodiments, the solubility-based layer material causes a reduction in the inflammatory response and / or a reduction in tissue necrosis. In some embodiments, the solubility-based layer material is not PAA and induces a reduction in the inflammatory response and / or a reduction in tissue necrosis compared to PAA. In some embodiments, the solubility-based layer material has a pH similar to the pH of the biological barrier being dissolved, e.g., a pH of about 4.0 to about 8.0.

[0085] Non-limiting examples of materials that can be used in the fabrication of the solubility-based layer include gelatin (e.g., hydrolyzed gelatin), polyethylene glycol (PEG), sucrose, low-viscosity carboxymethyl cellulose (CMC), polyvinylpyrrolidone (PVP), polyvinyl alcohol (PVA), hyaluronate, maltose, and / or methylcellulose. In some embodiments, the solubility-based layer comprises one, two, three, four, five, six, seven, eight, or more (e.g., all) of gelatin, polyethylene glycol (PEG), sucrose, carboxymethyl cellulose (CMC), polyvinylpyrrolidone (PVP), polyvinyl alcohol (PVA), hyaluronate, maltose, and methylcellulose at a concentration of, for example, about 1% to about 75% (e.g., about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 15%, about 20%, about 25%, about 30%, or about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, or about 75%). In some embodiments, the solubility-based layer does not contain the therapeutic agent described herein.

[0086] In some embodiments, the solubility-based layer comprises about 10% to about 70% gelatin (e.g., hydrolyzed gelatin) (e.g., about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, or about 70% gelatin).

[0087] In some embodiments, the solubility-based layer comprises about 1% to about 70% polyethylene glycol (PEG) (e.g., about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, or about 70% PEG).

[0088] In some embodiments, the solubility-based layer comprises about 1% to about 35% sucrose (e.g., about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 15%, about 20%, about 25%, about 30%, or about 35% sucrose).

[0089] In some embodiments, the solubility base comprises from about 1% to about 35% CMC (e.g., about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 15%, about 20%, about 25%, about 30%, or about 35% CMC).

[0090] In some embodiments, the solubility base comprises from about 10% to about 70% PVP (e.g., about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, or about 70% PVP).

[0091] In some embodiments, the solubility base comprises from about 1% to about 35% PVA (e.g., about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 15%, about 20%, about 25%, about 30%, or about 35% PVA).

[0092] In some embodiments, the solubility base comprises from about 1% to about 75% (e.g., about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 15%, about 20%, about 25%, about 30%, or about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, or about 75% hyaluronate).

[0093] In some embodiments, the solubility base comprises from about 1% to about 75% (e.g., about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 15%, about 20%, about 25%, about 30%, or about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, or about 75% maltose).

[0094] In some embodiments, the solubility base comprises from about 1% to about 75% (e.g., about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 15%, about 20%, about 25%, about 30%, or about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, or about 75% methylcellulose).

[0095] In some embodiments, the soluble base layer comprises 40% hydrolyzed gelatin and 10% sucrose w / v in DI water. Optionally, the base layer may include 1% low-viscosity carboxymethyl cellulose (CMC) which can reduce brittleness. In some embodiments, the soluble base layer may comprise polyvinylpyrrolidone (PVP) with a MW of 10 kD at up to 50% w / v in DI water; polyvinyl alcohol (PVA) with a MW of 13 kD hydrolyzed by 87% at up to 20% in DI water; or CMC at up to 10% in DI water. The following combinations, namely, 30% PVP and 10% PVA; 37% PVP, 5% PVA, and 15% sucrose; or various other ratios of PVP, PVA, and sucrose may be suitable for use in fabricating the soluble base layer.

[0096] In some embodiments, the soluble base layer is a square approximately 12 mm in size and 0.75 mm thick. In some embodiments, the soluble base layer can cover the entire patch. In some embodiments, the dimensions of the base layer can be a circle with a diameter of 12 mm, or a square of 12×12 mm.

[0097] Transplantable sustained-release tip In embodiments, the implantable sustained release tip can be fabricated from silk fibroin and can contain a vaccine, antigen, and / or immunogen (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccine) as described herein. In some embodiments, since the population of antigen-presenting cells in the dermis is much higher than in the subcutaneous space, the implantable sustained release tip may be designed to be deployed in the dermal layer of the skin (e.g., rather than in the subcutaneous space). In humans, the thickness of the dermis ranges from about 1000 to 2000 μm (e.g., about 1 to 2 mm), depending on location and the age and health of the patient. In rodents, the dermis is much thinner (e.g., about 100 to 300 μm in mice and about 800 to 1200 μm in rats). Without wishing to be bound by theory, an implantable sustained release tip can be deployed at a depth between about 100 μm and about 600 μm using a 650-μm high microneedle to achieve controlled or sustained release of a vaccine, antigen, and / or immunogen (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccine) as described herein.

[0098] While not wishing to be bound by theory, the molecular weight of the silk fibroin solution used in the fabrication of the microneedles described herein can function as a controlling factor for regulating the controlled or sustained release of vaccines, antigens, and / or immunogens (e.g., influenza vaccines and / or coronavirus vaccines, e.g., mRNA-based vaccines) from the tip. In some embodiments, a higher molecular weight silk fibroin solution may advantageously act for a slower controlled or sustained release (e.g., reducing the amount of the initial burst (e.g., the amount released on day 0) by at least about 10% and then releasing additional antigen for at least the next about 4 days). In some embodiments, the controlled or sustained release of vaccines, antigens, and / or immunogens (e.g., influenza vaccines and / or coronavirus vaccines, e.g., mRNA-based vaccines) from the tip can occur over at least about 4 days (e.g., about 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14 days or more, e.g., from about 4 days to about 14 days, e.g., about 1 to 2 weeks, about 1 to 3 weeks, or about 1 to 4 weeks). In some embodiments, the controlled or sustained release occurs over about 1 week to about 2 weeks.

[0099] In an embodiment, the silk fibroin solution used in the production of the microneedles described herein may be a low molecular weight silk fibroin composition containing a population of silk fibroin fragments having a range of molecular weights, wherein no more than 15% of the total number of silk fibroin fragments in the population have a molecular weight exceeding 200 kDa, and at least 50% of the total number of silk fibroin fragments in the population have a molecular weight within a specific range, the specific range being between about 3.5 kDa and about 120 kDa, or between about 5 kDa and about 125 kDa. In other words, the silk fibroin solution used in the production of the microneedles described herein may contain a population of silk fibroin fragments having a range of molecular weights, wherein no more than 15% of the total moles of silk fibroin fragments in the population have a molecular weight exceeding 200 kDa, and at least 50% of the total moles of silk fibroin fragments in the population have a molecular weight within a specific range, the specific range being between about 3.5 kDa and about 120 kDa or between about 5 kDa and about 125 kDa (see, for example, WO 2014 / 145002, which is incorporated herein by reference).

[0100] Exemplary silk fibroin (e.g., regenerated silk fibroin) solutions can have different molecular weight profiles, as determined, for example, by size exclusion chromatography (SEC) methods (see, e.g., FIG. 5). In some embodiments, the silk fibroin solution may be prepared, for example, according to established methods. In some embodiments, pieces of cocoons from Bombyx mori are first boiled in 0.02 M Na2CO3 to remove sericin proteins present in untreated natural silk prior to SEC analysis. In some embodiments, the silk fibroin composition is a composition or mixture produced by refining cocoons from Bombyx mori at atmospheric boiling temperature for about 480 minutes or less, such as less than 480 minutes, less than 400 minutes, less than 300 minutes, less than 200 minutes, less than 180 minutes, less than 120 minutes, less than 100 minutes, less than 60 minutes, less than 50 minutes, less than 40 minutes, less than 30 minutes, less than 20 minutes, less than 10 minutes, or even shorter times. In one embodiment, the silk fibroin composition is a composition or mixture produced by refining silk cocoons in an aqueous sodium carbonate solution at atmospheric boiling temperature for about 480 minutes or less, such as less than 480 minutes, less than 400 minutes, less than 300 minutes, less than 200 minutes, less than 180 minutes, less than 120 minutes, less than 100 minutes, less than 60 minutes, less than 50 minutes, less than 40 minutes, less than 30 minutes, less than 20 minutes, less than 10 minutes, or even shorter times.

