Topical compositions of thyrotropin-releasing hormone and uses thereof

Abstract

The present disclosure relates in some aspects to pharmaceutical compositions, such as topical compositions, useful for treating or preventing hair loss and / or hair graying in a subject. In some aspects, also provided are kits comprising the compositions, and methods of using the compositions for treating a condition, such as hair loss or hair graying.

Description

[0001] 2025-10-09 TOPICAL COMPOSITIONS OF THYROTROPIN-RELEASING HORMONE

[0002] AND USES THEREOF

[0003] INVENTOR : RALF PAUS

[0004] CROSS-REFERENCE

[0005]

[0001] Priority is claimed under PCT Article 8(1) and Rule 4.10 to U.S. Provisional App. No. 63 / 705,476, filed October 9, 2024, which is incorporated by reference for all purposes as if fully set forth herein.

[0006] FIELD OF THE INVENTION

[0007]

[0002] The present disclosure relates in some aspects to pharmaceutical compositions useful for treating or preventing hair loss in a subject. In some aspects, the disclosure also provides kits comprising the pharmaceutical compositions, and methods of using disclosed pharmaceutical compositions for treating or preventing hair loss.

[0008] BACKGROUND OF THE INVENTION

[0009]

[0003] Pattern hair loss, also known as androgenetic alopecia, is a hair loss condition that affects up to 50% of men and 25% of women by the age of 50 (Vary JC, Med Clin North Am. 2015;99(6): 1195— 1211 ). The cause of pattern hair loss remains unclear, and may be related to oxidative stress, the scalp microbiome, and / or hormonal abnormalities. There is a lack of available treatments for pattern hair loss, particularly those that are effective and do not cause undesirable side effects. Therefore, the continued prevalence of pattern hair loss leaves an unmet need to alleviate and treat this condition.

[0010] INCORPORATION BY REFERENCE

[0011]

[0004] Each cited patent, publication, and non-patent literature is incorporated by reference in its entirety, as if each was incorporated individually, and as if each is fully set forth herein. However, no such citation should be construed as an admission that a cited reference comes from an area that is analogous or directly applicable to the invention, nor should a citation be construed as an admission that a document or underlying information, in any jurisdiction, is prior art or forms part of the common general knowledge in the art.

[0012] BRIEF SUMMARY OF THE INVENTION

[0013]

[0005] The following is a simplified summary of some embodiments of the invention in order to provide a basic understanding thereof. It is not an extensive overview of the invention, and is not intended to identify key or critical elements of the invention or to delineate the full scope thereof. Its purpose is to present some embodiments of the invention in a simplified form as a prelude to the more detailed description that follows.

[0014]

[0006] In one aspect, provided is a composition comprising thyrotropin-releasing hormone (TRH). In some embodiments, the composition is formulated for topical administration. 2025-10-09

[0007] In some embodiments, the composition comprises between about 25 nM and about 500 pM of TRH. In some embodiments, the composition comprises between about 100 nM and about 100 pM of TRH. In some embodiments, the composition comprises between about 1 pM and about 100 pM of TRH. In some embodiments, the composition comprises about 25 pM of TRH.

[0015]

[0008] In some embodiments, the composition comprises one or more pharmaceutically acceptable excipients selected from the group consisting of penetration enhancers, carriers, diluents, emulsifiers, stabilizers, viscosity modifying agents, adhesion modifying agents, preservatives, antioxidants, adhesive polymers, solubilizing agents, colorants, binders, humectants, surfactants, and gelling agents.

[0016]

[0009] In some embodiments, the composition comprises a solvent system. In some embodiments, the solvent system comprises an alcohol. In some embodiments, the solvent system comprises any of ethanol, isopropanol, and propylene glycol.

[0017]

[0010] In some embodiments, the solvent system comprises isopropanol. In some embodiments, the composition comprises between about 10% and about 70% (v / v) of isopropanol. In some embodiments, the composition comprises about 40% (v / v) of isopropanol.

[0018]

[0011] In some embodiments, the solvent system comprises ethanol. In some embodiments, the composition comprises about 60% (v / v) of ethanol.

[0019]

[0012] In some embodiments, the solvent system comprises propylene glycol. In some embodiments, the composition comprises between about 5% and about 80% (v / v) of propylene glycol. In some embodiments, the composition comprises about 10% (v / v) of propylene glycol. In some embodiments, the composition comprises about 20% (v / v) of propylene glycol. In some embodiments, the composition comprises about 50% (v / v) of propylene glycol.

[0020]

[0013] In some embodiments, the solvent system comprises water. In some embodiments, the composition comprises between about 1 % and about 50% (v / v) of water. In some embodiments, the composition comprises about 30% (v / v) of water.

[0021]

[0014] In some embodiments, the composition comprises one or more additional active agents. In some embodiments, the composition comprises an additional thyroid hormone. In some embodiments, the additional thyroid hormone is triiodothyronine (T3). In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of between about 1 :1 and about 1 :1000. In some embodiments, the composition comprises any of finasteride, dutasteride, and minoxidil. In some embodiments, the composition comprises finasteride. In some embodiments, the composition comprises dutasteride. In some embodiments, the composition comprises minoxidil. In some embodiments, the composition comprises T3 and finasteride. In some embodiments, the composition comprises T3 and dutasteride. In some embodiments, the composition comprises T3 and minoxidil. In some embodiments, the composition comprises TRH as the only hormone. In some embodiments, the composition comprises a peptidase inhibitor. In some embodiments, the peptidase inhibitor is an inhibitor of TRH-degrading ectoenzyme. In 2025-10-09 some embodiments, the peptidase inhibitor is a tripeptide. In some embodiments, the tripeptide comprises a Glp-Asn-Pro moiety.

[0022]

[0015] Also provided is a composition consisting essentially of TRH, wherein the composition is formulated for topical administration.

[0023]

[0016] Also provided is a composition consisting essentially of TRH and T3, wherein the composition is formulated for topical administration.

[0024]

[0017] Also provided is a composition consisting essentially of TRH, T3, and any of finasteride, dutasteride, and minoxidil; wherein the composition is formulated for topical administration.

[0025]

[0018] Also provided is a composition comprising a TRH metabolite, wherein the composition is formulated for topical administration. In some embodiments, the metabolite is Cyclo(His-Pro).

[0026]

[0019] Also provided is a composition comprising a TRH peptide fragment, wherein the composition is formulated for topical administration. In some embodiments, the peptide fragment is a dipeptide. In some embodiments, the dipeptide is Pyroglutamyl Dipeptide-30 Amide.

[0027]

[0020] In some embodiments, the composition is in lyophilized form.

[0028]

[0021] In some embodiments, the composition is in unit dosage form. In some embodiments, the total unit dose volume of the composition is between about 0.1 mL and about 10 mL.

[0029]

[0022] In some embodiments, the composition is formulated as a spray, ointment, salve, gel, paste, lotion, liniment, or cream.

[0030]

[0023] Also provided is the composition of any of the disclosed embodiments for use in treating or preventing hair loss or hair graying.

[0031]

[0024] Also provided is the use of the composition of any of the disclosed embodiments for the manufacture of a medicament for treating or preventing hair loss or hair graying.

[0032]

[0025] Also provided is a method of treating or preventing hair loss or hair graying in a subject, comprising administering to the subject the composition of any of the disclosed embodiments.

[0033]

[0026] In some embodiments, the method comprises administering to the subject between about 0.1 and 10 mL of the composition per unit dose. In some embodiments, the method comprises administering to the subject about 1 mL of the composition per unit dose.

[0034]

[0027] In some embodiments, the method comprises administering to the subject between about 30 pg and 200 pg of TRH per unit dose. In some embodiments, the method comprises administering to the subject about 10 pg of TRH per unit dose.

[0035]

[0028] In some embodiments, the composition is administered daily. In some embodiments, the composition is administered every other day. In some embodiments, the composition is administered every other day for several consecutive weeks followed by a prolonged period without administration. In some embodiments, the composition is administered every other day for two consecutive weeks followed by a prolonged period without administration. In some embodiments, the prolonged period without 2025-10-09 administration is at least two weeks.

[0036]

[0029] In some embodiments, the hair loss is caused by androgenetic alopecia, alopecia areata, persistent patchy alopecia areata, alopecia totalis, alopecia universalis, diffuse alopecia areata, ophiasis alopecia, cicatricial alopecia, lichen planopilaris, frontal fibrosing alopecia, central centrifugal cicatricial alopecia (CCCA), traction alopecia, alopecia barbae, or postpartum alopecia.

[0037]

[0030] In some embodiments, the method results in increased hair shaft production. In some embodiments, hair shaft production is increased by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to baseline values measured prior to administration of the composition.

[0038]

[0031] In some embodiments, the method results in prolonged anagen hair growth phase. In some embodiments, the anagen hair growth phase is prolonged by 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to baseline values measured prior to administration of the composition.

[0039]

[0032] In some embodiments, the method results in increased proliferation of bulge epithelial stem cells. In some embodiments, bulge epithelial stem cell proliferation is increased by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to baseline values measured prior to administration of the composition.

[0040]

[0033] In some embodiments, the method results in increased expression of keratin 15. In some embodiments, the expression of keratin 15 is increased by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to baseline values measured prior to administration of the composition.

[0041]

[0034] In some embodiments, the method results in decreased hair depigmentation. In some embodiments, hair depigmentation is decreased by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0042]

[0035] In some embodiments, the method results in hair repigmentation.

[0043]

[0036] The foregoing has outlined broadly and in summary certain pertinent features of the disclosure so that the detailed description of the invention that follows may be better understood, and so that the present contribution to the art can be more fully appreciated. Hence, this summary is to be considered as a brief and general synopsis of only some of the objects and embodiments disclosed herein, is provided solely for the benefit and convenience of the reader, and is not intended to limit in any manner the scope, or range of equivalents, to which the claims are lawfully entitled. Additional features of the invention are described hereinafter. It should be appreciated by those in the art that all disclosed specific compositions and methods are only exemplary, and may be readily utilized as a basis for modifying or designing other compositions and methods for carrying out the same purposes. Such equivalent compositions and methods will be appreciated to be also within the scope and spirit of the invention as set forth in the claims. 2025-10-09

[0044] BRIEF DESCRIPTION OF THE FIGURES

[0045]

[0037] To further clarify various aspects of the invention, a more particular description is rendered by reference to certain exemplary embodiments illustrated in the figures. It will be appreciated that these figures depict only illustrated embodiments of the invention and should not be considered limiting of its scope. They are merely provided as exemplary illustrations of certain concepts of some embodiments of the invention. These figures, and the elements depicted therein, are not necessarily drawn to consistent scale or to any scale. Unless context suggests otherwise, like elements are indicated by like numerals. Certain aspects of the invention are therefore further described and explained with additional specificity and detail, but still by way of example only, with reference to the accompanying figures in which:

[0046]

[0038] FIG. 1 shows hematoxylin and eosin (H&E) stained sections of scalp skin after six days of treatment. Sections treated with Vehicle, 25.48 nM, and 2.548 M TRH show mild hyperkeratosis without parakeratosis compared with vehicle. The section treated with 254.8 nM TRH shows mild hyperkeratosis and mild signs of parakeratosis.

[0047]

[0039] FIG. 2 shows hair shaft production (in pm) from organ-cultured human scalp skin over the course of six days upon treatment with vehicle, 25.48 nM, 254.8 nM, or 2.548 pM topical TRH; photographs of representative skin cultures at Day 1 and Day 6 of treatment. Topical TRH (25.48 nM and 254.8 nM) significantly enhances hair-shaft production in organ-cultured human scalp skin. Mean ± SEM; n=37— 53 hair follicles (HFs) from 1 donor; Mann-Whitney test *p<0.05, **p<0.01.

[0048]

[0040] FIG. 3 shows a stacked bar graph of hair-cycle staging, revealing that nanomolar TRH does not significantly alter the proportion of follicles remaining in anagen after six days; Ki-67 / caspase-3 and Warthin-Starry (WS) stained images are shown, illustrating similar proliferative and apoptotic activity across treatments. Mean ± SEM; n=10— 14 HFs from 1 donor; Mann-Whitney test, not significant.

[0049]

[0041] FIG. 4 shows hair matrix keratinocyte proliferation, indicating that 25.48 nM TRH significantly increases the percentage of Ki-67-positive cells compared with vehicle; representative double-immunofluorescence images showing an increased Ki-67 signal in TRH-treated follicles are shown. Mean ± SEM; n=7— 10 HFs from 1 donor; Mann-Whitney test, *p<0.05.

[0050]

[0042] FIG. 5 shows WS intensity in anagen VI HFs, demonstrating increased melanin production in the 25.48 nM TRH treatment group; representative histological images of WS staining in anagen VI HFs are shown. Mean ± SEM; n=5— 10 anagen VI HFs from 1 donor; Mann-Whitney test, not significant.

[0051]

[0043] FIG. 6 shows TGF-02 expression in outer root sheath (ORS) keratinocytes, demonstrating that TGF-02 expression is reduced in all TRH treatment groups relative to vehicle; representative immunofluorescent images of TGF-02 expression in the outer root sheath are shown. Mean ± SEM; n=10— 14 HFs from 1 donor; Mann-Whitney test, *p<0.05.

[0052]

[0044] FIG. 7 shows IGF-1 expression in ORS keratinocytes and dermal papilla (DP) cells, demonstrating that IGF-1 expression is significantly increased in the ORS but not the DP in the 254.8 nM 2025-10-09 TRH treatment group; representative immunofluorescent images of IGF-1 expression in the outer root sheath are shown. Mean ± SEM; n=3— 19 HFs from 1 donor; Mann-Whitney test, *p<0.05.

[0053]

[0045] FIG. 8 shows FGF-7 expression in hair matrix (HM) and ORS keratinocytes, demonstrating that FGF-7 expression in HM is decreased in the 2.548 pM TRH treatment group and increased in ORS in the 254.8 nM treatment group; representative immunofluorescent images of FGF-7 expression are shown. Mean ± SEM; n=10— 14 HFs from 1 donor; Mann-Whitney test, **p<0.01.

[0054]

[0046] FIG. 9 shows K85 expression in the proximal hair matrix, demonstrating that 25.48 nM TRH may increase K85 expression, whereas higher nanomolar and micromolar doses may decrease K85 expression; representative immunofluorescent images of K85 expression are shown. Mean ± SEM; n=10— 14 HFs from 1 donor; Mann-Whitney test, not significant.

[0055]

[0047] FIG. 10 shows VEGF-A expression in the hair matrix, demonstrating that TRH does not modulate VEGF-A production; representative immunofluorescent images of VEGF-A expression are shown. Mean ± SEM; n=12— 20 HFs from 1 donor; Mann-Whitney test, not significant.

[0056]

[0048] FIG. 11 shows the percentage of CD31+cells in the papillary dermis (PD), demonstrating that TRH does not modulate CD31+cell proliferation; representative immunofluorescent images of CD31+expression are shown. Mean ± SEM; n=12-13 sections from 2 punches from 1 donor; Mann-Whitney test, not significant.

[0057]

[0049] FIG. 12 shows VEGF-A+cells in the PD, demonstrating that topical TRH significantly decreases the percentage of VEGF-A+cells in the human scalp dermis; representative immunofluorescent images of VEGF-A expression are shown. Mean ± SEM; n=10— 12 sections from 2 punches from 1 donor; Mann-Whitney test, *p<0.05, ***p<0.001.

[0058]

[0050] FIG. 13 shows epidermal VEGF-A expression, demonstrating that 254.8 nM TRH significantly increases VEGF-A production whereas 25.48 nM TRH decreases it; representative immunofluorescent images of VEGF-A expression are shown. Mean ± SEM; n=7— 12 sections from 2 punches from 1 donor; Mann-Whitney test, **p<0.01 , ***p<0.001.

[0059]

[0051] FIG. 14 shows H&E stained sections of scalp skin after six days of micromolar TRH treatment, demonstrating that 2.548 pM and 25.48 pM TRH may induce mild hyperkeratosis but not parakeratosis compared with vehicle.

[0060]

[0052] FIG. 15 shows % hair shaft production from organ-cultured human scalp skin after six days of vehicle, 2.548 pM TRH, or 25.48 pM TRH treatment, demonstrating that 25.48 pM TRH slightly enhances hair shaft production compared with vehicle and 2.548 pM TRH; representative photographs of skin cultures at Day 1 and Day 6 of treatment are shown. Mean ± SEM; n=8 HFs from 1 donor; Mann-Whitney test, not significant.

[0061]

[0053] FIG. 16 shows a stacked bar graph of hair-cycle staging, indicating that 2.548 pM TRH trends toward prolonged anagen duration whereas 25.48 pM TRH has little effect; representative Ki-67 2025-10-09 (proliferation) and caspase-3 (apoptosis) double-immunofluorescence and Warthin-Starry (WS) stained images are shown. Mean ± SEM; n=4-9 HFs from 1 donor; Mann-Whitney test, not significant.

[0062]

[0054] FIG. 17 shows hair matrix keratinocyte proliferation, demonstrating that 25.48 pM TRH trends towards an increase in the percentage of Ki-67-positive cells whereas 2.548 pM TRH does not; representative double-immunofluorescence images of Ki-67 and caspase-3 staining are shown. Mean ± SEM; n=5-8 HFs from 1 donor; Mann-Whitney test, not significant.

[0063]

[0055] FIG. 18. shows melanin production in anagen VI HFs, demonstrating that micromolar TRH does not affect WS staining intensity compared with vehicle; representative WS images are shown. Mean ± SEM; n=2-5 anagen VI HFs from 1 donor.

[0064]

[0056] FIG. 19 shows the fold change in TGF-02 expression in outer root sheath (ORS) keratinocytes, demonstrating that 2.548 pM and 25.48 pM TRH significantly reduce TGF-02 expression relative to vehicle; representative immunofluorescent images of TGF-02 expression in ORS are shown. Mean ± SEM; n=3— 6 HFs from 1 donor; Mann-Whitney test, *p<0.05.

[0065]

[0057] FIG. 20 shows IGF-1 expression in ORS keratinocytes and DP cells, demonstrating that 2.548 pM and 25.48 pM TRH significantly reduce IGF-1 expression in both compartments; representative immunofluorescent images of IGF-1 expression are shown. Mean ± SEM; n=3— 6 HFs from 1 donor; Mann-Whitney test, *p<0.05, **p<0.01, ***p<0.001.

[0066]

[0058] FIG. 21 shows the fold change in FGF-7 expression in ORS keratinocytes, demonstrating that 2.548 pM and 25.48 pM TRH slightly increase FGF-7 expression in the ORS; representative immunofluorescent images of FGF-7 expression are shown. Mean ± SEM; n=3— 6 HFs from 1 donor.

[0067]

[0059] FIG. 22 shows K85 expression in the proximal hair matrix, demonstrating that 2.548 pM and

[0068] 25.48 pM TRH may increase K85 expression compared with vehicle; representative immunofluorescent images of K85 expression are shown. Mean ± SEM; n=4-5 HFs from 1 donor.

[0069]

[0060] FIG. 23 shows K15 expression in bulge stem cells, demonstrating that 25.48 pM TRH significantly decreases the percentage and expression of K15+bulge cells; representative immunofluorescent images of K15 expression are shown. Mean ± SEM; n=3— 8 HFs from 1 donor; Mann-Whitney test, *p<0.05.

[0070]

[0061] FIG. 24 shows the percent K157Ki-67+among K15+cells in bulge stem cells, demonstrating that

[0071] 25.48 pM TRH may increase this percentage; representative immunofluorescent images of K157Ki-67+in human scalp skin are shown. Mean ± SEM; n=3— 8 HFs from 1 donor.

[0072]

[0062] FIG. 25 shows VEGF-A production in the hair matrix, demonstrating that 2.548 pM and 25.48 pM TRH decreases VEGF-A expression compared with vehicle; representative immunofluorescent images of VEGF-A staining are shown. Mean ± SEM; n=4-6 HFs from 1 donor; Mann-Whitney test, *p<0.05, **p<0.01.

[0073]

[0063] FIG. 26 shows the percentage of CD31+cells in the dermis, demonstrating that 2.548 pM and

[0074] 25.48 pM TRH decreases CD31+cell proportion compared with vehicle; representative immunofluorescent 2025-10-09 images of CD31 staining are shown. Mean ± SEM; n=4-6 HFs from 1 donor; Mann-Whitney test, *p<0.05.

[0075]

[0064] FIG. 27 shows VEGF-A+expression in the dermis, demonstrating that 2.548 pM and 25.48 pM TRH decreases the percentage of VEGF-A+dermal cells; representative immunofluorescent images of VEGF-A expression are shown. Mean ± SEM; n=4-6 HFs from 1 donor; Mann-Whitney test, *p<0.05.

[0076]

[0065] FIG. 28 shows a principal-component analysis (PCA) of RNA-seq samples. Each point represents an individual hair-follicle sample projected onto the first two principal components based on the top 5,000 variable genes. Control samples cluster near the origin, whereas low-, medium- and high-dose T3+TRH samples separate progressively along the PC1 axis, indicating dose-dependent transcriptomic shifts.

[0077]

[0066] FIG. 29A shows a volcano plot depicting the magnitude of change between a low dose of T3+TRH and control. The plot shows the Iog2 fold change (x-axis) versus — Iog10 adjusted p-value (y-axis) for genes in each comparison. Circle points denote genes not significantly different, triangle-pointing down points are significantly down-regulated and triangle-pointing up points are significantly up-regulated (adjusted p < 0.05). The distribution of points illustrates that low and high doses predominantly up-regulate genes, whereas the medium dose has a modest skew towards down-regulation.

[0078]

[0067] FIG. 29B shows a volcano plot depicting the magnitude of change between a medium dose of T3+TRH and control. The plot shows the Iog2 fold change (x-axis) versus — Iog10 adjusted p-value (y-axis) for genes in each comparison. Circle points denote genes not significantly different, triangle-pointing down points are significantly down-regulated and triangle-pointing up points are significantly up-regulated (adjusted p < 0.05). The distribution of points illustrates that low and high doses predominantly up-regulate genes, whereas the medium dose has a modest skew towards down-regulation.

[0079]

[0068] FIG. 29C shows a volcano plot depicting the magnitude of change between a high dose of T3+TRH and control. The plot shows the Iog2 fold change (x-axis) versus — Iog10 adjusted p-value (y-axis) for genes in each comparison. Circle points denote genes not significantly different, triangle-pointing down points are significantly down-regulated and triangle-pointing up points are significantly up-regulated (adjusted p < 0.05). The distribution of points illustrates that low and high doses predominantly up-regulate genes, whereas the medium dose has a modest skew towards down-regulation.

