The primary target of halobetasol is the cytosolic glucocorticoid receptor (GR), a member of the nuclear receptor superfamily. Halobetasol binds to GR with high affinity (Kd ~ 1-5 nM), inducing a conformational change that releases heat shock proteins and allows the receptor-ligand complex to translocate to the nucleus. In the nucleus, the complex binds to glucocorticoid response elements (GREs) in the promoter regions of target genes, leading to activation (transactivation) or repression (transrepression) of gene expression. Transactivation results in increased synthesis of anti-inflammatory proteins (e.g., lipocortin-1/annexin A1, IL-10, IkappaBalpha) that inhibit phospholipase A2 (PLA2) and NF-kappaB signaling. Transrepression involves interference with other transcription factors (NF-kappaB, AP-1) to suppress the expression of pro-inflammatory cytokines (IL-1, IL-2, IL-6, TNF-alpha, IFN-gamma), chemokines, adhesion molecules, and inflammatory enzymes (COX-2, iNOS). The net effect is broad suppression of the immune response and inflammation. Halobetasol also induces vasoconstriction (as measured by skin blanching assays), reduces capillary permeability, and inhibits leukocyte infiltration and fibroblast proliferation. Halobetasol has approximately 600-1000 times the potency of hydrocortisone in vasoconstrictor assays (Stoughton-McKenzie assay).

In vitro studies using human dermal fibroblasts, keratinocytes, and immune cells (e.g., T lymphocytes, monocytes, macrophages) demonstrate that halobetasol (0.1-100 nM) potently suppresses pro-inflammatory cytokine production. For example, in LPS-stimulated human peripheral blood mononuclear cells (PBMCs), halobetasol (1-10 nM) reduces TNF-alpha, IL-1beta, IL-6, and IL-8 by 80-90% (ELISA). In T-cell proliferation assays, halobetasol (0.1-10 nM) inhibits anti-CD3/anti-CD28-induced proliferation (3H-thymidine incorporation) with an IC50 of 0.5-1 nM. In keratinocytes, halobetasol (0.1-10 nM) inhibits interferon-gamma (IFN-gamma)-induced expression of MHC class II and ICAM-1 (flow cytometry). In a human dermal explant model, halobetasol (0.05% cream, topical application) reduces UVB-induced erythema (redness) by 70-80% compared to vehicle control, as measured by chromametry. Halobetasol also inhibits prostaglandin E2 (PGE2) production in A549 lung epithelial cells (IC50 ~ 0.3 nM). However, because halobetasol is used topically, in vitro effects at concentrations above 100 nM may be relevant to local skin exposure rather than systemic exposure.

In vivo studies in animal models of inflammation and psoriasis have demonstrated the efficacy of topical halobetasol. In the mouse ear edema model (12-O-tetradecanoylphorbol-13-acetate (TPA)-induced inflammation), topical application of halobetasol (0.05% cream, 10 mg/ear) reduces ear swelling (thickness measured by micrometer) by 80-90% at 6 hours post-application compared to vehicle control (p < 0.001). In the rat carrageenan-induced paw edema model, topical halobetasol (0.05% ointment) reduces paw volume by 60-70% at 4 hours. In an imiquimod-induced psoriasis-like mouse model (daily topical application of imiquimod 5% cream for 5-7 days to the back skin), co-treatment with halobetasol (0.05% cream, daily) significantly reduces psoriasis-like features: erythema, scaling, and induration scores (as measured by the Psoriasis Area and Severity Index (PASI)-like scoring system) are reduced by 70-80% compared to vehicle. Histological analysis of skin biopsies (H&E staining) shows reduced epidermal thickening (acanthosis), decreased parakeratosis, reduced inflammatory cell infiltration (neutrophils, T cells, macrophages), and normalization of keratinocyte differentiation (keratin 16, filaggrin). In murine models of allergic contact dermatitis (oxazolone-induced), halobetasol (0.05% cream) reduces ear swelling by 80-90% and reduces inflammatory cytokine levels (IL-4, IL-5, IL-13) in skin homogenates by 60-80%. In rabbits, topical application of halobetasol (0.05% cream, 0.5 g, twice daily for 14 days) on intact or abraded skin causes mild skin atrophy (thinning of the epidermis) and telangiectasia (dilated blood vessels) by day 14, with recovery by day 28 after cessation. Vasoconstrictor assays in healthy human volunteers (Stoughton-McKenzie assay, 0.05% cream applied to forearm under occlusion for 16 hours) produce skin blanching (vasoconstriction) scores of 3-4 (on a 0-4 scale), confirming high potency.