[0101] In some embodiments, the silk fibroin solution may be a silk fibroin solution boiled for 10 minutes (10MB), 60 minutes (60MB), 120 minutes (120MB), 180 minutes (180MB), or 480 minutes (480MB) (see, e.g., FIG. 5). In some embodiments, the influenza vaccine, antigen, and / or immunogen can be formulated in a 1% w / v to about 10% w / v (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10% w / v) 10MB silk fibroin solution. In some embodiments, the influenza vaccine, antigen, and / or immunogen can be formulated in a 1% w / v to about 10% w / v (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10% w / v) 60MB silk fibroin solution. In some embodiments, the influenza vaccine, antigen, and / or immunogen can be formulated in a 1% w / v to about 10% w / v (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10% w / v) 120MB silk fibroin solution.

[0102] In some embodiments, the influenza vaccine, antigen, and / or immunogen can be formulated in a 1% w / v to about 10% w / v (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10% w / v) 180MB silk fibroin solution. In some embodiments, the influenza vaccine, antigen, and / or immunogen can be formulated in a 1% w / v to about 10% w / v (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10% w / v) 480MB silk fibroin solution.

[0103] While not wishing to be bound by theory, the main tunability of the implantable sustained release tip is the crystallinity measured via the β-sheet content (intermolecular and intramolecular β-sheets). This affects both the solubility of the silk tip matrix and the antigen retention ability. As the β-sheet content increases, the mechanical strength of the tip also becomes higher. A specific vaccine release profile can be achieved by modulating the crystallinity and diffusivity of the silk matrix. This is accomplished by both the silk input material and formulation, and post-treatment (e.g., water annealing, methanol / solvent annealing) to increase crystallinity. In some embodiments, the implantable controlled release or sustained release microneedle tip contains a beta-sheet content between about 10% and about 60% (e.g., about 10%, about 20%, about 30%, about 40%, about 50%, about 60%), based on, for example, a "crystallinity index" such as those known in the art. In some embodiments, the implantable controlled release or sustained release microneedle tip can be formulated as particles (e.g., microparticles and / or nanoparticles).

[0104] Dimensions of the Implantable Sustained Release Tip The methods provided herein can be used, for example, to fabricate silk fibroin-based implantable sustained release tips having any dimension in the range of a height / length of from about 75 μm to about 800 μm (e.g., about 75, about 100 μm, about 125 μm, about 150 μm, about 250 μm to about 300 μm, about 300 μm to about 350 μm, about 350 μm to about 400 μm, about 400 μm to about 450 μm, about 450 μm to about 500 μm, about 500 μm to about 550 μm, about 550 μm to about 600 μm, about 600 μm to about 650 μm, about 650 μm to about 700 μm, about 700 μm to about 750 μm, about 750 μm to about 800 μm) and / or a tip radius of about 10 μm or less (e.g., from about 1 μm to about 10 μm, e.g., about 1 μm or less, about 2 μm or less, about 3 μm or less, about 4 μm or less, about 5 μm or less, about 6 μm or less, about 7 μm or less, about 8 μm or less, about 9 μm or less, or about 10 μm or less). In some embodiments, the implantable tip can have a diameter of any size, e.g., based on the type of biological barrier (e.g., skin layer) intended to be pierced by the tip. In embodiments, the tip can have a dimension (e.g., diameter) in the range of from about 50 nm to about 50 μm (e.g., from about 50 nm to about 250 nm, from about 250 nm to about 500 nm, from about 500 to about 750 nm, from about 750 nm to about 1 μm, from about 1 μm to about 5 μm, from about 5 μm to about 10 μm, from about 10 μm to about 15 μm, from about 15 μm to about 20 μm, from about 20 μm to about 25 μm, from about 25 μm to about 30 μm, from about 30 μm to about 35 μm, from about 35 μm to about 40 μm, from about 40 μm to about 45 μm, or from about 45 μm to about 50 μm). It can be appreciated that there are no fundamental limitations preventing the sustained release tip from having an even smaller diameter (e.g., the limits of silk replica casting are demonstrated at resolutions of tens of nm. See, e.g., Perry et al., 20 Adv. Mat. 3070 (2008)).

[0105] In some embodiments, the sharpness of the tip of the implantable sustained release tip is described herein with respect to the tip radius. The molds used to fabricate the microneedles described herein are designed to have a tip radius between about 0.5 μm and about 10 μm (e.g., about 0.5 μm, 0.6 μm, 0.7 μm, 0.8 μm, 0.9 μm, 1 μm, 2 μm, 3 μm, 4 μm, 5 μm, 6 μm, 7 μm, 8 μm, 9 μm, or 10 μm). In some embodiments, the tip radius is between about 20 μm and about 25 μm (e.g., about 20 μm, 21 μm, 22 μm, 23 μm, 24 μm, or 25 μm). Without being bound by theory, it will be understood that blunter needles may require more force to penetrate the epidermis. In embodiments, other dimensions of the implantable sustained release tip may be controlled by the shape of the mold and the fill volume. In some embodiments, the implantable sustained release tip may have an included angle between about 5 degrees and about 45 degrees (e.g., about 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, or 45 degrees). In some embodiments, the implantable sustained release tip may have an included angle between about 15 degrees and 45 degrees (e.g., about 15 degrees, about 16 degrees, about 17 degrees, about 18 degrees, about 19 degrees, about 20 degrees, about 21 degrees, about 22 degrees, about 23 degrees, about 24 degrees, about 25 degrees, about 26 degrees, about 27 degrees, about 28 degrees, about 29 degrees, about 30 degrees, about 31 degrees, about 32 degrees, about 33 degrees, about 34 degrees, about 35 degrees, about 36 degrees, about 37 degrees, about 38 degrees, about 39 degrees, about 40 degrees, about 41 degrees, about 42 degrees, about 43 degrees, about 44 degrees, or about 45 degrees).

[0106] In embodiments, the height of the implantable sustained release tip depends on the formulation and fill volume, which can affect surface tension and drying kinetics. In some embodiments, the height of the implantable sustained release tip may extend up to half of the total height of the microneedle. In some embodiments, the height of the implantable sustained release tip is between about 75 μm and about 475 μm (e.g., about 75, about 100 μm, about 125 μm, about 150 μm, about 175 μm, about 200 μm, about 225 μm, about 250 μm, about 275 μm, about 300 μm, about 325 μm, about 375 μm, about 400 μm, about 425 μm, or about 475 μm). In some embodiments, the base of the tip includes a thin "shell"-like layer having a thickness between about 5 and 10 μm (e.g., a thickness of about 5, 6, 7, 8, 9, or 10 μm). In some embodiments, the implantable sustained release tip can become a stiffer structure with a dried minimal "shell", and the height can be close to 150 μm (e.g., between about 50 μm and about 200 μm) and the thickness >50 μm (e.g., between about 25 μm and about 75 μm).