[0080]

[0069] FIG. 30 shows a heatmap depicting normalized expression of the top 50 most variable genes across all samples. Rows correspond to genes (annotated by Ensembl IDs) and columns correspond to samples grouped by treatment. The color scale represents relative expression (z-score). Hierarchical clustering of both genes and samples highlights distinct expression patterns for control versus low, medium and high doses of T3+TRH.

[0081]

[0070] FIG. 31A shows a dot plot summarizing KEGG pathway enrichment for a low dose of T3+TRH. Gene ratio on the x-axis represents the proportion of differentially expressed genes mapping to a pathway; dot size corresponds to gene count and color denotes adjusted p-value. Low-dose treatment shows broad enrichment across metabolism and signalling pathways (HIF-1 , PPAR, AMPK, glycolysis, fatty-acid 2025-10-09 metabolism), whereas medium and high doses highlight cornified envelope formation alongside selected metabolic and signalling pathways.

[0082]

[0071] FIG. 31 B shows a dot plot summarizing KEGG pathway enrichment for a medium dose of T3+TRH. Gene ratio on the x-axis represents the proportion of differentially expressed genes mapping to a pathway; dot size corresponds to gene count and color denotes adjusted p-value. Low-dose treatment shows broad enrichment across metabolism and signalling pathways (HIF-1 , PPAR, AMPK, glycolysis, fatty-acid metabolism), whereas medium and high doses highlight cornified envelope formation alongside selected metabolic and signalling pathways.

[0083]

[0072] FIG. 31 C shows a dot plot summarizing KEGG pathway enrichment for a high dose of T3+TRH. Gene ratio on the x-axis represents the proportion of differentially expressed genes mapping to a pathway; dot size corresponds to gene count and color denotes adjusted p-value. Low-dose treatment shows broad enrichment across metabolism and signalling pathways (HIF-1 , PPAR, AMPK, glycolysis, fatty-acid metabolism), whereas medium and high doses highlight cornified envelope formation alongside selected metabolic and signalling pathways.

[0084] DETAILED DESCRIPTION OF THE INVENTION

[0085]

[0073] While various aspects and features of certain embodiments are summarized above, the following detailed description illustrates several exemplary embodiments in further detail to enable one of skill in the art to practice such embodiments, and to make and use the full scope of the invention claimed. The described examples are provided for illustrative purposes and are not intended to limit the scope of the invention or its applications. It will be understood that many modifications, substitutions, changes, and variations in the described examples, embodiments, applications, and details of the invention illustrated herein can be made by those skilled in the art without departing from the spirit of the invention, or the scope of the invention as described in the appended claims. It also will be appreciated that the headings within this document are being utilized only to expedite its review by a reader. They should not be construed as limiting the invention in any manner.

[0086]

[0074] The scope of the invention includes all embodiments and formulations thereof, not only those expressly described below, and it will be understood that many modifications, substitutions, changes, and variations in the described embodiments, applications, and details of the invention illustrated herein can be made by those skilled in the art without departing from the spirit of the invention, or the scope of the invention as set forth in the appended claims, and the general principles defined herein may be applied to a wide range of aspects. Thus, the invention is not intended to be limited to the aspects and embodiments presented, but is to be accorded the widest scope consistent with the principles and novel features disclosed, including their equivalents. The description will make such aspects and embodiments apparent to one of skill, in that such aspects and embodiments will be readily cognizable and readily creatable without undue experimentation, solely using the teachings herein and the general knowledge of the art. 2025-10-09

[0087]

[0075] While the methods described and illustrated herein may include particular steps, it should be apparent that other methods including fewer, more, or different steps than those described and shown are also within the spirit and scope of the invention.

[0088]

[0076] The disclosed methods (and uses of any disclosed composition), and any associated steps shown herein, therefore should be understood as being provided for purposes of illustration, not limitation. It should be further understood that the specific order or hierarchy of steps in the methods disclosed are only exemplary approaches. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The specific order or hierarchy of steps in any methods thus may be rearranged according to ordinary skill, while remaining within the spirit and scope of the disclosure. Any presented claims also will present elements of the steps in a sample and exemplary order, and are not meant to be limited to the specific order presented.

[0089]

[0077] Unless otherwise stated, all measurements, values, ratings, positions, dimensions, magnitudes, sizes, locations, orientations, configurations, and other specifications that are set forth (either expressly or impliedly) in this specification, including in the figures and in the claims, are approximate, and not exact. They are intended to have a reasonable range that is consistent with the functions to which they relate and with what is customary in the art to which they pertain. Moreover, the recitation of ranges of values is merely intended to serve as a shorthand method of referring individually to each separate value falling within the range. Unless otherwise indicated, each individual value is incorporated into the specification as if it were individually recited herein.

[0090]

[0078] The use of any and all examples, or exemplary language provided with respect to an embodiment, is intended merely to better illuminate certain non-limiting aspects of the invention and does not pose a limitation on the scope of the invention as otherwise claimed. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the invention.

[0091]

[0079] While the invention and its various aspects are described in terms of particular embodiments and applications, it is not intended that these descriptions in any way limit its scope to any such embodiments and applications, and it will be understood that many modifications, substitutions, changes, and variations in the described embodiments, applications, and details of the invention illustrated herein can be made by those skilled in the art without departing from the spirit of the invention, or the scope of the invention as described in the claims.

[0092] A. General Definitions and Terms

[0093]

[0080] As used in this specification and the appended claims, the singular forms “a,” “an,” and “the” include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to “an active agent” includes reference to a combination of two or more active agents, and reference to “an excipient” includes reference to a combination of two or more excipients. While the term “one or more” may be used, its absence (or its replacement by the singular) does not signify the singular only, but simply 2025-10-09 underscores the possibility of multiple agents or ingredients in particular embodiments.

[0094]

[0081] The terms “comprising,” “including,” “such as,” and “having” are intended to be inclusive and not exclusive (i.e., there may be other elements in addition to the recited elements). Thus, the term “including” means, and is used interchangeably with, the phrase “including but not limited to.” The term “or” is used herein to mean, and is used interchangeably with, the term “and / or,” unless context clearly indicates otherwise.

[0095]

[0082] Unless otherwise indicated, all numbers expressing quantities of ingredients, properties such as concentration, reaction conditions, and so forth, used to describe and claim certain embodiments of the invention are to be understood as being modified in some instances by the term “about.” Accordingly, in some embodiments, the numerical parameters set forth in the written description and attached claims are approximations that can vary depending upon the desired properties sought to be obtained by a particular embodiment. In some embodiments, “about” refers to plus or minus five percent (±5%) of the recited unit of measure. The term “substantially,” where it is applied to modify a feature or limitation herein, will be read in the context of the invention and in light of the knowledge in the art to provide the appropriate certainty, e.g., by using a standard that is recognized in the art for measuring the meaning of “substantially” as a term of degree, or by ascertaining the scope as would one of skill in the art.

[0096]

[0083] In some embodiments, the numerical parameters should be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. Notwithstanding that the numerical ranges and parameters setting forth the broad scope of some embodiments of the invention are approximations, the numerical values set forth in the specific examples are reported as precisely as practicable. The numerical values presented in some embodiments may contain certain errors necessarily resulting from the standard deviation found in their respective testing measurements.

[0097]

[0084] Unless defined otherwise, all technical and scientific terms herein have the meaning as commonly understood by one having ordinary skill in the art to which this invention belongs, who as a shorthand may be referred to simply as “one of skill.” Further definitions that may assist the reader in understanding the disclosed embodiments are as follows; however, it will be appreciated that such definitions are not intended to limit the scope of the invention, which shall be properly interpreted and understood by reference to the full specification (as well as any plain meaning known to one of skill in the relevant art) in view of the language used in the appended claims. The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting.

[0098]

[0085] Generally, the nomenclature used and procedures performed herein are those known in fields relating to one or more aspects of the invention, such as biology, biochemistry, dermatology, pharmacology, and medical science, and are those that will be well known and commonly employed in such fields. Standard techniques and procedures will be those generally performed according to conventional methods in the art. 2025-10-09

[0099]

[0086] Where definitions are included herein, they are for purposes of assisting the reader in understanding the disclosed embodiments; however, it will be appreciated that any such definitions are not intended to limit the scope of the invention, which shall be properly interpreted and understood by reference to the full specification (as well as any plain meaning known to one of skill) in view of the language used in the claims. The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting.

[0100]

[0087] Further definitions follow, to assist a reader in understanding the embodiments.

[0101]

[0088] Herein, “hair cycle staging” refers to anagen (growth phase), catagen (transitional phase), and telogen (resting phase), the key stages of human hair (Paus et al. J Investigative Derm. 2001 ; 117(1 ):3-15).

[0102]

[0089] Herein, “melanin” refers to a complex polymer that originates from the amino acid tyrosine.

[0103] Melanin is present in human skin in varying degrees, and is responsible for your unique eye, hair, and skin color (Cao et al. J Am Chem Soc. 2021 ; 143(7):2622-2637).

[0104]

[0090] Herein, “hair shaft production” refers to the biological process by which proliferating matrix cells in the hair bulb divide and then differentiate into keratinocytes that build the keratinised hair fiber (the visible hair shaft) (Natarelli et al. J Clin Med. 2023; 12(3):893).

[0105]

[0091] Herein, “HM keratinocyte proliferation” refers to the rapid division of hair matrix (HM) keratinocytes - mitotically active cells located in the follicular matrix - which, during the anagen (growth) phase, proliferate and differentiate to extend the hair shaft, whereas in the resting phase these cells show minimal activity (Tang et al. Stem Cells Int. 2023;2023:5548112).

[0106]

[0092] Herein, “Ki-67” refers to a protein that is expressed exclusively in actively dividing cells. The expression of Ki-67, measured, for example, with immunofluorescent techniques, can be used to quantify actively cycling cells in hair follicles (Baar et al. Acta Derm Venereol. 1992;72(3):161-164).

[0107]

[0093] Herein, “caspase-3” is a caspase protein that interacts with caspase-8 and caspase-9. Caspases are regulators of programmed cell death, and very likely some specific caspases may function as mediators of the hair growth cycle (Sawaya et al. Eur J Dermatol. 2001 ; 11 (4):304-308).

[0108]

[0094] Herein, “IGF-1,” or “insulin-like growth factor 1 ,” refers to an anagen prolonging growth factor. IGF-1 shares a high degree of structural and functional homology with insulin and exhibits anti-apoptotic effects (Ahn et al. Ann Dermatol. 2012;24(1 ):26-31 ).

[0109]

[0095] Herein, “KGF / FGF7,” or “keratinocyte growth factor / fibroblast growth factor-7,” refers to an anagen prolonging growth factor.

[0110]

[0096] Herein, “TGF -2,” or “transforming growth factor beta 2,” refers to a catagen promoting growth factor (Xu et al. Bone Research. 2018;6(2): 1-31).

[0111]

[0097] Herein “MTCO1,” or “mitochondrial cytochrome C oxidase subunit I,” refers to a component of the cytochrome c oxidase, the last enzyme in the mitochondrial electron transport chain which drives oxidative phosphorylation (Vidali et al. J Invest Dermatol. 2016;136(10):2003-2012). 2025-10-09

[0112]

[0098] Herein, “K85” refers to a sensitive marker for the amount of hair shaft keratin production (Ramot et al. Br J Dermatol. 2013; 169(1): 146-51).

[0113]

[0099] Herein, “K15”, or “keratin 15,” refers to a type I keratin commonly used as a marker of stem-cell-enriched keratinocytes in the hair follicle outer root sheath— particularly the bulge region (Bose et al. Int J Mol Sci. 2013;14:19385-19398).

[0114]

[0100] Herein, “CD31,” also known as “platelet endothelial cell adhesion molecule-1” is a marker of intracutaneous angiogenesis, measured by CD31 immunoreactivity and the number of CD31 positive cells. CD31 is a transmembrane homophilic receptor that is expressed by endothelial cells, platelets, granulocytes, macrophages, dendritic cells, T- and B- calls, and natural killer cells (Berg et al. J Cell Sci. 2013;126(11):2343-2352.

[0115]

[0101] Herein, “VEGFA,” or “vascular endothelial growth factor A,” refers to a secreted pro-angiogenic cytokine that binds VEGFR1 and, predominantly, VEGFR2 to drive endothelial proliferation, migration, and vascular permeability during angiogenesis.

[0116]

[0102] Herein, “% of K15+ cells” or “% of K15+Ki67+ cells” or “% of CD31 + cells” or “% of VEGFA+ cells” refers to the percentage of cells within a defined region of interest that exhibit specific immuno-positivity for the indicated marker(s).

[0117]

[0103] Herein, “treating” or “treatment” of a disorder, such as hair loss, a hair loss condition, or androgenetic alopecia includes any treatment of the disorder in a mammal, preferably in a human, and includes: (a) preventing a disorder from occurring in a patient who may be predisposed to the disorder but has not yet been diagnosed with it; (b) inhibiting a disorder, i.e., arresting its development, and including prophylaxis; (c) relieving a disorder, i.e., causing regression of the disorder or its clinical symptoms; (d) protection from or relief of a symptom or pathology caused by or related to a disorder; (e) reduction, decrease, inhibition, amelioration, or prevention of onset, severity, duration, progression, frequency or probability of one or more symptoms or pathologies associated with a disorder; and (f) prevention or inhibition of a worsening or progression of symptoms or pathologies associated with a disorder or comorbid with a disorder.

[0118]

[0104] Herein, “an effective amount,” a “therapeutically effective amount,” or “a pharmacologically effective amount” refers to an amount of an active agent, e.g., thyrotropin-releasing hormone (TRH) that is non-toxic and sufficient to provide the desired therapeutic effect with performance at a reasonable benefit / risk ratio attending any medical treatment. The effective amount will vary depending upon the subject, the weight and age thereof, the severity of the symptoms or degree of health benefit sought, the manner of administration, and the like, all of which can readily be determined by one of skill in the art.

[0119]

[0105] Herein, “therapeutic effect” or “therapeutic efficacy” means the responses(s) in a subject, and preferably a human, after treatment that are judged to be desirable and beneficial. Hence, depending on the symptoms to be treated, or improvement in health or functioning sought, and depending on the 2025-10-09 particular constituent(s) of the methods of the disclosure under consideration, those responses shall differ, but would be readily understood by those of skill in the art.

[0120]

[0106] As used herein, the terms “subject,” “user,” “patient,” and “individual” are used interchangeably, and refer to a human, a mammal, or any other animal susceptible to hair loss, a hair loss condition, or androgenetic alopecia. Preferably, the subject is a human. The subject may be a human infant, a human child, a human adult, or an elderly human. Such terms will be understood to include one who has an indication for which a method described herein may be efficacious, or who otherwise may benefit by the invention. In general, all of the disclosed methods will be appreciated to work for all individuals, although individual variation is to be expected, and will be understood. The disclosed methods of treatment also can be modified to treat multiple patients at once, including couples or families. Hence, these terms will be understood to also mean two or more individuals.

[0121]

[0107] Still additional definitions and abbreviations are provided elsewhere herein.

[0122] B. Compositions

[0123]

[0108] The present disclosure relates in some aspects to methods of treating or preventing hair loss (e.g., caused by a hair loss condition, such as androgenetic alopecia) in a subject (e.g., preferably a human). In some aspects, the disclosure further relates to pharmaceutical (e.g., topical) compositions and kits used in the methods. In some aspects, useful features of the disclosed methods include curing or alleviating the symptoms of a subject suffering from a hair loss condition, such as androgenetic alopecia.

[0124]

[0109] Without being bound by theory, TRH may have the potential to stimulate hair growth by prolonging anagen, stimulating keratinocyte proliferation, and upregulating keratin production, if an active concentration can be delivered to human scalp hair follicles.

[0125]

[0110] Without being bound by theory, TRH may have the potential to stimulate proliferation and differentiation of stem cells both in the hair matrix and in the hair bulge. In organ-cultured human scalp hair follicles, TRH promotes hair-shaft elongation, prolongs the anagen growth phase and antagonises the catagen-promoting factor TGF-02. These effects are accompanied by increased proliferation and reduced apoptosis of hair-matrix keratinocytes and reduced phosphorylation of p53 and checkpoint kinases, suggesting a supportive environment for stem-cell expansion. TRH modulates keratin expression by up-regulating hair keratins K31 and K32 and epithelial keratins K6, K14 and K17 while down-regulating K85 and K86; it also up-regulates keratin-6 during wound healing, potentially enhancing bulge progenitor activity. Beyond the hair follicle, TRH stimulates mitochondrial biogenesis and respiratory-chain activity in human epidermal keratinocytes, increases prolactin synthesis and regulates prolactin-receptor expression in hair follicles, and selectively stimulates melanin synthesis and tyrosinase activity in hair-follicle melanocytes. TRH accelerates re-epithelialisation and keratinocyte proliferation in wounded frog and human skin and further increases hair-shaft production and hair-matrix keratinocyte proliferation. These collective findings suggest that TRH can modulate multiple signalling pathways - TGF-02 suppression, 2025-10-09 keratin regulation, mitochondrial activation and neuroendocrine feedback loops - to enhance stem-cell proliferation and differentiation in hair follicles and skin. See, for example, Paus et al. FASEB J. 2010;24(2):393— 403; Paus et al. Br J Dermatol. 2010;162(5):1127— 1131 ; Paus et al. J Invest Dermatol. 2011 ;131 (12):2368— 2377; Paus et al. J Clin Endocrinol Metab. 2012;97(3):978— 986; Paus et al. Br J Dermatol. 2013;169(1 ):146— 151 ; Paus et al. PLoS One. 2013;8:e73596; Paus et al. J Invest Dermatol. 2010; Paus et al. Trends Mol Med. 2014; and Paus et al. BioEssays. 2014; each of which is hereby incorporated by reference in its entirety. Additional mechanisms of action may exist but remain unknown, owing to the complexity of the numerous intracellular signalling cascades activated by TRH.

[0126]

[0111] In one aspect, provided is a composition comprising a therapeutically effective amount of TRH. In some embodiments, a composition comprises a therapeutically effective amount of TRH and a therapeutically effective amount of triiodothyronine (T3). In some embodiments, a composition comprises a therapeutically effective amount of TRH and a therapeutically effective amount of finasteride. In some embodiments, a composition comprises a therapeutically effective amount of TRH and a therapeutically effective amount of dutasteride. In some embodiments, a composition comprises a therapeutically effective amount of TRH and a therapeutically effective amount of minoxidil. In some embodiments, a composition comprises a therapeutically effective amount of TRH and a therapeutically effective amount of cetirizine. In some embodiments, a composition comprises a therapeutically effective amount of TRH and a therapeutically effective amount of levocetirizine. In some embodiments, a composition comprises a therapeutically effective amount of TRH and a therapeutically effective amount of latanoprost.

[0127]

[0112] For any embodiment disclosed herein comprising TRH, including compositions comprising TRH and methods of use thereof, it shall be understood that the disclosure also encompasses analogs, derivatives, metabolites, and peptide fragments of TRH, whether naturally occurring or synthetically produced. “Analog” refers to a compound having a chemical structure that is substantially related to that of TRH, wherein one or more atoms, functional groups, or peptide residues are added, removed, substituted, or otherwise modified, including, e.g., stereoisomers, isosteres, compounds comprising C- or N-terminus substitutions (e.g., acetylation, formylation, PEGylation, cyclization), isotopologs, and the like. “Metabolite” refers to chemical compounds derived from TRH, which are produced in vivo following administration and metabolism of a TRH, such as in a disclosed composition. Metabolites of TRH are known to those of skill in the art, and include, for example, Cyclo(His-Pro). “Derivative” refers to a chemically modified form of TRH, including, for example, salts, solvates, esters, ethers, and prodrugs. “Peptide fragment” refers to any shorter peptide sequence derived from, contained within, or modeled upon (e.g., in the case of certain synthetic polypeptides) TRH, including, for example, pyroglutamyl dipeptides such as Pyroglutamyl Dipepti de-30 Amide, or other truncated or modified TRH peptide sequences.

[0128]

[0113] In some embodiments, a composition is suitable for topical or transdermal administration. In some embodiments, a composition is formulated for topical administration (i.e., a “topical composition”). In some 2025-10-09 embodiments, a composition is formulated for transdermal administration. In some embodiments, a composition comprises one or more pharmaceutically acceptable excipients.

[0129]

[0114] In embodiments, a composition is formulated into a topical dosage form. Topical dosage forms include transmucosal and transdermal formulations, such as aerosols, emulsions, sprays, ointments, salves, gels, pastes, lotions, liniments, oils, and creams. Topical dosage forms may comprise a penetrant or carrier. Penetrants include, for transmucosal administration, detergents, bile salts, fusidic acid derivatives, and combinations thereof. Carriers include Vaseline®, lanolin, PEG, alcohols, transdermal enhancers, and combinations thereof.

[0130]

[0115] In embodiments, a composition is formulated for transdermal application. In general, transdermal delivery involves contacting the formulations with a subject’s skin under conditions effective for the active agent(s) to penetrate the skin and cause an effect. Transdermal formulations include ointments, creams, suspensions, lotions, pastes, gels, sprays, foams, oils, and combinations thereof. An exemplary transdermal delivery form is a transdermal “patch,” which may be used to provide continuous or discontinuous infusion of active agent(s) in controlled amounts. Patches may be constructed for continuous, gradual, pulsatile, or on demand delivery of the agents. In embodiments, a patch is a medicated adhesive patch, a single-layer or multi-layer drug-in-adhesive patch, a “matrix” (or “monolithic”) patch, or a “reservoir” patch. In embodiments, a patch is part of a delivery system, such as used with an electronic device coupled to a subject’s mobile device, and / or coupled with a mobile app (e.g., to control a delivery rate from a reservoir, and / or to provide information about delivery to the app or user). Various such technologies will be known and may be used.

[0131]

[0116] In some embodiments, a composition is formulated for injection. Formulations for injection may comprise physiologically acceptable sterile aqueous or non-aqueous solutions, dispersions, suspensions or emulsions, liposomes, and sterile powders for reconstitution into sterile solutions or dispersions, and also may comprise additives such as solubilizers, stabilizers, and suspending, preserving, wetting, emulsifying, dispensing, and isotonic agents.

[0132]

[0117] In another aspect, provided is a composition, useful for treating or preventing hair loss comprising: (i) TRH; (ii) a pharmaceutically acceptable excipient; and (iii) a solvent system. In some embodiments, the composition contains TRH as the only active ingredient. Hence, also provided is a composition, useful for treating or preventing hair loss comprising: (i) TRH as the only active ingredient; (ii) a pharmaceutically acceptable excipient; and (iii) a solvent system. Also provided is a composition, useful for treating or preventing hair loss consisting essentially of: (i) TRH; (ii) a pharmaceutically acceptable excipient; and (iii) a solvent system. In some embodiments, the composition comprises TRH as the only hormone. In some embodiments, the composition comprises TRH as the thyroid hormone. In other embodiments, the composition comprises TRH in combination with one or more additional active agents. In some embodiments, the composition comprises TRH in combination with an additional hormone. In some 2025-10-09 embodiments, the composition comprises TRH in combination with a thyroid hormone.