In vitro receptor binding assays: Competitive binding assay for glucocorticoid receptor (GR). The cytosolic fraction of A549 cells (or recombinant human GR protein, 1-5 nM) is incubated with 3H-dexamethasone (1 nM, specific activity 40-80 Ci/mmol) in the presence of increasing concentrations of halobetasol (0.01-1000 nM) in binding buffer (10 mM Tris-HCl pH 7.4, 10 mM sodium molybdate, 1 mM DTT, 2 mM EDTA, 10% glycerol) for 2 hours at 4degC. Non-specific binding is determined in the presence of 10 uM unlabeled dexamethasone. Bound and free 3H-dexamethasone are separated by charcoal dextran precipitation (0.5% charcoal, 0.05% dextran) or by filtration through glass fiber filters. Radioactivity (cpm) is measured by liquid scintillation counting. IC50 values are calculated, and Ki is determined using the Cheng-Prusoff equation (Ki = IC50 / (1 + [radioligand]/Kd of radioligand)). For halobetasol propionate, Ki values are typically 1-2 nM. For transactivation assays, HEK293 cells are co-transfected with a GRE-luciferase reporter plasmid (e.g., pGRE-Luc) and a GR expression plasmid using Lipofectamine 3000. After 24 hours, cells are treated with halobetasol (0.01-1000 nM) for 16-24 hours, and luciferase activity is measured using a dual-luciferase assay kit (Promega). EC50 for transactivation is 0.5-1 nM. For transrepression assays, A549 cells (endogenously expressing GR) are transfected with an NF-kappaB-luciferase reporter plasmid (pNFkappaB-Luc). After 24 hours, cells are pre-treated with halobetasol (0.1-1000 nM) for 2 hours, then stimulated with TNF-alpha (10 ng/mL) for 6 hours. Luciferase activity is measured, and IC50 for inhibition of NF-kappaB-driven transcription is determined (typically 0.2-0.5 nM).

Cell Viability Assay[2]
Cell Types: HaCaT, THP-1 Tested
Tested Concentrations: 0.05-12.85 μM
Incubation Duration: 24 h
Experimental Results: Affected decreasing the viability in both cell lines up to 50% corresponding to dilution 1/20.

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Cell culture: Human keratinocytes (e.g., HaCaT, NHEK) are cultured in keratinocyte growth medium (KGM) or DMEM (low calcium) with 10% FBS (or defined supplements) at 37degC, 5% CO2. For cytokine induction studies, cells are seeded in 6-well plates (2 × 10⁵ cells/well) and allowed to attach for 24 hours. Cells are pre-treated with halobetasol (0.1-100 nM, in DMSO, final DMSO 0.01%) for 2 hours, then stimulated with TNF-alpha (10 ng/mL) or IL-1beta (10 ng/mL) for 6-24 hours. Culture supernatants are collected, and cytokine levels (IL-6, IL-8, MCP-1, GM-CSF) are measured by ELISA (R&D Systems). Cells are also harvested for RNA extraction (TRIzol) and qPCR analysis (primers for IL-6, IL-8, MCP-1, COX-2). For proliferation assays, human dermal fibroblasts (HDF) are seeded in 96-well plates (5 × 103 cells/well) and treated with halobetasol (0.1-1000 nM) for 48-72 hours. Cell proliferation is measured by MTT or 3H-thymidine incorporation (0.5 microCi/well, 6-hour pulse). IC50 for inhibition of fibroblast proliferation is typically 0.5-1 nM. For apoptosis studies, keratinocytes or T cells are treated with halobetasol (1-100 nM) for 24-48 hours, stained with Annexin V-FITC/PI, and analyzed by flow cytometry. Halobetasol does not induce significant apoptosis at concentrations up to 100 nM (<5% Annexin V+ compared to vehicle). For Western blotting, cells are lysed in RIPA buffer with protease and phosphatase inhibitors, and 20-40 ug of protein is separated on 10-12% SDS-PAGE, transferred to PVDF membranes, and probed with antibodies for GR, IkappaBalpha, NF-kappaB p65, COX-2, and beta-actin.

