Improved delivery of large agents

Abstract

To provide methods, compositions, and devices for enhancing transdermal delivery and / or bioavailability of large agents.SOLUTION: A method comprises a step of applying an emulsion composition comprising a large agent having a molecular weight of 100,000 Da or greater to a site in combination with microneedle skin conditioning (MSC) of the site with a microneedle array having a microneedle density within a range of about 2 to about 50 microneedles / cm2.SELECTED DRAWING: None

Description

[Technical field]

[0001] (CROSS REFERENCE TO RELATED APPLICATIONS) This application claims priority to U.S. Provisional Application Nos. 62 / 774,677 (filed December 3, 2018), 62 / 789,407 (filed January 7, 2019), and 62 / 808,274 (filed February 20, 2019), the entire contents of all of which are incorporated herein by reference. [Background technology]

[0002] Significant resources have been invested in the development of effective transdermal delivery technologies. Those skilled in the art are well aware of the challenges associated with achieving effective transdermal delivery, especially for large drugs. As molecular size increases, transdermal penetration decreases until it becomes negligible or nonexistent. Summary of the Invention

[0003] Transdermal administration has been the subject of research to provide an alternative route of drug administration that does not have the undesirable consequences associated with injection and oral delivery in general.For example, needles often cause local pain, bleeding and subcutaneous bleeding, and may expose patients to infectious diseases; oral administration may suffer from low bioavailability of medicines due to the extremely acidic environment of the patient's stomach.In some embodiments, transdermal delivery has a more uniform, regular and / or consistent pharmacokinetic profile compared to other routes of administration.

[0004] Although it has many advantages, transdermal drug delivery poses many logistical problems. Only a limited number of drugs have been shown to be administrable by this route. It has been difficult to deliver active agents transdermally, including but not limited to hydrophilic molecules, large molecular structures (e.g., greater than several hundred daltons), gene therapy, vaccines, etc. Prausnitz, MR & Langer, R. "Transdermal drug delivery," Nat Biotechnol. 26(11): 1261-1268 (2008).

[0005] The present disclosure provides an improved technique for transdermal delivery of target drugs.In some embodiments, the present disclosure teaches that the combination of a specific microneedling technique and topical application of a composition containing a large drug can enhance and / or otherwise improve the delivery of topical drugs.In some embodiments, such compositions can be or include emulsions (e.g., nanoemulsions), such as those that contain large drugs.In some embodiments, such compositions can or do not include emulsions, and can be formulated as emulsions and / or, for example, liquids, gels, creams, or other formulations suitable for topical application.

[0006] Recent techniques have been developed that achieve various advantages by combining emulsion and microneedling techniques for transdermal delivery of target drugs (see, e.g., International Application No. PCT / US17 / 53333, incorporated herein by reference); in some embodiments, these techniques have been shown to achieve particularly surprising improvements for the transdermal delivery of large molecular structures.

[0007] The present disclosure discloses that even greater benefits are achieved when using a microneedling approach having a particular microneedle density and / or a particular microneedle puncture size. Surprisingly, the present disclosure teaches that particularly desirable results are achieved with a microneedling approach having a relatively low density and / or a relatively small puncture size.

[0008] Various microneedling techniques have been developed that may be useful for administering a particular target drug. Microneedling may avoid certain drawbacks (e.g., pain and / or degree of bleeding) that are often associated with the use of larger needles (e.g., standard injection techniques). Microneedling techniques may utilize one or more (e.g., arrays) of hollow or solid microneedles. Target drugs may be placed in or on (i.e., on the surface of) the microneedles (e.g., when the microneedles are hollow and / or when the drug is incorporated into the microneedle material) and / or may be applied to a skin site before, during or after microneedling of the site. Drugs in or on the microneedles may be released after application to the site, for example, by diffusion or expulsion from the microneedles, or by destruction and / or disintegration of the microneedle material.

[0009] In some embodiments, the present disclosure provides a strategy in which microneedling is used to "condition" the skin (especially to precondition the skin between administration of large drugs), such as microneedle skin conditioning (MSC) where a transdermal product has been applied, is being applied, or will be applied. The present disclosure provides the finding that such microneedle conditioning (e.g., microneedle conditioning with relatively low density and / or relatively small puncture size) described herein surprisingly provides a significant advantage in improving the transdermal delivery of large drugs (e.g., molecular weight of about 100 KDa or more) compared to microneedle conditioning approaches with higher density and / or larger puncture size. In some embodiments, the present disclosure teaches that microneedling can provide a surprising improvement in the delivery of large drugs. In some embodiments, the present disclosure demonstrates that microneedling treatment can be particularly advantageous for the delivery of large drugs in emulsion compositions (e.g., nanoemulsions).

[0010] Studies analyzing transdermal delivery of small molecules (especially short hydrophilic peptides with molecular weights in the range of 400-1000 Da) have found that "skin penetration of peptides depends on their molecular weight and decreases as the molecular weight increases," and therefore previous reports have found that microneedle conditioning strategies are likely to be useful only for drugs with small molecular weights. Zhang, S., et al., "Enhanced delivery of hydrophilic peptides in vitro by transdermal microneedle pretreatment." Acta Pharmaceutica Sinica B. 4(1):100-104 (2014). Furthermore, studies evaluating the effect of microneedle density, microneedle length, and / or microneedle puncture size on such small molecule delivery have demonstrated that microneedle density does not affect small molecule delivery and / or bioavailability, and that relatively longer microneedle lengths and larger microneedle puncture sizes improve small molecule delivery and / or bioavailability. An early report by Yan (Yan, G., et al., "Evaluation needle length and density of microneedle arrays in the pretreatment of skin for transdermal drug delivery", International Journal of Pharmaceutics, 391: 7-12, 2010) stated that "when microneedles with a sufficiently long needle length (>600 μm) were used, a lower needle density (<2000 needles / cm) was achieved." 2Although Yan stated that microneedle arrays with a diameter of 400 needles / cm were "more effective at enhancing drug flow," later studies identified flaws in the approach used by Yan, including that the assay used by Yan was "prone to artifacts" and could result in biological changes in the levels of skin tension and / or hydration (see Donnelly, RF, et al., Optical coherence tomography is a valuable tool in the study of the effects of microneedle geometry on skin penetration characteristics and in-skin dissolution, Journal of Controlled Release, 147: 333-341, 2010). Furthermore, the lowest microneedle density studied by Yan (400 needles / cm) 2 ) was relatively high. Furthermore, Yan's own data shows that needle density does not have a significant effect on small molecule delivery (i.e., "drug flux" through the skin) for needle lengths less than 1100 μm.

[0011] The Donnelly study mentioned above reported that microneedle (MN) penetration depth (rather than density or other factors) is the most important factor in determining efficient drug penetration. Donnelly also demonstrated that "changes in MN spacing do not affect the depth of penetration achieved."

[0012] Thus, prior to the present disclosure, the art has shown that changes in microneedle density are not expected to affect the delivery of drugs across the skin, at least at the density levels studied in the literature. In the first principle, one might naturally expect that if microneedling improves delivery, a relatively higher microneedle density (and / or a relatively larger microneedle puncture size) might be expected to be more effective. As the factors of microneedle density and microneedle diameter increase, it is expected that the total surface area of ​​the skin punctured increases, thereby allowing more active ingredients to permeate the skin transdermally. However, the present disclosure shows the surprising discovery that, at least for one and / or more large drugs in an emulsion (e.g., a nanoemulsion), a relatively lower microneedle density and / or a relatively smaller microneedle puncture size achieves better results. That is, it has been surprisingly found that the bioavailability of large drugs in an emulsion applied to the skin increases as the total surface area of ​​the skin punctured by the microneedles decreases. In some embodiments, the bioavailability of large drugs in an emulsion applied to the skin increases from about 2 to about 50 microneedles / cm. 2 Microneedle densities in the range of about 100 to about 60,000 μm can significantly improve transdermal delivery and / or bioavailability, even when compared to microneedling with relatively higher microneedle densities. Further, the present disclosure surprisingly provides a method for the preparation of a microneedle having a density in the range of about 100 to about 60,000 μm. 2 The present disclosure further discloses that microneedle conditioning of the skin using microneedle puncture sizes in the range of 0.1 to 1.0 μm can achieve significant transdermal delivery and / or bioavailability. The present disclosure also discloses that smaller microneedle puncture sizes (e.g., about 100 to about 30,000 μm) can achieve significant transdermal delivery and / or bioavailability. 2It is disclosed that microneedle conditioning of the skin using a microneedle having a diameter of about 100 to about 30,000 μm (range of microneedles) can significantly improve transdermal delivery and / or bioavailability, even when compared to microneedling using a relatively larger microneedle puncture size. In some embodiments, the microneedle puncture size is about 100 to about 30,000 μm. 2 / Microneedle range.

[0013] Prior to the present disclosure, one of skill in the art would have understood from the literature that microneedle conditioning of the skin with any particular density of microneedles would not be expected to improve transdermal delivery of even small molecules, let alone large drugs. The present disclosure surprisingly provides a method for the preparation of a drug with a density of microneedles between about 2 and about 50 microneedles / cm. 2 The present disclosure surprisingly discloses that microneedle conditioning of the skin with microneedle densities in the range of about 100 to about 30,000 μm can significantly enhance the transdermal delivery of drugs such as botulinum toxin, which has a molecular weight of about 150,000 Da. Standard antibodies also have a similar molecular weight. Moreover, the present disclosure surprisingly discloses that microneedle conditioning of the skin with microneedle densities in the range of about 100 to about 30,000 μm can significantly enhance the transdermal delivery of drugs such as botulinum toxin, which has a molecular weight of about 150,000 Da. Standard antibodies also have a similar molecular weight. 2 We disclose that microneedle conditioning of the skin using microneedle puncture sizes in the range of 100 / microneedles can significantly enhance the transdermal delivery of such large drugs.

[0014] Those skilled in the art who read this disclosure will logically understand that, in addition to reducing the total surface area punctured by reducing the density of microneedles and / or reducing the puncture size of the microneedles, the same effect can be achieved by minimizing (e.g., reducing) the number of impressions made by or with the microneedle array on the skin of the treatment area. Consistent with this understanding, and in light of the surprising observation above that the reduction in the total surface area punctured (e.g., may be achieved at least in part by the puncture size of the smaller microneedles, which may have additional concomitant advantages), the present disclosure further shows that relatively smaller impressions of the microneedle array may result in higher bioavailability than relatively more impressions. In some embodiments, the impact of the microneedle array is about 1 impression / cm. 2 ~5 pressure / cm 2 Microneedle impacts in the range of 100 nm to 150 nm may achieve significant transdermal delivery and / or bioavailability.

[0015] This disclosure provides a force of approximately 1 force / cm 2 ~Approximately 4 pressure / cm 2 This disclosure shows that microneedle conditioning of the skin using microneedle pressures in the range of about 1 pressure to about 20 pressures at the treatment site can significantly improve transdermal delivery and / or bioavailability, even when compared to microneedling with a relatively higher number of microneedle pressures. In some embodiments, microneedle pressures in the range of about 1 pressure to about 20 pressures at the treatment site can achieve significant transdermal delivery and / or bioavailability. Furthermore, this disclosure shows that microneedle conditioning of the skin using microneedle pressures in the range of about 1 pressure to about 13 pressures at the treatment site can significantly improve transdermal delivery and / or bioavailability, even when compared to microneedling with a relatively higher number of microneedle pressures.

[0016] Furthermore, one of skill in the art reading this disclosure will understand that reducing the total pierced area over which a large drug is applied by reducing the density of microneedles, reducing the puncture size of the microneedles, and / or reducing the pressure of the microneedle array, compared to the relatively higher pierced surface, results in a reduction in the total product volume (i.e., product formulations containing large drugs) administered to the skin area pierced by the microneedles. Thus, the present disclosure teaches one of skill in the art that improved transdermal delivery and / or higher bioavailability of large drugs can be achieved by reducing the total product volume applied (e.g., of formulations containing large drugs).

[0017] Absent the teachings embodied herein, one skilled in the art would typically expect that administering more product would increase the biological effect. Thus, prior to the present disclosure, it would have been expected that applying increasing amounts of product containing a biologically active agent (e.g., large drug), including when such application is in combination with MSCs and / or with product compositions containing emulsions (e.g., nanoemulsions), would achieve increasingly greater biological effects. However, the present disclosure surprisingly shows that beyond a certain "critical" or "threshold" product volume, further increasing the volume of product composition applied to a given skin treatment area in combination with MSCs reduces (rather than increases) the effect. Thus, beyond a certain critical product volume, administration of increasing volumes of product composition containing a biologically active agent (e.g., large drug, particularly including in the case of product compositions containing emulsions) (e.g., in combination with MSCs) reduces the biological effect, despite the increased volume of biologically active agent administered.

[0018] In some embodiments, the skin is administered at 1 drop / cm 2 ~About 5 drops / cm 2By applying a product volume (e.g., of a composition containing a large drug) in the range of about 1 / 100 of a volume in combination with conditioning the skin with one or more impressions of a microneedle array, significant transdermal delivery and / or bioavailability can be achieved. In some embodiments, the skin is applied with a volume of about 1 drop / cm. 2 ~About 4 drops / cm 2 By applying a product volume (e.g., of a composition containing a large drug) in the range of about 1 / 100 of a volume in combination with conditioning the skin with one or more impressions of a microneedle array, significant transdermal delivery and / or bioavailability can be achieved. In some embodiments, the skin is applied with a volume of about 1 drop / cm. 2 ~About 3 drops / cm 2 By applying a product volume (e.g., of a composition containing a large drug) in the range of about 1 / 100 of a volume in combination with conditioning the skin with one or more impressions of a microneedle array, significant transdermal delivery and / or bioavailability can be achieved. In some embodiments, the skin is applied with a volume of about 1 drop / cm. 2 ~About 2.5 drops / cm 2 By applying a product volume (e.g., of a composition containing a large drug) in the range of about 1 / 100 of a drop per cm2 of the skin in combination with conditioning the skin with one or more impressions of a microneedle array, significant transdermal delivery and / or bioavailability can be achieved. 2 ~About 2 drops / cm 2 We disclose that applying a product volume (e.g., of a composition containing a large drug) in the range of about 1 / 100 of the original volume, in combination with conditioning the skin with one or more impressions of a microneedle array, can significantly improve transdermal delivery and / or bioavailability, even when compared to a relatively higher product volume (and / or dose) (e.g., of a large drug).

[0019] In some embodiments, the skin is administered at about 0.0001 mls / cm 2 ~about 0.04mls / cm 2By applying a product volume (e.g., of a composition containing a large drug) in the range of about 0.0001 mls / cm2 in combination with conditioning the skin with one or more impressions of the microneedle array, significant transdermal delivery and / or bioavailability can be achieved. In some embodiments, the skin is applied with a volume of about 0.0001 mls / cm2. 2 ~about 0.05mls / cm 2 By applying a product volume (e.g., of a composition containing a large drug) in the range of about 0.0001 mls / cm2 in combination with conditioning the skin with one or more impressions of the microneedle array, significant transdermal delivery and / or bioavailability can be achieved. In some embodiments, the skin is applied with a volume of about 0.0001 mls / cm2. 2 ~about 0.06mls / cm 2 By applying a product volume (e.g., of a composition containing a large drug) in the range of about 0.0001 mls / cm2 in combination with conditioning the skin with one or more impressions of the microneedle array, significant transdermal delivery and / or bioavailability can be achieved. In some embodiments, the skin is applied with a volume of about 0.0001 mls / cm2. 2 ~about 0.065mls / cm 2 By applying a product volume (e.g., of a composition containing a large drug) in the range of about 0.0025 mls / cm2 in combination with conditioning the skin with one or more impressions of the microneedle array, significant transdermal delivery and / or bioavailability can be achieved. Further, the present disclosure provides a method for applying a product volume of about 0.0025 mls / cm2 to the skin. 2 ~about 0.07mls / cm 2 We disclose that applying a product volume in the range (e.g., of a composition containing a large drug) in combination with conditioning the skin with one or more impressions of a microneedle array can significantly improve transdermal delivery and / or bioavailability, even when compared to relatively higher product volumes (and / or doses) of the large drug.

[0020] Among the advantages offered by certain embodiments of the present disclosure (e.g., those that reduce the volume of product composition applied) is the reduction in administration time (e.g., rub-in time) of topical formulations. As previously mentioned, the present disclosure surprisingly demonstrates that a reduction in the volume (and / or in some embodiments, a reduction in the dose) of a topically applied drug can actually achieve greater delivery (e.g., bioavailability) of the drug across the skin, particularly (but not limited to) when the drug is applied in an emulsion (e.g., nanoemulsion) formulation.

[0021] As mentioned above, the study of microneedle length in small molecule delivery has demonstrated that a relatively longer microneedle length improves small molecule delivery and / or bioavailability. See Yan (Yan, G., et al., "Evaluation needle length and density of microneedle arrays in the pretreatment of skin for transdermal drug delivery", International Journal of Pharmaceutics, 391: 7-12, 2010). However, those skilled in the art who read this disclosure will understand that a reduction in the total area punctured will improve transdermal delivery and / or bioavailability of active substances (e.g., large drugs) compared to a relatively larger total area punctured. Therefore, those skilled in the art who read this disclosure will understand that improved transdermal delivery and / or higher bioavailability of large drugs can be logically achieved by reducing the length of the microneedles of the microneedle skin conditioning array. Those skilled in the art will understand that the longer the microneedle, the larger the base of the microneedle must be to structurally support the increased length, and the larger the base of the needle, the larger the puncture area created by the needle. Those skilled in the art will understand that the longer the microneedle, the larger the surface area of ​​the tissue that is punctured due to the increased surface area of ​​the microneedle itself. The present disclosure surprisingly discloses that microneedle skin conditioning using a relatively shorter microneedle length can result in greater bioavailability than using a relatively longer microneedle length. In some embodiments, microneedle lengths in the range of about 1 μm to about 900 μm can achieve significant transdermal delivery and / or bioavailability. In some embodiments, microneedle lengths of less than 1400 μm, and in some embodiments, less than about 1100 μm or less than 1000 μm are desirable. Indeed, the present disclosure specifically demonstrates the surprising effectiveness of microneedle lengths of about 800 μm or less.In some embodiments, microneedle lengths in the range of about 15 μm to about 800 μm can achieve significant transdermal delivery and / or bioavailability. In some embodiments, the present disclosure shows the surprising effectiveness of microneedle lengths of about 500 μm or less. In some embodiments, microneedle lengths in the range of about 15 μm to about 500 μm can achieve significant transdermal delivery and / or bioavailability.

[0022] In some embodiments, the length of the microneedle may range from about 50 μm to about 900 μm. In some embodiments, the present disclosure discloses that microneedle conditioning of the skin with a microneedle length in the range of about 50 μm to about 900 μm, or about 100 μm to about 700 μm, performed at the treatment site, can significantly improve transdermal delivery and / or bioavailability, even when compared to microneedling with a relatively longer microneedle length. In some embodiments, microneedle conditioning of the skin with a microneedle length in the range of about 100 μm to about 800 μm, performed at the treatment site, can significantly improve transdermal delivery and / or bioavailability, even when compared to microneedling with a relatively longer microneedle length. In some embodiments, microneedle conditioning of the skin at the treatment site using a microneedle length in the range of about 15 μm to about 500 μm can significantly improve transdermal delivery and / or bioavailability, even when compared to microneedling with a relatively longer microneedle length. In some embodiments, microneedle conditioning of the skin at the treatment site using a microneedle length of less than about 800 μm can significantly improve transdermal delivery and / or bioavailability, even when compared to microneedling with a relatively longer microneedle length.

[0023] Among the benefits provided by the unexpected discovery presented herein that shorter needle lengths can achieve more effective delivery (i.e., described as "increased bioavailability") is reduced pain to subjects undergoing microneedle skin conditioning for administration of large drugs (e.g., in combination with and / or as part of a topical treatment). Microneedles of various lengths (including, for example, within the range of about 500 to about 1400 μm) are readily available and have been described as particularly useful because bleeding can be minimized or avoided at these lengths. However, the present disclosure recognizes that even in the absence of bleeding, significant pain may be experienced, especially at the 1400 μm length, and sometimes even at shorter lengths. The present disclosure by demonstrating the effectiveness of microneedle lengths significantly shorter than 1400 μm, and in some embodiments, less than 1100 μm, less than 1000 μm, less than 900 μm or even shorter, offers significant advantages in avoiding or reducing pain in the local administration of large drugs, particularly (but not limited to) those administered in the context of an emulsion (or nanoemulsion) formulation.

[0024] In particular, the present disclosure demonstrates that microneedling techniques (e.g., microneedle conditioning of the skin using relatively lower needle density, relatively smaller microneedle puncture size (e.g., puncture size per microneedle), relatively fewer microneedle impacts, relatively smaller product volume (and / or dose), and relatively shorter needle length) can significantly enhance the transdermal delivery of large drugs, particularly (but not limited to) in emulsion compositions (e.g., macroemulsion compositions and / or nanoemulsion compositions). As illustrated, for example, a microneedle density of about 31 microneedles / cm prior to administration of a large drug (botulinum toxin) can be achieved. 2Preconditioning the skin by applying microneedles with a microneedle array having a microneedle density of less than 100 nm surprisingly improved the delivery of large drugs across the skin. Specific examples included herein demonstrate such improved delivery under various conditions and / or circumstances (e.g., different skin sites, application times, etc.). Those skilled in the art will be aware of other variations (e.g., application sites, administration times, etc.) that are within the scope of the present disclosure.

[0025] Nanoemulsion compositions of particular interest include water-in-oil and oil-in-water nanoemulsions characterized by droplet sizes ranging from about 10 nm to about 300 nm in diameter, an aqueous dispersion medium to oil ratio ranging from about 0.01:1 to about 20:1; an oil to surfactant ratio ranging from about 0.1 to about 40, and / or a zeta potential ranging from about -80 mV to about +80 mV (see, e.g., the nanoemulsion composition descriptions in one or more of PCT / US2006 / 26918; PCT / US06 / 46236; PCT / US2012 / 22276; and PCT / US2012 / 22279, the disclosures of each of which are incorporated herein by reference in their entirety).

[0026] In light of reports that transdermal delivery of solid nanoparticles of a size (e.g., 105±2.92 nm) comparable to that of the droplets in the nanoemulsion compositions utilized herein effectively delivers (or enhances) even small molecule drugs transdermally across the skin, International Application No. PCT / US17 / 53333 has already described certain surprising features achieved by the combination of microneedling and emulsion (e.g., nanoemulsion) technologies. For example, Gomaa et al. described a study in which a solution of rhodamine dye (molecular weight 479 Da) encapsulated in PLGA nanoparticles was applied to skin preconditioned by microneedling to evaluate skin penetration. See Gomaa, Y., et al, "Effect of microneedle treatment on the skin permeation of a nanoencapsulated dye." J Pharm Pharmacol. 2012 November; 64(11): 1592-1602. The data showed that very little dye began to penetrate the skin after 6 hours of continuous application; no significant increase in penetration was observed until the skin was treated continuously for 24 hours. The researchers explained that "there is an emerging consensus that NPs [nanoparticles] can be well deposited in hair follicles, but typically cannot penetrate the stratum corneum." Thus, prior to the present disclosure, one skilled in the art would have expected that the use of microneedling techniques with nano-sized vehicles would not be able to effectively deliver even small molecule drugs (e.g., rhodamine dyes) transdermally; certainly, improved delivery and / or bioavailability of larger drugs would have been thought impossible. International Application No. PCT / US17 / 53333 shows that microneedling can significantly improve the transdermal delivery of large drugs, especially when utilized in conjunction with emulsion (e.g., nanoemulsion) systems. The present disclosure further demonstrates that microneedling can be used to deliver large drugs at a concentration of about 2 to about 50 microneedles / cm. 2These results indicate that microneedling using a microneedle density in the range of about 40 microneedles / cm can significantly enhance the transdermal delivery and / or improve the bioavailability of large drugs compared to using relatively higher microneedle densities, particularly when used in conjunction with emulsion systems (e.g., nanoemulsion systems). In some embodiments, microneedling using a microneedle density in the range of about 40 microneedles / cm can significantly enhance the transdermal delivery and / or improve the bioavailability of large drugs compared to using relatively higher microneedle densities. 2 Less than (e.g., about 2 to about 40 microneedles / cm 2 Microneedling with a microneedle density in the range of 0.01 μm to 0.2 μm, or approximately 35 microneedles / cm 2 Less than (e.g., about 2 to about 35 microneedles / cm 2 Better microneedling with a microneedle density in the range of 0.01 to 0.2 mm, or about 32 microneedles / cm 2 Less than (e.g., about 2 to about 32 microneedles / cm 2 Microneedling with a microneedle density in the range of 0.01 μm to 0.2 μm, or approximately 31 microneedles / cm 2 Less than (e.g., about 2 to about 31 microneedles / cm 2 Microneedling with a microneedle density in the range of 0.01 μm to 0.1 μm, or approximately 30 microneedles / cm 2 Less than (e.g., about 2 to about 30 microneedles / cm 2 Microneedling with a microneedle density in the range of 0.01 μm to 0.2 μm, or approximately 29 microneedles / cm 2 Less than (e.g., about 2 to about 29 microneedles / cm 2 Microneedling with a microneedle density in the range of 0.01 μm to 0.1 μm, or approximately 28 microneedles / cm 2 Less than (e.g., about 2 to about 28 microneedles / cm 2 Microneedling using microneedle densities in the range of 0.1 to 1.0 μm can significantly enhance the transdermal delivery and / or improve the bioavailability of large drugs compared to using relatively higher microneedle densities, especially when utilized in conjunction with emulsion systems (e.g., nanoemulsion systems).

[0027] Furthermore, the present disclosure surprisingly provides a method for producing a cellulose acylate having a thickness of about 100 to about 60,000 μm. 2 The present disclosure discloses that microneedle conditioning of the skin using microneedle puncture sizes in the range of about 100 to about 30,000 μm / microneedle can achieve significant transdermal delivery and / or bioavailability. 2 It is disclosed that microneedling using smaller microneedle puncture sizes in the range of about 50,000 μm / microneedle can significantly enhance the transdermal delivery of large drugs and / or improve their bioavailability, especially when used in conjunction with emulsion systems (e.g., nanoemulsion systems), as compared to using relatively larger microneedle puncture sizes. In some embodiments, the microneedling method is described in detail below. 2 / Less than microneedle (e.g., about 100 to about 50,000 μm 2 Microneedling using a microneedle puncture size of approximately 45,000 μm 2 / Less than microneedle (e.g. about 100 to about 45,000 μm 2 / Microneedle range) refers to microneedling using a microneedle puncture size of approximately 40,000 μm 2 / Less than microneedle (e.g., about 100 to about 40,000 μm 2 Microneedling using a microneedle puncture size of about 35,000 μm / microneedle range 2 / Less than microneedle (e.g. about 100 to about 35,000 μm 2 Better microneedling with a microneedle puncture size of approx. 30,000 μm / microneedle range 2 / Less than microneedle (e.g. about 100 to about 30,000 μm 2 Microneedling using a microneedle puncture size of about 25,000 μm / microneedle range 2 / Less than microneedle (e.g. about 100 to about 25,000 μm 2 Microneedling using microneedle puncture sizes in the range of 0.1 mm / microneedle can significantly enhance the transdermal delivery of large drugs and / or improve their bioavailability compared to using relatively larger microneedle puncture sizes, particularly when utilized in conjunction with emulsion systems (e.g., nanoemulsion systems).

[0028] In particular, the present disclosure discloses that the microneedling techniques described herein can improve transdermal delivery (e.g., of large drugs, especially from macroemulsion or nanoemulsion compositions) when other disruptive agents (i.e., chemical penetration enhancers and other techniques that disrupt or puncture skin structures) are not utilized. Previous studies of transdermal delivery of drugs similar in size to botulinum toxin (i.e., about 150 kDa) using microneedles have reported that delivery fails unless further treatment is applied to disrupt the skin. For example, US Patent Application Publication No. 2010 / 0196445 reports that botulinum toxin is not effectively delivered from pre-coated microneedles unless skin digestive enzymes are also applied so that skin structures are disrupted at the site of microneedling.

[0029] In some embodiments, the present disclosure provides a technique for achieving improved transdermal delivery and / or improved bioavailability of large drugs (e.g., botulinum toxin, antibodies, etc.) by utilizing the microneedling techniques described herein without the need for additional penetration enhancers.Alternatively or in addition, in some embodiments, the present disclosure provides a technique for achieving improved transdermal delivery and / or improved bioavailability of large drugs (e.g., botulinum toxin, antibodies, etc.) by utilizing microneedling techniques without other disruption strategies.Therefore, the provided techniques can achieve effective delivery and / or improved bioavailability without the inflammation, irritation, and / or allergic reactions that are often associated with the use of disruption agents.

[0030] Alternatively or additionally, the present disclosure identifies the source of problems with certain previous approaches to conjugating large drugs, particularly large protein drugs (e.g., botulinum toxin, antibodies, etc.), in or on microneedle structures. Typically, such conventional conjugation strategies utilize a liquid solution of the relevant drug that is applied to the microneedle and air-dried. Such a strategy was utilized to coat microneedles with botulinum toxin in the above-mentioned US Patent Application Publication No. 2010 / 0228225. US Patent Application Publication No. 2017 / 0209553 describes microneedles that are loaded with botulinum. The present disclosure understands that the botulinum coating or loading material produced thereby is not stable and therefore not commercially viable when used to make products. Indeed, even when such liquids are produced from powder materials, the present disclosure understands that for many large drugs (e.g., botulinum toxin), the powder and other solid materials not formed by the freeze-drying process can be extremely unstable. For example, Johnson, E. et al., "Botulinum toxin is highly susceptible to denaturation due to surface denaturation, heat and alkaline conditions. Freeze-drying of botulinum toxin is the most economically sound and practical method of distributing the product in a form that is stable and easily used by clinicians." U.S. Patent No. 5,512,547. Similarly, such an approach would not work for the administration of therapeutic antibodies, which have their own stability and storage challenges. The present disclosure provides the discovery that the use of the emulsion compositions described herein (e.g., in some embodiments, nanoemulsion compositions, and / or in some embodiments, macroemulsion compositions) can protect or otherwise improve the stability of large drugs, particularly large protein drugs (including botulinum toxin and / or antibody drugs in particular), due to their association with microneedles.

[0031] The present disclosure surprisingly provides an effective technique for improving the transdermal delivery and / or bioavailability of large drugs. In particular, the present disclosure teaches that the transdermal delivery of such drugs can be significantly improved by the use of certain microneedling techniques. Those skilled in the art who read the present disclosure will understand that the teachings may be applicable to any topical formulation of large drugs. In some embodiments, the present disclosure teaches that particularly advantageous results are achieved when the microneedling technique is combined with an emulsion composition (e.g., in some embodiments, a nanoemulsion composition, and / or in some embodiments, a macroemulsion composition). In some embodiments, the microneedling technique is combined with a lotion, cream, or liquid composition, which in turn can be or include an emulsion composition (e.g., in some embodiments, using a nanoemulsion embodiment, and / or in some embodiments, using a macroemulsion composition). In some embodiments, the provided technique does not utilize skin-disrupting techniques, such as chemical penetration enhancers. [Brief description of the drawings]

[0032] [Figure 1] FIG. 1 shows the effect of varying microneedle array density on the bioavailability of a botulinum nanoemulsion formulation following MSCs ("microneedle skin conditioning") as determined by viability in a rat study.

[0033] [Diagram 2] FIG. 2 shows the effect of varying microneedle puncture size on the bioavailability of a botulinum nanoemulsion formulation following MSCs as determined by survival in a rat study. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0034] (definition) In this application, unless otherwise clear from the context, (i) as used herein, the term "a" may be understood to mean "at least one"; (ii) the term "or" may be understood to mean "and / or"; (iii) the terms "comprising" and "including" may be understood to encompass the recited elements or steps, whether presented by themselves or with one or more additional elements or steps; and (iv) the terms "about" and "approximately" may be understood to allow for standard variations understood by those of ordinary skill in the art; and (v) when ranges are provided, the endpoints are included.

