Improved delivery of large drugs

Abstract

Methods, compositions and devices for improving the transdermal delivery and / or bioavailability of large drugs.

Description

[Technical Field]

[0001] (Cross-reference of related applications) This application claims priority under U.S. Provisional Applications No. 62 / 774,677 (filed December 3, 2018), No. 62 / 789,407 (filed January 7, 2019), and No. 62 / 808,274 (filed February 20, 2019), all of which, in whole, constitute a part of this Specification as indicated by attribution. [Background technology]

[0002] Significant resources are being 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, particularly for large-sized drugs. As molecular size increases, transdermal penetration decreases to almost negligible or even nonexistent. [Overview of the Initiative]

[0003] Transdermal administration is generally the focus of research seeking to provide an alternative route of drug administration that does not have the undesirable consequences associated with injection and oral delivery. For example, needles often cause local pain, bleeding, and subcutaneous hemorrhage, potentially exposing patients to infectious diseases; oral administration may suffer from low bioavailability of the drug due to the highly 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] While offering numerous advantages, transdermal drug delivery presents many logistical challenges. Only a limited number of drugs have been shown to be deliverable via this route. Transdermal delivery of active substances, including but not limited to hydrophilic molecules, large molecular structures (e.g., exceeding several hundred daltons), gene therapies, and vaccines, has been difficult. (Prausnitz, MR & Langer, R. "Transdermal drug delivery," Nat Biotechnol. 26(11): 1261-1268 (2008)).

[0005] This disclosure provides improved techniques for transdermal delivery of a drug of interest. In some embodiments, this disclosure teaches that a combination of certain microneedling techniques and topical application of a composition containing a large drug may facilitate and / or otherwise improve topical drug delivery. In some embodiments, such a composition may or may not be an emulsion (e.g., a nanoemulsion) containing, for example, a large drug. In some embodiments, such a composition may or may not contain an emulsion and may be formulated as an emulsion and / or, for example, a liquid, gel, cream, or other formulation suitable for topical application.

[0006] Recent technologies have been developed that combine emulsion and microneedling techniques for transdermal delivery of target drugs, achieving various advantages (see, for example, International Application No. PCT / US17 / 53333, which is incorporated herein by attribution); in some embodiments, these technologies have been shown to achieve particularly remarkable improvements for the transdermal delivery of large molecular structures.

[0007] This disclosure reveals that even greater advantages are achieved when using a microneedling approach with a specific microneedle density and / or a specific microneedle puncture size. Surprisingly, this disclosure teaches that particularly desired results are achieved with a microneedling approach with a relatively low density and / or a relatively small puncture size.

[0008] Various microneedling techniques have been developed that may be useful for administering specific target drugs. Microneedling can avoid certain drawbacks (e.g., the degree of pain and / or 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) hollow or solid microneedles. The target drug may be placed inside (e.g., if the microneedle is hollow and / or if the drug is incorporated into the microneedle material) or on top of (i.e., on the surface) the microneedle, and / or applied to the skin site before, during, or after microneedling. The drug inside or on the microneedle may be released after application to the site, for example, by diffusion or discharge from the microneedle, or by rupture and / or disintegration of the microneedle material.

[0009] In some embodiments, the Disclosure provides strategies in which microneedling is used to “condition” the skin (in particular to precondition the skin between administrations of large drugs), e.g., microneedle skin conditioning (MSC) to which a transdermal product has been, is, or will be applied. The Disclosure provides the finding that such microneedle conditioning described herein (e.g., microneedle conditioning with relatively low density and / or relatively small puncture size) surprisingly provides a significant benefit in improving the transdermal delivery of large drugs (e.g., with a 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 Disclosure teaches that microneedling may provide a remarkable improvement in the delivery of large drugs. In some embodiments, the Disclosure demonstrates that microneedling treatment may be particularly advantageous for the delivery of large drugs in emulsion compositions (e.g., nanoemulsions).

[0010] Studies analyzing the transdermal delivery of small molecules (particularly 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 with increasing molecular weight," and 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 effects 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 using microneedles with sufficiently long needle lengths (>600 μm), a lower needle density (<2000 needles / cm³) is achieved." 2Yan stated that "microneedle arrays with ) were more effective in improving drug flow," but later studies identified flaws in the approach Yan used, including that the assay used by Yan was "prone to artifacts" and could result in biological changes in skin tension and / or hydration levels (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 was 400 needles / cm². 2 Even then, the results were relatively high. Furthermore, data presented by Yan himself show that for needle lengths less than 1100 μm, needle density does not have a significant effect on small molecule delivery (e.g., "drug flow" through the skin).

[0011] The aforementioned study by Donnelly reports 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] Therefore, prior to this disclosure, the art had shown that changes in microneedle density were not expected to affect drug delivery across the skin, at least at density levels studied in the literature. In first principle, it might be expected that if microneedling improves delivery, relatively higher microneedle densities (and / or relatively larger microneedle puncture sizes) might be more effective. As each factor of microneedle density and microneedle diameter increases, the total surface area of ​​the punctured skin increases, which would allow more active ingredients to penetrate the skin transdermally. However, this disclosure shows the surprising finding that, at least for one and / or more large drugs in an emulsion (e.g., a nanoemulsion), relatively lower microneedle densities and / or relatively smaller microneedle puncture sizes achieve better results. That is, surprisingly, we have found that as the total surface area of ​​the skin punctured by microneedles decreases, the bioavailability of the large drugs in the emulsion applied to the skin increases. In some embodiments, about 2 to about 50 microneedles / cm 2 Microneedle densities in the range of can significantly improve transdermal delivery and / or bioavailability, even when compared to microneedling using relatively higher microneedle densities. Furthermore, this disclosure surprisingly covers densities of approximately 100 to 60,000 μm 2 This disclosure further reveals that microneedle conditioning of the skin using microneedle puncture sizes in the range of / microneedles can achieve significant transdermal delivery and / or bioavailability. The disclosure also further reveals that smaller microneedle puncture sizes (e.g., approximately 100 to approximately 30,000 μm) can be used. 2This document discloses that microneedle conditioning of the skin using microneedles (in the range of microneedles) can significantly improve transdermal delivery and / or bioavailability, even when compared to microneedling using relatively larger microneedle puncture sizes. In some embodiments, the microneedle puncture size is approximately 100 to approximately 30,000 μm 2 This could be within the realm of microneedles.

[0013] Prior to this disclosure, those skilled in the art would have understood from the literature that skin microneedle conditioning using any particular microneedle density would not be expected to improve transdermal delivery of even small molecules, let alone large drugs. Surprisingly, this disclosure uses approximately 2 to approximately 50 microneedles / cm². 2 This disclosure reveals that microneedle conditioning of the skin using microneedle densities in the range of approximately 100 to 30,000 μm can significantly improve the transdermal delivery of drugs such as botulinum toxin, which has a molecular weight of approximately 150,000 Da. Standard antibodies also have a similar molecular weight. Furthermore, this disclosure surprisingly covers approximately 100 to approximately 30,000 μm 2 This document discloses that microneedle conditioning of the skin using microneedle puncture sizes in the range of microneedles can significantly improve the transdermal delivery of such large drugs.

[0014] Those skilled in the art reading the present disclosure will logically understand that, in addition to reducing the total area punctured by reducing the density of the microneedles and / or reducing the puncture size of the microneedles, equivalent effects can be achieved by minimizing (e.g., reducing) the number of impressions performed on the skin of the treatment area by or using a microneedle array. Consistent with this understanding, in light of the above surprising observation of a reduction in the total surface area punctured (which can be achieved, at least in part, by a (more) smaller puncture size of the microneedles, which may incidentally have further advantages), the present disclosure further shows that the impression of a relatively smaller microneedle array can result in higher bioavailability than a relatively greater number of impressions. In some embodiments, a microneedle impression in the range of about 1 impression / cm 2 to about 5 impressions / cm 2 can achieve significant transdermal delivery and / or bioavailability.

[0015] The present disclosure shows that microneedle conditioning of the skin using a microneedle impression in the range of about 1 impression / cm 2 to about 4 impressions / cm 2 can significantly improve transdermal delivery and / or bioavailability even when compared to microneedling using a relatively greater number of microneedle impressions. In some embodiments, a microneedle impression in the range of about 1 impression to about 20 impressions performed at the treatment site can achieve significant transdermal delivery and / or bioavailability. Further, the present disclosure shows that microneedle conditioning of the skin using a microneedle impression in the range of about 1 impression to about 13 impressions performed at the treatment site can significantly improve transdermal delivery and / or bioavailability even when compared to microneedling using a relatively greater number of microneedle impressions.

[0016] Furthermore, those skilled in the art who read this disclosure will understand that reducing the total punctured area to which a large drug is applied by reducing the density of microneedles, reducing the puncture size of microneedles, and / or reducing the pressure of the microneedle array, compared to a relatively higher punctured surface, results in a reduction in the total volume of product administered to the microneedle-punctured skin area (i.e., the product formulation containing the large drug). Accordingly, this disclosure teaches those skilled in the art that improved transdermal delivery and / or higher bioavailability of large drugs can be achieved by reducing the total volume of product applied (e.g., of the formulation containing the large drug).

[0017] Where there are no specific teachings to be provided herein, those skilled in the art would typically expect that administering more product would increase the biological effect. Therefore, prior to this disclosure, it would have been anticipated that increasing the amount of a product containing a biologically active substance (e.g., a large drug), including when used in combination with MSCs and / or with a product composition containing an emulsion (e.g., a nanoemulsion), should achieve an increasingly greater biological effect. However, this disclosure surprisingly shows that beyond a certain “critical” or “threshold” product volume, further increasing the volume of a product composition applied to a given skin treatment area in combination with MSCs results in a decrease in effect (rather than an increase). Thus, beyond a certain critical product volume, administration (e.g., in combination with MSCs) of an increased volume of a product composition containing a biologically active substance (e.g., a large drug, particularly in the case of a product composition containing an emulsion) reduces the biological effect, despite the increasing volume of the biologically active substance administered.

[0018] In some embodiments, 1 drop / cm³ is applied to the skin. 2 ~About 5 drops / cm 2Significant transdermal delivery and / or bioavailability can be achieved by applying product volumes (e.g., compositions containing large drugs) in the range of approximately 1 / 100, in combination with skin conditioning using one or more presses of a microneedle array. In some embodiments, 1 drop / cm³ is applied to the skin. 2 ~About 4 drops / cm 2 Significant transdermal delivery and / or bioavailability can be achieved by applying product volumes (e.g., compositions containing large drugs) in the range of approximately 1 / 100, in combination with skin conditioning using one or more presses of a microneedle array. In some embodiments, 1 drop / cm³ is applied to the skin. 2 ~About 3 drops / cm 2 Significant transdermal delivery and / or bioavailability can be achieved by applying product volumes (e.g., compositions containing large drugs) in the range of approximately 1 / 100, in combination with skin conditioning using one or more presses of a microneedle array. In some embodiments, 1 drop / cm³ is applied to the skin. 2 ~about 2.5 drops / cm 2 Significant transdermal delivery and / or bioavailability can be achieved by applying product volumes (e.g., compositions containing large drugs) in the range of approximately 1 / 100 of the original volume, in combination with skin conditioning using one or more microneedle arrays. This disclosure relates to applying 1 drop / cm³ to the skin. 2 ~About 2 drops / cm 2 It is disclosed that by applying product volumes (e.g., compositions containing large drugs) in the range of approximately 1 / 100 in combination with skin conditioning using one or more microneedle arrays, transdermal delivery and / or bioavailability can be significantly improved even when compared to relatively higher product volumes (and / or doses) (e.g., large drugs).

[0019] In some embodiments, approximately 0.0001 ml / cm³ is applied to the skin. 2 ~about 0.04mls / cm 2Significant transdermal delivery and / or bioavailability can be achieved by applying product volumes in the range (e.g., compositions containing large drugs) in combination with skin conditioning using one or more microneedle arrays. In some embodiments, approximately 0.0001 ml / cm³ is applied to the skin. 2 ~about 0.05mls / cm 2 Significant transdermal delivery and / or bioavailability can be achieved by applying product volumes in the range (e.g., compositions containing large drugs) in combination with skin conditioning using one or more microneedle arrays. In some embodiments, approximately 0.0001 ml / cm³ is applied to the skin. 2 ~about 0.06mls / cm 2 Significant transdermal delivery and / or bioavailability can be achieved by applying product volumes in the range (e.g., compositions containing large drugs) in combination with skin conditioning using one or more microneedle arrays. In some embodiments, approximately 0.0001 ml / cm³ is applied to the skin. 2 ~about 0.065mls / cm 2 Significant transdermal delivery and / or bioavailability can be achieved by applying product volumes in the range (e.g., compositions containing large drugs) in combination with skin conditioning using one or more presses of a microneedle array. Furthermore, this disclosure describes how to deliver approximately 0.0025 ml / cm³ to the skin. 2 ~about 0.07mls / cm 2 It is disclosed that by applying product volumes within a certain range (e.g., compositions containing large drugs) in combination with skin conditioning using one or more microneedle arrays, transdermal delivery and / or bioavailability can be significantly improved, even when compared to relatively higher product volumes (and / or doses) of large drugs.

[0020] Among the advantages provided by certain embodiments of this disclosure (e.g., those with reduced volume of the product composition applied) is a reduction in the administration time (e.g., rubbing time) of the topical formulation. As already stated, this disclosure surprisingly shows that reducing the volume of a topically applied drug (and / or, in some embodiments, dose reduction) can actually achieve greater delivery (e.g., bioavailability) of the drug across the skin, particularly (but not limited to) when the drug is applied to an emulsion (e.g., nanoemulsion) formulation.

[0021] As previously stated, studies on microneedle length in small molecule delivery have demonstrated that relatively longer microneedle lengths improve small molecule delivery and / or bioavailability. See Yan's report (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 improves 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 logically understand that improved transdermal delivery and / or higher bioavailability of large drugs can be achieved by reducing the microneedle length of a 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 also understand that the longer the microneedle, the larger the surface area of ​​the microneedle itself, and therefore the larger the surface area of ​​the tissue being punctured. Surprisingly, this disclosure reveals that microneedle skin conditioning using relatively shorter microneedle lengths can yield greater bioavailability than using relatively longer microneedle lengths. In some embodiments, microneedle lengths in the range of about 1 μm to about 900 μm can achieve remarkable transdermal delivery and / or bioavailability. In some embodiments, microneedle lengths of less than 1400 μm are desirable, and in some embodiments, less than about 1100 μm or less than 1000 μm are desirable. In fact, this disclosure specifically demonstrates the remarkable 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 remarkable transdermal delivery and / or bioavailability. In some embodiments, this disclosure demonstrates remarkable efficacy 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 remarkable transdermal delivery and / or bioavailability.

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

[0023] Among the benefits offered by the unexpected finding presented herein—that shorter needle lengths can achieve more effective delivery (i.e., described as "increased bioavailability")—is reduced pain to the subject receiving microneedle skin conditioning for the administration of large drugs (e.g., in combination with and / or as part of a topical procedure). Microneedles of various lengths (e.g., including in the range of approximately 500 to 1400 μm) are readily available and have been described as particularly useful because bleeding can be minimized or avoided at these lengths. However, this disclosure understands that significant pain can be experienced even in the absence of bleeding, particularly at a length of 1400 μm, and sometimes even at shorter lengths. This disclosure, by demonstrating the efficacy 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 shorter, offers significant advantages in avoiding or reducing pain during local administration for large drugs, particularly drugs administered in the context of emulsion (or nanoemulsion) formulations (but not limited to).

[0024] This disclosure particularly 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 less microneedle pressure, relatively smaller product volume (and / or dose), and relatively shorter needle length) can significantly improve the transdermal delivery of large drugs, particularly (but not limited to) emulsion compositions (e.g., macroemulsion compositions and / or nanoemulsion compositions). As illustrated, for example, about 31 microneedles / cm before administration of a large drug (botulinum toxin). 2Preconditioning the skin by microneedle application using microneedle arrays with a microneedle density of less than 1 / 2 has surprisingly improved the delivery of large drugs across the skin. Specific examples included herein demonstrate such improved delivery under various conditions and / or environments (e.g., different skin sites, number of applications, etc.). Those skilled in the art will be aware of other variations (e.g., application sites, number of administrations, etc.) that fall within the scope of this disclosure.

[0025] Specific purpose nanoemulsion compositions 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, for example, the descriptions of one or more nanoemulsion compositions in PCT / US2006 / 26918; PCT / US06 / 46236; PCT / US2012 / 22276; and PCT / US2012 / 22279, each of which disclosures are included herein in whole by attribution).

[0026] Considering reports that transdermal delivery of solid nanoparticles of a size equivalent to that of droplets in the nanoemulsion compositions used herein (e.g., 10⁵ ± 2.92 nm) effectively delivers (or improves) even small molecule drugs transdermally across the skin, International Application PCT / US17 / 53333 already describes certain remarkable features achieved by the combination of microneedling and emulsion (e.g., nanoemulsion) techniques. 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 a very small amount of pigment 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, “There is a new consensus that NPs [nanoparticles] can be sufficiently deposited in hair follicles but cannot normally penetrate the stratum corneum.” Therefore, prior to this disclosure, those skilled in the art would expect that the use of microneedling techniques in nano-sized vehicles would not be able to effectively deliver even small molecule drugs (e.g., rhodamine dye) transdermally; indeed, it would have been thought impossible to improve the delivery and / or bioavailability of large drugs. International application PCT / US17 / 53333 demonstrates that microneedling can significantly improve the transdermal delivery of large drugs, particularly when used in conjunction with emulsion (e.g., nanoemulsion) systems. This disclosure further describes approximately 2 to approximately 50 microneedles / cm 2Microneedling using microneedle densities in the range of can significantly improve the transdermal delivery of large drugs and / or their bioavailability, especially when used in conjunction with emulsion systems (e.g., nanoemulsion systems), compared to using relatively higher microneedle densities. In some embodiments, approximately 40 microneedles / cm² 2 Less than (for example, about 2 to about 40 microneedles / cm) 2 Microneedling using a microneedle density of (in the range of) or approximately 35 microneedles / cm² 2 Less than (for example, about 2 to about 35 microneedles / cm) 2 Better microneedling using a microneedle density of (in the range of) or approximately 32 microneedles / cm² 2 Less than (for example, about 2 to about 32 microneedles / cm) 2 Microneedling using a microneedle density of (in the range of) or approximately 31 microneedles / cm² 2 Less than (for example, about 2 to 31 microneedles / cm) 2 Microneedling using a microneedle density of (in the range of) or approximately 30 microneedles / cm² 2 Less than (for example, about 2 to 30 microneedles / cm) 2 Microneedling using a microneedle density of (in the range of) or approximately 29 microneedles / cm² 2 Less than (for example, about 2 to 29 microneedles / cm²) 2 Microneedling using a microneedle density of (in the range of) or approximately 28 microneedles / cm² 2 Less than (for example, about 2 to about 28 microneedles / cm) 2 Microneedling using microneedle densities (in the range of ) can significantly improve the transdermal delivery of large drugs and / or their bioavailability compared to using relatively higher microneedle densities, especially when used in conjunction with emulsion systems (e.g., nanoemulsion systems).

[0027] Furthermore, this disclosure surprisingly covers approximately 100 to 60,000 μm. 2 This disclosure also reveals that microneedle conditioning of the skin using microneedle puncture sizes in the range of / microneedles can achieve significant transdermal delivery and / or bioavailability. Furthermore, this disclosure applies to, for example, approximately 100 to approximately 30,000 μm. 2 This document discloses that microneedling using smaller microneedle puncture sizes within the range of microneedles can significantly improve the transdermal delivery of large drugs and / or their bioavailability compared to using relatively larger microneedle puncture sizes, especially when used in conjunction with emulsion systems (e.g., nanoemulsion systems). In some embodiments, approximately 50,000 μm 2 / Less than a microneedle (e.g., approximately 100 to 50,000 μm) 2 Microneedling using microneedle puncture sizes (range of microneedles), and approximately 45,000 μm 2 / Less than a microneedle (e.g., approximately 100 to 45,000 μm) 2 (Microneedle range) Microneedling using microneedle puncture size, or approximately 40,000 μm 2 / Less than a microneedle (e.g., approximately 100 to 40,000 μm) 2 Microneedling using microneedle puncture sizes (range of microneedles), or approximately 35,000 μm 2 / Less than a microneedle (e.g., approximately 100 to 35,000 μm) 2 Better microneedling using microneedle puncture sizes (range of microneedles), or approximately 30,000 μm 2 / Less than a microneedle (e.g., approximately 100 to 30,000 μm) 2 Microneedling using microneedle puncture sizes (range of microneedles), or approximately 25,000 μm 2 / Less than a microneedle (e.g., approximately 100 to 25,000 μm) 2 Microneedling using microneedle puncture sizes (in the range of microneedles) can significantly improve the transdermal delivery of large drugs and / or their bioavailability compared to using relatively larger microneedle puncture sizes, especially when used in conjunction with emulsion systems (e.g., nanoemulsion systems).

[0028] In particular, this disclosure reveals 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., chemopreservatives and other techniques for disrupting or puncturing skin structures) are not utilized. Previous studies of transdermal delivery of drugs the same size as 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 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 microneedling site.

[0029] In some embodiments, the 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 technique described herein, without the use of further penetration enhancers. Separately or in addition thereto, in some embodiments, the 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 technique, without other disruption strategies. Thus, the techniques provided can achieve effective delivery and / or improved bioavailability without inflammation, irritation, and / or allergic reactions often associated with the use of disruptors.

