Variables Affecting Delivery of Glycopyrronium Tosylate Through Human Skin In Vitro.

BACKGROUND: Hyperhidrosis is a condition characterized by excessive sweating beyond what is required for normal thermal regulation. It can involve multiple body areas including the axillae, palms, soles, or craniofacial regions. Glycopyrronium tosylate (GT) is a topical anticholinergic approved by the FDA (2018) for treatment of primary axillary hyperhidrosis in patients 9 years and older. OBJECTIVE: Gain insight into variables (anatomical sites, occlusion, exposure time) affecting GT delivery into human skin. METHODS: Human skin from different anatomical regions (palmar, plantar, axillary, and abdominal skin) was mounted into flow-through diffusion cells (MedFlux-HT®). GT solution (2.4%) was applied at 10 mg/cm2 and the receiving fluid was collected every 2 hours, for 24 hours. GT penetration was determined using LC/MS/MS. The effect of occlusion was assessed by covering the skin with either parafilm or saran wrap, and the effect of exposure time was assessed by incubating the skin for 5, 15, or 60 minutes before washing off the GT from the surface. RESULTS: GT delivery through palmar and plantar skin was up to 40-fold lower compared to delivery through axillary or abdominal skin. Occlusion increased GT delivery up to 10-fold. Reducing exposure time from 24 hours to either 5, 15, or 60 minutes, decreased GT flux by 90%. However, occlusion during these varied exposure times was able to restore GT delivery to levels found in the 24-hour exposed, non-occluded control group. CONCLUSION: These in vitro skin penetration studies showed that skin thickness, exposure time, and occlusion substantially influenced GT delivery, potentially impacting clinical trial design. J Drugs Dermatol. 2020;19(11): doi:10.36849/JDD.2020.5062.

Olumacostat glasaretil, a novel topical sebum inhibitor, in the treatment of acne vulgaris: A phase IIa, multicenter, randomized, vehicle-controlled study.

BACKGROUND: Olumacostat glasaretil (OG) inhibits acetyl-coenzyme A carboxylase, the enzyme responsible for the first, rate-limiting step in de novo fatty acid synthesis. OG inhibited in vitro human sebocyte lipid production and reduced in vivo sebaceous gland size in hamster ears. OBJECTIVES: Safety and efficacy of OG 7.5% gel were evaluated in patients with moderate to severe facial acne vulgaris. METHODS: Patients were randomized (1:1) to twice-daily application of OG or vehicle for 12 weeks. Efficacy was measured through changes in lesion counts and improvement in acne severity scores. RESULTS: A total of 108 patients received OG (n = 53) or vehicle (n = 55); these groups had mean baseline counts of 29.7 and 28.6 inflammatory and 40.9 and 38.8 noninflammatory lesions, respectively. At week 12, OG treatment showed greater reductions from baseline in inflammatory lesions (-63.9% vs -45.9%; P = .0006) and noninflammatory lesions (-48.1% vs -28.8%; P = .0025), and more patients with greater than or equal to 2-grade improvement in investigator global assessment score (24.5% vs 7.3%; P = .0070) than vehicle. Application-site adverse events (typically mild or moderate intensity) were more common with OG. LIMITATIONS: Larger trials are needed to optimize OG dosing and confirm the current results. CONCLUSION: OG was well tolerated and showed evidence of efficacy, suggesting further development is warranted.

Paired Transcriptomic and Proteomic Analysis Implicates IL-1ß in the Pathogenesis of Papulopustular Rosacea Explants.

Papulopustular rosacea (PPR) is a chronic inflammatory skin disease with limited treatment options. Although multiple pathways have been described to be upregulated in PPR, a mechanistic understanding of the key drivers and interaction between pathways in PPR pathology is lacking. In this study, we utilized PPR skin biopsy explants to integrate both differentially expressed genes and differentially expressed proteins in paired nonlesional and lesional PPR tissue (n = 5 patients). The results of this study identified 92 differentially expressed genes and 20 differentially expressed proteins between paired PPR lesional and nonlesional explants. MAPK and TNF signaling pathways were the most significantly upregulated pathways in PPR lesional tissue and aligned with differently expressed proteins identified in this study. Both MAPK and TNF signaling pathways highlighted IL-1ß as a potential central mediator for PPR pathogenesis. In support of this, stimulation of nonlesional explants with IL-1ß resulted in transcriptomic and proteomic profiles similar to those of lesional PPR. In this integrative transcriptomic and quantitative protein analysis, we identified several inflammatory genes, proteins, and pathways, which may be contributing to PPR, as well as highlighted a potential role of IL-1ß in driving inflammation in PPR.