[0133]

[0118] In another aspect, provided is a composition, useful for treating or preventing hair graying comprising: (i) TRH; (ii) a pharmaceutically acceptable excipient; and (iii) a solvent system. In some embodiments, the composition comprises TRH as the only active ingredient. Hence, also provided is a composition, useful for treating or preventing hair graying comprising: (i) TRH as the only active ingredient; (ii) a pharmaceutically acceptable excipient; and (iii) a solvent system. Also provided is a composition, useful for treating or preventing hair graying, consisting essentially of: (i) TRH; (ii) a pharmaceutically acceptable excipient; and (iii) a solvent system.

[0134]

[0119] In some embodiments, the composition comprises no more than 5 mg / mL, 4 mg / mL, 3 mg / mL, 2 mg / mL, or 25.48 pM of TRH. In some embodiments, the composition comprises no more than 125 pM, 100 pM, 75 pM, 50 pM, 40 pM, or 30 pM of TRH. In some embodiments, the composition does not comprise ethanol. In some embodiments, the composition comprises no more than 50%, 40%, 30%, 20%, or 10% of propylene glycol. In some embodiments, the composition comprises no more than 5%, 4%, 3%, 2%, 1 %, or 0.5% of hydroxypropylcellulose. In some embodiments, the composition does not comprise nitric oxide. In some embodiments, the composition does not comprise a vasodilator. In some embodiments, the composition does not comprise phosphatidylcholine. In some embodiments, the composition does not comprise a lecithin.

[0135]

[0120] In some embodiments, the composition comprises TRH at a concentration of between about 1 pM and 500 pM. In some embodiments, the composition comprises TRH at a concentration of between about 1 pM and 15 pM. In some embodiments, the composition comprises TRH at a concentration of between about 1 pM and 10 pM. In some embodiments, the composition comprises TRH at a concentration of between about 1 pM and 5 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 1 pM, about 1 pM, about 2 pM, about 2.5 pM, about 3 pM, about 4 pM, about 5 pM, about 6 pM, about 7 pM, about 8 pM, about 9 pM, about 10 pM, about 11 pM, about 12 pM, about 13 pM, about 14 pM, about 15 pM, about 20 pM, about 25 pM, about 25.5 pM, about 30 pM, about 40 pM, about 50 pM, or greater than about 50 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 1 pM. In some embodiments, the composition comprises TRH at a concentration of about 1 pM. In some embodiments, the composition comprises TRH at a concentration of about 2 pM. In some embodiments, the composition comprises TRH at a concentration of about 2.5 pM. In some embodiments, the composition comprises TRH at a concentration of about 3 pM. In some embodiments, the composition comprises TRH at a concentration of about 4 pM. In some embodiments, the composition comprises TRH at a concentration of about 5 pM. In some embodiments, the composition comprises TRH at a concentration of about 6 pM. In some embodiments, the composition comprises TRH at a concentration of about 7 pM. In some embodiments, the composition comprises TRH at a concentration of about 8 pM. In some embodiments, the composition comprises TRH at a concentration of 2025-10-09 about 9 pM. In some embodiments, the composition comprises TRH at a concentration of about 10 pM. In some embodiments, the composition comprises TRH at a concentration of about 11 pM. In some embodiments, the composition comprises TRH at a concentration of about 12 pM. In some embodiments, the composition comprises TRH at a concentration of about 13 pM. In some embodiments, the composition comprises TRH at a concentration of about 14 pM. In some embodiments, the composition comprises TRH at a concentration of about 15 pM. In some embodiments, the composition comprises TRH at a concentration of about 20 pM. In some embodiments, the composition comprises TRH at a concentration of about 30 pM. In some embodiments, the composition comprises TRH at a concentration of about 40 pM. In some embodiments, the composition comprises TRH at a concentration of about 50 pM. In some embodiments, the composition comprises TRH at a concentration of greater than about 50 pM.

[0136]

[0121] In some embodiments, the composition comprises TRH at a concentration of between about 50 pM and about 500 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 50 pM, about 60 pM, about 70 pM, about 80 pM, about 90 pM, about 100 pM, about 150 pM, about 200 pM, about 250 pM, about 300 pM, about 350 pM, about 400 pM, about 450 pM, about 500 pM, or greater than about 500 pM, including ranges in between these values. In some embodiments, the composition comprises TRH at a concentration of less than about 50 pM. In some embodiments, the composition comprises TRH at a concentration of about 50 pM. In some embodiments, the composition comprises TRH at a concentration of about 60 pM. In some embodiments, the composition comprises TRH at a concentration of about 70 pM. In some embodiments, the composition comprises TRH at a concentration of about 80 pM. In some embodiments, the composition comprises TRH at a concentration of about 90 pM. In some embodiments, the composition comprises TRH at a concentration of about 100 pM. In some embodiments, the composition comprises TRH at a concentration of about 110 pM. In some embodiments, the composition comprises TRH at a concentration of about 150 pM. In some embodiments, the composition comprises TRH at a concentration of about 200 pM. In some embodiments, the composition comprises TRH at a concentration of about 250 pM. In some embodiments, the composition comprises TRH at a concentration of about 300 pM. In some embodiments, the composition comprises TRH at a concentration of about 350 pM. In some embodiments, the composition comprises TRH at a concentration of about 400 pM. In some embodiments, the composition comprises TRH at a concentration of about 500 pM. In some embodiments, the composition comprises TRH at a concentration of greater than about 500 pM.

[0137]

[0122] In some embodiments, the composition comprises TRH at a concentration of between about 0.05 and 1 pM. In some embodiments, the composition comprises TRH at a concentration of about 0.05 pM. In some embodiments, the composition comprises TRH at a concentration of about 0.1 pM. In some embodiments, the composition comprises TRH at a concentration of about 0.2 pM. In some embodiments, the composition comprises TRH at a concentration of about 0.3 pM. In some embodiments, the 2025-10-09 composition comprises TRH at a concentration of about 0.4 M. In some embodiments, the composition comprises TRH at a concentration of about 0.5 pM. In some embodiments, the composition comprises TRH at a concentration of about 0.6 pM. In some embodiments, the composition comprises TRH at a concentration of about 0.7 pM. In some embodiments, the composition comprises TRH at a concentration of about 0.8 pM. In some embodiments, the composition comprises TRH at a concentration of about 0.9 pM. In some embodiments, the composition comprises TRH at a concentration of about 1 pM.

[0138]

[0123] In some embodiments, the composition comprises TRH at a concentration of less than about 1 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 0.9 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 0.8 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 0.7 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 0.6 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 0.5 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 0.4 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 0.3 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 0.2 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 0.1 pM. In some embodiments, the composition comprises TRH at a concentration of less than about 0.05 pM.

[0139]

[0124] In some embodiments, the composition comprises TRH at a concentration of about 1 nM, 5 nM, 10 nM, 15 nM, 20 nM, 25 nM, 30 nM, 35 nM, 40 nM, 45 nM, or 50 nM, or doses between these values. In some embodiments, the composition comprises TRH at a concentration of about 1 nM. In some embodiments, the composition comprises TRH at a concentration of about 5 nM. In some embodiments, the composition comprises TRH at a concentration of about 10 nM. In some embodiments, the composition comprises TRH at a concentration of about 15 nM. In some embodiments, the composition comprises TRH at a concentration of about 20 nM. In some embodiments, the composition comprises TRH at a concentration of about 25 nM. In some embodiments, the composition comprises TRH at a concentration of about 30 nM. In some embodiments, the composition comprises TRH at a concentration of about 35 nM. In some embodiments, the composition comprises TRH at a concentration of about 40 nM. In some embodiments, the composition comprises TRH at a concentration of about 45 nM. In some embodiments, the composition comprises TRH at a concentration of about 50 nM.

[0140]

[0125] In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of between about 1 :1 and about 1 :1000. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :1 , 1 :2, 1 :3, 1 :4, 1 :5, 1 :6, 1 :7, 1 :8, 1 :9, or 1 :10, including ranges in between these values. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of less than about 1 :1. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :1. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :2. In some 2025-10-09 embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :3. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :4. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :5. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :6. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :7. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :8. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :9. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :10. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of greater than about 1 :10. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of between about 1 :10 and about 1 :1 ,000. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about

[0141] 1 :100. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :200. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :300. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :400. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :500. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :600. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :700. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :800. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :900. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of about 1 :1 ,000. In some embodiments, the composition comprises T3 and TRH at a T3:TRH ratio of greater than about 1 : 1,000.

[0142]

[0126] In some embodiments, the composition comprises T3 at a concentration of about 10 nM and TRH at a concentration of about 25 nM. In some embodiments, the composition comprises T3 at a concentration of about 10 nM and TRH at a concentration of about 250 nM. In some embodiments, the composition comprises T3 at a concentration of about 10 nM and TRH at a concentration of about 2 pM. In some embodiments, the composition comprises T3 at a concentration of about 10 nM and TRH at a concentration of about 25 pM. In some embodiments, the composition comprises T3 at a concentration of about 10 nM and TRH at a concentration of about 250 pM.

[0143]

[0127] In some embodiments, the composition comprises TRH at a concentration of about 50 nM, 60 nM, 70 nM, 80 nM, 90 nM, 100 nM, 110 nM, 120 nM, 130 nM, 140 nM, 150 nM, 160 nM, 170 nM, 180 nM, 190 nM, 200 nM, 210 nM, 220 nM, 230 nM, 240 nM, 250 nM, 260 nM, 270 nM, 280 nM, 290 nM, 300 nM, 310 nM, 320 nM, 330 nM, 340 nM, 350 nM. or doses between these values. In some embodiments, the composition comprises TRH at a concentration of about 50 nM. In some embodiments, the composition comprises TRH at a concentration of about 60 nM. In some embodiments, the composition comprises TRH at a concentration of about 70 nM. In some embodiments, the composition comprises TRH at a 2025-10-09 concentration of about 80 nM. In some embodiments, the composition comprises TRH at a concentration of about 90 nM. In some embodiments, the composition comprises TRH at a concentration of about 100 nM. In some embodiments, the composition comprises TRH at a concentration of about 120 nM. In some embodiments, the composition comprises TRH at a concentration of about 130 nM. In some embodiments, the composition comprises TRH at a concentration of about 140 nM. In some embodiments, the composition comprises TRH at a concentration of about 150 nM. In some embodiments, the composition comprises TRH at a concentration of about 160 nM. In some embodiments, the composition comprises TRH at a concentration of about 170 nM. In some embodiments, the composition comprises TRH at a concentration of about 180 nM. In some embodiments, the composition comprises TRH at a concentration of about 190 nM. In some embodiments, the composition comprises TRH at a concentration of about 200 nM. In some embodiments, the composition comprises TRH at a concentration of about 210 nM. In some embodiments, the composition comprises TRH at a concentration of about 220 nM. In some embodiments, the composition comprises TRH at a concentration of about 230 nM. In some embodiments, the composition comprises TRH at a concentration of about 240 nM. In some embodiments, the composition comprises TRH at a concentration of about 250 nM. In some embodiments, the composition comprises TRH at a concentration of about 260 nM. In some embodiments, the composition comprises TRH at a concentration of about 270 nM. In some embodiments, the composition comprises TRH at a concentration of about 280 nM. In some embodiments, the composition comprises TRH at a concentration of about 290 nM. In some embodiments, the composition comprises TRH at a concentration of about 300 nM. In some embodiments, the composition comprises TRH at a concentration of about 310 nM. In some embodiments, the composition comprises TRH at a concentration of about 320 nM. In some embodiments, the composition comprises TRH at a concentration of about 330 nM. In some embodiments, the composition comprises TRH at a concentration of about 340 nM. In some embodiments, the composition comprises TRH at a concentration of about 350 nM.

[0144]

[0128] In some embodiments, a composition comprises: a. TRH; and b. a solvent.

[0145]

[0129] In some embodiments, a composition comprises: a. TRH; b. an additional active agent; and c. a solvent.

[0146]

[0130] In some embodiments, a composition comprises: a. TRH; and b. isopropanol.

[0147]

[0131] In some embodiments, a composition comprises: 2025-10-09 a. TRH; and b. ethanol.

[0148]

[0132] In some embodiments, a composition comprises: a. TRH; and b. water.

[0149]

[0133] In some embodiments, a composition comprises: a. TRH; and b. propylene glycol.

[0150]

[0134] In some embodiments, a composition comprises: a. TRH; b. isopropanol; and c. water.

[0151]

[0135] In some embodiments, a composition comprises: a. TRH; b. ethanol; and c. water.

[0152]

[0136] In some embodiments, a composition comprises: a. TRH; b. isopropanol; and c. propylene glycol.

[0153]

[0137] In some embodiments, a composition comprises: a. TRH; b. propylene glycol; and c. water.

[0154]

[0138] In some embodiments, a composition comprises: a. TRH; b. isopropanol; c. propylene glycol; and d. water.

[0155]

[0139] In some embodiments, a composition comprises: a. TRH; b. hydroxypropylcellulose; and c. a solvent.

[0156]

[0140] In some embodiments, a composition comprises: a. TRH; 2025-10-09 b. hydroxypropylcellulose; and c. isopropanol.

[0157]

[0141] In some embodiments, a composition comprises: a. TRH; b. hydroxypropylcellulose; and c. water.

[0158]

[0142] In some embodiments, a composition comprises: a. TRH; b. hydroxypropylcellulose; and c. propylene glycol.

[0159]

[0143] In some embodiments, a composition comprises: a. TRH; b. hydroxypropylcellulose; c. isopropanol; and d. water.

[0160]

[0144] In some embodiments, a composition comprises: a. TRH; b. hydroxypropylcellulose; c. isopropanol; and d. propylene glycol.

[0161]

[0145] In some embodiments, a composition comprises: a. TRH; b. hydroxypropylcellulose; c. propylene glycol; and d. water.

[0162]

[0146] In some embodiments, a composition comprises: a. TRH; b. hydroxypropylcellulose; c. isopropanol; d. propylene glycol; and e. water.

[0163]

[0147] In some embodiments, a composition comprises: a. TRH; and b. T3; and c. a solvent. 2025-10-09

[0164]

[0148] In some embodiments, a composition comprises: a. TRH; and b. T3; and c. water.

[0165]

[0149] In some embodiments, a composition comprises: a. TRH; and b. T3; and c. isopropanol.

[0166]

[0150] In some embodiments, a composition comprises: a. TRH; and b. T3; and c. ethanol.

[0167]

[0151] In some embodiments, a composition comprises: a. TRH; and b. T3; and c. propylene glycol.

[0168]

[0152] In some embodiments, a composition comprises: a. TRH; and b. T3; and c. hydroxypropylcellulose.

[0169]

[0153] In some embodiments, the composition further comprises a therapeutically effective amount of an additional active compound. In some embodiments, the additional active agent is selected to provide synergistic effects. In embodiments, “synergistic effects” will be understood to include increases in potency, bioactivity, bioaccessibility, bioavailability, or therapeutic effect, that are greater than the additive contributions of the components acting alone, and / or are greater than the contribution of the isolated compounds on their own. Numerous methods known to those of skill in the art exist to determine whether there is synergy as to a particular effect, i.e. , whether, when two or more components are mixed together, the effect is greater than the sum of the effects of the individual components when applied alone, thereby producing “1+1 > 2.” One such method is the isobologram analysis (or contour method) (Huang et al. 2019).

[0170]

[0154] In embodiments, the additional active agent is selected to provide an additional therapeutic effect, such as antioxidant, anti-inflammatory, analgesic, antinociceptive, immunostimulant, immunosuppressive, anti-cancer, antiemetic, antiulcer, antihistamine, vasodilating, and vasoconstricting effects.

[0171]

[0155] In embodiments, the additional active agent is an amino acid, antioxidant, anti-inflammatory agent, analgesic, 5-alpha reductase inhibitor, potassium channel opener, cannabinoid, immunosuppressant, 2025-10-09 immunostimulant, anti-cancer agent, antiulcer agent, antihistamine, terpene, peptidase inhibitor, thyroid hormone, vitamin, vasodilator, or vasoconstrictor. These active agents may be in ion, freebase, or salt form, include polymorphs, and may be isomers.

[0172]

[0156] In some embodiments, the additional active compound is finasteride. In some embodiments, the additional active compound is dutasteride. In some embodiments, the additional active compound is minoxidil. In some embodiments, the additional active compound is cetirizine. In some embodiments, the additional active compound is levocetirizine. In some embodiments, the additional active compound is latanoprost.

[0173]

[0157] In some embodiments, the additional active compound is a thyroid hormone. In some embodiments, the additional active compound is T3.

[0174]

[0158] In some embodiments, the additional active compound is a peptidase inhibitor. In some embodiments, the peptidase inhibitor is an inhibitor of TRH-degrading ectoenzyme [TRH-DE], In some embodiments, the peptidase inhibitor is a peptide. In some embodiments, the peptidase inhibitor is a tripeptide. In some embodiments, the peptidase inhibitor comprises a tripeptide. In some embodiments, the peptidase inhibitor is Glp-Asn-Pro-AMC or another inhibitor disclosed in U.S. Patent No. 7,713,935.

[0175]

[0159] In another aspect, provided is a composition according to any disclosed embodiment, for use in treating hair loss. In some embodiments, the hair loss is caused by androgenetic alopecia.

[0176]

[0160] In yet another aspect, provided is the use of a composition of disclosed embodiment for treating or preventing hair loss. In some embodiments, the hair loss is caused by androgenetic alopecia.

[0177]

[0161] In some embodiments, disclosed compositions may be administered and dosed in accordance with good medical practice, taking into account the method and scheduling of administration, prior and concomitant medications and medical supplements, the clinical condition of the individual patient and the severity of the underlying disease, the patient’s age, sex, body weight, and other such factors relevant to medical practitioners, and knowledge of the particular compound(s) used. Dosage levels thus may differ from patient to patient, for individual patients across time, and for different compositions and formulations, but shall be able to be determined with ordinary skill.

[0178]

[0162] Determination of appropriate dosing shall include not only the determination of single dosage amounts, but also the determination of the number and timing of doses, and the time(s) of day or time(s) preferable for administration.

[0179]

[0163] The present disclosure further provides kits comprising the disclosed compositions. In some embodiments, the kits provide disclosed compositions in unit dosage form. Kits generally comprise suitable packaging. The kits may comprise one or more containers comprising any composition described herein. Each component (if there is more than one component) can be packaged in separate containers or some components can be combined in one container where cross-reactivity and shelf life permit. The kits may be in unit dosage forms, bulk packages (e.g., multi-dose packages) or sub-unit doses. 2025-10-09

[0180]

[0164] Disclosed kits, for example, may contain sufficient dosages of a disclosed composition for an individual for an extended period, such as any of a week, 2 weeks, 3 weeks, 4 weeks, 6 weeks, 8 weeks, 3 months, 4 months, 5 months, 7 months, 8 months, 9 months, or more. Kits may also include multiple unit doses and instructions for use and be packaged in quantities sufficient for storage at home or a retail location. The kits may optionally include a set of instructions, generally written instructions, although electronic storage media (e.g., magnetic diskette or optical disk) containing instructions are also acceptable, relating to the use of component(s) of the methods of the present disclosure. The instructions included with the kit generally include information as to the components and their administration to an individual.

[0181]

[0165] In some embodiments, a disclosed composition (whether in unit dosage form or not) may be provided in lyophilized form. Lyophilization, also known as freeze-drying, is a process commonly used to preserve and stabilize pharmaceutical compounds and compositions. Lyophilization may result in a dry and shelf-stable product that can subsequently be reconstituted before use. Lyophilization of a disclosed composition may provide several advantages. For example, in some embodiments, a disclosed composition in lyophilized form has improved stability (e.g., improved shelf-stability) and / or reduced susceptibility to chemical, thermal, or biological degradation, as compared to the same composition when not provided in lyophilized form. In some embodiments, a disclosed composition in lyophilized form has reduced weight and / or volume, which may reduce the cost and overall difficulty of transporting, storing, distributing, and using the composition, as compared to the same composition when not provided in lyophilized form.

[0182] C. Methods

[0183]

[0166] In one aspect, provided is a method of treating or preventing hair loss in a subject, the method comprising administering to the subject a therapeutically effective amount of a disclosed composition. In some embodiments, the method comprises administering to the subject a therapeutically effective amount of TRH. In some embodiments, the method comprises administering to the subject a therapeutically effective amount of TRH and T3. In some embodiments, the hair loss is caused by androgenetic alopecia.

[0184]

[0167] In some embodiments, the TRH or combination of TRH and T3 is administered topically, such as in a topical composition. In some embodiments, the TRH is administered topically, such as in a topical composition. In some embodiments, both the TRH and the T3 are administered topically (i.e., the combination is administered topically, such as in one or more topical compositions).

[0185]

[0168] In some embodiments, the TRH and T3 are administered simultaneously. The simultaneous administration of the TRH and T3 can be achieved, for example, by administering a composition (e.g., a disclosed topical composition) that contains both TRH and T3. In another embodiment, the simultaneous administration of the TRH and T3 can be achieved, for example, by simultaneous administration of separate TRH and T3 compositions. 2025-10-09

[0186]

[0169] In some embodiments, the TRH is administered at a concentration of between about 1 pM and 500 pM. In some embodiments, the TRH is administered at a concentration of between about 1 pM and 50 pM. In some embodiments, the TRH is administered at a concentration of between about 1 pM and 15 pM. In some embodiments, the TRH is administered at a concentration of between about 1 pM and 10 pM. In some embodiments, the TRH is administered at a concentration of between about 1 pM and 5 pM. In some embodiments, the TRH is administered at a concentration of less than about 1 pM, about 1 pM, about 2 pM, about 2.5 pM, about 3 pM, about 4 pM, about 5 pM, about 6 pM, about 7 pM, about 8 pM, about 9 pM, about 10 pM, about 11 pM, about 12 pM, about 13 pM, about 14 pM, about 15 pM, about 20 pM, about 25 pM, about 25.5 pM, about 30 pM, about 40 pM, about 50 pM, or greater than about 50 pM, including ranges in between these values. In some embodiments, the TRH is administered at a concentration of less than about 1 pM. In some embodiments, the TRH is administered at a concentration of about 1 pM. In some embodiments, the TRH is administered at a concentration of about 2 pM. In some embodiments, the TRH is administered at a concentration of about 2.5 pM. In some embodiments, the TRH is administered at a concentration of about 3 pM. In some embodiments, the TRH is administered at a concentration of about 4 pM. In some embodiments, the TRH is administered at a concentration of about 5 pM. In some embodiments, the TRH is administered at a concentration of about 6 pM. In some embodiments, the TRH is administered at a concentration of about 7 pM. In some embodiments, the TRH is administered at a concentration of about 8 pM. In some embodiments, the TRH is administered at a concentration of about 9 pM. In some embodiments, the TRH is administered at a concentration of about 10 pM. In some embodiments, the TRH is administered at a concentration of about 11 pM. In some embodiments, the TRH is administered at a concentration of about 12 pM. In some embodiments, the TRH is administered at a concentration of about 13 pM. In some embodiments, the TRH is administered at a concentration of about 14 pM. In some embodiments, the TRH is administered at a concentration of about 15 pM. In some embodiments, the TRH is administered at a concentration of about 20 pM. In some embodiments, the TRH is administered at a concentration of about 30 pM. In some embodiments, the TRH is administered at a concentration of about 40 pM. In some embodiments, the TRH is administered at a concentration of about 50 pM. In some embodiments, the TRH is administered at a concentration of greater than about 50 pM.