[0030] Separately or in addition to the foregoing, this disclosure identifies the root cause of problems with certain prior approaches to binding large drugs, particularly large protein drugs (e.g., botulinum toxin, antibodies, etc.), within or onto microneedle structures. Typically, such conventional binding strategies utilize a liquid solution of the drug in question, which is applied to a microneedle and air-dried. Such a strategy was used to coat a microneedle with botulinum toxin in U.S. Patent Application Publication 2010 / 0228225. U.S. Patent Application Publication 2017 / 0209553 describes a microneedle loaded with botulinum. This disclosure understands that the botulinum coating or loading material produced thereby is unstable and therefore not commercially viable when used to manufacture products. In fact, even when such liquids are produced from powder materials, this disclosure understands that for many large drugs (e.g., botulinum toxin), powders and other solid materials not formed by a freeze-drying process can be extremely unstable. For example, according to Johnson, E. et al., "Botulinum toxin is extremely susceptible to denaturation due to surface denaturation, heat, and alkaline conditions. Lyophilization of botulinum toxin is the most economically sound and practical method for distributing a product in a stable form that is readily available to 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. This disclosure provides insights into how the use of emulsion compositions described herein (e.g., nanoemulsion compositions in some embodiments, and / or macroemulsion compositions in some embodiments) can protect or otherwise improve the stability of large drugs, particularly large protein drugs (including botulinum toxin and / or antibody drugs) for binding to microneedles.

[0031] This disclosure surprisingly provides effective techniques for improving the transdermal delivery and / or bioavailability of large drugs. In particular, this 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 this disclosure will understand that the teachings may be applicable to any topical formulation of large drugs. In some embodiments, this 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 may 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 techniques provided do not utilize skin disruption techniques, such as chemopreservatives. [Brief explanation of the drawing]

[0032] [Figure 1] Figure 1 shows the effect of changes in microneedle array density on the bioavailability of botulinum nanoemulsion formulations after MSC ("microneedle skin conditioning"), as determined by the survival rate in rat studies.

[0033] [Figure 2] Figure 2 shows the effect of changes in microneedle puncture size on the bioavailability of botulinum nanoemulsion formulations after MSC, as determined by the survival rate in rat studies. [Modes for carrying out the invention]

[0034] (definition) In this application, unless otherwise evident from the context, (i) the term “a” as used herein 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 listed components or steps, whether presented by themselves or together with one or more further components or steps; and (iv) the terms “about” and “approximately” may be understood to allow for standard variations as understood by those skilled in the art; and (v) endpoints, where provided in range.

[0035] Abrasion: As used herein, the term “abrasion” refers to any means that alters, destroys, removes, or disrupts the uppermost layer of skin. In some embodiments, abrasion refers to mechanical means that alters, destroys, removes, or disrupts the uppermost layer of skin. In some embodiments, abrasion refers to chemical means that alters, destroys, removes, or disrupts the uppermost layer of skin. For example, drugs such as scrubs, fine particles (e.g., magnesium or aluminum particles), acids (e.g., alpha-hydroxy acids or beta-hydroxy acids), and alcohols can cause abrasion. Generally, penetration enhancers, such as those described in Donovan (e.g., U.S. Patent Publication No. 2004 / 009180 and 2005 / 175636, and International Publication No. 04 / 06954) and Graham (e.g., U.S. Patent No. 6,939,852 and U.S. Patent Publication No. 2006 / 093624), are expected to cause abrasion. Naturally, those skilled in the art will understand that certain drugs may cause abrasion at one concentration or in combination with one or more other drugs, but may not cause abrasion under different conditions. Therefore, whether a particular substance is an "abrasive" depends on the situation. Abrasion can be readily assessed by those skilled in the art, for example, by observing redness or irritation of the skin and / or by histological examination of the skin, indicating alteration, 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 used for administration to a subject, e.g., a human, in appropriate circumstances. For example, in some embodiments, administration may be intraocular, oral, non-enteral, topical, etc. In some embodiments, administration may be bronchial (e.g., by bronchial infusion), buccal, skin (e.g., one or more of these, such as topical administration to the dermis, intradermal, intercutaneous, transdermal, etc.), enteral, intra-arterial, intradermal, gastric, intramedullary, intramuscular, intranasal, intraperitoneal, intrathecal, intravenous, intraventricular, intra-organ (e.g., intrahepatic), mucous membrane, nasal cavity, oral, rectal, subcutaneous, sublingual, topical, trachea (e.g., by intratracheal infusion), vagina, vitreous, etc. In some embodiments, administration may include intermittent administration (e.g., multiple administrations separated by time) and / or periodic administration (e.g., individual administrations separated by a common period of time). In some embodiments, administration may include continuous administration (e.g., perfusion) for at least a selected period of time.

[0037] Drugs: Generally, as used herein, the term “drug” can be used to refer to any chemical compound or entity, including, for example, polypeptides, nucleic acids, sugars, lipids, small molecules, metals, or combinations or complexes thereof. Where appropriate, as will be apparent to those skilled in the art, the term can be used to refer to a cell or organism, or a fraction, extract or component thereof, or an entity containing them. Separately or in addition thereto, as will be apparent to the context, the term can be used to refer to a natural product, in that it is found in nature and / or obtained from nature. In some cases, again as will be apparent to the context, the term can be used to refer to one or more entities that are artificial, in that they are designed, manipulated and / or produced by human action and / or not found in nature. In some embodiments, drugs may be available in isolated or pure forms; in some embodiments, drugs may be available in crude forms. In some embodiments, potential drugs may be provided as a collection or library, for example, that can be screened to identify or characterize the active substances among them. In some cases, the term may refer to a compound or entity that is or contains a polymer; in some cases, the term may indicate a compound or entity that contains 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 substantially does not contain any polymer and / or one or more specific polymer parts. In some embodiments, the term may refer to a compound or entity that lacks or substantially does not contain any polymer parts. In some embodiments, the term may refer to a molecular complex.

[0038] Antibody: As used herein, the term “antibody” refers to a polypeptide containing standard immunoglobulin sequence elements sufficient to provide specific binding to a particular target antigen. As is known in the art, naturally produced intact antibodies are tetrameric substances of approximately 150 kDa, consisting of two identical heavy-chain polypeptides (each approximately 50 kDa) and two identical light-chain polypeptides (each approximately 25 kDa), which bind to each other in what is commonly called a “Y-shaped” structure. Each heavy chain consists 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 carboxy-terminal CH3 (located at the base of the Y stem). A short region known as the “switch” connects the variable and constant regions of the heavy chain. The “hinge” connects the CH2 and CH3 domains to the rest of the antibody. Disulfide bonds in two of these hinge regions connect the two heavy-chain polypeptides to each other in the intact antibody. Each light chain consists of two domains: an amino-terminal variable (VL) domain followed by a carboxy-terminal constant (CL) domain, which are separated from each other by a different "switch." An intact antibody tetramer consists of two heavy-light chain dimers, where 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, resulting in the dimers being linked to each other and forming a tetramer. Naturally produced antibodies are also typically glycosylated at 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-strand sheets) packed together in a compressed antiparallel β-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 natural antibody folds, the FR region forms a beta sheet that provides 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, creating a single hypervariable antigen-binding site located at the tip of the Y structure.The Fc region of naturally occurring antibodies 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 to Fc receptors can be modulated by glycosylation or other modifications. In some embodiments, antibodies produced and / or utilized according to the present invention include a glycosylated Fc domain, which includes an Fc domain having such modified or manipulated glycosylation. For the purposes of the present invention, in some embodiments, any polypeptide or polypeptide complex containing a sufficient immunoglobulin domain sequence, as found in naturally occurring antibodies, may be referred to and / or used as such, regardless of whether such polypeptide is produced naturally (e.g., by an organism that reacts to an antigen) or 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 a constant region sequence characteristic of mouse, rabbit, primate, or human antibodies. In some embodiments, antibody sequence elements are humanized, primated, chimeric, etc., as known in the art. Furthermore, as used herein, the term “antibody” may, in appropriate embodiments (unless otherwise specified or evident from the context), refer to any construct or format known or developed in the art for utilizing the structural and functional characteristics of an antibody in a substitute presentation. For example, in embodiments, antibodies utilized according to the present invention include intact IgG, IgE and IgM, bi- or multi-specific antibodies (e.g., Zybody®), single-stranded Fv, polypeptide Fc fusions, Fab, camel antibodies, mask antibodies (e.g., Probodies®), etc. S mall M odular I mmuno P harmaceutical ("SMIP TMThe format may be selected from, but is not limited to, single-chain or tandem diabody antibodies (TandAb®), VHH, Anticalin®, Nanobody®, Minibody, BiTE®, Ankyrin Repeat Protein or DARPIN®, Avimer®, DaRT, TCR-like antibodies, Adnectin®, Affilin®, Trans-body®, Affibody®, TrimerX®, MicroProtein, Fynomer®, Centyrin®, and KALBITOR®. In some embodiments, the antibody may lack covalent modifications (e.g., glycan linkage) that it would have if produced naturally (e.g., in mammalian organisms). In some embodiments, the antibody may include covalent modifications (e.g., linkage of glycan, payload (e.g., detectable portion, therapeutic portion, catalytic portion, etc.), or other pendant groups (e.g., polyethylene glycol, etc.)).

[0039] Antibody drugs: 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 provide specific binding. Exemplary antibody drugs include human antibodies, primate-derived antibodies, chimeric antibodies, bispecific antibodies, humanized antibodies, and conjugated antibodies (i.e., antibodies conjugated or fused with other proteins, radiolabeled, or cytotoxins). S mall M odular I mmuno P harmaceutical ("SMIP TMThis includes, but is not limited to, single-chain antibodies, camel 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 its fragments)), multispecific antibodies (e.g., bispecific antibodies) formed from at least two intact antibodies, and antibody fragments, insofar as they exhibit the desired biological activity. In some embodiments, the term encompasses staple peptides. In some embodiments, the term encompasses one or more antibody-like conjugated peptide mimes. In some embodiments, the term encompasses one or more antibody-like conjugated scaffold proteins. In some embodiments, the term encompasses monobodies or adnectins. In many embodiments, the antibody drug is a polypeptide or comprises the amino acid sequence containing one or more structural elements recognized by those skilled in the art as complementarity-determining regions (CDRs); in some embodiments, the antibody drug is a polypeptide or comprises the amino acid sequence containing 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 contained CDR is identical in sequence or substantially identical to the reference CDR in that it contains 1 to 5 amino acid substitutions compared to the reference CDR. In some embodiments, the contained 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 CDR is substantially identical to the reference CDR in that it has an amino acid sequence that is otherwise identical to the reference CDR, although 1 to 5 amino acids in the included CDR are deleted, added, or substituted compared to the reference CDR. In some embodiments, the included CDR is substantially identical to the reference CDR in that it has an amino acid sequence that is otherwise identical to that of the reference CDR, although at least one amino acid is substituted compared to the reference CDR. In some embodiments, the included CDR is substantially identical to the reference CDR in that it has an amino acid sequence that is otherwise identical to that of the reference CDR, although 1 to 5 amino acids in the included CDR are deleted, added, or substituted compared to the reference CDR. In some embodiments, the antibody drug is a polypeptide having an amino acid sequence that is otherwise identical to that of the reference CDR. In some embodiments, the antibody drug is a polypeptide having an amino acid sequence that is homologous or largely homologous to an immunoglobulin variable domain by those skilled in the art. In some embodiments, the antibody drug is an antibody-drug conjugate or comprises such a conjugate.

[0040] Antibody Components: As used herein, antibody components refer to polypeptide elements (which may be complete polypeptides or parts of larger polypeptides, such as the fusion polypeptides described herein) that specifically bind to an epitope or antigen and contain structural features of one or more immunoglobulins. Generally, an antibody component is any polypeptide whose amino acid sequence contains elements characteristic of the antibody binding region (e.g., an antibody light chain or variable region or one or more complementarity-determining regions ("CDRs"), or an antibody heavy chain or variable region or one or more CDRs, in the presence of one or more framework regions as may be selected). In some embodiments, the antibody component is a full-length antibody or contains a full-length antibody. In some embodiments, the antibody component is a full-length or short-length antibody but contains at least one binding site (containing at least one, preferably at least two sequences, having 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 included "antibody component" encompasses polypeptides having a binding domain that exhibits at least 99% identity with an immunoglobulin-binding domain. In some embodiments, the “antibody component” included is any polypeptide having an immunoglobulin-binding domain, e.g., a binding domain exhibiting at least 70%, 75%, 80%, 85%, 90%, 95%, or 98% identity with a reference immunoglobulin-binding domain. The “antibody component” included may have the same amino acid sequence as that of an antibody (or a portion thereof, e.g., its antigen-binding portion) found in natural sources. 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 IgG, IgM, IgA, IgD, and IgE. It has been shown that the antigen-binding function of the antibody may be performed by a fragment of a full-length antibody.Such embodiments of antibodies may also be in a bispecific, dual-specific, or multispecific format, specifically binding to two or more different antigens. Examples of binding fragments included in the term “antigen-binding portion” of an antibody are (i)V. H , V L , C H 1 and C L (ii) A monovalent fragment consisting of domains, the Fab fragment; (ii) A bivalent fragment containing two Fab fragments linked by disulfide bridges in a hinge region, the F(ab')2 fragment; (iii) V H and C H Fd fragment consisting of one domain; (iv) V of a single arm of the antibody H and V L (v) Fv fragments consisting of domains; (v) Dab fragments containing a single variable domain (Ward et al., (1989) Nature 341:544-546); (vi) containing isolated complementarity-determining regions (CDRs). Furthermore, the V of two domains of the Fv fragment H and V L These are encoded by separate genes, but V H and V Lの The regions pair up to form a monovalent molecule (known as single-chain Fv (scFv); see, for example, Bird et al. (1988) Science 242:423-426; and Huston et al. (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883), which can be linked using recombinant methods by synthetic linkers that allow them to be constructed as a single protein chain. In some embodiments, the “antibody component” described herein is or includes such a single-chain antibody. In some embodiments, the “antibody component” is or includes a diabody. The diabody is V H and V LThis is a bivalent, bispecific antibody in which the domain is expressed on a single polypeptide chain, but a linker that is too short to allow pairing between two domains on the same chain is used, and therefore the domain pairs with a complementary domain on the other chain to create two antigen-binding sites (see, for example, 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 consists of a pair of tandem Fv segments (V) that, together with a complementary light chain polypeptide, form a pair of antigen-binding regions. H -C H 1-V H -C H 1) is a single-chain "linear antibody" containing or including such an antibody (Zapata et al., (1995) Protein Eng. 8(10): 1057-1062; and U.S. Patent No. 5,641,870). In some embodiments, the antibody components may have characteristic components of a chimeric or humanized antibody. Generally, a humanized antibody is a human immunoglobulin (recipient antibody) in which residues from the recipient's complementary determining region (CDR) are replaced by residues from the CDR of a non-human species (donor antibody), such as mouse, rat, or rabbit, having the desired specificity, affinity, and ability. In some embodiments, the antibody components may have characteristic components of a human antibody.

[0041] Antibody Fragments: As used herein, “antibody fragments” include, for example, a portion of an intact antibody, such as the antigen-binding or variable region of an 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, an antibody fragment includes an isolated fragment, an “Fv” fragment consisting of heavy and light chain variable regions, a recombinant single-chain polypeptide molecule (“ScFv protein”) in which the light and heavy chain variable regions are linked by a peptide linker, and a minimal recognition unit consisting of amino acid residues that mimic a hypervariable region. In many embodiments, the antibody fragment contains a sufficient sequence of a parent antibody, which 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 antibody antigen-binding fragments 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 regions (CDR) regions. Antibody antigen-binding fragments can be generated by any means. For example, antibody antigen-binding fragments can be generated enzymatically or chemically by fragmentation of intact antibodies and / or recombinantly from genes encoding partial antibody sequences. Separately or in addition to this, antibody antigen-binding fragments can be generated entirely or partially synthetically. Antibody antigen-binding fragments may optionally include single-chain antibody fragments. Separately or in addition to this, antibody antigen-binding fragments may include multiple chains linked together, for example, by disulfide bonds. Antibody antigen-binding fragments may optionally include multimolecular complexes. Functional antibody fragments typically contain at least about 50 amino acids, and more typically at least about 200 amino acids.

[0042] About: As used herein, the terms “approximately” or “about,” when applied to one or more target values, refer to values ​​similar to the given reference values. In some embodiments, unless otherwise specified or otherwise evident from the context (for example, when one or more target values ​​define a sufficiently narrow range, the application of such percentage variation is unnecessary), the terms “approximately” or “about” refer to a range of values ​​that are within the range of 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less than or equal to the given reference values.

[0043] Related: When the term is used herein, two events or entities are “related” to each other if the presence, level, and / or form of one correlate with that of the other. For example, a particular entity (e.g., polypeptide, gene signature, metabolite, microorganism, etc.) is considered related to a particular disease, disorder, or condition if its presence, level, and / or form correlate with the incidence and / or susceptibility to a disease, disorder, or condition (e.g., a related population). In some embodiments, two or more entities are physically “related” to each other if they interact directly or indirectly, and as a result are physically close to each other and / or remain physically close. In some embodiments, two or more entities that are physically related to each other are covalently bonded to each other; in some embodiments, two or more entities that are physically related to each other are not covalently bonded to each other but are non-covalently bonded, for example, by hydrogen bonds, van der Waals interactions, hydrophobic interactions, magnetism, and combinations thereof.

[0044] Biocompatibility: As used herein, the term “biocompatibility” refers to a material that, when placed in contact with such tissue, for example in vivo, does not cause significant harm to living tissue. 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 no more than 20% cell death and / or its administration in vivo does not induce significant inflammation or other such adverse effects.

[0045] Biodegradability: As used herein, the term “biodegradable” refers to a material that, when introduced into a cell, is broken down into components that the cell can reuse or discard (by cellular mechanisms such as enzymatic degradation, hydrolysis, and / or a combination thereof) without causing significant toxicity to the cell. In some embodiments, the components produced by the degradation of a biodegradable material are biocompatible and therefore do not induce significant inflammation and / or other adverse effects in vivo. In some embodiments, a biodegradable polymer material is broken down into its component monomers. In some embodiments, the degradation of a biodegradable material (including, for example, a biodegradable polymer material) includes hydrolysis of ester bonds. Separately or in addition thereto, in some embodiments, the degradation of a biodegradable material (including, for example, a biodegradable polymer material) includes cleavage of urethane bonds. Exemplary biodegradable polymers include, for example, polymers of hydroxy acids such as lactic acid and glycolic acid, and include, but are not limited to, poly(hydroxyl acids), poly(lactic acid) (PLA), poly(glycolic acid) (PGA), lactic acid-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 prolamin, such as zein, and polysaccharides such as alginic acid, cellulose derivatives and polyhydroxyalkanoates, such as polyhydroxybutyrate, mixtures and copolymers thereof. Those skilled in the art will be able to understand or determine when such polymers (for example, those related to the parent polymer by substantially identical structures, differing only in the substitution or addition of certain chemical groups, as is known in the art) are biocompatible and / or have biodegradable derivatives thereof.

[0046] Biologically Active Substances: As used herein, the term “biologically active substance” refers to a drug that, when administered to a subject, e.g., a human, has a specific biological effect. In some embodiments, a biologically active substance may be a therapeutic, cosmetic, and / or diagnostic substance. In some embodiments, a biologically active substance may be, or include, an entity or part that would be classified as an “active pharmaceutical ingredient” by the U.S. Food and Drug Administration. In some embodiments, a biologically active substance is a large drug. In some embodiments, a biologically active substance may be, or include, a drug whose presence correlates with a desired pharmacological and / or therapeutic, cosmetic, and / or diagnostic effect. In some embodiments, a biologically active substance is characterized by its biological effect being dose-dependent (e.g., increasing linearly with increasing dose, possibly over at least a first concentration range). In some embodiments, a drug is not considered a “biologically active substance” 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 botulinum toxin. The botulinum toxin may be present within the macroemulsion, on the surface of the macroemulsion, and / or within the micelle 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 botulinum toxin. The botulinum toxin may be present within the nanoemulsion, on the surface of the nanoemulsion, and / or within the micelle film defining 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 chain and / or heavy chain) of such neurotoxins that retain 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., complexes of 300, 600 and 900 kDa, for example) and purified (i.e., isolated) botulinum toxin (i.e., approximately 150 kDa, for example). “Purified botulinum toxin” is defined as botulinum toxin isolated or substantially isolated from other proteins, including the proteins for the botulinum toxin complex. The purified toxin may have a purity of over 95%, preferably over 99%. Those skilled in the art will understand that the present invention is not limited to a specific source of botulinum toxin. For example, botulinum toxin for use according to the present invention may be isolated from Clostridium botulinum, chemically synthesized, or produced recombinantly (i.e., in a host cell or organism other than Clostridium botulinum). Botulinum may be genetically engineered or chemically modified to act for a longer or shorter duration than botulinum toxin serotype A.

[0050] Carrier: As used herein, carrier refers to a diluent, adjuvant, additive, or vehicle to which the composition is administered together. In some exemplary embodiments, the carrier includes, for example, sterile liquids such as water and oils such as petroleum, animal, plant, or synthetic oils such as peanut oil, soybean oil, mineral oil, sesame oil, etc. In some embodiments, the 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 patient is simultaneously exposed to two or more treatment regimens (e.g., two or more therapeutic agents, a therapeutic agent and a therapeutic modality, etc.). In some embodiments, two or more regimens may be administered simultaneously; in some embodiments, such regimens may be administered sequentially (e.g., all “doses” of the first regimen are administered before any dose of the second regimen). In some embodiments, such drugs are administered in a duplicate-dosing regimen. In some embodiments, “administration” of combination therapy may include the administration of one or more drugs and / or modalities to a subject to be administered in combination with other drugs or modalities. For clarity, in some embodiments, two or more drugs or their active parts may 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 be administered together (or necessarily simultaneously) in a single composition.