Inhibition of Sebum Production with the Acetyl Coenzyme A Carboxylase Inhibitor Olumacostat Glasaretil.

Olumacostat glasaretil (OG) is a small molecule inhibitor of acetyl coenzyme A (CoA) carboxylase (ACC), the enzyme that controls the first rate-limiting step in fatty acid biosynthesis. Inhibition of ACC activity in the sebaceous glands is designed to substantially affect sebum production, because over 80% of human sebum components contain fatty acids. OG inhibits de novo lipid synthesis in primary and transformed human sebocytes. TrueMass Sebum Panel analyses showed a reduction in saturated and monounsaturated fatty acyl chains across lipid species, including di- and triacylglycerols, phospholipids, cholesteryl esters, and wax esters in OG-treated sebocytes. There was no shift to shorter acyl chain lengths observed, suggesting that the fatty acid chain elongation process was not affected. OG is a pro-drug of the ACC inhibitor 5-(tetradecyloxy)-2-furoic acid and was designed to enhance delivery in vivo. Topical application of OG but not 5-(tetradecyloxy)-2-furoic acid significantly reduced hamster ear sebaceous gland size, indicating that this pro-drug approach was critical to obtain the desired activity in vivo. High-performance liquid chromatography analyses of hamster ear extracts showed that OG treatment increased ACC levels and the ratio of acetyl-CoA to free CoA in these animals, indicating increased fatty acid oxidation. These changes are consistent with ACC inhibition. Matrix-assisted laser desorption/ionization imaging showed that OG applied onto Yorkshire pig ears accumulated in sebaceous glands relative to the surrounding dermis. Sebaceous gland ACC represents an attractive therapeutic target given its central role in formation of sebum, a key factor in acne pathogenesis.

Early Activation of Th2/Th22 Inflammatory and Pruritogenic Pathways in Acute Canine Atopic Dermatitis Skin Lesions.

Determining inflammation and itch pathway activation in patients with atopic dermatitis (AD) is fraught with the inability to precisely assess the age of skin lesions, thus affecting the analysis of time-dependent mediators. To characterize inflammatory events occurring during early experimental acute AD lesions, biopsy samples were collected 6, 24, and 48 hours after epicutaneous application of Dermatophagoides farinae house dust mites to sensitized atopic dogs. The skin transcriptome was assessed using a dog-specific microarray and quantitative PCR. Acute canine AD skin lesions had a significant up-regulation of genes encoding T helper (Th) 2 (e.g., IL4, IL5, IL13, IL31, and IL33), Th9 (IL9), and Th22 (IL22) cytokines as well as Th2-promoting chemokines such as CCL5 and CCL17. Proinflammatory (e.g., IL6, LTB, and IL18) cytokines were also up-regulated. Other known pruritogenic pathways were also activated: there was significant up-regulation of genes encoding proteases cathepsin S (CTSS), mast cell chymase (CMA1), tryptase (TPS1) and mastin, neuromedin-B (NMB), nerve growth factor (NGF), and leukotriene-synthesis enzymes (ALOX5, ALOX5AP, and LTA4H). Experimental acute canine house dust mite-induced AD lesions exhibit an activation of innate and adaptive immune responses and pruritogenic pathways similar to those seen in humans with acute AD, thereby validating this model to test innovative therapeutics modalities for this disease.

Development of a Topical Treatment for Psoriasis Targeting RORγ: From Bench to Skin.