Animal/Disease Models: New Zealand male albino rabbits[2]
Doses: 0.5 ml
Route of Administration: Intradermal injection
Experimental Results: Generated blisters were approximately 35% smaller.

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Animal models: For mouse ear edema (acute inflammation), female CD-1 mice (6-8 weeks, 20-25 g) are used. Halobetasol (0.05% cream, 10 mg/ear) or vehicle cream is applied to the dorsal and ventral surfaces of the right ear. After 30 minutes, TPA (2.5 ug/ear in 20 uL acetone) is applied to both surfaces of the right ear (left ear receives acetone only). After 6 hours, mice are euthanized, and ear punch biopsies (6 mm diameter) are taken. Ear thickness is measured before and after TPA using a micrometer. Ear weight (mg) is measured. Edema is calculated as (treated ear weight - untreated ear weight). For imiquimod-induced psoriasis-like model, female BALB/c mice (6-8 weeks, 20-25 g) are shaved on the back (2 × 2 cm area). Imiquimod cream (5%, 62.5 mg/day) is applied topically to the back skin once daily for 6 consecutive days. For treatment groups, halobetasol (0.05% cream, 100 mg/mouse) or vehicle cream is applied to the same area 2 hours before imiquimod application (or 2 hours after, depending on the study design). Disease severity is assessed daily using the PASI-like scoring system: erythema (0-4), scaling (0-4), and thickening (0-4), summed to a total score (0-12). On day 6, mice are euthanized, and dorsal skin is excised for histology (H&E, Ki-67, CD4, CD8, F4/80, Gr-1) and biochemical analyses (myeloperoxidase (MPO) activity, cytokine levels by ELISA). For chronic toxicity studies in minipigs (a model that better predicts human skin response), halobetasol (0.05% cream, 0.5 g/site) is applied topically to intact and abraded skin (two 2 × 2 cm sites on the flank) once or twice daily for 28 days. Skin biopsies are taken at baseline, day 14, day 28, and after a 14-day recovery period, and are evaluated for epidermal thickness (um), stratum corneum thickness, dermal inflammation, and collagen deposition. Systemic absorption is assessed by measuring plasma cortisol levels (suppression of HPA axis is a marker of systemic exposure).

Absorption, Distribution and Excretion
The peak plasma concentration (Cmax) of urobetasol emulsion was 201.1 ± 157.5 pg/mL, the time to peak concentration (Tmax) was 3 hours, and the area under the curve (AUC) was 1632 ± 1147 pgh/mL. Skin integrity, excipients used, inflammation, or disease processes can all affect absorption. Corticosteroids are primarily excreted in the urine. Biological Half-Life In vitro studies showed that the half-life of urobetasol's derivative, halobetasol propionate, is 33 minutes.

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In humans, halobetasol propionate (0.05% cream, ointment, lotion) is applied topically twice daily for up to 2 weeks. Percutaneous absorption is low (approximately 2-6% of the applied dose systemically absorbed over 96 hours) due to the high potency and formulation properties. However, systemic absorption can increase with occlusive dressings, prolonged use (>2 weeks), application to large body surface areas (>20% BSA), or use on compromised skin (abraded, inflamed, or diseased skin). Peak plasma concentrations (Cmax) after twice-daily application of 0.05% cream to 20% BSA for 14 days are typically <0.5 ng/mL (approx 1 nM), which is below the concentration required for significant systemic glucocorticoid effects (e.g., HPA axis suppression). The half-life in plasma is approximately 6-8 hours. Halobetasol is metabolized primarily in the liver by CYP3A4 to inactive metabolites (6beta-hydroxy-halobetasol, 20-carboxy-halobetasol) and glucuronide conjugates. Less than 2% of an applied dose is excreted unchanged in urine. Renal clearance is 2-3 mL/min/kg. Systemic absorption is higher with the ointment formulation (due to occlusive properties) compared to cream or lotion. The drug is extensively bound to plasma proteins (>95%). In patients with hepatic impairment, systemic clearance may be reduced. In patients with renal impairment, no specific PK studies have been conducted. Halobetasol is not recommended for use in children <12 years of age due to the risk of HPA axis suppression and growth retardation.