[0035] Abrasion: As used herein, the term "abrasion" refers to any means that alters, destroys, removes or disrupts the top layer of skin. In some embodiments, abrasion refers to mechanical means that alters, destroys, removes or disrupts the top layer of skin. In some embodiments, abrasion refers to chemical means that alters, destroys, removes or disrupts the top layer of skin. Drugs such as scrubbing agents, microparticles (e.g. magnesium or aluminum particles), acids (e.g. alpha hydroxy acid or beta hydroxy acid), alcohol, to name a few, can cause abrasion. In general, penetration enhancers, such as those described in Donovan (e.g. US Patent Application Publication Nos. 2004 / 009180 and 2005 / 175636, and WO 04 / 06954), and Graham (e.g. US Patent Application Publication No. 6,939,852 and US Patent Application Publication No. 2006 / 093624), are expected to cause abrasion. Of course, those skilled in the art will understand that a particular drug may cause wear when present at one concentration or in combination with one or more other drugs, but may not cause wear under different conditions.Therefore, whether a particular substance is an "wearing agent" depends on the situation.Wear can be easily evaluated by those skilled in the art, for example, by observing redness or irritation of the skin and / or histological examination of the skin, which indicates the change, destruction, removal or erosion of the stratum corneum.

[0036] Administration: As used herein, the term "administration" typically refers to the administration of a composition to a subject or system. Those skilled in the art will be aware of the various routes that may be utilized for administration to a subject, e.g., a human, in the appropriate circumstances. For example, in some embodiments, administration may be intraocular, oral, parenteral, topical, etc. In some embodiments, administration may be bronchial (e.g., by bronchial infusion), buccal, dermal (e.g., one or more of topical administration to the dermis, intradermal, interdermal, transdermal, etc.), enteral, intraarterial, intradermal, intragastric, intramedullary, intramuscular, intranasal, intraperitoneal, intrathecal, intravenous, intraventricular, in a particular organ (e.g., intrahepatic), mucosal, nasal, oral, rectal, subcutaneous, sublingual, topical, tracheal (e.g., by intratracheal infusion), vaginal, vitreous, etc. In some embodiments, administration may include administering intermittently (e.g., multiple administrations separated by time) and / or periodically (e.g., individual administrations separated by a common period of time). In some embodiments, administration can include continuous administration (eg, perfusion) for at least a selected period of time.

[0037] Drug: In general, the term "drug" as used herein may be used to refer to any chemical class of compound or entity, including, for example, polypeptides, nucleic acids, saccharides, lipids, small molecules, metals, or combinations or complexes thereof. In appropriate circumstances, as will be clear to those skilled in the art from the context, the term may be used to refer to an entity that is or includes a cell or organism, or a fraction, extract, or component thereof. Alternatively or additionally, as will be clear from the context, the term may be used to refer to a natural product, in that it is found in nature and / or is obtained from nature. In some cases, again as will be clear from the context, the term may be used to refer to one or more entities that are artificial, in that they are designed, engineered, and / or produced by the action of the human hand, and / or are not found in nature. In some embodiments, the drug may be used in an isolated or pure form; in some embodiments, the drug may be used in a crude form. In some embodiments, potential drugs may be provided as a collection or library that can be screened, for example, to identify or characterize active substances therein. In some cases, the term may refer to a compound or entity that is or comprises a polymer; in some cases, the term may refer to a compound or entity that comprises one or more polymer molecules.In some embodiments, the term "drug" may refer to a compound or entity that is not a polymer and / or does not substantially comprise any polymer and / or one or more specific polymeric moieties.In some embodiments, the term may refer to a compound or entity that lacks or does not substantially comprise any polymeric moieties.In some embodiments, the term may refer to a molecular complex.

[0038] Antibody: As used herein, the term "antibody" refers to a polypeptide that contains sufficient standard immunoglobulin sequence elements to confer specific binding to a particular target antigen. As is known in the art, naturally produced intact antibodies are tetrameric entities of approximately 150 kDa composed of two identical heavy chain polypeptides (each approximately 50 kDa) and two identical light chain polypeptides (each approximately 25 kDa) that bind to each other in what is commonly referred to as a "Y-shaped" structure. Each heavy chain is composed of at least four domains (each approximately 110 amino acids long) - an amino-terminal variable (VH) domain (located at the tip of the Y structure), followed by three constant domains CH1, CH2, and a carboxy-terminal CH3 (located at the base of the stem of the Y). A short region known as the "switch" connects the variable and constant regions of the heavy chain. A "hinge" connects the CH2 and CH3 domains to the rest of the antibody. Disulfide bonds in two of the hinge regions connect the two heavy chain polypeptides to each other in the intact antibody. Each light chain is composed of two domains - an amino-terminal variable (VL) domain followed by a carboxy-terminal constant (CL) domain, which are separated from each other by another "switch". An intact antibody tetramer is composed of two heavy-light chain dimers in which the heavy and light chains are linked to each other by a single disulfide bond; two other disulfide bonds connect the heavy chain hinge regions to each other, resulting in the dimers being connected to each other and forming a tetramer. Naturally produced antibodies are also typically glycosylated in the CH2 domain. Each domain of a natural antibody has a structure characterized by an "immunoglobulin fold" formed from two beta sheets (e.g., 3-, 4- or 5-stranded sheets) packed together in a compressed antiparallel beta barrel. Each variable domain contains three hypervariable loops known as "complement determining regions" (CDR1, CDR2 and CDR3) and four somewhat invariant "framework" regions (FR1, FR2, FR3 and FR4). When a native antibody folds, the FR regions form beta sheets that provide the structural framework for the domain, and the CDR loop regions of both the heavy and light chains are brought together in three-dimensional space to create a single hypervariable antigen-binding site located at the tip of a Y-structure.The Fc region of a naturally occurring antibody binds to elements of the complement system and also to receptors on effector cells, including, for example, effector cells that mediate cytotoxicity. As is known in the art, the affinity and / or other binding properties of the Fc region for an Fc receptor can be modulated by glycosylation or other modifications. In some embodiments, the antibodies produced and / or utilized in accordance with the present invention comprise a glycosylated Fc domain, including Fc domains with such glycosylation that have been modified or engineered. For purposes of the present invention, in some embodiments, any polypeptide or polypeptide complex that comprises a sufficient immunoglobulin domain sequence as found in a natural antibody may be referred to and / or used as an "antibody", regardless of whether such polypeptide is produced naturally (e.g., produced by an organism in response to an antigen) or produced by recombinant engineering, chemical synthesis or other artificial systems or methods. In some embodiments, the antibody is polyclonal; in some embodiments, the antibody is monoclonal. In some embodiments, the antibody has constant region sequences characteristic of mouse, rabbit, primate or human antibodies. In some embodiments, the antibody sequence elements are humanized, primatized, chimeric, etc., as known in the art. Furthermore, the term "antibody" as used herein, in appropriate embodiments (unless otherwise stated or clear from the context), may refer to any of the constructs or formats known or developed in the art for utilizing the structural and functional features of antibodies in surrogate presentation. For example, in embodiments, the antibodies utilized in accordance with the present invention may be intact IgG, IgE and IgM, bi- or multispecific antibodies (such as Zybody®), single chain Fv, Fc fusions of polypeptides, Fab, camelid antibodies, masked antibodies (such as Probodies®), etc. S mall M odd I mmuno P harmaceutical ("SMIP TM"), single chain or tandem diabody antibodies (TandAb®), VHH, Anticalin®, Nanobody®, minibody, BiTE®, ankyrin repeat protein or DARPIN®, Avimer®, DaRT, TCR-like antibody, Adnectin®, Affilin®, Trans-body®, Affibody®, TrimerX®, MicroProtein, Fynomer®, Centyrin® and KALBITOR®. In some embodiments, the antibody may lack a covalent modification (e.g., attachment of a glycan) that it would have if it were produced naturally (e.g., in a mammalian organism). In some embodiments, the antibody may include a covalent modification (e.g., attachment of a glycan, a payload (e.g., a detectable moiety, a therapeutic moiety, a catalytic moiety, etc.), or other pendant group (e.g., polyethylene glycol, etc.).

[0039] Antibody drug: As used herein, the term "antibody drug" refers to a drug that specifically binds to a particular antigen. In some embodiments, the term encompasses any polypeptide or polypeptide complex that contains sufficient immunoglobulin structural elements to confer specific binding. Exemplary antibody drugs include human antibodies, primatized antibodies, chimeric antibodies, bispecific antibodies, humanized antibodies, conjugated antibodies (i.e., antibodies conjugated or fused to other proteins, radiolabels, cytotoxins), S mall M odd I mmuno P harmaceutical ("SMIP TM"), single chain antibodies, camelid antibodies and antibody fragments. As used herein, the term "antibody drug" also includes intact monoclonal antibodies, polyclonal antibodies, single domain antibodies (e.g., shark single domain antibodies (e.g., IgNAR or fragments thereof)), multispecific antibodies (e.g., bispecific antibodies) formed from at least two intact antibodies, and antibody fragments, so long as they exhibit the desired biological activity. In some embodiments, the term includes stapled peptides. In some embodiments, the term includes one or more antibody-like binding peptidomimetics. In some embodiments, the term includes one or more antibody-like binding scaffold proteins. In some embodiments, the term includes monobodies or adnectins. In many embodiments, the antibody drug is or comprises a polypeptide whose amino acid sequence comprises one or more structural elements recognized by those skilled in the art as a complementarity determining region (CDR); in some embodiments, the antibody drug is or comprises a polypeptide whose amino acid sequence comprises at least one CDR (e.g., at least one heavy chain CDR and / or at least one light chain CDR) that is substantially identical to that found in a reference antibody. In some embodiments, the included CDR is substantially identical to the reference CDR in that it is identical in sequence or contains 1-5 amino acid substitutions compared to the reference CDR. In some embodiments, the included CDR is substantially identical to the reference CDR in that it exhibits at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity with the reference CDR. In some embodiments, the included CDR is substantially identical to the reference CDR in that it exhibits at least 96%, 96%, 97%, 98%, 99% or 100% sequence identity with the reference CDR. In some embodiments, the included CDR is substantially identical to the reference CDR in that at least one amino acid in the included CDR is deleted, added or substituted compared to the reference CDR, but the included CDR has an amino acid sequence that is otherwise identical to that of the reference CDR.In some embodiments, the included CDRs are substantially identical to the reference CDRs in that 1-5 amino acids in the included CDRs are deleted, added or substituted compared to the reference CDRs, but have an amino acid sequence that is otherwise identical to that of the reference CDRs. In some embodiments, the included CDRs are substantially identical to the reference CDRs in that at least one amino acid is substituted compared to the reference CDRs, but have an amino acid sequence that is otherwise identical to that of the reference CDRs. In some embodiments, the included CDRs are substantially identical to the reference CDRs in that 1-5 amino acids in the included CDRs are deleted, added or substituted compared to the reference CDRs, but have an amino acid sequence that is otherwise identical to that of the reference CDRs. In some embodiments, the antibody drug is or comprises a polypeptide whose amino acid sequence comprises structural elements recognized by those skilled in the art as an immunoglobulin variable domain. In some embodiments, the antibody drug is a polypeptide protein having a binding domain that is homologous or largely homologous to an immunoglobulin binding domain. In some embodiments, the antibody drug is or comprises an antibody-drug conjugate.

[0040] Antibody component: As used herein, an antibody component refers to a polypeptide element (which may be a complete polypeptide or a part of a larger polypeptide, such as, for example, a fusion polypeptide as described herein) that specifically binds to an epitope or antigen and contains one or more immunoglobulin structural features. In general, an antibody component is any polypeptide whose amino acid sequence contains elements characteristic of an antibody binding region (e.g., an antibody light chain or variable region or one or more complementarity determining regions ("CDRs") thereof, or an antibody heavy chain or variable region or one or more CDRs thereof, optionally in the presence of one or more framework regions). In some embodiments, an antibody component is or comprises a full-length antibody. In some embodiments, an antibody component is short in full length but contains at least one binding site (containing at least one, preferably at least two sequences with the structure of a known antibody "variable region"). In some embodiments, the term "antibody component" encompasses any protein having a binding domain that is homologous or largely homologous to an immunoglobulin binding domain. In certain embodiments, the "antibody component" encompassed encompasses a polypeptide having a binding domain that exhibits at least 99% identity to an immunoglobulin binding domain. In some embodiments, the "antibody component" included is any polypeptide having an immunoglobulin binding domain, e.g., a binding domain that exhibits at least 70%, 75%, 80%, 85%, 90%, 95% or 98% identity with a reference immunoglobulin binding domain. The "antibody component" included may have an amino acid sequence identical to that of an antibody (or a portion thereof, e.g., an antigen-binding portion thereof) found in a natural source. The antibody component may be monospecific, bispecific or multispecific. The antibody component may include structural elements characteristic of any immunoglobulin class, including any of the human classes of IgG, IgM, IgA, IgD and IgE. It has been shown that the antigen-binding function of an antibody can be performed by fragments of a full-length antibody.Such antibody embodiments may also be bispecific, dual specific or multispecific formats that specifically bind to two or more different antigens. Examples of binding fragments encompassed by the term "antigen-binding portion" of an antibody include (i) V. H , V L , C H 1 and C L (ii) a bivalent fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region, called F(ab') 2 Fragment; (iii) V H and C H (iv) a single-arm V of an antibody; H and V L (v) a Dab fragment containing a single variable domain (Ward et al., (1989) Nature 341:544-546); (vi) an isolated complementarity determining region (CDR). In addition, the V of the two domains of the Fv fragment may be H and V L are encoded by separate genes, but V H and V Lの The domains may be recombinantly linked by synthetic linkers that allow them to be made as a single protein chain, where they pair to form monovalent molecules (known as single-chain Fvs (scFvs); see, e.g., Bird et al. (1988) Science 242:423-426; and Huston et al. (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883). In some embodiments, the "antibody component" described herein is or comprises such a single-chain antibody. In some embodiments, the "antibody component" is or comprises a diabody. Diabodies are a type of antibody that is composed of a V H and V LBivalent, bispecific antibodies are those in which the domains are expressed on a single polypeptide chain, but with a linker that is too short to allow pairing between the two domains on the same chain, and thus the domains pair with complementary domains on the other chain, creating two antigen-binding sites (see, e.g., Holliger, P., et al., (1993) Proc. Natl. Acad. Sci. USA 90:6444-6448; Poljak, RJ, (1994) Structure 2(12):1121-1123). Such antibody binding moieties are known in the art (Kontermann and Dubel, eds., Antibody Engineering (2001) Springer-Verlag. New York. 790 pp. (ISBN 3-540-41354-5)). In some embodiments, the antibody component comprises a pair of tandem Fv segments (Vs) that, together with complementary light chain polypeptides, form a pair of antigen-binding regions. H -C H 1-V H -C H 1) (Zapata et al., (1995) Protein Eng. 8(10): 1057-1062; and U.S. Pat. No. 5,641,870). In some embodiments, the antibody component may have the characteristic components of chimeric or humanized antibodies. In general, humanized antibodies are human immunoglobulins (recipient antibody) in which residues from a complementarity determining region (CDR) of the recipient are replaced by residues from a CDR of a non-human species (donor antibody) such as mouse, rat or rabbit having the desired specificity, affinity, and capacity. In some embodiments, the antibody component may have the characteristic components of human antibodies.

[0041] Antibody fragment: As used herein, "antibody fragment" includes a portion of an intact antibody, such as, for example, the antigen-binding or variable region of the antibody. Examples of antibody fragments include Fab, Fab', F(ab')2 and Fv fragments; triabodies; tetrabodies; linear antibodies; single-chain antibody molecules; and multispecific antibodies formed from antibody fragments. For example, antibody fragments include isolated fragments, "Fv" fragments consisting of the variable regions of the heavy and light chains, recombinant single-chain polypeptide molecules in which the light and heavy chain variable regions are connected by a peptide linker ("ScFv protein"), and minimal recognition units consisting of amino acid residues that mimic the hypervariable region. In many embodiments, an antibody fragment includes sufficient sequence of a parent antibody that it is a fragment that binds to the same antigen as the parent antibody; in some embodiments, the fragment binds to the antigen with an affinity comparable to that of the parent antibody and / or competes with the parent antibody for binding to the antigen. Examples of antigen-binding fragments of antibodies include, but are not limited to, Fab fragments, Fab' fragments, F(ab')2 fragments, scFv fragments, Fv fragments, dsFv diabodies, dAb fragments, Fd' fragments, Fd fragments, and isolated complementarity determining region (CDR) regions. Antigen-binding fragments of antibodies can be produced by any means. For example, antigen-binding fragments of antibodies can be enzymatically or chemically produced by fragmentation of an intact antibody and / or recombinantly produced from a gene encoding a partial antibody sequence. Alternatively or additionally, antigen-binding fragments of antibodies can be wholly or partially synthetically produced. Antigen-binding fragments of antibodies can optionally include single-chain antibody fragments. Alternatively or additionally, antigen-binding fragments of antibodies can include multiple chains linked together, for example, by disulfide bonds. Antigen-binding fragments of antibodies can optionally include multimolecular complexes. Functional antibody fragments typically include at least about 50 amino acids, more typically include at least about 200 amino acids.

[0042] About: As used herein, the term "approximately" or "about" refers to a value that is similar to the stated reference value when applied to one or more target values.In some embodiments, the term "approximately" or "about" refers to a range of values ​​that is within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greater or smaller) of the stated reference value, unless otherwise stated or otherwise clear from the context (e.g., when one or more target values ​​define a sufficiently narrow range, the application of such percentage variations is unnecessary).

[0043] Associated: Two events or entities are "associated" with each other, as the term is used herein, when the presence, level and / or form of one correlates with that of the other. For example, a particular entity (e.g., a polypeptide, a gene signature, a metabolite, a microorganism, etc.) is considered to be associated with a particular disease, disorder or condition when its presence, level and / or form correlates with the incidence of and / or susceptibility to the disease, disorder or condition (e.g., an associated population). In some embodiments, two or more entities are physically "associated" with each other when they directly or indirectly interact with each other, and thus are in and / or remain in physical proximity to each other. In some embodiments, two or more entities that are physically associated with each other are covalently bound to each other; in some embodiments, two or more entities that are physically associated with each other are not covalently bound to each other, but are non-covalently bound, for example, by hydrogen bonds, van der Waals interactions, hydrophobic interactions, magnetism, and combinations thereof.

[0044] Biocompatible: As used herein, the term "biocompatible" refers to a material that does not cause significant harm to living tissue when placed in contact with such tissue, for example in vivo. In some embodiments, a material is "biocompatible" if it is not toxic to cells. In some embodiments, a material is "biocompatible" if its addition to cells in vitro results in 20% or less cell death, and / or its administration in vivo does not induce significant inflammation or other such adverse effects.

[0045] Biodegradable: As used herein, the term "biodegradable" refers to a material that, when introduced into cells, is degraded (e.g., by cellular mechanisms such as enzymatic degradation, hydrolysis, and / or a combination thereof) into components that the cells can reuse or discard without significant toxic effects on the cells. In some embodiments, the components generated by the degradation of the biodegradable material are biocompatible and therefore do not induce significant inflammation and / or other adverse effects in vivo. In some embodiments, the biodegradable polymeric materials degrade into their component monomers. In some embodiments, the degradation of the biodegradable material (including, for example, the biodegradable polymeric material) includes hydrolysis of ester bonds. Alternatively or additionally, in some embodiments, the degradation of the biodegradable material (including, for example, the biodegradable polymeric material) includes cleavage of urethane bonds. Exemplary biodegradable polymers are polymers of hydroxy acids, such as, for example, lactic acid and glycolic acid, including, but not limited to, poly(hydroxyl acids), poly(lactic acid) (PLA), poly(glycolic acid) (PGA), lactic-glycolic acid copolymers (PLGA), and copolymers with PEG, polyanhydrides, poly(ortho)esters, polyesters, polyurethanes, poly(butyric acid), poly(valeric acid), poly(caprolactone), poly(hydroxyalkanoates), lactide-caprolactone copolymers, mixtures and copolymers thereof. Many naturally occurring polymers are also biodegradable, including, for example, proteins, such as albumin, collagen, gelatin, and prolamines, such as zein, and polysaccharides, such as alginic acid, cellulose derivatives, and polyhydroxyalkanoates, such as polyhydroxybutyrate, mixtures and copolymers thereof. One of ordinary skill in the art will understand or be able to determine when such a polymer (e.g., related to the parent polymer by substantially identical structure differing only in the substitution or addition of certain chemical groups, as known in the art) is biocompatible and / or a biodegradable derivative thereof.

[0046] Biologically active agent: As used herein, the term "biologically active agent" refers to a drug that has a specific biological effect when administered to a subject, e.g., a human. In some embodiments, the biologically active agent may be a therapeutically active agent, a cosmetically active agent, and / or a diagnostically active agent. In some embodiments, the biologically active agent may be or include an entity or moiety that would be classified as an "active pharmaceutical ingredient" by the U.S. Food and Drug Administration. In some embodiments, the biologically active agent is a large drug. In some embodiments, the biologically active agent may be or include a drug whose presence correlates with a desired pharmacological and / or therapeutic, cosmetic and / or diagnostic effect. In some embodiments, the biologically active agent is characterized in that its biological effect is dose-dependent (e.g., increases linearly with increasing dose, sometimes over at least a first concentration range). In some embodiments, a drug is not considered to be a "biologically active agent" if it merely enhances the delivery of a different drug that actually achieves a desired effect.

[0047] Botulinum macroemulsion composition: As used herein, the term "botulinum macroemulsion composition" refers to any macroemulsion composition in which at least one macroemulsion contains a botulinum toxin. The botulinum toxin may be present within the macroemulsion, on the surface of the macroemulsion, and / or within the micellar membrane that defines the macroemulsion.

[0048] Botulinum nanoemulsion composition: As used herein, the term "botulinum nanoemulsion composition" refers to any nanoemulsion composition in which at least one nanoemulsion contains a botulinum toxin. The botulinum toxin may be present within the nanoemulsion, on the surface of the nanoemulsion, and / or within the micellar membrane that defines the nanoemulsion.

[0049] Botulinum toxin: As used herein, the term "botulinum toxin" refers to any neurotoxin produced by Clostridium botulinum. Unless otherwise specified, the term includes fragments or portions (e.g., light and / or heavy chains) of such neurotoxins that retain the appropriate activity (e.g., muscle relaxant activity). The term "botulinum toxin" includes botulinum toxin serotypes A, B, C, D, E, F and G. As used herein, botulinum toxin also includes both botulinum toxin complexes (i.e., e.g., 300, 600 and 900 kDa complexes) and purified (i.e., e.g., isolated) botulinum toxins (i.e., e.g., about 150 kDa). A "purified botulinum toxin" is defined as a botulinum toxin that has been isolated or substantially isolated from other proteins, including proteins for the botulinum toxin complex. A purified toxin can be greater than 95% pure, and preferably greater than 99% pure. Those skilled in the art will appreciate that the present invention is not limited to a particular source of botulinum toxin. For example, botulinum toxins for use according to the present invention can be isolated from Clostridium botulinum, chemically synthesized, or produced recombinantly (i.e., in a host cell or organism other than Clostridium botulinum). Botulinum can be genetically engineered or chemically modified to act with a longer or shorter duration than botulinum toxin serotype A.

[0050] Carrier: as used herein, carrier refers to the diluent, adjuvant, additive or vehicle that the composition is administered together with.In some exemplary embodiments, carrier comprises, for example, sterile liquid, such as water and oil, such as oil of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil, etc.In some embodiments, carrier is or comprises one or more solid components.

[0051] Combination therapy: As used herein, the term "combination therapy" refers to a situation in which a subject is exposed to two or more therapeutic regimens (e.g., two or more therapeutic agents, therapeutic agents and therapeutic modalities, etc.) at the same time. In some embodiments, two or more regimens can be administered simultaneously; in some embodiments, such regimens can be administered sequentially (e.g., all "doses" of a first regimen are administered before administration of any dose of a second regimen). In some embodiments, such drugs are administered in overlapping dosing regimens. In some embodiments, "administration" of a combination therapy can include administration of one or more drugs and / or modalities to a subject that is administered the other drug or modality in combination. For clarity, in some embodiments, two or more drugs or active portions thereof can be administered in a combination composition or in a combination compound (e.g., as part of a single chemical complex or covalent entity), but combination therapy does not require that the individual drugs are administered together (or even necessarily at the same time) in a single composition.

[0052] Equivalent: As used herein, the term "equivalent" refers to two or more drugs, entities, situations, sets of conditions, etc., which may not be identical to each other, but are sufficiently similar to allow comparison between them, so that a person skilled in the art will understand that a conclusion can be reasonably drawn based on the differences or similarities observed. In some embodiments, an equivalent set of conditions, environments, individuals, or populations is characterized by a number of substantially identical characteristics and one or a few different characteristics. A person skilled in the art will understand how much identity is required in a given environment in order for two or more such drugs, entities, situations, sets of conditions, etc. to be considered equivalent in context. For example, a person skilled in the art will understand that a set of situations, individuals, or populations is equivalent to each other when it is characterized by a sufficient number and type of substantially identical characteristics to ensure a reasonable conclusion that the differences between the results obtained or the phenomena observed under or with the various sets of environments, individuals, or populations are caused by or indicate the alteration in these characteristics that have changed.

[0053] Composition: Those skilled in the art will appreciate that the term "composition" as used herein may be used to refer to a separate physical entity that includes one or more specific components. Generally, unless otherwise specified, a composition may be in any form, such as a gas, gel, liquid, solid, etc.

[0054] Comprising: Compositions or methods described herein as "comprising" one or more specified components or steps are open-ended, meaning that the specified components or steps are essential, but that other components or steps may be added within the scope of the composition or method. To avoid redundancy, it is also understood that any composition or method described as "comprising" (or "comprises") one or more specified components or steps also describes a corresponding, more limited composition or method "consisting essentially of" (or "consist essentially of") the same specified components or steps, meaning that the composition or method includes the specified essential components or steps, and may also include additional components or steps that do not materially affect the basic and novel characteristics of the composition or method. It is also understood that any composition or method described herein as "comprising" or "consisting essentially of" one or more specified components or steps also describes the corresponding, more limited, close-ended composition or method "consisting of" (or "consist of") the specified components or steps, excluding other unspecified components or steps. In any composition or method disclosed herein, known or disclosed equivalents of any required specified component or step may be substituted for that component or step.

[0055] Dosage form or unit dosage form: Those skilled in the art will understand that the term "dosage form" can be used to refer to a physically discrete unit of an active agent (e.g., a therapeutic or diagnostic agent) for administration to a subject. Typically, each such unit contains a predetermined amount of the active agent. In some embodiments, such amount is a unit dosage (or a whole fraction thereof) appropriate for administration according to a dosing regimen (i.e., a therapeutic dosing regimen) determined to correlate with a desired or beneficial outcome when administered to a relevant population. Those skilled in the art will understand that the total amount of a therapeutic composition or drug to be administered to a particular subject is determined by one or more physicians and may include administration of multiple dosage forms.

[0056] Dosing regimen: Those skilled in the art will understand that the term "dosing regimen" can be used to refer to a series of unit doses (typically multiple) that are administered individually to a subject and typically separated by a period of time. In some embodiments, a given therapeutic agent has a recommended dosing regimen, which can include one or more doses. In some embodiments, the dosing regimen includes multiple doses, each of which is separated in time from the other doses. In some embodiments, the individual doses are separated from each other by periods of the same length; in some embodiments, the dosing regimen includes multiple doses, and at least two different periods that separate the individual doses. In some embodiments, all doses in the dosing regimen are the same unit dose. In some embodiments, the different doses in the dosing regimen are different amounts. In some embodiments, the dosing regimen includes a first administration of a first dose, followed by one or more further administrations of a second dose that is different from the first dose. In some embodiments, the dosing regimen includes a first administration of a first dose, followed by one or more further administrations of a second dose that is the same as the first dose. In some embodiments, the dosing regimen correlates with a desired or beneficial outcome when administered to an entire relevant population (ie, is a therapeutic dosing regimen).

[0057] Emulsion: The term "emulsion" is used herein consistent with its understanding in the art as "a system consisting of a liquid dispersed, with or without an emulsifying agent, in an immiscible liquid, usually in droplets greater than colloidal size." See, e.g., Medline Plus Online Medical Dictionary, Merriam Webster (2005).

[0058] Additives: As used herein, additives refer to non-therapeutic substances that may be included in a pharmaceutical composition, for example, to provide or contribute to a desired consistency or stabilizing effect. Suitable pharmaceutical additives include, for example, starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, dried skim milk, glycerol, propylene, glycol, water, ethanol, etc.

[0059] Human: In some embodiments, the human is an embryo, fetus, infant, child, teenager, adult, or elderly.

[0060] Hydrophilic: As used herein, the terms "hydrophilic" and / or "polar" refer to the tendency to mix with or dissolve readily in water.

[0061] Hydrophobic: As used herein, the terms "hydrophobic" and / or "non-polar" refer to the tendency to repel, not mix with, or not readily dissolve in water.

[0062] Improve, increase or decrease: As used herein or its grammatical equivalents, the terms "improvement," "increase," or "decrease" refer to a value relative to a baseline measurement, e.g., a measurement in the same individual before the initiation of a treatment described herein, or a measurement in a control individual (or control individuals) in the absence of a treatment described herein. In some embodiments, a "control individual" is an individual suffering from the same form of disease or disorder as the individual being treated.

[0063] Macromolecule: The term "macromolecule" is generally used herein to describe a molecule that is greater than about 100 kilodaltons (KDa) in size. In some embodiments, the macromolecule is greater than about 110 kDa, about 120 kDa, about 130 kDa, about 140 kDa, about 150 kDa, about 160 kDa, about 170 kDa, about 180 kDa, about 190 kDa, about 200 kDa, about 250 kDa, about 300 kDa, about 400 kDa or about 500 kDa. In some embodiments, the macromolecule is a polymer or comprises a polymer moiety or entity. In some embodiments, the macromolecule is or comprises a polypeptide. In some embodiments, the macromolecule is or comprises a nucleic acid.

[0064] Large drug: As used herein, the term "large drug" generally refers to a drug with a molecular weight of more than about 100 kilodaltons (KDa). In some embodiments, the large molecule is greater than about 110 kDa, about 120 kDa, about 130 kDa, about 140 kDa, about 150 kDa, about 160 kDa, about 170 kDa, about 180 kDa, about 190 kDa, about 200 kDa, about 250 kDa, about 300 kDa, about 400 kDa or about 500 kDa. In some embodiments, the large drug is a biologically active substance. In some embodiments, the large drug is or comprises one or more large molecules. In some embodiments, the large drug is or comprises one or more molecular complexes. In some embodiments, the large drug is or comprises a polypeptide. In some embodiments, the large drug is or comprises a polypeptide complex. In some embodiments, the large drug is or comprises a bacterial toxin (e.g., a botulinum toxin). In some embodiments, the large drug is or comprises an antibody drug.

[0065] Macroemulsion: As used herein, the term "macroemulsion" refers to an emulsion in which at least some of the droplets have a diameter in the size range of hundreds of nanometers to several micrometers. As will be understood by those skilled in the art, macroemulsions are characterized by droplets with a diameter of more than 300 nm. In some embodiments, the macroemulsion composition utilized in accordance with the present disclosure includes one or more larger drugs or one or more biologically active substances. In some embodiments, the large drug included in the macroemulsion composition can be a biologically active substance. It will be understood by those skilled in the art that the macroemulsion composition for use in accordance with the present disclosure can be manufactured according to any available means, including, for example, chemical or mechanical means. In some embodiments, the droplets in the macroemulsion have a size in the range of about 301 nm to about 1000 μm. In some embodiments, the macroemulsion has a droplet size distribution of about 301 nm to about 1000 μm. In some embodiments, the droplets in the macroemulsion have a size in the range of about 500 nm to about 5000 μm. In some embodiments, the macroemulsions have a droplet size distribution of about 500 nm to about 5000 μm.