BACKGROUND: Psoriasis is a chronic inflammatory skin disorder involving marked immunological changes. IL-17-targeting biologics have been successful in reducing the disease burden of psoriasis patients with moderate-to-severe disease. Unfortunately, the stratum corneum prevents penetration of large molecule weight proteins, including monoclonal antibodies. Thus, for the majority of psoriasis patients ineligible for systemic treatments, a small molecule targeting RORγt, the master regulator of IL-17 family cytokines, may represent an alternative topical medicine with biologic-like efficacy. METHODS AND FINDINGS: The preclinical studies described in this manuscript bridge the gap from bench to bedside to provide the scientific foundation for a compound entering clinical trials for patients with mild to moderate psoriasis. In addition to several ex vivo reporter assays, primary T cell cultures, and the imiquimod mouse model, we demonstrate efficacy in a newly developed human ex vivo skin assay, where Th17-skewed cytokine expression is induced from skin-resident immune cells. Importantly, the skin barrier remains intact allowing for the demonstration of topical drug delivery. With the development of this novel assay, we demonstrate potent compound activity in the target tissue: human skin. Finally, target engagement by this small molecule was confirmed in ex vivo lesional psoriatic skin. CONCLUSIONS: Our work describes a progressive series of assays to demonstrate the potential clinical value of a novel RORγ inverse agonist small molecule with high potency and selectivity, which will enter clinical trials in late 2015 for psoriasis patients.

Effects of topically applied acitretin in reconstructed human epidermis and the rhino mouse.

Oral acitretin is currently indicated for the treatment of severe psoriasis in adults, but its use is limited by systemic side effects and teratogenicity. Topical administration of acitretin may lessen the risk of systemic toxicity while increasing local bioavailability in the skin. The effects of topical acitretin on reconstructed human epidermis (RHE) and Rhino mice were investigated and compared to those of currently marketed topical retinoids: tretinoin and tazarotene. In acitretin-treated RHE cultures, there was a reduction in keratohyalin granules and filaggrin expression in the stratum granulosum, a loss of keratin 10 expression in the stratum spinosum, and an increase in keratin 19 expression in all viable cell layers. All retinoids showed similar signs of activity in RHE cultures. Furthermore, the release of pro-inflammatory cytokines IL-1alpha and IL-8 in RHE cultures was less pronounced with acitretin compared to tretinoin- and tazarotene-containing formulations, suggesting that acitretin may be less irritating. In Rhino mice, acitretin induced a local, dose-dependent reduction in utricle diameter after seven daily dermal doses. A similar effect was observed in tretinoin- and tazarotene-treated mice. Our data suggest that topical application of acitretin may have a therapeutic benefit in the local management of keratinization disorders.

Mouse skin models for carcinogenic hazard identification: utilities and challenges.

This report addresses 1) the predictability of mouse skin models for carcinogenic hazard identification, 2) the association between early changes in the skin and later tumorigenic responses, and 3) the relative sensitivity of three mouse models of skin tumorigenesis; i.e. the genetically-initiated Tg.AC and RasH2 lines and the SENCAR mouse model. All three mouse models responded similarly, with mild inflammation and epidermal hyperplasia, to several weeks of treatment with a topical agent. Based on our previous research experience, we hypothesized that inflammation, irritation, proliferation, and/or hyperplasia in the skin would precede and predict the appearance of tumors in these sensitive mouse skin models. Consistent with our hypothesis, the test agent caused a low but significant tumorigenic response in Tg.AC mice. We propose that inflammation, irritation, and hyperplasia are sensitive predictors of a later tumorigenic response in Tg.AC mice. Further studies are needed, however, to better determine the relative sensitivity of these 3 models to a wider variety of agents.

Folate-targeted gene transfer in vivo.

Nonviral systemic delivery is one of the most attractive approaches for cancer gene therapy. To achieve this goal, various laboratories have developed cationic liposomes. However, when injected intravenously, cationic lipid-DNA complexes accumulate mostly into and transfect lung tissue. Here, we describe a method by which these complexes can be targeted to tumors using folic acid. Adding polyethylene glycol (PEG)-lipids to the complexes dramatically reduced both lung accumulation and gene transfer to lungs and tumors after intravenous administration. The presence of folic acid at the distal end of the PEG-lipid did not modify tumor accumulation of the complexes. However, with folate-targeted complexes, gene transfer activity was restored in tumors while the activity in lungs was reduced by 50- to 100-fold compared with nontargeted lipid-DNA complexes. This approach provides a first in vivo proof of concept to achieve targeted tumor gene delivery.