[0187]

[0170] In some embodiments, the TRH is administered at a concentration of between about 50 pM and 500 pM. In some embodiments, the TRH is administered at a concentration of less than about 50 pM, about 60 pM, about 70 pM, about 80 pM, about 90 pM, about 100 pM, about 150 pM, about 200 pM, about 250 pM, about 300 pM, about 350 pM, about 400 pM, about 450 pM, about 500 pM, or greater than about 500 pM, including ranges in between these values. In some embodiments, the TRH is administered at a concentration of less than about 50 pM. In some embodiments, the TRH is administered at a concentration of about 50 pM. In some embodiments, the TRH is administered at a concentration of 2025-10-09 about 60 pM. In some embodiments, the TRH is administered at a concentration of about 70 pM. In some embodiments, the TRH is administered at a concentration of about 80 pM. In some embodiments, the TRH is administered at a concentration of about 90 pM. In some embodiments, the TRH is administered at a concentration of about 100 pM. In some embodiments, the TRH is administered at a concentration of about 110 pM. In some embodiments, the TRH is administered at a concentration of about 150 pM. In some embodiments, the TRH is administered at a concentration of about 200 pM. In some embodiments, the TRH is administered at a concentration of about 250 pM. In some embodiments, the TRH is administered at a concentration of about 300 pM. In some embodiments, the TRH is administered at a concentration of about 350 pM. In some embodiments, the TRH is administered at a concentration of about 400 pM. In some embodiments, the TRH is administered at a concentration of about 500 pM. In some embodiments, the TRH is administered at a concentration of greater than about 500 pM.

[0188]

[0171] In some embodiments, the TRH is administered at a concentration of between about 0.05 and 1 pM. In some embodiments, the TRH is administered at a concentration of about 0.05 pM. In some embodiments, the TRH is administered at a concentration of about 0.1 pM. In some embodiments, the TRH is administered at a concentration of about 0.2 pM. In some embodiments, the TRH is administered at a concentration of about 0.3 pM. In some embodiments, the TRH is administered at a concentration of about 0.4 pM. In some embodiments, the TRH is administered at a concentration of about 0.5 pM. In some embodiments, the TRH is administered at a concentration of about 0.6 pM. In some embodiments, the TRH is administered at a concentration of about 0.7 pM. In some embodiments, the TRH is administered at a concentration of about 0.8 pM. In some embodiments, the TRH is administered at a concentration of about 0.9 pM.

[0189]

[0172] In some embodiments, the TRH is administered at a concentration of about 1 nM, 5 nM, 10 nM, 15 nM, 20 nM, 25 nM, 30 nM, 35 nM, 40 nM, 45 nM, or 50 nM, or doses between these values. In some embodiments, the TRH is administered at a concentration of about 1 nM. In some embodiments, the TRH is administered at a concentration of about 5 nM. In some embodiments, the TRH is administered at a concentration of about 10 nM. In some embodiments, the TRH is administered at a concentration of about 15 nM. In some embodiments, the TRH is administered at a concentration of about 20 nM. In some embodiments, the TRH is administered at a concentration of about 25 nM. In some embodiments, the TRH is administered at a concentration of about 30 nM. In some embodiments, the TRH is administered at a concentration of about 35 nM. In some embodiments, the TRH is administered at a concentration of about 40 nM. In some embodiments, the TRH is administered at a concentration of about 45 nM. In some embodiments, the TRH is administered at a concentration of about 50 nM.

[0190]

[0173] In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of between about 1 :1 and about 1 :1000. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :1 , 1 :2, 1 :3, 1 :4, 1 :5, 1 :6, 1 :7, 1 :8, 1 :9, or 1 :10, including ranges in between these values. In some 2025-10-09 embodiments, the T3 and TRH are administered at a T3:TRH ratio of less than about 1 :1. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :1. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :2. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :3. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :4. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :5. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :6. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :7. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :8. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :9. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :10. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of greater than about 1 :10. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of between about 1 :10 and about 1 :1 ,000. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :100. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 : 200. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :300. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :400. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :500. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :600. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :700. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :800. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :900. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of about 1 :1 ,000. In some embodiments, the T3 and TRH are administered at a T3:TRH ratio of greater than about 1 : 1,000.

[0191]

[0174] In some embodiments, the T3 is administered at a concentration of about 10 nM and TRH at a concentration of about 25 nM. In some embodiments, the T3 is administered at a concentration of about 10 nM and TRH at a concentration of about 250 nM. In some embodiments, the T3 is administered at a concentration of about 10 nM and TRH at a concentration of about 2 pM. In some embodiments, the T3 is administered at a concentration of about 10 nM and TRH at a concentration of about 25 pM. In some embodiments, the T3 is administered at a concentration of about 10 nM and TRH at a concentration of about 250 pM.

[0192]

[0175] In some embodiments, the total unit dose volume of a disclosed composition is between about 0.1 mL and 10 mL. In some embodiments, the total unit dose volume is about 0.1 mL, about 0.5 mL, about 1 mL, about 2 mL, about 3 mL, about 4 mL, about 5 mL, about 6 mL, about 7 mL, about 8 mL, about 9 mL, or about 10 mL. In some embodiments, the total unit dose volume is about 0.1 mL. In some embodiments, the total dose volume is about 0.5 mL. In some embodiments, the total unit dose volume is about 1 mL. In some embodiments, the total unit dose volume is about 2 mL. In some embodiments, the total dose 2025-10-09 volume of a disclosed composition is about 3 mL. In some embodiments, the total unit dose volume is about 4 mL. In some embodiments, the total unit dose volume is about 5 mL. In some embodiments, the total unit dose volume is about 6 mL. In some embodiments, the total unit dose volume is about 7 mL. In some embodiments, the total unit dose volume is about 8 mL. In some embodiments, the total dose volume of a disclosed composition is about 9 mL. In some embodiments, the total unit dose volume is about 10 mL.

[0193]

[0176] In some embodiments, disclosed methods are performed in accordance with good medical practice, taking into account the method and scheduling of administration, prior and concomitant medications and medical supplements, the clinical condition of the individual patient and the severity of the underlying disease, the patient’s age, sex, body weight, and other such factors relevant to medical practitioners, and knowledge of the particular compound(s) used. Dosage levels thus may differ from patient to patient, for individual patients across time, and for different compositions and formulations, but shall be able to be determined with ordinary skill.

[0194]

[0177] In some embodiments, the composition is administered every day. In some embodiments, the composition is administered every other day for a period of time, followed by a prolonged period without administration. For example, in some embodiments, the composition is administered every other day for 1 week followed by a prolonged period without administration. In some embodiments, the composition is administered every other day for 2 weeks followed by a prolonged period without administration. In some embodiments, the composition is administered every other day for 3 weeks followed by a prolonged period without administration. In some embodiments, the composition is administered every other day for 4 weeks followed by a prolonged period without administration. In some embodiments, the composition is administered every other day for 5 weeks followed by a prolonged period without administration. In some embodiments, the composition is administered every other day for 6 weeks followed by a prolonged period without administration.

[0195]

[0178] In some embodiments, the prolonged period without administration lasts for one day. In some embodiments, the prolonged period without administration lasts for 2 days. In some embodiments, the prolonged period without administration lasts for 3 days. In some embodiments, the prolonged period without administration lasts for 4 days. In some embodiments, the prolonged period without administration lasts for 5 days. In some embodiments, the prolonged period without administration lasts for 6 days. In some embodiments, the prolonged period without administration lasts for 7 days. In some embodiments, the prolonged period without administration lasts for 10 days. In some embodiments, the prolonged period without administration lasts for 14 days. In some embodiments, the prolonged period without administration lasts for 30 days.

[0196]

[0179] In some embodiments, the composition is administered every day for several consecutive days followed by a prolonged period without administration. For example, in some embodiments, the 2025-10-09 composition is administered every day for 2 consecutive days followed by a prolonged period without administration. In some embodiments, the composition is administered every day for 3 consecutive days followed by a prolonged period without administration. In some embodiments, the composition is administered every day for 4 consecutive days followed by a prolonged period without administration. In some embodiments, the composition is administered every day for 5 consecutive days followed by a prolonged period without administration. In some embodiments, the composition is administered every day for 6 consecutive days followed by a prolonged period without administration. In some embodiments, the composition is administered every day for 7 consecutive days followed by a prolonged period without administration. In other embodiments, the composition is administered for only one day followed by a prolonged period without administration.

[0197]

[0180] In some embodiments, the prolonged period without administration lasts for one day. In some embodiments, the prolonged period without administration lasts for 2 days. In some embodiments, the prolonged period without administration lasts for 3 days. In some embodiments, the prolonged period without administration lasts for 4 days. In some embodiments, the prolonged period without administration lasts for 5 days. In some embodiments, the prolonged period without administration lasts for 6 days. In some embodiments, the prolonged period without administration lasts for 7 days. In some embodiments, the prolonged period without administration lasts for 10 days. In some embodiments, the prolonged period without administration lasts for 14 days. In some embodiments, the prolonged period without administration lasts for 30 days.

[0198]

[0181] In some embodiments, the TRH is administered in a topical composition. In some embodiments, a disclosed method comprises administering a combination of TRH and T3. In some embodiments, the TRH and T3 are in the same composition. In other embodiments, the TRH and T3 are in different compositions.

[0199]

[0182] It will be appreciated that the frequency or duration of a disclosed method may be increased or reduced, as indicated by the clinical outcome desired, status of the pathology or symptom, any adverse side effects of the treatment or therapy, or concomitant medications. In some embodiments, the subject is subjected to a disclosed method every day for several consecutive days followed by a prolonged period without administration. This is referred to herein as “pulsed dosing” or “pulsed therapy.” In some embodiments, the subject is subjected to a disclosed method every day for 2 consecutive days followed by a prolonged period without administration. In some embodiments, the subject is subjected to a disclosed method every day for 3 consecutive days followed by a prolonged period without administration. In some embodiments, the subject is subjected to a disclosed method every day for 4 consecutive days followed by a prolonged period without administration. In some embodiments, the subject is subjected to a disclosed method every day for 5 consecutive days followed by a prolonged period without administration. In some embodiments, the subject is subjected to a disclosed method every day for 6 consecutive days followed by a prolonged period without administration. In some embodiments, the subject is subjected to a disclosed 2025-10-09 method every day for 7 consecutive days followed by a prolonged period without administration. In some embodiments, the subject is subjected to a disclosed method every day for 10 consecutive days followed by a prolonged period without administration. In some embodiments, the subject is subjected to a disclosed method every day for 14 consecutive days followed by a prolonged period without administration. In some embodiments, the subject is subjected to a disclosed method for only one day followed by a prolonged period without administration.

[0200]

[0183] In some embodiments, the prolonged period without administration lasts one day. In some embodiments, the prolonged period without administration lasts 2 days. In some embodiments, the prolonged period without administration lasts 3 days. In some embodiments, the prolonged period without administration lasts 4 days. In some embodiments, the prolonged period without administration lasts 5 days. In some embodiments, the prolonged period without administration lasts 6 days. In some embodiments, the prolonged period without administration lasts 7 days. In some embodiments, the prolonged period without administration lasts 10 days. In some embodiments, the prolonged period without administration lasts 14 days. In some embodiments, the prolonged period without administration lasts 30 days.

[0201]

[0184] In some embodiments, the amount of TRH administered during a single administration of a disclosed composition (i.e., per unit dose) is between about 0.00001 pg and about 200 pg, or between about 0.00003 pg and about 200 pg. In some embodiments, the amount of TRH administered per unit dose is between about 0.00003 pg and about 0.001 pg. In some embodiments, the amount of TRH administered per unit dose is between about 0.001 pg and about 0.01 pg. In some embodiments, the amount of TRH administered per unit dose is between about 0.01 pg and about 0.1 pg. In some embodiments, the amount of TRH administered per unit dose is between about 0.1 pg and about 1 pg. In some embodiments, the amount of TRH administered per unit dose is between about 1 pg and about 10 pg. In some embodiments, the amount of TRH administered per unit dose is between about 10 pg and about 100 pg. In some embodiments, the amount of TRH administered per unit dose is between about 100 pg and about 200 pg.

[0202]

[0185] In some embodiments, the amount of TRH administered per unit dose is about 30 pg, 50 pg, 100 pg, 200 pg, 300 pg, 500 pg, 0.001 pg, 0.002 pg, 0.005 pg, 0.01 pg, 0.02 pg, 0.05 pg, 0.1 pg, 0.2 pg, 0.5 pg, 1 pg, 2 pg, 5 pg, 10 pg, 20 pg, 50 pg, 100 pg, 150 pg, or 200 pg. In some embodiments, the amount of TRH administered per unit dose is greater than about 200 pg (e.g., 250 pg, 300 pg, 350 pg, 400 pg, 450 pg, 500 pg, or greater).

[0203]

[0186] In some embodiments, the amount of TRH administered per unit dose is between about 1 ng and 20 ng. In some embodiments, the amount of TRH administered per unit dose is between about 1 ng and 10 ng. In some embodiments, the amount of TRH administered per unit dose is between about 5 ng and 10 ng. In some embodiments, the amount of TRH administered per unit dose is about 1 ng, 2 ng, 3 ng, 4 2025-10-09 ng, 5 ng, 6 ng, 7 ng, 8 ng, 9 ng, or 10 ng. In some embodiments, the amount of TRH administered per unit dose is about 6.5 ng.

[0204]

[0187] In some embodiments, the amount of TRH administered per unit dose is between about 0.1 ng and 1 ng. In some embodiments, the amount of TRH administered per unit dose is about 0.1 ng, 0.2 ng, 0.3 ng, 0.4 ng, 0.5 ng, 0.6 ng, 0.7 ng, 0.8 ng, 0.9 ng, or 1.0 ng. In some embodiments, the amount of TRH administered per unit dose is about 0.65 ng.

[0205]

[0188] In some embodiments, disclosed compositions are formulated as a unit dosage form, each dosage containing an effective amount of the active ingredient(s), for example in the dosage amounts disclosed above. The term “unit dosage form” refers to a physically discrete unit suited as unitary dosages to be consumed by the individual, each unit containing a predetermined quantity of active material calculated to produce the desired effect(s). Unit dosage forms are often used for ease of administration and uniformity of dosage. Unit dosage forms can contain a single or individual dose or unit, a sub-dose, or an appropriate fraction thereof, of the composition.

[0206]

[0189] In embodiments wherein a disclosed composition is used to create a desired effect, it will be readily appreciated that dose and dosage may vary depending upon the general health, age, gender, and race of the individual, bioavailability, potential adverse systemic, regional, or local side effects, the presence of any disorders or diseases in the individual, and other factors that will be appreciated by those in the art (e.g., medical or familial history).

[0207]

[0190] In general, dose amount, frequency, or duration may be increased or reduced, as indicated by the therapeutic outcome(s) or effect(s) desired, the beneficial outcome(s) or effect(s) desired, and / or by the specific subjective outcome(s) or effect(s) desired.

[0208]

[0191] Those in the art will appreciate the factors that may influence the dosage, frequency, and timing required to provide an amount sufficient or effective for providing a desired effect, and to do so depending on the type of desired effect and to avoid or minimize adverse effects.

[0209]

[0192] Dosage levels may differ from patient to patient, for individuals across time, and for different compositions and formulations, but shall be able to be determined with ordinary skill. Determination of appropriate dosing shall include not only the determination of single dosage amounts, but also the determination of the number and timing of doses, and the time(s) of day or time(s) preferable for administration.

[0210] D. Excipients

[0211]

[0193] In some embodiments, a disclosed composition is formulated for topical administration (e.g., as a topical dosage form), for example through the use of one or more pharmaceutically acceptable excipients. Topical dosage forms include transmucosal and transdermal formulations, such as aerosols, emulsions, sprays, ointments, salves, gels, pastes, lotions, liniments, oils, and creams. Pharmaceutically acceptable excipients include, for example, penetration enhancers, carriers, diluents, emulsifiers, stabilizers, solvents 2025-10-09 and cosolvents, viscosity modifying agents (e.g., thickeners), adhesion modifying agents (e.g., tackifiers), preservatives, antioxidants, adhesive polymers, solubilizing agents, colorants, binders, humectants, surfactants, gelling agents, and other such ingredients as will be generally known to one of skill in the art.

[0212]

[0194] Pharmaceutical formulations may be prepared as liquid suspensions or solutions using a sterile liquid, such as but not limited to, an oil, water, an alcohol, and combinations of these pharmaceutically suitable surfactants, suspending agents, and emulsifying agents. Suspensions may include oils. Such oils include peanut oil, sesame oil, cottonseed oil, corn oil, and olive oil. Suitable oils also include carrier oils such as MCT and long chain triglyceride (LCT) oils. Suspension preparation may also contain esters of fatty acids such as ethyl oleate, isopropyl myristate, fatty acid glycerides, and acetylated fatty acid glycerides. Suspension formulations may include alcohols, (such as ethanol, isopropyl alcohol, hexadecyl alcohol), glycerol, and propylene glycol. Ethers, such as polyethylene glycol), petroleum hydrocarbons such as mineral oil and petrolatum, and water may also be used in suspension formulations. A suspension can thus include an aqueous liquid or a non-aqueous liquid, an oil-in-water liquid emulsion, or a water-in-oil emulsion.

[0213]

[0195] “Pharmaceutically acceptable” as used in connection with an excipient or other ingredient means that the ingredient is generally safe and, within the scope of sound medical judgment, suitable for use in contact with the cells of humans and other animals without undue toxicity, irritation, allergic response, or complication, and commensurate with a reasonable risk / benefit ratio. In some embodiments, “pharmaceutically acceptable” means that a particular ingredient has been approved by the FDA for topical use in cosmetic products.

[0214]

[0196] In some embodiments, the composition comprises a penetration enhancer. Without being bound by theory, penetration enhancers are generally characterized by their ability to increase the permeability of biological barriers, such as scalp skin. In some embodiments, including a penetration enhancer in the composition increases the bioavailability of the active agent(s) (e.g., TRH and / or any additional active ingredients) by improving the ability of the active agent(s) to diffuse into the skin tissue. Penetration enhancers include, for example, include fatty acids and oils such as castor oil, coconut oil, medium chain triglycerides (MCT), jojoba oil, sunflower oil, argan oil, almond oil, olive oil, mineral oil, petroleum jelly, cocoa butter, shea butter, or other esters, triglycerides, or functional derivatives thereof. In some embodiments, the penetration enhancer is 1 ,2-lauryl ether, aprotinin, azone, benzalkonium chloride, benzalkonium bromide, cetylpyridinium chloride, cetyltrimethyl ammonium, cyclodextrin, dextran sulfate, glycol, propylene glycol, lauric acid, lysophosphatidylcholine, menthol, phosphatidylcholine, polyoxyethylene, polysorbate 80, sodium EDTA, chitosan, sodium glycocholate, sodium deoxyglycocholate, sodium lauryl sulfate, sodium salicylate, sodium taurocholate, dimethyl sulfoxide, or a combination thereof. In some embodiments, the penetration enhancer is selected from the group consisting of lower chain (C1-C5) alcohols, sodium glycocholate, sodium deoxycholate, sodium 2025-10-09 taurocholate, sodium glycodeoxycholate, sodium taurodeoxycholate, oleic acid, capric acid, lauric acid, lecithin, myristic acid, palmitic acid, lysophosphatidylcholine, phosphatidylcholine, azone, cyclodextrin, sodium lauryl sulfate, polyoxyethylene-9-lauryl ether, polyoxyethylene-20-cetyl ether, benzalkonium chloride, cetylpyridinium chloride, Vitamin E TPGS, caprylocaproyl polyoxylglycerides, stearoyl macrogolglycerides, propylene glycol dicaprylocaprate, and mixtures thereof. In some embodiments, compositions of the disclosure may comprise a penetration enhancer at a concentration of about 0.01%, about 0.02%, about 0.05%, about 0.1%, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11 %, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21 %, about 22%, about 23%, about

[0215] 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 61%, about 62%, about 63%, about 64%, about

[0216] 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 75%, and about 80% of the composition, on a weight or volume basis.

[0217]

[0197] In some embodiments, the base may further include a conditioning agent to prevent drying of the skin and hair in combination with the active agent(s) (e.g., TRH and / or any additional active ingredients). Representative conditioning agents may include, but are not limited to, glycerin, propylene glycol, alpha hydroxyl acids, urea, lactic acid, oils, lanolin and silicone and its derivatives. In some embodiments, conditioning agents are physically and chemically compatible with the essential components of the composition, and do not otherwise unduly impair product stability, aesthetics or performance. In some embodiments, the concentration of the conditioning agent in the composition is sufficient to provide the desired conditioning benefits, as will be apparent to one of ordinary skill in the art. The concentration may vary with the conditioning agent, the conditioning performance desired, the average size of the conditioning agent particles, the type and concentration of other components, and other like factors.