[0066] Microneedle: As used herein, the term "microneedle" generally refers to an elongated structure of suitable length, diameter and shape to penetrate the skin. In some embodiments, the microneedle is arranged and constructed (by itself or within a device) to create an efficient pathway for drug delivery while minimizing contact with nerves when inserted into the skin. In some embodiments, the microneedle has a diameter that is constant along the length of the microneedle. In some embodiments, the microneedle has a diameter that varies along the length of the microneedle. In some embodiments, the microneedle has a diameter that tapers along the length of the microneedle. In some embodiments, the diameter of the microneedle is narrowest at the tip that penetrates the skin. In some embodiments, the microneedle can be solid. In some embodiments, the microneedle can be hollow. In some embodiments, the microneedle can be tubular. In some embodiments, the microneedle can be sealed at one end. In some embodiments, multiple microneedles are utilized. In some embodiments, multiple microneedles are utilized in an array format. In some embodiments, the microneedle may have a length in the range of about 1 μm to about 4,000 μm. In some embodiments, the microneedle may have a length of about 1 μm to about 2,000 μm. In some embodiments, the microneedle may have a length of about 50 μm to about 400 μm. In some embodiments, the microneedle may have a length of about 800 μm to about 1500 μm.

[0067] Pressing of microneedle array: As used herein, the term "pressing of microneedle array" refers to microneedle pressing achieved by pressing microneedles and / or microneedle arrays onto skin and then removing them from the skin. In some embodiments, the microneedle array can be stamped onto the skin (e.g., using a microneedle array stamp). In some embodiments, the microneedle array can be rolled onto the skin (e.g., using a microneedle array roller).

[0068] Microneedle density: As used herein, the term "microneedle density" refers to the number of microneedles per area measurement (e.g., square centimeter). In some embodiments, the microneedle density is evaluated as the number of microneedles per area of ​​a microneedle array; in some embodiments, the microneedle density is evaluated as the number of microneedle punctures per area of ​​the site to be microneedled; in some embodiments, the microneedle density is evaluated as the number of microneedles per area that simultaneously achieves maximum or near maximum skin penetration possible for the microneedles in the array. In any case, those skilled in the art will understand that microneedle density can be expressed regardless of whether the relevant area is flat (e.g., a microneedle array stamp), curved (e.g., a microneedle array roller) or irregular. Those skilled in the art will appreciate that assessing the density of microneedles as microneedle punctures per area of ​​the microneedled site can be particularly useful, for example, when an array has needles of different lengths and / or when the site to be microneedled has topological diversity such that not all needles are actually able to puncture the skin when the array is applied to the site.

[0069] Microneedle puncture size: As used herein, the term "microneedle puncture size" or "microneedle puncture hole size" refers to the calculated puncture area made by each microneedle of a microneedle array, which is achieved after pressing the microneedle and / or microneedle array onto the skin and removing it from the skin. In many embodiments, the microneedle puncture size is calculated as the area of ​​the base of the microneedle.

[0070] Nanoemulsion: As used herein, the term "nanoemulsion" refers to an emulsion in which at least some of the droplets have diameters in the nanometer size range. As will be understood by those of skill in the art, nanoemulsions are characterized by droplets with diameters of 300 nm or less. In some embodiments, the nanoemulsion compositions utilized in accordance with the present disclosure include one or more large drugs or one or more biologically active substances. In some embodiments, the large drugs included in the nanoemulsion composition can be biologically active substances. It will be understood by those of skill in the art that the nanoemulsion compositions for use in accordance with the present disclosure can be manufactured according to any available means, including, for example, chemical or mechanical means. In some embodiments, the droplets in the nanoemulsion have a size in the range of about 1 nm to about 300 nm. In some embodiments, the nanoemulsion has droplets with a size distribution of about 1 nm to about 300 nm.

[0071] Nanoparticle: The term "nanoparticle" as used herein refers to a solid particle having a diameter of less than 300 nm, as defined by the National Science Foundation.In some embodiments, nanoparticles have a diameter of less than 100 nm, as defined by the National Institutes of Health.

[0072] Patient: As used herein, the term "patient" refers to any organism to which the provided compositions are or can be administered, for example, for experimental, diagnostic, prophylactic, cosmetic and / or therapeutic purposes. Typical patients include animals (e.g., mammals, such as mice, rats, rabbits, non-human mammals and / or humans). In some embodiments, the patient is a human. In some embodiments, the patient is suffering from or susceptible to one or more disorders or conditions. In some embodiments, the patient exhibits one or more symptoms of a disorder or condition. In some embodiments, the patient has been diagnosed with one or more disorders or conditions. In some embodiments, the disorder or condition is or includes the presence of one or more tumors. In some embodiments, the patient is undergoing or has undergone a particular therapy to diagnose and / or treat a disease, disorder or condition.

[0073] Penetration enhancer: As used herein, the term "penetration enhancer" refers to a substance whose presence or level correlates with increased penetration of a target drug across the skin compared to that observed in its absence. In some embodiments, the penetration enhancer is characterized by its ability to degrade and / or destroy skin structure. In some embodiments, the penetration enhancer is or includes a chemical agent (e.g., a chemical or enzyme). For example, chemical agents that can damage, destroy and / or degrade one or more components of the stratum corneum can include, for example, alcohols, such as short-chain alcohols, long-chain alcohols or polyalcohols; amines and amides, such as urea, amino acids or their esters, amides, AZONE®, derivatives of AZONE®, pyrrolidones or derivatives of pyrrolidones; terpenes and derivatives of terpenes; fatty acids and their esters; macrocyclic compounds; tensides; or sulfoxides (e.g., dimethyl sulfoxide (DMSO), decyl methyl sulfoxide, etc.); surfactants, such as anionic, cationic and nonionic surfactants; polyols; essential oils; and / or hyaluronidase. In some embodiments, the penetration enhancer may be irritating in that it causes inflammatory and / or allergic reactions when the substance is applied to the skin. In some embodiments, the penetration enhancer is not irritating. In some embodiments, the penetration enhancer may be or include a chemical agent that does not damage, destroy or degrade skin structure, but whose presence or level nevertheless correlates with increased penetration of the target drug across the skin compared to that observed in its absence. In some embodiments, the copeptide, carrier molecule and carrier peptide may be a penetration enhancer that does not damage, destroy and / or degrade skin structure. In some embodiments, the copeptide, carrier molecule and carrier peptide may be a penetration enhancer that does not irritate the skin. The term "penetration enhancer" does not include mechanical devices (e.g., needles, scalpels, etc.) or equivalents thereof (e.g., other damaging procedures).Additionally, one of skill in the art will understand that structures such as nanoparticles or emulsions are not chemical agents and therefore are not chemical penetration enhancers, even if their presence correlates with improved skin penetration of a desired drug that may be associated with the structure.

[0074] Pharmaceutical composition: As used herein, the term "pharmaceutical composition" refers to a composition in which an active agent is formulated together with one or more pharma- ceutical acceptable carriers. In some embodiments, the active agent is present in a unit dose suitable for administration in a treatment regimen that shows a statistically significant probability of achieving a predetermined therapeutic effect when administered to a relevant population. In some embodiments, the pharmaceutical composition can be specially formulated for administration in solid or liquid form, including those adapted for local administration, such as gel, cream, ointment or controlled release patch or spray applied to the skin, lung or oral cavity; for example, as pessary, cream or foam applied to the vagina or rectum; sublingual; intraocular; transdermal; or applied to the nasal, pulmonary and other mucosal surfaces, sterile liquid or suspension or sustained release formulation.

[0075] Pharmaceutically acceptable: As used herein, the term "pharmaceutical acceptable" applied to carriers, diluents, or excipients used to formulate the compositions described herein, means that the carrier, diluent, or excipient must be compatible with the other ingredients of the composition and not deleterious to the recipient thereof.

[0076] Pharmaceutically acceptable carrier: As used herein, the term "pharmaceutical acceptable carrier" refers to a pharma- ceutically acceptable material, composition, or vehicle, such as a liquid or solid excipient, diluent, additive, or solvent that encapsulates a material, involved in carrying or transporting a compound of interest from one organ or part of the body to another. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not harmful to the subject or patient. Some examples of substances which may serve as pharma- ceutically acceptable carriers include sugars such as lactose, glucose and sucrose; starches such as corn starch and potato starch; cellulose and its derivatives such as sodium carboxymethylcellulose, ethylcellulose and cellulose acetate; powdered tragacanth; malt; gelatin; talc; additives such as cocoa butter and suppository wax; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, medium chain triglycerides and soybean oil; glycols such as propylene glycol; polyols such as glycerin, sorbitol, mannitol and polyethylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffers such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol; pH buffers; polyesters, polycarbonates and / or polyanhydrides; and other non-toxic, compatible substances used in pharmaceutical formulations.

[0077] Premix: As used herein, the term "premix" refers to any combination of ingredients that are subsequently used to generate a nanoemulsion composition or in accordance with the present invention. For example, a premix is ​​any collection of ingredients that, when subjected to high shear forces, generate a nanoemulsion in accordance with the present invention. In some embodiments, a premix is ​​a collection of ingredients that, when subjected to high shear forces, generate a nanoemulsion composition, such as a uniform nanoemulsion composition. A premix often contains a liquid dispersion medium and sufficient other ingredients to generate a nanoemulsion within the dispersion medium. According to some embodiments of the present disclosure, one or more large drugs may be included in the premix. According to some embodiments of the present disclosure, one or more biological drugs may be included in the premix. According to the present invention, a botulinum toxin may be included in the premix. According to the present invention, one or more antibodies may be included in the premix. In some embodiments, a premix may include one or more surfactants, penetration enhancers, and / or other substances. In some embodiments, a premix comprises a solution. In some embodiments in which the premix comprises a botulinum toxin, an antibody, another biologically active agent, and / or a penetration enhancer, the botulinum toxin, the antibody, another biologically active agent, and / or the penetration enhancer are in solution prior to applying high shear forces to the premix.

[0078] Prevent or prophylaxis: As used herein, when used in relation to the occurrence of a disease, disorder, and / or condition, prevent or prophylaxis refers to reducing the risk of developing a disease, disorder, and / or condition and / or delaying the onset of one or more characteristics or symptoms of a disease, disorder, or condition. When the onset of a disease, disorder, or condition has been delayed for a certain period of time, prevention may be considered complete.

[0079] Protein: As used herein, the term "protein" refers to a polypeptide (i.e., a string of at least two amino acids linked together by peptide bonds). Proteins may contain moieties other than amino acids (e.g., glycoproteins, proteoglycans, etc.) and / or may be otherwise processed or modified. Those skilled in the art will understand that a "protein" may be a complete polypeptide chain as produced by a cell (with or without a signal sequence) or a characteristic portion thereof. Those skilled in the art will understand that a protein may contain multiple polypeptide chains, for example, linked by one or more disulfide bonds or linked by other means. Polypeptides may contain L-amino acids, D-amino acids, or both, and may contain any of a variety of amino acid modifications or analogs known in the art. Useful modifications include, for example, terminal acetylation, amidation, methylation, etc. In some embodiments, proteins may contain natural amino acids, unnatural amino acids, synthetic amino acids, and combinations thereof. The term "peptide" is generally used to refer to polypeptides having a length of less than about 100 amino acids, less than about 50 amino acids, less than 20 amino acids, or less than 10 amino acids. In some embodiments, the protein is an antibody, an antibody fragment, a biologically active portion thereof, and / or a characteristic portion thereof.

[0080] Polypeptide: As used herein, the term "polypeptide" generally has its art-recognized meaning of a polymer of at least three amino acids. Those of skill in the art will appreciate that the term "polypeptide" is intended to encompass not only polypeptides having the complete sequences described herein, but also polypeptides that represent functional fragments of such complete polypeptides (i.e., fragments that retain at least one activity). Moreover, those of skill in the art will appreciate that protein sequences generally tolerate some substitutions without destroying activity. Thus, polypeptides that retain activity and share at least 30-40%, often more than 50%, 60%, 70% or 80% overall sequence identity with polypeptides of the same class, and further include at least one region of much higher identity, often more than 90%, and even more than 95%, 96%, 97%, 98% or 99%, in one or more highly conserved regions, usually comprising at least 3-4, and often 20 or more amino acids, are included in the related term "polypeptide" as used herein. Polypeptides may contain L-amino acids, D-amino acids, or both, and may contain any of a variety of amino acid modifications or analogs known in the art. Useful modifications include, for example, terminal acetylation, amidation, methylation, and the like. In some embodiments, proteins may contain natural amino acids, unnatural amino acids, synthetic amino acids, and combinations thereof. The term "peptide" is generally used to refer to a polypeptide having a length of less than about 100 amino acids, less than about 50 amino acids, less than 20 amino acids, or less than 10 amino acids. In some embodiments, the protein is an antibody, an antibody fragment, a biologically active portion thereof, and / or a characteristic portion thereof.

[0081] Reference: As used herein, reference describes a standard or control against which comparison is performed. For example, in some embodiments, a drug, animal, individual, population, sample, regimen, sequence or value of interest is compared to a drug, animal, individual, population, sample, regimen, sequence or value of a reference or control. In some embodiments, the reference or control is tested and / or determined substantially simultaneously with the test or determination of interest. In some embodiments, the reference or control is a historical reference or control, and is optionally embodied in a tangible medium. Typically, as understood by those skilled in the art, a reference or control is determined or characterized under conditions or circumstances equivalent to those being evaluated. Those skilled in the art will understand when there is sufficient similarity to determine reliance and / or comparison to a particular possible reference or control.

[0082] Self-administration: As used herein, the term "self-administration" refers to the situation in which a subject has the ability to administer a composition to himself or herself without the need for medical supervision.In some embodiments of the present invention, self-administration can be performed outside of a clinical setting.For example, in some embodiments of the present invention, a facial cosmetic cream can be administered by a subject in his or her own home.

[0083] Small molecule: Generally, a "small molecule" is understood in the art to be an organic molecule that is less than about 5 Daltons (Kd) in size. In some embodiments, the small molecule is less than about 3Kd, 2Kd, or 1Kd. In some embodiments, the small molecule is less than about 800 Daltons (D), 600D, 500D, 400D, 300D, 200D, or 100D. In some embodiments, the small molecule is non-polymeric. In some embodiments, the small molecule is not a protein, peptide, or amino acid. In some embodiments, the small molecule is not a nucleic acid or nucleotide. In some embodiments, the small molecule is not a saccharide or polysaccharide.

[0084] Subject: As used herein, "subject" refers to an organism, typically a mammal (e.g., a human, including in some embodiments prenatal human forms). In some embodiments, the subject is suffering from the relevant disease, disorder or condition. In some embodiments, the subject is susceptible to the disease, disorder or condition. In some embodiments, the subject exhibits one or more symptoms or characteristics of the disease, disorder or condition. In some embodiments, the subject does not exhibit any symptoms or characteristics of the disease, disorder or condition. In some embodiments, the subject is a person who has one or more characteristics characteristic of susceptibility or risk for a disease, disorder, or condition. In some embodiments, the subject is a patient. In some embodiments, the subject is an individual who is being and / or has been diagnosed and / or treated.

[0085] Substantially: As used herein, the term "substantially" refers to the qualitative condition of the total or near total extent or degree of a desired feature or characteristic. Those skilled in the art of biology understand that biological and chemical phenomena rarely, if ever, proceed to completion and / or perfection or achieve or avoid absolute results. Thus, the term "substantially" is used herein to capture the potential lack of completeness inherent in many biological and chemical phenomena.

[0086] Therapeutic Agent: As used herein, the term "therapeutic agent" generally refers to any drug that induces a desired pharmacological effect when administered to an organism. In some embodiments, a drug is considered to be a therapeutic agent if it shows a statistically significant effect across an appropriate population. In some embodiments, the appropriate population can be a population of model organisms. In some embodiments, the appropriate population can be defined by various criteria, such as a specific age group, sex, genetic background, pre-existing clinical conditions, etc. In some embodiments, a therapeutic agent is a substance that can be used to alleviate, ameliorate, mitigate, prevent, delay the onset of, reduce the severity of, and / or reduce the incidence of one or more symptoms or characteristics of a disease, disorder, and / or condition. In some embodiments, a "therapeutic agent" is a drug that has been approved or needs to be approved by a government agency before it can be sold for administration to humans. In some embodiments, a "therapeutic agent" is a drug that requires a prescription for administration to humans. In some embodiments, a drug is not considered to be a "therapeutic agent" if it merely enhances the delivery of a different drug that actually achieves a desired effect.

[0087] Therapeutically effective amount: as used herein, a therapeutically effective amount refers to an amount that produces the desired effect for which it is administered. In some embodiments, the term refers to an amount that is sufficient when administered to a population suffering from or susceptible to a disease, disorder, and / or condition according to a therapeutic administration regimen to treat the disease, disorder, and / or condition. In some embodiments, a therapeutically effective amount is one that reduces the incidence and / or severity and / or delays the onset of one or more symptoms of a disease, disorder, and / or condition. Those skilled in the art will appreciate that the term "therapeutically effective amount" does not actually require that successful treatment be achieved in a particular individual. Rather, a therapeutically effective amount may be an amount that provides a particular desired pharmacological response in a significant number of subjects when administered to a patient in need of such treatment. In some embodiments, a reference to a therapeutically effective amount may be a reference to an amount measured in one or more specific tissues (e.g., tissues affected by a disease, disorder, or condition) or bodily fluids (e.g., blood, saliva, serum, sweat, tears, urine, etc.). Those skilled in the art will appreciate that in some embodiments, a therapeutically effective amount of a particular drug or therapy may be formulated and / or administered in a single dose. In some embodiments, a therapeutically effective drug may be formulated and / or administered in multiple doses, for example as part of a dosing regimen.

[0088] Therapeutic regimen: As used herein, the term "therapeutic regimen" refers to a dosing regimen whose administration across a relevant population can be correlated with a desired or beneficial therapeutic outcome.

[0089] Treatment: As used herein, the term "treatment" ("treat" or "treating") refers to any administration of a therapy that partially or completely relieves, improves, alleviates, delays the onset of, reduces the severity of, and / or reduces the incidence of one or more symptoms, characteristics, and / or causes of a particular disease, disorder, and / or condition. In some embodiments, such treatment may be of subjects who do not show signs of the associated disease, disorder, and / or condition, and / or of subjects who show only early signs of the disease, disorder, and / or condition. Alternatively or additionally, such treatment may be of subjects who show one or more established signs of the associated disease, disorder, and / or condition. In some embodiments, treatment may be of subjects who have been diagnosed as suffering from the associated disease, disorder, and / or condition. In some embodiments, treatment may be of subjects who are known to have one or more susceptibility factors that are statistically correlated with an increased risk of developing the associated disease, disorder, and / or condition.

[0090] Uniform: The term "uniform" as used herein with respect to nanoemulsion compositions refers to nanoemulsion compositions in which the individual droplets have a particular range of droplet diameter sizes. For example, in some embodiments, a uniform nanoemulsion composition is one in which the difference between the smallest diameter and the largest diameter is no more than about 300, 250, 200, 150, 100, 90, 80, 70, 60, 50 nm, or less. In some embodiments, the droplets (e.g., large drug-containing droplets) within the uniform large drug nanoemulsion compositions of the invention have diameters less than about 300, 250, 200, 150, 130, 120, 115, 110, 100, 90, 80 nm, or less. In some embodiments, the droplets (e.g., large drug-containing droplets) within the uniform large drug nanoemulsion compositions of the invention have diameters within the range of about 10 nm to about 300 nm. In some embodiments, the droplets in the large uniform drug nanoemulsion compositions of the invention have diameters in the range of about 10-300, 10-200, 10-150, 10-130, 10-120, 10-115, 10-110, 10-100, or 10-90 nm. In some embodiments, the droplets (e.g., large drug-containing droplets) in the large drug nanoemulsion compositions of the invention have an average droplet size of less than about 300, 250, 200, 150, 130, 120, 115, 110, 100, or 90 nm. In some embodiments, the average droplet size is in the range of about 10-300, 50-250, 60-200, 65-150, 70-130 nm. In some embodiments, the average droplet size is about 80-110 nm. In some embodiments, the average droplet size is about 90-100 nm. In some embodiments, a majority of the droplets (e.g., large drug-containing droplets) within a homogeneous nanoemulsion composition of the invention have a diameter less than a specified size or within a specified range, hi some embodiments, the majority is 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9% or more of the droplets in the composition.In some embodiments of the present invention, a homogenous nanoemulsion composition is achieved by microfluidization of the sample.

[0091] Variant: As used herein, the term "variant" refers to an entity that exhibits significant structural identity with a reference entity, but is structurally different from the reference entity in the presence or level of one or more chemical moieties compared to the reference entity. In many embodiments, variants are also functionally different from the reference entity. In general, whether a particular entity is properly considered to be a "variant" of a reference entity is based on the degree of structural identity with the reference entity. As will be understood by those skilled in the art, every biological or chemical reference entity has certain characteristic structural elements. Variants are, by definition, distinct chemical entities that share one or more such characteristic structural elements. To give some examples, small molecules can have a characteristic core structural element (e.g., a macrocyclic core) and / or one or more characteristic pendant moieties, such that variants of the small molecule share the core structural element and the characteristic pendant moieties but differ in other pendant moieties and / or bond types within the core (single vs. double, E vs. Z, etc.), polypeptides can have characteristic sequence elements composed of multiple amino acids that have designated positions relative to each other in linear or three-dimensional space and / or that contribute to a particular biological function, and nucleic acids can have characteristic sequence elements composed of multiple nucleotide residues that have designated positions relative to each other in linear or three-dimensional space. For example, variant polypeptides can differ from a reference polypeptide as a result of one or more differences in amino acid sequence and / or one or more differences in chemical moieties (e.g., carbohydrates, lipids, etc.) that are covalently attached to the polypeptide backbone. In some embodiments, the mutant polypeptide shows an overall sequence identity with the reference polypeptide that is at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97% or 99%, optionally excluding conservative amino acid substitutions.Alternatively or in addition, in some embodiments, the mutant polypeptide does not share at least one characteristic sequence element with the reference polypeptide.In some embodiments, the reference polypeptide has one or more biological activities.In some embodiments, the mutant polypeptide shares one or more biological activities of the reference polypeptide.In some embodiments, a mutant polypeptide lacks one or more biological activities of a reference polypeptide. In some embodiments, a mutant polypeptide exhibits a reduced level of one or more biological activities compared to a reference polypeptide. In many embodiments, a target polypeptide is considered to be a "variant" of a parent or reference polypeptide when the target polypeptide has an amino acid sequence identical to that of the parent but with a small number of sequence changes at certain positions. Typically, less than 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2% of the residues of the mutant are substituted compared to the parent. In some embodiments, a mutant has 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1 substituted residues compared to the parent. In many cases, a mutant has a very small number (e.g., less than 5, 4, 3, 2 or 1) of functional residues (i.e., residues involved in a particular biological activity) substituted. Furthermore, a mutant typically has no more than 5, 4, 3, 2 or 1 additions or deletions, and often no additions or deletions compared to the parent. Moreover, any additions or deletions are typically less than about 25, about 20, about 19, about 18, about 17, about 16, about 15, about 14, about 13, about 10, about 9, about 8, about 7, about 6, and generally less than about 5, about 4, about 3 or about 2 residues. In some embodiments, the parent or reference polypeptide is one found in nature.

[0092] DETAILED DESCRIPTION OF SPECIFIC EMBODIMENTS Transdermal Drug Delivery In some embodiments, the present invention provides a technology for improving the transdermal delivery and / or bioavailability of large drugs (e.g., botulinum toxin, antibodies). In some embodiments, the present disclosure teaches that particularly advantageous results are achieved when microneedling technology is combined with an emulsion composition. In some embodiments, microneedling technology is combined with a lotion, cream or liquid composition, which in turn can be or can include an emulsion composition (e.g., a macroemulsion composition and / or a nanoemulsion composition). In some embodiments, the provided technology does not utilize a penetration enhancer. In some embodiments, the provided technology does not utilize a chemical penetration enhancer that damages, destroys and / or breaks down the skin. In some embodiments, the provided technology does not utilize a chemical penetration enhancer.

[0093] Human skin comprises the dermis and the epidermis. The epidermis has several layers of tissue: the stratum corneum, the stratum lucidum, the stratum granulosum, the stratum spinosum, and the stratum basale (identified from the outer surface of the skin inwards).

[0094] The stratum corneum presents the greatest hurdle in transdermal delivery in general, and perhaps in particular, of large drugs. It is typically about 10-15 μm thick and consists of flattened keratinized cells (keratinocytes) arranged in several layers. The spaces between the keratinocytes are filled with lipid structures, which may play an important role in the penetration of substances through the skin (Bauerova et al., 2001, European Journal of Drug Metabolism and Pharmacokinetics, 26:85).

[0095] The remaining epidermis, below the stratum corneum, is about 150 μm thick. The dermis, about 1-2 mm thick, lies below the epidermis. The dermis is innervated by various capillaries and nerve processes.

[0096] Transdermal administration has been the subject of research to provide an alternative route of administration that does not have the undesirable consequences commonly associated with injections and oral delivery. For example, needles often cause local pain and can expose the patient receiving the injection to blood-borne diseases. Oral administration can suffer from poor bioavailability of pharmaceuticals due to the extremely acidic environment of the patient's stomach.

[0097] In an attempt to overcome these shortcomings by providing non-invasive administration, efforts have been made to develop transdermal administration techniques for certain pharmaceuticals. In general, it is desirable for transdermal administration to minimize damage to the patient's skin. Thus, transdermal administration can reduce or eliminate the pain associated with injections, reduce the possibility of blood contamination, and improve the bioavailability of the drug when incorporated into the systemic environment.

[0098] Traditionally, attempts at transdermal administration have focused on the destruction and / or decomposition of the stratum corneum. Some attempts include the use of chemical penetration enhancers. Penetration enhancers can function to decompose and / or destroy skin structure. In some embodiments, the penetration enhancer is or includes a chemical agent (e.g., a chemical or enzyme that can destroy and / or decompose one or more components of the stratum corneum). In some embodiments, the penetration enhancer can be irritating in that it causes inflammatory and / or allergic reactions when the drug is applied to the skin.

[0099] "However, the main limitation of penetration enhancers is that their efficacy is often closely correlated with the occurrence of skin irritation." Alkilani, AZ, et al., "Transdermal drug delivery: Innovative pharmaceutical developments based on disruption of the barrier properties of the stratum corneum." Pharmaceutics. 7:438-470 (2015). Penetration enhancers tend to have poor efficacy and safety profiles. "They do not achieve the desired skin breakdown and their ability to increase transport across the skin is low and variable." Ibid.

[0100] Some attempts have included the use of mechanical devices to bypass or ablate portions of the stratum corneum. Others have included the use of ultrasound or iontophoresis to facilitate the penetration of pharmaceuticals through the skin. In most cases, the goal is to allow drugs, typically small molecules, to pass into the capillary bed of the dermis, where they can be systemically incorporated into the subject to achieve a therapeutic effect. These methods are limited by the amount of energy that can be applied to the skin without causing discomfort and / or skin damage.

[0101] Transdermal Delivery of Macromolecules Microneedling technology has been shown to improve transdermal delivery of a variety of small drugs, such as calcein (about 623 Da), desmopressin (about 1070 Da), diclofenac (about 270 Da), methyl nicotinate (about 40 Da), bischloroethylnitrosourea (about 214 Da), insulin (about 5.8 kDa), bovine serum albumin (about 66.5 kDa) and ovalbumin (about 45 kDa), but until the present disclosure improving the delivery and / or bioavailability of larger drugs, especially those above 100 kDa, remained problematic.

[0102] It is recognized that the transdermal delivery of large molecules brings about large challenges.Until the present disclosure, microneedling, particularly microneedle skin preconditioning using a relatively low microneedle density and / or a relatively small microneedle puncture size (e.g., puncture size per microneedle), was not thought to affect or be effective in the transdermal administration of large drugs.For example, a study of the use of solid microneedles for the delivery of four hydrophilic peptides, low molecular weight tetrapeptide-3 (456.6 Da); hexapeptide (498.6 Da); acetyl hexapeptide-3 (889 Da); and oxytocin (1007.2 Da), as well as L-carnitine (161.2 Da), showed that while microneedle pretreatment significantly increases the penetration of each of these peptides, the skin penetration of peptides depends on their molecular weight and decreases as molecular weight increases. Zhang, S., et al., "Enhanced delivery of hydrophilic peptides in vitro by transdermal microneedle pretreatment." Acta Pharmaceutica Sinica B. 4(1):100-104 (2014).

[0103] When sandpaper abrasion, tape stripping and single puncture subcutaneous needle models of MSCs were compared in a study of the effect of molecular size of larger FITC (fluorescein isothiocyanate) conjugated molecules on transdermal delivery, all methods were found to be similar and, when tested on untreated skin, transdermal drug delivery was again shown to decrease as the size of the tested molecules increased (4.3, 9.6 and 42.0 KDa FITC conjugates). Tape stripping was found to be the most effective technique, while sandpaper abrasion was found to be the most damaging to the skin. Wu, X., et al., "Effects of pretreatment of needle puncture and sandpaper abrasion on the in vitro skin permeation of fluorescein isothiocyanate (FITC)-dextran." International Journal of Pharmaceutics. 316:102-108 (2006).

[0104] Other studies have attempted to deliver even larger molecules: Cascade Blue (CB, MW 538), Dextran-Cascade Blue (DCB, MW 10 kDa) and FITC-conjugated dextran (FITC-Dex, MW 72 kDa). In those studies, microneedles of various lengths (300, 550, 700 or 900 μm) were used to puncture lamina human skin and assess the diffusion of each of the above compounds. Transport of each compound was seen with all but the 300 μm microneedle array, although degradation of DCB and FITC-Dex was observed.

[0105] As the prior art shows, as molecular size increases, transdermal penetration using MSCs ("microneedle skin conditioning") decreases to the point where it becomes minimal or even non-existent. Even when some minimal penetration is observed, larger molecules are observed to degrade and become biologically inactive. Recent techniques (see, for example, International Application No. PCT / US17 / 53333) have been developed that achieve various advantages by combining emulsion and microneedling techniques for transdermal delivery of large drugs of interest; in some embodiments, these techniques have shown that they can achieve particularly surprising improvements in transdermal delivery of large molecular structures without the use of mechanical or chemical penetration enhancing means. For example, in some embodiments, these techniques have achieved transdermal delivery of Clostridium botulinum, which is about 150 KDa and more than twice the size of FITC-Dex. The present disclosure discloses that the microneedling techniques described herein can surprisingly further enhance the transdermal delivery and / or improve the bioavailability of large drugs (e.g., those with molecular weights greater than 100 kDa (e.g., botulinum). Clostridium botulinum is a complex protein, and three regions or functional parts must be intact for the protein to be biologically active. Thus, damage to any one of the three regions of the protein renders the protein biologically inactive. According to Johnson, E. et al., "Botulinum toxin is highly susceptible to denaturation due to surface denaturation, heat and alkaline conditions." U.S. Patent Publication No. 5,512,547. Thus, a significant level of degradation and inactivation of Clostridium botulinum is expected under the microneedling conditions described by Wu.

[0106] In particular, the present disclosure discloses that the microneedling techniques described herein can enhance transdermal delivery and / or improve the bioavailability of (e.g., of large drugs, particularly from macroemulsion or nanoemulsion compositions) when not utilizing other penetration enhancers, particularly disruptive agents (i.e., chemical penetration enhancers and other techniques that disrupt or puncture skin structures).

[0107] Microneedling The present disclosure provides a surprising discovery that the MSCs described herein can surprisingly improve the transdermal delivery of large drugs. In some embodiments, the large drugs can be formulated as creams and / or lotions. In some embodiments, the large drugs can be combined with one or more biologically active substances. In some embodiments, the large drugs can be formulated as or in emulsion (e.g., macroemulsion or nanoemulsion) compositions. In some embodiments, the emulsions containing one or more large drugs can be formulated as creams and / or lotions.