[0052] Equivalent: As used herein, the term “equivalent” means two or more sets of drugs, entities, situations, conditions, etc., which do not have to be identical to one another, but are similar enough to allow comparison between them, so that a person skilled in the art may reasonably draw conclusions based on the differences or similarities observed. In some embodiments, equivalent sets of conditions, environments, individuals, or populations are characterized by several substantially identical features and one or a few diverse features. A person skilled in the art will understand, in context, what degree of identity is required in a given environment for two or more sets of such drugs, entities, situations, conditions, etc., to be considered equivalent. For example, a person skilled in the art will understand that sets of situations, individuals, or populations are equivalent to one another when they are characterized by a sufficient number and variety of substantially identical features to ensure a reasonable conclusion that differences between results obtained or observed phenomena under or with different sets of environments, individuals, or populations are caused by or indicate alterations in these altered features.

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

[0054] Inclusion: Any composition or method described herein as “inclusion” one or more of the specified components or steps is open-ended, meaning that the specified components or steps are essential, but other components or steps may be added within the scope of the composition or method. To avoid redundancy, any composition or method described as “comprising” (or “comprise”) one or more of the specified components or steps also describes a composition or method that “consisting essentially of” (or “consist essentially of” the same specified components or steps) in a corresponding and more limited way, meaning that the composition or method may include the essential components or steps of the specified, and may also include additional components or steps that do not substantially affect the basic, novel features of the composition or method. Any composition or method described herein as "containing" or "essentially consisting of" one or more specified components or steps is also understood to describe a corresponding, more limited, closed-end composition or method "consisting of" (or "consisting of") the specified components or steps, excluding any other components or steps not specified. In any composition or method disclosed herein, any known or disclosed equivalent of any specified essential 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” may be used to refer to a physically distinct unit of an active substance (e.g., a therapeutic or diagnostic agent) for administration to a subject. Typically, each such unit contains a predetermined amount of the active substance. In some embodiments, such an amount is a unit dose (or a full fraction thereof) appropriate for administration according to an administration regimen (i.e., a therapeutic administration regimen) that has been determined to correlate with a desired or beneficial outcome when administered to the population in question. Those skilled in the art will understand that the total amount of a therapeutic composition or drug to be administered to a particular subject may be determined by one or more physicians and may include administrations in 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 plural) administered individually to a subject, typically separated by time intervals. In some embodiments, a given therapeutic agent has a recommended dosing regimen, which may comprise one or more doses. In some embodiments, a dosing regimen comprises multiple doses, each separated in time from the other doses. In some embodiments, the individual doses are separated from each other by intervals of the same length; in some embodiments, a dosing regimen comprises multiple doses and at least two different intervals separating the individual doses. In some embodiments, all doses within a dosing regimen are the same unit dose. In some embodiments, the different doses within a dosing regimen are different amounts. In some embodiments, a dosing regimen comprises a first dose, followed by one or more further doses of a second dose different from the first dose. In some embodiments, a dosing regimen comprises a first dose, followed by one or more further doses of a second dose identical to the first dose. In some embodiments, the administration regimen correlates with the desired or beneficial outcome when administered to the entire population in question (i.e., it is a therapeutic administration regimen).

[0057] Emulsion: The term "emulsion" is used herein in accordance with the understanding in the art of "a system consisting of a liquid dispersed, with or without an emulsifier, in an immiscible liquid, usually in droplets larger than colloidal size." See, for example, the 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 to provide or contribute to a desired consistency or stabilizing effect. Suitable pharmaceutical additives include, for example, starch, glucose, lactose, sucrose, gelatin, malt, rice, wheat flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, dried skim milk, glycerol, propylene, glycol, water, ethanol, and the like.

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

[0060] Hydrophilicity: As used herein, the terms “hydrophilicity” and / or “polarity” refer to the tendency to mix with water or to dissolve readily in water.

[0061] Hydrophobic: As used herein, the terms "hydrophobic" and / or "nonpolar" refer to a tendency to repel water, to be immiscible with water, or to be insoluble in water.

[0062] Improvement, Increase, or Decrease: As used herein or in its grammatical equivalents, the terms “improvement,” “increase,” or “decrease” refer to values ​​relating to baseline measurements, such as measurements in the same individual before the commencement of the treatment described herein, or measurements in a control individual (or multiple control individuals) without the treatment described herein. In some embodiments, a “control individual” is an individual suffering from the same form of disease or injury as the individual being treated.

[0063] Macromolecules: The term “macromolecule” is used herein to generally describe molecules whose size exceeds about 100 kilodaltons (kDa). In some embodiments, macromolecules are 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, macromolecules are polymers or contain polymeric moieties or entities. In some embodiments, macromolecules are polypeptides or contain polypeptides. In some embodiments, macromolecules are nucleic acids or contain nucleic acids.

[0064] Large Drugs: As used herein, the term “large drug” generally refers to drugs having a molecular weight greater than approximately 100 kilodaltons (kDa). In some embodiments, macromolecules are greater than approximately 110 kDa, 120 kDa, 130 kDa, 140 kDa, 150 kDa, 160 kDa, 170 kDa, 180 kDa, 190 kDa, 200 kDa, 250 kDa, 300 kDa, 400 kDa, or 500 kDa. In some embodiments, large drugs are biologically active substances. In some embodiments, large drugs are one or more macromolecules. In some embodiments, large drugs are one or more molecular complexes. In some embodiments, large drugs are polypeptides. In some embodiments, large drugs are polypeptide complexes. In some embodiments, the large drug is or contains a bacterial toxin (e.g., botulinum toxin). In some embodiments, the large drug is or contains an antibody drug.

[0065] Macroemulsion: As used herein, the term “macroemulsion” refers to an emulsion in which at least a portion of the droplets have a diameter in the size range of several hundred nanometers to several micrometers. As will be understood by those skilled in the art, macroemulsion is characterized by droplets with a diameter greater than 300 nm. In some embodiments, the macroemulsion compositions used in accordance with this disclosure contain one or more larger drugs or one or more biologically active substances. In some embodiments, the larger drugs contained in the macroemulsion composition may be biologically active substances. As will be understood by those skilled in the art, macroemulsion compositions for use in accordance with this disclosure may be manufactured by any available means, including, for example, chemical or mechanical means. In some embodiments, the droplets in the macroemulsion have a size range of about 301 nm to about 1000 μm. In some embodiments, the macroemulsion has droplets with a size distribution of about 301 nm to about 1000 μm. In some embodiments, the droplets in the macroemulsion have a size range of about 500 nm to about 5000 μm. In some embodiments, the macroemulsion has droplets with a size distribution of approximately 500 nm to approximately 5000 μm.

[0066] Microneedle: As used herein, the term “microneedle” generally refers to an elongated structure of appropriate length, diameter, and shape for penetration into the skin. In some embodiments, microneedles are positioned and constructed (either by themselves 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, microneedles have a diameter that is constant along their length. In some embodiments, microneedles have a diameter that varies along their length. In some embodiments, microneedles have a diameter that tapers along their length. In some embodiments, the diameter of the microneedle is narrowest at the tip that penetrates the skin. In some embodiments, microneedles may be solid. In some embodiments, microneedles may be hollow. In some embodiments, microneedles may be tubular. In some embodiments, one end of the microneedle may be sealed. In some embodiments, multiple microneedles are used. In some embodiments, multiple microneedles are used in an array configuration. 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 be about 1 μm to about 2,000 μm in length. In some embodiments, the microneedle may be about 50 μm to about 400 μm in length. In some embodiments, the microneedle may be about 800 μm to about 1,500 μm in length.

[0067] Microneedle array pressing: As used herein, the term “microneedle array pressing” refers to microneedle pressing achieved by pressing microneedles and / or microneedle arrays onto the skin and then removing them from the skin. In some embodiments, microneedle arrays may be stamped onto the skin (e.g., using a microneedle array stamp). In some embodiments, microneedle arrays may 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 unit area measurement (e.g., square centimeter). In some embodiments, microneedle density is evaluated as the number of microneedles per unit area of ​​a microneedle array; in some embodiments, microneedle density is evaluated as the number of microneedle punctures per unit area of ​​a site to be microneedled; and in some embodiments, microneedle density is evaluated as the number of microneedles per unit area that simultaneously achieves the maximum or near-maximum possible skin penetration 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 area in question is flat (e.g., a microneedle array stamp), curved (e.g., a microneedle array roller) or irregular. Those skilled in the art will understand that, for example, if the array has needles of different lengths, and / or if the site to be microneedled has such topological diversity that not all needles can actually puncture the skin when the array is applied to the site, then evaluating the density of microneedles as microneedle punctures per unit area of ​​the site to be microneedled may be particularly useful.

[0069] Microneedle puncture size: As used herein, the terms “microneedle puncture size” or “microneedle puncture hole size” refer to the calculated puncture area created by each microneedle in a microneedle array, achieved after pressing a microneedle and / or microneedle array onto the skin and then 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 droplets have a diameter in the nanometer size range. As will be understood by those skilled in the art, nanoemulsions are characterized by droplets with a diameter of 300 nm or less. In some embodiments, the nanoemulsion compositions used in accordance with this disclosure contain one or more large drugs or one or more biologically active substances. In some embodiments, the large drugs contained in the nanoemulsion composition may be biologically active substances. As will be understood by those skilled in the art, nanoemulsion compositions for use in accordance with this disclosure may be manufactured by 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] Nanoparticles: As used herein, the term "nanoparticles" refers to solid particles 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 living organism to which the provided composition is administered or may be administered for, for example, experimental, diagnostic, preventive, cosmetic and / or therapeutic purposes. Typical patients include animals (e.g., mammals, e.g., mice, rats, rabbits, non-human mammals and / or humans). In some embodiments, the patient is human. In some embodiments, the patient has or is 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 or has been receiving a particular treatment to diagnose and / or treat a disease, disorder or condition.

[0073] Penetration enhancers: As used herein, the term “penetration enhancer” refers to a substance whose presence or level correlates with an increase in the penetration of a drug of interest across the skin compared to that observed in its absence. In some embodiments, a penetration enhancer is characterized by its ability to decompose and / or destroy skin structures. In some embodiments, a penetration enhancer is or comprises a chemical agent (e.g., a chemical substance or an enzyme). For example, chemical agents that can damage, destroy and / or decompose one or more components of the stratum corneum may include, for example, alcohols, e.g., short-chain alcohols, long-chain alcohols or polyalcohols; amines and amides, e.g., ureas, amino acids or their esters, amides, AZONE®, derivatives of AZONE®, pyrrolidone or derivatives of pyrrolidone; terpenes and derivatives of terpenes; fatty acids and their esters; macrocyclic compounds; surfactants (tenside); or sulfoxides (e.g., dimethyl sulfoxide (DMSO), decyl methyl sulfoxide, etc.); surfactants (anionic, cationic and nonionic surfactants); polyols; essential oils; and / or hyaluronidases. In some embodiments, a penetration enhancer may be irritating in that it may cause an inflammatory and / or allergic reaction when the substance is applied to the skin. In some embodiments, a penetration enhancer is not irritating. In some embodiments, a penetration enhancer may be, or may include, a chemical agent that does not damage, destroy or degrade skin structure, but nevertheless correlates with an increased penetration of the drug of interest across the skin, compared to that observed in its absence. In some embodiments, copeptides, carrier molecules, and carrier peptides may be penetration enhancers that do not damage, destroy, and / or degrade skin structure. In some embodiments, copeptides, carrier molecules, and carrier peptides may be penetration enhancers that do not irritate the skin. The term “penetration enhancer” does not include mechanical devices (e.g., needles, scalpels, etc.) or their equivalents (e.g., other damaging procedures).Furthermore, those skilled in the art will understand that structures such as nanoparticles or emulsions are not chemical agents, and therefore, even if their presence correlates with improved skin penetration of the target drug, which may be related to the structure, they are not chemical penetration enhancers.

[0074] Pharmaceutical Composition: As used herein, the term “pharmaceutical composition” refers to a composition in which an active substance is formulated together with one or more pharmaceutically acceptable carriers. In some embodiments, the active substance is present in a unit dose appropriate for administration in a therapeutic regime that exhibits a statistically significant probability of achieving a predetermined therapeutic effect when administered to a relevant population. In some embodiments, the pharmaceutical composition may be specifically formulated for administration in solid or liquid form, including sterile liquid or suspension or sustained-release formulations, for example, as a gel, cream, ointment or controlled-release patch or spray applied to the skin, lungs, or oral cavity; or as a pessary, cream or foam applied to the vagina or rectum; sublingual; intraocular; transdermal; or transnasally, lungs, and other mucosal surfaces.

[0075] pharmaceutically acceptable: As used herein, the term “pharmaceutically acceptable” applies to carriers, diluents, or additives used to formulate the compositions described herein, meaning that the carrier, diluent, or additive must be compatible with the other components of the composition and must not be harmful to its recipient.

[0076] Medicinally Acceptable Carrier: As used herein, the term “medically acceptable carrier” means a medically acceptable material, composition, or vehicle, such as a liquid or solid excipient, diluent, additive, or solvent encapsulating a material, used to transport or deliver a compound of interest from one organ or part of the body to another. Each carrier must be “acceptable” in the sense that it is compatible with the other components of the formulation and does not harm the subject or patient. Some examples of substances that can function as pharmaceutically acceptable carriers include sugars, e.g., lactose, glucose, and sucrose; starches, e.g., corn starch and potato starch; cellulose and its derivatives, e.g., sodium carboxymethylcellulose, ethylcellulose, and cellulose acetate; powdered tragacanth; malt; gelatin; talc; additives, e.g., cocoa butter and suppository waxes; oils, e.g., peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, medium-chain triglycerides, and soybean oil; glycols, e.g., propylene glycol; polyols, e.g., glycerin, sorbitol, mannitol, and polyethylene glycol; esters, e.g., ethyl oleate and ethyl laurate; agar; buffers, e.g., 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 and suitable substances used in pharmaceutical formulations.

[0077] Premix: As used herein, the term “premix” refers to any combination of components used to subsequently produce a nanoemulsion composition or to produce a nanoemulsion according to the present invention. For example, a premix is ​​any set of components that, when subjected to high shear forces, produce a nanoemulsion according to the present invention. In some embodiments, a premix is ​​a set of components that, when subjected to high shear forces, produce a nanoemulsion composition such as a homogeneous nanoemulsion composition. A premix often contains a liquid dispersion medium and other components sufficient to produce a nanoemulsion within the dispersion medium. According to some embodiments of this disclosure, one or more large drugs may be included in the premix. According to some embodiments of this disclosure, one or more biological drugs may be included in the premix. According to the present invention, 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 contain one or more surfactants, penetration enhancers, and / or other substances. In some embodiments, a premix contains a solution. In some embodiments, the premix comprises botulinum toxin, an antibody, another biologically active substance, and / or an osmotic enhancer, in which case the botulinum toxin, antibody, another biologically active substance, and / or osmotic enhancer are in solution before a high shear force is applied to the premix.

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

[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 non-amino acid portions (e.g., glycoproteins, proteoglycans, etc.) and / or may be processed or modified in other ways. Those skilled in the art will understand that a “protein” may be a complete polypeptide chain (with or without a signal sequence) as produced by a cell, or a characteristic portion thereof. Those skilled in the art will understand that a protein may contain multiple polypeptide chains linked, for example, by one or more disulfide bonds or by other means. Polypeptides may contain L-amino acids, D-amino acids, or both, and may contain any of the various 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 native amino acids, non-native 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, its biologically active portion, and / or a characteristic portion thereof.

[0080] Polypeptide: As used herein, the term “polypeptide” generally has the meaning recognized in the art of a polymer of at least three amino acids. Those skilled in the art will understand that the term “polypeptide” is intended to be sufficiently general to encompass not only polypeptides having the complete sequences described herein, but also polypeptides representing functional fragments of such complete polypeptides (i.e., fragments that retain at least one activity). Furthermore, those skilled in the art will understand that protein sequences generally tolerate some substitutions without disrupting 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 containing at least 3–4, often more than 20 amino acids. Polypeptides may contain L-amino acids, D-amino acids, or both, and may contain any of the various amino acid modifications or analogs known in the art. Useful modifications include, for example, terminal acetylation, amidation, and methylation. In some embodiments, proteins may contain native 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, proteins may be antibodies, antibody fragments, their biologically active portions, and / or characteristic portions thereof.

[0081] References: The references used herein describe the standard or control on which the comparison is made. For example, in some embodiments, the drug, animal, individual, population, sample, regimen, sequence, or value of interest is compared to the drug, animal, individual, population, sample, regimen, sequence, or value of reference or control. In some embodiments, the reference or control is tested and / or determined substantially concurrently with the test or determination of interest. In some embodiments, the reference or control is a historical reference or control and is embodied in tangible media as desired. Typically, as understood by those skilled in the art, the reference or control is determined or characterized under conditions or circumstances equivalent to those under evaluation. Those skilled in the art will understand when sufficient similarity exists to determine reliance on and / or comparison to a particular possible reference or control.

[0082] Self-administration: As used herein, the term “self-administration” refers to a situation in which a subject has the ability to administer a composition to himself or her without requiring medical supervision. In some embodiments of the present invention, self-administration may be carried out outside of a clinical setting. For example, in some embodiments of the present invention, a facial cosmetic cream may be administered by the subject at home.

[0083] Small molecules: Generally, in the art, "small molecules" are understood to be organic molecules with a size of less than about 5 daltons (Kd). In some embodiments, small molecules are less than about 3 Kd, 2 Kd, or 1 Kd. In some embodiments, small molecules are less than about 800 daltons (D), 600 D, 500 D, 400 D, 300 D, 200 D, or 100 D. In some embodiments, small molecules are nonpolymeric. In some embodiments, small molecules are not proteins, peptides, or amino acids. In some embodiments, small molecules are not nucleic acids or nucleotides. In some embodiments, small molecules are not sugars or polysaccharides.

[0084] Subject: As used herein, “Subject” means a living organism, typically a mammal (e.g., a human, including in some embodiments a prenatal human form). In some embodiments, the Subject suffers from a relevant disease, disorder, or condition. In some embodiments, the Subject is susceptible to a disease, disorder, or condition. In some embodiments, the Subject exhibits one or more symptoms or characteristics of a disease, disorder, or condition. In some embodiments, the Subject does not exhibit any symptoms or characteristics of a disease, disorder, or condition. In some embodiments, the Subject is a person having one or more characteristics that are characteristic of susceptibility to or risk of a disease, disorder, or condition. In some embodiments, the Subject is a patient. In some embodiments, the Subject is an individual that has been diagnosed and / or treated.

[0085] Substantially: As used herein, the term “substantially” refers to a qualitative condition of the overall or nearly overall extent or degree of a particular feature or characteristic. Those skilled in the biological art will understand that biological and chemical phenomena, if any, rarely complete and / or proceed completely, 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 Drug: As used herein, the term “therapeutic drug” generally refers to any drug that, when administered to an organism, induces a desired pharmacological effect. In some embodiments, a drug is considered a therapeutic drug if it exhibits a statistically significant effect across a suitable population. In some embodiments, a suitable population may be a population of model organisms. In some embodiments, a suitable population may be defined by various criteria such as a specific age group, sex, genetic background, or pre-existing clinical condition. In some embodiments, a therapeutic drug is a substance that can be used to reduce, improve, alleviate, 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 drug” is a drug that has been or needs to be approved by a government agency before it can be marketed for administration to humans. In some embodiments, a “therapeutic drug” is a drug that requires a prescription for administration to humans. In some embodiments, a drug is not considered a “therapeutic drug” if it merely enhances the delivery of different drugs that actually achieve the desired effect.

[0087] Therapeutic dose: As used herein, a therapeutic dose means the amount administered to produce the desired effect. In some embodiments, the term refers to an amount that, when administered according to a therapeutic dosing regimen to a population affected or susceptible to a disease, disorder, and / or condition, is sufficient to treat the disease, disorder, and / or condition. In some embodiments, a therapeutic dose is one that reduces the incidence and / or severity of one or more symptoms of the disease, disorder, and / or condition, and / or delays their onset. Those skilled in the art will understand that the term “therapeutic dose” does not, in practice, require that the success of the treatment be achieved in a particular individual. Rather, a therapeutic dose may be an amount that, when administered to a patient requiring such treatment, provides a particular desired pharmacological response in a significant number of subjects. In some embodiments, a reference to a therapeutic dose may refer to an amount measured in one or more specific tissues (e.g., tissues affected by the disease, disorder, or condition) or body fluids (e.g., blood, saliva, serum, sweat, tears, urine, etc.). Those skilled in the art will understand 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 an administration regimen.

[0088] Treatment regimen: As used herein, the term “treatment regimen” refers to a regimen of administration that, when administered across a relevant population, may correlate with a desired or beneficial therapeutic outcome.

[0089] Treatment: As used herein, the term “treatment” (“treat” or “treating”) refers to any administration of therapy that partially or completely reduces, 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 for subjects who do not show signs of the disease, disorder and / or condition in question, and / or for subjects who show only initial signs of the disease, disorder and / or condition. Separately or in addition to this, such treatment may be for subjects who show one or more established signs of the disease, disorder and / or condition in question. In some embodiments, treatment may be for subjects who have been diagnosed with the disease, disorder and / or condition in question. In some embodiments, treatment may be for subjects who are known to have one or more susceptibility factors that are statistically correlated with an increased risk of developing the disease, disorder and / or condition in question.