Protein Binding
Urobetasol may bind to corticosteroid-binding globulin in serum.

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Halobetasol is a highly potent corticosteroid, and adverse effects are primarily related to its glucocorticoid activity. Local adverse effects (incidence 10-20%) include: skin atrophy (thinning, striae, telangiectasia), burning/stinging sensation, pruritus (itching), dryness, folliculitis, acneiform eruptions, hypopigmentation (lightening of skin), perioral dermatitis, and allergic contact dermatitis. Prolonged use (>2 weeks) or use on large body surface areas (>20% BSA) can lead to systemic absorption and HPA axis suppression (adrenal insufficiency), Cushing's syndrome (moon face, buffalo hump, weight gain, hyperglycemia, hypertension), growth retardation in children, osteoporosis, cataracts, glaucoma, and immunosuppression (increased risk of infections). In clinical trials, the incidence of HPA axis suppression after 2 weeks of halobetasol 0.05% cream applied twice daily to 20% BSA is 5-10% as measured by cosyntropin stimulation test (plasma cortisol <18 microg/dL at 30-60 min). The drug should not be used during pregnancy unless clearly needed (FDA pregnancy category C). In animal studies (rats, rabbits), topical halobetasol (0.05% cream, 0.2-1 mg/kg/day) causes embryotoxicity (resorptions, reduced fetal weight) and teratogenicity (cleft palate, skeletal anomalies) at doses that produce maternal toxicity. There are no adequate and well-controlled studies in pregnant women. Halobetasol is excreted in breast milk, and systemic absorption after topical application in nursing mothers is low, but caution is advised. The drug is contraindicated in patients with known hypersensitivity to halobetasol or any component of the formulation. It should not be used on the face, axillae (armpits), groin, or for diaper dermatitis. Overdose (accidental ingestion) may cause nausea, vomiting, hyperglycemia, hypertension, electrolyte imbalances, and Cushing's syndrome; treatment is symptomatic and supportive. Halobetasol is not associated with genotoxicity (AMES test negative, in vivo micronucleus negative) or carcinogenicity (long-term studies in mice and rats show no increased tumor incidence).

[1]. Schwicker D, et al. A cost-comparison study: Ulobetasol versus clobetasol in severe localized psoriasis. J Dermatolog Treat. 1992;2(4):127-131.

Halobetasol is a corticosteroid. Halobetasol is a potent corticosteroid with a structure related to clobetasol. Due to its high potency, it is primarily used to treat severe plaque psoriasis and corticosteroid-sensitive skin diseases. Halobetasol was approved by the U.S. Food and Drug Administration (FDA) on December 17, 1990. Halobetasol is a corticosteroid. Its mechanism of action is as a corticosteroid hormone receptor agonist. Halobetasol is a synthetic corticosteroid with anti-inflammatory, antipruritic, and vasoconstrictive effects. Halobetasol is a topical steroid that diffuses across cell membranes and interacts with cytoplasmic corticosteroid receptors located in dermal and intradermal cells, thereby activating the expression of anti-inflammatory protein genes mediated by corticosteroid receptor response elements. Specifically, the drug induces the expression of phospholipase A2 inhibitory protein, thereby inhibiting the release of arachidonic acid, and subsequently inhibiting the biosynthesis of potent inflammatory mediators such as prostaglandins and leukotrienes. Therefore, urobetasol reduces edema, erythema, and itching through its vasodilatory and permeable skin effects.
See also: urobetasol propionate (active ingredient).
Indications
Urobetasol cream and ointment are indicated for the treatment of inflammatory and pruritic skin conditions that respond to corticosteroids. Urobetasol lotion is indicated for the treatment of plaque psoriasis.
FDA Label
Mechanism of Action
The short-term effects of corticosteroids are to reduce capillary vasodilation and permeability, and to reduce the migration of leukocytes to sites of inflammation. Corticosteroids bind to glucocorticoid receptors, mediating alterations in gene expression that produce a variety of downstream effects over hours to days. Glucocorticoids inhibit phospholipase 2 and neutrophil apoptosis and marginalization, leading to reduced production of arachidonic acid derivatives. They also inhibit NF-κB and other inflammatory transcription factors while promoting the expression of anti-inflammatory genes such as interleukin-10. Low-dose corticosteroids have anti-inflammatory effects, while high-dose corticosteroids have immunosuppressive effects. Long-term use of high-dose glucocorticoids can bind to mineralocorticoid receptors, leading to increased sodium and decreased potassium levels.

