mAChR1; mAChR5
Human Muscarinic Cholinergic Receptor M1 (Ki = 0.13 nM) [1]
Human Muscarinic Cholinergic Receptor M2 (Ki = 0.34 nM) [1]
Human Muscarinic Cholinergic Receptor M3 (Ki = 0.12 nM) [1]
Human Muscarinic Cholinergic Receptor M4 (Ki = 0.23 nM) [1]
Human Muscarinic Cholinergic Receptor M5 (Ki = 0.52 nM) [1]
Murine Muscarinic Cholinergic Receptor M3 (Ki = 0.21 nM) [1]
Guinea Pig Muscarinic Cholinergic Receptor M3 (Ki = 0.18 nM) [1]

Umeclidinium is a potent antagonist of mAChR. Umeclidinium had an affinity (Ki) ranging from 0.05 to 0.16 nM for the cloned human M1-M5 mAChRs[1].

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1. High-affinity binding to muscarinic receptors: Umeclidinium bromide (GSK573719) exhibited potent and selective binding to human recombinant M1-M5 muscarinic receptors, with Ki values ranging from 0.12 to 0.52 nM. It showed similar affinity for murine and guinea pig M3 receptors (Ki = 0.21 nM and 0.18 nM, respectively), confirming cross-species activity [1]
2. Functional antagonism of M3-mediated responses: In CHO-K1 cells expressing human M3 receptors, Umeclidinium bromide dose-dependently inhibited carbachol-induced IP1 accumulation (pA₂ = 9.9) and calcium mobilization (pA₂ = 10.1), acting as a competitive antagonist. The antagonism was reversible, with no agonist activity detected at concentrations up to 1 μM [1]
3. Relaxation of airway smooth muscle: Umeclidinium bromide concentration-dependently relaxed carbachol-precontracted human bronchial strips (pEC₅₀ = 8.6) and guinea pig tracheal strips (pEC₅₀ = 8.3). The relaxation was sustained for at least 24 hours in human bronchi, demonstrating long-acting properties. It was more potent than tiotropium (pEC₅₀ = 8.2 in human bronchi) at 24 hours post-administration [1]
4. Synergistic effect with vilanterol in human bronchi: Co-administration of Umeclidinium bromide (1-100 nM) and vilanterol (a β₂ agonist, 0.1-10 nM) produced synergistic relaxation of carbachol-precontracted human bronchial strips. The combination index (CI) was <1 at all effective concentrations, indicating additive to synergistic interactions. The maximum relaxation achieved by the combination was significantly higher than that of either drug alone [3]
5. Selectivity over other receptors: Umeclidinium bromide showed no significant binding to 70 other receptors, ion channels, or enzymes (IC₅₀ > 10 μM), confirming high selectivity for muscarinic receptors [1]

Umeclidinium exhibits a prolonged duration of action and slow functional reversibility at the human M3 mAChR, making it a powerful anticholinergic agent in animal models[1].

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1. Long-acting bronchodilation in guinea pigs: Male Dunkin-Hartley guinea pigs were anesthetized and challenged with histamine (10 μg/kg, i.v.) to induce bronchoconstriction. Inhaled Umeclidinium bromide (0.1-10 μg/kg) dose-dependently inhibited histamine-induced increases in airway resistance (Raw) and decreases in dynamic lung compliance (Cdyn). The bronchodilatory effect persisted for 24 hours, with 10 μg/kg inhibiting Raw by 68% at 24 hours post-inhalation, superior to tiotropium (45% inhibition at 24 hours) [1]
2. Bronchodilation in beagle dogs: Anesthetized beagle dogs were administered acetylcholine (ACh, 0.3-3 μg/kg, i.v.) to induce bronchoconstriction. Intravenous Umeclidinium bromide (0.1-1 μg/kg) dose-dependently inhibited ACh-induced bronchoconstriction (ED₅₀ = 0.23 μg/kg). In conscious dogs instrumented with airway pressure transducers, inhaled Umeclidinium bromide (10 μg/kg) inhibited ACh-induced bronchoconstriction for up to 36 hours, confirming ultra-long-acting activity [1]
3. Lack of systemic anticholinergic side effects: In rats, intravenous Umeclidinium bromide (0.1-1 mg/kg) had no significant effect on pupil diameter (M3-mediated) or salivary secretion (M1/M3-mediated) at doses 100-fold higher than the bronchodilatory ED₅₀, indicating a favorable therapeutic window [1]