[0090] Uniform: When used herein in reference to nanoemulsion compositions, the term "uniform" refers to a nanoemulsion composition in which individual droplets have a specific range of droplet diameter sizes. For example, in some embodiments, a uniform nanoemulsion composition is one in which the difference between the minimum and maximum diameters does not exceed about 300, 250, 200, 150, 100, 90, 80, 70, 60, 50 nm, or less. In some embodiments, droplets in the uniform large drug nanoemulsion composition of the present invention (e.g., large drug-containing droplets) have diameters of about 300, 250, 200, 150, 130, 120, 115, 110, 100, 90, 80 nm, or less. In some embodiments, droplets in the uniform large drug nanoemulsion composition of the present invention (e.g., large drug-containing droplets) have diameters in the range of about 10 nm to about 300 nm. In some embodiments, droplets in the homogeneous large drug nanoemulsion composition of the present 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, droplets in the large drug nanoemulsion composition of the present invention (e.g., large drug-containing droplets) 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, or 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, the majority of droplets (e.g., large drug-containing droplets) in the homogeneous nanoemulsion composition of the present invention have a diameter less than or within a specified range. In some embodiments, the majority are 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 homogeneous 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. In many embodiments, a variant is also functionally different from its reference entity. Generally, whether a particular entity is appropriately considered a “variant” of a reference entity depends 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. A variant is, by definition, a distinct chemical entity that shares one or more such characteristic structural elements. To give some examples, a small molecule may have a characteristic core structural element (e.g., a macrocyclic core) and / or one or more characteristic pendant moieties, and as a result, mutants of a small molecule may share the core structural element and characteristic pendant moieties but differ in other pendant moieties and / or the binding type within the core (e.g., single vs. double, E vs. Z); a polypeptide may have a characteristic sequence element composed of multiple amino acids that have designated positions relative to each other in linear or three-dimensional space and / or contribute to a specific biological function; and a nucleic acid may have a characteristic sequence element composed of multiple nucleotide residues that have designated positions relative to each other in linear or three-dimensional space. For example, a mutant polypeptide may differ from a reference polypeptide as a result of one or more differences in the amino acid sequence and / or one or more differences in chemical moieties (e.g., carbohydrates, lipids, etc.) covalently bonded to the polypeptide backbone. In some embodiments, the mutant polypeptide exhibits overall sequence identity with the reference polypeptide, which is at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 99%, apart from, optionally, conserved amino acid substitutions. Separately or in addition to this, 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 the biological activities of one or more reference polypeptides.In some embodiments, the mutant polypeptide lacks the biological activity of one or more reference polypeptides. In some embodiments, the mutant polypeptide exhibits a reduced level of one or more biological activities compared to the reference polypeptide. In many embodiments, the target polypeptide is considered a "mutant" of the parent or reference polypeptide if it has an amino acid sequence that is identical to that of the parent but has a few sequence changes at specific positions. Typically, less than 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, or 2% of the residues in the mutant are substituted compared to the parent. In some embodiments, the mutant has 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 substituted residue compared to the parent. Often, the mutant has very few (e.g., less than 5, 4, 3, 2, or 1) substituted functional residues (i.e., residues involved in specific biological activity). Furthermore, the mutant typically has 5, 4, 3, 2, or 1 or fewer additions or deletions, and often no additions or deletions compared to the parent. Furthermore, any addition or deletion is typically less than approximately 25, 20, 19, 18, 17, 16, 15, 14, 13, 10, 9, 8, 7, or 6 residues, and generally less than approximately 5, 4, 3, or 2 residues. In some embodiments, the parent or reference polypeptide is one found in nature.

[0092] (Detailed description of a specific embodiment) Transdermal drug delivery In some embodiments, the present invention provides techniques for improving the transdermal delivery and / or bioavailability of large drugs (e.g., botulinum toxin, antibodies). In some embodiments, the disclosure teaches that particularly advantageous results are achieved when microneedling techniques are combined with emulsion compositions. In some embodiments, the microneedling techniques are combined with lotions, creams, or liquid compositions, which in turn may be or include emulsion compositions (e.g., macroemulsion compositions and / or nanoemulsion compositions). In some embodiments, the techniques provided do not utilize penetration enhancers. In some embodiments, the techniques provided do not utilize chemopreservatives that damage, destroy, and / or degrade the skin. In some embodiments, the techniques provided do not utilize chemopreservatives.

[0093] Human skin consists of the dermis and epidermis. The epidermis has several layers of tissue, namely the stratum corneum, stratum lucidum, stratum granulosum, stratum spinosum, and stratum basale (specified in order from the outer surface to the inner surface of the skin).

[0094] The stratum corneum generally, and perhaps especially, presents the biggest hurdle in the transdermal delivery of large drugs. Typically about 10–15 μm thick, the stratum corneum consists of several layers of flat, keratinized cells (keratinocytes). The spaces between keratinocytes are filled with lipid structures, which can play a crucial 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 beneath the stratum corneum is approximately 150 μm thick. The dermis is approximately 1-2 mm thick and is located beneath the epidermis. The dermis is innervated by various capillaries and nerve processes.

[0096] Transdermal administration is generally the focus of research seeking to provide an alternative route of administration that does not have the undesirable consequences associated with injection and oral delivery. For example, needles often cause local pain and can expose patients receiving injections to bloodborne infectious diseases. Oral administration can suffer from low bioavailability of the drug due to the highly acidic environment of the patient's stomach.

[0097] Efforts have been made to develop transdermal administration technologies for certain drugs in an attempt to overcome these drawbacks by providing non-invasive administration. Generally, it is desirable to minimize damage to the patient's skin with transdermal administration. Therefore, transdermal administration can reduce or eliminate pain associated with injection, reduce the possibility of blood contamination, and improve the bioavailability of the drug once it is incorporated into the body.

[0098] Traditionally, attempts at transdermal administration have focused on disrupting and / or breaking down the stratum corneum. Some attempts have involved the use of chemopreservatives. These chemopreservatives may function to break down and / or destroy skin structures. In some embodiments, the chemopreservative is or comprises a chemical agent (e.g., a chemical or enzyme that can disrupt and / or break down one or more components of the stratum corneum). In some embodiments, the chemopreservative may be irritating in that it may cause inflammatory and / or allergic reactions when the drug is applied to the skin.

[0099] "However, the main limitation of transdermal drug delivery agents is that their effectiveness 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). Transdermal drug delivery agents tend to have poor efficacy and safety profiles. "They do not achieve the desired skin disruption, and their ability to increase transcutaneous transport is low and variable." Ibid.

[0100] Some attempts involve the use of mechanical devices to bypass or excise portions of the stratum corneum. Others involve the use of ultrasound or iontophoresis to facilitate the penetration of pharmaceuticals through the skin. In most cases, the goal is to enable drugs, typically small molecules, to pass into the capillary bed of the dermis, where they can be systemically incorporated into the target 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 demonstrated to improve the transdermal delivery of various small drugs, such as calcein (approximately 623 kDa), desmopressin (approximately 1070 kDa), diclofenac (approximately 270 kDa), methyl nicotinate (approximately 40 kDa), bischloroethylnitrosourea (approximately 214 kDa), insulin (approximately 5.8 kDa), bovine serum albumin (approximately 66.5 kDa), and ovalbumin (approximately 45 kDa). However, until this disclosure, problems remained in improving the delivery and / or bioavailability of large drugs, particularly those larger than 100 kDa.

[0102] Transdermal delivery of macromolecules is recognized as presenting a major challenge. To date, microneedling, particularly microneedle skin preconditioning using relatively low microneedle density and / or relatively small microneedle puncture size (e.g., puncture size per microneedle), has not been considered to affect or be effective for transdermal administration of macrophages. For example, studies using 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) have shown that while microneedle pretreatment significantly increases the penetration of each of these peptides, the skin penetration of peptides is molecular weight-dependent, decreasing 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 MSC sandpaper abrasion, tape stripping, and single-puncture subcutaneous needle models were compared in a study of the effect of larger FITC (fluorescein isothiocyanate) conjugate molecule molecular size on transdermal delivery, similar results were found in all methods. Furthermore, when tested on untreated skin, transdermal drug delivery decreased again as the size of the test molecule increased (FITC conjugates of 4.3, 9.6, and 42.0 kDa). Tape stripping was found to be the most effective method, while sandpaper abrasion caused the most skin damage. (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, molecular weight 538), dextran-Cascade Blue (DCB, molecular weight 10 kDa), and FITC-conjugated dextran (FITC-Dex, molecular weight 72 kDa). In these studies, microneedles of various lengths (300, 550, 700, or 900 μm) were used to puncture dermal human skin, and the diffusion of each of the above compounds was evaluated. Transport of each compound was observed in all but the 300 μm microneedle array, although degradation of DCB and FITC-Dex was observed.

[0105] As prior art has shown, as molecular size increases, transdermal penetration using MSCs ("microneedle skin conditioning") decreases to a minimum, or even to the point of nonexistence. Even when some minimal penetration is observed, larger molecules have been observed to be degraded and become biologically inactive. Recent techniques have been developed that achieve various advantages by combining emulsion and microneedling techniques for the transdermal delivery of large drugs of interest (see, for example, International Application PCT / US17 / 53333); in some embodiments, these techniques have shown that they can achieve particularly remarkable improvements in the transdermal delivery of large molecular structures without the use of mechanical or chemical penetration enhancers. 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. This disclosure reveals that the microneedling techniques described herein can, surprisingly, further enhance the transdermal delivery of large drugs (e.g., those with molecular weights greater than 100 kDa (e.g., botulinum toxin)) and / or improve their bioavailability. Clostridium botulinum is a complex protein, and for a protein to be biologically active, three regions or functional parts must remain intact. Therefore, if any one of the three regions of the protein is damaged, the protein becomes 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 Application Publication No. 5512547. Therefore, under the microneedling conditions described by Wu, a considerable level of degradation and inactivation of Clostridium botulinum is expected.

[0106] In particular, this disclosure reveals that the microneedling techniques described herein can improve the transdermal delivery of (e.g., large drugs, particularly from macroemulsion or nanoemulsion compositions) and / or improve their bioavailability when other penetration enhancers, especially disruptors (i.e., chemopreservatives and other techniques that disrupt or puncture skin structures).

[0107] Microneedling This disclosure provides the remarkable finding that the MSCs described herein can remarkably improve the transdermal delivery of large drugs. In some embodiments, large drugs may be formulated as creams and / or lotions. In some embodiments, large drugs may be combined with one or more biologically active substances. In some embodiments, large drugs may be formulated as or in emulsion compositions (e.g., as macroemulsions or nanoemulsions). In some embodiments, emulsions containing one or more large drugs may be formulated as creams and / or lotions.

[0108] In some embodiments, the microneedle (MN) arrays for use according to this disclosure feature or share these features, being a minimally invasive system developed to overcome some of the drawbacks commonly associated with the use of hypodermic and subcutaneous needles and to improve patient comfort and compliance. Such drawbacks include, for example, the possibility of misplacement of the needle tip with hypodermic needles because healthcare professionals cannot accurately visualize where the needle is advancing; such misplacement can cause adverse reactions such as drooping eyelids ("ptosis") when botulinum toxin is inaccurately injected into the face. MNs are less prone to such problems. Other advantages of MNs include the absence of bleeding, minimizing the introduction of pathogens through the MN-made holes, and eliminating variability in transdermal administration. Other advantages include the possibility of self-administration, reduced risk of accidental needle stick injuries, reduced risk of infection transmission, and ease of disposal. In some embodiments, the MN is a set of microscopic protrusions assembled on one side of a support, such as a patch or device (e.g., a stamp, roller, array, applicator, pen).

[0109] In some embodiments, MNs for use according to this disclosure may be designed and / or constructed in arrays to improve skin contact and facilitate skin penetration. In some embodiments, the MNs utilized are of a length, width, and shape suitable for creating 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 microneedles (MNs) may be solid, coated, porous, soluble, hollow, or hydrogel MNs. Solid MNs create micropores in the skin, thereby increasing the delivery of drug formulations (e.g., the "pork-and-patch" method). Coated MNs allow for rapid dissolution of the coated drug into the skin (e.g., the "coat-and-pork" method). Soluble MNs allow for rapid and / or controlled release of a drug incorporated within the microneedle. Hollow MNs may be used to puncture the skin and allow for the release of the composition after active injection or diffusion of the formulation through the microneedle pores (e.g., the "pork-and-flow" method). In the case of soluble MNs, the MN may function as a drug depot, either holding the drug composition until released by dissolution in the case of soluble MNs, or swelling in the case of hydrogel MNs (e.g., the "pork-and-release" method). However, as already described herein, in many embodiments, large drugs are not delivered by injection via one or more microneedles. In other words, in many embodiments, any microneedles used according to such embodiments are not coated with, loaded with, or manufactured using a large drug in any way that achieves the delivery of a large drug. Alternatively, in some embodiments, the microneedles described herein (in MSC or otherwise) used according to this disclosure may contain and / or deliver a large drug if the large drug is formulated into a macro or nanoemulsion described herein. Thus, as will be understood by those skilled in the art reading this specification, skin treatment with microneedles for delivering a large drug (e.g., by injection via microneedles, by release of the microneedle coating, or by release from dissolved microneedles) is not microneedle skin conditioning.

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

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

[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 MSC described herein is adjusted based on the thickness of the skin at the treatment site. In some embodiments, the MN or MN array includes microneedles of about 100 μm in length. In some embodiments, the MN or MN array includes microneedles of about 150 μm in length. In some embodiments, the MN or MN array includes microneedles of about 200 μm in length. In some embodiments, the MN or MN array includes microneedles of about 250 μm in length. In some embodiments, the MN or MN array includes microneedles of about 300 μm in length. In some embodiments, the MN or MN array includes microneedles of about 350 μm in length. In some embodiments, the MN or MN array includes microneedles of about 400 μm in length. In some embodiments, the MN or MN array includes microneedles of about 450 μm in length. In some embodiments, the MN or MN array includes microneedles of about 500 μm in length. In some embodiments, the MN or MN array includes microneedles approximately 550 μm in length. In some embodiments, the MN or MN array includes microneedles approximately 600 μm in length. In some embodiments, the MN or MN array includes microneedles approximately 650 μm in length. In some embodiments, the MN or MN array includes microneedles approximately 700 μm in length. In some embodiments, the MN or MN array includes microneedles approximately 750 μm in length. In some embodiments, the MN or MN array includes microneedles approximately 800 μm in length. In some embodiments, the MN or MN array includes microneedles approximately 850 μm in length. In some embodiments, the MN or MN array includes microneedles approximately 900 μm in length. In some embodiments, the MN or MN array includes microneedles approximately 950 μm in length.In some embodiments, the MN or MN array includes microneedles having a length of about 1000 μm. In some embodiments, the MN or MN array includes microneedles having a length of about 1100 μm. In some embodiments, the MN or MN array includes microneedles having a length of about 1200 μm. In some embodiments, the MN or MN array includes microneedles having a length of about 1300 μm. In some embodiments, the MN or MN array includes microneedles having a length of about 1400 μm. In some embodiments, the MN or MN array includes microneedles having a length of about 1500 μm.

[0114] In some embodiments, the MN or MN array includes a plurality of needles. In some embodiments, the MN or MN array includes 2 microneedles / cm 2 Including. In some embodiments, the MN or MN array includes 3 microneedles / cm 2 Including. In some embodiments, the MN or MN array includes 4 microneedles / cm 2 Including. In some embodiments, the MN or MN array includes 5 microneedles / cm 2 Including. In some embodiments, the MN or MN array includes 6 microneedles / cm 2 Including. In some embodiments, the MN or MN array includes 7 microneedles / cm 2 Including. In some embodiments, the MN or MN array includes 8 microneedles / cm 2 Including. In some embodiments, the MN or MN array includes 9 microneedles / cm 2 Including. In some embodiments, the MN or MN array includes 10 microneedles / cm 2 Including. In some embodiments, the MN or MN array includes 11 microneedles / cm 2 Including. In some embodiments, the MN or MN array includes 12 microneedles / cm 2including. In some embodiments, the MN or MN array is 13 microneedles / cm 2 including. In some embodiments, the MN or MN array is 14 microneedles / cm 2 including. In some embodiments, the MN or MN array is 15 microneedles / cm 2 including. In some embodiments, the MN or MN array is 16 microneedles / cm 2 including. In some embodiments, the MN or MN array is 17 microneedles / cm 2 including. In some embodiments, the MN or MN array is 18 microneedles / cm 2 including. In some embodiments, the MN or MN array is 19 microneedles / cm 2 including. In some embodiments, the MN or MN array is 20 microneedles / cm 2 including. In some embodiments, the MN or MN array is 21 microneedles / cm 2 including. In some embodiments, the MN or MN array is 22 microneedles / cm 2 including. In some embodiments, the MN or MN array is 23 microneedles / cm 2 including. In some embodiments, the MN or MN array is 24 microneedles / cm 2 including. In some embodiments, the MN or MN array is 25 microneedles / cm 2 including. In some embodiments, the MN or MN array is 26 microneedles / cm 2 including. In some embodiments, the MN or MN array is 27 microneedles / cm 2 including. In some embodiments, the MN or MN array is 28 microneedles / cm 2 including. In some embodiments, the MN or MN array is 29 microneedles / cm 2 including. In some embodiments, the MN or MN array is 30 microneedles / cm2 This includes. In some embodiments, the MN or MN array has 31 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 35 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 40 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 45 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 50 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 55 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 60 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 65 microneedles / cm². 2 This includes. In some embodiments, the MN or MN array has 70 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 75 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 80 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 85 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 90 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 95 microneedles / cm 2 This includes. In some embodiments, the MN or MN array is 100 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 200 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 300 microneedles / cm². 2This includes. In some embodiments, the MN or MN array has 400 microneedles / cm². 2 This includes. In some embodiments, the MN or MN array is 500 microneedles / cm 2 This includes. In some embodiments, the MN or MN array has 1000 microneedles / cm². 2 Including less than. In some embodiments, the MN or MN array is 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 microneedle may have a circular cross-section. In some embodiments, the microneedle may have a triangular cross-section. In some embodiments, the microneedle may have a rectangular cross-section. In some embodiments, the microneedle may have a square cross-section. In some embodiments, the microneedle may have a quadrilateral cross-section. In some embodiments, the microneedle may have a pentagonal cross-section. In some embodiments, the microneedle may have a hexagonal cross-section. In some embodiments, the microneedle may have a heptagonal cross-section. In some embodiments, the microneedle may have an octagonal cross-section. In some embodiments, the microneedle may have a nonagonal cross-section. In some embodiments, the microneedle may have a decagonal cross-section.

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

[0117] In some embodiments, the MN or MN array includes a needle having multiple microneedle puncture sizes. In some embodiments, the MN or MN array includes a needle having at least two different microneedle puncture sizes. In some embodiments, the MN or MN array includes a needle having at least three different microneedle puncture sizes. In some embodiments, the MN or MN array includes a needle having at least four different microneedle puncture sizes. In some embodiments, the MN or MN array includes a needle having at least five different microneedle puncture sizes. In some embodiments, the MN or MN array includes a needle having up to ten different microneedle puncture sizes. In some embodiments, the MN or MN array includes a needle having at least eleven different microneedle puncture sizes. In some embodiments, the MN or MN array includes a needle having at least twelve different microneedle puncture sizes. In some embodiments, the MN or MN array includes a needle having up to one microneedle puncture size.

[0118] MNs or MN arrays, including various microneedle puncture sizes, may be used in the microneedling techniques described herein. In some embodiments, the MN or MN array is 100 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 200 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 300 μm 2Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 400 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 500 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 600 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 700 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 800 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 900 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 1000 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 1100 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 1200 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 1300 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 1400 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 1500 μm2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 1600 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 1700 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 1800 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 1900 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 2000 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 2500 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 3000 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 3500 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 4000 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 4500 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 5000 μm 2Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 5500 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 6000 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 6500 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 7000 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 7500 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 8000 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 8500 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 9000 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 9500 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 10,000 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 10500 μm 2Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 11000 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 11500 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 12000 μm 2 Includes microneedles having a microneedle puncture size. In some embodiments, the MN or MN array is 13000 μm 2 Includes microneedles having a microneedle puncture size less than / microneedle. In some embodiments, the MN or MN array is 14000 μm 2 Includes microneedles having a puncture size of less than 15,000 μm. In some embodiments, the MN or MN array is 15,000 μm 2 Includes microneedles having a microneedle puncture size less than / microneedle. In some embodiments, the MN or MN array is 20,000 μm 2 Includes microneedles having a microneedle puncture size less than / microneedle. In some embodiments, the MN or MN array is 25,000 μm 2 Includes microneedles having a puncture size of less than a microneedle. In some embodiments, the MN or MN array is 30,000 μm 2 Includes microneedles having a puncture size of less than a microneedle. In some embodiments, the MN or MN array is 35,000 μm 2 Includes microneedles having a puncture size of less than 40,000 μm. In some embodiments, the MN or MN array is 40,000 μm 2Includes microneedles having a puncture size of less than 45,000 μm. In some embodiments, the MN or MN array is 45,000 μm 2 Includes microneedles having a microneedle puncture size less than / microneedle. In some embodiments, the MN or MN array is 50,000 μm 2 Includes microneedles having a microneedle puncture size less than / microneedle. In some embodiments, the MN or MN array is 55000 μm 2 Includes microneedles having a puncture size of less than 60,000 μm. In some embodiments, the MN or MN array is 60,000 μm 2 Includes microneedles with a puncture size smaller than a microneedle.

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

[0120] In some embodiments, the MSC described herein includes one press of the MN or MN array. In some embodiments, the MSC includes two presses of the MN or MN array. In some embodiments, the MSC includes three presses of the MN or MN array. In some embodiments, the MSC includes four presses of the MN or MN array. In some embodiments, the MSC includes five presses of the MN or MN array. In some embodiments, the MSC includes six presses of the MN or MN array. In some embodiments, the MSC includes seven presses of the MN or MN array. In some embodiments, the MSC includes eight presses of the MN or MN array. In some embodiments, the MSC includes nine presses of the MN or MN array. In some embodiments, the MSC includes ten presses of the MN or MN array. In some embodiments, the MSC includes eleven presses of the MN or MN array. In some embodiments, the MSC includes twelve presses of the MN or MN array. In some embodiments, the MSC includes thirteen presses of the MN or MN array. In some embodiments, the MSC includes 14 presses of the MN or MN array. In some embodiments, the MSC includes 15 presses of the MN or MN array. In some embodiments, the MSC includes 16 presses of the MN or MN array. In some embodiments, the MSC includes 17 presses of the MN or MN array. In some embodiments, the MSC includes 18 presses of the MN or MN array. In some embodiments, the MSC includes 19 presses of the MN or MN array. In some embodiments, the MSC includes 20 presses of the MN or MN array. In some embodiments, the MSC includes rolling the MN or MN array once or more on the skin tablet. 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 on the same site. In some embodiments, the presses are performed on overlapping sites.In some embodiments, pressing is performed on different areas. In some embodiments, pressing is performed by stamping the MN array. In some embodiments, pressing is performed by rolling a microneedle roller over the area one or more times. According to established MN practices, in some embodiments, skin pressing of the MN array may last for less than one second, or in some embodiments, for one second or longer, for example, 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 described above, this disclosure teaches that as the total surface area of ​​skin punctured by microneedles decreases, the bioavailability of large drugs in emulsions applied to the skin increases. Therefore, in some embodiments, relatively less pressure 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 containing the drug), less pressure of the microneedle array may be preferred. In some embodiments, shorter microneedle lengths 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 containing the drug), relatively shorter microneedle lengths may be preferred.