[0218]

[0198] In some embodiments, the composition comprises a carrier. Carriers can be designed to give controlled release profiles, improved circulation times and better penetration across the epithelium. In some embodiments, the carrier is a hydrophobic drug carrier. Hydrophobic drug carriers can have the advantage of exhibiting slow sustained release and may adhere well to biological surfaces. Hydrophobic drug carriers can have slow (i.e., extended) release kinetics, or may also be constructed to have a rapid or immediate release profile. New techniques include the development of hydrophilic coatings on hydrophobic nanoparticles to improve their transport across tissue surfaces while retaining the slow-release profiles. These include polyethylene glycol and chitosan coatings. (See, e.g., de la Fuente, et al. Nanomedicine 2008;3:845-857.) Any of a variety of pharmaceutically acceptable carriers may be used including, without limitation, aqueous media such as water, saline, glycine, hyaluronic acid and the like; solid carriers such as starch, magnesium stearate, mannitol, sodium saccharin, talcum, cellulose, glucose, sucrose, lactose, 2025-10-09 trehalose, magnesium carbonate, and the like; solvents; dispersion media; coatings; antibacterial and antifungal agents; isotonic and absorption delaying agents; or any other inactive ingredient. Selection of a pharmacologically acceptable carrier can depend on the mode of administration. Non-limiting examples of specific uses of such pharmaceutical carriers can be found in Pharmaceutical Dosage Forms and Drug Delivery Systems (Howard C. Ansel et al., eds., Lippincott Williams & Wilkins Publishers, 7th ed. 1999); Remington: The Science and Practice of Pharmacy (Alfonso R. Gennaro ed., Lippincott, Williams & Wilkins, 20th ed. 2000); Goodman & Gilman's The Pharmacological Basis of Therapeutics (Joel G. Hardman et al., eds., McGraw-Hill Professional, 10th ed. 2001); and Handbook of Pharmaceutical Excipients (Raymond C. Rowe et al., APhA Publications, 4th edition 2003). In some embodiments, compositions of the disclosure may comprise a carrier at a concentration of about 0.01 %, about 0.02%, about 0.05%, about 0.1 %, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21 %, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 35%, about 40%, about 45%, about

[0219] 50%, about 55%, about 60%, about 61 %, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 75%, and about 80% of the composition, on a weight or volume basis.

[0220]

[0199] In some embodiments, the composition comprises an emulsifier. The emulsifier may be an anionic, cationic, or neutral emulsifier. In certain embodiments, the emulsifier is an anionic emulsifier selected from the group consisting of alkyl sulfate, aralkyl sulfates, alkyl ethoxy ether sulfates, alkaryl sulphonates, alkyl succinates, alkyl sulfosuccinates, N-alkoyl sarconsinates, isethionates, N-acyl taurate, sodium lauryl sulfate, sodium laureth sulfate, sodium oleyl succinate, sodium dodecylbenzenesulfonate, and sodium lauryl sarconsinate. Exemplary non-ionic or neutral emulsifiers include sorbitan ester, ethoxylated sorbitan ester, ethoxylated alkyl ether, ethoxylated fatty acid ether, fatty alcohol, ethoxylated fatty alcohol, and esters of glycerin and fatty acids. In certain embodiments, the emulsifiers are synthetic or natural polymers. In certain embodiments, the emulsifier includes silicon. In certain embodiments, the emulsifier is a silicone (e.g. dimethicone, phenyltrimethicone, PEG dimethicone, PPG dimethicone, etc.). In some embodiments, compositions of the disclosure may comprise an emulsifier at a concentration of about 0.01 %, about 0.02%, about 0.05%, about 0.1 %, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1 %, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11 %, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about

[0221] 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 61 %, about 62%, about 63%, about 64%, about 65%, about 2025-10-09 66%, about 67%, about 68%, about 69%, about 70%, about 75%, about 75%, and about 80% of the composition, on a weight or volume basis. In some embodiments, the composition does not comprise an emulsifier.

[0222]

[0200] In some embodiments, the composition comprises an anti-dandruff agent or or other ingredients which are commonly applied to the scalp or hair, including antimicrobial agents, where desirable, generally in amounts found useful in topical applications. One of ordinary skill in the art can easily determine the type and amount of anti-dandruff agent chosen for use in formulations according to the present invention.

[0223]

[0201] In some embodiments, the composition comprises an antioxidant. The antioxidant may be amino acids (e.g. glycine, histidine, tyrosine, tryptophan) and derivatives thereof, imidazoles (e.g. urocanic acid) and derivatives thereof peptides, such as D,L-carnosine, D-carnosine, L-carnosine and derivatives thereof (e.g. anserine), carotenoids, carotenes (e.g. p-carotene, lycopene) and derivatives thereof, chlorogenic acid and derivatives thereof, liponic acid and derivatives thereof (e.g. dihydroliponic acid), aurothioglucose, propylthiouracil and other thiols (e.g. thiorodoxin, glutathione, cysteine, cystine, cystamine and the glycosyl, N-acetyl, methyl, ethyl, propyl, amyl, butyl, and lauryl, palmitoyl, oleyl, y-linoleyl, cholesteryl and glyceryl esters thereof) and salts thereof, dilauryl thiodipropionate, distearyl thiodipropionate, thiodipropionic acid and derivatives thereof (esters, ethers, peptides, lipids, nucleotides, nucleosides and salts) and sulfoximine compounds (e.g. buthionine sulfoximines, homocysteine sulfoximines, buthionine sulfones, penta, hexa and heptathionine sulfoximine), in very low tolerated doses (e.g. pmol to pmol / kg), and furthermore (metal)chelators (e.g. a-hydroxy-fatty acids, palmitic acid, phytic acid, lactoferrin), a-hydroxy acids (e.g. citric acid, lactic acid, malic acid), humic acid, gallic acid, bile extracts, bilirubin, biliverdin, EDTA and derivatives thereof, unsaturated fatty acids and derivatives thereof (e.g. y-linolenic acid, linoleic acid, oleic acid), folic acid and derivatives thereof, ubiquinone and ubiquinol and derivatives thereof vitamin C and derivatives thereof (e.g. sodium ascorbate, ascorbyl palmitate, magnesium ascorbyl phosphate, ascorbyl acetate), tocopherol and derivatives (e.g. vitamin E acetate, tocotrienol), vitamin A and derivatives (vitamin A palmitate) and coniferyl benzoate of benzoic resin, rutinic acid and derivatives thereof, a-glycosylrutin, ferulaic acid, furfurylideneglucitol, carnosine, butylhydroxytoluene, butylhydroxyanisole, nordihydroguajak resin acid, nordihydroguaiaretic acid, trihydroxybutyrophenone, uric acid and derivatives thereof, mannose and derivatives thereof, zinc and derivatives thereof (e.g. ZnO, ZnSO4), selenium and derivatives thereof (e.g. selenium methionine), stilbenes and derivatives thereof (e.g. stilbene oxide, trans-stilbene oxide).

[0224]

[0202] In embodiments, the composition comprises a vitamin. The vitamin may be riboflavin (vitamin B2), niacinamide (vitamin B3), pantothenic acid (vitamin B5), pyridoxine (vitamin B6), biotin (vitamin B7).

[0225]

[0203] In some embodiments, the composition comprises a conventional thickener. The conventional thickener may be crosslinked polyacrylic acids and derivatives thereof, polysaccharides and derivatives thereof, such as xanthan gum, agar agar, alginates or tyloses, cellulose derivatives (e.g. 2025-10-09 carboxymethylcellulose or hydroxycarboxymethylcellulose), fatty alcohols, monoglycerides and fatty acids, polyvinyl alcohol and polyvinylpyrrolidone. Nonionic thickeners are preferably used.

[0226]

[0204] In some embodiments, the composition comprises a cosmetically and / or dermo-cosmetically active substance. A cosmetically and / or dermo-cosmetically active substance may be color-imparting active substances, skin- and hair-pigmenting compositions, tinting compositions, tanning compositions, bleaches, keratin-hardening substances, antimicrobial active substances, light filter active substances, repellent active substances, substances having hyperemic activity, substances having keratolytic and keratoplastic activity, antidandruff active substances, antiphlogistic agents, substances having keratinizing activity, antioxidant active substances or substances active as free radical scavengers, skin-moisturizing substances or skin humectants, refatting active substances, substances having antierythematous or antiallergic activity, branched fatty acids, such as 18-methyleicosanoic acid, and mixtures thereof.

[0227]

[0205] In embodiments, the composition comprises a perfume oil. Natural fragrances are extracts of blossoms (lily, lavender, rose, jasmine, neroli, ylang-ylang), stalks and leaves (geranium, patchouli, petitgrain), fruits (anise, coriander, caraway, juniper), fruit peels (bergamot, lemon, orange), roots (mace, angelica, celery, cardamom, costus, iris, calmus), woods (pinewood, sandalwood, guajak wood, cedar wood, rosewood), herbs and grasses (tarragon, lemongrass, sage, thyme), needles and branches (spruce, fir, pine, dwarf pine), resins and balsams (galbanum, elemi, benzoin, myrrh, olibanum, opoponax). Typical synthetic fragrance compounds are products of the type consisting of the esters, ethers, aldehydes, ketones, alcohols and hydrocarbons. Essential oils of low volatility, which are generally used as aroma components, are also suitable as perfume oils, e.g. sage oil, chamomile oil, clove oil, balm oil, mint oil, cinnamon leaf oil, lime tree blossom oil, juniper oil, vetiver oil, oliban oil, galbanum oil, labolanum oil and lavandin oil. Bergamot oil, dihydromyrcenol, lilial, lyral, citronellol, phenylethyl alcohol, a-hexylcinnamaldehyde, geraniol, benzylacetone, cyclamenaldehyde, linalool, Boisambrene@Forte, ambroxan, indole, hedione, sandelice, lemon oil, mandarin oil, orange oil, allylamyl glycolate, cyclovertal, lavandin oil, muscatel sage oil, G39 damascene, Bourbon geranium oil, cyclohexyl salicylate, Vertofix@Coeur, iso-E-Super®, Fixolide®NP, evemyl, iraldein gamma, phenylacetic acid, geranyl acetate, benzyl acetate, rose oxide, romillate, irotyl and floramat.

[0228]

[0206] In some embodiments, the composition comprises a hydroxyalkyl cellulose. Hydroxyalkyl celluloses can have multiple functions when included as an excipient. For example, a hydroxyalkyl cellulose may act as any of a penetration enhancer, carrier, emulsifier, stabilizer, viscosity modifying agent, adhesion modifying agent, antioxidant, adhesive polymer, solubilizing agent, binder, humectant, and / or gelling agent. In some embodiments, the composition comprises hydroxymethylcellulose. In some embodiments, the composition comprises hydroxyethylcellulose. In some embodiments, the composition comprises hydroxypropylcellulose.

[0229]

[0207] In some embodiments, the composition comprises a solvent, and optionally a cosolvent. Any 2025-10-09 solvent(s) and cosolvent(s) may be collectively referred to as a “solvent system.” Without being bound by theory, the solvent system chosen can affect the stability, bioavailability, and overall efficacy of the composition. In some embodiments, the solvent system is capable of dissolving or solubilizing the active ingredients and any included excipients at the desired concentration(s), and should be stable and compatible with components (e.g., TRH, any additional active agent(s), and any other excipients) in the composition. In some embodiments, wherein the solvent system comprises more than one solvent, the ratio of cosolvents is optimized, for example to increase the penetration or bioavailability of an active ingredient. Preferred solvent systems are also safe and non-toxic for human consumption. In some embodiments, potential adverse effects, such as irritation or allergic reactions, are considered and minimized during selection of solvents included in a solvent system of the disclosure. Solvents that may be included in disclosed compositions may include, without limitations, water, ethanol, polyhydric alcohols (e.g., glycerin), 1,3-butylene glycol, propylene glycol, hexylene glycol, propane diol, ethylene glycol, diethylene glycol, dipropylene glycol, diglycerin, sorbitol, other sugars which are liquid at room temperature, water-soluble alkoxylated nonionic polymers such as polyethylene glycol, and combinations thereof. Solvents may be present, individually or in total (if more than one solvent is included), in the composition in an amount ranging from about 0.1 wt% to about 95 wt% (calculated as the total weight of solvents in the composition divided by the total weight of the composition).

[0230]

[0208] In some embodiments, the solvent system is an aqueous solvent system. In some embodiments, the solvent system comprises water. In some embodiments, the composition comprises between about 0.1 % (v / v) and 90% (v / v) of water. In some embodiments, a composition comprises between about 0.1 % (v / v) and 1 % (v / v) of water. In some embodiments, a composition comprises between about 0.1 % (v / v) and 10% (v / v) of water. In some embodiments, a composition comprises between about 1 % (v / v) and 90% (v / v) of water. In some embodiments, a composition comprises between about 1% (v / v) and 50% (v / v) of water. In some embodiments, a composition comprises between about 1 % (v / v) and 30% (v / v) of water. In some embodiments, the solvent system comprises ethanol. In some embodiments, a composition comprises between about 5% (v / v) and 50% (v / v) of water. In some embodiments, a composition comprises between about 5% (v / v) and 30% (v / v) of water. In some embodiments, a composition comprises between about 5% (v / v) and 20% (v / v) of water. In some embodiments, a composition comprises between about 10% (v / v) and 20% (v / v) of water. In some embodiments, a composition comprises about 10% (v / v) of water. In some embodiments, a composition comprises about 15% (v / v) of water. In some embodiments, a composition comprises about 20% (v / v) of water.

[0231]

[0209] In some embodiments, the solvent system comprises ethanol. In some embodiments, a composition comprises about 60% (v / v) of ethanol. In some embodiments, a composition comprises between about 10% (v / v) and 70% (v / v) of ethanol. In some embodiments, a composition comprises between about 10% (v / v) and 60% (v / v) of ethanol. In some embodiments, the solvent system comprises 2025-10-09 ethanol. In some embodiments, a composition comprises between about 15% (v / v) and 55% (v / v) of ethanol. In some embodiments, a composition comprises between about 20% (v / v) and 50% (v / v) of ethanol. In some embodiments, a composition comprises between about 20% (v / v) and 40% (v / v) of ethanol. In some embodiments, a composition comprises between about 25% (v / v) and 35% (v / v) of ethanol. In some embodiments, a composition comprises abou 25% (v / v) of ethanol. In some embodiments, a composition comprises about 30% (v / v) of ethanol. In some embodiments, a composition comprises about 35% (v / v) of ethanol.

[0232]

[0210] In some embodiments, the solvent system comprises isopropanol. In some embodiments, a disclosed composition comprises about 40% (v / v) of isopropanol. In some embodiments, a disclosed composition comprises between about 10% (v / v) and 70% (v / v) of isopropanol. In some embodiments, a disclosed composition comprises between about 10% (v / v) and 60% (v / v) of isopropanol. In some embodiments, a disclosed composition comprises between about 15% (v / v) and 55% (v / v) of isopropanol. In some embodiments, a disclosed composition comprises between about 20% (v / v) and 50% (v / v) of isopropanol. In some embodiments, a disclosed composition comprises between about 20% (v / v) isopropanol. In some embodiments, a disclosed composition comprises between about 25% isopropanol. In some embodiments, a disclosed composition comprises about 30% (v / v) of isopropanol. In some embodiments, a disclosed composition comprises about 35% (v / v) of isopropanol. In some embodiments, a disclosed composition comprises about 40% (v / v) of isopropanol.

[0233]

[0211] In some embodiments, the solvent system comprises propylene glycol. In some embodiments, a disclosed composition comprises about 10% (v / v) of propylene glycol. In some embodiments, a disclosed composition comprises between about 1 % (v / v) and 50% (v / v) of propylene glycol. In some embodiments, a disclosed composition comprises between about 5% (v / v) and 40% (v / v) of propylene glycol. In some embodiments, a disclosed composition comprises between about 9% (v / v) and 30% (v / v) of propylene glycol. In some embodiments, a disclosed composition comprises about 25% (v / v) of propylene glycol. In some embodiments, a disclosed composition comprises about 20% (v / v) of propylene glycol. In some embodiments, a disclosed composition comprises about 15% (v / v) of propylene glycol. In some embodiments, a disclosed composition comprises about 10% (v / v) of propylene glycol.

[0234]

[0212] In some embodiments, a composition comprises a viscosity modifying agent. In some embodiments, the viscosity modifying agent is a thickener. Common thickeners include but are not limited to: acrylates, carbomers, cellulose matrices, silicones, carrageenans, gums, resins, polysaccharides, and high melting point waxes and oils such as beeswax, coconut oil, palm oil, soybean oil, stearic acid, rapeseed, cocoa butter, shea butter, gums, rosins, resins, paraffins, and petroleum jelly. In some embodiments, the viscosity modifying agent is a carbohydrate. Exemplary carbohydrates include monosaccharides, disaccharides, oligosaccharides, and polysaccharides. Exemplary polysaccharides include cellulose, methylcellulose, hydroxypropylmethylcellulose, chitin, galactoarabinan, polygalactose, 2025-10-09 and polyarabinose. Exemplary glycerides includes hydroxystearic acid monoglyceride, hydroxystearic acid diglyceride, isostearic acid monoglyceride, isostearic acid diglyceride, oleic acid monoglyceride, oleic acid diglyceride, ricinoleic acid monoglyceride, ricinoleic acid diglyceride, linoleic acid monoglyceride, linoleic acid diglyceride, linolenic acid monoglyceride, linolenic acid diglyceride, erucic acid monoglyceride, erucic acid diglyceride, tartaric acid monoglyceride, tartaric acid diglyceride, citric acid monoglyceride, citric acid diglyceride, malic acid monoglyceride, malic acid monoglyceride, malic acid diglyceride, and mixture thereof. In some embodiments, the viscosity modifying agent is a polymer. The polymer may be a natural or synthetic polymer. Natural polymers include polysaccharides, nucleic acid, and proteins. Synthetic polymers include polyesters, polyureas, polycarbonates, polyvinyl alcohol, polyamides, polyethers, polyesters, polyamines, polytyrosines, polyanhydrides, polyphosphazenes, polyacrylamides, polyacrylates, polymethacrylates, polyvinylpyrrolidone, etc. Exemplary thickening agents include alginate derivatives, preneutralized carbomer 430, hydrophilic silicas, polysaccharides, xanthan gum, guar guar, agar agar, carboxymethylcellulose, hydroxyethylcellulose, polyacrylates, polyacrylamides, polyvinylpyrrolidone, and salts. In some embodiments, compositions of the disclosure may comprise a viscosity modifying agent at a concentration of about 0.01 %, about 0.02%, about 0.05%, about 0.1 %, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11 %, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 61 %, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 75%, about 75%, and about 80% of the composition, on a weight or volume basis.

[0235]

[0213] For example, in some embodiments, a composition comprises hydroxypropylcellulose as a viscosity modifying agent. It will be appreciated, however, that hydroxypropylcellulose or another disclosed excipient may perform multiple functions when included in a composition, as noted elsewhere herein. In some embodiments, a composition comprises between about 0.1 % (w / v) and about 10% (w / v) of hydroxypropylcellulose. In some embodiments, a composition comprises about 0.1 %, about 0.5%, about 1 %, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, or about 10% (w / v) of hydroxypropylcellulose.

[0236]

[0214] In some embodiments, a disclosed composition comprises about 2% (w / v) of hydroxypropylcellulose. In some embodiments, the composition comprises less than about 1 % (w / v) of hydroxypropylcellulose, such as about 0.75%, 0.5%, or 0.125% (w / v) of hydroxypropylcellulose. In some embodiments, a composition does not comprise a hydroxyalkylcellulose.

[0237]

[0215] In some embodiments, a composition comprises an adhesion modifying agent. In some embodiments, a composition comprises an adhesive polymer. Adhesive polymers have physicochemical 2025-10-09 properties that allow prolonged binding to tissue surfaces. In some embodiments, inclusion of an adhesive polymer in the composition increases the amount of time that an active agent is in contact with, and can diffuse across, a barrier (e.g., scalp skin). In some embodiments, the adhesive polymer is chitosan, gelatin guar gum, lectins, sodium alginate, soluble starch, tragacanth, xanthan gum deacetylated gum, polyacrylic acid, polyvinyl alcohol, hydroxypropylmethylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, sodium carboxymethylcellulose, a thiomer, polycarbophil, hyaluronic acid, dermatan sulfate, or a combination thereof. In some embodiments, the adhesion modifying agent is a tackifier. Common tackifiers include but are not limited to gums, resins (natural or modified), carbomers, or other natural or synthetic polymers. In some embodiments, compositions of the disclosure may comprise a adhesion modifying agent at a concentration of about 0.01 %, about 0.02%, about 0.05%, about 0.1%, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1 %, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about

[0238] 21 %, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 61 %, about

[0239] 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 75%, about 75%, and about 80% of the composition, on a weight or volume basis.

[0240]

[0216] In some embodiments, a composition comprises a preservative. Preservatives can be used to inhibit microbial growth or increase stability of the composition, thereby prolonging the shelf life of the composition. Suitable preservatives are known in the art and include EDTA, EGTA, benzalkonium chloride or benzoic acid or benzoates (e.g., sodium benzoate), vitamin A, vitamin C (ascorbic acid), citric acid, vitamin E, and tocopherol.

[0241]

[0217] In some embodiments, a composition comprises an antioxidant. Without being bound by theory, antioxidants generally can delay or inhibit the oxidative decomposition of components of disclosed compositions (e.g., active agents, such as TRH or any other active ingredients), which may thereby improve the stability and extend the shelf-life of disclosed compositions. In some embodiments, the antioxidant is a-tocopherol, ascorbyl palmitate, butylated hydroxyanisole, butylated hydroxytoluene, methionine, citric acid, ascorbic acid, sodium ascorbate, sodium thiosulfate, sodium bisulfite, sodium metabisulfite, ascorbyl palmitate, thioglycerol, propyl gallate, cysteine, or a combination thereof. In some embodiments, the antioxidant is a cyclodextrin, D-a-tocopherol, rosmarinic acid, or a combination thereof. In some embodiments, compositions of the disclosure may comprise an antioxidant at a concentration of about 0.01 %, about 0.02%, about 0.05%, about 0.1 %, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1 %, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11 %, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21 %, about 22%, about 2025-10-09 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 61 %, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 75%, about 75%, and about 80% of the composition, on a weight or volume basis.

[0242]

[0218] In some embodiments, the composition may include one or more vitamins. Any vitamin having a property, for example, to nourish the hair, inhibit hair loss and / or enhance hair growth may be used. Representatively, suitable vitamins may include, but are not limited to, essential B vitamins such as thiamine, riboflavin, niacin, vitamin B6, folic acid, vitamin B12, biotin and pantothenic acid. In some embodiments, the concentration of the vitamin in the composition is sufficient to provide the desired benefit of treating hair loss and / or promoting hair growth while remaining compatible with disclosed compositions (e.g., active agents, such as TRH, or any other active ingredients). Such concentration can vary with the vitamin selected, the effect desired and the type and concentration of other components, and other like factors. Representatively, the composition may include between 1 mg and 200 mg of a vitamin(s), and in some embodiments between 50 mg and 250 mg of the vitamin(s).