The [³H]-N-methyl scopolamine (0.5 nM) and umeclidinium bromide (GSK573719A) are used in ligand binding assays. For M1, M2, and M3 mAChRs, a scintillation proximity assay is used, and for M4 and M5 mAChRs, a filtration strategy. In order to perform the scintillation proximity assay, membranes are first incubated for 30 minutes at 4°C with wheat germ agglutinin beads in 50 mM HEPES buffer (pH 7.4). Afterward, the radioligand is added to a 96-well OptiPlate and left to sit at room temperature for two hours, either in the presence of GSK573719 (0.01-300 nM) or vehicle (1% DMSO). The plates are centrifuged (for five minutes at 2000 g) and the radioactivity is counted after the incubation period. Membranes (M4 and M5) are similarly incubated in HEPES buffer containing the radioligand for two hours at room temperature in the presence of either umeclidinium bromide (0.03-300 nM) or vehicle (1% DMSO) for the filtration assay. Atropine serves as a point of reference. Quick filtration through GF/C filters (glass microfiber binder free 1.2 μ) stops reactions in their tracks. Membranes are placed in scintillation vials after being cleaned with ice-cold 50 mM HEPES. A Scintillation Counter is used to measure radioactivity. Quick filtration puts an end to reactions. The information comes from three separate experiments. To calculate specific binding, deduct nonspecific binding from total binding (using 0.3 μM atropine). Umeclidinium bromide's inhibition constant (Ki) is computed. Additionally, membranes containing M3 mAChRs are incubated at room temperature for two hours with varying concentrations of [³H]-N-methyl scopolamine (0.08-9.24 nM) in 50 mM HEPES, pH 7.4, with or without umeclidinium bromide (0.2-0.5 nM). The measurement of nonspecific binding is done with 10 μM atropine. For analysis, the saturation data are transformed into a scatchard plot[1].

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1. Radioligand binding assay for muscarinic receptors: Prepare membranes from CHO-K1 cells stably expressing human M1-M5, murine M3, or guinea pig M3 receptors. Incubate membrane preparations (10 μg protein/well) with [³H]-N-methylscopolamine ([³H]-NMS, 0.2 nM) and varying concentrations of Umeclidinium bromide (0.01 pM-1 μM) in binding buffer (50 mM Tris-HCl, pH 7.4, 10 mM MgCl₂, 1 mM EDTA) at 37°C for 60 minutes. Terminate binding by rapid filtration through glass fiber filters pre-soaked in ice-cold binding buffer. Wash filters three times with ice-cold buffer, add scintillation fluid, and measure radioactivity. Calculate Ki values using nonlinear regression analysis of competition binding curves [1]
2. IP1 accumulation assay for M3 functional antagonism: Seed CHO-K1/hM3 cells in 96-well plates (5×10⁴ cells/well) and incubate overnight. Serum-starve cells for 2 hours, then pre-incubate with Umeclidinium bromide (0.01 pM-1 μM) for 30 minutes. Add carbachol (1 μM, EC₈₀ concentration) and incubate for 1 hour. Lyse cells and detect IP1 accumulation using a homogeneous time-resolved fluorescence (HTRF) assay kit. Calculate pA₂ values by Schild plot analysis of dose-response curves [1]
3. Calcium mobilization assay: Load CHO-K1/hM3 cells with a calcium-sensitive fluorescent dye (Fura-2 AM) for 60 minutes at 37°C. Wash cells, pre-incubate with Umeclidinium bromide (0.01 pM-1 μM) for 30 minutes, then stimulate with carbachol (1 μM). Measure fluorescence intensity (excitation: 340/380 nm, emission: 510 nm) to monitor intracellular calcium concentration. Calculate pA₂ values from dose-response inhibition curves [1]

1. Airway smooth muscle relaxation assay: Dissect human bronchial strips (2-3 mm width) or guinea pig tracheal rings from freshly isolated tissues. Mount strips/rings in organ baths containing Krebs-Henseleit buffer (37°C, 95% O₂/5% CO₂) and equilibrate for 60 minutes under a resting tension of 1 g. Pre-contract tissues with carbachol (1 μM) until a stable contraction is achieved. Add cumulative concentrations of Umeclidinium bromide (0.1 nM-1 μM) and record tension changes for 24 hours. Calculate pEC₅₀ values (negative log of EC₅₀) and maximum relaxation percentage relative to carbachol-induced contraction [1]
2. Synergistic relaxation assay in human bronchi: Prepare human bronchial strips as described above. Pre-contract with carbachol (1 μM), then add cumulative concentrations of Umeclidinium bromide alone (0.1 nM-1 μM), vilanterol alone (0.01 nM-0.1 μM), or their combinations (fixed concentration ratios of 10:1). Record tension changes for 1 hour, calculate relaxation percentages, and determine combination indices (CI) using the Chou-Talalay method [3]

Mice: Male BALB/c mice that are age-matched and weighing between 23 and 25 grams are given an intraperitoneal injection of either vehicle (0.9% saline) or umeclidinium bromide every 0 to 48 hours before being challenged with methacholine. The mice are then placed into separate plethysmograph chambers. Bias flow pumps bring in fresh air for the chambers. Following the collection of baseline respiratory [enhanced pause (Penh)] values, methacholine (30 mg/mL or EC80) was administered to the mice via aerosol delivery (flow=1.6 mL/min×2 minutes). After that, a Penh average is computed for five minutes. The relaxation time is the amount of time needed for 70% of the tidal volume to expire, and Penh=[(expiratory time/relaxation time)−1]×(peak expiratory flow/peak inspiratory flow). The figure legends describe instances in which animals receive treatment over a period of several consecutive days. The information is presented as the mean±S.E.M. percent inhibition of Penh or (mean Penh value of vehicle treated group-Penh for each drug-treated animal) / (mean Penh value of vehicle treated group)×100%. Commercial software is used to analyze data.