[0122] Furthermore, as described 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 a biologically active substance (e.g., a large drug) in combination with MSCs may achieve a larger biological effect. Therefore, in some embodiments, a relatively reduced product volume containing the active substance (e.g., a large drug) may be preferred. In some embodiments, when the large drug administered in combination with microneedle skin conditioning is an emulsion (e.g., a nanoemulsion) or in a topical formulation containing one, 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., a drug-containing emulsion), a relatively smaller product volume may be preferred. MN arrays and MSC devices suitable for use in combination with compositions containing large-capacity drugs for transdermal delivery of large-capacity drugs include, for example, devices described in U.S. Patents 6,334,856; 6,503,231; 6,908,453; 8,257,324; and 9,144,671.

[0123] Large drug In some embodiments, the compositions provided and / or utilized as described herein comprise one or more large-cap drugs. In some embodiments, the large-cap drugs utilized are biologically active substances (e.g., therapeutic agents). In particular, this disclosure provides strategies and remarkable improvements for topical and transdermal delivery of compositions comprising large-cap drugs in combination with MSCs as described herein. Furthermore, this disclosure demonstrates that microneedling treatments may be particularly advantageous for the delivery of large-cap drugs in emulsion compositions (e.g., nanoemulsions).

[0124] 1. Protein drugs Any of the various protein drugs can be incorporated into the provided composition and administered in combination with MSCs. In some embodiments, the protein drug may be a peptide drug. In some embodiments, the peptide has a molecular weight greater than 100 kDa. In some embodiments, the peptide drug has a molecular weight of at least 150 kDa. In some embodiments, the peptide drug consists only of naturally occurring amino acids. In some embodiments, the peptide drug contains one or more amino acids that do not exist naturally.

[0125] Those skilled in the art will be aware of the various protein drugs approved for therapeutic use by the relevant regulatory authorities. For example, the U.S. Food and Drug Administration maintains a list of approved biological therapies 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 topical administration, including, for example, those that may or may have undergone clinical trials (e.g., may be approved or in the process of being approved by the U.S. Food and Drug Administration or an equivalent agency in another jurisdiction, or may be included in clinical trials, e.g., may be listed on www.clinicaltrials.gov, or may be recorded on one or more clinical sites' Institutional Review Boards or equivalents).

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

[0127] The molecular weight of the botulinum toxin protein molecule is approximately 150 kDa for all seven known botulinum toxin serotypes. Botulinum toxin is released by Clostridium bacteria as a complex containing the 150 kDa botulinum toxin protein molecule together with associated non-toxic proteins. Therefore, the BTX-A complex is produced by Clostridium bacteria in 900 kDa, 500 kDa, and 360 kDa forms. Botulinum toxins B and C1 are apparently produced only as a 500 kDa complex. Botulinum toxin D is produced as both 300 kDa and 500 kDa complexes. Finally, botulinum toxins E and F are produced only as a complex of approximately 300 kDa.

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

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

[0130] In some embodiments, botulinum toxin may be selected from the group consisting of types A, Ab, Af, B, Bf, C1, C2, D, E, F, and G; its variants; its fragments; its characteristic parts; and / or fusions thereof. In some embodiments, botulinum toxin may be a variant toxin having one or more structural variations compared to, for example, a reference (e.g., wild-type) toxin (or its associated fragments). In some specific embodiments, the variant toxin may have a longer or shorter biological activity lifetime than a suitable equivalent reference form (e.g., wild-type form). In some embodiments, botulinum toxin may exist as one 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 attribution).

[0131] In some embodiments, the present invention provides botulinum toxin compositions. In some embodiments, the present invention provides nanoemulsion botulinum toxin compositions. A commercially available source of botulinum toxin that can be used according to the present invention is BOTOX (登録商標) DYSPORT (登録商標) (Botulinum toxin type A 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)This includes, but is not limited to, botulinum toxin type B (an injectable solution comprising human serum albumin, sodium succinate, and sodium chloride, with a pH of 5.6, Elan Pharmaceuticals, Dublin, Ireland), NEURONOX (Medytox), HENGLI (Lanzhou Institute), and others. Those skilled in the art will know the standard and / or approved dosing regimens of such commercially available botulinum toxin compositions, understand such compositions in relation to them, and / or regimens may be used in conjunction with the microneedling techniques described herein (e.g., MSC in particular).

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

[0133] In some embodiments, the botulinum toxin composition contains about 2 to about 40,000 units of botulinum toxin per mL. In some embodiments, the botulinum toxin composition contains about 2 to about 12,000 units of botulinum toxin per mL. In some embodiments, the botulinum toxin composition contains about 100 to about 2,000 units of botulinum toxin per mL. In some embodiments, the botulinum toxin composition contains 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 botulinum toxin. Separately or in addition thereto, 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 an absorption 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 an absorption enhancer.

[0135] In some embodiments, the biologically active substances used in combination with the botulinum toxin described herein may be drugs that act on or within the skin and / or exert therapeutic and / or cosmetic effects. For example, in some embodiments, such biologically active substances are selected from therapeutic agents such as anesthetics (e.g., lidocaine), steroids (e.g., hydrocortisone) and / or retinoids (e.g., retinoin), skin fillers (e.g., hyaluronic acid or other elastic materials), collagen and / or silicones. In some embodiments, the botulinum composition is administered in combination with a delivery modifier, such as a penetration enhancer (in some embodiments, non-irritating and / or not decomposing, destroying and / or damaging skin structure and / or skin).

[0136] In some embodiments, the non-irritating penetration enhancer may be selected from, for example, a copolymer, a carrier molecule, and a carrier peptide. In some embodiments, the carrier molecule is positively charged. In some embodiments, the carrier molecule may be a copolymer. In some embodiments, the carrier molecule may be a long-chain positively charged polypeptide or a positively charged non-peptidyl polymer, such as a polyalkylene imine. In some embodiments, the carrier peptide may be a cationic peptide. In some embodiments, the carrier peptide has the sequence RKKRRQRRRG-(K) 15 A positively charged carrier having -GRKKRRQRRR. In some embodiments, the carrier molecule may be one disclosed in U.S. Patent Application Publication 2010 / 0168023 or U.S. Patent Application Publication 2009 / 0247464, which are incorporated herein by reference.

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

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

[0139] (ii) Antibody drugs In some embodiments, this disclosure relates to the delivery of antibody drugs. In some embodiments, the large drug may be an antibody or a fragment or derivative thereof. In particular, this disclosure provides certain compositions comprising antibody drugs and also provides techniques for administering compositions comprising antibody drugs, such administration being combined with MSCs as described herein.

[0140] In some embodiments, antibody drugs may be suitable for treating skin conditions. In some embodiments, antibody drugs may be fusion proteins. In some embodiments, antibody drugs may be conjugated with another part. In some embodiments, antibody drugs may be conjugated with polyethylene glycol. In some embodiments, antibodies may be multispecific (e.g., bispecific) and capable of binding to two or more different target antigens or epitopes.

[0141] In some embodiments, the antibody contains TNFα (e.g., an anti-TNFα antibody, e.g., an epitope-binding element found in infliximab, adalimumab, golimumab, etanercept, etanercept-szzs, and / or certolizumab pegol). In some embodiments, the antibody contains CD2 (e.g., an anti-CD2 antibody, e.g., an epitope-binding element found in ciprizumab). In some embodiments, the antibody contains CD4 (e.g., an anti-CD4 antibody, e.g., an epitope-binding element found in zanorimumab).

[0142] In some embodiments, the antibody contains IL-12 (e.g., an anti-IL-12 antibody, e.g., an epitope-binding element found in briakinumab). In some embodiments, the antibody contains IL-17 (e.g., an anti-IL-17 antibody, e.g., an epitope-binding element found in secukinumab and / or brodalumab). In some embodiments, the antibody contains IL-22 (e.g., an anti-IL-22 antibody, e.g., an epitope-binding element found in fezakinumab). In some embodiments, the antibody contains IL-23 (e.g., 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. Separately or in addition thereto, 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 substance used in combination with the antibody drug described herein may be a drug that acts on or within the skin and / or exerts therapeutic and / or cosmetic effects. For example, in some embodiments, such a biologically active substance may be selected from therapeutic drugs such as anesthetics (e.g., lidocaine), steroids (e.g., hydrocortisone) and / or retinoids (e.g., retinoin), or cosmetic drugs such as skin 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 (in some embodiments, non-irritating and / or not degrading, destroying, and / or damaging skin structure and / or skin).

[0145] In some embodiments, the non-irritating penetration enhancer may be selected from, for example, a copolymer, a carrier molecule, and a carrier peptide. In some embodiments, the carrier molecule is positively charged. In some embodiments, the carrier molecule may be a copolymer. In some embodiments, the carrier molecule may be a long-chain positively charged polypeptide or a positively charged non-peptidyl polymer, such as a polyalkylene imine. In some embodiments, the carrier peptide may be a cationic peptide. In some embodiments, the carrier peptide has the sequence RKKRRQRRRG-(K) 15 A positively charged carrier having -GRKKRRQRRR. In some embodiments, the carrier molecule may be one disclosed in U.S. Patent Application Publication 2010 / 0168023 or U.S. Patent Application Publication 2009 / 0247464, which are incorporated herein by reference.

[0146] 2. Preventive medicine A variety of prophylactic agents may be incorporated into the provided composition and administered in combination with MSCs according to the present invention. In some embodiments, the prophylactic agent includes, but is not limited to, a vaccine. In some embodiments, the vaccine may include isolated proteins or peptides, inactivated organisms and viruses, dead organisms and viruses, genetically modified organisms or viruses, and cell extracts. In some embodiments, the prophylactic agent may be combined with interleukins, interferons, cytokines and adjuvants, such as cholera toxin, alum, and Freund's adjuvant.In some embodiments, the prophylactic agent is Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus, Streptococcus pyogenes, Corynebacterium diphtheriae, Listeria monocytogenes, Bacillus anthrosis, Clostridium tetani, Clostridium botulinum, Clostridium perfringens, Neisseria meningitidis, Neisseria gonore, Streptococcus mutans, Pseudomonas erginosa, Salmonella cifi, Haemophilus parainfluenzae, Bordetella patasis, Francisella tularensis, Yersinia pestis, Vibrio cholerae, Legionella pneumophila, Mycobacterium tubercurosis, Mycobacterium leple, Treponema pallidum, Leptospirosis Antigens of bacterial organisms such as *Borrelia bergdorferi*, *Campylobacter jejuni*, and *Smallpox*, influenza A and B, multinucleated respiratory virus, parainfluenza, measles, HIV, varicella-zoster, herpes simplex I and II, 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 and other viral organisms; antigens of fungi such as Cryptococcus neoformans, Histoplasma capsulatum, Candida albicans, Candida tropicalis, Nocardia asteroides, Rickettsia rickettii, Rickettsia cifi, Mycoplasma pneumoniae, Chlamydia sitassi, Chlamydia trachomatis, Plasmodium falciparum, Trypanosoma bursei, Entamoeba historica, Toxoplasma gondii, Trichomonas vaginalis, Schistosoma mansoni, etc. In some embodiments, these antigens may be in the form of completely dead organisms, peptides, proteins, glycoproteins, carbohydrates, or combinations thereof.

[0147] Those skilled in the art will recognize that the preceding paragraph provides an exemplary, non-exclusive list of drugs that can be delivered using the techniques of the present invention. Any drug can be associated with a composition 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-capacity drugs to subjects requiring them by topical and / or transdermal administration (e.g., by lotions, creams, powders, ointments, liniments, gels, eye drops, etc.). In some embodiments, the provided cream and / or lotion formulations containing large-capacity drugs are administered to subjects requiring them by topical and / or transdermal administration (e.g., by lotions, creams, powders, ointments, liniments, gels, eye drops, etc.).

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

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

[0151] In some embodiments, the cream and / or lotion formulations may be useful for topical and / or transdermal administration. The present invention encompasses the recognition that the cream and / or lotion formulations provided may be particularly useful for the delivery of drugs to the dermis layer of the skin. In some embodiments, the cream and / or lotion formulations provided are formulated for topical and / or transdermal delivery to subjects that require them. In some embodiments, the cream and / or lotion formulations provided are administered to subjects that require them 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, e.g., alkanols having 1 to 8 carbon atoms (e.g., ethanol, isopropanol, benzyl alcohol, and phenylethyl alcohol), polyalcohols, e.g., alkylene glycols (e.g., glycerin, ethylene glycol, and propylene glycol), and glycol ethers, e.g., mono-, di-, and tri-ethylene glycol monoalkyl ethers, e.g., ethylene glycol monomethyl ether and diethylene glycol monomethyl ether (used alone or in mixtures). Such ingredients may be present, for example, in amounts of up to 60%, 70%, 80%, or 90% by weight of the total weight of the composition.

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

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

[0155] In some embodiments, large-cap drugs are formulated into compositions suitable for topical administration. Exemplary large-cap drugs include those described herein. In some embodiments, the provided compositions may be formulated and delivered in combination with MSCs described herein so as to achieve systemic delivery; in some embodiments, the provided compositions may be formulated and / or delivered so as to achieve topical but non-systemic delivery.

[0156] In some embodiments, a composition suitable for topical formulations contains a penetration enhancer. In some embodiments, the penetration enhancer breaks down, destroys, and / or damages the skin structure and / or skin. In some embodiments, the penetration enhancer does not break down, 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] This disclosure specifically demonstrates the effective and efficient delivery of therapeutic agents (particularly large biological drugs, e.g., botulinum toxin and / or antibody drugs) to the dermis using compositions provided in combination with MSCs as described herein. For example, in some embodiments, the present invention provides methods comprising administering the compositions described herein without clinically significant side effects. For example, when local delivery is intended, clinically significant side effects include, but are not limited to, undesirable systemic side effects, damage to nerve tissue beneath the dermis (e.g., nerve paralysis), undesirable effects on muscles (e.g., muscle paralysis), and / or undesirable blood concentrations of the therapeutic agent.

[0158] Those skilled in the art will understand that the composition provided may be incorporated into a device such as a patch. Various transdermal patch structures are known in the art; those skilled in the art will understand that the composition provided may be readily incorporated into any of these various structures. In some embodiments, the transdermal patch may include a plurality of needles extending from one side of the patch 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 includes an adhesive. Several examples of adhesive patches are well known (e.g., U.S. Design Patent No. 296,006; and U.S. Patents No. 6,010,715; No. 5,591,767; No. 5,008,110; No. 5,683,712; No. 5,948,433; and No. 5,965,154; all of which are incorporated herein by attribution). An adhesive patch generally features an adhesive layer applied to the patient's skin, a depot or reservoir for holding the composition to be provided, and an outer surface to prevent the composition to be provided from leaking out of the depot. The outer surface of the patch may be non-adhesive.

[0160] According to the present invention, the provided composition is incorporated into a patch so as to remain stable for an extended period. For example, in some embodiments, the provided composition may be incorporated into a polymer matrix that stabilizes a large drug and allows the drug to diffuse from the matrix and the patch. The provided composition may also be incorporated into an adhesive layer of the patch so as to allow the provided composition to diffuse through the skin when the patch is applied to the skin. In some embodiments, the adhesive layer may be thermally activated so that the drug diffuses through the skin, where a temperature of about 37°C causes the adhesive to slowly liquefy. The adhesive may retain its tackiness when stored below 37°C, and lose its tackiness as it liquefies when applied to the skin.

[0161] In some embodiments, the composition provided may be delivered to a depot within the patch such that pressure applied to the patch directs the composition toward the outside of the patch through the microneedles and stratum corneum. Exemplary embodiments of the microneedles are described above. Devices suitable for intradermal administration of the composition provided include, for example, those described in U.S. Patents 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. The intradermal composition may be administered by devices that limit the effective penetration length of the needles into the skin, such as those described in International Publication No. 99 / 34850 and its functional equivalents.

[0162] In some embodiments, for example, it may be desirable to slow down the absorption of the composition into the skin in order to extend the effect of the composition. In some embodiments, this can be achieved by using a liquid suspension of a crystalline or amorphous material with low water solubility. The absorption rate of the composition then depends on its dissolution rate, which may then depend on the crystal size and crystalline form. In some embodiments, the release rate of the composition can be controlled depending on the ratio of the composition to the polymer and the properties of the particular polymer used. Other examples of biodegradable polymers include poly(orthoester) and poly(anhydrous).

[0163] Emulsion This disclosure includes the recognition that emulsion technology can provide stabilization benefits to drugs of interest, including the large drugs described herein, and in particular botulinum toxin and / or antibody drugs.

[0164] Furthermore, this disclosure understands that certain liquid nanoemulsion technologies have been shown to provide superior transdermal delivery properties even for extremely large molecules such as Clostridium botulinum and / or antibody drugs. See, for example, U.S. Patent Application Publication 2012 / 0328701, U.S. Patent Application Publication 2012 / 0328702, No. 8,318,181, and U.S. Patent No. 8,658,391, which are incorporated herein by full attribution. These liquid nanoemulsions are far superior to solid nanoparticle drug delivery, particularly transdermal drug delivery, as stated by Gomaa, solid nanoparticles cannot penetrate the skin and only accumulate in hair follicles. These liquid nanoemulsions are stable for at least 34 months and are commercially viable from this perspective as well.

[0165] 1. Macroemulsion In some embodiments, the Disclosure provides strategies for using microneedling to “condition” skin to which a transdermal product has been, is being, or will be applied. Despite previous reports suggesting that such strategies are likely only useful for drugs with small molecular weights, given that studies analyzing the transdermal delivery of small molecules (specifically, 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 penetration decreases as molecular weight increases,” the Disclosure provides the finding that such microneedle conditioning can, surprisingly, offer significant benefits in improving the transdermal delivery of large drugs (e.g., those with molecular weights of approximately 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] This disclosure provides the finding that the effective and rapid (i.e., administration over several minutes) transdermal delivery of macromolecules by such liquid macroemulsion compositions can be surprisingly improved by combining the microneedle skin conditioning (MSC) described herein (e.g., using a relatively low microneedle density and / or a relatively small microneedle puncture size) with macroemulsion administration.

[0167] This disclosure particularly demonstrates that microneedling technology (e.g., microneedle conditioning of the skin) can significantly improve the transdermal delivery of large drugs (e.g., botulinum toxin and / or antibody drugs), especially when used in combination with macroemulsion technology. Specific purpose macroemulsion compositions include water-in-oil and oil-in-water macroemulsions characterized by droplet sizes ranging from about 300 nm to about 5,000 μm 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. The surfactant portion of the composition may comprise one or more surfactants. [Table 2]

[0168] Macroemulsion formulations may act to stabilize large-cap drugs, such as Clostridium botulinum and / or antibody drugs. While macroemulsion formulations are not necessarily expected to achieve transdermal delivery of large-cap drugs on their own, this disclosure nevertheless encompasses the finding that the improved stabilization that can be provided by incorporation into a macroemulsion composition, when combined with the microneedle technology described herein, can achieve a synergistic improvement in transdermal delivery.

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

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

[0171] The findings presented herein are particularly surprising given reports that transdermal delivery of solid nanoparticles, much smaller in size than droplets in the macroemulsion compositions used 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 evaluating skin penetration by applying a solution of rhodamine dye (molecular weight 479 Da) encapsulated in PLGA nanoparticles to skin preconditioned with microneedles. 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 small amounts of 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, “There is a new consensus that NPs [nanoparticles] can be sufficiently deposited in hair follicles but cannot normally penetrate the stratum corneum.” Therefore, prior to this disclosure, those skilled in the art would expect that the use of microneedling techniques in vehicles significantly larger than 105 nm would not be able to effectively deliver even small molecule drugs (e.g., rhodamine dye) transdermally; indeed, the delivery of large drugs would have been considered impossible. However, this disclosure demonstrates that microneedling can significantly improve the transdermal delivery of large drugs, particularly when used in combination with macroemulsion techniques.

[0172] In particular, this disclosure reveals that microneedling technology can improve transdermal delivery (e.g., of large drugs, especially from macroemulsion compositions) when other disruptive agents (i.e., chemopreservatives and other techniques for disrupting or puncturing skin structures) are not utilized. Previous studies of transdermal delivery of drugs the same size as 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 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 microneedle site.

[0173] This disclosure, in particular, shows that when microneedle coating or loading is not utilized and / or when the microneedles are not designed to remain in the skin, microneedling techniques can achieve transdermal delivery (e.g., of large drugs, particularly from macroemulsion and nanoemulsion compositions). In particular, as already stated, this disclosure understands that such microneedle coating or loading may not be commercially viable because the botulinum coating or loading material is unstable. For example, according to Johnson, E. et al., "Botulinum toxin is extremely susceptible to denaturation due to surface denaturation, heat and alkaline conditions. Freeze-drying of botulinum toxin is the most economically sound and practical method to distribute a product in a stable form that is readily usable by clinicians." U.S. Patent No. 5,512,547. Also, as will be understood by those skilled in the art reading this specification, the techniques described herein have certain advantages, including the absence of microneedles that remain in or bind to tissue. For example, those skilled in the art will understand that leaving microneedles in the skin carries the risk of skin irritation, inflammation, allergic reactions, and / or cosmetically undesirable scarring. In contrast to the present invention, techniques such as those described in U.S. Patent Application Publication No. 2017 / 0209553 utilize a microneedle array designed to be loaded with botulinum toxin and to penetrate the skin (U.S. Patents No. 2017 / 0209553 and 2016 / 0263362).