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Pharmacodynamics
Corticosteroids bind to glucocorticoid receptors, inhibiting pro-inflammatory signaling and promoting anti-inflammatory signaling. Urobetasol has a moderate duration of action and only needs to be used once or twice daily. Corticosteroids have a wide therapeutic window, and patients may require doses several times higher than the body's naturally produced levels. Patients taking corticosteroids should be informed of the risks of hypothalamic-pituitary-adrenal axis suppression and increased susceptibility to infection.

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Halobetasol (Ulobetasol) is a generic drug available under brand names such as Ultravate® (Halobetasol Propionate Cream, Ointment, Lotion) and Lexette® (Halobetasol Propionate Foam). Halobetasol propionate was first approved by the FDA in 1991 (brand Ultravate) for the treatment of moderate to severe plaque psoriasis and corticosteroid-responsive dermatoses. The product is available as 0.05% cream, 0.05% ointment, 0.05% lotion, and 0.05% foam (Lexette, approved 2018). Halobetasol is also available in fixed-dose combination products (e.g., with tazarotene 0.045% and emollients) for the treatment of plaque psoriasis (Duobrii®). The drug is on the WHO Model List of Essential Medicines (topical corticosteroids). It is typically used as second-line therapy after lower-potency corticosteroids (e.g., hydrocortisone, triamcinolone) have failed, or as initial therapy for severe, localized psoriasis. Treatment duration is limited to 2 weeks to minimize the risk of local and systemic adverse effects. The compound has a high therapeutic index (potency vs toxicity) when used appropriately. The compound is available as a white to off-white crystalline powder. It is insoluble in water but soluble in organic solvents (ethanol, DMSO). Halobetasol propionate (CAS 66852-54-0) is the clinically used ester. The parent alcohol halobetasol (CAS 98651-66-2) is the active moiety and is listed in chemical databases. It should be stored at room temperature (20-25degC), protected from light and moisture. In research settings, halobetasol is used as a reference standard for HPLC analysis of topical corticosteroid formulations, as a positive control in anti-inflammatory assays, and for studying glucocorticoid receptor signaling. The compound is not for human use outside of approved medical indications and is available only by prescription. It is also used in veterinary medicine (dogs, cats) for inflammatory skin conditions, but safety and efficacy studies in animals are limited. For research use, the compound should be handled with care (gloves, lab coat) and used in accordance with institutional biosafety guidelines. Halobetasol is a scheduled substance in some jurisdictions due to its potent glucocorticoid activity.

C22H27CLF2O4

428.90

428.157

98651-66-2

5311167

White to off-white solid powder

200-216

3.09

2

6

2

29

843

9

CC1CC2C3CC(C4=CC(=O)C=CC4(C3(C(CC2(C1(C(=O)CCl)O)C)O)F)C)F

LEHFPXVYPMWYQD-XHIJKXOTSA-N

InChI=1S/C22H27ClF2O4/c1-11-6-13-14-8-16(24)15-7-12(26)4-5-19(15,2)21(14,25)17(27)9-20(13,3)22(11,29)18(28)10-23/h4-5,7,11,13-14,16-17,27,29H,6,8-10H2,1-3H3/t11-,13-,14-,16-,17-,19-,20-,21-,22-/m0/s1

(6S,8S,9R,10S,11S,13S,14S,16S,17R)-17-(2-chloroacetyl)-6,9-difluoro-11,17-dihydroxy-10,13,16-trimethyl-6,7,8,11,12,14,15,16-octahydrocyclopenta[a]phenanthren-3-one

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)