[0108] In some embodiments, the microneedle (MN) array for use according to the present disclosure features or shares the features of a minimally invasive system developed to overcome some of the drawbacks commonly associated with the use of hypodermic and subcutaneous needles and improve patient comfort and compliance. Such drawbacks include, for example, the possibility of misplacement of the needle tip with hypodermic needles, as medical professionals cannot visualize exactly where the needle is going; such needle misplacement can cause adverse reactions, such as drooping eyelids ("ptosis"), when botulinum is injected inaccurately into the face. MNs are less prone to such problems. Other advantages of MNs are that they do not cause bleeding, minimize the introduction of pathogens through MN-created holes, and eliminate variability in transdermal administration. Other advantages are the possibility of self-administration, reduced risk of accidental needlestick injury, reduced risk of infectious disease transmission, and ease of disposal. In some embodiments, the MNs are a plurality of microscopic projections assembled on one side of a support, such as a patch or device (eg, a stamp, roller, array, applicator, pen).

[0109] In some embodiments, MNs for use according to the present disclosure can be designed and / or constructed in an array to improve skin contact and promote skin penetration. In some embodiments, the MNs utilized are of suitable length, width and shape to create an efficient pathway for drug delivery while minimizing contact with nerves when inserted into the skin. Alkilani, AZ, et al., "Transdermal drug delivery: Innovative pharmaceutical developments based on disruption of the barrier properties of the stratum corneum." Pharmaceutics. 7:438-470 (2015).

[0110] In some embodiments, suitable MNs can be solid, coated, porous, dissolving, hollow or hydrogel MNs. Solid MNs create microscopic holes in the skin, thereby increasing the transport of drug formulations (e.g., the "poke and patch" method). Coated MNs allow for rapid elution of coated drugs into the skin (e.g., the "coat and poke" method). Dissolving MNs allow for rapid and / or controlled release of drugs incorporated within the microneedles. Hollow MNs can be used to puncture the skin and allow for the release of the composition after active injection or diffusion of the formulation through the pores of the microneedles (e.g., the "poke and flow" method). In the case of dissolving MNs, the MNs can function as drug depots, holding the drug composition until released by dissolution in the case of dissolving MNs, or swelling in the case of hydrogel MNs (e.g., the "poke and release" method). However, as previously described herein, in many embodiments, large drugs are not delivered by injection through one or more microneedles. That is, in many embodiments, any microneedles utilized according to such embodiments are not coated with, loaded with, or manufactured with large drugs in any manner that achieves delivery of large drugs. Alternatively, in some embodiments, MNs described herein (in MSCs or otherwise) utilized according to the present disclosure can contain and / or deliver large drugs when the large drugs are formulated into macro- or nanoemulsions described herein. Thus, as will be understood by those skilled in the art upon reading this specification, treatment of skin with microneedles that deliver large drugs (e.g., by injection through the microneedles, by release of microneedle coatings, or by release from dissolved microneedles) is not microneedle skin conditioning.

[0111] In some embodiments, the microneedle has a diameter that is constant throughout the length of the microneedle. In some embodiments, the diameter of the microneedle is greatest at the base end of the microneedle. In some embodiments, the microneedle tapers to a point at the end distal to the base of the microneedle. In some embodiments, the microneedle can be solid. In some embodiments, the microneedle can be hollow. In some embodiments, the microneedle can be tubular. In some embodiments, one end of the microneedle can be sealed. In some embodiments, the microneedle is part of an array of microneedles. In some embodiments, the microneedle can be about 1 μm to about 4,000 μm long. In some embodiments, the microneedle can be about 1 μm to about 2,000 μm long. In some embodiments, the microneedle can be about 50 μm to about 400 μm long. In some embodiments, the microneedle can be about 50 μm to about 500 μm long. In some embodiments, the microneedle can be about 50 μm to about 600 μm long. In some embodiments, the microneedles may be from about 50 μm to about 700 μm in length. In some embodiments, the microneedles may be from about 50 μm to about 800 μm in length. In some embodiments, the microneedles may be from about 800 μm to about 1500 μm in length. In some embodiments, the microneedles may be less than about 1400 μm in length. In some embodiments, the microneedles may be less than about 1100 μm in length. In some embodiments, the microneedles may be less than about 1000 μm in length. In some embodiments, the microneedles may be less than about 800 μm in length. In some embodiments, the microneedles may be from about 100 μm to about 800 μm in length.

[0112] In some embodiments, the microneedling described herein comprises applying to the skin multiple microneedles of a common length (e.g., a microneedle array); the microneedling described herein comprises applying to the skin multiple microneedles of different lengths (e.g., a microneedle array).

[0113] Microneedles of various lengths may be used in the microneedling techniques described herein. In some embodiments, the length of the microneedles used in the MSCs described herein is adjusted based on the thickness of the skin at the treatment site. In some embodiments, the MN or MN array comprises microneedles of about 100 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 150 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 200 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 250 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 300 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 350 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 400 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 450 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 500 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 550 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 600 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 650 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 700 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 750 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 800 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 850 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 900 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 950 μm in length.In some embodiments, the MN or MN array comprises microneedles of about 1000 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 1100 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 1200 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 1300 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 1400 μm in length. In some embodiments, the MN or MN array comprises microneedles of about 1500 μm in length.

[0114] In some embodiments, the MN or MN array comprises a plurality of needles. In some embodiments, the MN or MN array comprises more than 2 microneedles / cm 2 In some embodiments, the MN or MN array comprises 3 microneedles / cm 2 In some embodiments, the MN or MN array comprises 4 microneedles / cm 2 In some embodiments, the MN or MN array comprises 5 microneedles / cm 2 In some embodiments, the MN or MN array comprises 6 microneedles / cm 2 In some embodiments, the MN or MN array comprises 7 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 8 microneedles / cm 2 In some embodiments, the MN or MN array comprises 9 microneedles / cm 2 In some embodiments, the MN or MN array comprises 10 microneedles / cm 2 In some embodiments, the MN or MN array comprises 11 microneedles / cm 2 In some embodiments, the MN or MN array comprises 12 microneedles / cm 2In some embodiments, the MNs or MN arrays comprise 13 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 14 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 15 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 16 microneedles / cm 2 In some embodiments, the MN or MN array comprises 17 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 18 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 19 microneedles / cm 2 In some embodiments, the MN or MN array comprises 20 microneedles / cm 2 In some embodiments, the MN or MN array comprises 21 microneedles / cm 2 In some embodiments, the MN or MN array comprises 22 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 23 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 24 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 25 microneedles / cm 2 In some embodiments, the MN or MN array comprises 26 microneedles / cm 2 In some embodiments, the MN or MN array comprises 27 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 28 microneedles / cm 2 In some embodiments, the MN or MN array comprises 29 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 30 microneedles / cm2 In some embodiments, the MN or MN array comprises 31 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 35 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 40 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 45 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 50 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 55 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 60 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 65 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 70 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 75 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 80 microneedles / cm 2 In some embodiments, the MN or MN array comprises 85 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 90 microneedles / cm 2 In some embodiments, the MN or MN array comprises 95 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 100 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 200 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 300 microneedles / cm 2In some embodiments, the MNs or MN arrays comprise 400 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 500 microneedles / cm 2 In some embodiments, the MNs or MN arrays comprise 1000 microneedles / cm 2 In some embodiments, the MN or MN array comprises less than 2000 microneedles / cm. 2 Includes less than.

[0115] Microneedles of any shape may be used in the microneedling techniques described herein. In some embodiments, the microneedles may have a circular cross-section. In some embodiments, the microneedles may have a triangular cross-section. In some embodiments, the microneedles may have a rectangular cross-section. In some embodiments, the microneedles may have a square cross-section. In some embodiments, the microneedles may have a quadrangular cross-section. In some embodiments, the microneedles may have a pentagonal cross-section. In some embodiments, the microneedles may have a hexagonal cross-section. In some embodiments, the microneedles may have a heptagonal cross-section. In some embodiments, the microneedles may have an octagonal cross-section. In some embodiments, the microneedles may have a nonagonal cross-section. In some embodiments, the microneedles may have a decagonal cross-section.

[0116] Microneedles of various cross-sectional areas can be used in the microneedling techniques described herein. The cross-sectional area of ​​each microneedle in the MN array used for MSCs ("microneedle skin conditioning") described herein can then define the puncture size (e.g., puncture size per microneedle) of the microneedle in the MN array used for MSCs. In some embodiments, the puncture size of the microneedle is about 100 to about 60,000 μm 2In some embodiments, the puncture size of the microneedle may range from about 100 to about 30,000 μm. 2 / Microneedle range.

[0117] In some embodiments, the MN or MN array comprises needles having a plurality of microneedle puncture sizes. In some embodiments, the MN or MN array comprises needles having at least two different microneedle puncture sizes. In some embodiments, the MN or MN array comprises needles having at least three different microneedle puncture sizes. In some embodiments, the MN or MN array comprises needles having at least four different microneedle puncture sizes. In some embodiments, the MN or MN array comprises needles having at least five different microneedle puncture sizes. In some embodiments, the MN or MN array comprises needles having at most ten different microneedle puncture sizes. In some embodiments, the MN or MN array comprises needles having at least eleven different microneedle puncture sizes. In some embodiments, the MN or MN array comprises needles having at least twelve different microneedle puncture sizes. In some embodiments, the MN or MN array comprises needles having at most one microneedle puncture size.

[0118] MNs or MN arrays containing various microneedle puncture sizes can be used in the microneedling techniques described herein. In some embodiments, the MNs or MN arrays are 100 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 200 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 300 μm. 2In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 400 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 500 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 600 μm. 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 700 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 800 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 900 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 1000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 1100 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 1200 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 1300 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 1400 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 1500 μm2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 1600 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 1700 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 1800 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 1900 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 2000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 2500 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 3000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 3500 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 4000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 4500 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 5000 μm 2In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 5500 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 6000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 6500 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 7000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 7500 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 8000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 8500 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 9000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 9500 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 10,000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 10,500 μm 2In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 11000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 11,500 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 12000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a microneedle puncture size of 13,000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a puncture size of less than 14000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a puncture size of less than 15,000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a puncture size of less than 20,000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a puncture size of less than 25,000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a puncture size of less than 30,000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a puncture size of less than 35,000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a puncture size of less than 40,000 μm 2In some embodiments, the MN or MN array comprises a microneedle having a puncture size of less than 45,000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a puncture size of less than 50,000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a puncture size of less than 55,000 μm 2 In some embodiments, the MN or MN array comprises a microneedle having a puncture size of less than 60,000 μm 2 / Microneedles having a puncture size smaller than a microneedle.

[0119] In some embodiments, MNs for use according to the present disclosure may be manufactured from different materials using techniques including, but not limited to, micromolding processes or lasers. In some embodiments, MNs may be manufactured using various types of biocompatible materials, including polymers, metals, ceramics, semiconductors, organics, composites, or silicon. Unless they are designed to penetrate and dissolve in the skin, in some embodiments, microneedles have the mechanical strength to remain intact and deliver drugs or collect biological fluids while being inserted into the skin and / or removed from the skin after insertion. In some embodiments, MNs can remain in place for up to several days before being completely removed. In some embodiments, microneedles can be sterilizable using standard techniques. In some embodiments, MNs are biodegradable. In some embodiments, MNs comprise polymeric materials. In some embodiments, the polymeric material comprises poly-L-lactic acid, poly-glycolic acid, polycarbonate, polylactic-co-glycolic acid (PLGA), polydimethylsiloxane, polyvinylpyrrolidone (PVP), copolymers of methyl vinyl ether and maleic anhydride, sodium hyaluronate, carboxymethylcellulose, maltose, dextrin, galactose, starch, gelatin, or combinations thereof.

[0120] In some embodiments, the MSCs described herein comprise one press of MN or MN arrays. In some embodiments, the MSCs comprise two presses of MN or MN arrays. In some embodiments, the MSCs comprise three presses of MN or MN arrays. In some embodiments, the MSCs comprise four presses of MN or MN arrays. In some embodiments, the MSCs comprise five presses of MN or MN arrays. In some embodiments, the MSCs comprise six presses of MN or MN arrays. In some embodiments, the MSCs comprise seven presses of MN or MN arrays. In some embodiments, the MSCs comprise eight presses of MN or MN arrays. In some embodiments, the MSCs comprise nine presses of MN or MN arrays. In some embodiments, the MSCs comprise ten presses of MN or MN arrays. In some embodiments, the MSCs comprise eleven presses of MN or MN arrays. In some embodiments, the MSCs comprise twelve presses of MN or MN arrays. In some embodiments, the MSCs comprise thirteen presses of MN or MN arrays. In some embodiments, the MSCs comprise 14 presses of the MN or MN array. In some embodiments, the MSCs comprise 15 presses of the MN or MN array. In some embodiments, the MSCs comprise 16 presses of the MN or MN array. In some embodiments, the MSCs comprise 17 presses of the MN or MN array. In some embodiments, the MSCs comprise 18 presses of the MN or MN array. In some embodiments, the MSCs comprise 19 presses of the MN or MN array. In some embodiments, the MSCs comprise 20 presses of the MN or MN array. In some embodiments, the MSCs comprise rolling the MN or MN array with the skin tablet one or more times. In some embodiments, the MN array is rotated between presses. In some embodiments, the MN array is not rotated between presses. In some embodiments, the presses are performed at the same site. In some embodiments, the presses are performed at overlapping sites.In some embodiments, the pressing is performed on different sites. In some embodiments, the pressing is performed by stamping the MN array. In some embodiments, the pressing is performed by rolling a microneedle roller on the site one or more times. In accordance with established MN practice, in some embodiments, the pressing of the MN array on the skin can last for less than 1 second, or in some embodiments, for 1 second or more, such as 30 seconds or more, 60 seconds or more, 2 minutes or more, 5 minutes or more, 10 minutes or more, 30 minutes or more, etc.

[0121] As mentioned above, the present disclosure teaches that the bioavailability of large drugs in emulsions applied to skin increases as the total surface area of ​​skin pierced by microneedles decreases.Therefore, in some embodiments, a relatively smaller pressing force may be preferred.In some embodiments, when the large drug administered in conjunction with microneedle skin conditioning is in a topical formulation that is not (or does not) contain an emulsion (e.g., an emulsion that contains a drug), a smaller pressing force of the microneedle array may be preferred.In some embodiments, a shorter microneedle length may be preferred.In some embodiments, when the large drug administered in conjunction with microneedle skin conditioning is in a topical formulation that is not (or does not) contain an emulsion (e.g., an emulsion that contains a drug), a relatively shorter microneedle length may be preferred.

[0122] Furthermore, as mentioned above, those skilled in the art who read this disclosure will understand that in some embodiments, the application of a relatively reduced amount of product containing biologically active substances (e.g. large drugs) in combination with MSCs can achieve a larger biological effect.Therefore, in some embodiments, a relative reduction in the product volume containing active substances (e.g. large drugs) may be preferred.In some embodiments, when the large drug administered in combination with microneedle skin conditioning is in a topical formulation that is or contains an emulsion (e.g. nanoemulsion), a relatively smaller product volume may be preferred.In some embodiments, when the large drug administered in combination with microneedle skin conditioning is in a topical formulation that is not (or does not contain) an emulsion (e.g. an emulsion that contains a drug), a relatively smaller product volume may be preferred. MN arrays and MSC devices suitable for use in combination with compositions containing large drugs for transdermal delivery of large drugs include devices such as those described in U.S. Patent Nos. 6,334,856; 6,503,231; 6,908,453; 8,257,324; and 9,144,671.

[0123] Blockbuster drugs In some embodiments, the composition provided and / or utilized as described herein comprises one or more large drugs. In some embodiments, the utilized large drug is a biologically active substance (e.g., a therapeutically active substance). In particular, the present disclosure provides a strategy and surprising improvements for topical administration and transdermal delivery of compositions comprising large drugs in combination with MSCs as described herein. Furthermore, the present disclosure demonstrates that microneedling treatment can be particularly advantageous for the delivery of large drugs in emulsion compositions (e.g., nanoemulsions).

[0124] 1. Protein drugs Any of various protein drugs can be incorporated into the provided compositions and administered in combination with MSCs.In some embodiments, the protein drug can be a peptide drug.In some embodiments, the peptide has a molecular weight of more than 100 kDa.In some embodiments, the peptide drug has a molecular weight of at least 150 kDa.In some embodiments, the peptide drug is composed of only naturally occurring amino acids.In some embodiments, the peptide drug contains one or more non-naturally occurring amino acids.

[0125] Those skilled in the art will be aware of various protein drugs that have been approved for therapeutic use by relevant regulatory authorities. For example, the US Food and Drug Administration maintains a list of approved biological therapeutics, organized by the year of approval, which can be found at www.fda.gov / biologicsbloodvaccines / developmentapprovalprocess / biologicalapprovalsbyyear / ucm547553.htm. Those skilled in the art reading this disclosure will understand that the teachings may be applicable to a variety of such drugs. Of particular interest are those intended and / or formulated for local administration, including, for example, those that may be undergoing or have undergone clinical trials (e.g., may be approved or being approved by the US Food and Drug Administration or an equivalent in another jurisdiction, or may be included in clinical trials, e.g., may be included in the list at www.clinicaltrials.gov, or may be on file with one or more clinical facility's Institutional Review Boards or equivalent).

[0126] (i) Botulinum toxin In some embodiments, the large drug may be a botulinum toxin. Botulinum toxin (BTX) is naturally produced by the anaerobic gram-positive bacterium Clostridium botulinum and is a potent polypeptide neurotoxin. Most notably, BTX causes a neuroparalytic disease in humans and animals called botulism. BTX can apparently cross the lining of the intestine and attack peripheral motor nerves. Symptoms of botulinum toxin poisoning may progress from difficulty walking, swallowing, and speaking to paralysis of the respiratory muscles and death.

[0127] The molecular weight of the botulinum toxin protein molecule is approximately 150 kDa for all seven known botulinum toxin serotypes. Botulinum toxins are released by Clostridium bacteria as complexes containing the 150 kDa botulinum toxin protein molecule together with associated non-toxin proteins. Thus, the BTX-A complex is produced by Clostridium bacteria as 900 kDa, 500 kDa and 360 kDa forms. Botulinum toxin types B and C 1 Toxin type 1 is apparently produced only as a 500 kDa complex. Botulinum toxin type D is produced as both a 300 kDa and a 500 kDa complex. Finally, toxin types E and F are produced only as approximately 300 kDa complexes.

[0128] BTX complexes (i.e., those compositions having a molecular weight greater than about 150 kDa) are believed to contain a non-toxin hemagglutinin protein as well as a non-toxin non-toxic non-hemagglutinin protein. These two non-toxin proteins, which together with the botulinum toxin molecule constitute the associated neurotoxin complex, may act to provide stability to the botulinum toxin molecule against denaturation and protection against digestive acids when the toxin is ingested.

[0129] Either the BTX protein or the BTX complex may be utilized in accordance with the present invention as a known therapeutic agent and / or an independently active biologically active substance. Indeed, it will be understood by those skilled in the art that any portion or fragment of the BTX protein or complex that retains the appropriate activity may be utilized as described herein.

[0130] In some embodiments, the botulinum toxin may be selected from the group consisting of types A, Ab, Af, B, Bf, C1, C2, D, E, F and G; mutants thereof; variants thereof; fragments thereof; characteristic portions thereof; and / or fusions thereof. In some embodiments, the botulinum toxin may be a variant toxin, e.g., having one or more structural mutations compared to a reference (e.g., wild-type) toxin (or a related fragment thereof). In some particular embodiments, the variant toxin may have a longer or shorter biological activity lifetime than the appropriate comparable reference form (e.g., wild-type form). In some embodiments, the botulinum toxin exists as any of the subtypes described in Sakaguchi, 1982, Pharmacol. Ther., 19:165; and / or Smith et al., 2005, Infect. Immun., 73:5450, both of which are incorporated herein by reference.

[0131] In some embodiments, the present invention provides a botulinum toxin composition. In some embodiments, the present invention provides a nanoemulsion botulinum toxin composition. Commercially available sources of botulinum toxin that may be utilized in accordance with the present invention include BOTOX®, botulinum toxin 1000 mg ... (登録商標) , DYSPORT (登録商標) (botulinum type A toxin hemagglutinin complex with human serum albumin and lactose; Ispen Limited, Berkshire UK), Xeomin (登録商標) , PurTox (登録商標) , Medy-Tox, NT-201 (Merz Pharmaceuticals), and / or MYOBLOC (登録商標)(An injectable solution consisting of botulinum toxin type B, human serum albumin, sodium succinate and sodium chloride, pH 5.6, Elan Pharmaceuticals, Dublin, Ireland), NEURONOX (Medytox), HENGLI (Lanzhou Institute), and the like. Those skilled in the art will know the standard and / or approved administration regimens of such commercially available botulinum toxin compositions and will understand the relevant such compositions and / or regimens may be utilized in conjunction with the microneedling techniques described herein (e.g., particularly MSCs).

[0132] In some embodiments, compositions provided that include a botulinum toxin composition and are formulated as a cream and / or lotion contain about 1 to about 200,000 units of botulinum toxin per mL. In some embodiments, compositions provided that include a botulinum toxin composition and are formulated as a cream and / or lotion contain about 1 to about 100,000 units of botulinum toxin per mL. In some embodiments, compositions provided that include a botulinum toxin composition and are formulated as a cream and / or lotion contain about 1 to about 50,000 units of botulinum toxin per mL. In some embodiments, compositions provided that include a botulinum toxin composition and are formulated as a cream and / or lotion contain about 500 to about 20,000 units of botulinum toxin per mL. In some embodiments, compositions provided that include a botulinum toxin composition and are formulated as a cream and / or lotion contain about 100 to about 2,000 units of botulinum toxin per mL. In some embodiments, compositions provided that include a botulinum toxin composition and are formulated as a cream and / or lotion contain about 50 to about 500 units of botulinum toxin per mL. In some embodiments, compositions provided that include a botulinum toxin composition and are formulated as a cream and / or lotion contain about 25 to about 400 units of botulinum toxin per mL.

[0133] In some embodiments, the botulinum toxin composition comprises about 2 to about 40,000 units of botulinum toxin per mL. In some embodiments, the botulinum toxin composition comprises about 2 to about 12,000 units of botulinum toxin per mL. In some embodiments, the botulinum toxin composition comprises about 100 to about 2,000 units of botulinum toxin per mL. In some embodiments, the botulinum toxin composition comprises about 50 to about 1,000 units of botulinum toxin per mL.

[0134] In some embodiments, the botulinum toxin composition comprises at least one biologically active substance other than the botulinum toxin. Alternatively or additionally, in some embodiments, the botulinum composition is administered in combination with at least one other composition other than such biologically active substance. In some embodiments, the botulinum composition is administered in combination with a penetration enhancer. In some embodiments, the botulinum composition is administered in combination with another biologically active substance. In some embodiments, the botulinum composition is administered in combination with another biologically active substance and a penetration enhancer.

[0135] In some embodiments, the biologically active agent utilized in combination with the botulinum toxin described herein can be a drug that acts on or in the skin and / or has a therapeutic and / or cosmetic effect.For example, in some embodiments, such biologically active agents are selected from therapeutic agents such as anesthetics (e.g., lidocaine), steroids (e.g., hydrocortisone) and / or retinoids (e.g., retinoin), cosmetic agents such as dermal fillers (e.g., hyaluronic acid or other elastic materials), collagen and / or silicone.In some embodiments, the botulinum composition is administered in combination with a delivery modifier, such as a penetration enhancer (which in some embodiments is not irritating and / or does not degrade, destroy and / or damage skin structure and / or skin).

[0136] In some embodiments, the non-irritating penetration enhancer may be selected from, for example, copeptides, carrier molecules, and carrier peptides. In some embodiments, the carrier molecule is positively charged. In some embodiments, the carrier molecule may be a copeptide. In some embodiments, the carrier molecule may be a long chain positively charged polypeptide or a positively charged non-peptidyl polymer, such as a polyalkyleneimine. In some embodiments, the carrier peptide may be a cationic peptide. In some embodiments, the carrier peptide may be a peptide having the sequence RKKRRQRRRG-(K) 15 -GRKKRRQRRR is a positively charged carrier. In some embodiments, the carrier molecule can be one disclosed in U.S. Patent Application Publication No. 2010 / 0168023 or U.S. Patent Application Publication No. 2009 / 0247464, the contents of which are incorporated herein by reference in their entirety.

[0137] In some embodiments, compositions provided, including both a botulinum toxin nanoemulsion composition and a cream and / or lotion formulation, contain from about 1 to about 100,000 units of botulinum toxin per mL. In some embodiments, compositions provided, including both a botulinum toxin nanoemulsion composition and a cream and / or lotion formulation, contain from about 1 to about 100,000 units of botulinum toxin per mL. In some embodiments, compositions provided, including both a botulinum toxin nanoemulsion composition and a cream and / or lotion formulation, contain from about 1 to about 50,000 units of botulinum toxin per mL. In some embodiments, compositions provided, including both a nanoemulsion composition and a cream and / or lotion formulation, contain from about 500 to about 20,000 units of botulinum toxin per mL. In some embodiments, provided compositions, including both a nanoemulsion composition and a cream and / or lotion formulation, contain from about 100 to about 2,000 units of botulinum toxin per mL. In some embodiments, provided compositions, including both a botulinum toxin nanoemulsion composition and a cream and / or lotion formulation, contain from about 50 to about 500 units of botulinum toxin per mL. In some embodiments, provided compositions, including both a botulinum toxin nanoemulsion composition and a cream and / or lotion formulation, contain from about 25 to about 400 units of botulinum toxin per mL.

[0138] In some embodiments, the botulinum toxin nanoemulsion composition comprises about 2 to about 40,000 units of botulinum toxin per mL. In some embodiments, the botulinum toxin nanoemulsion composition comprises about 2 to about 12,000 units of botulinum toxin per mL. In some embodiments, the botulinum toxin nanoemulsion composition comprises about 100 to about 2,000 units of botulinum toxin per mL. In some embodiments, the botulinum toxin nanoemulsion composition comprises about 50 to about 1,000 units of botulinum toxin per mL.

[0139] (ii) Antibody drugs In some embodiments, the present disclosure relates to the delivery of antibody drugs.In some embodiments, the large drug can be an antibody or its fragment or derivative.In particular, the present disclosure provides certain compositions that include antibody drugs, and also provides techniques for the administration of compositions that include antibody drugs, and such administration is combined with MSCs as described herein.

[0140] In some embodiments, the antibody drug may be suitable for treating skin conditions. In some embodiments, the antibody drug may be a fusion protein. In some embodiments, the antibody drug may be conjugated with another moiety. In some embodiments, the antibody drug may be conjugated with polyethylene glycol. In some embodiments, the antibody may be multispecific (e.g., bispecific) and may bind to two or more different target antigens or epitopes.

[0141] In some embodiments, the antibody comprises TNFα (e.g., comprises an epitope binding element found in anti-TNFα antibodies such as infliximab, adalimumab, golimumab, etanercept, etanercept-szzs, and / or certolizumab pegol). In some embodiments, the antibody comprises CD2 (e.g., comprises an epitope binding element found in anti-CD2 antibodies such as siplizumab). In some embodiments, the antibody comprises CD4 (e.g., comprises an epitope binding element found in anti-CD4 antibodies such as zanolimumab).

[0142] In some embodiments, the antibody comprises IL-12 (e.g., comprises an epitope binding element found in an anti-IL-12 antibody, such as briakinumab). In some embodiments, the antibody comprises IL-17 (e.g., comprises an epitope binding element found in an anti-IL-17 antibody, such as secukinumab and / or brodalumab). In some embodiments, the antibody comprises IL-22 (e.g., comprises an epitope binding element found in an anti-IL-22 antibody, such as fezakinumab). In some embodiments, the antibody comprises IL-23 (e.g., comprises an epitope binding element found in ustekinumab and / or guselkumab).

[0143] In some embodiments, the antibody drug composition comprises at least one biologically active substance other than the antibody drug. Alternatively or additionally, in some embodiments, the antibody drug composition is administered in combination with at least one other composition comprising such a biologically active substance. In some embodiments, the antibody drug composition is administered in combination with a penetration enhancer. In some embodiments, the antibody drug composition is administered in combination with another biologically active substance. In some embodiments, the antibody drug composition is administered in combination with another biologically active substance and a penetration enhancer. In some embodiments, the antibody drug composition is a nanoemulsion. In some embodiments, the antibody drug composition is a cream and / or lotion formulation.

[0144] In some embodiments, the biologically active agent utilized in combination with the antibody drug described herein can be a drug that acts on or in the skin and / or has a therapeutic and / or cosmetic effect.For example, in some embodiments, such biologically active agent is selected from therapeutic agents such as anesthetics (e.g., lidocaine), steroids (e.g., hydrocortisone) and / or retinoids (e.g., retinoin), cosmetic agents such as dermal fillers (e.g., hyaluronic acid or other elastic materials), collagen and / or silicone.In some embodiments, the antibody drug composition is administered in combination with a delivery modifier, such as a penetration enhancer (which in some embodiments is not irritating and / or does not degrade, destroy and / or damage skin structure and / or skin).

[0145] In some embodiments, the non-irritating penetration enhancer may be selected from, for example, copeptides, carrier molecules, and carrier peptides. In some embodiments, the carrier molecule is positively charged. In some embodiments, the carrier molecule may be a copeptide. In some embodiments, the carrier molecule may be a long chain positively charged polypeptide or a positively charged non-peptidyl polymer, such as a polyalkyleneimine. In some embodiments, the carrier peptide may be a cationic peptide. In some embodiments, the carrier peptide may be a peptide having the sequence RKKRRQRRRG-(K) 15 -GRKKRRQRRR is a positively charged carrier. In some embodiments, the carrier molecule can be one disclosed in U.S. Patent Application Publication No. 2010 / 0168023 or U.S. Patent Application Publication No. 2009 / 0247464, the contents of which are incorporated herein by reference in their entirety.

[0146] 2. Preventive medicines Any of a variety of prophylactic agents can be incorporated into the provided compositions and administered in combination with the MSCs of the present invention.In some embodiments, the prophylactic agent includes, but is not limited to, a vaccine.In some embodiments, the vaccine can include isolated proteins or peptides, inactivated organisms and viruses, killed organisms and viruses, genetically modified organisms or viruses, and cell extracts.In some embodiments, the prophylactic agent can be combined with interleukins, interferons, cytokines, and adjuvants, such as cholera toxin, alum, and Freund's adjuvant.In some embodiments, the prophylactic agent is selected from the group consisting of Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus, Streptococcus pyogenes, Corynebacterium diphtheriae, Listeria monocytogenes, Bacillus anthracis, Clostridium tetani, Clostridium botulinum, Clostridium perfringens, Neisseria meningitidis, Neisseria gonorrhoeae, Streptococcus mutans, Pseudomonas aeruginosa, Salmonella typhi, Haemophilus parainfluenzae, Bordetella pertussis, Francisella tularensis, Yersinia pestis, Vibrio cholerae, Legionella pneumophila, Mycobacterium tuberculosis, Mycobacterium leprae, Treponema pallidum, Leptospira interrogans, and the like. interrogans), Borrelia burgdorferi, Campylobacter jejuni; antigens of bacterial organisms such as smallpox, influenza A and B, respiratory syncytial virus, parainfluenza, measles, HIV, varicella zoster, herpes simplex 1 and 2, cytomegalovirus, Epstein-Barr virus, rotavirus, rhinovirus, adenovirus, papillomavirus, poliovirus, mumps, rabies, rubella, coxsackievirus, equine encephalitis, Japanese encephalitis, yellow fever, Rift Valley fever, hepatitis A, B, C, D and E antigens of viruses such as Cryptococcus neoformans, Histoplasma capsulatum, Candida albicans, Candida tropicalis, Nocardia asteroides, Rickettsia rickettsii, Rickettsia typhi, Mycoplasma pneumoniae, Chlamydia psittaci, Chlamydia trachomatis, Plasmodium falciparum, Trypanosoma brucei, Entamoeba histolytica, Toxoplasma gondii, Trichomonas vaginalis, Schistosoma mansoni, and other fungal, protozoan, and parasitic antigens. In some embodiments, these antigens can be in the form of whole killed organisms, peptides, proteins, glycoproteins, carbohydrates, or combinations thereof.