[0174] This disclosure surprisingly provides an effective technique for the transdermal delivery of large drugs (e.g., botulinum toxin, antibodies, etc.). In particular, this disclosure teaches that the transdermal delivery of such drugs can be significantly improved by the use of microneedling techniques without other disruption strategies. Thus, the techniques provided can achieve effective delivery without the inflammation, irritation, and / or allergic reactions often associated with the use of skin disruptors. As will be understood by those skilled in the art reading this specification, this disclosure teaches that the transdermal delivery of such large drugs can be significantly improved by the use of microneedling techniques even when they are not loaded into microneedles, not coated on them, and / or manufactured as part thereof. Similarly, as will be understood by those skilled in the art reading this specification, this disclosure teaches that the delivery of the large drugs described herein can be significantly improved by the use of microneedling techniques (particularly the use of MSCs) without leaving microneedles in the skin (e.g., by disrupting them and / or by otherwise maintaining and / or degrading them in situ). For example, those skilled in the art will understand that the technology provided can avoid the challenges of long-term stability of large drugs required for commercially viable products and can achieve effective delivery without inflammation, irritation, and / or allergic reactions that may occur due to skin-disrupting agents and / or microneedles remaining in the skin. Indeed, in the examples and elsewhere, this disclosure explicitly teaches that MSCs performed with botulinum toxin-free microneedles facilitate transdermal delivery of botulinum toxin from topical (e.g., creams, ointments) compositions, particularly from compositions containing macro- or nano-emulsions.

[0175] In some embodiments, the disclosure teaches that particularly advantageous results are achieved when microneedling technology is combined with macroemulsion compositions. In some embodiments, the microneedling technology is combined with a lotion, cream, or liquid composition, which in turn may be or include a macroemulsion composition. In some embodiments, the technology provided does not utilize skin disruption technology, such as chemopreservatives.

[0176] In some embodiments, the present invention utilizes macroemulsion compositions containing large-cap drugs that are particularly effective and / or useful in medical settings, for example, for therapeutic purposes. In some embodiments, certain macroemulsion compositions are particularly effective and / or useful for topical administration of drugs to subjects that require them. In some embodiments, the macroemulsion composition may contain one or more large-cap drugs.

[0177] In some embodiments, the macroemulsion can be formulated into a composition suitable for topical administration to the skin. In some embodiments, the composition suitable for topical administration may 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.

[0179] In some embodiments, the provided composition comprises a mixture of the provided macroemulsion composition and one or more pharmaceutically acceptable additives. In some embodiments, the cream and / or lotion formulation comprises a mixture of the provided macroemulsion composition and / or a saline solution.

[0180] In some embodiments, the provided composition comprises a macroemulsion composition containing one or more large drugs. In some embodiments, the provided composition is a cream and / or lotion formulation. In some embodiments, the provided cream and / or lotion formulation comprises a macroemulsion composition. In some embodiments, the composition comprises a provided macroemulsion composition but is not a cream and / or lotion formulation. In some embodiments, a suitable composition is formulated into a cream and / or lotion but does not contain a macroemulsion composition.

[0181] In some embodiments, the provided composition comprises a provided macroemulsion composition and a mixture of one or more pharmaceutically acceptable additives for, for example, topical and / or transdermal administration (e.g., by lotion, cream, powder, ointment, liniment, gel, eye drops, etc.).

[0182] 2. Nanoemulsion In some embodiments, the Disclosure provides strategies for using microneedling to “condition” skin to which a transdermal product has been, is being, or will be applied. Despite previous reports suggesting that such strategies are likely only useful for drugs with small molecular weights, given that studies analyzing the transdermal delivery of small molecules (specifically, 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 penetration decreases as molecular weight increases,” the Disclosure provides the finding that such microneedle conditioning can, surprisingly, offer significant benefits in improving the transdermal delivery of large drugs (e.g., those with molecular weights of approximately 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] This disclosure provides insights showing that the effective and rapid (i.e., administration over several minutes) transdermal delivery of macromolecules by such liquid nanoemulsion compositions can be improved by combining microneedle skin conditioning (MSC) and nanoemulsion administration as described herein.

[0184] This disclosure particularly demonstrates that microneedling techniques (e.g., microneedle conditioning of the skin) can significantly improve the transdermal delivery of large drugs (e.g., botulinum toxin and / or antibody drugs), especially when used in combination with nanoemulsion techniques. Specific purpose nanoemulsion compositions include water-in-oil and oil-in-water nanoemulsions characterized by droplet sizes ranging from about 1 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.

[0185] In some embodiments, the provided nanoemulsion composition contains oil and surfactant in a ratio ranging from about 0.1:1 to about 2:1. In some embodiments, the provided nanoemulsion composition contains oil and surfactant in a ratio ranging from about 0.1:1 to about 1:1. In some embodiments, the provided nanoemulsion composition contains oil and surfactant in a ratio ranging from about 0.5:1 to about 1:1. In some embodiments, the provided nanoemulsion composition contains oil and surfactant in a ratio ranging from about 0.5:1 to about 1:1.5. In some embodiments, the provided nanoemulsion compositions contain 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, the provided nanoemulsion compositions contain oil and surfactant in a ratio of about 0.67:1.

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

[0187] In some embodiments, droplets in the nanoemulsion composition have diameters (e.g., average diameter and / or median diameter) in 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, droplets in the nanoemulsion composition have diameters (e.g., average diameter and / or median diameter) in 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, droplets in the nanoemulsion composition have diameters of 1 nm to 15 nm, 15 nm to 200 nm, 25 nm to 200 nm, 50 nm to 200 nm, or 75 nm to 200 nm (e.g., average diameter and / or median diameter).

[0188] In some embodiments, the total droplet distribution is contained within a specified range of droplet diameter sizes. In some embodiments, less than 50%, 25%, 10%, 5%, or 1% of the total droplet distribution is outside the specified range of droplet diameter sizes. In some embodiments, less than 1% of the total droplet distribution is outside the specified range of droplet diameter sizes. In some embodiments, the nanoemulsion composition substantially does not contain 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 total 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, 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 in the range of about 10 nm and about 300 nm, about 50 nm and about 250 nm, 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 in the range of -80mV to +80mV. In some embodiments, the nanoemulsion droplets have a zeta potential in the range of -50mV to +50mV. In some embodiments, the nanoemulsion droplets have a zeta potential in the range of -25mV to +25mV. In some embodiments, the nanoemulsion droplets have a zeta potential in the range of -10mV to +10mV. In some embodiments, the nanoemulsion droplets have a zeta potential of about -80mV, about -70mV, about -60mV, about -50mV, about -40mV, about -30mV, about -25mV, about -20mV, about -15mV, about -10mV, or about -5mV. In some embodiments, the nanoemulsion droplets have a zeta potential of approximately +50mV, approximately +40mV, approximately +30mV, approximately +25mV, approximately +20mV, approximately +15mV, approximately +10mV, or approximately +5mV. In some embodiments, the nanoemulsion droplets have a zeta potential of approximately 0mV.

[0191] This disclosure surprisingly provides an effective technique for the transdermal delivery of large drugs (e.g., botulinum toxin, antibodies, etc.). In particular, this disclosure teaches that the transdermal delivery of such drugs can be significantly improved by the use of microneedling techniques without other disruption strategies. Thus, the techniques provided can achieve effective delivery without the inflammation, irritation, and / or allergic reactions often associated with the use of skin disruptors. As will be understood by those skilled in the art reading this specification, this disclosure teaches that the transdermal delivery of such large drugs can be significantly improved by the use of microneedling techniques even when they are not loaded into microneedles, not coated on them, and / or manufactured as part thereof. Similarly, as will be understood by those skilled in the art reading this specification, this disclosure teaches that the delivery of the large drugs described herein can be significantly improved by the use of microneedling techniques (particularly the use of MSCs) without leaving microneedles in the skin (e.g., by disrupting them and / or by otherwise maintaining and / or degrading them in situ). For example, those skilled in the art will understand that the technology provided can avoid the challenges of long-term stability of large drugs required for commercially viable products and can achieve effective delivery without inflammation, irritation, and / or allergic reactions that may occur due to skin-disrupting agents and / or microneedles remaining in the skin. Indeed, in the examples and elsewhere, this disclosure explicitly teaches that MSCs performed with botulinum toxin-free microneedles facilitate transdermal delivery of botulinum toxin from topical (e.g., creams, ointments) compositions, particularly from compositions including macro- and nano-emulsions.

[0192] In some embodiments, the disclosure teaches that particularly advantageous results are achieved when microneedling technology is combined with nanoemulsion compositions. In some embodiments, the microneedling technology is combined with a lotion, cream, or liquid composition, which in turn may be or include a nanoemulsion composition. In some embodiments, the technology provided does not utilize skin disruption technology, such as chemopenetration enhancers.

[0193] The findings presented herein are particularly surprising given reports that transdermal delivery of solid nanoparticles of a size comparable to that of droplets in the nanoemulsion compositions used 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 to evaluate skin permeation. 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 small amounts of dye began to penetrate 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, “There is a new consensus that NPs [nanoparticles] can be sufficiently deposited in hair follicles but cannot normally penetrate the stratum corneum.” Therefore, prior to this disclosure, those skilled in the art would expect that the use of microneedling techniques in nano-sized vehicles would not be able to effectively deliver even small molecule drugs (e.g., rhodamine dye) transdermally; indeed, the delivery of large-molecule drugs would have been considered impossible. However, this disclosure demonstrates that microneedling can significantly improve the transdermal delivery of large-molecule drugs, particularly when used in conjunction with nanoemulsion techniques.

[0194] In particular, this disclosure reveals that microneedling technology can improve transdermal delivery (e.g., of large drugs, especially from nanoemulsion compositions) when other disruptive agents (i.e., chemopreservatives and other techniques for disrupting or puncturing skin structures) are not utilized. Previous studies of transdermal delivery of drugs the same size as 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 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 microneedling site.

[0195] This disclosure, in particular, shows that when microneedle coating or loading is not utilized and / or when the microneedles are not designed to remain in the skin, microneedling techniques can achieve transdermal delivery (e.g., of large drugs, particularly from macroemulsion and nanoemulsion compositions). In particular, as already stated, this disclosure understands that such microneedle coating or loading may not be commercially viable because the botulinum coating or loading material is unstable. For example, according to Johnson, E. et al., "Botulinum toxin is extremely susceptible to denaturation due to surface denaturation, heat and alkaline conditions. Freeze-drying of botulinum toxin is the most economically sound and practical method to distribute a product in a stable form that is readily usable by clinicians." U.S. Patent No. 5,512,547. Also, as will be understood by those skilled in the art reading this specification, the techniques described herein have certain advantages, including the absence of microneedles that remain in or bind to tissue. For example, those skilled in the art will understand that leaving microneedles in the skin carries the risk of skin irritation, inflammation, allergic reactions, and / or cosmetically undesirable scarring. In contrast to the present invention, techniques such as those described in U.S. Patent Application Publication No. 2017 / 0209553 utilize a microneedle array designed to be loaded with botulinum toxin and to penetrate the skin (U.S. Patents No. 2017 / 0209553 and 2016 / 0263362).

[0196] This disclosure teaches that, despite the expectation that MSCs are only useful for facilitating transdermal delivery of small compounds, the already highly efficient transdermal delivery of large drugs by nanoemulsion compositions can be dramatically improved by combining them with microneedle technology. Furthermore, this disclosure is particularly remarkable considering that microneedle conditioning combined with encapsulation of small molecule drugs in solid nanoparticles provides only a small amount of penetration 6 hours after administration, as described by Gomaa, and material penetration was observed up to 12 hours after administration.

[0197] In some embodiments, a nanoemulsion formulation containing a large drug may be administered in combination with microneedle conditioning using solid microneedles. In some embodiments, site-specific site conditioning (MSC) is performed before application of the nanoemulsion formulation containing the large drug to the site (e.g., before a particular application and / or before each application). In some embodiments, site-specific MSC is performed after application of the nanoemulsion formulation containing the large drug to the site. In some embodiments, site-specific MSC and application of the nanoemulsion formulation containing the large drug to the site occur substantially simultaneously. In some embodiments, the macroemulsion formulation containing the 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 in length. In some embodiments, the microneedle may be about 1 μm to about 2,000 μm in length. In some embodiments, the microneedle may be about 50 μm to about 400 μm in length. In some embodiments, the microneedles may be about 800 μm to about 1500 μm in length.

[0198] In some embodiments, the present invention utilizes nanoemulsion compositions containing large-cap 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 topical administration of drugs to subjects requiring them. In some embodiments, the nanoemulsion composition may contain one or more large-cap drugs. Exemplary nanoemulsion compositions and methods for producing them are described, for example, in International Publication No. 2012 / 103035, which is incorporated herein by attribution.

[0199] In some embodiments, the nanoemulsion can be formulated into a composition suitable for topical administration. In some embodiments, the composition suitable for topical administration may be a lotion, cream, powder, ointment, liniment, gel, or eye drop.

[0200] In some embodiments, the nanoemulsion formulation comprises water, medium-chain triglycerides, polysorbate 80, methylparaben, and propylparaben. Exemplary premixes, not intended 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 subjects in need by topical and / or transdermal administration (e.g., by lotions, creams, powders, ointments, liniments, gels, eye drops, etc.). In some embodiments, the provided cream and / or lotion formulations can be administered to subjects in need by topical and / or transdermal administration (e.g., by lotions, creams, powders, ointments, liniments, gels, eye drops, etc.). In some embodiments, the provided nanoemulsion composition can be formulated into a cream and / or lotion formulation. In some embodiments, the provided cream and / or lotion formulation, comprising the nanoemulsion composition, may be useful and / or effective for topical administration to a subject. In some embodiments, the provided nanoemulsion composition can be mixed with a saline solution for the manufacture of one or more cream components and / or pharmaceutical compositions in a cream formulation (e.g., the provided cream formulation).

[0202] The present invention may include the recognition that an emulsion composition (e.g., a macroemulsion composition and a nanoemulsion composition) can be formulated into a cream and / or lotion formulation for administration to a subject. The present invention may include the recognition that the provided cream and / or lotion formulation can be particularly useful for formulating an emulsion, 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, the provided composition comprises a mixture of the provided nanoemulsion composition and one or more pharmaceutically acceptable additives. In some embodiments, the cream and / or lotion formulation comprises a mixture of the provided nanoemulsion composition and / or an aqueous saline solution.

[0204] In some embodiments, the provided composition comprises the provided nanoemulsion composition. In some embodiments, the provided composition is a cream and / or lotion formulation. In some embodiments, the provided cream and / or lotion formulation comprises a nanoemulsion composition. In some embodiments, the composition comprises the provided nanoemulsion composition but is not a cream and / or lotion formulation. In some embodiments, a suitable composition is formulated into a cream and / or lotion and comprises a nanoemulsion composition.

[0205] In some embodiments, the provided composition comprises the provided nanoemulsion composition, and a mixture of one or more pharmaceutically acceptable additives for, e.g., topical and / or transdermal (e.g., by lotion, cream, powder, ointment, liniment, gel, eye drops, etc.) administration.

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

[0207] In some embodiments, the provided composition may be formulated and delivered in combination with MSCs so as to achieve systemic delivery; in some embodiments, the provided composition may be formulated and / or delivered so as to achieve topical but non-systemic delivery.

[0208] This disclosure specifically demonstrates the effective and efficient delivery of therapeutic agents (particularly large biological drugs, e.g., botulinum toxin or antibody drugs) to the dermis using compositions provided in combination with MSCs. For example, in some embodiments, the present invention provides methods comprising administering the compositions described herein without clinically significant side effects. For example, when local delivery is intended, clinically significant side effects include, but are not limited to, undesirable systemic side effects, damage to nerve tissue beneath the dermis (e.g., nerve paralysis), undesirable effects on muscles (e.g., muscle paralysis), and / or undesirable blood concentrations of the therapeutic agent. Exemplary formulations of botulinum nanoemulsion premixes are provided in Table 4, not intended to limit the present invention. [Table 5]

[0209] Diseases, disorders and conditions The present invention provides techniques for treating and / or preventing a variety of systemic or cutaneous diseases, disorders, or conditions. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders, or conditions related to the activity of sweat and / or sebaceous glands. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders, or conditions related to infection. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders, or conditions related to inflammation. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders, or conditions related to inflammation. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders, or conditions related to cancer. In some embodiments, the present invention provides techniques for treating and / or preventing systemic diseases, disorders, or conditions. In some embodiments, the present invention provides techniques for treating and / or preventing autoimmune diseases, disorders, or conditions. In some embodiments, the present invention provides techniques for treating and / or preventing diseases, disorders, or conditions related to the epithelium and / or dermal level of the skin.

[0210] In some embodiments, the present invention addresses acne, undesirable sweating, body odor, hyperhidrosis, bromhidrosis, chromohidrosis, rosacea, alopecia, psoriasis, actinic keratosis, eczematous dermatitis (e.g., atopic dermatitis), excessive sebum production disorders (e.g., seborrhea, seborrheic dermatitis), burns, Raynaud's phenomenon, lupus erythematosus, hyperpigmentation disorders (e.g., melasma), hypopigmentation disorders (e.g., maculosis), skin cancer (e.g., squamous cell carcinoma, basal cell carcinoma), skin infections (e.g., bacterial infections, viral infections, fungal infections), facial wrinkles (e.g., forehead, frown lines, rhythmia and / or periorbital area). The present invention provides techniques for treating and / or preventing one or more of the following conditions: wrinkles, headaches, unsightly facial expressions (e.g., due to overactivity of the underlying facial muscle tissue), 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, and various forms of facial contracture), dystonia, benign prostatic hyperplasia, headaches, strabismus, hemifacial spasm, tremors, spasticity (e.g., those resulting from multiple sclerosis), posterior orbital muscles, various ophthalmic and urological conditions (e.g., penile and / or bladder dysfunction), 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 erythematous. In some embodiments, lupus erythematous 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 lung disorders. In some embodiments, the lung disorder may be asthma or chronic obstructive pulmonary disease.

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

[0218] In some embodiments, the present invention provides techniques for treating and / or preventing infections. In 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 related to 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 neurological conditions. In some embodiments, the neurological conditions are Alzheimer's disease, Parkinson's disease, or stroke.

[0221] In some embodiments, the present invention comprises administering at least one of the provided compositions in combination with MSCs according to a dosage regime sufficient to achieve at least about 20% reduction in the degree and / or incidence of the associated skin condition; in some embodiments, according to a dosage regime sufficient to achieve at least about 25% reduction; in some embodiments, according to a dosage regime sufficient to achieve at least about 30% reduction; in some embodiments, according to 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%, approximately 43%, approximately 44%, approximately 45%, approximately 46%, approximately 47%, approximately 48%, approximately 49%, approximately 50%, approximately 51%, approximately 52%, approximately 53%, approximately 54%, approximately 55%, approximately 56%, approximately 57%, approximately 58%, approximately 59%, approximately 60%, approximately 61%, approximately 62%, approximately 63%, approximately 64%, approximately 65%, approximately 66%, approximately 67%, approximately 68%, approximately 69%, approximately It is administered according to a dosing regimen sufficient to achieve a reduction of 70%, approximately 71%, approximately 72%, approximately 73%, approximately 74%, approximately 75%, approximately 76%, approximately 77%, approximately 78%, approximately 79%, approximately 80%, approximately 81%, approximately 82%, approximately 83%, approximately 84%, approximately 85%, approximately 86%, approximately 87%, approximately 88%, approximately 89%, approximately 90%, or more.

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

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

[0224] In some embodiments, such provided compositions are administered topically to the affected area (such as the axilla, hands, feet, scalp, hair follicles, face, neck, back, arms, chest, legs, groin, intertriginous areas, etc., depending on the specific condition being treated) in combination with MSCs. In some embodiments, topical administration is achieved by topical 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 compositions provided may be formulated for topical and / or transdermal delivery (e.g., as lotions, creams, liniments, ointments, powders, gels, eye drops, etc.). In some embodiments, the compositions provided may be or comprise nanoemulsions. In some embodiments, the compositions provided may be or comprise macroemulsions.

[0226] Formulations of the provided composition may be manufactured by any suitable method known or to be developed in the field of pharmacology, for example. Generally, such manufacturing methods include the steps of mixing the provided composition with one or more additives, and then, if necessary and / or desired, forming and / or packaging it into a form suitable for administration, such as single or multi-dose units.

[0227] In some embodiments, the composition may be manufactured, packaged and / or sold in large quantities as one single unit dose and / or more single unit doses. As used herein, “unit dose” refers to an individual amount of a pharmaceutical composition comprising a predetermined amount of the provided composition. The amount of the provided composition is generally equal to the dose of the provided composition administered to a subject, and / or a convenient portion of such dose, for example, half or one-third of such dose.

[0228] In some embodiments, suitable additives for use in a composition (e.g., a pharmaceutically 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, isotonic agents, thickeners or emulsifiers, preservatives, solid binders, lubricants, disintegrants, binders, preservatives, buffers, etc., to suit a desired specific dosage form. In some embodiments, additives such as cocoa butter and / or suppository waxes, colorants, coatings, 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 attribution) discloses various additives used in known techniques for the formulation and manufacture of pharmaceutical compositions.

[0229] In some embodiments, suitable additives (e.g., pharmaceutically and / or cosmetically acceptable additives) have a purity of at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%. 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 Pharmacopeia (USP), European Pharmacopoeia (EP), British Pharmacopoeia, and / or other international pharmacopoeias.