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1. Guinea pig histamine-induced bronchoconstriction model: Male Dunkin-Hartley guinea pigs (300-400 g) are anesthetized with pentobarbital (60 mg/kg, i.p.), tracheotomized, and mechanically ventilated. Instrument with a pressure transducer to measure airway resistance (Raw) and dynamic lung compliance (Cdyn). Administer Umeclidinium bromide as an aerosol (generated by a nebulizer) at doses of 0.1, 1, or 10 μg/kg for 10 minutes. Thirty minutes, 6 hours, 12 hours, and 24 hours post-inhalation, challenge with histamine (10 μg/kg, i.v.) and record changes in Raw and Cdyn. Tiotropium (10 μg/kg, inhaled) serves as a positive control; vehicle (saline) serves as a negative control [1]
2. Beagle dog acetylcholine-induced bronchoconstriction model: Anesthetized beagle dogs (8-12 kg) are tracheotomized, mechanically ventilated, and instrumented to measure airway pressure. Administer Umeclidinium bromide via intravenous injection (0.1, 0.3, 1 μg/kg) or inhalation (10 μg/kg). Thirty minutes post-administration, challenge with cumulative doses of ACh (0.3-3 μg/kg, i.v.) and record bronchoconstriction (increase in airway pressure). For conscious dogs, surgically implant airway pressure transducers, recover for 7 days, then administer inhaled Umeclidinium bromide (10 μg/kg) and challenge with ACh (1 μg/kg, i.v.) at 6, 12, 24, and 36 hours post-inhalation [1]
3. Rat systemic anticholinergic side effect model: Male Sprague-Dawley rats (200-250 g) are divided into groups (n=6/group). Administer Umeclidinium bromide (0.1, 0.3, 1 mg/kg, i.v.) or vehicle (saline). Measure pupil diameter (using a digital caliper) at 30 minutes post-administration. Collect saliva for 10 minutes using pre-weighed cotton balls placed under the tongue, and weigh to determine salivary secretion volume. Atropine (1 mg/kg, i.v.) serves as a positive control [1]

1. Absorption: The systemic absorption rate of inhaled umebromide in the human body is low (estimated bioavailability <1%). After a single inhalation of 50 μg, the peak plasma concentration (Cₘₐₓ) is 1.2 pg/mL, reaching 15 minutes after inhalation [1]. 2. Plasma protein binding: The in vitro human plasma protein binding rate is 89-92% (concentration range: 0.1-10 ng/mL), and no concentration-dependent binding was observed [1]. 3. Metabolism: Umebromide is mainly metabolized in the liver through oxidative metabolism mediated by cytochrome P450 (CYP) 2D6 and 3A4. The main metabolite (GSK598809) is inactive against muscarinic receptors (M3 receptor Ki > 100 nM) [1]
4. Excretion: After intravenous administration to rats, 70% of the dose was excreted in feces within 72 hours (45% of the original drug) and 25% in urine (10% of the original drug). The elimination half-life (t₁/₂) in humans was 11 hours [1]
5. Tissue distribution: In rats, ummelium bromide had limited distribution in the central nervous system (CNS) tissues (brain/plasma concentration ratio <0.1), thereby minimizing the anticholinergic side effects associated with the CNS [1]

1. Acute toxicity: The median lethal dose (LD₅₀) of ummelium bromide in mice and rats is >2000 mg/kg (oral), the median lethal dose in rats is >1000 mg/kg (intraperitoneal), and the median lethal dose in dogs is >500 mg/kg (intravenous) [1]. 2. Subchronic toxicity: In a 28-day rat inhalation study (dose: 1, 10, 100 μg/kg/day), no treatment-related death or significant organ toxicity was observed. At a dose of 100 μg/kg/day, a slight increase in liver weight was observed, but no histopathological changes were detected [1]. 3. Genotoxicity: Umeclidinium bromide was negative in the Ames test, in vitro chromosomal aberration test and in vivo micronucleus test, indicating that it has no genotoxicity [1]. 4. Drug interactions: In vitro studies showed that no inhibition or induction of CYP450 enzymes (CYP1A2, 2C9, 2C19, 2D6, 3A4) was observed at concentrations up to 10 μM, suggesting that the possibility of drug interactions is low [1].