[0107] Furthermore, the microneedles of the present invention may utilize techniques known in the art that have been developed, for example, to functionalize silk fibroin (such as active agents like dyes and sensors). For example, U.S. Patent No. 6,287,340, Bioengineered anterior cruciate ligament; International Publication No. WO 2004 / 000915, Silk Biomaterials & Methods of Use Thereof; International Publication No. WO 2004 / 001103, Silk Biomaterials & Methods of Use Thereof; International Publication No. WO 2004 / 062697, Silk Fibroin Materials & Use Thereof; International Publication No. WO 2005 / 000483, Method for Forming inorganic Coatings; International Publication No. WO 2005 / 012606, Concentrated Aqueous Silk Fibroin Solution & Use Thereof; International Publication No. WO 2011 / 1005381, Vortex-Induced Silk fibroin Gelation for Encapsulation & Delivery; International Publication No. WO 2005 / 123114, Silk-Based Drug Delivery System; International Publication No. WO 2006 / 076711, Fibrous Protein Fusions & Uses Thereof in the Formation of Advanced Organic / lnorganic Composite Materials; U.S. Patent Application Publication No. 2007 / 0212730, Covalently immobilized protein gradients in three-dimensional porous scaffolds; International Publication No. WO 2006 / 042287, Method for Producing Biomaterial Scaffolds, International Publication No. WO 2007 / 016524, Method for Stepwise Deposition of Silk Fibroin Coatings;International Publication No. WO 2008 / 085904, "Biodegradable Electronic Devices"; International Publication No. WO 2008 / 1118133, "Silk Microspheres for Encapsulation & Controlled Release"; International Publication No. WO 2008 / 1108838, "Microfluidic Devices & Methods for Fabricating Same"; International Publication No. WO 2008 / 1127404, "Nanopattemed Biopolymer Device & Method of Manufacturing Same"; International Publication No. WO 2008 / 1118211, "Biopolymer Photonic Crystals & Method of Manufacturing Same"; International Publication No. WO 2008 / 1127402, "Biopolymer Sensor & Method of Manufacturing Same"; International Publication No. WO 2008 / 1127403, "Biopolymer Optofluidic Device & Method of Manufacturing the Same"; International Publication No. WO 2008 / 1127401, "Biopolymer Optical Wave Guide & Method of Manufacturing Same"; International Publication No. WO 2008 / 1140562, "Biopolymer Sensor & Method of Manufacturing Same"; International Publication No. WO 2008 / 1127405, "Microfluidic Device with Cylindrical Microchannel & Method for Fabricating Same"; International Publication No. WO 2008 / 1106485, "Tissue-Engineered Silk Organs"; International Publication No. WO 2008 / 1140562, "Electroactive Bioploymer Optical & Electro-Optical Devices & Method of Manufacturing Same"; International Publication No. WO 2008 / 1150861, "Method for Silk Fibroin Gelation Using Sonication"See International Publication No. WO 2007 / 1103442, Biocompatible Scaffolds & Adipose-Derived Stem Cells; International Publication No. WO 2009 / 1155397, Edible Holographic Silk Products; International Publication No. WO 2009 / 1100280, 3-Dimensional Silk Hydroxyapatite Compositions; International Publication No. WO 2009 / 061823, Fabrication of Silk Fibroin Photonic Structures by Nanocontact Imprinting; International Publication No. WO 2009 / 1126689, System & Method for Making Biomaterial Structures;

[0108] In various embodiments, the silk fibroin-based microneedle tip can further comprise at least one additional therapeutic agent, which can be dispersed throughout the microneedle or form at least a portion of the microneedle tip. In some embodiments, the additional therapeutic agent is useful for treating viral infections as described herein. Optionally, the silk fibroin-based microneedle tip can further comprise excipients and / or adjuvants as described herein.

[0109] Viruses, Antigens, and Immunogens In some embodiments, the present invention provides for the administration of various therapeutic agents such as vaccines, antigens, and / or immunogens derived from viruses that are members of the Orthomyxoviridae family, e.g., controlled or sustained administration, by, for example, the formulations, compositions, articles, devices, formulations, microneedles, and / or microneedle devices (e.g., microneedle patches) described herein, and / or according to the methods described herein. In some embodiments, the vaccines, microneedles, and / or microneedle devices (e.g., microneedle patches) described herein may comprise negative sense ssRNA viruses and / or RNA viruses (e.g., influenza virus). In some embodiments, the vaccine, antigen, and / or immunogen comprises nucleic acid (e.g., DNA and / or RNA) derived from an influenza virus. In some embodiments, the vaccine, antigen, and / or immunogen comprises amino acids (e.g., peptides and / or proteins) derived from an influenza virus. In some embodiments, the influenza vaccine, antigen, and / or immunogen comprises inactivated and / or live attenuated virions, or split virions of an influenza virus. In some embodiments, the vaccine and / or microneedle comprises a non-replicating viral antigen.

[0110] In particular, the present invention contemplates vaccines, microneedles, and / or microneedle devices (e.g., microneedle patches) comprising an influenza virus vaccine, antigen, and / or immunogen. The influenza virus is an RNA virus (e.g., a linear negative sense single-stranded RNA virus). There are four known genera of influenza virus, each containing a single type (e.g., influenza A, B, C, and D). The influenza virus can change continuously and is subject to both antigenic drift and antigenic shift. Exemplary influenza strains are further described in the Examples (e.g., see Tables 1 and 2).

[0111] Influenza A can be classified into subtypes based on two proteins, hemagglutinin (HA) and neuraminidase (NA), on the surface of the virus. Influenza A includes 18 known HA subtypes, referred to herein as H1 to H18, and 11 known NA subtypes, referred to herein as N1 to N11. Many different combinations of HA and NA proteins may be found on the surface of influenza A viruses. For example, the "H1N1 virus" refers to an influenza A virus subtype that includes the H1 protein and the N1 protein. Exemplary influenza A virus subtypes that have been confirmed to infect humans include, but are not limited to, H1N1, H3N2, H2N2, H5N1, H7N7, H1N2, H9N2, H7N2, H7N3, H10N7, and H7N9. The H1N1 virus and the H3N2 virus are currently generally widespread among humans.

[0112] Exemplary influenza B viruses can belong to, for example, the B / Yamagata lineage and / or the B / Victoria lineage.

[0113] Vaccine Non-limiting examples of influenza vaccines for use with the microneedles and microneedle devices (e.g., microneedle patches) described herein can include commercially available vaccines such as seasonal vaccines, pandemic vaccines, and / or universal vaccines; egg-based vaccines, cell culture-based vaccines; recombinant vaccines; live attenuated, inactivated whole virus, split virion, and / or protein subunit vaccines; and adjuvanted vaccines. Various commercially available influenza vaccines are listed below. Further, influenza vaccines containing mRNA, DNA, viral vectors, and / or virus-like particles (VLPs) are suitable for use with the microneedles and microneedle devices (e.g., microneedle patches) described herein. In some embodiments, the influenza vaccine can target matrix protein 1, matrix protein 2 (M2e), and / or nucleoprotein (NP) of the influenza virus.

[0114]

Table 1

[0115] Vaccine formulations and compositions for controlled or sustained release At least one vaccine, antigen, and / or immunogen described herein (e.g., at least one vaccine, antigen, and / or immunogen derived from an influenza virus described herein) can be incorporated into various formulations, compositions, articles, devices, and / or preparations for administration to achieve, for example, controlled release and / or sustained release. More specifically, at least one vaccine, antigen, and / or immunogen described herein (e.g., at least one vaccine, antigen, and / or immunogen derived from an influenza virus described herein) can be formulated into a formulation, composition, article, device, and / or preparation by combining it with a suitable pharmaceutically acceptable carrier or diluent, and formulated into the preparation in semi-solid, solid, or liquid form. In some embodiments, the formulations, compositions, articles, devices, and / or preparations described herein contain silk fibroin. Exemplary formulations, compositions, articles, devices, and / or preparations include microneedles (e.g., microneedle devices described herein, e.g., microneedle patches), implantable devices (e.g., pumps, e.g., subcutaneous pumps), injectable formulations, depots, gels (e.g., hydrogels), implants, and particles (e.g., microparticles and / or nanoparticles). Thus, administration of the composition can be achieved by various methods including intradermal, intramuscular, transdermal, subcutaneous, or intravenous administration. Further, the formulations, compositions, articles, devices, and / or preparations can be formulated and / or administered to achieve controlled release and / or sustained release of at least one vaccine, antigen, and / or immunogen described herein (e.g., at least one vaccine, antigen, and / or immunogen derived from an influenza virus described herein).