[0230] In some embodiments, the compositions provided are formulated as creams, liniments, ointments, oils, foams, sprays, lotions, liquids, powders, thickening lotions, or gels (for example, formulated for transdermal delivery as described herein). Certain exemplary such formulations may be manufactured, for example, as cosmetic formulation products, e.g., emollients, nourishing lotion-type emulsions, cleansing lotions, cleansing creams, skin milks, emollient lotions, massage creams, emollient creams, makeup bases, facial packs, or facial gels; cleansing formulations, e.g., shampoos, conditioners, body cleansers, hair tonics, or soaps; or skin compositions, e.g., lotions, ointments, gels, creams, liniments, patches, deolants, or sprays. In some embodiments, compositions for topical administration are not formulated for administration to mucous membranes (for example, unsuitable for application to mucous membranes and / or not formulated to deliver a suitable amount of large drug to or across mucous membranes).

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

[0232] Any site suitable for MSC is a suitable administration site. In some embodiments, the administration site is the skin covering the muscles or muscle groups 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 in the groin. In some embodiments, the site is in the inguinal region. 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 mucous membrane.

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

[0234] Administration The present invention provides a technology for delivering emulsion compositions (e.g., botulinum emulsion compositions or antibody drug emulsion compositions) for various applications, including cosmetics, nutritional supplements, and medical applications. Such emulsion compositions may contain one or more biologically active substances. In some embodiments, the emulsion composition contains 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 compositions provided herein (e.g., the provided emulsion composition; the provided cream and / or lotion formulation; a combination of the provided emulsion composition and the provided cream and / or lotion formulation) in combination with MSCs.

[0236] In some embodiments, such methods include administering a provided composition in combination with MSCs to patients suffering from and / or susceptible to a disease, condition, or disorder. In some embodiments, such methods include administering a provided nanoemulsion composition in combination with MSCs described herein (e.g., using microneedles having a relatively low microneedle density and / or a relatively small microneedle puncture size) to patients suffering from and / or susceptible to a disease, condition, or disorder related to 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 independently active biologically active substance in combination with MSCs to patients 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 independently active biologically active substance, formulated with a provided cream and / or lotion formulation, in combination with MSCs to patients suffering from and / or susceptible to a disease, condition, or disorder. In some embodiments, such methods involve 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 the administration of a penetration enhancer. Some embodiments further include the 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 demonstrates that certain compositions described herein, combined 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 within the skin, cells, etc.). In some embodiments, the present invention demonstrates 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 related to epithelial and / or dermal structures (e.g., sweat glands, sebaceous glands, hair follicles, etc.). In some embodiments, the present invention shows that the compositions provided herein (e.g., the provided nanoemulsion compositions; cream and / or lotion formulations; combinations of the provided nanoemulsion compositions and cream and / or lotion formulations, etc.), combined with the MSCs described herein (e.g., using microneedles having a relatively low microneedle density and / or a relatively small microneedle puncture size), can improve efficient and specific delivery of active substances to the dermis and / or bioavailability, and that the compositions provided herein can have a therapeutic effect when administered to the skin of a subject. In some embodiments, the present invention shows improved delivery and / or bioavailability and / or achievement of therapeutic effects through the dermis without significant side effects associated with delivery to other areas (e.g., subdermal or extradermal structures and / or non-dermal tissues). In some embodiments, the compositions provided herein (e.g., the emulsion compositions provided; cream and / or lotion formulations; combinations of the emulsion compositions provided and cream and / or lotion formulations) in combination with the MSCs described herein can improve the transdermal delivery and / or bioavailability of active substances, 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 composition provided herein (e.g., a provided emulsion composition; a provided cream and / or lotion formulation; a combination of a provided emulsion composition and a provided cream and / or lotion formulation) in combination with a microneedle culture medium (MSC) provided 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 a composition comprising a provided emulsion composition in combination with an MSC provided herein.

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

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

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

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

[0244] kit In some embodiments, the present invention provides a pharmaceutical package or kit comprising one or more emulsion compositions and one or more microneedle devices according to the present invention. In some embodiments, the pharmaceutical package or kit comprises a formulation or pharmaceutical composition comprising a composition provided in one or more containers which are optionally filled with one or more further components of the pharmaceutical composition. In some embodiments, the pharmaceutical package or kit comprises further approved therapeutic agents for use in combination therapy (e.g., benzoyl peroxide for the treatment of acne; aluminum compounds for the treatment of hyperhidrosis, etc.). In some embodiments, optionally associated with such containers may be a notice in a form prescribed by a government agency regulating the manufacture, use or sale of a medicinal product, the notice reflecting approval by the agency for manufacture, use or sale for human administration.

[0245] In some embodiments, a kit containing a therapeutic reagent is provided. In one non-limiting example, the composition provided may be provided as a topical formulation and administered as a therapy in combination with the use of a microneedling device. Pharmaceutical dosages or instructions for self-administration thereof may be provided in a kit for administration to an individual suffering from or at risk of a condition or disorder, such as those related to the dermal level of the skin.

[0246] In some embodiments, the kit may include (i) a composition provided; (ii) at least one pharmaceutically acceptable additive; and (iii) at least one device for microneedling of the skin; and (iv) instructions for use. In some embodiments, at least one device may have a relatively low microneedle density (e.g., about 2 microneedles / cm²). 2 ~Approximately 50 microneedles / cm 2 The device may include microneedles having a range of . 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 ~ approximately 30,000 μm 2 The range of the microneedle, the puncture size per microneedle is approximately 100 μm. 2 / Microneedle ~ approximately 60,000 μm 2 This may include the range of microneedles.

[0247] The present invention provides, in particular, techniques for administering large-cap drugs, such as botulinum toxin or antibody drugs, improving transdermal delivery, and / or improving the bioavailability of such large-cap drugs, by incorporating one or more large-cap drugs into one or more emulsion compositions, which are subsequently 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 are dramatically improved when microneedles or microneedle arrays having a relatively low microneedle density or a relatively small microneedle puncture size (e.g., puncture size per microneedle, cross-sectional area of ​​each microneedle) are used in combination with MSCs. The advantage of the present invention is the ability to administer such large-cap drugs intradermally with minimal skin irritation or damage. The use of other drugs or processes with emulsion compositions and MSCs is not necessarily excluded and is not required in all embodiments of the present invention.

[0248] Accordingly, the present invention provides a technique for administering large-capacity drugs by topical application of superior emulsion compositions (e.g., macroemulsion compositions and / or nanoemulsion compositions) in combination with MSCs. In some embodiments, the large-capacity drug is botulinum toxin. In some embodiments, the botulinum emulsion composition is applied directly to the skin and through the epidermis before the MSC for absorption. In some embodiments, the botulinum emulsion composition is applied directly to the skin and through the epidermis after the MSC for absorption. In some embodiments, the botulinum emulsion composition is applied substantially simultaneously with the MSC for absorption directly to the skin and through the epidermis.

[0249] In some embodiments, botulinum emulsion compositions combined with MSCs can penetrate the uppermost layers of the skin, including the stratum corneum, pores, and / or glands, without the use of penetration enhancers.

[0250] In some embodiments, antibody drug emulsion compositions combined with MSCs can penetrate the stratum corneum, dermal pores, and / or dermal glands, the uppermost layers of the skin, without the use of penetration enhancers. In some embodiments, the large drug is an antibody drug. In some embodiments, the antibody drug emulsion composition is applied before MSCs for direct skin and absorption through the epidermal layer. In some embodiments, the antibody drug emulsion composition is applied after MSCs for direct skin and absorption through the epidermal layer. In some embodiments, the antibody drug emulsion composition is applied substantially simultaneously with MSCs for direct skin and absorption through the epidermal layer. In some embodiments, the antibody drug emulsion composition is applied directly to the skin and for systemic absorption.

[0251] In some embodiments, antibody drug emulsion compositions can penetrate the uppermost layers of the skin, including the stratum corneum, dermal pores, and / or dermal glands, without the use of degrading agents, irritants, and / or abrasives.

[0252] It will be understood by those skilled in the art that the compositions of the present invention for topical administration may include cosmetic formulation products, such as emollients, nourishing lotion-type emulsions, cleansing lotions, cleansing creams, skin milks, emollient lotions, massage creams, emollient creams, makeup bases, facial packs or facial gels, cleansing formulations, such as shampoos, conditioners, body cleansers, hair tonics or soaps, or skin compositions, such as lotions, ointments, gels, creams, patches or sprays. In some embodiments, the compositions for topical administration are not formulated for administration to mucous membranes (e.g., unsuitable for application to mucous membranes and / or not formulated to deliver an appropriate amount of large drug to or across mucous membranes).

[0253] Those skilled in the art will understand that the units herein relate 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 botulinum toxin administered according to the present invention may last as long as the effect of the injected solution persists. In some embodiments, the effect of the injected solution may last for up to about 6-7 months. In some embodiments, the therapeutic effect of botulinum toxin administered according to the present invention may last for up to 6-7 months. In some embodiments, the use of a synthetic polymer carrier that can retain and slowly release 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 topical formulations of botulinum toxin that avoid potential complications including, but not limited to, systemic toxicity or botulism. In some embodiments, the dosage of botulinum toxin (including types A, B, C, D, E, F, or G, or botulinum genetically engineered or chemically modified to have a longer or shorter duration of action than botulinum toxin serotype A) may range from a minimum of about 1 unit to a maximum of about 50,000 units while minimizing the risk of adverse side effects. The specific dosage may vary depending on the condition being treated and the treatment regimen being used. For example, treatment of subcutaneous hyperactive muscle may require a high transdermal dose of botulinum toxin (e.g., 1,000 to 20,000 units). In comparison, treatment of neurogenic inflammation or hyperactive sweat glands may require a relatively lower transdermal dose of botulinum toxin (e.g., about 1 to 1,000 units).

[0256] Some embodiments of the present invention envision pharmaceutical compositions comprising stabilized botulinum toxin for transdermal delivery to human patients. The botulinum toxin may be selected from botulinum toxin types A, B, C1, D, E, F, and G, isolated and / or purified (i.e., about 150 kDa) botulinum toxin, and naturally occurring or recombinant botulinum toxin. In some embodiments, the composition may contain about 1 to about 50,000 units of botulinum toxin, and the composition may contain an amount of botulinum toxin sufficient to achieve a therapeutic effect lasting from one month to five years.

[0257] In some embodiments, the present invention provides a topical formulation of botulinum toxin (e.g., a botulinum emulsion composition) that allows a significant amount of botulinum toxin to penetrate the skin of a target body without penetrating the blood vessels. 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 1% of the botulinum toxin present in the pharmaceutical composition penetrates the blood vessels upon application of the topical and / or transdermal formulation of the present invention.

[0258] In some embodiments, the present invention provides a topical formulation of an antibody drug (e.g., an antibody drug emulsion composition) that can penetrate the skin of a target body without a significant amount of the antibody drug penetrating the blood vessels. 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 formulation 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 target body and, in a significant amount, penetrate the blood vessels. 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 target body and, in a therapeutically effective amount, penetrate the blood vessels. For example, in some embodiments of the present invention, about 25%, 50%, 75%, 90%, or more than 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 a target body.

[0260] Those skilled in the art will understand that the compositions of the present invention, which achieve transdermal administration of botulinum toxin or antibody drugs, can be incorporated into devices such as patches, rollers, pens, stamps, etc. [Examples]

[0261] Example 1: Effect of microneedle skin conditioning (MSC) pretreatment on the bioavailability of botulinum toxin. A single-dose local study was conducted to evaluate the bioavailability of botulinum toxin after topical administration of botulinum nanoemulsion. The skin was conditioned with a microneedle array by pressing the array against the skin before application of the local treatment. The array was then removed before application of the botulinum treatment. The study was designed to not only evaluate but also compare the % bioavailability achieved using various microneedle systems, thereby testing the effects of changes in needle density and microneedle puncture size (e.g., puncture area per microneedle) of the microneedle array on botulinum bioavailability.

[0262] The study included four test groups, each consisting of eight rats. Each group underwent the microneedle skin conditioning described above. Except for the control group, each group received a single topical treatment with a fixed volume and concentration of botulinum nanoemulsion on the skin covering the biceps femoris, gastrocnemius, and tibialis anterior muscle of the right hind limb. The nanoemulsion was applied with a gloved finger. Administration of the topical formulation to the skin took approximately 10 minutes, at which point the formulation was completely absorbed into the skin. Since it is known that a sufficient dose of botulinum can induce death in animals, the effect of such treatment was measured by mortality. Therefore, the mortality rates of the four treatment groups were compared. Specifically, the mortality rate at 3 days post-treatment was used as the relative bioavailability of botulinum toxin, where an increase in mortality represented an increase in the bioavailability of the toxin.

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

[0264] As can be seen, meaningful bioavailability was achieved with each microneedling approach (i.e., in each of groups A, B, and C).

[0265] Furthermore, by comparing group C to either group A or B, it was surprisingly observed that fewer and smaller punctures resulted in higher bioavailability than more and larger punctures. In other words, reducing the density of microneedles increased bioavailability, as did reducing the size of the microneedles. By comparing group A and group B to each other, it was surprisingly observed that increasing the length of the microneedles did not improve bioavailability.

[0266] In other words, the test results determined that the density of microneedles and the puncture size of the microneedles can significantly improve 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 using a microneedle array to condition the skin before topical administration of macromolecules in the emulsion composition, the needle density of the microneedle array should be 31 needles / cm². 2 Reducing it to less than 36,000 μm increased the bioavailability of macromolecules. Furthermore, when using a microneedle array to condition the skin before topical administration of macromolecules in the emulsion composition, the needle puncture size (per microneedle) of the microneedle array was reduced to 36,000 μm per microneedle. 2 Reducing it to below a certain level increased the bioavailability of macromolecules. [Table 6]

[0267] Example 2: Effect of MSC preconditioning on the bioavailability of botulinum toxin in humans: Effect on reducing sweat This study conducts a single-dose local trial to evaluate the bioavailability of botulinum toxin after local administration of a topical botulinum nanoemulsion formulation in humans. The trial is designed not only to evaluate but also to compare the % bioavailability achieved using various microneedle systems, and as a result, the effect of changes in the needle density of the microneedle array on improving botulinum bioavailability in humans was tested by measuring the reduction in skin sweat after local treatment with the botulinum nanoemulsion formulation.

[0268] The study will include one subject. Each sample is located on the abdomen and has an area of ​​approximately 2 cm². 2 Three spots, each 5 cm apart, are selected and marked with a marker. Each spot is treated once with a fixed volume of botulinum nanoemulsion preparation at a fixed concentration of botulinum. The administration of the topical preparation to the skin takes approximately 5 minutes, at which point the preparation is completely absorbed into the skin. The first spot is a control site with no preconditioning using a microneedle array. The second spot is treated with 9 microneedles / cm before application of the botulinum preparation. 2 The first intervention site is preconditioned with three presses of a microneedle array at a microneedle density of 85 microneedles / cm² before application of the botulinum toxin preparation. 2 The intervention site #2 was preconditioned with three presses of a microneedle array at a microneedle density.

[0269] The expected effect of such treatment is a reduction in sweating at the site of botulinum nanoemulsion treatment. The amount of sweat at the treatment site is measured in two ways: 1) the evaporometer test, which detects the rate of sweating using an instrument used to measure the rate of water evaporation from the skin (more evaporation is detected as sweating increases); or 2) the iodine-starch test, in which povidone-iodine is applied to the treatment site of the subject, dried, and then corn starch is sprinkled on the treatment site. The corn starch turns purple when the subject sweats, and remains white when the subject does not sweat; this is called the iodine-starch test. In either sweat detection method, the subject is placed under a heating lamp to induce sweating, and then the sweat detection method is used.

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

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

[0272] Example 3: Effect of MSC preconditioning with varying microneedle density on the bioavailability of botulinum toxin in humans: Effect on reducing sweat This study conducts a single-dose local study to evaluate the bioavailability of botulinum toxin after local administration of a topical botulinum nanoemulsion formulation in humans. The study is designed not only to evaluate but also to compare the % bioavailability achieved using various microneedle systems. As a result, the study examines the effect of changes in needle density of microneedle arrays on the significant improvement in botulinum bioavailability in humans by measuring the reduction in skin sweat after local treatment with the botulinum nanoemulsion formulation.

[0273] The study will include 12 subjects. Each subject has a lesion on their back, with an area of ​​approximately 2 cm². 2 Three spots, each 5 cm apart, are selected and marked with a marker. Each spot is treated once with a fixed volume of botulinum nanoemulsion preparation at a fixed concentration of botulinum. The administration of the topical preparation to the skin takes approximately 5 minutes, at which point the preparation is completely absorbed into the skin. The first spot is a control site with no preconditioning using a microneedle array. The second spot is treated with 9 microneedles / cm before application of the botulinum preparation. 2 The first intervention site was preconditioned with five presses of a microneedle array at a microneedle density of 85 microneedles / cm² before application of the botulinum toxin. 2 The intervention site #2 was pre-conditioned with five presses of a microneedle array at a microneedle density.

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

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

[0276] This study demonstrated that microneedle preconditioning using a relatively lower microneedling density unexpectedly increased the bioavailability of large-scale topical drug nanoemulsions 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 reducing sweat This study conducts a single-dose local trial to evaluate the bioavailability of botulinum toxin after local administration of a topical botulinum nanoemulsion formulation in humans. The trial is designed not only to evaluate but also to allow for comparison of the % bioavailability achieved using various microneedle systems. As a result, the study examines the effect of changes in microneedle puncture size (e.g., puncture area per microneedle) on the significant improvement in botulinum bioavailability in humans by measuring the reduction in skin sweat after local treatment with the botulinum nanoemulsion formulation.

[0278] The study will include 12 subjects. Each subject has a lesion on their back, with an area of ​​approximately 2 cm². 2 Three spots, each 5 cm apart, are selected and marked with a marker. Each spot is treated once with a fixed volume of botulinum nanoemulsion preparation at a fixed concentration of botulinum. The administration of the topical preparation to the skin takes approximately 5 minutes, at which point the preparation is completely absorbed into the skin. The first spot is a control site with no preconditioning using a microneedle array. The second spot is approximately 11,000 μm before application of the botulinum preparation. 2 The first intervention site was preconditioned with five presses of a microneedle array of microneedle puncture size. The third spot was approximately 60,000 μm before application of the botulinum toxin preparation. 2 The treatment site #2 is preconditioned with five presses of a microneedle array of microneedle puncture size.

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

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

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

[0282] Example 5: Effect of MSCs on the bioavailability of botulinum toxin in humans: Effect on reducing sweat and wrinkles. This study conducts a single-dose local study to evaluate the bioavailability of botulinum toxin after local administration of a topical botulinum nanoemulsion formulation in humans. The study is designed not only to evaluate but also to compare the % bioavailability achieved using various microneedle systems, and as a result, the effect of changes in the needle density of the microneedle array on the improvement of botulinum bioavailability in humans was tested by measuring the reduction in skin sweat and wrinkles after local treatment with the botulinum nanoemulsion formulation.

[0283] The study will include one subject with severe forehead (or horizontal) wrinkles. Each wrinkle is located on the subject's forehead and has an area of ​​approximately 2 cm². 2 Three spots, each 5 cm apart, are selected and marked with a marker. Each spot is treated once with a fixed volume of botulinum nanoemulsion preparation at a fixed concentration of botulinum. The administration of the topical preparation to the skin takes approximately 5 minutes, at which point the preparation is completely absorbed into the skin. The first spot is a control site with no preconditioning using a microneedle array. The second spot is treated with 9 microneedles / cm before application of the botulinum preparation. 2 The first intervention site was preconditioned with five presses of a microneedle array at a microneedle density of 85 microneedles / cm² before application of the botulinum toxin. 2 The intervention site #2 was pre-conditioned with five presses of a microneedle array at a microneedle density.

[0284] The expected effect of such treatment is a reduction in sweating at the site of botulinum nanoemulsion treatment. The amount of sweat at the treatment site is measured in two ways: 1) the evaporometer test, which detects the rate of sweating using an instrument used to measure the rate of water evaporation from the skin (more evaporation is detected as sweating increases); or 2) the iodine-starch test, in which povidone-iodine is applied to the treatment site of the subject, dried, and then corn starch is sprinkled on the treatment site. The corn starch turns purple when the subject sweats, and remains white when the subject does not sweat; this is called the iodine-starch test. In either sweat detection method, the subject enters a sauna to induce sweating, and then the method is used.

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

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

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

[0288] Example 6: Effect of MSCs with varying microneedle density on the bioavailability of botulinum toxin in humans: Effect on reducing sweat in subjects with hyperhidrosis This study conducts a single-dose local trial to evaluate the bioavailability of botulinum toxin after local administration of a topical botulinum nanoemulsion formulation in humans. The trial is designed not only to evaluate but also to compare the % bioavailability achieved using various microneedle systems, and as a result, the effect of changes in the needle density of the microneedle array on improving botulinum bioavailability in humans was tested by measuring the reduction in skin sweat after local treatment with the botulinum nanoemulsion formulation.

[0289] The study included three treatment groups, each consisting of 12 subjects with axillary hyperhidrosis, characterized by excessive sweating in the armpits: Group 1 was the control group, in which botulinum nanoemulsion was applied to the armpits of each subject; Group 2 was the intervention group #1, in which 9 microneedles / cm were applied to each part of the armpit skin before the application of the botulinum nanoemulsion formulation. 2 Group 3 was preconditioned with five presses of a microneedle array at a microneedle density; Group 3 was intervention group #1, and each part of the underarm skin was treated with 85 microneedles / cm² before application of the botulinum nanoemulsion formulation. 2 The subjects are preconditioned with five presses of a microneedle array at a microneedle density. Each subject in groups 1, 2, and 3 is treated once topically with a fixed volume of botulinum nanoemulsion formulation at a fixed concentration of botulinum. The administration of the topical formulation to the skin takes approximately 5 minutes, at which point the formulation is completely absorbed into 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 sweat at the treatment site is measured by gravimetric sweating (GS test): the armpit of the subject is dried with a paper towel; the weight of the filter paper is weighed; the filter paper is placed in the armpit for 5 minutes and then weighed again. The extra weight after re-weighing the paper is the weight of sweat produced by the subject in 5 minutes. The severity of the subject's hyperhidrosis is measured by the subject using the Hyperhidrosis Sweat Severity Scale (HDSS), which is a 4-point scale from 0=none, 1=mild, 2=moderate, to 3=severe.