[0243]

[0219] In some embodiments, a composition comprises a solubilizing agent. Without being bound by theory, solubilizing agents generally form complexes with active ingredients which can have different physicochemical properties than the active ingredient alone. The properties of the complexes can increase the solubility of TRH, or any other active ingredient(s) in the composition. In some embodiments, the solubilizing agent is a water-soluble organic solvent, a non-ionic surfactant, a water insoluble lipid, an organic liquid, a cyclodextrin, or a phospholipid. In some embodiments the solubilizing agent is a water-soluble enhancing agent. In some embodiments, the water-soluble enhancing agent is polyethylene glycol 300, polyethylene glycol 400, ethanol, propylene glycol, xanthan gum, glycerin, N-methyl-2-pyrrolidone, dimethylacetamide, dimethylsulfoxide, or a combination thereof. In some embodiments, the solubilizing agent is propylene glycol. In some embodiments, the solubilizing agent is xanthan gum. In some embodiments the solubilizing agent is a non-ionic surfactant. In some embodiments, the non-ionic surfactant is Cremophor EL, Cremophor RH 40, Cremophor RH 60, d-tocopherol polyethylene glycol 1000 succinate, polysorbate 20, polysorbate 80, Solutol HS 15, sorbitan monooleate, poloxamer 407, Labrafil M-1944CS, Labrafil M-2125CS, Labrasol, Gellucire 44 / 14, Softigen 767, mono- and di-fatty acid esters of PEG 300, 400, or 1750, or a combination thereof. In some embodiments the solubilizing agent is an organic liquid. In some embodiments, the organic liquid is beeswax, d-alpha-tocopherol, oleic acid, or a medium-chain mono- or diglyceride. In some embodiments the solubilizing agent is a cyclodextrin. In some embodiments, the solubilizing agent is alpha-cyclodextrin, beta-cyclodextrin, gamma-cyclodextrin, hydroxypropyl-beta-cyclodextrin, and sulfobutylether-beta-cyclodextrin. In some embodiments, the solubilizing agent is alpha-cyclodextrin. In some embodiments, the solubilizing agent is beta-cyclodextrin. In some embodiments, the solubilizing 2025-10-09 agent is gamma-cyclodextrin. In some embodiments, the solubilizing agent is hydroxypropyl-beta-cyclodextrin. In some embodiments, the solubilizing agent is sulfobutylether-beta-cyclodextrin. In some embodiments the solubilizing agent is a phospholipid. In some embodiments, the phospholipid is hydrogenated soy phosphatidylcholine, distearoylphosphatidylglycerol, L-alpha-dimyristoylphosphatidylcholi ne, or L-alpha-dimyristoylphosphatidylglycerol. In some embodiments, the solubilizing agent is lecithin. In some embodiments, compositions of the disclosure may comprise solubilizing agent at a concentration of about 0.01 %, about 0.02%, about 0.05%, about 0.1 %, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1 %, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about

[0244] 20%, about 21 %, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about

[0245] 61 %, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 75%, about 75%, or about 80% of the composition, on a weight or volume basis.

[0246]

[0220] In some embodiments, a disclosed composition comprises a colorant. Suitable colorants and / or dyes and / or pigments may include, but not be limited to, colors such as e.g., white, black, yellow, blue, green, pink, red, orange, violet, indigo, brown, and combinations thereof, pigments such as, e.g., Timica Extra Large Sparkles, titanium dioxide and chromium oxide greens, ultramarine blues and pinks and ferric oxides. Colorants and / or dyes and / or pigments may be present, individually or in total (if more than one colorant and / or dye and / or pigment is included), in disclosed compositions in an amount ranging from about 0.01 wt% to about 5 wt% (calculated as the total weight of colorants and / or dyes and / or pigments in the composition divided by the total weight of the composition). Colorants may be present, individually or in total (if more than one colorant is included), in disclosed compositions in an amount ranging from about 0.01 wt% to about 5 wt% (calculated as the total weight of colorants in the composition divided by the total weight of the composition).

[0247]

[0221] In some embodiments, a disclosed composition comprises a binder. Suitable binders include, without limitations, polyvinylpyrrolidone (PVP), marine colloids, carboxyvinyl polymers, starches, cellulosic polymers such as hydroxyethylcellulose, carboxymethylcellulose (carmellose), hydroxypropylmethylcellulose, hydroxyethylpropylcellulose, hydroxybutyl methyl cellulose, and salts thereof (e.g., carmellose sodium), natural gums such as karaya, xanthan, carrageenans, gellan gum, locust bean gum, gum arabic and tragacanth, chitosan, colloidal magnesium aluminum silicate, and colloidal silica. Binders may be present, individually or in total (if more than one binder is included), in disclosed compositions in an amount ranging from about 0.01 wt% to about 5 wt% (calculated as the total weight of binders in the composition divided by the total weight of the composition).

[0248]

[0222] In some embodiments, a composition comprises a humectant. Humectants, such as low molecular 2025-10-09 weight polyethylene glycol (e.g., PEG6-PEG12), may be present, individually or in total (if more than one humectant is included), in the composition in an amount of up to about 10 wt%, up to about 5 wt%, up to about 3 wt%, up to about 1 wt%, or up to about 0.1 wt% (calculated as the total weight of humectants in the composition divided by the total weight of the composition).

[0249]

[0223] In some embodiments, a composition comprises a surfactant. The surfactants that can be included in the composition may be anionic, nonionic, or amphoteric compounds. Suitable examples of anionic surfactants are one or more of higher alkyl sulfates such as potassium or sodium lauryl sulfate, higher fatty acid monoglyceride monosulfates, such as the salt of the monosulfated monoglyceride of hydrogenated coconut oil fatty acids, alkyl sulfonates such as sodium dodecyl benzene sulfonate, higher fatty sulfoacetates, higher fatty acid esters of 1 ,2 dihydroxypropane sulfonate. Examples of water soluble nonionic surfactants are condensation products of ethylene oxide with various hydrogen-containing compounds that are reactive therewith and have long hydrophobic chains (e.g., aliphatic chains of about 12 of 20 carbon atoms), which condensation products contain hydrophilic polyoxyethylene moieties, such as condensation products of poly (ethylene oxide) with fatty acids, fatty alcohols, fatty amides and other fatty moieties, and with propylene oxide and polypropylene oxides, e.g., Pluronic materials such as Pluronic F127. Exemplary suitable alkyl polyglycoside (APG) surfactant(s) that may be used in the composition may comprise APG C8-C10, APG C10-C16, decyl glucoside, coco-glucoside, anionic APG carboxylate, sodium lauryl glucose carboxylate, lauryl glucoside, D-glucopyranose (oligomeric, CIO-16 glycosides, carboxymethyl ethers, sodium salts), C12-C16 fatty alcohol glycoside, and combinations thereof. Exemplary APG surfactant(s) that may be used may have an industry designation of Plantaren® 2000 N UP / MB, Plantapon® LGC Sorb, Plantaren® 1200 N UP / MB, and Plantaren® 818 UP / MB. Surfactants may be present, individually or in total (if more than one surfactant is included) in the composition in an amount ranging from about 0.01 wt% to about 10 wt% (calculated as the total weight of surfactants in the composition divided by the total weight of the composition).

[0250]

[0224] In some embodiments, a composition comprises a gelling agent. Exemplary gelling agent(s) that may be issued in disclosed compositions may comprise pectins, starches, and gelatin forms derived from animals or from plants (e.g., pork gelatin). The pectin in the composition may include, e.g., high methoxyl pectin, low methoxyl pectin, or a combination thereof. In some embodiments, the pectin is amidated pectin. In other embodiments, the pectin is non-amidated pectin. In certain embodiments, the pectin is a combination of amidated pectin and non-amidated pectin. The gelatin in the composition may include Type A gelatin, Type B gelatin, a hide or skin gelatin (e.g., calf skin, pig skin) and / or a bone gelatin (e.g., calf bone, pig bone) used alone or in combination. Gelling agent(s) may be present, individually or in total (if more than one gelling agent is included) in the composition in an amount ranging from about 0.1 wt% to about 20 wt% (calculated as the total weight of gelling agents in the composition divided by the total weight of the composition). In some embodiments, the composition does not comprise a gelling agent. 2025-10-09

[0251]

[0225] Depending on unit dosage volume and total volume, the composition may be provided as a final packaged product (e.g., in a bottle or any other suitable container). In some embodiments, the bottle is a dropper bottle, a fine mist spray bottle, a pump bottle, a glass bottle, or a plastic bottle. In some embodiments, the bottle is a dropper bottle. In some embodiments, the bottle is a fine mist spray bottle. In some embodiments, the bottle is a pump bottle. In some embodiments, the bottle is a glass bottle. In some embodiments, the bottle is a plastic bottle. In some embodiments, the bottle is between about 15 mL and 90 mL. In some embodiments, the bottle is about 15 mL (i.e., about 0.5 ounces). In some embodiments, the bottle is about 20 mL. In some embodiments, the bottle is about 30 mL (i.e., about 1 ounce) . In some embodiments, the bottle is about 40 mL. In some embodiments, the bottle is about 50 mL. In some embodiments, the bottle is about 60 mL (i.e., about 2 ounces). In some embodiments, the bottle is about 70 mL. In some embodiments, the bottle is about 80 mL. In some embodiments, the bottle is about 90 mL (i.e., about 3 ounces). In some embodiments, the bottle is greater than 90 mL.

[0252]

[0226] In some embodiments, a disclosed excipient may perform more than one function when included in a composition. For example, hydroxypropylcellulose may function to modify the viscosity of the composition, while also affecting the composition’s adhesive properties, or stabilizing an active agent in the composition, promoting emulsification, or another function as described herein.

[0253]

[0227] One of ordinary skill in the art appreciates that the selection of a suitable excipient for use in a disclosed composition may depend on a variety of factors. Relevant factors in the selection of the appropriate excipient(s), include, for example, compatibility of the excipient with the active agents (e.g., T3, T4, any additional active agent(s)), desired penetration kinetics of the active agents, processing parameters, biocompatibility, and user preferences.

[0254] E. Exemplary Features of Disclosed Compositions and Methods

[0255]

[0228] In some embodiments, disclosed compositions and methods produce fewer adverse effects (e.g., side effects) than a comparative method or composition, such as the standard of care for a particular hair loss condition (e.g., androgenetic alopecia). Without being bound by theory, it is proposed that TRH could be hypothetically mostly absorbed by the epidermis and dermis before entering the bloodstream. This would localize potential side effects and likely be mitigated further by pulsing the therapy, as described in various embodiments herein.

[0256]

[0229] In some embodiments, the hair loss is caused by alopecia. In some embodiments, the hair loss is caused by androgenetic alopecia, alopecia areata, persistent patchy alopecia areata, alopecia totalis, alopecia universalis, diffuse alopecia areata, ophiasis alopecia, cicatricial alopecia, lichen planopilaris, frontal fibrosing alopecia, central centrifugal cicatricial alopecia (CCCA), traction alopecia, alopecia barbae, or postpartum alopecia. In some embodiments, the hair loss is caused by androgenetic alopecia. In some embodiments, the hair loss is male pattern baldness. In some embodiments, the hair loss is female pattern baldness. 2025-10-09

[0257]

[0230] In some embodiments, a disclosed composition or method is particularly suitable for administration to a subject with certain qualities (e.g., biomarkers) indicative of high likelihood of treatment success, or low risk of treatment (e.g., reduced risk of side effects). For example, in some embodiments, only subjects below a threshold thyroid hormone level (e.g., level of TRH, TSH, T3, or T4) or another biomarker are subjected to a disclosed method or composition. In some embodiments, a subject is subjected to bloodwork before treatment with a disclosed method or composition.

[0258]

[0231] The present disclosure further provides kits for carrying out the methods of the disclosure, which comprises TRH described herein, or a pharmaceutical composition comprising TRH with additional active ingredients as described herein.

[0259]

[0232] In some embodiments, administration of a disclosed composition increases the level of a biomarker in the subject. In some embodiments, administration of a disclosed composition decreases the level of a biomarker in the subject. The "level" of a biomarker refers to a measurable quantity, quality, or characteristic of a biomarker, including concentration, amount, presence, frequency, activity, or expression. For example, the "level" of a biomarker may refer to its concentration in a biological sample (e.g., blood, plasma, serum, tissue), its rate of production or degradation, its expression in a cell or tissue (e.g., as determined by gene expression assays such as RT-PCR, RNA sequencing, or microarrays), its activity or functional state, or any other measurable parameter indicative of the biomarker's presence or effect.

[0260]

[0233] In some embodiments, the biomarker is associated with hair follicle activity, cycling, or miniaturization. In some embodiments, increases and decreases of a measure (e.g., duration of a hair style stage, hair shaft production, hair follicle length, rate of hair follicle growth, rate of decreasing hair follicle length, repigmentation, depigmentation) are determined by comparison with the measure in the subject prior to treatment with a disclosed composition or method. In some embodiments, increases and decreases of a measure are determined by comparison with a different subject (i.e., a control subject) who has not been treated with a disclosed composition or method. In some embodiments, increases and decreases of a measure are determined by comparison with an average of the measure in a population of subjects who have not been treated with a disclosed composition or method.

[0261]

[0234] In embodiments, administration of a disclosed composition increases hair shaft production. Hair shaft production refers to the amount of hair produced by a hair follicle. In some embodiments, the composition increases the hair shaft production by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%. In some embodiments, the composition increases the hair shaft production by greater than 100%.

[0262]

[0235] In embodiments, administration of a disclosed composition prolongs the anagen hair growth phase. The anagen phase is the first of three hair growth stages, during which hair is actively produced by a hair follicle. In embodiments, the composition prolongs the anagen phase by 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%. 2025-10-09

[0263]

[0236] In embodiments, administration of a disclosed composition increases the proliferation of bulge epithelial stem cells. Bulge epithelial stem cells are located within the bulge region of hair follicles, have high proliferative capacity, and multipotency to regenerate keratinocytes, sebaceous glands, and epidermis tissue. In embodiments, the composition increases the proliferation of bulge stem cells by 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0264]

[0237] In embodiments, administration of a disclosed composition increases the proliferation of hair matrix keratinocytes. In some embodiments, hair matrix keratinocyte proliferation is measured by the expression of the protein Ki-67. In embodiments, the composition increases the proliferation of hair matrix keratinocytes by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0265]

[0238] In embodiments, administration of a disclosed composition increases melanin production. In embodiments, the composition increases melanin production by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0266]

[0239] In embodiments, administration of a disclosed composition increases expression of FGF7. FGF7 is a growth factor which promotes hair growth by regulating the initiation and prolongation of the anagen hair growth stage. In embodiments, the composition increases the expression of FGF7 by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0267]

[0240] In embodiments, administration of a disclosed composition decreases the expression of TGF0-2. In adult hair cycling, TGF0-2 is a catagen-promoting cytokine that suppresses the anagen hair growth stage and enhances apoptotic signaling in hair follicles. In embodiments, the composition decreases the expression of TGFp-2 by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0268]

[0241] In embodiments, administration of a disclosed composition increases expression of IGF-1. In embodiments, the composition increases the expression of IGF-1 by 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0269]

[0242] In embodiments, administration of a disclosed composition increases expression of epidermal VEGF-A. In embodiments, the composition increases the expression of epidermal VEGF-A by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0270]

[0243] In embodiments, administration of a disclosed composition increases expression of keratin 15 (K15). Keratin 15 is a widely used biomarker for bulge epithelial stem cells. The increase of keratin 15 indicates an increased presence or maintenance of bulge epithelial stem cells. In embodiments, the composition increases the expression of keratin 15 by 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0271]

[0244] In embodiments, administration of a disclosed composition increases the percentage of K15+cells in hair follicles. Keratin 15 marks bulge epithelial stem and progenitor keratinocytes, and an increase in K15+cells indicates enrichment or maintenance of this compartment. In embodiments, the composition increases the percentage of K15+cells by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 2025-10-09

[0272] 100%.

[0273]

[0245] In embodiments, administration of a disclosed composition increases the percentage of K15+Ki67+cells. K15+Ki67+double-positive cells indicate proliferating bulge-lineage keratinocytes, and an increase in K15+Ki67+cells indicates activation and expansion of this compartment. In embodiments, the composition increases the percentage of K15+Ki67+cells by 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0274]

[0246] In embodiments, administration of a disclosed composition decreases the percentage of K15+Casp-3+cells. K15+Casp-3+double-positive cells indicate apoptotic bulge-lineage keratinocytes, and a decrease indicates reduced apoptotic signaling in this compartment. In embodiments, the composition decreases the percentage of K15+Casp-3+cells by 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%. In some embodiments, the percentage of K15+Casp-3+cells shows no material change.

[0275]

[0247] In embodiments, administration of a disclosed composition increases the expression of MTCO1. MTCO1 encodes a core subunit of mitochondrial cytochrome c oxidase (complex IV), and increased MTCO1 indicates enhanced mitochondrial respiratory activity supportive of the anagen hair growth stage. In embodiments, the composition increases the expression of MTCO1 by 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0276]

[0248] In embodiments, administration of a disclosed composition increases the expression of keratin 85 (K85 or KRT85). Keratin 85 is a hair-shaft keratin expressed by matrix-derived cortical and cuticular keratinocytes during anagen, and increased keratin 85 indicates enhanced hair-shaft differentiation and fiber formation. In embodiments, the composition increases the expression of keratin 85 by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0277]

[0249] In embodiments, administration of a disclosed composition decreases the rate of hair graying, otherwise known as depigmentation, in a subject. Melanin content in the hair shaft and hair-follicle pigmentary unit is used as the primary metric of pigmentation, and an increase in melanin content indicates repigmentation and slowing of graying. Hair graying (depigmentation) may be caused by dysfunction of differentiated melanocytes and is generally temporarily reversible until hair-follicle melanocyte stem cells are depleted, after which depigmentation can become irreversible. Without being bound by theory, a disclosed composition increases melanin synthesis or retention in the hair-follicle pigmentary unit, thereby promoting repigmentation and reducing the rate of graying.

[0278]

[0250] In embodiments, a disclosed composition decreases depigmentation by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%. In embodiments, the composition increases repigmentation by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%. In some embodiments, the composition increases repigmentation by greater than 100%.

[0279]

[0251] In embodiments, a disclosed composition prevents the initiation of depigmentation in hair follicles. In embodiments, the composition prevents 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 2025-10-09 100% of hair follicles from initiating depigmentation in a subject. In embodiments, the composition increases the amount of hair follicles producing pigment by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0280] F. Examples

[0281]

[0252] The following examples are included for illustrative purposes only and are not intended to limit the scope of the invention.

[0282] EXAMPLE 1 : Human Skin Organ Culture Studies

[0283]

[0253] Purpose: The study described herein was tests the effects of administration of disclosed topical compositions on human scalp skin in organ culture. Prior work (e.g., Paus, et al. FASEB J. 2010;24(2):393— 403) tested applying thyrotropin-releasing hormone (TRH) to hair follicles extracted from the scalp through a solution infused into the culture itself. The current study was conducted by applying disclosed topical compositions to scalp skin. In this work, TRH promoted hair-shaft elongation and prolonged the anagen phase of the hair follicle cycle, increased hair matrix keratinocyte proliferation, reduced apoptosis, antagonized TGF0-2-induced catagen, and stimulated follicular melanogenesis.

[0284]

[0254] Methods:

[0285]

[0255] Skin Biopsy: Skin biopsies (4 mm) containing terminal hair follicles were prepared and placed at air liquid interface (with the subcutis and dermis located in the medium to avoid the epidermis falling into the culture medium during the culture) within serum-free supplemented William’s E media and incubated at 37°C in a humidified atmosphere of 5% CO. After 24 hours of culture for equilibration, skin biopsies were treated topically with 2 pL of viscous formulation either containing William’s E medium for the vehicle, or a combination of isopropanol, propylene glycol, hydroxypropylcellulose, and TRH (25.48nM - 25.48pM), without and with a specific reverse peptidase inhibitor (PI) against TRH-degrading ectoenzyme [TRH-DE], Glp-Asn-Pro-AMC (1 mM).

[0286]

[0256] Medium change and topical administration of test compounds (any leftover test compound was removed before applying fresh TRH + / - PI) were performed every other day. At Day 0, 1 , 6, and during every medium change, the surface of each 4 mm skin fragment was imaged to perform hair shaft production analysis. At the end of the six days of culture, samples were embedded in OCT and snap-frozen in liquid nitrogen before being stored at -80°C until further analyses.

[0287]

[0257] OCT-embedded hairy scalp skin was cryosectioned (7 pm thickness) with a Crysostar NX50 (Thermo Scientific, Epredia) or an M630 (Medite Medical GmbH) cryostat.

[0288]

[0258] Immunochemistrv: For immunofluorescence staining, all samples / sections were fixed and pre-incubated for 30 minutes at RT, followed by incubation with a primary antibody (previously used and characterized) overnight at 4°C where indicated below. After three 5-minute washes in PBS, sections were incubated with the appropriate fluorescently tagged secondary antibody for 45 minutes at room 2025-10-09 temperature and then embedded in DAPI / Fluoromount (Electron Microscopy Sciences). Negative control for the primary antibody was used by omitting the primary antibody.

[0289]

[0259] Ki-67 + Caspase-3. To evaluate proliferating and apoptotic keratinocytes in the hair matrix, cryosections were fixed with 4% paraformaldehyde in PBS and pre-incubated in 10% goat serum, followed by incubation with a mouse anti-Ki-67 antibody (1 :800; Cell Signalling Technology, 9449S) and a rabbit anti-caspase-3 antibody (1 :400; Cell Signalling Technology, 9661) overnight at 4°C. After three 5-minute washes in PBS, sections were incubated with goat anti-mouse IgG - Rhodamine (1 :200; Jackson ImmunoResearch) and goat anti-rabbit IgG - Alexa Fluor 488 (1 : 400; Life Technologies) for 45 minutes at RT.

[0290]

[0260] IGF-1 . To evaluate IGF-1 protein expression, tissue cryosections were fixed in acetone (10 min at -20°C) followed by incubation with a rabbit anti-human IGF-1 antibody (1 :250; Novus Biologicals, NBP2-16299) overnight at 4°C. After three 5-minute washes in PBS, secondary incubation was performed for 45 minutes at RT using a goat anti-rabbit IgG - Alexa Fluor 555 antibody (1 :400; Life Technologies).

[0291]

[0261] TGF -2. To evaluate TGF0-2 protein expression, tissue cryosections were fixed in methanol (10 min at -20°C) and pre-incubated in 10% goat serum in TBS, followed by incubation with a rabbit anti-human TGF-02 antibody (1 :400; Abeam, ab113670) overnight at 4°C. After three 5-minute washes in PBS, secondary incubation was performed for 45 minutes at RT using a goat anti-rabbit IgG - Rhodamine antibody (1 :200; Jackson ImmunoResearch).

[0292]

[0262] FGF7. To evaluate FGF7 protein expression, tissue cryosections were fixed in acetone (10 min at -20°C) and pre-incubated in 10% goat serum in TBS, followed by incubation with a rabbit anti-KGF / FGF-7 antibody (1 :100; Abeam, ab90259) overnight at 4°C. After three 5-minute washes in PBS, sections were incubated with a goat anti-rabbit IgG - Alexa Fluor 488 antibody (1 :400; Life Technologies) for 45 minutes at RT.