[0116] In some embodiments, a vaccine (e.g., an influenza vaccine) is administered, for example, over a period of time, or at least for a period of 1, 5, 10, 15, 30, 45 minutes; over a period of time, or at least for a period of 1, 2, 3, 4, 5, 10, 24 hours; over a period of time, or at least for a period of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 days; over a period of time, or at least for a period of 1, 2, 3, 4, 5, 6, 7, 8 weeks; over a period of time, or at least for a period of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 months; over a period of time, or at least for a period of 1 year, 2 years, 3 years, 4 years, 5 years, or more, and is, for example, substantially sustained. In one embodiment, a vaccine (e.g., an influenza vaccine) is administered as a controlled release or sustained release formulation, dosage form, or device. In certain embodiments, a vaccine (e.g., an influenza vaccine) is formulated for continuous administration, e.g., intradermal, intramuscular, and / or intravenous continuous administration. In some embodiments, a composition or device for controlled or sustained release of a vaccine is selected from the following, namely, microneedles (e.g., microneedle devices, e.g., microneedle patches), implantable devices (e.g., pumps, e.g., subcutaneous pumps), injectable formulations, depots, gels (e.g., hydrogels), implants, or particles (e.g., microparticles and / or nanoparticles). In one embodiment, a vaccine (e.g., an influenza vaccine) is a silk-based controlled or extended release dosage form or formulation (e.g., the microneedles described herein). In one embodiment, a vaccine (e.g., an influenza vaccine) is administered via an implantable device, e.g., a pump (e.g., a subcutaneous pump), an implant, an implantable tip of a microneedle, or a depot.The administration method can be optimized such that the dose (e.g., standard dose) of the vaccine described in this specification (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccine) is administered and / or maintained in a subject for a predetermined period (e.g., over a certain period, or at least 1, 5, 10, 15, 30, 45 minutes; 1, 2, 3, 4, 5, 10, 24 hours; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 days; 1, 2, 3, 4, 5, 6, 7, 8 weeks; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 months; 1, 2, 3, 4, 5 years, or more). Substantially sustained or extended release of the vaccine (influenza vaccine) can be used over a period of time, days, weeks, months, or years for the prevention or treatment of viral infection (e.g., influenza virus infection).

[0117] In some embodiments, the formulations, compositions, articles, devices, and / or preparations of the present invention can be formulated and / or configured to provide controlled or sustained release of at least one vaccine, antigen, and / or immunogen (e.g., at least one vaccine, antigen, and / or immunogen derived from an influenza virus as described herein) in an amount (e.g., dose) and / or over a period sufficient to generate an immune response (e.g., cellular and / or humoral immune response) against a virus, e.g., an influenza virus, in a subject.

[0118] In some embodiments, the formulations, compositions, articles, devices, and / or preparations of the present invention can be formulated and / or configured for controlled or sustained release of at least one vaccine, antigen, and / or immunogen (e.g., at least one vaccine, antigen, and / or immunogen derived from an influenza virus as described herein) in an amount (e.g., dose) and / or over a period sufficient to provide broad-spectrum immunity in a subject.

[0119] Substantially continuous or sustained release administration or formulation of a vaccine (e.g., an influenza vaccine) can be used for the prevention or treatment of viral infections (e.g., influenza virus infections) for hours, days, weeks, months, or years.

[0120] In some embodiments, at least one vaccine, antigen, and / or immunogen described herein can be added to a silk fibroin solution, for example, before forming a silk fibroin micro-needle or micro-needle device described herein. In embodiments, the silk fibroin solution is mixed with a vaccine, antigen, and / or immunogen and then used, for example, in a process of filling and / or casting, drying and / or annealing, to produce a micro-needle having any of the desired material properties for the production of a transplantable micro-needle tip.

[0121] Without being bound by theory, the ratio of silk fibroin to vaccine, antigen, and / or immunogen at the transplantable tip of the micro-needle affects their release. In some embodiments, an increase in silk concentration at the transplantable tip aids in the delayed release and / or excellent retention of the antigen within the tip. Any concentration of silk may be used as long as it is printable and has sufficient mechanical strength to penetrate the skin.

[0122] In some embodiments, silk fibroin can be used at a concentration ranging from about 1% w / v to about 10% w / v (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10% w / v) in the production of micro-needles or their components as described herein.

[0123] Exemplary excipients Furthermore, the formulations, compositions, articles, devices, and / or preparations can be formulated for administration by intradermal, intramuscular, transdermal, subcutaneous, or intravenous routes using common excipients, diluents, or carriers. In some embodiments, the formulations, compositions, articles, devices, and / or preparations may be administered, for example, transdermally and can be formulated as controlled release or sustained release dosage forms. The formulations, compositions, articles, devices, and / or preparations described herein can be administered alone, in combination with each other, or used in combination with other known therapeutic agents.

[0124] Formulations suitable for use in the present invention are described in Remington´s Pharmaceutical Sciences (1985). Further, for an overview of drug administration methods, see Langer (1990) Science 249:1527-1533. The formulations, compositions, articles, devices, and / or preparations described herein can be made by methods known to those skilled in the art, for example, by mixing, dissolving, granulating, tabletting, triturating, emulsifying, encapsulating, entrapping, or lyophilization processes. The following methods and excipients are merely illustrative and are in no way limiting.

[0125] The silk fibroin formulations used in the production of the microneedles described herein may contain excipients. In embodiments, the inclusion of excipients is for the purpose of improving the stability of the incorporated vaccine, antigen, and / or immunogen; increasing the porosity and diffusibility of the silk matrix of the vaccine, antigen, and / or immunogen from the microneedle tip (e.g., for the microneedle, formulation, composition, article, device, preparation, and / or); and / or increasing the crystallinity / beta-sheet content of the silk matrix to reduce the solubility of the silk material.

[0126] Exemplary excipients include, but are not limited to, sugars or sugar alcohols (e.g., sucrose, trehalose, sorbitol, mannitol, or combinations thereof), divalent cations (e.g., Ca2+ , Mg 2+ , Mn 2+ , and Cu 2+ ) and / or a buffer solution. In some embodiments, the concentration of the excipient can be used to modify the porosity of the matrix. For example, sucrose is the most common excipient used for this purpose. The excipient may also be added to promote the self - organization of silk to form a beta - sheet secondary structure. Such excipients can generally achieve this effect by participating in hydrogen bonding or charge interactions with silk. Non - limiting examples of excipients that can be used to promote the self - organization of silk into a beta - sheet secondary structure include sodium glutamate (e.g., L - glutamic acid), lysine, sugar alcohols (e.g., sorbitol and / or glycerol), and solvents (e.g., DMSO, methanol, and / or ethanol).

[0127] In some embodiments, the sugar or sugar alcohol is sucrose present in an amount of less than 70% (w / v), less than 60% (w / v), less than 50% (w / v), less than 40% (w / v), less than 30% (w / v), less than 20% (w / v), less than 10% (w / v), less than 9% (w / v), less than 8% (w / v), less than 7% (w / v), less than 6% (w / v), or less than or equal to 5% (w / v), for example, immediately before drying.

[0128] In some embodiments, the sugar or sugar alcohol is sucrose present in an amount of about 1% (w / v) to about 10% (w / v), about 2% (w / v) to about 8% (w / v), about 2.2% (w / v) to about 6% (w / v), about 2.4% (w / v) to about 5.5% (w / v), about 2.5 to about 5%, or about 2.4% (w / v), about 2.5%, or about 5% (w / v), for example, immediately before drying.

[0129] In some embodiments, the sugar or sugar alcohol is trehalose present in an amount of, for example, about 1% (w / v) to about 10% (w / v), about 2% (w / v) to about 8% (w / v), about 2.2% (w / v) to about 6% (w / v), about 2.4% (w / v) to about 5.5% (w / v), about 2.5 to about 5%, or about 2.4% (w / v), about 2.5%, or about 5% (w / v), for example, immediately before drying.