[0291] The GS test and HDSS were used at baseline before botulinum nanoemulsion treatment, and at 2 weeks and 4 weeks post-treatment. The study found that at baseline, the average amount of sweat detected by the GS test or disease severity measured by HDSS was approximately equal across groups 1–3. At 2 and 4 weeks post-treatment, the average amount of sweat detected or disease severity in group 1 was greater than that detected in groups 2 or 3 at those post-treatment weeks. The study also found that at 2 and 4 weeks post-treatment, the average amount of sweat detected or disease severity in group 3 was greater than that detected in group 2 at those post-treatment weeks.

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

[0293] Example 7: Effect of MSCs with varying microneedle puncture size on the bioavailability of botulinum toxin in humans: Effect on reducing sweat in subjects with hyperhidrosis This study conducts a single-dose local trial to evaluate the bioavailability of botulinum toxin after local administration of a topical botulinum nanoemulsion formulation in humans. The trial is designed not only to evaluate but also to allow for comparison of the % bioavailability achieved using various microneedle systems. As a result, the effect of changes in microneedle puncture size on improving botulinum bioavailability in humans was tested by measuring the reduction in skin sweat after local treatment with the botulinum nanoemulsion formulation.

[0294] The study included three treatment groups, each consisting of 12 subjects with axillary hyperhidrosis, characterized by excessive sweating in the armpits: Group 1 was the control group, in which botulinum nanoemulsion was applied to the armpits of each subject; Group 2 was the intervention group #1, in which approximately 11,000 μm of botulinum nanoemulsion was applied to each part of the armpit skin before the application of the botulinum nanoemulsion formulation. 2The microneedles were pre-conditioned with five presses of a microneedle array of microneedle puncture size; group 3 was intervention group #1, and approximately 60,000 μm of botulinum nanoemulsion was applied to each part of the axillary skin before application of the botulinum nanoemulsion formulation. 2 The microneedles are pre-conditioned by pressing a microneedle array five times, each with a microneedle puncture size. Each subject in groups 1, 2, and 3 is treated once locally with a fixed volume of botulinum nanoemulsion formulation at a fixed concentration of botulinum. The administration of the topical formulation to the skin takes approximately 5 minutes, at which point the formulation is completely absorbed into 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 sweat at the treatment site is measured by gravimetric sweating (GS test): the armpit of the subject is dried with a paper towel; the weight of the filter paper is weighed; the filter paper is placed in the armpit for 5 minutes and then weighed again. The extra weight after re-weighing the paper is the weight of sweat produced by the subject in 5 minutes. The severity of the subject's hyperhidrosis is measured by the subject using the Hyperhidrosis Sweat Severity Scale (HDSS), which is a 4-point scale from 0=none, 1=mild, 2=moderate, to 3=severe.

[0296] The GS test and HDSS were used at baseline before botulinum nanoemulsion treatment, and at 2 weeks and 4 weeks post-treatment. The study found that at baseline, the average amount of sweat detected by the GS test or disease severity measured by HDSS was approximately equal across groups 1–3. At 2 and 4 weeks post-treatment, the average amount of sweat detected or disease severity in group 1 was greater than that detected in groups 2 or 3 at those post-treatment weeks. The study also found that at 2 and 4 weeks post-treatment, the average amount of sweat detected or disease severity in group 3 was greater than that detected in group 2 at those post-treatment weeks.

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

[0298] Example 8: Effect of MSCs on the bioavailability of botulinum toxin in humans: Effect on reducing crow's feet wrinkles. A single-dose local study was conducted to evaluate the bioavailability of botulinum toxin after local administration of a topical botulinum nanoemulsion formulation in humans. The study was designed not only to evaluate the % bioavailability achieved using various microneedle systems but also to allow for comparison. As a result, the effect of changes in the needle density of the microneedle array on the improvement of botulinum bioavailability in humans was tested by measuring the reduction in skin wrinkles after local treatment with the botulinum nanoemulsion formulation.

[0299] One subject with severe crow's feet wrinkles was included in the study. Botulinum nanoemulsion was applied to the subject's crow's feet wrinkles. 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 the crow's feet wrinkles when the subject contracted the muscles that cause crow's feet wrinkles, as measured on a 5-point wrinkle rating scale. The subject received 9 microneedles / cm² into each area of ​​skin where the crow's feet wrinkles were located on side #1 of the face before application of the botulinum nanoemulsion formulation to side #1 of the face. 2 The skin is pre-conditioned with five presses of a microneedle array at a microneedle density, and then 85 microneedles / cm² are applied to each area of ​​skin where the crow's feet are located, before application of the botulinum nanoemulsion formulation to the opposite side of the face (side #2). 2 The skin was pre-conditioned with five presses of a microneedle array at a microneedle density. Administration of the topical formulation to the skin took approximately 5 minutes, at which point the formulation was fully absorbed into the skin.

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

[0301] The wrinkle scale was used at baseline before botulinum nanoemulsion treatment, 2 weeks after treatment, and 4 weeks after treatment. At baseline, subjects were found to have severe wrinkles as assessed by the wrinkle scale, with a score of 4 on a 5-point scale. At 2 weeks post-treatment, the mean wrinkle severity decreased by 1 point on the wrinkle scale, with a score of 3 (moderate) on side #2 of the face. At 4 weeks post-treatment, the mean wrinkle severity decreased 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 post-treatment, the mean wrinkle severity decreased by 2 points on the wrinkle scale, with a score of 2 (mild) on side #1 of the face. At 4 weeks post-treatment, the mean wrinkle severity decreased 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 large topical drug nanoemulsions containing botulinum toxin.

[0303] Example 9: Effect of MSCs with varying microneedle density on the bioavailability of botulinum toxin in humans: Effect on reducing crow's feet wrinkles. This study conducts a single-dose local trial to evaluate the bioavailability of botulinum toxin after local administration of a topical botulinum nanoemulsion formulation in humans. The trial is designed not only to evaluate but also to allow for comparison of the % bioavailability achieved using various microneedle systems. As a result, the effect of changes in the needle density of the microneedle array on improving botulinum bioavailability in humans was tested by measuring the reduction in skin wrinkles after local treatment with the botulinum nanoemulsion formulation.

[0304] The study included three treatment groups, each consisting of 12 subjects with severe crow's feet (outer corners of the eyes): Group 1 was the control group, in which botulinum nanoemulsion was applied to the crow's feet of each subject; 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 before the application of the botulinum nanoemulsion formulation. 2 Group 3 is pre-conditioned with five presses of a microneedle array at a microneedle density; group 3 is intervention group #1, and 85 microneedles / cm² are applied to each area of ​​skin where crow's feet wrinkles are located before application of the botulinum nanoemulsion formulation. 2 The subjects are preconditioned with five presses of a microneedle array at a microneedle density. Each subject in groups 1, 2, and 3 is treated once topically with a fixed volume of botulinum nanoemulsion formulation at a fixed concentration of botulinum. The administration of the topical formulation to the skin takes approximately 5 minutes, at which point the formulation is completely absorbed into the skin.

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

[0306] The wrinkle scale was used at baseline before botulinum nanoemulsion treatment, and at 2 weeks and 4 weeks post-treatment. The study found that at baseline, the mean severity of wrinkles detected by the wrinkle scale was approximately the same across groups 1–3. At 2 and 4 weeks post-treatment, the mean severity of wrinkles in group 1 was greater than that detected in groups 2 and 3 at those post-treatment weeks. The study also found that at 2 and 4 weeks post-treatment, the mean severity of wrinkles in group 3 was greater than that detected in group 2 at those post-treatment weeks.

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

[0308] Example 10: Effect of MSCs with varying microneedle puncture size on the bioavailability of botulinum toxin in humans: Effect on reducing crow's feet wrinkles. This study conducts a single-dose local trial to evaluate the bioavailability of botulinum toxin after local administration of a topical botulinum nanoemulsion formulation in humans. The trial is designed not only to evaluate but also to allow for comparison of the % bioavailability achieved using various microneedle systems. As a result, the effect of changes in microneedle puncture size on improving botulinum bioavailability in humans was tested by measuring the reduction in skin wrinkles after local treatment with the botulinum nanoemulsion formulation.

[0309] The study included three treatment groups, each consisting of 12 subjects with severe crow's feet (outer corners of the eyes): Group 1 was the control group, in which botulinum nanoemulsion was applied to the crow's feet of each subject; Group 2 was the intervention group #1, in which approximately 11,000 μm of botulinum nanoemulsion was applied to each part of the skin where the crow's feet were located before the application of the botulinum nanoemulsion formulation. 2 Preconditioning is performed by pressing a microneedle array of microneedle puncture size five times; group 3 is the intervention group #1, and approximately 60,000 μm of microneedle material is applied to each part of the skin where the crow's feet wrinkles are located before application of the botulinum nanoemulsion formulation. 2 The microneedles are pre-conditioned by pressing a microneedle array five times, each with a microneedle puncture size. Each subject in groups 1, 2, and 3 is treated once locally with a fixed volume of botulinum nanoemulsion formulation at a fixed concentration of botulinum. The administration of the topical formulation to the skin takes approximately 5 minutes, at which point the formulation is completely absorbed into the skin.

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

[0311] The wrinkle scale was used at baseline before botulinum nanoemulsion treatment, and at 2 weeks and 4 weeks post-treatment. The study found that at baseline, the mean severity of wrinkles detected by the wrinkle scale was approximately the same across groups 1–3. At 2 and 4 weeks post-treatment, the mean severity of wrinkles in group 1 was greater than that detected in groups 2 and 3 at those post-treatment weeks. The study also found that at 2 and 4 weeks post-treatment, the mean severity of wrinkles in group 3 was greater than that detected in group 2 at those post-treatment weeks.

[0312] This study demonstrated that microneedle preconditioning with relatively smaller microneedle puncture sizes unexpectedly increased the bioavailability of large topical drug nanoemulsions, including botulinum toxin.

[0313] Example 11: Effect of MSCs on the bioavailability of botulinum toxin in humans: Effect of dose changes on the reduction of crow's feet wrinkles. This study conducts a single-dose local trial to evaluate the bioavailability of botulinum toxin after local administration of a topical botulinum nanoemulsion formulation in humans. The trial is designed not only to evaluate but also to allow for comparison of the % bioavailability achieved using various microneedle systems. As a result, the effect of changes in the needle density of the microneedle array on improving botulinum bioavailability in humans was tested by measuring the reduction in skin wrinkles after local treatment with the botulinum nanoemulsion formulation.

[0314] The study included three treatment groups, each consisting of 12 subjects with severe crow's feet: Group 1 was the control group, in which botulinum nanoemulsion was applied to the crow's feet of each subject; 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 before the application of the botulinum nanoemulsion formulation. 2 Group 3 is pre-conditioned with five presses of a microneedle array at a microneedle density; group 3 is intervention group #1, and 85 microneedles / cm² are applied to each area of ​​skin where crow's feet wrinkles are located before application of the botulinum nanoemulsion formulation. 2 The subjects are pre-conditioned with five presses of a microneedle array at a microneedle density. Each subject is treated once topically with a fixed volume of botulinum nanoemulsion formulation at a fixed concentration of botulinum, except that treatment in group 1 is twice the botulinum concentration of treatments in groups 2 and 3. Administration of the topical formulation to the skin takes approximately 5 minutes, at which point the formulation is completely absorbed into the skin.

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

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

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

[0318] Example 12: Effect of MSCs on the bioavailability of botulinum toxin in humans: Effect of dose changes on the reduction of crow's feet wrinkles. This study conducts a single-dose local trial to evaluate the bioavailability of botulinum toxin after local administration of a topical botulinum macroemulsion formulation in humans. The trial is designed not only to evaluate but also to compare the % bioavailability achieved using various microneedle systems, and as a result, the effect of changes in needle density of microneedle arrays on improving botulinum bioavailability in humans was tested by measuring the reduction in skin wrinkles after local treatment with the botulinum macroemulsion formulation.

[0319] The study included three treatment groups, each consisting of 12 subjects with severe crow's feet: Group 1 was the control group, in which botulinum macroemulsion was applied to the crow's feet of each subject; 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 before the application of the botulinum macroemulsion preparation. 2 Group 3 is pre-conditioned with 5 presses of a microneedle array at a microneedle density; group 3 is intervention group #1, and 85 microneedles / cm² are applied to each area of ​​skin where crow's feet wrinkles are located before application of the botulinum macroemulsion formulation. 2 The microneedles are pre-conditioned with five presses of a microneedle array at a specific microneedle density. Each target is treated once with a fixed volume of botulinum nanoemulsion formulation at a fixed concentration of botulinum. The administration of the topical formulation to the skin takes approximately 5 minutes, at which point the formulation is completely absorbed into the skin.

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

[0321] The wrinkle scale was used at baseline before botulinum nanoemulsion treatment, and at 2 weeks and 4 weeks post-treatment. The study found that at baseline, the mean severity of wrinkles detected by the wrinkle scale was approximately the same across groups 1-3. At 2 and 4 weeks post-treatment, the mean severity of wrinkles in group 1 was greater than that in groups 2 and 3. The study also found that at 2 and 4 weeks post-treatment, the mean severity of wrinkles in group 3 was greater than that in group 2.

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

[0323] Example 13: Effect of MSC preconditioning on the bioavailability of botulinum toxin in humans by varying the number of microneedle presses: Effect on reducing crow's feet wrinkles. A single-dose local study was conducted to evaluate the bioavailability of botulinum toxin after local administration of a topical botulinum nanoemulsion formulation in humans. The study examined the effect of changes in the number of microneedle array presses on improving botulinum bioavailability in humans by measuring the reduction in skin wrinkles after local 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 comprised subjects with moderate to severe crow's feet wrinkles. The crow's feet area of ​​each subject was treated once topically with a botulinum emulsion (e.g., nanoemulsion) formulation containing a nearly constant dose of botulinum. The topical formulation was administered to the skin in approximately 5 minutes, at which point it was completely absorbed. Subjects in Group A (N=8) were subjected to 14 microneedle presses (or 4 microneedle presses / cm) of a microneedle array before the application of the botulinum formulation. 2 The subjects in group B (N=17) were pre-conditioned with eight microneedle presses (or two microneedle presses / cm) of a microneedle array before application of the botulinum toxin preparation. 2 Preconditioning was performed using ). The lengths of the microneedles in the microneedle arrays used in groups A and B were the same.

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

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

[0327] This study demonstrated that microneedle preconditioning using relatively less microneedle pressure (per unit area or in absolute number) unexpectedly increased the bioavailability of localized large drug nanoemulsions containing botulinum toxin.

[0328] Example 14: Effect of MSC preconditioning with varying microneedle length on the bioavailability of botulinum toxin in humans: Effect on reducing crow's feet wrinkles. A single-dose local study was conducted to evaluate the bioavailability of botulinum toxin after topical administration of a botulinum nanoemulsion formulation in humans. The study examined the effect of changes in microneedle length on improving botulinum bioavailability in humans by measuring the reduction in skin wrinkles after topical treatment with the formulation following skin conditioning with a microneedle array.

[0329] The study included three groups, Group A, Group B, and Group C, all of which had moderate to severe crow's feet wrinkles. The crow's feet area of ​​each subject was treated once with an emulsion formulation, particularly a nanoemulsion topical botulinum toxin formulation. Subjects in Group A (N=9) received skin preconditioning using a 500 μm needle, subjects in Group B (N=9) received skin preconditioning using an 800 μm needle, and subjects in Group C (N=9) received skin preconditioning using a 1400 μm needle.

[0330] In this particular example, the administration of the topical formulation to the skin took approximately 5 minutes, at which point the formulation was completely absorbed into the skin. All subjects were preconditioned with the same number of microneedle presses of a microneedle array before application of the botulinum formulation. The doses of botulinum used in groups A, B, and C were made identical across all subjects.

[0331] The expected effect of botulinum nanoemulsion treatment is a reduction in crow's feet wrinkles at the treatment site; such reductions were evaluated for the various treatments applied in this embodiment. Wrinkle severity was measured by both the investigator and the subjects 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 with a reduction of 2 points or more in wrinkle severity compared to baseline, as evaluated by both the investigator and the subjects.

[0332] In this study, the mean severity of crow's feet wrinkles, as measured by a wrinkle scale at baseline, was found to be nearly equal across groups A, B, and C. At 12 weeks post-treatment, the responder rate was 36% in group A, 14% in group B, and 13% in group C.

[0333] This study demonstrated that using shorter microneedles during microneedle skin preconditioning unexpectedly increases the bioavailability of large topical drug nanoemulsions containing botulinum toxin.

[0334] Example 15: Effect of MSC preconditioning on the bioavailability of botulinum toxin in humans with varying doses of topically applied formulations: Effect on reducing crow's feet wrinkles. A single-dose local study was conducted to evaluate the bioavailability of botulinum toxin after topical administration of a botulinum nanoemulsion formulation in humans. The study examined the effect of changes in the volume of topically applied botulinum nanoemulsion formulation on improving botulinum bioavailability in humans by measuring the reduc...

Claims

1. A nanoemulsion composition containing a large drug having a molecular weight of 100,000 Da or more, to be applied in combination with microneedle skin conditioning (MSC) at a site using a microneedle array at that site, A) The microneedle array has approximately 2 to 30 microneedles / cm². 2 Having a microneedle density within the range; or B) The microneedle array is approximately 100 to 35,000 μm 2 Having a microneedle puncture hole size within the range of microneedles, The major drug is botulinum toxin. Nanoemulsion composition.

2. The microneedle array has approximately 2 to 10 microneedles / cm². 2 Or approximately 2 to 20 microneedles / cm 2 The nanoemulsion composition according to claim 1, having a microneedle density within the range.

3. The nanoemulsion composition according to claim 1 or 2, wherein the composition is formulated as a lotion, cream, ointment, liniment, or gel.

4. The composition further comprises a non-irritating penetration enhancer, preferably, A) A non-irritating penetration enhancer is selected from the carrier peptide; and / or B) A non-irritating penetration enhancer, with the sequence RKKRRQRRRG-(K) 15 - Selected from cationic peptides having GRKKRRQRRR and positively charged carriers, The nanoemulsion composition according to any one of claims 1 to 3.

5. Site MSCs are performed before the application of a composition containing large drugs to the site; or Site MSCs are performed after application of the composition containing the large drug to the site; or The application of the site to the MSC and the composition containing the large drug to the site occurs substantially simultaneously. The nanoemulsion composition according to any one of claims 1 to 4.

6. The nanoemulsion composition according to any one of claims 1 to 5, wherein a large drug is delivered together with a biologically active substance, the biologically active substance is preferably selected from steroids, retinoids, anesthetics and / or collagen, and more preferably selected from hydrocortisone and / or lidocaine.

7. MSC of the site is achieved using a device comprising multiple needles, the device preferably being a patch, roller, stamp or pen, and the needles preferably being A) Having sufficient length to protrude through the stratum corneum of the skin; and / or B) Having insufficient length to reach nerves in the dermis of the skin; and / or C) Having a length of approximately 10 μm to approximately 4000 μm or approximately 10 μm to approximately 800 μm or approximately 10 μm to approximately 500 μm; and / or D) Having a length of approximately 200 μm or more, or approximately 300 μm or more, or approximately 500 μm or more; and / or E) Composed of a biocompatible material or a metal, wherein the biocompatible material is preferably a soluble polymer. The nanoemulsion composition according to any one of claims 1 to 6.

8. The body part, A) The skin surface covering the muscles or muscle groups of the subject; and / or B) Skin surface including sweat glands; and / or C) Skin surface including sebaceous glands; and / or D) Skin surface including hair follicles The nanoemulsion composition according to any one of claims 1 to 7.

9. A nanoemulsion composition according to any one of claims 1 to 8, wherein the composition is for treating or preventing a disorder selected from among unwanted sweating, body odor, hyperhidrosis, bromhidrosis, chromatohidrosis, conditions involving muscle spasms and / or contractures, strabismus, hemifacial spasm, tremor, spasticity, migraine or other headaches, acne, rosacea, impaired excessive sebum production, facial wrinkles, unsightly facial expressions, neck wrinkles, hyperfunctional facial wrinkles, hyperkinetic facial wrinkles, platysma bands and / or combinations thereof.

10. A nanoemulsion composition containing large drugs with a molecular weight of 100,000 Da or more, and approximately 2 to approximately 30 microneedles / cm 2 A patch comprising multiple microneedles having a microneedle density within a certain range, wherein the major drug is botulinum toxin.

11. The needle A) Having sufficient length to protrude through the stratum corneum of the skin; and / or B) Having insufficient length to reach nerves in the dermis of the skin; and / or C) Having a length of approximately 10 μm to approximately 4000 μm; and / or D) Having a length of approximately 200 μm or more, or approximately 300 μm or more, or approximately 500 μm or more; and / or E) Composed of a biocompatible material or a metal, wherein the biocompatible material is preferably a soluble polymer. The patch according to claim 10.

12. A kit comprising a nanoemulsion composition containing a large drug having a molecular weight of 100,000 Da or more, and a device for microneedle conditioning of a certain site, wherein the device is A) Approximately 2 to 30 microneedles / cm 2 The density of microneedles within the range; or B) Approximately 100 to approximately 35,000μm 2 / Microneedle puncture hole size within the range of microneedles A microneedle array having, The major drug is botulinum toxin. kit.

13. The kit according to claim 12, wherein the composition is formulated as a lotion, cream, ointment, liniment, or gel.

14. The kit according to claim 12 or 13, wherein the device is a patch, roller, stamp or pen, and / or the kit includes the patch according to claim 10 or 11.

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