[0293]

[0263] K15. To evaluate K15 protein expression, tissue cryosections were fixed in 4% paraformaldehyde in PBS and pre-incubated in 10% goat serum, followed by incubation with a mouse anti-human CK15-FITC conjugated antibody (1 :200; Novus Biologicals, NBP2-54462F) for 2 hours at 37°C. After three 5-minute washes in PBS, sections were mounted as above (no secondary required).

[0294]

[0264] K85. To evaluate K85 protein expression, tissue cryosections were fixed in acetone (10 min at -20°C) followed by incubation with a guinea pig anti-human CK85 antibody (1 :1000; Progen, GP-HHB5) overnight at 4°C. After three 5-minute washes in PBS, secondary incubation was performed for 45 minutes at RT using a goat anti-guinea pig IgG - Alexa Fluor 594 antibody (1 :400; Life Technologies).

[0295]

[0265] CD31. To evaluate CD31 protein expression, tissue cryosections were fixed in acetone (10 min at -20°C) followed by incubation with a mouse anti-human CD31 antibody (1 :50; Agilent, IS610) overnight at 4°C. After three 5-minute washes in PBS, sections were incubated with a goat anti-mouse IgG - Alexa Fluor 555 antibody (1 :400; Life Technologies) for 45 minutes at RT. 2025-10-09

[0296]

[0266] MTCO1. To evaluate MTCO1 protein expression, tissue cryosections were fixed in 4% paraformaldehyde and permeabilized and blocked in 10% goat serum + 0.3% Triton X-100 in PBS, followed by incubation with a rabbit anti-human MTCO1 antibody [EPR19628] (1 :50; Abeam, ab203912) overnight at 4°C. After three 5-minute washes in PBS, sections were incubated with a goat anti-rabbit IgG-FITC antibody (1 :200; Jackson ImmunoResearch, 111-095-144) for 45 minutes at RT, followed by an amplification step with an anti-FITC Alexa Fluor 488 antibody (1 :700; Life Technologies, A11096) for 30 minutes at RT.

[0297]

[0267] VEGFA. To evaluate VEGFA protein expression, tissue cryosections were fixed in 4% paraformaldehyde in PBS (10 min at RT), followed by incubation with a rabbit anti-human Alexa Fluor 488 anti-VEGFA antibody [EP1176Y] (1 :500; Abeam, ab206886). After three 5-minute washes in PBS, sections were mounted as above (no secondary required).

[0298]

[0268] Melanin. Warthin-Starry histochemistry was carried out to assess the melanin content of the HFs according to the previously established protocol (Joly-Tonetti et al. 2016; Lai and Healy 2016; Samra et al. 2023; Sevilla et al. 2023). Briefly, slides were fixed in 4% paraformaldehyde and then washed with PBS and distilled water. The cryosections were next stained with 0.5% silver nitrate solution and incubated at 50 °C for 6 minutes. A reducing agent solution, consisting of 4% acidulated gelatin, 2% acidulated silver nitrate, and 0.1 % hydroquinone, was added to each section and incubated at 50 °C for 5 minutes. The sections were washed in PBS, counterstained with hematoxylin, and dehydrated in 100% 2-propanol before embedding with VectaMount Express Mounting Medium (Vector Laboratories).

[0299]

[0269] Quantitative (Immuno-)Histomorohometrv and Microscopy: Images were taken using a BZ-X800 All-in-one Fluorescence Microscope (Keyence Corporation) and Image Analysis software Bz-800 Analyzer (Keyence Corporation) at 200x magnification, with a constant exposure time maintained throughout imaging to facilitate later normalization. Images were then analyzed using NIH Imaged software (National Institute of Health), with defined reference areas (dotted area shown in the vehicle images). For staining intensity, the mean intensity was measured. The Imaged cell counter function was used for counting single and double-positive cells.

[0300]

[0270] For Hair cycle staging and hair cycle score, Masson-Fontana histochemistry and Ki-67 / Caspase-3 immunostainings were used for accurately determining hair cycle staging ex vivo, using the well-defined classification criteria described in detail (Kloepper et al. 2010; Oh et al. 2016).

[0301]

[0271] Statistical analysis: All data are presented as fold change of the mean ± SEM. We employed the Student’s t-test or the Mann-Whitney test for statistical analysis, depending on whether the data adhered to a Gaussian or non-Gaussian distribution, respectively (as determined by the d'Agostino and Pearson omnibus normality test). This analysis used Graph Pad Prism 9 software (GraphPad Software, San Diego, CA, USA). Statistical significance was set at p<0.05. 2025-10-09

[0302]

[0272] Vehicle Formulation and Experimental Design: The two vehicle formulations chosen for topically applying TRH peptide hormone compositions were designed on the basis of their likelihood to enhance penetration and transfollicular absorption of TRH. Dissolution of the ingredients was tested; TRH was fully soluble at the highest concentration tested (25.48 pM).

[0303]

[0273] The concentration of the specific peptidase inhibitor (against TRH-degrading ectoenzyme |TRH-DE], which is the principal enzyme responsible for terminating the action of TRH) to increase the action of TRH, namely Glp-Asn-Pro-AMC (1 mM) was based on data found in the literature, which shows a significant increase of TRH release even under basal conditions with a Ki=0.87mM (Kelly JA et al, Biochem J 2005). Furthermore, this inhibitor is a reversible inhibitor, which may limit its potential adverse effects. 74] The concentrations of TRH in Formulation A were varied as follows:

[0304] 275] The experiments of this example were conducted according to the following timeline: 2025-10-09

[0305] 276] Human scalp skin was sourced from three donors:

[0306]

[0277] Donor 1 : Scalp skin, Female, 51 yr-old, Caucasian

[0307]

[0278] Donor 2: Scalp Skin, Female, 66 yr-old, Middle East

[0308]

[0279] Donor 3: Scalp Skin, Male, 36 yr-old, Caucasian

[0309]

[0280] Frontotemporal and occipital human scalp skin biopsies from healthy female and male patients undergoing facelift surgery were collected for this study. Punch biopsies were then placed in culture in 5% CO2at air / liquid interphase. 2 pL of treatment (vehicle T3 or T4) was applied on day 1 , day 3, and day 5 after previously removing any left over of previous application. On day 6, the skin biopsies were embedded in cryomatrix and snap-frozen in liquid nitrogen before being processed for quantitative-(immuno)histomorphometry.

[0310]

[0281] Results:

[0311]

[0282] Histology and Hyperkeratosis. Topical application of TRH produced minimal epidermal changes. At nanomolar concentrations, the histological sections of scalp skin showed a modest thickening of the stratum corneum but no evidence of parakeratosis (retention of nuclei in the cornified layer). In particular, 25.48 nM and 254.8 nM TRH occasionally increased desquamation yet the epidermis remained 2025-10-09 well-differentiated. FIG. 1 illustrates these findings: hematoxylin-eosin (H&E) sections from Day 6 skin specimens show comparable epidermal architecture across vehicle, 25.48 nM, 254.8 nM and 2.548 pM TRH. The dashed green line in FIG. 1 emphasizes that only the 254.8 nM dose shows mild focal parakeratosis.

[0312]

[0283] At micromolar concentrations, epidermal effects were slightly more pronounced. FIG. 14 depicts Day 6 H&E sections following topical 2.548 pM and 25.48 pM TRH. While 2.548 pM TRH produced a modestly thicker stratum corneum, the highest dose (25.48 pM) resulted in more noticeable hyperkeratosis but still lacked parakeratosis. These observations suggest that topical TRH, even at high micromolar doses, is not overtly irritant, though refinement of the vehicle may minimize mild desquamation.

[0313]

[0284] Hair Shaft Production. The effect of TRH on hair-shaft growth was dose dependent. When applied at 25.48 nM or 254.8 nM, TRH significantly increased hair shaft production over six days of organ culture. The bar graph in FIG. 2 shows that hair shafts lengthened by 75-80 pm in the 25.48 nM group and approximately 60 pm in the 254.8 nM group compared with ~30 pm in vehicle controls; photomicrographs of Day 1 and Day 6 punches on the right demonstrate visually fuller hair shafts. In contrast, the 2.548 pM dose increased shaft growth only slightly.

[0314]

[0285] At higher concentrations, TRH showed a smaller yet still positive effect. In FIG. 15, topical 25.48 pM TRH increased hair-shaft production by -10 percentage points relative to vehicle over days 1-6, whereas 2.548 pM TRH showed little or no change. Combined, these data indicate that nanomolar doses of TRH optimally stimulate hair shaft growth whereas micromolar doses confer only a modest benefit.

[0315]

[0286] Anagen Duration. Hair cycle staging revealed that low-dose TRH did not significantly alter the proportion of follicles remaining in anagen after six days. FIG. 3 presents stacked bar graphs showing that ~70 % of hair follicles (HFs) remained in anagen regardless of treatment. Immunostaining for Ki-67 (green) and cleaved caspase-3 (red) and Warthin-Starry staining (WS) on the right confirm similar proliferative and apoptotic activity across vehicle, 25.48 nM, 254.8 nM and 2.548 pM groups.

[0316]

[0287] In contrast, high-dose TRH prolonged anagen. FIG. 16 demonstrates that 2.548 pM TRH increased the percentage of HFs in anagen from ~80 % to nearly 90 %, while 25.48 pM TRH showed no appreciable effect. These results echo prior microdissected follicle studies demonstrating that TRH prolongs anagen and likely reduces androgenetic alopecia-associated telogen effluvium.

[0317]

[0288] Hair Matrix Keratinocyte Proliferation. Proliferation of hair matrix (HM) keratinocytes was markedly enhanced by low doses of TRH. The bar graph in FIG. 4 indicates that 25.48 nM TRH increased the percentage of Ki-67A+ HM keratinocytes from ~5 % to -10 %; 254.8 nM and 2.548 pM doses produced only minor changes. Ki-67 / caspase-3 double-immunofluorescence images show more numerous red Ki-67A+ nuclei in the 25.48 nM group. This confirms earlier ex vivo findings that TRH promotes hair matrix proliferation when delivered systemically.

[0318]

[0289] At micromolar doses, a similar pattern emerged. FIG. 17 shows that 25.48 pM TRH increased 2025-10-09 Ki-67A+ HM cells to ~20 %, whereas 2.548 pM TRH had little effect. Thus, both low-dose (25.48 nM) and high-dose (25.48 pM) TRH promote matrix keratinocyte proliferation, suggesting that TRH’s mitogenic effect is concentration dependent but not limited to a narrow window.

[0319]

[0290] Melanin Production. Melanin production in anagen VI hair follicles responded differentially to TRH dose. FIG. 5 demonstrates that 25.48 nM TRH elevated Warthin-Starry (WS) staining intensity by -15 % relative to vehicle, whereas 254.8 nM and 2.548 pM doses did not alter pigmentation. This replicates earlier reports that very low concentrations of TRH stimulate melanogenesis.

[0320]

[0291] In the micromolar range, no significant changes in melanin were detected. The bar graph in FIG. 18 shows that both 2.548 pM and 25.48 pM TRH maintained WS intensity near vehicle levels. This suggests that only nanomolar TRH enhances pigment production; higher concentrations may not reach the pigmentary unit effectively or could activate counter-regulatory pathways.

[0321]

[0292] TGF- 2 Expression. Transforming growth factor 02 (TGF-02) is a catagen-inducing cytokine. Topical TRH robustly suppressed TGF-02 expression in outer root sheath keratinocytes at all low doses tested. FIG. 6 shows that 25.48 nM, 254.8 nM and 2.548 pM TRH reduced TGF-02 immunoreactivity to -80 % of vehicle levels, with representative confocal images highlighting diminished green signal in treated follicles. This confirms that TRH antagonizes TGF-02-induced premature catagen induction.

[0322]

[0293] High-dose TRH had a similar effect. In FIG. 19, both 2.548 pM and 25.48 pM TRH reduced TGF-02 expression to -20 % of vehicle. These data underscore that TRH’s hair-growth-promoting action is mediated primarily through suppression of catagen-inducing TGF-02 signaling across a wide concentration range.

[0323]

[0294] IGF-1 Expression. Insulin-like growth factor 1 (IGF-1) is a known hair-growth factor. Surprisingly, topical TRH at low doses did not appreciably raise IGF-1 levels. FIG. 7 reveals that IGF-1 expression in dermal papilla (DP) cells remained near vehicle levels across all nanomolar doses, while 254.8 nM TRH significantly increased IGF-1 in the outer root sheath to -1 .3-fold. The text accompanying this figure notes that the hair-growth effect of 25.48 nM TRH is therefore not driven by IGF-1 induction. At micromolar concentrations TRH decreased IGF-1. As illustrated in FIG. 20, both 2.548 pM and 25.48 pM TRH reduced IGF-1 immunoreactivity in DP and ORS cells to -30-40 % of vehicle, despite concomitant increases in hair shaft production and anagen duration. This further supports the notion that TRH promotes hair growth primarily via TGF-02 suppression rather than IGF-1 up-regulation.

[0324]

[0295] FGF-7 (Keratinocyte Growth Factor) Expression. Fibroblast growth factor 7 (FGF-7 / KGF) stimulates hair matrix keratinocyte activity and hair shaft production. Low-dose TRH decreased FGF-7 in hair matrix keratinocytes yet modestly increased it in the ORS. FIG. 8 shows that 25.48 nM and 2.548 pM TRH reduced FGF-7 expression in the hair matrix to -80 % of vehicle, whereas 254.8 nM TRH slightly increased ORS FGF-7 (right panel). These data implicate TRH in modulating intrafollicular KGF signaling primarily by suppressing TGF-02. 2025-10-09

[0325]

[0296] Micromolar doses tended to increase FGF-7. In FIG. 21, both 2.548 pM and 25.48 pM TRH increased FGF-7 immunoreactivity in the ORS to ~1.2-fold, while hair matrix levels remained unchanged. Although statistical significance is not yet established due to small sample size, these findings suggest that high-dose TRH may promote hair shaft keratin synthesis via KGF up-regulation.

[0326]

[0297] Keratin 85 Expression. Keratin 85 (K85) is a marker of hair shaft keratin production. As shown in FIG. 9, 25.48 nM TRH increased K85 expression in the proximal hair matrix to ~1.2-fold compared with vehicle, whereas higher nanomolar and low micromolar doses reduced K85. Fluorescent images reveal increased red K85 signal within the evaluation region (green dashed square) for the 25.48 nM group.

[0327]

[0298] In the micromolar range, both 2.548 pM and 25.48 pM TRH enhanced K85 expression. FIG. 22 demonstrates that K85 immunoreactivity increased to roughly 1.5-fold of vehicle in both dose groups. These data align with the hair-shaft production results, indicating that TRH stimulates keratin synthesis particularly at very low and high concentrations.

[0328]

[0299] Keratin 15 and Bulge Stem Cells. K15A+ cells mark the bulge epithelial stem cell niche. Nanomolar TRH did not markedly change K15 levels. However, high-dose TRH exerted distinct effects on bulge stem cells. FIG. 23 shows that 25.48 pM TRH significantly decreased both the number and expression of K15A+ cells (to ~60 % of vehicle), whereas 2.548 pM TRH had a modest effect. Conversely, FIG. 24 reveals that 25.48 pM TRH increased the fraction of proliferative K15+Ki-67+ cells from ~0.5 % to -2.5 %, suggesting replenishment of the stem cell pool despite overall reduced K15 expression. These findings may reflect accelerated differentiation of bulge stem cells into CD200+ / CD34+ progenitor cells rather than depletion of the niche.

[0329]

[0300] Angiogenesis and VEGF-A / CD31. Vascular endothelial growth factor A (VEGF-A) and CD31 mark angiogenic activity. At nanomolar doses, TRH did not alter VEGF-A expression in the hair matrix (see FIG. 10) and did not change the number of CD31A+ endothelial cells in the papillary dermis (FIG. 11). Nevertheless, FIG. 12 indicates that all nanomolar doses significantly reduced the percentage of VEGF-AA+ cells in the dermis (from -8 % in vehicle to -4-6 % in treated groups), hinting at a potential anti-angiogenic effect. Interestingly, FIG. 13 shows that 254.8 nM TRH increased epidermal VEGF-A production to -1.3-fold of vehicle, whereas 25.48 nM TRH decreased it; this may support epidermal rejuvenation effects.

[0330]

[0301] Micromolar TRH consistently decreased angiogenic markers. FIG. 25 demonstrates that 2.548 pM and 25.48 pM TRH reduced VEGF-A expression in the hair matrix by -30 % compared with vehicle. Similarly, FIG. 26 shows that the percentage of CD31A+ cells in dermal sections fell from -60 % in controls to -40 % in both micromolar groups. The number of VEGF-AA+ dermal cells also declined sharply (see FIG. 27). Together these observations suggest that high-dose TRH exerts an unexpected anti-angiogenic effect that warrants further investigation.

[0331]

[0302] Discussion. Collectively, these data demonstrate that topical TRH exerts 2025-10-09 concentration-dependent effects on human scalp hair follicles. Low nanomolar doses (25.48 nM) robustly promote hair shaft production, increase hair matrix keratinocyte proliferation, stimulate melanin production and up-regulate K85 expression. These benefits occur without significant changes in anagen duration or in angiogenic markers. The principal mechanism appears to be suppression of TGF-02-induced catagen induction, since IGF-1 and FGF-7 are not markedly elevated. The slightly higher nanomolar dose (254.8 nM) retains some pro-growth activity but with reduced efficacy and an unexpected increase in epidermal VEGF-A production.

[0332]

[0303] At micromolar concentrations (2.548 pM and 25.48 pM), TRH continues to suppress TGF-02 and increases hair matrix proliferation and keratin synthesis; however, only the highest dose significantly prolongs anagen. Melanin production is unaffected and IGF-1 expression is reduced, indicating that TGF-02 suppression remains the major pathway. Importantly, both micromolar doses decrease VEGF-A and CD31A+ cells in the dermis, suggesting an anti-angiogenic effect that could have long-term consequences for hair follicle nutrition. The highest dose also reduces K15 expression while increasing proliferation of K15A+ cells, implying accelerated differentiation of bulge stem cells. Future work should clarify which dose has the most favorable effects on humans in vivo.

[0333] Summary of TRH Effects on Human Hair Follicles 2025-10-09

[0334] Arrows indicate the direction of change relative to vehicle t increase, | decrease, «-► no change).

[0335] EXAMPLE 2: Human Hair Follicle Organ Culture

[0336]

[0304] Purpose: The study described herein was tests the effects of administration of disclosed topical compositions on human hair follicular units, with surrounding scalp skin, in organ culture. Prior work (e.g., (Paus, et al. FASEB J. 2010;24(2):393— 403)) tested applying thyrotropin-releasing hormone (TRH) as the only active to hair follicles extracted from the scalp through a solution infused into the culture itself. In this work, novel TRH and T3 combinations promoted H1 F-1 signaling, carbon metabolism, and fatty acid metabolism in human hair follicles surrounded by scalp skin.

[0337]

[0305] Methods:

[0338]

[0306] Follicular Unit Bioosv: Hair follicular units biopsies containing terminal hair follicles were prepared and placed in serum-free supplemented William’s E media and incubated at 37°C in a humidified atmosphere of 5% CO. After 24 hours of culture for equilibration, follicular unit biopsies were treated topically with 1 mL of supplemented William’s E media prepared in our laboratory, either containing William’s E medium alone for the vehicle, or William’s E Media with TRH (25.48nM - 2.548 uM) and T3 (10nM).

[0339]

[0307] From Day 1 - Day 7, medium change was performed every other day. At every medium change, each follicular unit was imaged to perform hair shaft length and width analysis. At the end of the seven 2025-10-09 days of culture, samples were snap-frozen in liquid nitrogen before being stored at -80°C before being delivered to a third-party laboratory for RNA-sequencing.

[0340]

[0308] RNA-Seauencing. For RNA-sequencing, ten frozen tissue samples (human hair follicles) were submitted for nucleic-acid extraction and quality control. Total RNA was isolated and assessed on an Agilent TapeStation system, which provided RNA integrity numbers (RINe) and 28S / 18S area ratios; for example, sample SGL16960 had a RIN of 5.9 and a 28S / 18S ratio of 2.8 with an RNA concentration of 360 ng / l, while sample SGL16964 had a RIN of 7.2, a 3.4 ratio, and 110 ng / l concentration. After confirming that RIN values ranged from approximately 5.9— 7.3 and that sufficient RNA was available, messenger RNA was enriched using poly(A) selection specific for eukaryotic transcripts; ERCC RNA spike-in controls and unique molecular identifier (UMI) adapters were added during library construction to enable accurate normalization and duplicate removal. Libraries were prepared from the enriched RNA, qualified and quantified, and then sequenced on an Illumina platform. Sequencing employed paired-end reads and a depth of about 50 million reads per sample, producing raw FASTQ data for downstream bioinformatic processing. Standard data processing included adapter trimming, alignment to the human reference genome and quantification of gene expression to support differential expression analysis.

[0341]

[0309] Statistical Analysis.

[0342] RNA Sequencing FASTQ files were aligned using STAR algorithm. Gene level raw count matrices were analyzed using DESeq2. Library size normalization and dispersion estimation was conducted according to the DESEq2 framework with design matrix specifying our four experimental groups: control, low dose T3+TRH, medium dose T3+TRH and high dose T3+TRH, exact dosages can be found in Table 1. Differential expression was assessed using the Wald test on group contrasts. P values were adjusted for multiple testing using BH false discovery rate. The cut off for the adjusted p-value was p<0.05. Functional interpretation was performed using KEGG pathway enrichment for Homo sapiens.

[0343]

[0310] Formulation and Experimental Design: The combinations of TRH and T3 were designed based on their ability to shift hair follicle RNA expression to a pro-growth state.

[0344]

[0311] The formulations of TRH and T3 were as follows: 12] The experiments of this example were conducted according to the following timeline: 2025-10-09

[0345]

[0314] Donor 1 : Scalp skin, Male.

[0346]

[0315] Results.

[0347]

[0316] Global transcriptional changes: FIG. 28 depicts a principal-component analysis (PCA) revealing a strong separation between control samples and T3+TRH-treated samples along the first principal component. Control replicates clustered together, whereas treated samples formed a dose-dependent gradient: low-dose samples were closer to controls, medium-dose samples occupied an intermediate position and high-dose samples were the furthest from baseline. This indicates that the combination of T3 and TRH induces broad transcriptional shifts in hair follicles and that the magnitude of change scales with TRH concentration. FIG. 30 depicts a heatmap of the top 50 most variable genes across all samples, highlighting distinct expression patterns for control versus low, medium and high doses of T3+TRH.

[0348]

[0317] Differential expression analysis quantified these differences. Compared with vehicle, low-dose treatment yielded «532 significantly differentially expressed genes (FIG. 29A), with a bias towards 2025-10-09 up-regulation. High-dose treatment produced «237 differentially expressed genes with a similar up-regulation bias (FIG. 29C), whereas the medium-dose produced —216 differentially expressed genes but skewed towards down-regulation (FIG. 29B). These results suggest that both low and high doses stimulate transcriptional activation, whereas intermediate dosing may trigger homeostatic or negative feedback responses.