[0130] In some embodiments, the sugar or sugar alcohol is sorbitol present in an amount of, for example, about 1% (w / v) to about 10% (w / v), about 2% (w / v) to about 8% (w / v), about 2.2% (w / v) to about 6% (w / v), about 2.4% (w / v) to about 5.5% (w / v), about 2.5 to about 5%, or about 2.4% (w / v), about 2.5%, or about 5% (w / v), for example, immediately before drying.

[0131] In some embodiments, the sugar or sugar alcohol is glycerol present in an amount of, for example, about 1% (w / v) to about 10% (w / v), about 2% (w / v) to about 8% (w / v), about 2.2% (w / v) to about 6% (w / v), about 2.4% (w / v) to about 5.5% (w / v), about 2.5 to about 5%, or about 2.4% (w / v), about 2.5%, or about 5% (w / v), for example, immediately before drying.

[0132] In some embodiments, the vaccine preparation further comprises a divalent cation. In some embodiments, the divalent cation is selected from the group consisting of Ca 2+ , Mg 2+ , Mn 2+ , and Cu 2+ . In some embodiments, the divalent cation is present in the preparation in an amount of, for example, 0.1 mM to 100 mM, for example, immediately before drying. In some embodiments, the divalent cation is present in the preparation in an amount of, for example, 10 -7 moles to 10 -4 moles per standard dose of the viral immunogen, for example, immediately before drying. In some embodiments, the divalent cation is present in the preparation in an amount of 10 -10 moles to 2×10 -3 moles, for example, immediately before drying.

[0133] In some embodiments, the vaccine preparation further comprises poly(lactic-co-glycolic acid) (PGLA).

[0134] In some embodiments, the viral vaccine preparation further comprises, for example, a buffer immediately prior to drying. In some embodiments, the buffer has a buffering capacity at pH 3 to pH 8, pH 4 to pH 7.5, or pH 5 to pH 7. In some embodiments, the buffer is selected from the group consisting of HEPES and CP buffer. In some embodiments, the buffer is present in the preparation in an amount of 0.1 mM to 100 mM, for example, immediately prior to drying. In some embodiments, the buffer is present in an amount of 10 -7 moles to 10 -4 moles per standard dose of the viral immunogen. In some embodiments, the buffer is present in an amount of 10 -10 moles to 2×10 -3 moles.

[0135] Furthermore, the vaccine can also be formulated as a depot, gel, or hydrogel preparation. Such long-acting formulations can be administered by implantation (e.g., subcutaneously or intramuscularly) or by intramuscular injection. Thus, for example, the vaccine can be formulated using a suitable polymer or hydrophobic material (e.g., as an emulsion in an acceptable oil) or an ion exchange resin, or as a poorly soluble derivative (e.g., a poorly soluble salt).

[0136] In one embodiment, the vaccine is administered via an implantable infusion device, such as a pump (e.g., a subcutaneous pump), an implant, or a depot. An implantable infusion device typically includes a housing containing a liquid reservoir that can be filled percutaneously by a subcutaneous needle piercing the infusion port septum. The dosing reservoir is generally connected via an internal flow path to a device outlet port for administering the liquid to a body site of the patient through a catheter. A typical infusion device also includes a fluid transfer mechanism, such as a controller and a pump or valve, for moving the liquid from the reservoir through the internal flow path to the outlet port of the device.

[0137] In some embodiments, the vaccine can be packaged and / or formulated as particles, for example, as microparticles and / or nanoparticles. Typically, the diameter of the nanoparticles is 10, 15, 20, 25, 30, 35, 45, 50, 75, 100, 150 or 200 nm or 200 to 1,000 nm, for example, 10, 15, 20, 25, 30, 35, 45, 50, 75, 100, 150, or 200, or 20 or 30 or 50 to 400 nm. The smaller the particle, the more rapidly it tends to be eliminated from the system. The therapeutic agent containing the vaccine may be encapsulated within the nanoparticles or may be linked to the nanoparticles, for example, covalently linked to the nanoparticles or, in other situations, adhered to the nanoparticles.

[0138] Lipid- or oil-based nanoparticles, such as liposomes and solid lipid nanoparticles, can be used to administer therapeutic agents (e.g., vaccines) as described herein. Solid lipid nanoparticles for the administration of therapeutic agents are described (see, e.g., Serpe et al. (2004) Eur. J. Pharm. Bioparm. 58:673-680, and Lu et al. (2006) Eur. J. Pharm. Sci. 28: 86-95). Polymer-based nanoparticles, such as PLGA-based nanoparticles, can be used to administer the agents described herein. These rely on a biodegradable backbone having a therapeutic agent (with or without a covalent bond to the polymer) inserted into the polymer matrix. PLGA is widely used in polymeric nanoparticles (see, e.g., Hu et al. (2009) J. Control. Release 134:55-61; Cheng et al. (2007) Biomaterials 28:869-876, and Chan et al. (2009) Biomaterials 30:1627-1634). PEGylated PLGA-based nanoparticles can also be used to administer therapeutic agents (see, e.g., Danhier et al. (2009) J. Control. Release 133:11-17, Gryparis et al (2007) Eur. J. Pharm. Biopharm. 67:1-8). Metal-based nanoparticles, such as gold-based nanoparticles, can also be used to administer therapeutic agents. Protein-based nanoparticles, such as albumin-based nanoparticles, can be used to administer the therapeutic agents described herein. In some embodiments, the therapeutic agent can bind to nanoparticles of human albumin.

[0139] A wide range of nanoparticles are known in the art. Exemplary approaches include those described in WO 2010 / 005726, WO 2010 / 005723, WO 2010 / 005721, WO 2010 / 121949, WO 2010 / 0075072, WO 2010 / 068866, WO 2010 / 005740, WO 2006 / 014626, US Patent No. 7,820,788, and US Patent No. 7,780,984, the entire contents of which are incorporated herein by reference.

[0140] Dosage For example, when administered by the microneedles of the present invention, any dosage (e.g., standard dosage and / or divided dosage) of a vaccine, antigen, and / or immunogen that can induce an immune response (e.g., immunogenicity and / or broad-spectrum immunity) in a subject can be used according to the methods described herein. In some embodiments, the dosage of a vaccine, antigen, and / or immunogen (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccine), e.g., a standard dosage (e.g., human dosage), is from about 0.1 μg to about 65 μg (e.g., from about 0.1 μg to about 10 μg, from about 0.1 μg to about 1 μg, from about 0.5 μg to about 5 μg, from about 5 μg to about 10 μg, from about 10 μg to about 20 μg, from about 20 μg to about 30 μg, from about 30 μg to about 40 μg, from about 40 μg to about 50 μg, from about 50 μg to about 65 μg, e.g., about 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, or 65 μg). In some embodiments, the dosage of a vaccine described herein (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccine), e.g., a standard human dosage, is from about 1 μg to about 30 μg per strain, e.g., from about 5 μg to about 30 μg per strain (e.g., about 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 μg per strain).In some embodiments, the dose of the vaccine described herein (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccine), e.g., the divided dose, is less than or equal to 1 / X of the total dose (e.g., the standard dose), where X is any number, e.g., X is 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, 60, 70, 80, 90, or 100 or more. In the art, it is known that there are preclinical examples of dose sparing when administering influenza vaccine into the intradermal space (e.g., Fluzone ID), and this dose is about 9 μg per strain. Thus, in some embodiments, the total dose of influenza vaccine (e.g., Fluzone ID) that can be administered by the microneedles of the present invention can be between about 5 μg and 13 μg (e.g., about 5 μg, about 6 μg, about 7 μg, about 8 μg, about 9 μg, about 10 μg, about 11 μg, about 12 μg, or about 13 μg).