[0147] Those skilled in the art will recognize that the preceding paragraphs provide an exemplary, not exhaustive, list of drugs that can be delivered using the techniques according to the present invention. Any drug can be associated with the compositions provided in accordance with the present invention.

[0148] Topical preparations The compositions described herein are particularly useful in that they can be used to deliver large drugs to a subject in need thereof by topical and / or transdermal (e.g., by lotions, creams, powders, ointments, liniments, gels, eye drops, etc.) administration. In some embodiments, the provided cream and / or lotion formulations containing large drugs are administered to a subject in need thereof by topical and / or transdermal (e.g., by lotions, creams, powders, ointments, liniments, gels, eye drops, etc.) administration.

[0149] In some embodiments, the cream and / or lotion formulation comprises purified water, methylparaben, mineral oil, isopropyl myristate, white petrolatum, emulsifying wax, and propylparaben.In some embodiments, the cream and / or lotion formulation comprises purified water, mineral oil, isopropyl myristate, white petrolatum, and emulsifying wax.

[0150] In some embodiments, the present invention provides certain cream and / or lotion formulations described herein. In some embodiments, the cream and / or lotion formulations provided comprise water. In some embodiments, the cream and / or lotion formulations provided comprise methylparaben. In some embodiments, the cream and / or lotion formulations provided comprise mineral oil. In some embodiments, the cream and / or lotion formulations provided comprise isopropyl myristate. In some embodiments, the cream and / or lotion formulations provided comprise white petrolatum. In some embodiments, the cream and / or lotion formulations provided comprise emulsifying wax. In some embodiments, the cream and / or lotion formulations provided comprise propylparaben. In some embodiments, the cream and / or lotion formulations provided are paraben-free. In some embodiments, the cream and / or lotion formulations provided are methylparaben-free. In some embodiments, the cream and / or lotion formulations provided are propylparaben-free. Exemplary lotion formulations are provided in Table 1. [Table 1]

[0151] In some embodiments, cream and / or lotion formulations can be useful for topical and / or transdermal administration.The present invention encompasses the recognition that the cream and / or lotion formulations provided can be particularly useful for delivering drugs to the dermal layer of the skin.In some embodiments, the cream and / or lotion formulations provided are formulated for topical and / or transdermal delivery to a subject in need thereof.In some embodiments, the cream and / or lotion formulations provided are administered to a subject in need thereof by topical and / or transdermal delivery.

[0152] In some embodiments, the provided composition is formulated with cosmetically acceptable ingredients.For example, in some embodiments, the provided composition is formulated with water and any cosmetically acceptable solvent, in particular monoalcohols, such as alkanols having 1-8 carbon atoms (e.g. ethanol, isopropanol, benzyl alcohol and phenylethyl alcohol), polyalcohols, such as alkylene glycols (e.g. glycerin, ethylene glycol and propylene glycol), and glycol ethers, such as mono-, di- and tri-ethylene glycol monoalkyl ethers, such as ethylene glycol monomethyl ether and diethylene glycol monomethyl ether (used alone or in mixture).Such ingredients can be present, for example, in a proportion of up to 60%, 70%, 80% or 90% by weight based on the weight of the total composition.

[0153] In some embodiments, the provided compositions for topical administration include one or more cosmetically acceptable ingredients that impart desirable or suitable appearance characteristics to the subject to which the composition is applied (e.g., a matte appearance, which may be particularly desirable or suitable for administration to subjects with oily skin).

[0154] In some embodiments, provided compositions are formulated with at least one cosmetically acceptable filler material to obtain a mattifying product, which may be particularly desirable, for example, for individuals with oily skin.

[0155] In some embodiments, the large drug is formulated into a composition suitable for local administration.Exemplary large drugs include those described herein.In some embodiments, the provided composition can be formulated and delivered in combination with MSCs described herein to achieve systemic delivery; in some embodiments, the provided composition can be formulated and / or delivered to achieve local but not systemic delivery.

[0156] In some embodiments, the composition suitable for topical formulation comprises a penetration enhancer. In some embodiments, the penetration enhancer degrades, destroys and / or damages the skin structure and / or skin. In some embodiments, the penetration enhancer does not degrade, destroy and / or damage the skin structure and / or skin. In some embodiments, the penetration enhancer is irritating. In some embodiments, the penetration enhancer is not irritating.

[0157] The present disclosure specifically demonstrates effective and efficient delivery of therapeutic agents (particularly large biological drugs, such as botulinum toxin and / or antibody drugs) to the dermis using the compositions provided in combination with the MSCs described herein. For example, in some embodiments, the present invention provides methods that include administration of the compositions described herein without clinically significant side effects. By way of example, when local delivery is contemplated, clinically significant side effects include, but are not limited to, undesirable systemic side effects, damage to nerve tissues underlying the dermis (e.g., nerve paralysis), undesirable effects on muscles (e.g., muscle paralysis), and / or undesirable blood levels of therapeutic agents.

[0158] Those skilled in the art will understand that the provided compositions can be incorporated into devices such as patches.Various transdermal patch structures are known in the art; those skilled in the art will understand that the provided compositions can be easily incorporated into any of such various structures.In some embodiments, the transdermal patch can include multiple needles extending from one side of the patch that is applied to the skin, where the needles extend from the patch and protrude through the stratum corneum of the skin.In some embodiments, the needles do not rupture blood vessels.In some embodiments, the needles do not penetrate deep enough to reach nerves in the dermis of the skin.

[0159] In some embodiments, the transdermal patch comprises an adhesive.Several examples of adhesive patches are known (e.g., U.S. Design Patent No. 296,006; and U.S. Patent Nos. 6,010,715; 5,591,767; 5,008,110; 5,683,712; 5,948,433; and 5,965,154; all of which are incorporated herein by reference).Adhesive patches are generally characterized by having an adhesive layer that is applied to the skin of a patient, a depot or reservoir for holding the composition provided, and an outer surface that prevents the composition provided from leaking from the depot.The outer surface of the patch can be non-adhesive.

[0160] According to the present invention, the provided composition is incorporated into the patch so that it remains stable for a long period of time.For example, in some embodiments, the provided composition can be incorporated into a polymer matrix that stabilizes large drugs and allows the drug to diffuse out of the matrix and the patch.The provided composition can also be incorporated into the adhesive layer of the patch so that the provided composition can diffuse through the skin when the patch is applied to the skin.In some embodiments, the adhesive layer can be heat-activated, where a temperature of about 37°C slowly liquefies the adhesive so that the drug diffuses through the skin.The adhesive can remain sticky when stored below 37°C, and lose its stickiness as it liquefies when applied to the skin.

[0161] In some embodiments, the provided composition can be provided in a depot in the patch, so that pressure applied to the patch directs the provided composition through the microneedle and the stratum corneum to the outside of the patch.Exemplary embodiments of microneedles are described above.Devices suitable for intradermal administration of the provided composition include, for example, those described in U.S. Patent Nos. 4,886,499; 5,190,521; 5,328,483; 5,527,288; 4,270,537; 5,015,235; 5,141,496; and 5,417,662.Intradermal compositions can be administered by devices that limit the effective penetration length of needles into the skin, such as those described in WO 99 / 34850 and their functional equivalents.

[0162] In some embodiments, it may be desirable to slow down the absorption of the provided composition into the skin, for example, to prolong the effect of the provided composition.In some embodiments, this can be achieved by using a liquid suspension of crystalline or amorphous material with low water solubility.Then, the absorption rate of the provided composition depends on its dissolution rate, which in turn can depend on the crystal size and crystalline form.In some embodiments, depending on the ratio of the provided composition to polymer and the nature of the specific polymer used, the release rate of the provided composition can be controlled.Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides).

[0163] Emulsions The present disclosure encompasses the recognition that emulsion technology can provide stabilization benefits to drugs of interest, including large drugs as described herein, and in particular, including botulinum toxins and / or antibody drugs.

[0164] Further, the present disclosure recognizes that certain liquid nanoemulsion technologies have been shown to provide superior transdermal delivery properties, even for extremely large molecules such as botulinum and / or antibody drugs. See, for example, U.S. Patent Application Publication Nos. 2012 / 0328701, 2012 / 0328702, 8,318,181, and 8,658,391, the disclosures of which are incorporated herein by reference in their entirety. These liquid nanoemulsions are far superior to solid nanoparticle drug delivery, especially transdermal drug delivery, which, as Gomaa states, cannot penetrate the skin and only accumulate in hair follicles. These liquid nanoemulsions are stable for at least 34 months, making them commercially viable from this perspective as well.

[0165] 1. Macroemulsion In some embodiments, the present disclosure provides a strategy of using microneedling to "condition" the skin to which a transdermal product has been applied, has been applied, or will be applied. Despite previous reports that such a strategy is likely to be useful only for drugs with small molecular weights, since a study analyzing the transdermal delivery of small molecules (specifically, short hydrophilic peptides with molecular weights in the range of 400-1000 Da) found that "skin penetration of peptides depends on their molecular weight, with penetration decreasing as molecular weight increases," the present disclosure provides the finding that such microneedle conditioning can surprisingly provide significant benefits in improving the transdermal delivery of large drugs (e.g., having a molecular weight of about 100 kDa or more). Zhang, S., et al., "Enhanced delivery of hydrophilic peptides in vitro by transdermal microneedle pretreatment." Acta Pharmaceutica Sinica B. 4(1):100-104 (2014).

[0166] The present disclosure provides the discovery that effective and rapid (i.e., administration over minutes) transdermal delivery of macromolecules by such liquid macroemulsion compositions can surprisingly be improved by combining macroemulsion administration with microneedle skin conditioning (MSC) as described herein (e.g., using a relatively low microneedle density and / or a relatively small microneedle puncture size).

[0167] The present disclosure in particular demonstrates that microneedling techniques (e.g., microneedle conditioning of the skin) can significantly enhance the transdermal delivery of large drugs (e.g., botulinum and / or antibody drugs), especially when utilized in conjunction with macroemulsion techniques. Particular macroemulsion compositions of interest include water-in-oil and oil-in-water macroemulsions characterized by droplet sizes ranging from greater than about 300 nm to about 5,000 μm in diameter, ratios of aqueous dispersion medium to oil ranging from about 0.01:1 to about 20:1; ratios of oil to surfactant ranging from about 0.1 to about 40, and / or zeta potentials ranging from about -80 mV to about +80 mV. The surfactant portion of the composition may include one or more surfactants. [Table 2]

[0168] Macroemulsion formulations can act to stabilize large drugs, such as botulinum and / or antibody drugs. Although macroemulsion formulations are not necessarily expected to achieve transdermal delivery of large drugs by themselves, the present disclosure nevertheless embraces the finding that the improved stabilization that can be provided by incorporation into macroemulsion compositions can achieve synergistic improvements in transdermal delivery when combined with the microneedle technology described herein.

[0169] The present disclosure teaches that, despite the expectation that MSCs are only useful for enhancing the transdermal delivery of small compounds, transdermal delivery of large drugs by nanoemulsion compositions may be possible in combination with microneedle technology. This disclosure is particularly surprising considering that microneedle conditioning combined with encapsulation of small molecule drugs in solid nanoparticles only provided a small amount of penetration 6 hours after administration, as explained by Gomaa, and penetration of the material was observed up to 12 hours after administration.

[0170] In some embodiments, the macroemulsion formulation comprising a large drug may be administered in combination with microneedle conditioning with solid microneedles. In some embodiments, MSC of the site is performed prior to application of the macroemulsion formulation comprising a large drug to the site (e.g., prior to the particular application and / or each application). In some embodiments, MSC of the site is performed after application of the macroemulsion formulation comprising a large drug to the site. In some embodiments, MSC of the site and application of the macroemulsion formulation comprising a large drug to the site occur substantially simultaneously. In some embodiments, the macroemulsion formulation comprising a large drug is not injected by one or more microneedles. In some embodiments, the microneedle is part of an array of microneedles. In some embodiments, the microneedle may be about 1 μm to about 4,000 μm long. In some embodiments, the microneedle may be about 1 μm to about 2,000 μm long. In some embodiments, the microneedle may be about 50 μm to about 400 μm long. In some embodiments, the microneedles may be from about 800 μm to about 1500 μm in length.

[0171] The findings presented herein are particularly surprising in light of reports that transdermal delivery of solid nanoparticles with sizes (e.g., 105±2.92 nm) much smaller than those of the droplets in the macroemulsion compositions utilized herein effectively delivers (or enhances) even small molecule drugs transdermally across the skin. For example, Gomaa et al. described a study in which a solution of rhodamine dye (molecular weight 479 Da) encapsulated in PLGA nanoparticles was applied to preconditioned skin by microneedles and skin permeation was evaluated. See Gomaa, Y., et al, "Effect of microneedle treatment on the skin permeation of a nanoencapsulated dye." J Pharm Pharmacol. 2012 November; 64(11): 1592-1602. The data showed that very little dye began to permeate the skin after 6 hours of continuous application; no significant increase in permeation was observed until the skin was continuously treated for 24 hours. The researchers explained that "there is an emerging consensus that NPs [nanoparticles] can be well deposited in hair follicles, but generally cannot penetrate the stratum corneum." Thus, prior to this disclosure, one skilled in the art would have expected that the use of microneedling technology with vehicles significantly larger than 105 nm would not be able to effectively deliver even small molecule drugs (e.g., rhodamine dyes) transdermally; indeed, delivery of large drugs would have been considered impossible. However, the present disclosure shows that microneedling can significantly enhance the transdermal delivery of large drugs, especially when combined with macroemulsion technology.

[0172] In particular, the present disclosure discloses that microneedling technology can improve transdermal delivery (e.g., of large drugs, especially from macroemulsion compositions) when other disruptive agents (i.e., chemical penetration enhancers and other techniques that disrupt or puncture skin structures) are not utilized. Previous studies of transdermal delivery of drugs similar in size to botulinum toxin (i.e., about 150 kDa) using microneedles have reported that delivery fails unless further treatment is applied to disrupt the skin. For example, U.S. Patent Application Publication No. 2010 / 0196445 reports that botulinum toxin is not effectively delivered from pre-coated microneedles unless skin digestive enzymes are also applied so that skin structures are disrupted at the site of the microneedle.

[0173] The present disclosure shows, inter alia, that microneedling techniques can achieve transdermal delivery (e.g., of large drugs, particularly from macroemulsion and nanoemulsion compositions) when no microneedle coating or loading is utilized and / or when the microneedles are not designed to be left in the skin. In particular, as previously noted, the present disclosure recognizes that such microneedle coating or loading may not be commercially viable due to the instability of at least the botulinum coating or loading material. For example, according to Johnson, E. et al., "Botulinum toxin is highly susceptible to denaturation due to surface denaturation, heat and alkaline conditions. Freeze-drying of botulinum toxin is the most economically sound and practical method of distributing the product in a form that is stable and easily used by clinicians." U.S. Patent No. 5,512,547. Also, as will be appreciated by those of skill in the art reading this specification, the techniques described herein have certain advantages, including the absence of the need for microneedles to be left in or associated with tissue. For example, one skilled in the art will appreciate that leaving microneedles in the skin risks skin irritation, inflammation, allergic reactions, and / or cosmetically undesirable scarring. In contrast to the present invention, techniques such as those described in US Patent Application Publication No. 2017 / 0209553 utilize microneedle arrays that are loaded with botulinum and designed to penetrate the microneedles into the skin (US Patent Nos. 2017 / 0209553 and 2016 / 0263362).

[0174] The present disclosure surprisingly provides an effective technique for transdermal delivery of large drugs (e.g., botulinum toxin, antibodies, etc.). In particular, the present disclosure teaches that the transdermal delivery of such drugs can be significantly improved by the use of microneedling technology without other disruptive strategies. Thus, the provided technology can achieve effective delivery without the inflammation, irritation, and / or allergic reactions that often accompany the use of skin disrupting agents. As will be understood by those skilled in the art upon reading this specification, the present disclosure teaches that the transdermal delivery of such large drugs can be significantly improved by the use of microneedling technology, even when not loaded into, coated on, and / or manufactured as part of, the microneedles. Similarly, as will be understood by those skilled in the art upon reading this specification, the present disclosure teaches that the delivery of large drugs described herein can be significantly improved by the use of microneedling technology (particularly the use of MSCs), without leaving microneedles in the skin (e.g., by breaking them and / or otherwise maintaining and / or degrading them in situ). For example, one of skill in the art will appreciate that the provided technology can avoid challenges with long-term stability of large drugs necessary for commercially viable products, and can achieve effective delivery without inflammation, irritation, and / or allergic reactions that may result from skin-disrupting agents and / or microneedles left in the skin. Indeed, in the Examples and elsewhere, the present disclosure expressly teaches that MSCs implemented with microneedles that do not contain botulinum toxin facilitate transdermal delivery of botulinum toxin from topical (e.g., creams, ointments) compositions, particularly from compositions that include macro- or nano-emulsions.

[0175] In some embodiments, the present disclosure teaches that when microneedling technology is combined with macroemulsion composition, particularly advantageous results are achieved.In some embodiments, microneedling technology is combined with lotion, cream or liquid composition, which in turn can be or contain macroemulsion composition.In some embodiments, the provided technology does not utilize skin disruption technology, such as chemical penetration enhancers.

[0176] In some embodiments, the present invention utilizes macroemulsion compositions that contain large drugs that are particularly effective and / or useful in medical situations, for example for therapeutic purposes.In some embodiments, certain macroemulsion compositions are particularly effective and / or useful for topical administration of drugs to subjects in need thereof.In some embodiments, macroemulsion compositions can contain one or more large drugs.

[0177] In some embodiments, the macroemulsion can be formulated into a composition suitable for topical administration on the skin. In some embodiments, the composition suitable for topical administration can be a lotion, cream, powder, ointment, liniment, gel, or eye drop.

[0178] In some embodiments, the macroemulsion formulation comprises water, medium chain triglycerides, Span 65, polysorbate 80, methylparaben, and propylparaben. In some embodiments, the macroemulsion formulation comprises water, medium chain triglycerides, Span 65, and polysorbate 80.

[0179] In some embodiments, the provided compositions comprise a mixture of the provided macroemulsion compositions and one or more pharma- ceutically acceptable excipients. In some embodiments, cream and / or lotion formulations comprise a mixture of the provided macroemulsion compositions and / or saline solution.

[0180] In some embodiments, the compositions provided comprise a macroemulsion composition comprising one or more large drugs. In some embodiments, the compositions provided are cream and / or lotion formulations. In some embodiments, the cream and / or lotion formulations provided comprise a macroemulsion composition. In some embodiments, the compositions provided comprise a macroemulsion composition but are not cream and / or lotion formulations. In some embodiments, the suitable compositions are formulated into cream and / or lotion but do not comprise a macroemulsion composition.

[0181] In some embodiments, provided compositions comprise a mixture of provided macroemulsion compositions and one or more pharma- ceutically acceptable excipients, e.g., for topical and / or transdermal administration (e.g., via lotions, creams, powders, ointments, liniments, gels, eye drops, etc.).

[0182] 2. Nanoemulsion In some embodiments, the present disclosure provides a strategy of using microneedling to "condition" the skin to which a transdermal product has been applied, has been applied, or will be applied. Despite previous reports that such a strategy is likely to be useful only for drugs with small molecular weights, since a study analyzing the transdermal delivery of small molecules (specifically, short hydrophilic peptides with molecular weights in the range of 400-1000 Da) found that "skin penetration of peptides depends on their molecular weight, with penetration decreasing as molecular weight increases," the present disclosure provides the finding that such microneedle conditioning can surprisingly provide significant benefits in improving the transdermal delivery of large drugs (e.g., having a molecular weight of about 100 kDa or more). Zhang, S., et al., "Enhanced delivery of hydrophilic peptides in vitro by transdermal microneedle pretreatment." Acta Pharmaceutica Sinica B. 4(1):100-104 (2014).

[0183] The present disclosure provides the discovery that effective and rapid (i.e., administration over minutes) transdermal delivery of macromolecules by such liquid nanoemulsion compositions can be improved by combining nanoemulsion administration with microneedle skin conditioning (MSC) as described herein.

[0184] The present disclosure specifically demonstrates that microneedling technology (e.g., microneedle conditioning of the skin) can significantly enhance the transdermal delivery of large drugs (e.g., botulinum and / or antibody drugs), especially when utilized in conjunction with nanoemulsion technology. Particular nanoemulsion compositions of interest include water-in-oil and oil-in-water nanoemulsions characterized by droplet sizes ranging from about 1 nm to about 300 nm in diameter, ratios of aqueous dispersion medium to oil ranging from about 0.01:1 to about 20:1; oil to surfactant ratios ranging from about 0.1 to about 40, and / or zeta potentials ranging from about -80 mV to about +80 mV.

[0185] In some embodiments, provided nanoemulsion compositions include an oil and a surfactant in a ratio ranging from about 0.1:1 to about 2:1. In some embodiments, provided nanoemulsion compositions include an oil and a surfactant in a ratio ranging from about 0.1:1 to about 1:1. In some embodiments, provided nanoemulsion compositions include an oil and a surfactant in a ratio ranging from about 0.5:1 to about 1:1. In some embodiments, provided nanoemulsion compositions include an oil and a surfactant in a ratio ranging from about 0.5:1 to about 1:1.5. In some embodiments, provided nanoemulsion compositions comprise an oil and surfactant in a ratio of about 0.1:1, about 0.15:1, about 0.2:1, about 0.25:1, about 0.3:1, about 0.35:1, about 0.4:1, about 0.45:1, about 0.5:1, about 0.5:1, about 0.55:1, about 0.6:1, about 0.65:1, about 0.7:1, about 0.75:1, about 0.8:1, about 0.85:1, about 0.9:1, about 0.95:1, or about 1:1. In some embodiments, provided nanoemulsion compositions comprise an oil and surfactant in a ratio of about 0.67:1.

[0186] In some embodiments, the aqueous dispersion medium (e.g., water, buffer, salt solution, etc.) and the surfactant are utilized in a ratio ranging from 0.01 to 20. In some embodiments, the aqueous dispersion medium (e.g., water, buffer, salt solution, etc.) and the surfactant are utilized in a ratio ranging from 0.1 to 20. In some embodiments, the aqueous dispersion medium (e.g., water, buffer, salt solution, etc.) and the surfactant are utilized in a ratio ranging from 0.5 to 10. In some embodiments, the aqueous dispersion medium (e.g., water, buffer, salt solution, etc.) and the surfactant are utilized in a ratio ranging from 0.5 to 1. In some embodiments, the ratio of aqueous dispersion medium (e.g., water, buffer, salt solution, etc.) to surfactant is about 0.01:1, about 0.02:1, about 0.03:1, about 0.04:1, about 0.05:1, about 0.06:1, about 0.07:1, about 0.08:1, about 0.0:1, about 0.1:1, about 0.2:1, about 0.3:1, about 0.4:1, about 0.5:1, about 1:1, about 2:1, about 3:1, about 4:1, about 5:1, about 6:1, about 7:1, about 8:1, about 9:1, or about 10:1. In some embodiments, the ratio of surfactant to water is about 0.5:1, about 1:1, about 2:1, about 3:1, about 4:1, about 5:1, about 6:1, about 7:1, about 8:1, about 9:1, about 10:1, about 11:1, about 12:1, about 13:1, about 14:1, about 15:1, about 16:1, about 17:1, about 18:1, about 19:1, or about 20:1. In some embodiments, the aqueous dispersion medium (e.g., water, buffer, salt solution, etc.) and the surfactant are utilized in a ratio ranging from 0.5 to 2. In some embodiments, the ratio of the aqueous dispersion medium (e.g., water, buffer, salt solution, etc.) to the surfactant is about 0.5:1, about 1:1, or about 2:1. In some embodiments, the ratio of the surfactant to the aqueous dispersion medium (e.g., water, buffer, salt solution, etc.) is about 0.5:1, about 1:1, or about 2:1. In some embodiments, the ratio of aqueous dispersion medium (e.g., water, buffer, salt solution, etc.) to surfactant is about 1: 1. In some embodiments, compositions utilizing such a ratio of aqueous dispersion medium (e.g., water, buffer, salt solution, etc.) to surfactant comprise a water-in-oil emulsion.

[0187] In some embodiments, the droplets in the nanoemulsion composition have a diameter (e.g., average diameter and / or median diameter) within the range of about 10 nm to about 300 nm, about 10 nm to about 200 nm, about 10 nm to about 150 nm, about 10 nm to about 130 nm, about 10 nm to about 120 nm, about 10 nm to about 115 nm, about 10 nm to about 110 nm, about 10 nm to about 100 nm, or about 10 nm to about 90 nm. In some embodiments, the droplets in the nanoemulsion composition have a diameter (e.g., average diameter and / or median diameter) within the range of 1 nm to 300 nm, 1 nm to 200 nm, 1 nm to 150 nm, 1 nm to 120 nm, 1 nm to 100 nm, 1 nm to 75 nm, 1 nm to 50 nm, or 1 nm to 25 nm. In some embodiments, the droplets within a nanoemulsion composition have a diameter (eg, average diameter and / or median diameter) between 1 nm and 15 nm, between 15 nm and 200 nm, between 25 nm and 200 nm, between 50 nm and 200 nm, or between 75 nm and 200 nm.

[0188] In some embodiments, the entire droplet distribution falls within the specified range of droplet diameter size. In some embodiments, less than 50%, 25%, 10%, 5% or 1% of the entire droplet distribution is outside the specified range of droplet diameter size. In some embodiments, less than 1% of the entire droplet distribution is outside the specified range of droplet diameter size. In some embodiments, the nanoemulsion composition is substantially free of droplets having a diameter greater than 300 nm, 250 nm, 200 nm, 150 nm, 120 nm, 100 nm, 75 nm, 50 nm or 25 nm. In some embodiments, less than 50%, 25%, 10%, 5% or 1% of the entire droplet distribution has a diameter greater than 300 nm, 250 nm, 200 nm, 150 nm, 120 nm, 100 nm, 75 nm, 50 nm or 25 nm.

[0189] In some embodiments, the droplets in the nanoemulsion composition have an average droplet size of about 300 nm, about 250 nm, about 200 nm, about 150 nm, about 130 nm, about 120 nm, about 115 nm, about 110 nm, about 100 nm, about 90 nm, or less than about 50 nm. In some embodiments, the average droplet size is within the range of about 10 nm and about 300 nm, about 50 nm and about 250, about 60 nm and about 200 nm, about 65 nm and about 150 nm, or about 70 nm and about 130 nm. In some embodiments, the average droplet size is about 80 nm and about 110 nm. In some embodiments, the average droplet size is about 90 nm and about 100 nm.

[0190] In some embodiments, the nanoemulsion droplets have a zeta potential ranging from -80 mV to +80 mV. In some embodiments, the nanoemulsion droplets have a zeta potential ranging from -50 mV to +50 mV. In some embodiments, the nanoemulsion droplets have a zeta potential ranging from -25 mV to +25 mV. In some embodiments, the nanoemulsion droplets have a zeta potential ranging from -10 mV to +10 mV. In some embodiments, the nanoemulsion droplets have a zeta potential of about -80 mV, about -70 mV, about -60 mV, about -50 mV, about -40 mV, about -30 mV, about -25 mV, about -20 mV, about -15 mV, about -10 mV, or about -5 mV. In some embodiments, the nanoemulsion droplets have a zeta potential of about +50 mV, about +40 mV, about +30 mV, about +25 mV, about +20 mV, about +15 mV, about +10 mV, or about +5 mV. In some embodiments, the nanoemulsion droplets have a zeta potential of about 0 mV.

[0191] The present disclosure surprisingly provides an effective technique for transdermal delivery of large drugs (e.g., botulinum toxin, antibodies, etc.). In particular, the present disclosure teaches that the transdermal delivery of such drugs can be significantly improved by the use of microneedling technology without other disruptive strategies. Thus, the provided technology can achieve effective delivery without the inflammation, irritation, and / or allergic reactions that often accompany the use of skin disrupting agents. As will be understood by those skilled in the art upon reading this specification, the present disclosure teaches that the transdermal delivery of such large drugs can be significantly improved by the use of microneedling technology, even when not loaded into, coated on, and / or manufactured as part of, the microneedles. Similarly, as will be understood by those skilled in the art upon reading this specification, the present disclosure teaches that the delivery of large drugs described herein can be significantly improved by the use of microneedling technology (particularly the use of MSCs), without leaving microneedles in the skin (e.g., by breaking them and / or otherwise maintaining and / or degrading them in situ). For example, one skilled in the art will appreciate that the provided technology can avoid challenges with long-term stability of large drugs necessary for commercially viable products, and can achieve effective delivery without inflammation, irritation, and / or allergic reactions that may result from skin disrupting agents and / or microneedles left in the skin. Indeed, in the Examples and elsewhere, the present disclosure expressly teaches that MSCs implemented with microneedles that do not contain botulinum toxin facilitate transdermal delivery of botulinum toxin from topical (e.g., creams, ointments) compositions, particularly from compositions that include macro- and nano-emulsions.

[0192] In some embodiments, the present disclosure teaches that particularly advantageous results are achieved when microneedling technology is combined with nanoemulsion compositions.In some embodiments, microneedling technology is combined with lotion, cream or liquid composition, which in turn can be or contain nanoemulsion compositions.In some embodiments, the provided technology does not utilize skin disrupting technology, such as chemical penetration enhancers.

[0193] The findings presented herein are particularly surprising in light of reports that transdermal delivery of solid nanoparticles of sizes comparable to those of the droplets in the nanoemulsion compositions utilized herein (e.g., 105±2.92 nm) effectively delivers (or enhances) even small molecule drugs transdermally across the skin. For example, Gomaa et al. described a study in which a solution of rhodamine dye (molecular weight 479 Da) encapsulated in PLGA nanoparticles was applied to skin preconditioned by microneedling and skin permeation was evaluated. See Gomaa, Y., et al, "Effect of microneedle treatment on the skin permeation of a nanoencapsulated dye." J Pharm Pharmacol. 2012 November; 64(11): 1592-1602. The data showed that very little dye began to penetrate the skin after 6 hours of continuous application; no significant increase in penetration was observed until the skin was continuously treated for 24 hours. The researchers explained that "there is an emerging consensus that NPs [nanoparticles] can be well deposited in hair follicles, but generally cannot penetrate the stratum corneum." Thus, prior to the present disclosure, one skilled in the art would have expected that the use of microneedling technology with nano-sized vehicles would not be able to effectively deliver even small molecule drugs (e.g., rhodamine dyes) transdermally; certainly, delivery of larger drugs would have been thought impossible. However, the present disclosure shows that microneedling can significantly enhance the transdermal delivery of larger drugs, especially when utilized in conjunction with nanoemulsion technology.

[0194] In particular, the present disclosure discloses that microneedling technology can improve transdermal delivery (e.g., of large drugs, especially from nanoemulsion compositions) when other disruptive agents (i.e., chemical penetration enhancers and other techniques that disrupt or puncture skin structures) are not utilized. Previous studies of transdermal delivery of drugs similar in size to botulinum toxin (i.e., about 150 kDa) using microneedles have reported that delivery fails unless further treatments are applied to disrupt the skin. For example, U.S. Patent Application Publication No. 2010 / 0196445 reports that botulinum toxin is not effectively delivered from pre-coated microneedles unless skin digestive enzymes are also applied so that skin structures are disrupted at the site of microneedling.