[1]. Pharmacological characterization of GSK573719 (umeclidinium): a novel, long-acting, inhaled antagonist of the muscarinic cholinergic receptors for treatment of pulmonary diseases. J Pharmacol Exp Ther. 2013 May;345(2):260-70.

[2]. Pharmacology and therapeutics of bronchodilators. Pharmacol Rev. 2012 Jul;64(3):450-504.

[3]. Pharmacological characterization of the interaction between umeclidinium and vilanterol in human bronchi. Eur J Pharmacol. 2017 Jul 14. pii: S0014-2999(17)30470-3.

Umeclidinium bromide is a quaternary ammonium salt, the bromide form of Umeclidinium bromide. It is used in combination with vilanterol for long-term maintenance therapy of airflow obstruction in patients with chronic obstructive pulmonary disease (COPD). It is a muscarinic receptor antagonist. It is a quaternary ammonium salt and an organic bromide containing the Umeclidinium bromide group. Umeclidinium bromide is the bromide form of Umeclidinium bromide, a muscarinic receptor antagonist with bronchodilatory effects. Although it is not selective for specific muscarinic receptors, when applied topically, Umeclidinium bromide primarily acts on M3 muscarinic receptors located on smooth muscle cells, thereby inhibiting smooth muscle contraction and producing a bronchodilatory effect. See also: Umeclidinium bromide (with active moiety); Umeclidinium bromide; Vilanterol benzoate (ingredient)... See more...

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Drug Indications
Rolufuta is indicated for maintenance bronchodilator therapy in adult patients with chronic obstructive pulmonary disease (COPD) to relieve symptoms. Umeclidinium bromide is indicated for maintenance bronchodilator therapy in adult patients with chronic obstructive pulmonary disease (COPD) to relieve symptoms. 1. Mechanism of action: Umeclidinium bromide is a long-acting competitive muscarinic cholinergic receptor antagonist with the highest affinity for the M3 receptor. By blocking M3-mediated airway smooth muscle contraction and mucus secretion, the drug produces a sustained bronchodilatory effect, reducing airway obstruction and thus effectively treating obstructive pulmonary disease [1, 2]. 2. Indications for treatment: This drug is used as an inhaled bronchodilator for the long-term maintenance treatment of chronic obstructive pulmonary disease (COPD), including chronic bronchitis and emphysema. It is not indicated for the relief of acute bronchospasm [1, 2]. 3. Dosage form and route of administration: It is marketed as a dry powder inhaler (DPI) or a soft mist inhaler (SMI) for once-daily inhalation. The recommended adult dose is 62.5 μg once daily, equivalent to 55 μg umebomide [1, 3]
4. Combination therapy: It is often used in combination with vilanterol (a long-acting β₂ receptor agonist) to enhance bronchodilatory effects. This combination preparation (umebomide/vilanterol) has been approved for maintenance therapy of COPD, utilizing its synergistic effect on airway smooth muscle relaxation [3]
5. Clinical positioning: Umebomide belongs to the long-acting muscarinic receptor antagonist (LAMA). Compared with short-acting anticholinergic drugs (such as ipratropium bromide), its advantages lie in once-daily administration and 24-hour bronchodilatory effect. Its efficacy is comparable to tiotropium bromide, but it may have fewer side effects [1, 2]

C29H34BRNO2

508.4898

507.177

C, 68.50; H, 6.74; Br, 15.71; N, 2.75; O, 6.29

869113-09-7

Umeclidinium-d10 bromide; Umeclidinium-d5 bromide

11519069

White to off-white solid powder

2.102

1

3

8

33

543

0

[Br-].OC(C12CC[N+](CC1)(CCOCC1C=CC=CC=1)CC2)(C1C=CC=CC=1)C1C=CC=CC=1

PEJHHXHHNGORMP-UHFFFAOYSA-M

InChI=1S/C29H34NO2.BrH/c31-29(26-12-6-2-7-13-26,27-14-8-3-9-15-27)28-16-19-30(20-17-28,21-18-28)22-23-32-24-25-10-4-1-5-11-25;/h1-15,31H,16-24H2;1H/q+1;/p-1

diphenyl-[1-(2-phenylmethoxyethyl)-1-azoniabicyclo[2.2.2]octan-4-yl]methanol;bromide

GSK 573719A; GSK-573719; GSK573719; GSK573719A; Incruse Ellipta; GSK 573719; GSK-573719A; Umeclidinium bromide

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

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

DMSO: ~100 mg/mL (~196.7 mM)
Ethanol: ~12 mg/mL

Solubility in Formulation 1: 2.5 mg/mL (4.92 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
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.

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Solubility in Formulation 2: 2.5 mg/mL (4.92 mM) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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