[0141] While not wishing to be bound by theory, the total dose (e.g., standard dose) of vaccine, antigen, and / or immunogen to be administered by the microneedles described herein may be divided among a plurality of microneedles (e.g., within a patch) such that the microneedle tip contains less than about 1% of the total dose (e.g., within an array containing about 121 microneedles), or at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, or 25% or more of the total dose. In some embodiments, the implantable microneedle tip described herein can contain from about 0.1 μg to about 65 μg (e.g., about 0.1 μg, about 0.2 μg, about 0.3 μg, about 0.4 μg, about 0.5 μg, about 0.6 μg, about 0.7 μg, about 0.8 μg, about 0.9 μg, about 1 μg, from about 1 μg to about 10 μg, from about 10 μg to about 20 μg, from about 20 μg to about 30 μg, from about 30 μg to about 40 μg, from about 40 μg to about 50 μg, from about 50 μg to about 65 μg) of the vaccine, antigen, and / or immunogen described herein.

[0142] In some embodiments, the vaccine dose loaded into the microneedle patch may be manipulated via the concentration of antigen in the formulation solution forming the tip of the needle, the volume of solution dispensed to the tip of each needle, and the total number of needles (the former two being more convenient means of changing the dose). The dose released to the skin is related to the deployment efficiency (the portion of the needle tip remaining in the skin after removal of the patch) as well as the release profile of the tip over time and the residence time of the tip within the skin. Due to the continuous desquamation of the skin from the epidermis, the depth of placement within the skin is related to the length of the residence time. As a result, it is desirable to maximize the penetration depth of the needle tip (up to the limit defined by the depth of pain receptors within the skin, e.g., a depth between about 100 μm and about 600 μm), and to have the antigen spatially concentrated towards the tip of the needle.

[0143] The formulations, compositions, articles, devices, and / or preparations described herein that include implantable sustained-release tip formulations are designed to release vaccine antigens, for example, over a period of time during which the tip is retained in the dermis and to maintain antigen stability during this period (e.g., at least about 1 to 2 weeks). In some embodiments, for example, about 95 to 100% of the total dose incorporated into the formulations, compositions, articles, devices, preparations, and / or microneedles described herein may be expected to be available for administration, for example, into a subject, for example, into a subject's tissue, such as skin, mucosa, organ tissue, oral cavity, tissue, or cell membrane. Without being bound by theory, successful placement of microneedles into the skin is at least about 50% of the array and may be 100% of the array (e.g., upon application, at least about 50%, 60%, 70%, 80%, 90% or more (e.g., 100%) of the total number of microneedles in the array are successfully placed, for example, in the skin, for controlled or sustained release of the vaccine antigen). In some embodiments, some of the antigen may not be released from the silk tip during the placement period.

[0144] Use The present invention also provides a method for administering a vaccine, antigen, and / or immunogen (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccine) across a biological barrier (e.g., skin). Such methods can include providing a formulation, composition, article, device, formulation, and / or microneedle described herein. For example, such methods can include providing at least one microneedle or at least one microneedle device described herein, where the microneedle or microneedle device can include a silk fibroin-based implantable tip having at least one vaccine, antigen, and / or immunogen (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccine), inserting the microneedle or microneedle device into a biological barrier (e.g., skin), and enabling the vaccine, antigen, and / or immunogen to be released from the implantable tip over at least about 4 days (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14 days or more, e.g., about 4 to about 14 days, e.g., about 1 to 2 weeks, about 1 to 3 weeks, or about 1 to 4 weeks). In some embodiments, the vaccine, antigen, and / or immunogen is released into the biological barrier by degradation and / or dissolution of the implantable microneedle tip. In some embodiments, the microneedle or microneedle device is configured to administer the vaccine, antigen, and / or immunogen in an amount and / or for a period of time that results in broad-spectrum immunity in the subject, e.g., immunity to one or more viral antigens not present in the implantable sustained-release tip, e.g., immunity to a drift strain not present in the implantable sustained-release tip.

[0145] The present invention also provides a method of providing broad-spectrum immunity against a virus, such as an influenza virus, in a subject, the method comprising administering a vaccine (such as an influenza vaccine and / or a coronavirus vaccine, such as an mRNA-based vaccine) in an amount (such as a dose) and / or for a period sufficient to elicit an immune response (such as a cellular and / or humoral immune response) against the virus that provides broad-spectrum immunity against the virus in the subject, such as against drifted strains of the virus. In some embodiments, the vaccine is administered in a composition for controlled or sustained release of the vaccine (such as controlled or sustained release of one or more viral antigens described herein). In some embodiments, the vaccine is administered by a device for controlled or sustained release of the vaccine (such as for controlled or sustained release of one or more viral antigens described herein). The vaccine can be administered to the subject in a tissue or cavity of the subject selected from, for example, the skin, mucosa, organ tissue, muscle tissue, or oral cavity.

[0146] In some embodiments, the methods described herein include administering, in an amount (e.g., dosage) and / or for a period sufficient to cause one or more of the following: (i) eliciting broad-spectrum immunity, e.g., exposing the subject to one or more antigens in the vaccine in an amount and / or for a period that elicits an immune response (e.g., a cellular and / or humoral immune response) against drifted strains of a virus in the subject; or (ii) the level of one or more antigens in the subject that is substantially stable, e.g., about 20%, 15%, 10%, 5%, or 1%, relative to the amount necessary to elicit an immune response (e.g., a cellular and / or humoral immune response) against one or more antigens, e.g., the minimum amount. In some embodiments, the composition or device for controlled or sustained release of the vaccine is selected from microneedles (e.g., microneedle devices, e.g., microneedle patches as described herein), implantable devices (e.g., pumps, e.g., subcutaneous pumps), injectable formulations, depots, gels (e.g., hydrogels), implants, or particles (e.g., microparticles and / or nanoparticles).

[0147] In some embodiments, the vaccine is administered to a subject, e.g., by a composition or device for controlled or sustained release of the vaccine, e.g., released, in order to maintain a vaccine dose (e.g., antigen concentration) for a sufficient period of time (e.g., this period is about 1 to 21 days, e.g., about 5 to about 10 days or about 5 to about 7 days, e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 21 days) to provide a broad-spectrum immunity, e.g., an immune response (e.g., a cellular immune response and / or a humoral immune response) against a drifted strain of a virus in the subject. The composition or device for controlled or sustained release of the vaccine can maintain the release and / or level of the antigen in the subject over a sustained period. In some embodiments, the composition or device for controlled or sustained release of the vaccine maintains the continuous or discontinuous release of the immunogen to the subject over the duration. The vaccine may be administered over a period including at least about 1 week, e.g., about 1 to 2 weeks, about 1 to 3 weeks, or about 1 to 4 weeks, e.g., released by a composition or device for controlled or sustained release. In some embodiments, the vaccine is administered over a period including at least about 4 days (e.g., about 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 days, or more, e.g., between about 4 days to about 2 weeks, between about 4 days to about 1 week), e.g., administered, e.g., released, by a composition or device for controlled or sustained release.

[0148] The vaccine may be administered in a dosage that contains between about 0.1 μg to about 65 μg per strain, for example, between 0.2 μg to about 50 μg per strain (e.g., about 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, or 65 μg per strain). In some embodiments, for example, at least about 1% of the dosage of the vaccine (e.g., at least about 0.5% to about 10%, at least about 5% to about 15%, at least about 10% to about 20%), which is released to the subject, for example, by a composition or device for controlled or sustained release of the vaccine, is maintained over a period of at least about 4 days (e.g., about 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14 days, or more, e.g., about 4 days to about 2 weeks, about 4 days to about 1 week).

[0149] In some embodiments, the vaccine is administered in a plurality of divided doses of the total dosage (e.g., standard dosage) over a period of time, for example, released by a composition or device for controlled or sustained release, such that an immune response and / or broad-spectrum immunity is achieved, where the amount of the vaccine administered in each of these divided doses is 1 / X or less, where X is any number, for example, X is 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, 60, 70, 80, 90, or 100 or more of the total dosage (e.g., standard dosage) of the vaccine.