[0195] The present disclosure shows, inter alia, that microneedling techniques can achieve transdermal delivery (e.g., of large drugs, particularly from macroemulsion and nanoemulsion compositions) when no microneedle coating or loading is utilized and / or when the microneedles are not designed to be left in the skin. In particular, as previously noted, the present disclosure recognizes that such microneedle coating or loading may not be commercially viable due to the instability of at least the botulinum coating or loading material. For example, according to Johnson, E. et al., "Botulinum toxin is highly susceptible to denaturation due to surface denaturation, heat and alkaline conditions. Freeze-drying of botulinum toxin is the most economically sound and practical method of distributing the product in a form that is stable and easily used by clinicians." U.S. Patent No. 5,512,547. Also, as will be appreciated by those of skill in the art reading this specification, the techniques described herein have certain advantages, including the absence of the need for microneedles to be left in or associated with tissue. For example, one skilled in the art will appreciate that leaving microneedles in the skin risks skin irritation, inflammation, allergic reactions, and / or cosmetically undesirable scarring. In contrast to the present invention, techniques such as those described in US Patent Application Publication No. 2017 / 0209553 utilize microneedle arrays that are loaded with botulinum and designed to penetrate the microneedles into the skin (US Patent Nos. 2017 / 0209553 and 2016 / 0263362).

[0196] Despite the expectation that MSCs would only serve to enhance the transdermal delivery of small compounds, the present disclosure teaches that the already highly efficient transdermal delivery of large drugs by nanoemulsion compositions can be dramatically improved by combination with microneedle technology. Moreover, the present disclosure is particularly surprising considering that microneedle conditioning combined with encapsulation of small molecule drugs in solid nanoparticles provided only a small amount of penetration after 6 hours of administration, as explained by Gomaa, while penetration of the material was observed up to 12 hours after administration.

[0197] In some embodiments, the nanoemulsion formulation comprising a large drug may be administered in combination with microneedle conditioning with solid microneedles. In some embodiments, MSC of the site is performed prior to application of the nanoemulsion formulation comprising a large drug to the site (e.g., prior to the particular application and / or each application). In some embodiments, MSC of the site is performed after application of the nanoemulsion formulation comprising a large drug to the site. In some embodiments, MSC of the site and application of the nanoemulsion formulation comprising a large drug to the site occur substantially simultaneously. In some embodiments, the macroemulsion formulation comprising a large drug is not injected by one or more microneedles. In some embodiments, the microneedle is part of an array of microneedles. In some embodiments, the microneedle may be about 1 μm to about 4,000 μm long. In some embodiments, the microneedle may be about 1 μm to about 2,000 μm long. In some embodiments, the microneedle may be about 50 μm to about 400 μm long. In some embodiments, the microneedles may be from about 800 μm to about 1500 μm in length.

[0198] In some embodiments, the present invention utilizes nanoemulsion compositions that contain large drugs that are particularly effective and / or useful in medical settings, for example, for therapeutic purposes. In some embodiments, certain nanoemulsion compositions are particularly effective and / or useful for local administration of drugs to subjects in need thereof. In some embodiments, nanoemulsion compositions can contain one or more large drugs. Exemplary nanoemulsion compositions and methods of manufacture are described, for example, in WO 2012 / 103035, the disclosure of which is incorporated herein by reference in its entirety.

[0199] In some embodiments, the nanoemulsions can be formulated into compositions suitable for topical administration, hi some embodiments, compositions suitable for topical administration can be lotions, creams, powders, ointments, liniments, gels, or eye drops.

[0200] In some embodiments, the nanoemulsion formulation comprises water, medium chain triglycerides, polysorbate 80, methylparaben, and propylparaben. In some embodiments, the nanoemulsion formulation comprises water, medium chain triglycerides, and polysorbate 80. Exemplary premixes, which are not meant to be limiting, are provided in Table 2. [Table 3]

[0201] These compositions are particularly useful in that they can be used to deliver drugs to a subject in need thereof by topical and / or transdermal (e.g., by lotions, creams, powders, ointments, liniments, gels, eye drops, etc.) administration. In some embodiments, the provided cream and / or lotion formulations can be administered to a subject in need thereof by topical and / or transdermal (e.g., by lotions, creams, powders, ointments, liniments, gels, eye drops, etc.) administration. In some embodiments, the provided nanoemulsion compositions can be formulated into cream and / or lotion formulations. In some embodiments, the provided cream and / or lotion formulations comprising the nanoemulsion compositions can be useful and / or effective for topical administration to a subject. In some embodiments, the provided nanoemulsion compositions can be mixed with one or more cream components in a cream formulation (e.g., a provided cream formulation) and / or with a saline solution for the preparation of a pharmaceutical composition.

[0202] The present invention may include the recognition that emulsion compositions (e.g., macroemulsion and nanoemulsion compositions) may be formulated into cream and / or lotion formulations for administration to a subject. The present invention may include the recognition that the cream and / or lotion formulations provided may be particularly useful for formulating emulsions, such as those described herein, for administration to a subject. Exemplary nanoemulsion formulations, which are not meant to be limiting, are provided in Table 3. [Table 4]

[0203] In some embodiments, provided compositions comprise a mixture of a provided nanoemulsion composition and one or more pharma- ceutically acceptable excipients. In some embodiments, cream and / or lotion formulations comprise a mixture of a provided nanoemulsion composition and / or saline solution.

[0204] In some embodiments, the compositions provided comprise the nanoemulsion compositions provided. In some embodiments, the compositions provided are cream and / or lotion formulations. In some embodiments, the cream and / or lotion formulations provided comprise the nanoemulsion compositions. In some embodiments, the compositions comprise the nanoemulsion compositions provided but are not cream and / or lotion formulations. In some embodiments, the suitable compositions are formulated into cream and / or lotion but comprise the nanoemulsion compositions.

[0205] In some embodiments, provided compositions comprise a mixture of provided nanoemulsion compositions and one or more pharma- ceutically acceptable excipients, e.g., for topical and / or transdermal administration (e.g., via lotions, creams, powders, ointments, liniments, gels, eye drops, etc.).

[0206] In some embodiments, for nanoemulsion compositions containing known therapeutic agents and / or independently active biologically active agents, such nanoemulsion compositions are configured, constructed, and administered in combination with MSCs to deliver a sufficient amount of therapeutic agent to the desired target site (e.g., epidermal and / or dermal structures) to treat a condition or disorder. In some embodiments, the provided nanoemulsion compositions are configured and constructed (e.g., by drug selection and / or combination, composition structure, etc.) to achieve a desired therapeutic effect upon administration to the skin. In some embodiments, the provided nanoemulsion compositions are configured and constructed so as not to induce undesirable clinical effects inside and / or outside the desired site of action (e.g., skin surface, dermis, etc.). In some embodiments, the provided nanoemulsion compositions are configured, constructed, and administered in combination with MSCs to have a systemic effect.

[0207] In some embodiments, the compositions provided can be formulated and delivered in combination with MSCs to achieve systemic delivery; in some embodiments, the compositions provided can be formulated and / or delivered to achieve local, but not systemic, delivery.

[0208] The present disclosure specifically demonstrates effective and efficient delivery of therapeutic agents (particularly large biological drugs, such as botulinum toxin or antibody drugs) to the dermis using the compositions provided in combination with MSCs. For example, in some embodiments, the present invention provides methods that include administration of the compositions described herein without clinically significant side effects. By way of example, when local delivery is contemplated, clinically significant side effects include, but are not limited to, undesirable systemic side effects, damage to nerve tissues underlying the dermis (e.g., nerve paralysis), undesirable effects on muscles (e.g., muscle paralysis), and / or undesirable blood levels of the therapeutic agent. Exemplary formulations of botulinum nanoemulsion premixes, which are not meant to be limiting, are provided in Table 4. [Table 5]

[0209] Diseases, Disorders and Conditions The present invention provides techniques for treating and / or preventing various systemic or skin diseases, disorders or conditions. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders or conditions associated with sweat and / or sebaceous gland activity. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders or conditions associated with infection. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders or conditions associated with inflammation. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders or conditions associated with inflammation. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders or conditions associated with cancer. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders or conditions that are systemic. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders or conditions that are autoimmune. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders or conditions associated with the epidermis and / or dermis level of the skin.

[0210] In some embodiments, the present invention provides a method for treating acne, unwanted sweating, body odor, hyperhidrosis, bromhidrosis, chromhidrosis, rosacea, alopecia, psoriasis, actinic keratosis, eczematous dermatitis (e.g., atopic dermatitis, etc.), disorders of excessive sebum production (e.g., seborrhea, seborrheic dermatitis, etc.), burns, Raynaud's phenomenon, lupus erythematosus, hyperpigmentation disorders (e.g., melasma, etc.), hypopigmentation disorders (e.g., plaques, etc.), skin cancer (e.g., squamous cell skin carcinoma, basal cell skin carcinoma, etc.), skin infections (e.g., bacterial infections, viral infections, fungal infections, etc.), facial wrinkles (e.g., forehead, glabella, rhytid and / or periorbital areas, etc.), and skin disorders (e.g., dermal blemishes, ... In one embodiment, the present invention provides techniques for treating and / or preventing one or more of: facial wrinkles (including facial wrinkles), headaches, unsightly facial expressions (e.g., due to overactivity of the underlying facial musculature), neck wrinkles, hyperfunctional facial wrinkles, hyperkinetic facial wrinkles, platysma bands, neuromuscular diseases and conditions involving muscle spasms and / or contractures (including facial paralysis, cerebral palsy, blepharospasm, various forms of facial contractures), dystonia, prostatic hyperplasia, headaches, strabismus, hemifacial spasms, tremors, spasticity, such as those caused by multiple sclerosis, retroorbital muscles, various ophthalmologic and urological conditions (e.g., penile and / or bladder disorders), and / or combinations thereof.

[0211] In some embodiments, the present invention provides techniques for treating and / or preventing rheumatoid arthritis.In some embodiments, the present invention provides techniques for treating and / or preventing psoriatic arthritis.In some embodiments, the present invention provides techniques for treating and / or preventing osteoarthritis.

[0212] In some embodiments, the present invention provides techniques for treating and / or preventing lupus erythematosus. In some embodiments, the lupus erythematosus is systemic, discoid, drug-induced, or neonatal.

[0213] In some embodiments, the present invention provides techniques for treating and / or preventing Crohn's disease. In some embodiments, the present invention provides techniques for treating and / or preventing inflammatory bowel disease. In some embodiments, the present invention provides techniques for treating and / or preventing ulcerative colitis.

[0214] In some embodiments, the present invention provides techniques for treating and / or preventing pulmonary disorders. In some embodiments, the pulmonary disorder may be asthma or chronic obstructive pulmonary disorder.

[0215] In some embodiments, the present invention provides techniques for treating and / or preventing amyloidosis. In some embodiments, the amyloidosis is systemic or cutaneous.

[0216] In some embodiments, the present invention provides techniques for treating and / or preventing cancer. In some embodiments, the cancer is of the skin, blood, breast, colon or lung.

[0217] In some embodiments, the present invention provides techniques for treating and / or preventing dyslipidemia. In some embodiments, the dyslipidemia is hypercholesterolemia.

[0218] In some embodiments, the present invention provides techniques for treating and / or preventing an infection, hi some embodiments, the infection is or is caused by C. difficile or Staphylococcus aureus.

[0219] In some embodiments, the present invention provides techniques for treating and / or preventing pain. In some embodiments, the pain is associated with arthritis. In some embodiments, the arthritis is rheumatoid arthritis, psoriatic arthritis or osteoarthritis.

[0220] In some embodiments, the present invention provides techniques for treating and / or preventing a neurological condition. In some embodiments, the neurological condition is Alzheimer's disease, Parkinson's disease, or stroke.

[0221] In some embodiments, the invention comprises administration of at least one provided composition administered in combination with MSCs according to a dosing regimen sufficient to achieve a reduction in the severity and / or prevalence of an associated skin condition by at least about 20%; in some embodiments, according to a dosing regimen sufficient to achieve a reduction of at least about 25%; in some embodiments, according to a dosing regimen sufficient to achieve a reduction of at least about 30%; in some embodiments, according to a dosing regimen sufficient to achieve a reduction of at least about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, and / or administered according to a dosing regimen sufficient to achieve a reduction of 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90% or more.

[0222] In some embodiments, the invention comprises administration of at least one provided composition administered in combination with MSCs according to a dosing regimen sufficient to achieve a reduction in the severity and / or prevalence of an associated skin condition of at least about 20% in a particular percentage of a patient population to which the composition is administered; in some embodiments, according to a dosing regimen sufficient to achieve a reduction in the severity and / or prevalence of an associated skin condition of at least about 25% in a particular percentage of a patient population to which the composition is administered; in some embodiments, according to a dosing regimen sufficient to achieve a reduction in the severity and / or prevalence of an associated skin condition of at least about 30% in a particular percentage of a patient population to which the composition is administered; in some embodiments, according to a dosing regimen sufficient to achieve a reduction in the severity and / or prevalence of an associated skin condition of at least about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 9 About 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65 %, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90% or more reduction. In some embodiments, the particular percentage of the patient population to which the composition is administered is at least about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95% or about 100%. To give some examples, in some embodiments, the invention includes administration of at least one provided composition according to a dosing regimen sufficient to achieve a reduction in the severity and / or prevalence of the associated skin condition in at least about 20% of the patient population to which the composition is administered.In some embodiments, the invention involves administration of at least one provided composition according to a dosing regimen sufficient to achieve a reduction in the severity and / or prevalence of the associated skin condition in at least about 30% of a patient population to which the composition is administered.

[0223] The present invention provides techniques for treating and / or preventing skin diseases, comprising administering a composition in combination with MSCs to a subject suffering from, susceptible to, and / or exhibiting symptoms of a skin disease. In some embodiments, the compositions provided for treating a skin disease described herein are formulated for any route of administration described herein. In some embodiments, the compositions provided are formulated for topical administration. In some embodiments, the compositions provided are formulated as creams, liniments, lotions, gels, shampoos, conditioners, sunscreens, deodorants and / or antiperspirants (e.g., as roll-ons, solid sticks, gels, creams, aerosols, etc.), etc., depending on the condition being treated.

[0224] In some embodiments, such provided compositions are administered locally in combination with MSCs to affected areas (e.g., armpits, hands, feet, scalp, hair follicles, face, neck, back, arms, chest, legs, groin, groin, etc., depending on the particular condition being treated). In some embodiments, local administration is achieved by local administration in combination with MSCs.

[0225] Compositions and Formulations As described herein, the present invention provides and / or utilizes compositions comprising one or more large drugs for administration in combination with MSCs.In some embodiments, the provided compositions can be formulated for topical and / or transdermal delivery (e.g., as lotions, creams, liniments, ointments, powders, gels, eye drops, etc.).In some embodiments, the provided compositions can be or include nanoemulsions.In some embodiments, the provided compositions can be or include macroemulsions.

[0226] Formulations of the provided compositions may be prepared by any suitable method, e.g., known or hereafter developed in the art of pharmacology. Generally, such preparation methods include mixing the provided compositions with one or more additives, and then, if necessary and / or desired, shaping and / or packaging the composition in a suitable form for administration, e.g., single or multiple dosage units.

[0227] In some embodiments, the composition can be manufactured, packaged and / or sold in bulk as one single unit dose and / or multiple single unit doses.As used herein, "unit dose" refers to a discrete amount of pharmaceutical composition that contains a predetermined amount of the composition provided.The amount of the composition provided is generally equal to the dosage of the composition provided that is administered to subject, and / or a convenient fraction of such dosage, for example, half or one third of such dosage.

[0228] In some embodiments, additives suitable for use in the composition (e.g., pharma- ceutically and / or cosmetically acceptable composition) may include, for example, one or more additives, such as solvents, dispersion media, granulation media, diluents or other liquid vehicles, dispersion or suspension aids, surfactants and / or emulsifiers, isotonicity agents, thickening or emulsifying agents, preservatives, solid binders, lubricants, disintegrants, binders, preservatives, buffers, etc., as appropriate for the particular administration form desired. In some embodiments, additives such as cocoa butter and / or suppository wax, colorants, coating agents, sweeteners, flavors and / or fragrances may be used. Remington's The Science and Practice of Pharmacy, 21 st Edition, AR Gennaro (Lippincott, Williams & Wilkins, Baltimore, MD, 2005; incorporated herein by reference) discloses various additives used in the known art for the formulation and manufacture of pharmaceutical compositions.

[0229] In some embodiments, suitable additives (e.g., pharma- ceutically and / or cosmetically acceptable additives) are at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% pure. In some embodiments, the additives are approved by the U.S. Food and Drug Administration. In some embodiments, the additives are pharmaceutical grade. In some embodiments, the additives meet the standards of the United States Pharmacopoeia (USP), the European Pharmacopoeia (EP), the British Pharmacopoeia, and / or other international pharmacopoeias.

[0230] In some embodiments, provided compositions are formulated as cream, liniment, ointment, oil, foam, spray, lotion, liquid, powder, thickened lotion or gel (e.g., formulated for transdermal delivery as described herein).Specific exemplary such formulations can be prepared as, for example, cosmetic formulation products, such as skin softener, nutritional lotion type emulsion, cleansing lotion, cleansing cream, skin milk, emollient lotion, massage cream, emollient cream, makeup base, facial pack or facial gel, cleaner formulation, such as shampoo, rinse, body cleanser, hair tonic or soap, or skin composition, such as lotion, ointment, gel, cream, liniment, patch, deolant or spray.In some embodiments, the composition for topical administration is not formulated for administration to mucosa (e.g., it is not suitable for application to mucosa and / or is not formulated to deliver a suitable amount of large drug to or across mucosa).

[0231] Treatment site The technology of the present invention is suitable for both human and veterinary use. Subjects suffering from any disorder that would benefit from the topical application of an active agent may be treated with the disclosed technology for transdermal drug delivery.

[0232] Any site suitable for MSCs is a suitable administration site. In some embodiments, the administration site is the skin overlying a muscle or muscle group of the subject. In some embodiments, the site is hairless. In some embodiments, the site is on the torso. In some embodiments, the site is on the back. In some embodiments, the site is on the chest. In some embodiments, the site is on the buttocks. In some embodiments, the site is on the groin. In some embodiments, the site is on the groin. In some embodiments, the site is on the head. In some embodiments, the site is on the scalp. In some embodiments, the site is on the face. In some embodiments, the site is on the neck. In some embodiments, the site is on the décolleté. In some embodiments, the site is in the armpit. In some embodiments, the site is in the axilla. In some embodiments, the site is on the hand. In some embodiments, the site is on the foot. In some embodiments, the site is on the arm. In some embodiments, the site is on the leg. In some embodiments, the site is not a mucous membrane.

[0233] In some embodiments, the site is affected by skin disease.In some embodiments, the site is the skin covering the muscle or muscle group affected by neuromuscular condition.In some embodiments, the length of the microneedle used with MSC is adjusted based on the thickness of the skin at the treatment site.

[0234] Administration The present invention provides a technique for delivering emulsion compositions (e.g., botulinum emulsion compositions or antibody drug emulsion compositions) for various applications, including, for example, cosmetics, dietary supplements, and medical applications. Such emulsion compositions may contain one or more biologically active substances. In some embodiments, the emulsion composition contains a botulinum toxin. In some embodiments, the emulsion composition contains an antibody drug. In some embodiments, the emulsion composition is a nanoemulsion composition and / or a macroemulsion composition.

[0235] The present invention provides techniques for treating a condition or disorder using any of the provided compositions described herein (e.g., provided emulsion compositions; cream and / or lotion formulations; combinations of provided emulsion compositions and cream and / or lotion formulations, etc.) in combination with MSCs.

[0236] In some embodiments, such methods include administering the provided compositions in combination with MSCs to a patient suffering from and / or susceptible to a disease, condition, or disorder. In some embodiments, such methods include administering the provided nanoemulsion compositions in combination with MSCs described herein (e.g., using microneedles with a relatively low microneedle density and / or a relatively small microneedle puncture size) to a patient suffering from and / or susceptible to a disease, condition, or disorder associated with the dermis layer of the skin. In some embodiments, such methods include administering an emulsion composition comprising at least one known therapeutic agent and / or an independently active biologically active agent in combination with MSCs to a patient suffering from and / or susceptible to a disease, condition, or disorder. In some embodiments, such methods include administering an emulsion composition and / or at least one known therapeutic agent and / or an independently active biologically active agent in combination with MSCs, formulated with a provided cream and / or lotion formulation, to a patient suffering from and / or susceptible to a disease, condition, or disorder. In some embodiments, such methods include administering the composition in combination with MSCs by topical and / or transdermal administration (e.g., by lotion, cream, powder, ointment, liniment, gel, eye drops, etc.). Some embodiments further include administration of a penetration enhancer. Some embodiments further include administration of a non-irritating penetration enhancer.

[0237] In some embodiments, the present invention provides techniques for treating any condition or disorder.In some embodiments, the present invention shows that certain compositions described herein in combination with MSCs described herein can achieve efficient and specific controlled and / or improved delivery of active substances to biologically relevant target sites (e.g., specific tissues, locations in the skin, cells, etc.).In some embodiments, the present invention shows controlled delivery and / or achievement of therapeutic effects at specific biologically relevant target sites without significant side effects associated with delivery to other areas.

[0238] In some embodiments, the present invention provides improved techniques for treating conditions or disorders associated with epidermal and / or dermal structures (e.g., sweat glands, sebaceous glands, hair follicles, etc.). In some embodiments, the present invention demonstrates that the compositions provided herein (e.g., the nanoemulsion compositions provided; cream and / or lotion formulations; the combination of the nanoemulsion compositions provided with cream and / or lotion formulations, etc.) in combination with the MSCs described herein (e.g., using microneedles with a relatively low microneedle density and / or a relatively small microneedle puncture size) can improve efficient and specific active agent delivery and / or bioavailability to the dermis, and that the compositions provided herein can have a therapeutic effect upon administration to the skin of a subject. In some embodiments, the present invention demonstrates improved delivery and / or bioavailability through the dermis and / or achievement of a therapeutic effect without significant side effects associated with delivery to other areas (e.g., subdermal or extradermal structures and / or tissues other than the dermis). In some embodiments, the compositions provided herein (e.g., provided emulsion compositions; cream and / or lotion formulations; combinations of provided emulsion compositions and cream and / or lotion formulations, etc.) in combination with the MSCs described herein can improve transdermal delivery and / or bioavailability of active agents, such as therapeutic agents (e.g., botulinum toxin, antibody drugs, etc.).

[0239] The present invention provides techniques for treating a condition or disorder by administering to a patient a provided composition as described herein (e.g., a provided emulsion composition; a cream and / or lotion formulation; a combination of a provided emulsion composition and a cream and / or lotion formulation, etc.) in combination with MSCs as described herein (e.g., using microneedles having a relatively low microneedle density and / or a relatively small microneedle puncture size). In some embodiments, the present invention provides techniques for treating a condition or disorder by topically administering to a patient a composition comprising a provided emulsion composition in combination with MSCs as described herein.

[0240] In some embodiments, the large drug penetrates the skin within about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 minutes of administration. In some embodiments, the large drug penetrates the skin within about 5 to about 60 minutes of administration. In some embodiments, the large drug penetrates the skin within about 5 to about 12 minutes of administration. In some embodiments, the large drug penetrates the skin within about 5 to about 15 minutes of administration. In some embodiments, the large drug penetrates the skin within about 15 to about 30 minutes of administration. In some embodiments, the large drug penetrates the skin within about 1 hour of administration. In some embodiments, the large drug penetrates the skin within about 2 hours of administration. In some embodiments, the large drug penetrates the skin within about 3 hours of administration. In some embodiments, the large drug penetrates the skin within about 4 hours of administration. In some embodiments, the large drug penetrates the skin within about 5 hours of administration. In some embodiments, the large drug penetrates the skin within about 6 hours of administration.

[0241] In some embodiments, the large drug penetrates the layers of the skin within about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 minutes of administration. In some embodiments, the large drug penetrates the layers of the skin within about 5 to about 60 minutes of administration. In some embodiments, the large drug penetrates the layers of the skin within about 5 to about 12 minutes of administration. In some embodiments, the large drug penetrates the layers of the skin within about 5 to about 15 minutes of administration. In some embodiments, the large drug penetrates the layers of the skin within about 15 to about 30 minutes of administration. In some embodiments, the large drug penetrates the layers of the skin within about 1 hour of administration. In some embodiments, the large drug penetrates the layers of the skin within about 2 hours of administration. In some embodiments, the large drug penetrates the layers of the skin within about 3 hours of administration. In some embodiments, the large drug penetrates the layers of the skin within about 4 hours of administration. In some embodiments, the large drug penetrates the layers of the skin within about 5 hours of administration. In some embodiments, the large drug penetrates the layers of the skin within about 6 hours of administration.

[0242] In some embodiments, the large drug penetrates the top layer of the skin within about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 minutes of administration. In some embodiments, the large drug penetrates the top layer of the skin within about 5 to about 60 minutes of administration. In some embodiments, the large drug penetrates the top layer of the skin within about 5 to about 12 minutes of administration. In some embodiments, the large drug penetrates the top layer of the skin within about 5 to about 15 minutes of administration. In some embodiments, the large drug penetrates the top layer of the skin within about 15 to about 30 minutes of administration. In some embodiments, the large drug penetrates the top layer of the skin within about 1 hour of administration. In some embodiments, the large drug penetrates the top layer of the skin within about 2 hours of administration. In some embodiments, the large drug penetrates the top layer of the skin within about 3 hours of administration. In some embodiments, the large drug penetrates the top layer of the skin within about 4 hours of administration. In some embodiments, the large drug penetrates the top layer of the skin within about 5 hours of administration. In some embodiments, the large drug penetrates the top layer of the skin within about 6 hours of administration.

[0243] In some embodiments, the large drug penetrates the top layer of the skin, including the stratum corneum, skin pores, and / or skin glands, within about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 minutes of administration. In some embodiments, the large drug penetrates the top layer of the skin, including the stratum corneum, skin pores, and / or skin glands, within about 5 to about 60 minutes of administration. In some embodiments, the large drug penetrates the top layer of the skin, including the stratum corneum, skin pores, and / or skin glands, within about 5 to about 12 minutes of administration. In some embodiments, the large drug penetrates the top layer of the skin, including the stratum corneum, skin pores, and / or skin glands, within about 5 to about 15 minutes of administration. In some embodiments, the large drug penetrates the top layer of the skin, including the stratum corneum, skin pores, and / or skin glands, within about 15 to about 30 minutes of administration. In some embodiments, the large drug penetrates the top layer of the skin, including the stratum corneum, skin pores, and / or skin glands, within about 1 hour of administration. In some embodiments, the large drug penetrates the top layer of the skin, including the stratum corneum, skin pores and / or skin glands, within about 2 hours of administration. In some embodiments, the large drug penetrates the top layer of the skin, including the stratum corneum, skin pores and / or skin glands, within about 3 hours of administration. In some embodiments, the large drug penetrates the top layer of the skin, including the stratum corneum, skin pores and / or skin glands, within about 4 hours of administration. In some embodiments, the large drug penetrates the top layer of the skin, including the stratum corneum, skin pores and / or skin glands, within about 5 hours of administration. In some embodiments, the large drug penetrates the top layer of the skin, including the stratum corneum, skin pores and / or skin glands, within about 6 hours of administration.

[0244] kit In some embodiments, the present invention provides a pharmaceutical package or kit comprising one or more emulsion compositions according to the present invention and one or more microneedle devices.In some embodiments, the pharmaceutical package or kit comprises a formulation or pharmaceutical composition comprising the composition provided in one or more containers, which are optionally filled with one or more additional components of the pharmaceutical composition.In some embodiments, the pharmaceutical package or kit comprises an additional approved therapeutic agent for use in combination therapy (e.g., benzoyl peroxide for treating acne; aluminum compound for treating hyperhidrosis, etc.).In some embodiments, such container can optionally be associated with a notice in the form prescribed by a government agency that regulates the manufacture, use, or sale of pharmaceutical products, which notice reflects the approval by the agency of manufacture, use, or sale for human administration.

[0245] In some embodiments, a kit is provided that includes therapeutic reagent.As a non-limiting example, the composition provided can be provided as a topical formulation and can be administered as a treatment in combination with the use of microneedling device.Medicinal dosage or instructions for self-administration can be provided in the kit for administering to individuals suffering from or at risk of conditions or disorders, such as those related to the dermis level of the skin.

[0246] In some embodiments, the kit may include (i) a provided composition; and (ii) at least one pharma- ceutically acceptable excipient; and (iii) at least one device for microneedling of the skin; and (iv) instructions for use. In some embodiments, the at least one device has a relatively low density of microneedles (e.g., about 2 microneedles / cm). 2 ~About 50 microneedles / cm 2 In some embodiments, for example, at least one device may include microneedles having a relatively small microneedle puncture size (e.g., a puncture size of about 100 μm per microneedle).2 / Microneedle ~ approx. 30,000μm 2 / Microneedle range, puncture size per microneedle is approximately 100μm 2 / Microneedle ~ approx. 60,000μm 2 / microneedle range).

[0247] The present invention provides, inter alia, a technique for administering, improving transdermal delivery, and / or improving the bioavailability of large drugs, such as botulinum toxin or antibody drugs, by incorporating one or more large drugs into one or more emulsion compositions that are then administered in combination with MSCs as described herein.The inventors have surprisingly found that the transdermal penetration and bioavailability of botulinum toxin or antibody drugs incorporated into nanoemulsion compositions is dramatically improved when using microneedles or microneedle arrays with relatively low microneedle density or relatively small microneedle puncture size (e.g., puncture size per microneedle, cross-sectional area of ​​each microneedle) in combination with MSCs.The advantage of the present invention is the ability to administer such large drugs intradermally while minimizing irritation or damage to the skin.The use of other drugs or processes with emulsion compositions and MSCs is not necessarily excluded or required in all embodiments of the present invention.

[0248] Thus, the present invention provides techniques for administering large drugs by topical application of enhanced emulsion compositions (e.g., macroemulsion and / or nanoemulsion compositions) in combination with MSCs. In some embodiments, the large drug is a botulinum toxin. In some embodiments, the botulinum emulsion composition is applied prior to the MSCs directly to the skin and for absorption through the epithelial layer. In some embodiments, the botulinum emulsion composition is applied after the MSCs directly to the skin and for absorption through the epithelial layer. In some embodiments, the botulinum emulsion composition is applied substantially simultaneously with the MSCs directly to the skin and for absorption through the epithelial layer.

[0249] In some embodiments, the botulinum emulsion composition in combination with MSCs can penetrate the top layers of the skin, including the stratum corneum, skin pores, and / or skin glands, without the use of penetration enhancers. In some embodiments, the botulinum emulsion composition in combination with MSCs can penetrate the top layers of the skin, including the stratum corneum, skin pores, and / or skin glands, without the use of disintegrating, irritating, and / or abrasive agents.

[0250] In some embodiments, the antibody drug emulsion composition in combination with MSCs can penetrate the top layer of the skin, including the stratum corneum, skin pores and / or skin glands, without the use of a penetration enhancer. In some embodiments, the large drug is an antibody drug. In some embodiments, the antibody drug emulsion composition is applied directly to the skin and for absorption through the epidermal layer before the MSCs. In some embodiments, the antibody drug emulsion composition is applied directly to the skin and for absorption through the epidermal layer after the MSCs. In some embodiments, the antibody drug emulsion composition is applied directly to the skin and for absorption through the epidermal layer substantially simultaneously with the MSCs. In some embodiments, the antibody drug emulsion composition is applied directly to the skin and for absorption through the epidermal layer for systemic absorption.

[0251] In some embodiments, the antibody drug emulsion composition may penetrate the uppermost layers of the skin, including the stratum corneum, skin pores, and / or skin glands, without the use of disintegrating, irritating, and / or abrasive agents.

[0252] It is understood by those skilled in the art that the composition of the present invention for topical administration can have cosmetic formulation products, such as skin softener, nutritional lotion type emulsion, cleansing lotion, cleansing cream, skin milk, emollient lotion, massage cream, emollient cream, makeup base, facial pack or facial gel, cleaner formulation, such as shampoo, rinse, body cleanser, hair tonic or soap, or skin composition, such as lotion, ointment, gel, cream, patch or spray.In some embodiments, the composition for topical administration is not formulated for administration to mucosa (e.g., it is not suitable for application to mucosa and / or is not formulated to deliver a suitable amount of large drug to or across mucosa).

[0253] Those skilled in the art will understand that the units herein refer to units that are biologically equivalent or bioactively equivalent to units defined by commercial manufacturers of botulinum toxin.