[0349]

[0318] Functional enrichment: KEGG pathway analysis of the differentially expressed genes highlighted several biological pathways. The low-dose group showed robust enrichment in hypoxia-inducible factor-1 (HIF-1) signalling, AMP-activated protein kinase (AMPK) signalling, PPAR signalling, gluconeogenesis / glycolysis, fatty-acid metabolism and ferroptosis (FIG. 31A). These pathways remained enriched at high dose (FIG. 31 C), whereas the medium-dose group (FIG. 31 B) shared only a subset (AMPK, PPAR and ferroptosis) and enriched cornified envelope formation pathways with high dose. The enrichment of HIF-1 signalling is notable because HIF-1a is suppressed in androgenetic alopecia (AGA) scalp tissue and its activation up-regulates trichogenic genes; thus, the observed up-regulation of HIF-1 targets may underlie the hair-growth effects of the combination treatment. Enrichment of PPAR signalling is also mechanistically relevant: deletion of PPARy in murine hair follicle stem cells causes a scarring-alopecia phenotype and reduced expression of lipid-metabolism genes, suggesting that PPAR activation supports maintenance of the epithelial stem-cell niche.

[0350]

[0319] The pathway analysis also revealed significant alterations in metabolic pathways. Human hair follicles are highly glycolytic mini-organs; the outer root sheath (ORS) stores glycogen and active glycolysis provides both energy and metabolic precursors for hair growth. Inhibition of glycogen phosphorylase prolongs anagen and promotes ex vivo hair growth, illustrating that metabolic reprogramming directly influences the hair-cycle. The observed enrichment of glycolysis / gluconeogenesis, pyruvate metabolism and glucagon signalling pathways indicates that T3+TRH alters carbon metabolism, possibly to meet the energetic demands of prolonged anagen. In addition, many fatty-acid-related pathways were enriched (fatty-acid metabolism, glycerolipid metabolism, adipocytokine signalling). Thyroid hormones are known regulators of lipid metabolism: they modulate the expression of lipogenic genes via nuclear receptors and can either promote fatty-acid p-oxidation or induce lipolysis depending on receptor isoform and tissue context. Thyroid hormones also promote lipolysis and up-regulate fatty-acid transporter proteins (FATPs) and fatty-acid binding proteins, increasing circulating free fatty acids. Enrichment of lipid-metabolism pathways suggests that T3+TRH co-treatment remodels lipid handling within hair follicles, which may provide energy for growth and support membrane biogenesis.

[0351]

[0320] Integration with TRH biology: The RNA-seq findings align with known biology of TRH and T3 in hair follicles. The TRH gene is expressed within human hair follicles, and exogenous TRH stimulates hair-shaft elongation, prolongs the anagen growth phase and modulates keratinocyte proliferation and apoptosis. Meanwhile, thyroid hormone receptors influence both carbohydrate and lipid metabolism. Our 2025-10-09 enrichment of HIF-1 and PPAR signalling pathways suggests that T3+TRH may act via both hypoxic and lipid-metabolism pathways. HIF-1 activation promotes trichogenic gene expression and hair induction, whereas PPARy maintains stem-cell function and its loss causes scarring alopecia. Collectively, the transcriptional response to T3+TRH is consistent with a pro-growth and metabolic-reprogramming effect, supporting the hypothesis that combining TRH with T3 enhances hair-growth signalling and may reverse AGA-associated down-regulation of HIF-1 and PPAR pathways.

[0352]

[0321] Discussion: The RNA-sequencing analysis of human hair follicular units treated with combined TRH and T3 provides mechanistic insights into how this co-therapy may promote hair growth. Principal-component separation and differential expression analyses revealed dose-dependent transcriptional changes, with low and high doses inducing robust gene up-regulation and the intermediate dose yielding more subdued responses. These results mirror known biology: hypoxia-inducible factor-1 (HIF-1) signalling is down-regulated in androgenetic alopecia (AGA) and its activation stimulates trichogenic genes. Up-regulation of HIF-1 targets by T3+TRH suggests the treatment counters hypoxic suppression and may extend the anagen phase. Similarly, enrichment of peroxisome proliferator-activated receptor (PPAR) pathways aligns with reports that PPARy maintains the lipid metabolism of follicular stem cells and that its loss causes scarring alopecia.

[0353]

[0322] Metabolic reprogramming was another hallmark of T3+TRH treatment. Pathways related to glycolysis, gluconeogenesis, pyruvate metabolism and fatty-acid turnover were consistently enriched. Hair follicles are highly glycolytic mini-organs that store glycogen in the outer root sheath and rely on aerobic glycolysis to fuel rapid keratinocyte proliferation. Thyroid hormones exert broad effects on carbohydrate and lipid metabolism; they modulate gluconeogenic enzymes, stimulate lipolysis and up-regulate fatty-acid transporter proteins. The concerted activation of carbon and lipid pathways therefore likely supplies energy and membrane precursors to support hair growth. TRH, meanwhile, is expressed in human hair follicles and independently prolongs anagen and stimulates hair-shaft elongation. Together, TRH and T3 may synergistically activate hypoxic, metabolic and lipid signalling networks to rejuvenate follicular growth.

[0354]

[0323] Despite these promising findings, the analysis has limitations: it relied on a set of ex vivo follicles from a single donor. More comprehensive studies in vivo with larger cohorts are needed to validate these results, and dissecting downstream HIF-1 and PPAR effectors could identify additional actionable targets. The combined TRH and T3 treatment nonetheless appears to induce a pro-growth, metabolically active state by activating hypoxic signalling, reprogramming carbohydrate and lipid metabolism, and supporting the follicular stem-cell niche. Further work in physiologically relevant in vivo models will determine whether this combination therapy can translate into durable hair regeneration in androgenetic alopecia and related disorders.

[0355] EXAMPLE 3: Clinical Study Design for Assessing Efficacy of Disclosed Topical Compositions

[0356]

[0324] Purpose: To evaluate the efficacy of the disclosed TRH-containing topical compositions and 2025-10-09 methods for treating or preventing hair loss. The study measures clinical endpoints (e.g., hair-shaft production, rate of hair loss) and molecular biomarkers of hair growth identified in ex vivo studies (e.g., TGF-02, IGF-1 , FGF-7, K85, K15, VEGF-A, CD31).

[0357]

[0325] Eligibility: All subjects undergo collection of medical history and physical examination. A treatment group includes individuals experiencing hair loss (e.g., as a result of androgenetic alopecia or another like condition). Hair loss, including various underlying causes thereof, is diagnosed in an individual according to diagnostic techniques known to those of skill. A control group is established which will be administered a topical composition that does not comprise an active agent (e.g., the vehicle formulation). Subjects will be over the age of 18 years. Subjects can withdraw from the study at any time, and for any reason. Subjects unwilling to participate in the study, or having a medical condition that contraindicates them for treatment with a disclosed topical composition, will also be excluded.

[0358]

[0326] Study design overview: The treatment group will self-administer a topical composition comprising (i) 25.48 nM thyrotropin-releasing hormone (TRH); (ii) 0.5 % (w / v) hydroxypropylcellulose; (iii) 60 % (v / v) ethanol; (iv) 20 % (v / v) propylene glycol; and about (v) 19.5 % (v / v) water. The topical composition will be administered every day for four weeks, followed by one week without administration. A higher dose cohort will receive 254.8 nM TRH formulated identically. A third cohort will receive 2.548 pM TRH in the same vehicle to evaluate dose dependency. The control formulation lacking TRH will be administered according to the same treatment cycles. At the end of each treatment cycle, overall hair growth will be assessed according to techniques known in the art (e.g., measuring hair-shaft production). Subjects will also be biopsied before and at the end of treatment. Biopsy samples will be analysed for biomarkers of hair growth, including hair-shaft production, melanin production and the expression of the proteins TGF-02, IGF-1 , FGF-7, K85, K15, VEGF-A and CD31 , as well as proliferation (Ki-67) and apoptosis markers (cleaved caspase-3). These biomarkers will be measured according to techniques described herein and otherwise known in the art.

[0359]

[0327] Results: Based on ex vivo organ-culture results, subjects in the low-dose TRH treatment group are expected to show increased hair-shaft growth relative to the control group. Subjects treated with nanomolar TRH may also exhibit reductions in TGF-02 expression and modest increases in FGF-7 and K85, together with increased hair-matrix keratinocyte proliferation. Micromolar TRH doses are expected to produce only slight increases in hair-shaft production but significant suppression of TGF-02 and up-regulation of K85 and bulge stem-cell proliferation, although they may reduce IGF-1 and FGF-7 and decrease VEGF-A and CD31 , signalling a potential anti-angiogenic effect. The study will compare the clinical data with these preclinical expectations to determine the efficacy and safety of each dose of TRH.

[0360] EXAMPLE 4: Individual Administration of Disclosed Topical Compositions for Treating Hair Loss

[0361]

[0328] Described herein are exemplary treatment protocols for treating or preventing hair loss in individuals in need thereof. The following illustrative cases highlight how TRH-containing compositions 2025-10-09 may be self-administered to promote hair growth. Each patient administers the topical composition to the scalp according to the general vehicle described in Example 2.

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[0329] Patient 1 : Patient 1 is diagnosed with androgenetic alopecia and suffers from hair loss. Patient 1 self-administers a topical composition comprising 25.48 nM TRH; 0.125 % (w / v) hydroxypropylcellulose; 60 % (v / v) ethanol; 20 % (v / v) propylene glycol; and about 18 % (v / v) water. The composition is administered every other day for two consecutive weeks, followed by one week without administration. After one treatment cycle, Patient 1 experiences increased hair-shaft production (e.g., thicker, longer hair shafts) and exhibits improvements in biomarkers indicative of successful treatment: suppressed TGF-02 expression, elevated K15 and increased Ki-67-positive hair-matrix cells. Patient 1 also shows a modest increase in hair pigment (melanin) and elevated K85 keratin expression.

[0363]

[0330] Patient 2: Patient 2 is diagnosed with alopecia areata and suffers from patchy hair loss. Patient 2 self-administers a topical composition comprising 254.8 nM TRH formulated as above. The composition is administered every other day for two consecutive weeks, followed by one week without administration. After four treatment cycles, Patient 2 experiences a decreased rate of hair loss and exhibits improvements in biomarkers: moderate suppression of TGF-02, slight increases in IGF-1 and FGF-7 in the outer root sheath, and increased K85 expression. Hair-matrix proliferation remains close to baseline, but hair-shaft production remains above that of the vehicle group. Patient 2 does not show significant changes in angiogenic markers (VEGF-A and CD31).

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[0331] Patient 3: Patient 3 is diagnosed with alopecia totalis and suffers from total hair loss. Patient 3 self-administers a topical composition comprising 2.548 M TRH; 2 % (w / v) hydroxypropylcellulose; 60 % (v / v) isopropanol; 20 % (v / v) propylene glycol; and about 18 % (v / v) water. The composition is administered daily for three consecutive days, followed by one week without administration. After two treatment cycles, Patient 3 experiences moderate improvement in hair-shaft production and displays strong suppression of TGF-02, increased K85 expression and increased proliferation of bulge epithelial stem cells (K15+Ki-67+). However, Patient 3 also exhibits decreased IGF-1 and FGF-7 expression and reduced VEGF-A and CD31 levels, indicating a potential anti-angiogenic effect; therefore, continued monitoring is advised.

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[0332] Patient 4: Patient 4 is diagnosed with persistent patchy alopecia areata and suffers from hair loss. Patient 4 self-administers a topical composition comprising 25.48 pM TRH; 0.5 % (w / v) hydroxypropylcellulose; 60 % (v / v) ethanol; and about 40 % (v / v) propylene glycol;The composition is administered every other day for two consecutive weeks, followed by one week without administration. After three treatment cycles, Patient 4 experiences only a slight increase in hair-shaft production but shows robust hair-matrix keratinocyte proliferation and up-regulation of K85. Patient 4 also exhibits suppressed TGF-02 and increased proliferation of bulge epithelial stem cells, but this comes with significant decreases in IGF-1 , FGF-7, VEGF-A and CD31, suggesting the high dose may inhibit angiogenesis. The potential benefits of enhanced proliferation must therefore be balanced against the risk of reduced nutrient supply to 2025-10-09 the follicle.

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[0333] Patient 5: Patient 5 is diagnosed with androgenetic alopecia and suffers from hair loss. Patient 4 self-administers a topical composition comprising 25.48 pM TRH; 10 nM T3; 0.125 % (w / v) hydroxypropylcellulose; 60 % (v / v) ethanol; and about 20 % (v / v) propylene glycol; and about 19 % (v / v) water. The composition is administered every day for three months. After two treatment cycles, Patient 5 experiences only a slight increase in hair-shaft production but shows robust hair-matrix keratinocyte proliferation and up-regulation of K85. Patient 5 also exhibits suppressed TGF-02 and increased proliferation of bulge epithelial stem cells, but this comes with significant decreases in IGF-1 , FGF-7, VEGF-A and CD31 , suggesting the high dose may inhibit angiogenesis. The potential benefits of enhanced proliferation must therefore be balanced against the risk of reduced nutrient supply to the follicle. EXAMPLE 5: Individual Administration of Disclosed Topical Compositions for Treating Hair Graying

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[0334] Described herein are exemplary treatment protocols for treating or preventing hair graying in individuals in need thereof. Organ-culture experiments indicate that low nanomolar TRH can modestly enhance melanin production in hair follicles. The following illustrative cases apply this finding to human subjects.

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[0335] Patient 6: Patient 6 is experiencing hair graying. Patient 6 self-administers a topical composition comprising 25.48 nM TRH; 1 % (w / v) hydroxypropylcellulose; 30 % (v / v) ethanol; 50 % (v / v) propylene glycol; and about 15 % (v / v) water. The composition is administered every other day for two consecutive weeks, followed by one week without administration. Beginning after one treatment cycle, Patient 6 experiences gradual repigmentation of their hair, consistent with the modest increase in melanogenesis observed ex vivo. Biomarker analysis shows suppressed TGF-02, slight increases in FGF-7, elevated K85, and stable IGF-1 and VEGF-A levels, indicating that low-dose TRH promotes pigment without inhibiting angiogenesis.

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[0336] Patient 7: Patient 7 is experiencing hair graying. Patient 7 self-administers a topical composition comprising 254.8 nM TRH; 1 % (w / v) hydroxypropylcellulose; 30 % (v / v) ethanol; 50 % (v / v) propylene glycol; and about 15 % (v / v) water. The composition is administered every other day for two consecutive weeks, followed by one week without administration. Beginning after one treatment cycle, Patient 7 experiences a decreased rate of hair graying (e.g., slower depigmentation) and shows improvements in biomarkers such as suppressed TGF-02, slight increases in IGF-1 and FGF-7, and elevated K85. As TRH doses increase, the anti-angiogenic effects observed in vitro (decreased VEGF-A and CD31) may become more pronounced; therefore, continued monitoring and adjustment of dose may be warranted to balance pigment restoration with vascular health.

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[0337] The foregoing description, for purposes of illustration, uses specific nomenclature to provide a thorough understanding of the invention. However, it will be apparent to one skilled in the art that specific details are not required in order to practise the invention. Thus, the foregoing description of specific 2025-10-09 embodiments is presented for purposes of illustration and description. It is not intended to be exhaustive or to limit the invention to the precise compositions, formulations, methods or the like disclosed; many modifications and variations are possible in view of the above teachings. The embodiments were chosen and described in order to best explain the principles of the invention and its practical applications, to thereby enable others skilled in the art to best utilise the invention and various embodiments with various modifications as are suited to the particular use contemplated, when such uses are beyond the specific examples disclosed. Accordingly, the scope of the invention shall be defined solely by the following claims and their equivalents.

Claims

2025-10-09CLAIMSThe invention claimed is :1 . A composition comprising thyrotropin-releasing hormone (TRH), wherein the composition is formulated for topical administration.

2. The composition of claim 1 , comprising between about 25 nM and about 500 M of TRH.

3. The composition of claim 2, comprising between about 100 nM and about 100 pM of TRH.

4. The composition of claim 3, comprising between about 1 pM and about 100 pM of TRH.

5. The composition of claim 4, comprising about 25 pM of TRH.

6. The composition of claim 1 , comprising one or more pharmaceutically acceptable excipients selected from the group consisting of penetration enhancers, carriers, diluents, emulsifiers, stabilizers, viscosity modifying agents, adhesion modifying agents, preservatives, antioxidants, adhesive polymers, solubilizing agents, colorants, binders, humectants, surfactants, and gelling agents.

7. The composition of claim 1 , comprising a solvent system.

8. The composition of claim 7, wherein the solvent system comprises an alcohol.

9. The composition of claim 8, wherein the solvent system comprises any of ethanol, isopropanol, and propylene glycol.

10. The composition of claim 9, wherein the solvent system comprises isopropanol.11 . The composition of claim 10, comprising between about 10% and about 70% (v / v) of isopropanol.

12. The composition of claim 11 , comprising about 40% (v / v) of isopropanol.

13. The composition of claim 9, wherein the solvent system comprises ethanol.

14. The composition of claim 13, comprising about 60% (v / v) of ethanol.

15. The composition of claim 9, wherein the solvent system comprises propylene glycol.

16. The composition of claim 15, comprising between about 5% and about 80% (v / v) of propylene glycol.2025-10-0917. The composition of claim 16, comprising about 10% (v / v) of propylene glycol.

18. The composition of claim 16, comprising about 20% (v / v) of propylene glycol.

19. The composition of claim 16, comprising about 50% (v / v) of propylene glycol.

20. The composition of claim 7, wherein the solvent system comprises water.21 . The composition of claim 20, comprising between about 1 % and about 50% (v / v) of water.

22. The composition of claim 21 , comprising about 30% (v / v) of water.

23. The composition of claim 1 , further comprising one or more additional active agents.

24. The composition of claim 23, comprising an additional thyroid hormone.

25. The composition of claim 24, wherein the additional thyroid hormone is triiodothyronine (T3).

26. The composition of claim 25, comprising T3 and TRH at a T3:TRH ratio of between about 1 : 1 and about 1 :1000.

27. The composition of claim 23, comprising any of finasteride, dutasteride, and minoxidil.

28. The composition of claim 27, comprising finasteride.

29. The composition of claim 27, comprising dutasteride.

30. The composition of claim 27, comprising minoxidil.31 . The composition of claim 23, comprising T3 and finasteride.

32. The composition of claim 23, comprising T3 and dutasteride.

33. The composition of claim 23, comprising T3 and minoxidil.

34. The composition of claim 1 , comprising TRH as the only hormone.

35. The composition of claim 23, comprising a peptidase inhibitor.

36. The composition of claim 35, wherein the peptidase inhibitor is an inhibitor of TRH-degrading ectoenzyme.

37. The composition of claim 36, wherein the peptidase inhibitor is a tripeptide.

38. The composition of claim 37, wherein the tripeptide comprises a Glp-Asn-Pro moiety.2025-10-0939. A composition consisting essentially of TRH, wherein the composition is formulated for topical administration.

40. A composition consisting essentially of TRH and T3, wherein the composition is formulated for topical administration.41 . A composition consisting essentially of TRH, T3, and any of finasteride, dutasteride, and minoxidil; wherein the composition is formulated for topical administration.

42. The composition of any of claims 1-41 , in lyophilized form.

43. The composition of any of claims 1-41 , in unit dosage form.

44. The composition of claim 43, wherein the total unit dose volume of the composition is between about 0.1 mL and about 10 mL.

45. The composition of any of claims 1-41 , wherein the composition is formulated as a spray, ointment, salve, gel, paste, lotion, liniment, or cream.

46. The composition of any of claims 1-41 for use in treating or preventing hair loss or hair graying.

47. Use of the composition of any of claims 1 -41 for the manufacture of a medicament for treating or preventing hair loss or hair graying.

48. A method of treating or preventing hair loss or hair graying in a subject, comprising administering to the subject the composition of any of claims 1 -41 .

49. The method of claim 48, comprising administering to the subject between about 0.1 and 10 mL of the composition per unit dose.

50. The method of claim 49, comprising administering to the subject about 1 mL of the composition per unit dose.51 . The method of claim 48, comprising administering to the subject between about 30 pg and 200 pg of TRH per unit dose.

52. The method of claim 51 , comprising administering to the subject about 10 pg of TRH per unit dose.

53. The method of claim 48, wherein the composition is administered daily.

54. The method of claim 48, wherein the composition is administered every other day.2025-10-0955. The method of claim 48, wherein the composition is administered every other day for several consecutive weeks followed by a prolonged period without administration.

56. The method of claim 55, wherein the composition is administered every other day for two consecutive weeks followed by a prolonged period without administration.

57. The method of claim 55 or 56, wherein the prolonged period without administration is at least two weeks.

58. The method of claim 48, wherein the hair loss is caused by androgenetic alopecia, alopecia areata, persistent patchy alopecia areata, alopecia totalis, alopecia universalis, diffuse alopecia areata, ophiasis alopecia, cicatricial alopecia, lichen planopilaris, frontal fibrosing alopecia, central centrifugal cicatricial alopecia (CCCA), traction alopecia, alopecia barbae, or postpartum alopecia.

59. The method of claim 48, resulting in increased hair shaft production.

60. The method of claim 59, wherein hair shaft production is increased by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to baseline values measured prior to administration of the composition.61 . The method of claim 48, resulting in prolonged anagen hair growth phase.

62. The method of claim 61 , wherein the anagen hair growth phase is prolonged by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to baseline values measured prior to administration of the composition.

63. The method of claim 48, resulting in increased proliferation of bulge epithelial stem cells.

64. The method of claim 63, wherein bulge epithelial stem cell proliferation is increased by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to baseline values measured prior to administration of the composition.

65. The method of claim 48, resulting in increased expression of keratin 15.

66. The method of claim 65, wherein the expression of keratin 15 is increased by 1 %, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% compared to baseline values measured prior to administration of the composition.

67. The method of claim 48, resulting in decreased hair depigmentation.

68. The method of claim 67, wherein hair depigmentation is decreased by 1 %, 5%, 10%, 20%, 30%,2025-10-0940%, 50%, 60%, 70%, 80%, 90%, or 100%.

69. The method of claim 67, resulting in hair repigmentation.

70. A composition comprising a TRH metabolite, wherein the composition is formulated for topical administration.71 . The composition of claim 70, wherein the metabolite is Cyclo(His-Pro).

72. A composition comprising a TRH peptide fragment, wherein the composition is formulated for topical administration.

73. The composition of claim 72, wherein the peptide fragment is a dipeptide.

74. The composition of claim 73, wherein the dipeptide is Pyroglutamyl Dipepti de-30 Amide.

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