[0150] In some embodiments, the vaccine is administered, for example, to the skin of a subject, e.g., by a composition or device for controlled or sustained release of the vaccine, e.g., released, in a plurality of doses corresponding to a percentage of the total dose (e.g., a percentage of the standard dose) over a period of time, e.g., such that broad-spectrum immunity is achieved, and the amount of vaccine administered in each of the plurality of doses is about X%, where X is any number, e.g., X is 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, 40, 50, 60, 70, 80, 90, 100, 125, 150, 175, 200, 300, 400, or 500 or more of the total dose of the vaccine (e.g., the standard dose).

[0151] The vaccine can be administered according to any method described herein, e.g., such that an immune response, such as a cellular and / or humoral immune response against a drifted strain, is achieved, e.g., such that broad-spectrum immunity is achieved.

[0152] While not wishing to be bound by theory, a subject exposed to and / or infected with a first influenza virus can mount an immune response (e.g., a cellular and / or humoral immune response), resulting in the production of antibodies against that first influenza virus. As antigenic changes (e.g., mutations) accumulate over time in the first influenza virus, the subject's antibodies produced against the first influenza virus may become unable to recognize drifted viruses (e.g., antigenically distinct strains). The methods, dosing regimens, microneedles, and microneedle devices described herein can confer broad-spectrum immunity to subjects exposed to, infected with, and / or at risk of infection with influenza virus. Further, the methods, dosing regimens, microneedles, and microneedle devices described herein can confer improved immunogenicity and / or broad-spectrum immunity to a subject as compared to, for example, a burst-release administration of a conventional vaccine. For example, the improved immunogenicity and / or broad-spectrum immunity detectable in a subject can be greater (e.g., 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 11-fold, 12-fold, 13-fold, 14-fold, or 15-fold or more) as compared to a burst-release administration of a conventional vaccine, e.g., a single-dose or bolus administration of a vaccine.

[0153] In some embodiments, the implantable sustained-release tip or vaccine comprises a first influenza strain, and administration of a dose of the first influenza strain (e.g., the first influenza A, B, C, and / or D type strains described herein) to a subject results in the development of broad-spectrum immunity against a second influenza strain (e.g., the drifted influenza A, B, C, and / or D type strains described herein) that is not present in either the implantable sustained-release tip or the vaccine.

[0154] In some embodiments, the subject (e.g., a human subject) is a pediatric subject, an adult subject, or an elderly subject. The subject may have been exposed to, infected with, and / or at risk of infection from an influenza virus (e.g., a specific strain of influenza virus). Such risks may be due to the subject's health status or age, and / or travel to an area where a particular influenza virus strain is prevalent.

[0155] In some embodiments, the present invention provides a method for providing controlled or sustained release of a vaccine in a subject. Controlled or sustained release of the vaccine can achieve improved immunogenicity and / or broad-spectrum immunity compared to bolus administration of conventional vaccines. Without wishing to be bound by theory, the method of administering the vaccines described herein, and / or the controlled or sustained release rate, e.g., by the compositions and / or microneedles described herein, that mimics the natural exposure pattern of a subject (e.g., a human subject) to a virus can provide enhanced and / or broad-spectrum immunity in the subject compared to conventional single-dose vaccine administration regimens.

[0156] In some embodiments, a desired amount of at least one vaccine, antigen, and / or immunogen (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccine) can be released in a sustained manner from the microneedles (e.g., implantable microneedle tip) described herein over a pre-defined period. In some embodiments, at least about 5% of the vaccine, antigen, and / or immunogen (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccine), e.g., at least about 10%, at least about 15%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, or at least about 97%, about 98%, or about 99%, or 100% can be released from the microneedles (e.g., tip of the implantable microneedle) over a pre-defined period. In such embodiments, a desired amount (e.g., a dose such as a standard dose of a vaccine) of the vaccine, antigen, and / or immunogen (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccine) can be released from the microneedles over seconds, minutes, hours, months, and / or years. In some embodiments, a desired amount (e.g., a dose such as a standard dose of a vaccine) of the vaccine, antigen, and / or immunogen (e.g., influenza vaccine and / or coronavirus vaccine, e.g., mRNA-based vaccine) can be released from the microneedles, e.g., within 5 seconds, within 10 seconds, within 30 seconds, within 1 minute, within 2 minutes, within 3 minutes, within 4 minutes, within 5 minutes, or more, upon insertion into a biological barrier.In some embodiments, a desired amount (e.g., a dosage such as a standard dosage of a vaccine) of a vaccine, antigen, and / or immunogen (e.g., an influenza vaccine and / or a coronavirus vaccine, e.g., an mRNA-based vaccine) can be released from the microneedles over a period of at least about 1 hour, at least about 2 hours, at least about 3 hours, at least about 6 hours, at least about 12 hours, at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 1 week, at least about 2 weeks, at least about 1 month, at least about 2 months, at least about 3 months, at least about 6 months or more. In some embodiments, a desired amount (e.g., a dosage such as a standard dosage of a vaccine) of a vaccine, antigen, and / or immunogen (e.g., an influenza vaccine and / or a coronavirus vaccine, e.g., an mRNA-based vaccine) can be released from the microneedles over a period of about 1 year or more.

[0157] In some embodiments, the present invention provides a method of enhancing an immune response against a virus in a subject. In some embodiments, the presence of a cell-mediated immunological response can be determined by any method recognized in the art, e.g., a proliferation assay (CD4+ T cells), a CTL (cytotoxic T lymphocyte) assay (see Burke, supra; Tigges, supra), or immunohistochemistry using tissue sections of the subject to determine the presence of activated cells such as monocytes and macrophages after immunogen administration. One of ordinary skill in the art can readily determine the presence of a humoral-mediated immunological response in a subject by any well-established method. For example, the level of antibodies produced in a biological sample such as blood can be measured by Western blot, ELISA, or other methods known for antibody detection. In some embodiments, an increase in the hemagglutination inhibition (HAI) antibody titer can be detected in the blood of the subject over the period of a complete influenza season after immunization.

[0158] In some embodiments, an immune response and / or broad spectrum immunity is a cellular and / or humoral immune response comprising: (i) an increase in hemagglutination inhibition (HAI) antibody titer detectable in the subject's blood, e.g., detectable for at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, and / or 30 or more weeks after immunization; (ii) an increase in anti-influenza IgG antibody titer detectable in the subject's blood, e.g., detectable for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, and / or 12 or more months after immunization; and / or (iii) a level of antibody-secreting plasma cells (ASCs) against a virus, e.g., influenza virus, detectable in the subject's bone marrow, e.g., detectable for at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, and / or 34 or more weeks after immunization. In some embodiments, the increased HAI antibody titer is against drift influenza A, B, C, and / or D strains. In some embodiments, the increased anti-influenza IgG titer is against drift influenza A, B, C, and / or D strains. In some embodiments, the immune response is a cellular immune response comprising, e.g., an increase in the level of IFNγ-secreting cells in the subject's blood by, e.g., at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, and / or 12 or more weeks after immunization, by, e.g., the microneedles described herein.

[0159] In some embodiments, the increased HAI antibody titer, increased anti-influenza IgG titer, level of antibody-secreting plasma cells (ASCs) against a virus, and / or level of IFNγ-secreting cells detectable in the subject is greater (e.g., 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 11-fold, 12-fold, 13-fold, 14-fold, or 15-fold or more) compared to a single dose administration or bolus administration of the vaccine.

[0160] In some embodiments, broad-spectrum immunity can be characterized by measuring the seroconversion rate in a subject. For example, broad-spectrum immunity can include, for example, a seroconversion rate of greater than about 20% (e.g., 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% or more, e.g., 100%) at about 6 months after immunization, based on, for example, an elevated HAI antibody titer detectable in the subject's blood. Such levels of seroconversion associated with broad-spectrum immunity conferred by the methods, dosing regimens, microneedles, and microneedle devices described herein can be greater (e.g., 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 11-fold, 12-fold, 13-fold, 14-fold, or 15-fold or more) compared to the levels of seroconversion obtained by conventional burst-release dosing of a vaccine, e.g., a single-dose administration or a bolus administration of a vaccine.