[0254] In some embodiments, the therapeutic effect of the botulinum toxin administered in accordance with the present invention may last as long as the effect of the injected solution. In some embodiments, the effect of the injected solution may last up to about 6-7 months. In some embodiments, the therapeutic effect of the botulinum toxin administered in accordance with the present invention may last up to 6-7 months. In some embodiments, the use of a synthetic polymer carrier capable of holding and slowly releasing the botulinum toxin may extend the effect for up to 5 years (U.S. Patent No. 6,312,708).

[0255] In some embodiments, the present invention provides a topical formulation of botulinum toxin that avoids potential complications, including but not limited to systemic toxicity or botulism poisoning. In some embodiments, the dosage of botulinum toxin (including botulinum types A, B, C, D, E, F or G, or botulinum genetically engineered or chemically modified to act longer or shorter than botulinum toxin serotype A) can range from as low as about 1 unit to as high as about 50,000 units, while minimizing the risk of adverse side effects. The specific dosage can vary depending on the condition being treated and the therapeutic regimen being utilized. For example, treatment of subcutaneous overactive muscles may require a high transdermal dosage (e.g., 1000 units to 20,000 units) of botulinum toxin. In comparison, treatment of neurogenic inflammation or overactive sweat glands may require a relatively low transdermal dosage (e.g., about 1 unit to about 1,000 units) of botulinum toxin.

[0256] Some embodiments of the present invention contemplate pharmaceutical compositions comprising stabilized botulinum toxins for transdermal delivery to a human patient. The botulinum toxins include botulinum toxins A, B, C, and the like. 1 The botulinum toxin may be selected from types A, B, C, D, E, F, and G, isolated and / or purified (i.e., about 150 kDa), and natural or recombinantly produced botulinum toxins. In some embodiments, the composition may contain from about 1 unit to about 50,000 units of a botulinum toxin, and the composition may contain a sufficient amount of a botulinum toxin to achieve a therapeutic effect lasting from one month to five years.

[0257] In some embodiments, the present invention provides topical formulations of botulinum toxin (e.g., botulinum emulsion compositions) that are capable of penetrating the skin of a subject without the botulinum toxin penetrating blood vessels in significant amounts. For example, in some embodiments of the present invention, less than about 25%, less than about 20%, less than about 15%, less than about 10%, less than about 5%, less than about 4%, less than about 3%, less than about 2%, or less than about 1% of the botulinum toxin present in the pharmaceutical composition penetrates blood vessels upon application of the topical and / or transdermal formulations of the present invention.

[0258] In some embodiments, the present invention provides topical formulations of antibody drugs (e.g., antibody drug emulsion compositions) in which the antibody drug can penetrate the skin of a subject without penetrating the blood vessels in significant amounts. For example, in some embodiments of the present invention, less than about 25% or less than about 5% of the antibody drug present in the pharmaceutical composition penetrates the blood vessels upon application of the topical and / or transdermal formulations of the present invention.

[0259] In some embodiments, the present invention provides a topical formulation of an antibody drug (e.g., an antibody drug emulsion composition) in which the antibody drug can penetrate the skin of a subject and penetrate the blood vessels in a significant amount. In some embodiments, the present invention provides a topical formulation of an antibody drug (e.g., an antibody drug emulsion composition) in which the antibody drug can penetrate the skin of a subject and penetrate the blood vessels in a therapeutically effective amount. For example, in some embodiments of the present invention, more than about 25%, 50%, 75%, 90% or 95% of the antibody drug present in the pharmaceutical composition penetrates the blood vessels upon application of the topical and / or transdermal formulation of the present invention. In some embodiments, the present invention provides a topical formulation of an antibody drug (e.g., an antibody drug emulsion composition) in which the antibody drug can have a systemic effect on the subject.

[0260] One of skill in the art will appreciate that the compositions of the invention that achieve transdermal administration of a botulinum toxin or antibody drug can be incorporated into a device, such as a patch, roller, pen, stamp, or the like. EXAMPLES

[0261] Example 1: Effect of Microneedle Skin Conditioning (MSC) Pretreatment on Botulinum Toxin Bioavailability A single dose topical study of the bioavailability of botulinum toxin following topical administration of botulinum nanoemulsion was conducted. Prior to application of the topical treatment, the skin was conditioned with a microneedle array by pressing the array against the skin. The array was then removed prior to application of the botulinum treatment. The study was designed to not only evaluate but also compare the % bioavailability achieved with various microneedle systems, thereby testing the effect of varying the density of needles in the microneedle array and the puncture size of the microneedles (e.g., puncture area per microneedle) on the bioavailability of botulinum.

[0262] The study included four test groups, each consisting of eight rats. Each group was subjected to microneedle skin conditioning as described above. Except for the control group, each group was topically treated once with a volume and concentration of botulinum nanoemulsion on the skin covering the biceps femoris, gastrocnemius, and tibialis anterior of the right hind leg. The nanoemulsion was applied with a gloved finger. Administration of the topical formulation to the skin took approximately 10 minutes, at which point the topical formulation was completely absorbed into the skin. It is also known that a dose of botulinum administered at a sufficient concentration can induce the death of animals, and therefore the effect of such treatment was measured by mortality. Thus, the mortality rates of the four treatment groups were compared. Specifically, the mortality rate at 3 days after treatment was used as the relative bioavailability of botulinum toxin, where an increase in mortality represents an increase in the bioavailability of the toxin.

[0263] Table 5 describes the characteristics of each type of microneedle used in each treatment, including needle density, needle length, and the puncture size each needle makes in the skin prior to treatment (the puncture size of the microneedles described herein). Table 5 also details the mortality rate for each group.

[0264] As can be seen, significant bioavailability was achieved with each microneedling approach (ie, in each of Groups A, B and C).

[0265] Moreover, by comparing group C with either group A or B, it is surprisingly observed that fewer and smaller puncture holes result in higher bioavailability than more and larger puncture holes.That is, reducing the density of microneedles increases bioavailability, as does reducing the puncture size of microneedles.By comparing group A and group B with each other, it is surprisingly observed that increasing the length of microneedles does not improve bioavailability.

[0266] That is, the results of the test determined that the density of the microneedles and the puncture size of the microneedles can significantly increase the bioavailability of botulinum when such botulinum is included in a nanoemulsion and applied to the skin after microneedle skin conditioning as described herein. Specifically, when a microneedle array is used to condition the skin prior to topical administration of a macromolecule in an emulsion composition, the needle density of the microneedle array can be increased to 31 needles / cm. 2 In addition, when a microneedle array is used to condition the skin prior to topical administration of a macromolecule in an emulsion composition, the needle puncture size (per microneedle) of the microneedle array is reduced to less than 36,000 μm per microneedle, which increases the bioavailability of the macromolecule. 2 Reducing the dose to less than 100 mg increased the bioavailability of the macromolecule. [Table 6]

[0267] Example 2: Effect of MSC preconditioning on the bioavailability of botulinum toxin in humans: Effect on sweat reduction A single-dose topical study of the bioavailability of botulinum toxin following topical administration of a topical botulinum nanoemulsion formulation in humans is conducted. The study is designed to not only evaluate but also compare the % bioavailability achieved using various microneedle systems, thereby testing the effect of varying needle density of the microneedle array on enhancing the bioavailability of botulinum in humans by measuring reduction in skin sweating following topical treatment with a botulinum nanoemulsion formulation.

[0268] One subject was included in the study. Each was located in the abdomen, each measuring approximately 2 cm 2 Three spots, each 5 cm apart from each other, are selected and marked with a marker. Each spot is topically treated once with a volume of the botulinum nanoemulsion formulation, which is a fixed concentration of botulinum. Administration of the topical formulation to the skin takes approximately 5 minutes, at which point the topical formulation is completely absorbed by the skin. The first spot is a control site, with no preconditioning with the microneedle array. The second spot is a control site, with 9 microneedles / cm prior to application of the botulinum formulation. 2 The third spot was preconditioned with three impressions of a microneedle array with a microneedle density of 85 microneedles / cm before application of the botulinum formulation, which is intervention site #1. 2 and intervention site #2, which was preconditioned with three impressions of a microneedle array with a microneedle density of 0.01 μm.

[0269] The expected effect of such treatment is a reduction in sweating at the site of botulinum nanoemulsion treatment. Sweating at the treatment site is measured in two ways: 1) an evaporometer test, where an instrument used to measure the rate of water evaporation from the skin is used to detect the rate of sweating (with increased sweating, more evaporation is detected); or 2) an iodine starch test, where povidone-iodine is applied to the treatment site of the subject, it is allowed to dry, and cornstarch is sprinkled on the treatment site, and when the subject sweats into the white cornstarch it turns purple and when the subject does not sweat it remains white, which is called the iodine starch test. For either sweat detection method, the subject goes under a heat lamp to induce sweating, and then the sweat detection method is used.

[0270] Sweat detection methods are used at baseline prior to botulinum nanoemulsion treatment, 2 weeks after treatment, and 4 weeks after treatment. The study found that at baseline, the mean amount of sweat detected by either the evaporometer test or the starch iodine test was approximately equal at the control and intervention sites. At 2 and 4 weeks after treatment, the mean amount of sweat detected at the control site by either the evaporometer test or the starch iodine test was greater than that detected at the intervention site at these post-treatment weeks. The study also found that at 2 and 4 weeks after treatment, the mean amount of sweat detected at intervention site #2 by either the evaporometer test or the starch iodine test was greater than that detected at intervention site #1 at these post-treatment weeks.

[0271] This study demonstrated that microneedle preconditioning using a relatively lower microneedle density unexpectedly increased the bioavailability of a topical large drug nanoemulsion containing botulinum toxin.

[0272] Example 3: Effect of MSC preconditioning with varying microneedle densities on botulinum toxin bioavailability in humans: Effect on sweat reduction A single-dose topical study of the bioavailability of botulinum toxin following topical administration of a topical botulinum nanoemulsion formulation in humans is conducted. The study is designed to not only evaluate but also compare the % bioavailability achieved using various microneedle systems, thereby testing the effect of varying the density of needles in a microneedle array on significantly enhancing the bioavailability of botulinum in humans by measuring the reduction in skin sweating following topical treatment with a botulinum nanoemulsion formulation.

[0273] Twelve subjects were included in the study, each with a 10 cm2 area on their backs. 2 Three spots, each 5 cm apart from each other, are selected and marked with a marker. Each spot is topically treated once with a volume of the botulinum nanoemulsion formulation, which is a fixed concentration of botulinum. Administration of the topical formulation to the skin takes approximately 5 minutes, at which point the topical formulation is completely absorbed by the skin. The first spot is a control site, with no preconditioning with the microneedle array. The second spot is a control site, with 9 microneedles / cm prior to application of the botulinum formulation. 2 The third spot was preconditioned with five impressions of a microneedle array with a microneedle density of 85 microneedles / cm before application of the botulinum formulation, which is intervention site #1. 2 and intervention site #2, which was preconditioned with five impressions of a microneedle array with a microneedle density of 0.01 μm.

[0274] The expected effect of such treatment is a reduction in sweating at the site of botulinum nanoemulsion treatment. The amount of sweating at the treatment site is measured in two ways: 1) an evaporometer test, where an instrument used to measure the rate of water evaporation from the skin is used to detect the rate of sweating (with increased sweating, more evaporation is detected); or 2) an iodine starch test, where povidone-iodine is applied to the treatment site of the subject, it is allowed to dry, and cornstarch is sprinkled on the treatment site, and when the subject sweats into the white cornstarch it turns purple and when the subject does not sweat it remains white, which is called the iodine starch test. For either sweat detection method, the subject enters a sauna to induce sweating, and then the sweat detection method is used.

[0275] Sweat detection methods are used at baseline prior to botulinum nanoemulsion treatment, 2 weeks after treatment, and 4 weeks after treatment. The study found that at baseline, the mean amount of sweat detected by either the evaporometer test or the starch iodine test was approximately equal at the control and intervention sites. At 2 and 4 weeks after treatment, the mean amount of sweat detected at the control site by either the evaporometer test or the starch iodine test was greater than that detected at the intervention site at these post-treatment weeks. The study also found that at 2 and 4 weeks after treatment, the mean amount of sweat detected at intervention site #2 by either the evaporometer test or the starch iodine test was greater than that detected at intervention site #1 at these post-treatment weeks.

[0276] This study demonstrated that microneedle preconditioning using a relatively lower microneedling density unexpectedly increased the bioavailability of a topical large drug nanoemulsion containing botulinum toxin.

[0277] Example 4: Effect of MSC preconditioning with varying microneedle puncture size on the bioavailability of botulinum toxin in humans: Effect on sweat reduction A single-dose topical study of the bioavailability of botulinum toxin following topical administration of a topical botulinum nanoemulsion formulation in humans is conducted. The study is designed to not only evaluate but also compare the % bioavailability achieved with various microneedle systems, thereby testing the effect of varying microneedle puncture size (e.g., puncture area per microneedle) on significantly enhancing the bioavailability of botulinum in humans by measuring reduction in skin sweating following topical treatment with a botulinum nanoemulsion formulation.

[0278] Twelve subjects were included in the study, each with a 10 cm2 area on their backs. 2 Three spots, each 5 cm apart from each other, are selected and marked with a marker. Each spot is topically treated once with a volume of the botulinum nanoemulsion formulation, which is a fixed concentration of botulinum. Administration of the topical formulation to the skin takes approximately 5 minutes, at which point the topical formulation is completely absorbed by the skin. The first spot is a control site, with no preconditioning with the microneedle array. The second spot is a control site, with approximately 11,000 μm of the botulinum formulation prior to application. 2 The first spot was preconditioned with five impressions of the microneedle array of microneedle puncture size / microneedle, which is intervention site #1. The third spot was approximately 60,000 μm before application of the botulinum formulation. 2 / preconditioned with five impresses of a microneedle array of microneedle puncture size, intervention site #2.

[0279] The expected effect of such treatment is a reduction in sweating at the site of botulinum nanoemulsion treatment. The amount of sweating at the treatment site is measured in two ways: 1) an evaporometer test, where an instrument used to measure the rate of water evaporation from the skin is used to detect the rate of sweating (with increased sweating, more evaporation is detected); or 2) an iodine starch test, where povidone-iodine is applied to the treatment site of the subject, it is allowed to dry, and cornstarch is sprinkled on the treatment site, and when the subject sweats into the white cornstarch it turns purple and when the subject does not sweat it remains white, which is called the iodine starch test. For either sweat detection method, the subject enters a sauna to induce sweating, and then the sweat detection method is used.

[0280] Sweat detection methods are used at baseline prior to botulinum nanoemulsion treatment, 2 weeks after treatment, and 4 weeks after treatment. The study found that at baseline, the mean amount of sweat detected by either the evaporometer test or the starch iodine test was approximately equal at the control and intervention sites. At 2 and 4 weeks after treatment, the mean amount of sweat detected at the control site by either the evaporometer test or the starch iodine test was greater than that detected at the intervention site at these post-treatment weeks. The study also found that at 2 and 4 weeks after treatment, the mean amount of sweat detected at intervention site #2 by either the evaporometer test or the starch iodine test was greater than that detected at intervention site #1 at these post-treatment weeks.

[0281] This study demonstrated that microneedle preconditioning using a relatively smaller microneedle puncture size unexpectedly increased the bioavailability of a topical large drug nanoemulsion containing botulinum toxin.

[0282] Example 5: Effect of MSCs on the bioavailability of botulinum toxin in humans: Effect on sweat and wrinkle reduction A single dose topical study of the bioavailability of botulinum toxin following topical administration of a topical botulinum nanoemulsion formulation in humans is conducted. The study is designed to not only evaluate but also compare the % bioavailability achieved with various microneedle systems, thereby testing the effect of varying needle density of the microneedle array on improving the bioavailability of botulinum in humans by measuring reduction in skin sweating and reduction in wrinkling following topical treatment with a botulinum nanoemulsion formulation.

[0283] One subject with severe forehead (or side) wrinkles on the forehead was included in the study. Each was located on the subject's forehead, each measuring approximately 2 cm in area. 2 Three spots, each 5 cm apart from each other, are selected and marked with a marker. Each spot is topically treated once with a volume of the botulinum nanoemulsion formulation, which is a fixed concentration of botulinum. Administration of the topical formulation to the skin takes approximately 5 minutes, at which point the topical formulation is completely absorbed by the skin. The first spot is a control site, with no preconditioning with the microneedle array. The second spot is a control site, with 9 microneedles / cm prior to application of the botulinum formulation. 2 The third spot was preconditioned with five impressions of a microneedle array with a microneedle density of 85 microneedles / cm before application of the botulinum formulation, which is intervention site #1. 2 and intervention site #2, which was preconditioned with five impressions of a microneedle array with a microneedle density of 0.01 μm.

[0284] The expected effect of such treatment is a reduction in sweating at the site of botulinum nanoemulsion treatment. Sweating at the treatment site is measured in two ways: 1) an evaporometer test, where an instrument used to measure the rate of water evaporation from the skin is used to detect the rate of sweating (with increased sweating, more evaporation is detected); or 2) an iodine starch test, where povidone-iodine is applied to the treatment site of the subject, it is allowed to dry, and cornstarch is sprinkled on the treatment site, and when the subject sweats into the white cornstarch it turns purple and remains white if the subject does not sweat, which is called the iodine starch test. For either sweat detection method, the subject enters a sauna to induce sweating, and then the method is used.

[0285] The predicted effect of botulinum nanoemulsion treatment is a reduction in forehead wrinkles at the botulinum nanoemulsion treatment sites. The severity of wrinkles is measured using a 4-point wrinkle scale (wrinkle scale): 0=none, 1=mild, 2=moderate, 3=severe.

[0286] Sweat detection methods are used at baseline prior to botulinum nanoemulsion treatment, 2 weeks after treatment, and 4 weeks after treatment. The study found that at baseline, the mean amount of sweat detected by either the evaporometer test or the starch iodine test was approximately equal between the control and intervention sites. At 2 and 4 weeks after treatment, the mean amount of sweat detected by either the evaporometer test or the starch iodine test was greater than that detected at the intervention sites at these post-treatment weeks. The study also found that at 2 and 4 weeks after treatment, the mean amount of sweat detected by either the evaporometer test or the starch iodine test was greater than that detected at intervention site #1 at these post-treatment weeks. The study found that at baseline, the mean severity of forehead wrinkles measured by the wrinkle scale was approximately equal between the control and intervention sites. At 2 and 4 weeks after treatment, the mean severity of forehead wrinkles measured by the wrinkle scale was greater than that detected at the intervention sites at these post-treatment weeks. The study also found that at 2 and 4 weeks post-treatment, the average severity of forehead wrinkles measured by the Wrinkle Scale at intervention site #2 was greater than that detected at intervention site #1 at these post-treatment weeks.

[0287] This study demonstrated that microneedle preconditioning using a relatively lower microneedle density unexpectedly increased the bioavailability of a topical large drug nanoemulsion containing botulinum toxin.

[0288] Example 6: Effect of MSCs with varying microneedle densities on the bioavailability of botulinum toxin in humans: Effect on sweat reduction in hyperhidrosis subjects A single-dose topical study of the bioavailability of botulinum toxin following topical administration of a topical botulinum nanoemulsion formulation in humans is conducted. The study is designed to not only evaluate but also compare the % bioavailability achieved using various microneedle systems, thereby testing the effect of varying needle density of the microneedle array on enhancing the bioavailability of botulinum in humans by measuring reduction in skin sweating following topical treatment with a botulinum nanoemulsion formulation.

[0289] The study included three treatment groups of 12 subjects each with axillary hyperhidrosis, characterized by excessive underarm sweating: Group 1 was the control group, in which botulinum nanoemulsion was applied to each subject's underarm; Group 2 was intervention group #1, in which 9 microneedles / cm were applied to each area of ​​underarm skin prior to application of the botulinum nanoemulsion formulation. 2 Group 3 was the intervention group #1, in which each area of ​​underarm skin was preconditioned with five impressions of a microneedle array with a microneedle density of 85 microneedles / cm2 before application of the botulinum nanoemulsion formulation. 2 The skin is preconditioned with five impressions of a microneedle array with a microneedle density of 0.01 μg / ml. Each subject in Groups 1, 2, and 3 is topically treated once with a volume of a botulinum nanoemulsion formulation at a fixed concentration of botulinum. Administration of the topical formulation to the skin takes approximately 5 minutes, at which point the topical formulation is completely absorbed by the skin.

[0290] The expected effect of such treatment is a reduction in sweating at the site of botulinum nanoemulsion treatment, which is the armpit. The amount of sweating at the treated site is measured by gravimetric measurement of sweating (GS test): the subject's armpit is dried with a paper towel; a filter paper is weighed; the filter paper is applied to the armpit for 5 minutes and then reweighed. The excess weight after reweighing the paper is the weight of sweat produced by the subject during the 5 minutes. The severity of the subject's hyperhidrosis condition is measured by the subject using the Hyperhidrosis Sweat Severity Scale (HDSS), in which a 4-point scale is rated by the subject: 0=none, 1=mild, 2=moderate, 3=severe.

[0291] The GS test and HDSS are used at baseline prior to botulinum nanoemulsion treatment, 2 weeks after treatment, and 4 weeks after treatment. The study found that at baseline, the mean amount of sweat detected by the GS test or disease severity measured by the HDSS was approximately equal in groups 1-3. At 2 and 4 weeks after treatment, the mean amount of sweat detected or disease severity in group 1 was greater than that detected in groups 2 or 3 at these post-treatment weeks. The study found that at 2 and 4 weeks after treatment, the mean amount of sweat detected or disease severity in group 3 was greater than that detected in group 2 at these post-treatment weeks.

[0292] This study demonstrated that microneedle preconditioning using a relatively lower microneedle density unexpectedly increased the bioavailability of a topical large drug nanoemulsion containing botulinum toxin.

[0293] Example 7: Effect of MSCs with varying microneedle puncture size on the bioavailability of botulinum toxin in humans: Effect on sweat reduction in subjects with hyperhidrosis A single-dose topical study of the bioavailability of botulinum toxin following topical administration of a topical botulinum nanoemulsion formulation in humans is conducted. The study is designed to not only evaluate but also compare the % bioavailability achieved using various microneedle systems, thereby testing the effect of varying microneedle puncture size on enhancing the bioavailability of botulinum in humans by measuring reduction in skin sweating following topical treatment with a botulinum nanoemulsion formulation.

[0294] The study included three treatment groups of 12 subjects each with axillary hyperhidrosis, characterized by excessive sweating in the armpits: Group 1 was the control group, in which botulinum nanoemulsion was applied to each subject's armpits; Group 2 was intervention group #1, in which approximately 11,000 μm of botulinum nanoemulsion was applied to each area of ​​underarm skin prior to application of the botulinum nanoemulsion formulation. 2Group 3 was the intervention group #1, which was preconditioned with 5 intrusions of a microneedle array of microneedle puncture size of about 60,000 μm on each part of the underarm skin before application of the botulinum nanoemulsion formulation. 2 The skin is preconditioned with five impressions of a microneedle array of microneedle puncture size. Each subject in groups 1, 2, and 3 is topically treated once with a volume of a botulinum nanoemulsion formulation at a fixed concentration of botulinum. Administration of the topical formulation to the skin takes approximately 5 minutes, at which point the topical formulation is completely absorbed by the skin.

[0295] The expected effect of such treatment is a reduction in sweating at the site of botulinum nanoemulsion treatment, which is the armpit. The amount of sweating at the treated site is measured by gravimetric measurement of sweating (GS test): the subject's armpit is dried with a paper towel; a filter paper is weighed; the filter paper is applied to the armpit for 5 minutes and then reweighed. The excess weight after reweighing the paper is the weight of sweat produced by the subject during the 5 minutes. The severity of the subject's hyperhidrosis condition is measured by the subject using the Hyperhidrosis Sweat Severity Scale (HDSS), in which a 4-point scale is rated by the subject: 0=none, 1=mild, 2=moderate, 3=severe.

[0296] The GS test and HDSS are used at baseline prior to botulinum nanoemulsion treatment, 2 weeks after treatment, and 4 weeks after treatment. The study found that at baseline, the mean amount of sweat detected by the GS test or disease severity measured by the HDSS was approximately equal in groups 1-3. At 2 and 4 weeks after treatment, the mean amount of sweat detected or disease severity in group 1 was greater than that detected in groups 2 or 3 at these post-treatment weeks. The study found that at 2 and 4 weeks after treatment, the mean amount of sweat detected or disease severity in group 3 was greater than that detected in group 2 at these post-treatment weeks.

[0297] This study demonstrated that microneedle preconditioning using a relatively smaller microneedle puncture size unexpectedly increased the bioavailability of a topical large drug nanoemulsion containing botulinum toxin.

[0298] Example 8: Effect of MSCs on the bioavailability of botulinum toxin in humans: Effect on reduction of crow's feet wrinkles A single-dose topical study of the bioavailability of botulinum toxin following topical administration of a topical botulinum nanoemulsion formulation in humans was conducted. The study was designed to not only evaluate but also compare the % bioavailability achieved using various microneedle systems, thereby testing the effect of varying needle density of the microneedle array on improving the bioavailability of botulinum in humans by measuring reduction in skin wrinkling following topical treatment with a botulinum nanoemulsion formulation.

[0299] The study included one subject with severe crow's feet beside the eyes. Botulinum nanoemulsion was applied to the subject's crow's feet. The dose of botulinum applied to the skin was approximately 15% of the effective dose when botulinum nanoemulsion was applied without microneedle skin preconditioning. The effective dose was defined as the dose that produced at least a 2 point improvement in the appearance of wrinkles when the subject contracted the muscles that cause crow's feet as measured by a 5 point wrinkle rating scale. The subject received 9 microneedles / cm2 of botulinum nanoemulsion formulation on each area of ​​skin where the crow's feet are located prior to application of the botulinum nanoemulsion formulation to side #1 of the face. 2 The skin was preconditioned with five impressions of a microneedle array with a microneedle density of 85 microneedles / cm2 onto each section of skin where the crow's feet are located prior to application of the botulinum nanoemulsion formulation to the other side of the face (Side #2). 2 The skin was preconditioned with five impressions of a microneedle array having a microneedle density of 0.01 μg / ml. Application of the topical formulation to the skin took approximately 5 minutes, at which point the topical formulation was completely absorbed into the skin.

[0300] The predicted effect of botulinum nanoemulsion treatment is a reduction in crow's feet wrinkles at the botulinum nanoemulsion treatment sites. Wrinkle severity was measured using a 5-point wrinkle scale (Wrinkle Scale): 0=none, 1=minimal, 2=mild, 3=moderate, 4=severe.

[0301] The wrinkle scale was used at baseline prior to botulinum nanoemulsion treatment, 2 weeks after treatment, and 4 weeks after treatment. The study found that at baseline, subjects had severe wrinkles as assessed by the wrinkle scale, with a score of 4 on a 5-point wrinkle scale. At 2 weeks after treatment, the mean severity of wrinkles was reduced by 1 point on the wrinkle scale, with a score of 3 (moderate) on side #2 of the face. At 4 weeks after treatment, the mean severity of wrinkles was reduced by 2 points on the wrinkle scale, with a score of 2 (mild) on side #2 of the face. The study also found that at 2 weeks after treatment, the mean severity of wrinkles was reduced by 2 points on the wrinkle scale, with a score of 2 (mild) on side #1 of the face. At 4 weeks after treatment, the mean severity of wrinkles was reduced by 3 points on the wrinkle scale, with a score of 1 (minimal) on side #1 of the face.

[0302] This study demonstrated that microneedle preconditioning using a relatively lower microneedle density unexpectedly increased the bioavailability of a topical large drug nanoemulsion containing botulinum toxin.

[0303] Example 9: Effect of MSCs with varying microneedle densities on the bioavailability of botulinum toxin in humans: Effect on reduction of crow's feet wrinkles A single dose topical study of the bioavailability of botulinum toxin following topical administration of a topical botulinum nanoemulsion formulation in humans is conducted. The study is designed to not only evaluate but also compare the % bioavailability achieved using various microneedle systems, thereby testing the effect of varying needle density of the microneedle array on enhancing the bioavailability of botulinum in humans by measuring reduction in skin wrinkling following topical treatment with a botulinum nanoemulsion formulation.

[0304] The study included three treatment groups of 12 subjects each with severe crow's feet around the eyes: Group 1 was the control group, in which botulinum nanoemulsion was applied to each subject's crow's feet; Group 2 was the intervention group #1, in which 9 microneedles / cm were applied to each area of ​​skin where the crow's feet were located prior to application of the botulinum nanoemulsion formulation. 2 Group 3 is the intervention group #1, which was preconditioned with five impressions of a microneedle array with a microneedle density of 85 microneedles / cm2 on each area of ​​skin where the crow's feet are located prior to application of the botulinum nanoemulsion formulation. 2 The skin is preconditioned with five impressions of a microneedle array with a microneedle density of 0.01 μg / ml. Each subject in Groups 1, 2, and 3 is topically treated once with a volume of a botulinum nanoemulsion formulation at a fixed concentration of botulinum. Administration of the topical formulation to the skin takes approximately 5 minutes, at which point the topical formulation is completely absorbed by the skin.

[0305] The predicted effect of botulinum nanoemulsion treatment is a reduction in crow's feet wrinkles at the botulinum nanoemulsion treated sites. The severity of wrinkles is measured using a 4-point wrinkle scale (wrinkle scale): 0=none, 1=mild, 2=moderate, 3=severe.

[0306] The wrinkle scale is used at baseline prior to botulinum nanoemulsion treatment, 2 weeks after treatment, and 4 weeks after treatment. The study found that at baseline, the mean severity of wrinkles detected by the wrinkle scale was approximately equal for groups 1-3. At 2 and 4 weeks after treatment, the mean severity of wrinkles in group 1 was greater than that detected in groups 2 and 3 at these post-treatment weeks. The study found that at 2 and 4 weeks after treatment, the mean severity of wrinkles in group 3 was greater than that detected in group 2 at these post-treatment weeks.

[0307] This study demonstrated that microneedle preconditioning using a relatively lower microneedle density unexpectedly increased the bioavailability of a topical large drug nanoemulsion containing botulinum toxin.

[0308] Example 10: Effect of MSCs with varying microneedle puncture size on the bioavailability of botulinum toxin in humans: Effect on reduction of crow's feet wrinkles A single dose topical study of the bioavailability of botulinum toxin following topical administration of a topical botulinum nanoemulsion formulation in humans is conducted. The study is designed to not only evaluate but also compare the % bioavailability achieved using various microneedle systems, thereby testing the effect of varying microneedle puncture size on enhancing botulinum bioavailability in humans by measuring reduction in skin wrinkling following topical treatment with a botulinum nanoemulsion formulation.

[0309] The study included three treatment groups of 12 subjects each with severe crow's feet around the eyes: Group 1 was the control group, in which botulinum nanoemulsion was applied to each subject's crow's feet; Group 2 was intervention group #1, in which approximately 11,000 μm 2 Group 3 is the intervention group #1, preconditioned with 5 application of a microneedle array of microneedle puncture size of the microneedle to each area of ​​the skin where the crow's feet are located prior to application of the botulinum nanoemulsion formulation. 2 The skin is preconditioned with five impressions of a microneedle array of microneedle puncture size. Each subject in groups 1, 2, and 3 is topically treated once with a volume of a botulinum nanoemulsion formulation at a fixed concentration of botulinum. Administration of the topical formulation to the skin takes approximately 5 minutes, at which point the topical formulation is completely absorbed by the skin.

[0310] The predicted effect of botulinum nanoemulsion treatment is a reduction in crow's feet wrinkles at the botulinum nanoemulsion treated sites. The severity of wrinkles is measured using a 4-point wrinkle scale (wrinkle scale): 0=none, 1=mild, 2=moderate, 3=severe.

[0311] The wrinkle scale is used at baseline prior to botulinum nanoemulsion treatment, 2 weeks after treatment, and 4 weeks after treatment. The study found that at baseline, the mean severity of wrinkles detected by the wrinkle scale was approximately equal for groups 1-3. At 2 and 4 weeks after treatment, the mean severity of wrinkles in group 1 was greater than that detected in groups 2 and 3 at these post-treatment weeks. The study found that at 2 and 4 weeks after treatment, the mean severity of wrinkles in group 3 was greater than that detected in group 2 at these post-treatment weeks.