[0161] Combination therapy The microneedles and microneedle devices (e.g., microneedle patches) described herein can be fabricated by precisely filling each of the individual microneedle tips in order to enable different patterns of vaccine administration, dosing regimens, and co - administration of vaccines and additional therapeutic agents. The immunization, vaccine administration, and dosing methods described herein can include co - administration of a vaccine and an additional therapeutic agent. In some embodiments, the additional therapeutic agent may be formulated at the same tip as the vaccine. In some embodiments, the additional therapeutic agent may be formulated with the vaccine. For example, an adjuvant for enhancing the immune response to a co - administered antigen may be administered at the same microneedle tip and / or with the vaccine. Without wishing to be bound by theory, such combination therapies may include adjuvants for driving a more potent cellular immune response and / or mucosal response. Further, additional influenza antigens can be administered for heterologous "prime / boost - like" immunization, for example, a primary immunization with the HA antigen of various influenza strains and a boost with a different antigen (e.g., a drifted strain, hemagglutinin stem, m2e protein, or NA) (e.g., provided via controlled or sustained release, or via a kinetic pattern different from the "prime").

[0162] Due to formulation compatibility, there may be limitations as to whether two given therapeutic agents can be co - formulated for dispensing at the same needle tip. If co - formulation is not possible, the manufacturing process may be adapted to dispense a first formulation into one part of the needle array and then a second formulation into a different part of the needle array. The different formulations can also be subject to different processing after filling. For example, if the first formulation is for controlled or sustained release and silk is rendered less soluble by hydro - annealing, while the second formulation is for burst release without annealing, the second formulation can be dispensed after the annealing step. Since the manufacturing method is flexible, other process sequences are possible.

[0163] In some embodiments, the present invention also provides a method of combination therapy that fabricates the micro-needles or micro-needle devices of the present invention to administer at least one additional therapeutic agent. When administered to a subject, various forms of therapeutic agents can be used that can be released from the micro-needles described herein into adjacent tissue or fluid. In some embodiments, additional therapeutic agents can be included within the base layer and / or within the implantable tip.

[0164] Examples of additional therapeutic agents that can be used in accordance with the methods of the present invention and that can, for example, be incorporated during fabrication into the micro-needles of the present invention include steroids and steroid esters (e.g., estrogen, progesterone, testosterone, androsterone, cholesterol, norethindrone, digoxigenin, cholic acid, deoxycholic acid, and chenodeoxycholic acid), boron-containing compounds (e.g., carborane), chemotherapeutic nucleotides, drugs (e.g., antibiotics, antiviral agents, antifungal agents), enediynes (e.g., calicheamicin, esperamicin, dynemicin, neocarzinostatin chromophore, and kedarcidin chromophore), heavy metal complexes (e.g., cisplatin), hormone antagonists (e.g., tamoxifen), non-specific (non-antibody) proteins (e.g., glyco-oligomers), oligonucleotides (e.g., mRNA sequences or antisense oligonucleotides that bind to target nucleic acid sequences), peptides, proteins, antibodies, photodynamic agents (e.g., rhodamine 123), radionuclides (e.g., I-131, Re-186, Re-188, Y-90, Bi-212, At-211, Sr-89, Ho-166, Sm-153, Cu-67, and Cu-64), toxins (e.g., ricin), and transcription-based pharmaceuticals.

[0165] Exemplary kits In certain embodiments, the present invention relates to a package or kit comprising a microneedle as described herein (e.g., a microneedle comprising a vaccine, antigen, and / or immunogen as described herein, e.g., an influenza virus). In some embodiments, the present invention relates to a package or kit comprising a vaccine as described herein (e.g., a vaccine, antigen, and / or immunogen as described herein, e.g., an influenza virus). In some embodiments, the kit can further comprise an additional therapeutic agent for combination therapy with the microneedle. In some embodiments, the kit can further comprise a disinfectant (e.g., an alcohol swab). In some embodiments, such packages and kits as described herein can be used for the purpose of vaccination, e.g., to achieve broad-spectrum immunity in a subject as described herein.

Claims

1. A patch and applicator system comprising: A drug patch including a flexible sheet material; An applicator for holding and applying the drug patch to a patient's skin, the applicator comprising: An outer body portion that is substantially cylindrical and has a hollow interior; A piston portion slidably connected to the hollow interior of the outer body portion; A compressible member disposed within the outer body and configured to apply a downward pressure to the piston; An actuator disposed below the piston and slidably engaged with the hollow interior, the actuator extending from the bottom of the outer body such that an upward pressure on the actuator pushes the piston upward and compresses the compressible member; An applicator comprising; Wherein the hollow interiors of the piston and the outer body portion are slidably connected to a protrusion and a cam path, and the upward pressure on the piston portion compresses the compressible member and moves the piston upward within the hollow interior of the outer body portion until the protrusion reaches the top of the cam path and engages the downward portion of the cam path, whereby the cam path forms a continuous loop such that the piston is released into the downward portion of the continuous loop and the piston is released and pushed downward; The drug patch is held in a ring holder at the bottom of the actuator, pushed through the ring holder, and pressed against a target that abuts the bottom surface of the ring holder.

2. The system of claim 1, wherein the compressible member includes a spring.

3. The system of claim 1 or 2, wherein the ring holder is removable from the actuator.

4. The system of claim 1 or 2, wherein the ring holder is integrally connected to the actuator.

5. The system according to any one of claims 1 to 4, wherein the hollow interiors of the piston and the outer body portion are slidably connected to at least one additional protrusion and at least one additional cam path.

6. The system according to any one of claims 1 to 4, wherein the protrusion and the cam path include a protrusion extending from a sidewall of the piston and a cam path formed along an inner wall of the outer body portion.

7. The system of claim 5, wherein each protrusion and cam path includes a protrusion extending from a side wall of the piston and a cam path formed along an inner wall of the outer body portion. **Claim 8** The system according to any one of claims 1 to 7, wherein the continuous loop of the cam path includes a loop having a substantially parallelogram shape. **Claim 9** The system according to any one of claims 1 to 8, wherein the actuator is slidably engaged with the hollow interior along a connection between at least one protrusion of the actuator and at least one cam path of the hollow interior. **Claim 10** The system of claim 9, wherein the at least one cam path of the actuator and the at least one cam path of the hollow interior are configured to allow substantially linear sliding of the actuator relative to the hollow interior without rotation of the actuator. **Claim 11** The system according to any one of claims 1 to 10, wherein the piston is rotatably movable within the hollow interior. **Claim 12** The system according to any one of claims 1 to 11, wherein the ring holder has a solid rim portion and an open central portion. **Claim 13** The system of claim 12, wherein the solid rim is substantially circular. **Claim 14** The system according to claim 12 or 13, wherein the drug patch and the ring holder are dimensioned such that the patch is larger than the open central portion in at least one dimension so that the drug patch can rest on top of the ring holder without passing through the central portion. **Claim 15** The system of claim 14, wherein the drug patch has a flexibility such that downward pressure on the drug patch caused by release of the piston causes the drug patch to bend sufficiently to allow the drug patch to pass through the central portion of the ring holder. **Claim 16** The system according to any one of claims 1 to 15, wherein the drug patch is a flat sheet having a circular, triangular, square, polygonal, or elliptical shape. **Claim 17** The system according to any one of claims 1 to 16, wherein the drug patch has one or more cuts along its perimeter to facilitate bending of the patch near its perimeter. **Claim 18** The system according to any one of claims 1 to 17, wherein the patch includes a microneedle patch. **Claim 19** The system according to claim 18, wherein the patch is a fibroin-based microneedle patch. **Claim 20** The system according to claim 18 or 19, wherein the microneedle patch includes biodegradable microneedles.

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