[0312] This study demonstrated that microneedle preconditioning with a relatively smaller microneedle puncture size unexpectedly increased the bioavailability of a topical large drug nanoemulsion containing botulinum toxin.

[0313] Example 11: Effect of MSC on the bioavailability of botulinum toxin in humans: Effect of varying dosing on the reduction of crow's feet A single dose topical study of the bioavailability of botulinum toxin following topical administration of a topical botulinum nanoemulsion formulation in humans is conducted. The study is designed to not only evaluate but also compare the % bioavailability achieved using various microneedle systems, thereby testing the effect of varying needle density of the microneedle array on enhancing the bioavailability of botulinum in humans by measuring reduction in skin wrinkling following topical treatment with a botulinum nanoemulsion formulation.

[0314] The study included three treatment groups of 12 subjects each with severe crow's feet: Group 1 was the control group, in which botulinum nanoemulsion was applied to each subject's crow's feet; Group 2 was the intervention group #1, in which 9 microneedles / cm were applied to each area of ​​skin where the crow's feet were located prior to application of the botulinum nanoemulsion formulation. 2 Group 3 is the intervention group #1, which was preconditioned with five impressions of a microneedle array with a microneedle density of 85 microneedles / cm2 on each area of ​​skin where the crow's feet are located prior to application of the botulinum nanoemulsion formulation. 2 Each subject is preconditioned with five impressions of a microneedle array with a microneedle density of 100 μg / mL. Each subject is topically treated once with a volume of a botulinum nanoemulsion formulation at a constant concentration of botulinum, except that the treatment in Group 1 has twice the botulinum concentration of the treatments in Groups 2 and 3. Administration of the topical formulation to the skin takes approximately 5 minutes, at which point the topical formulation is completely absorbed into the skin.

[0315] The predicted effect of botulinum nanoemulsion treatment is a reduction in crow's feet wrinkles at the botulinum nanoemulsion treated sites. The severity of wrinkles is measured using a 4-point wrinkle scale (wrinkle scale): 0=none, 1=mild, 2=moderate, 3=severe.

[0316] The wrinkle scale is used at baseline prior to botulinum nanoemulsion treatment, 2 weeks after treatment, and 4 weeks after treatment. The study found that at baseline, the mean severity of wrinkles as detected by the wrinkle scale was approximately equal for groups 1-3. At 2 and 4 weeks after treatment, the mean severity of wrinkles in groups 1 and 3 compared to baseline was reduced by approximately the same amount, even though group 1 was treated with twice the concentration of group 3. The study also found that at 2 and 4 weeks after treatment, the mean severity of wrinkles in group 2 compared to baseline was reduced significantly more than the mean severity of wrinkles in group 3 compared to baseline, even though groups 2 and 3 were treated with the same concentration of botulinum.

[0317] This study demonstrated that microneedle preconditioning using a lower relative microneedle density unexpectedly increased the bioavailability of a topical large drug nanoemulsion containing botulinum toxin, resulting in equivalent therapeutic efficacy using a lower dose of botulinum compared to patients who did not receive microneedle skin preconditioning or who received MSCs using a higher relative microneedle density.

[0318] Example 12: Effect of MSC on the bioavailability of botulinum toxin in humans: Effect of varying dosing on the reduction of crow's feet A single dose topical study of the bioavailability of botulinum toxin following topical administration of a topical botulinum macroemulsion formulation in humans is conducted. The study is designed to not only evaluate but also compare the % bioavailability achieved using various microneedle systems, thereby testing the effect of varying needle density of the microneedle array on enhancing the bioavailability of botulinum in humans by measuring reduction in skin wrinkles following topical treatment with a botulinum macroemulsion formulation.

[0319] The study included three treatment groups of 12 subjects each with severe crow's feet: Group 1 was the control group, in which botulinum macroemulsion was applied to each subject's crow's feet; Group 2 was the intervention group #1, in which 9 microneedles / cm were applied to each area of ​​skin where the crow's feet were located prior to application of the botulinum macroemulsion formulation. 2 Group 3 is the intervention group #1, which was preconditioned with five impressions of a microneedle array with a microneedle density of 85 microneedles / cm2 on the area of ​​skin where the crow's feet are located prior to application of the botulinum macroemulsion formulation. 2 Each subject is preconditioned with five impressions of a microneedle array with a microneedle density of 100 μg / mL. Each subject is topically treated once with a volume of a botulinum nanoemulsion formulation with a fixed concentration of botulinum. Administration of the topical formulation to the skin takes approximately 5 minutes, at which point the topical formulation is completely absorbed by the skin.

[0320] The predicted effect of botulinum nanoemulsion treatment is a reduction in crow's feet wrinkles at the botulinum nanoemulsion treated sites. The severity of wrinkles is measured using a 4-point wrinkle scale (wrinkle scale): 0=none, 1=mild, 2=moderate, 3=severe.

[0321] The wrinkle scale is used at baseline prior to botulinum nanoemulsion treatment, 2 weeks after treatment, and 4 weeks after treatment. The study found that at baseline, the mean severity of wrinkles detected by the wrinkle scale was approximately equal in groups 1-3. At 2 and 4 weeks after treatment, the mean severity of wrinkles in group 1 was greater than the mean severity of wrinkles in groups 2 and 3. The study found that at 2 and 4 weeks after treatment, the mean severity of wrinkles in group 3 was greater than the mean severity of wrinkles in group 2.

[0322] This study demonstrates that microneedle preconditioning using a relatively lower microneedle density unexpectedly increases the bioavailability of a topical large drug macroemulsion containing botulinum toxin compared to patients who did not receive microneedle skin preconditioning or who received MSCs using a relatively higher microneedle density.

[0323] Example 13: Effect of MSC preconditioning with varying microneedle impingement times on the bioavailability of botulinum toxin in humans: Effect on reduction of crow's feet wrinkles A single-dose topical study of the bioavailability of botulinum toxin following topical administration of a topical botulinum nanoemulsion formulation in humans was conducted. The study examined the effect of varying the number of microneedle array impacts on improving the bioavailability of botulinum in humans by measuring the reduction in skin wrinkling following topical treatment with the botulinum nanoemulsion formulation following skin conditioning with a microneedle array.

[0324] The study included two groups, Group A and Group B, which included subjects with moderate to severe crow's feet wrinkles. Each subject was topically treated once with a near-constant dose of botulinum, a botulinum emulsion (e.g., nanoemulsion) formulation in the crow's feet area. Administration of the topical formulation to the skin takes approximately 5 minutes, at which point the topical formulation is completely absorbed by the skin. Subjects in Group A (N=8) were treated with 14 microneedle impressions (or 4 microneedle impressions / cm) of the microneedle array prior to application of the botulinum formulation. 2 Subjects in Group B (N=17) were preconditioned with 8 microneedle presses (or 2 microneedle presses / cm) of the microneedle array prior to application of the botulinum formulation. 2 The lengths of the microneedles in the microneedle arrays used in groups A and B were the same.

[0325] The expected effect of the botulinum nanoemulsion treatment is a reduction in crow's feet wrinkles at the sites of botulinum nanoemulsion treatment. Wrinkle severity is measured by the investigator and the subject separately using a 5-point wrinkle scale (wrinkle scale) with 0=none, 1=minimal, 2=mild, 3=moderate, and 4=severe. Responders in this study were subjects who had a reduction in wrinkle severity of 1 point or more compared to baseline as assessed by both the investigator and the subject.

[0326] The study found that at baseline, the mean severity of crow's feet wrinkles, as measured by the wrinkle scale, was approximately equal in Groups A and B. Eight weeks after treatment, the responder rate in Group A was 25% and in Group B was 50%.

[0327] This study demonstrated that microneedle preconditioning using relatively fewer microneedle impacts (per unit area or in absolute number) unexpectedly increased the bioavailability of a topical large drug nanoemulsion containing botulinum toxin.

[0328] Example 14:...

Claims

1. An emulsion composition containing a large drug having a molecular weight of 100,000 Da or more is administered to a certain site at a rate of about 2 to about 50 microneedles / cm 2 in combination with microneedle skin conditioning (MSC) of the site using a microneedle array having a density of microneedles in the range of

2. The density of the microneedles is about 2 to about 10 microneedles / cm 2 The method of claim 1, wherein the range is

3. Microneedle density is about 2 to about 35 microneedles / cm 2 The method of claim 1, wherein the range is

4. 2. The method of claim 1, wherein the composition comprising the large drug comprises a nanoemulsion comprising the large drug.

5. 10. The method of claim 1, wherein the composition comprising a large drug comprises a macroemulsion comprising a large drug.

6. 6. The method of any one of claims 1 to 5, further comprising administration of a non-irritating penetration enhancer.

7. 7. The method of claim 6, wherein the non-irritating penetration enhancer is selected from a carrier peptide and a copeptide.

8. The non-irritating penetration enhancer has the sequence RKKRRQRRRG-(K) 15 The method according to any one of claims 6 to 7, wherein the cationic peptide having -GRKKRRQRRR and a positively charged carrier are selected from the group consisting of cationic peptides having -GRKKRRQRRR and positively charged carriers.

9. The method of any one of claims 1 to 8, wherein MSC of the site is performed prior to application of a composition comprising a large drug to the site.

10. The method of any one of claims 1 to 8, wherein MSC of the site is performed after application of a composition comprising a large drug to the site.

11. 9. The method of any one of claims 1 to 8, wherein application of the MSCs at the site and the composition comprising the large drug to the site occurs substantially simultaneously.

12. The method of any one of claims 1 to 11, wherein the large drug is a botulinum toxin.

13. 13. The method of claim 12, further comprising delivering a botulinum toxin with the biologically active agent.

14. 14. The method according to claim 13, wherein the biologically active substance is selected from steroids, retinoids, anesthetics, fillers, silicones and / or collagen.

15. 15. The method of claim 14, wherein the biologically active agent is selected from hydrocortisone, retin A and / or lidocaine.

16. The method of any one of claims 1 to 11, wherein the large drug is an antibody drug.

17. The method of claim 16, wherein the antibody drug is selected from an anti-TNFα antibody, an anti-CD2 antibody, an anti-CD4 antibody, an anti-IL-12 antibody, an anti-IL-17 antibody, an anti-IL-22 antibody, and an anti-IL-23 antibody.

18. 18. The method of any one of claims 16 to 17, wherein the antibody drug is selected from antibodies having an epitope binding member found in one or more of infliximab, adalimumab, golimumab, etanercept, etanercept-szzs, certolizumab pegol, siplizumab, zanolimumab, briakinumab, secukinumab, brodalumab, fezakinumab, ustekinumab and / or guselkumab.

19. The method of any one of claims 16 to 17, further comprising delivering an antibody drug with a biologically active agent.

20. 20. The method of any one of claims 16-19, further comprising delivering the antibody drug with a non-irritating penetration enhancer.

21. 21. The method of claim 20, wherein the non-irritating penetration enhancer is selected from a copeptide and a carrier peptide.

22. 22. The method of any one of claims 1 to 21, wherein MSC at the site is achieved using a device comprising multiple needles.

23. 23. The method of claim 22, wherein the device is a patch, roller, stamp or pen.

24. The method according to any one of claims 1 to 23, wherein the site is a skin surface overlying a muscle or muscle group of the subject.

25. The method according to any one of claims 1 to 24, wherein the site is a skin surface containing sweat glands.

26. The method according to any one of claims 1 to 25, wherein the site is a skin surface containing a sebaceous gland.

27. The method according to any one of claims 1 to 26, wherein the site is a skin surface containing hair follicles.

28. 28. The method of any one of claims 22 to 27, wherein the needles have a length sufficient to protrude through the stratum corneum of the skin.

29. 29. The method of any one of claims 22 to 28, wherein the needle has a length insufficient to reach nerves in the dermis of the skin.

30. 30. The method of any one of claims 22 to 29, wherein the length of the needle is from about 10 μm to about 4000 μm.

31. The method of any one of claims 22 to 30, wherein the length of the needle is from about 10 μm to about 800 μm.

32. The method of any one of claims 22 to 31, wherein the length of the needle is from about 10 μm to about 500 μm.

33. The method of any one of claims 22 to 30, wherein the length of the needle is about 200 μm or more.

34. The method of any one of claims 22 to 33, wherein the length of the needle is about 300 μm or more.

35. The method of any one of claims 22 to 34, wherein the length of the needle is about 500 μm or more.

36. The length of the needle is about 200 μm, about 300 μm, about 500 μm, about 800 μm, about 1000 μm, about 1100 μm, about 1200 μm, about 1300 μm, about 1400 μm, about 1500 μm, about 1600 μm, about 1700 μm, about 1800 μm, about 1900 μm, about 2000 μm, about 2100 μm, about 2200 μm, about 2300 μm, about 2400 μm, 31. The method of any one of claims 22 to 30, wherein the length is equal to or greater than about 2500 μm, about 2600 μm, about 2700 μm, about 2800 μm, about 2900 μm, about 3000 μm, about 3100 μm, about 3200 μm, about 3300 μm, about 3400 μm, about 3500 μm, about 3600 μm, about 3700 μm, about 3800 μm or about 3900 μm.

37. The method of any one of claims 22 to 36, wherein the needle is made of a biocompatible material.

38. The method of any one of claims 22 to 36, wherein the needle is made of metal.

39. 39. The method of any one of claims 1 to 38, wherein the MSC comprises administering 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 presses of MNs or MN arrays, each press being a number of consecutive applications of MNs to the skin.

40. The method of claim 39, wherein the MN array is rotated between one or more of the pressings.

41. The method of claim 39, wherein the MN array is not rotated between one or more pressings.

42. 42. The method according to any one of claims 39 to 41, wherein the pressing is performed at approximately the same site.

43. The method according to any one of claims 39 to 41, wherein the pressing is performed at an overlapping site.

44. The method according to any one of claims 39 to 41, wherein the pressing is performed at different sites.

45. The method of any one of claims 39 to 44, wherein the MN array is in the form of a stamp or roller.

46. 46. ​​The method of claim 45, wherein the pressing is performed by stamping or rolling.

47. 47. The method of any one of claims 1 to 46, wherein the large drug penetrates the skin within about 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 minutes of administration.

48. 47. The method of any one of claims 1 to 46, wherein the large drug penetrates the skin within about 5 to about 60 minutes, about 5 to about 12 minutes, about 5 to about 15 minutes, or about 15 to about 30 minutes of administration.

49. 47. The method of any one of claims 1 to 46, wherein the large drug penetrates the skin within about 1, 2, 3, 4, 5 or 6 hours of administration.

50. The method according to any one of claims 22 to 37, wherein the needle is composed of at least one soluble polymer.

51. Dosage: 50 microneedles / cm 2 51. The method of any one of claims 1 to 50, comprising administering a composition comprising a lower dose of a large drug compared to a reference treatment regimen with MSCs at a site using a reference microneedle array having a microneedle density of greater than 100 μg / ml.

52. 52. The method of any one of claims 1 to 51, comprising administering two or more doses of a composition comprising a large drug over time.

53. The dose should be 50 microneedles / cm to produce a comparable treatment effect. 2 53. The method of claim 52, comprising administering a composition comprising a smaller dose of a large drug over a fixed treatment dose compared to a reference treatment regimen with MSCs at a site using a reference microneedle array having a density of microneedles greater than 100 μg / mL.

54. 54. The method of claim 52 or 53, wherein the composition comprising each dose of the large drug is separated by a specific time.

55. At a specific time, 50 microneedles / cm 2 55. The method of claim 54, wherein the time is longer than a particular time for administering a reference treatment regimen with MSCs at a site using a reference microneedle array having a density of microneedles greater than 100 μg / mL.

56. The reference treatment regimen was 50 microneedles / cm 2 56. The method of any one of claims 51 to 55, comprising administering the composition comprising a large drug using a reference microneedle array having a density of microneedles greater than

57. 57. A method of treating a skin disorder comprising the method of any one of claims 1 to 56.

58. 58. The method of claim 57, wherein the skin disorder is selected from acne, unwanted sweating, body odor, hyperhidrosis, bromhidrosis, chromhidrosis, rosacea, alopecia, Raynaud's syndrome, psoriasis, actinic keratosis, eczematous dermatitis, excessive sebum production disorders, burns, lupus erythematosus, hyperpigmentation disorders, hypopigmentation disorders, skin cancer, skin infections, facial wrinkles, unsightly facial expressions, neck wrinkles, hyperfunctional facial wrinkles, hyperkinetic facial wrinkles, platysma bands, and / or combinations thereof.

59. 58. The method of claim 1, further comprising administering to the patient a therapeutically effective amount of ... A method for treating or preventing a disorder selected from: Bacillus difficile infection, Staphylococcus infection, dystonia, headache, pain, arthritis associated pain, rheumatoid arthritis associated pain, psoriatic arthritis associated pain, osteoarthritis associated pain, certain ophthalmological conditions, certain urological conditions, neuromuscular diseases, conditions involving muscle spasms and / or contractures, strabismus, hemifacial spasm, tremor, spasticity such as caused by multiple sclerosis, retroorbital muscles, neurological conditions, migraine or other headaches, Alzheimer's disease, Parkinson's disease, or stroke.

60. 60. The method of any one of claims 1 to 59, wherein the composition comprising the large drug is formulated as a lotion, cream, powder, ointment, liniment, gel or eye drops.

61. A composition containing a large drug and about 2 to about 50 microneedles / cm 2 A patch comprising a plurality of microneedles having a microneedle density in the range of

62. 62. The patch of claim 61, wherein the composition comprising the large drug comprises a nanoemulsion.

63. 62. The patch of claim 61, wherein the composition comprising the large drug comprises a macroemulsion.

64. The patch of any one of claims 61 to 63, wherein the needles have a length sufficient to protrude through the stratum corneum of the skin.

65. The patch of any one of claims 62-64, wherein the needles have a length insufficient to reach nerves in the dermis of the skin.

66. The patch of any one of claims 62 to 65, wherein the length of the needles is from about 10 μm to about 4000 μm.

67. The patch of any one of claims 62 to 66, wherein the needles have a length of about 200 μm or more.

68. The patch of any one of claims 62 to 67, wherein the needles have a length of about 300 μm or more.

69. The patch of any one of claims 62 to 68, wherein the needles have a length of about 500 μm or more.

70. The patch of any one of claims 62 to 69, wherein the needle is constructed from a biocompatible material.

71. The patch of any one of claims 62 to 70, wherein the needle is constructed from metal.

72. The patch of any one of claims 62 to 70, wherein the needle is composed of a dissolvable polymer.

73. The patch of any one of claims 62-72, wherein the large drug has a molecular weight of 100 KDa or more.

74. The patch of any one of claims 62 to 73, wherein the large drug is a botulinum toxin.

75. The patch of any one of claims 62 to 73, wherein the large drug is an antibody drug.

76. An emulsion composition containing a large drug having a molecular weight of 100,000 Da or more is applied to a certain site at a depth of about 100 to about 35,000 μm. 2 / Applying in combination with microneedle skin conditioning (MSC) of the site using a microneedle array having a microneedle puncture hole size within the range of the microneedles.

77. The dose is approximately 35,000 μm 2 77. The method of claim 76, comprising administering a composition comprising a lower dose of a large drug compared to a reference treatment regimen with MSCs at a site using a reference microneedle array having a microneedle puncture hole size greater than the microneedle.

78. 78. The method of any one of claims 76 to 77, comprising administering two or more doses of a composition comprising a large drug over time.

79. The dose should be approximately 35,000 μm to produce a comparable therapeutic effect. 2 The method of claim 78, comprising administering a composition comprising a smaller dose of a large drug over a fixed treatment dose compared to a reference treatment regimen with MSCs at a site using a reference microneedle array having a microneedle puncture hole size greater than the microneedle.

80. 80. The method of claim 78 or 79, wherein the composition comprising each dose of the large drug is separated by a specific time.

81. A specific time is about 35,000 μm 2 / The method of claim 80, wherein the time is longer than the specified time for administering a reference treatment regimen with MSCs at a site using a reference microneedle array having a microneedle puncture hole size greater than the microneedle.

82. The reference treatment regimen was ca. 2 The method of any one of claims 77 to 81, comprising administering a composition comprising a large drug using a reference microneedle array having a microneedle puncture hole size larger than the microneedle.

83. A kit comprising a composition comprising a large drug and a device for microneedle conditioning of an area, the device comprising about 2 to about 50 microneedles / cm. 2 The kit is a microneedle array having a density of microneedles in the range of

84. 84. The kit of claim 83, wherein the composition comprising the large drug comprises a nanoemulsion.

85. 85. The kit of claim 83 or 84, wherein the composition comprising the large drug comprises a macroemulsion.

86. The kit of any one of claims 83 to 85, wherein the device is a patch, roller, stamp or pen.

87. A kit according to any one of claims 83 to 86, comprising a patch according to any one of claims 56 to 68.

88. 84. The kit of claim 83, wherein the composition comprising the large drug is formulated as a lotion, cream, powder, ointment, liniment, gel or eye drops.

89. 1. A method comprising applying to a site an emulsion composition comprising a large drug having a molecular weight of 100,000 Da or greater, the site comprising about 2 to about 50 microneedles / cm 2 The site is conditioned with microneedle skin conditioning (MSC) using a microneedle array having a density of microneedles in the range of.

90. 1. A method comprising the step of applying to a site an emulsion composition containing a large drug having a molecular weight of 100,000 Da or more, the site being between about 100 and about 35,000 μm 2 / The site is conditioned with microneedle skin conditioning (MSC) using a microneedle array having a microneedle puncture hole size within the range of the microneedles.

91. A kit comprising a composition comprising a large drug and a device for microneedle conditioning of a site, the device having a diameter of about 100 to about 35,000 μm. 2 / A kit which is a microneedle array having a microneedle puncture hole size within the range of microneedles.

92. 92. The kit of claim 91, wherein the composition comprising the large drug comprises a nanoemulsion.

93. 93. The kit of claim 91 or 92, wherein the composition comprising the large drug comprises a macroemulsion.

94. The kit of any one of claims 91 to 93, wherein the device is a patch, roller, stamp or pen.

95. A kit according to any one of claims 91 to 94, comprising a patch according to any one of claims 56 to 68.

96. 92. The kit of claim 91, wherein the composition comprising the large drug is formulated as a lotion, cream, powder, ointment, liniment, gel or eye drops.

97. A method comprising applying to a site an emulsion composition comprising a large drug having a molecular weight of 100,000 Da or greater in combination with microneedle skin conditioning (MSC) of the site using a microneedle array, wherein about 1 to about 13 application of the microneedle array is made to the site.

98. An emulsion composition containing a large drug having a molecular weight of 100,000 Da or more is applied to a certain site by pressing the microneedle array about once to about four times to a depth of 1 cm 2 of the site. 2 in combination with microneedle skin conditioning (MSC) of the site using a microneedle array per unit time.

99. Approximately 1 to 3 presses of the microneedle array are performed on 1 cm 2 99. The method of claim 98, wherein the method is performed per

100. Approximately 1 to 2 presses of the microneedle array are performed on 1 cm 2 99. The method of claim 98, wherein the method is performed per

101. A method comprising the step of applying to a site an emulsion composition comprising a large drug having a molecular weight of 100,000 Da or greater in combination with microneedle skin conditioning (MSC) of the site using a microneedle array having microneedle lengths in the range of about 1 μm to about 700 μm.

102. A method comprising the step of applying to a site an emulsion composition comprising a large drug having a molecular weight of 100,000 Da or greater in combination with microneedle skin conditioning (MSC) of the site using a microneedle array having microneedle lengths in the range of about 1 μm to about 500 μm.

103. At a site, 1 drop / cm containing a large drug with a molecular weight of 100,000 Da or more 2 ~About 2 drops / cm 2 applying about 1 / 100 of a product volume of the emulsion composition in combination with microneedle skin conditioning (MSC) of the site using a microneedle array.

104. At a certain site, approximately 0.0001 mls / cm containing a large drug with a molecular weight of 100,000 Da or more 2 ~Approx. 0.065mls / cm 2 applying a product volume of the emulsion composition in combination with microneedle skin conditioning (MSC) of the area using a microneedle array.

105. Product volume is approximately 0.0001mls / cm 2 ~Approx. 0.05mls / cm 2The method of claim 104, wherein the range is

106. 105. The method of any one of claims 101 to 104, wherein the composition comprising the large drug comprises a nanoemulsion comprising the large drug.

107. 105. The method of any one of claims 101 to 104, wherein the composition comprising the large drug comprises a macroemulsion comprising the large drug.

108. 108. The method of any one of claims 101-107, further comprising administration of a non-irritating penetration enhancer.

109. 109. The method of claim 108, wherein the non-irritating penetration enhancer is selected from a carrier peptide and a copeptide.

110. The non-irritating penetration enhancer has the sequence RKKRRQRRRG-(K) 15 The method according to any one of claims 108 to 109, wherein the cationic peptide having -GRKKRRQRRR and a positively charged carrier are selected from the group consisting of cationic peptides having -GRKKRRQRRR and positively charged carriers.

111. 111. The method of any one of claims 101 to 110, wherein MSC of the site is performed prior to application of a composition comprising a large drug to the site.

112. 111. The method of any one of claims 101 to 110, wherein MSC of the site is performed after application of a composition comprising a large drug to the site.

113. 111. The method of any one of claims 101 to 110, wherein application of the composition comprising the MSCs and the large drug to the site occurs substantially simultaneously.

114. The method of any one of claims 101 to 113, wherein the large drug is a botulinum toxin.

115. 115. The method of claim 114, further comprising delivering a botulinum toxin with the biologically active agent.

116. 116. The method of claim 115, wherein the biologically active agent is selected from steroids, retinoids, anesthetics, fillers, silicones and / or collagen.

117. 117. The method of claim 116, wherein the biologically active agent is selected from hydrocortisone, retin A, and / or lidocaine.

118. The method of any one of claims 101 to 113, wherein the large drug is an antibody drug.

119. The method of claim 118, wherein the antibody drug is selected from an anti-TNFα antibody, an anti-CD2 antibody, an anti-CD4 antibody, an anti-IL-12 antibody, an anti-IL-17 antibody, an anti-IL-22 antibody and an anti-IL-23 antibody.

120. 120. The method of any one of claims 118-119, wherein the antibody drug is selected from antibodies having an epitope binding member found in one or more of: infliximab, adalimumab, golimumab, etanercept, etanercept-szzs, certolizumab pegol, siplizumab, zanolimumab, briakinumab, secukinumab, brodalumab, fezakinumab, ustekinumab and / or guselkumab.

121. 121. The method of any one of claims 118-120, further comprising delivering an antibody drug with a biologically active agent.

122. 122. The method of any one of claims 118-121, further comprising delivering the antibody drug with a non-irritating penetration enhancer.

123. 123. The method of claim 122, wherein the non-irritating penetration enhancer is selected from a copeptide and a carrier peptide.

124. 124. The method of any one of claims 101 to 123, wherein MSC at the site is achieved using a device comprising multiple needles.

125. 125. The method of claim 124, wherein the device is a patch, roller, stamp or pen.

126. The method of any one of claims 101 to 125, wherein the site is a skin surface overlying a muscle or muscle group of the subject.

127. The method of any one of claims 101 to 126, wherein the site is a skin surface containing sweat glands.

128. The method according to any one of claims 101 to 127, wherein the site is a skin surface comprising a sebaceous gland.

129. The method of any one of claims 101 to 128, wherein the site is a skin surface containing hair follicles.

130. 130. The method of any one of claims 124 to 129, wherein the needles have a length sufficient to protrude through the stratum corneum of the skin.

131. 131. The method of any one of claims 124 to 130, wherein the needle has a length insufficient to reach nerves in the dermis of the skin.

132. The method of any one of claims 124 to 131, wherein the needle is constructed from a biocompatible material.

133. The method of any one of claims 124 to 131, wherein the needle is constructed from metal.

134. The method of any one of claims 101 to 133, wherein the MN array is rotated between one or more pressings.

135. The method of any one of claims 101 to 133, wherein the MN array is not rotated between one or more pressings.

136. 136. The method of any one of claims 134 to 135, wherein the pressing is performed at approximately the same site.

137. 136. The method of any one of claims 134 to 135, wherein the pressing is performed at an overlapping site.

138. The method according to any one of claims 134 to 135, wherein the pressing is performed at different sites.

139. The method of any one of claims 134 to 138, wherein the MN array is in the form of a stamp or roller.

140. 140. The method of claim 139, wherein the pressing is performed by stamping or rolling.

141. 141. The method of any one of claims 101-140, wherein the large drug penetrates the skin within about 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 minutes of administration.

142. 141. The method of any one of claims 101-140, wherein the large drug penetrates the skin within about 5 to about 60 minutes, about 5 to about 12 minutes, about 5 to about 15 minutes, or about 15 to about 30 minutes of administration.

143. 141. The method of any one of claims 101-140, wherein the large drug penetrates the skin within about 1, 2, 3, 4, 5 or 6 hours of administration.

144. The method of any one of claims 124 to 132, wherein the needle is comprised of at least one dissolvable polymer.

145. 145. The method of any one of claims 101, 106-144, wherein the administration comprises administration of a composition comprising a lower dose of a large drug compared to a reference treatment regimen with MSCs at a site using a reference microneedle array having a microneedle length of greater than about 700 μm.

146. 145. The method of any one of claims 102, 106-144, wherein the administration comprises administration of a composition comprising a lower dose of a large drug compared to a reference treatment regimen with MSCs at a site using a reference microneedle array having a microneedle length of greater than about 500 μm.

147. 147. The method of any one of claims 101 to 146, comprising administering two or more doses of a composition comprising a large drug over time.

148. The method of claim 147, wherein the administration comprises administration of a composition comprising a smaller dose of a large drug over a fixed treatment dose compared to a reference treatment regimen with MSCs at the site using a reference microneedle array.

149. 149. The method of claim 147 or 148, wherein the composition comprising each dose of the large drug is separated by a specific time.

150. The method of claim 149, wherein the specified time is longer than the specified time for administering a reference treatment regimen with MSCs at the site using the reference microneedle array.

151. 151. The method of any one of claims 145, 147-150, wherein the reference treatment regimen comprises administration of a composition comprising a large drug using a reference microneedle array having a microneedle length of greater than about 700 μm.

152. 151. The method of any one of claims 147 to 150, wherein the reference treatment regimen comprises administration of a composition comprising a large drug using a reference microneedle array having a microneedle length of greater than about 500 μm.

153. 153. A method of treating a skin disorder comprising the method of any one of claims 101 to 152.

154. 154. The method of claim 153, wherein the skin disorder is selected from acne, unwanted sweating, body odor, hyperhidrosis, bromhidrosis, chromhidrosis, rosacea, alopecia, Raynaud's syndrome, psoriasis, actinic keratosis, eczematous dermatitis, excessive sebum production disorders, burns, lupus erythematosus, hyperpigmentation disorders, hypopigmentation disorders, skin cancer, skin infections, facial wrinkles, unsightly facial expressions, neck wrinkles, hyperfunctional facial wrinkles, hyperkinetic facial wrinkles, platysma bands, and / or combinations thereof.

155. The method of any one of claims 101 to 153, comprising administering to a patient a therapeutically effective amount of ... A method of treating or preventing a disorder selected from: C. difficile infection, Staphylococcus infection, dystonia, headache, pain, arthritis associated pain, rheumatoid arthritis associated pain, psoriatic arthritis associated pain, osteoarthritis associated pain, certain ophthalmological conditions, certain urological conditions, neuromuscular diseases, conditions involving muscle spasms and / or contractures, strabismus, hemifacial spasm, tremor, spasticity such as those caused by multiple sclerosis, retroorbital muscles, neurological conditions, migraine or other headaches, Alzheimer's disease, Parkinson's disease, or stroke.

156. The method of any one of claims 101 to 155, wherein the composition comprising the large drug is formulated as a lotion, cream, powder, ointment, liniment, gel, or eye drops.

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