Intracellular interactions of umeclidinium and vilanterol in human airway smooth muscle.

BACKGROUND: Intracellular mechanisms of action of umeclidinium (UMEC), a long-acting muscarinic receptor antagonist, and vilanterol (VI), a long-acting β2 -adrenoceptor (β2 R) agonist, were investigated in target cells: human airway smooth-muscle cells (ASMCs).

MATERIALS AND METHODS: ASMCs from tracheas of healthy lung-transplant donors were treated with VI, UMEC, UMEC and VI combined, or control compounds (salmeterol, propranolol, ICI 118.551, or methacholine [MCh]). Cyclic adenosine monophosphate (cAMP) was measured using an enzyme-linked immunosorbent assay, intracellular free calcium ([Ca2+ ]i ) using a fluorescence assay, and regulator of G-protein signaling 2 (RGS2) messenger RNA using real-time quantitative polymerase chain reaction.

RESULTS: VI and salmeterol (10-12 -10-6 M) induced cAMP production from ASMCs in a concentration-dependent manner, which was greater for VI at all concentrations. β2 R antagonism by propranolol or ICI 118.551 (10-12 -10-4 M) resulted in concentration-dependent inhibition of VI-induced cAMP production, and ICI 118.551 was more potent. MCh (5×10-6 M, 30 minutes) attenuated VI-induced cAMP production ( P <0.05), whereas pretreatment with UMEC (10-8 M, 1 hour) restored the magnitude of VI-induced cAMP production. ASMC stimulation with MCh (10-11 -5×10-6 M) resulted in a concentration-dependent increase in [Ca2+ ]i , which was attenuated with UMEC pretreatment. Reduction of MCh-induced [Ca2+ ]i release was greater with UMEC + VI versus UMEC. UMEC enhanced VI-induced RGS2 messenger RNA expression.

CONCLUSION: These data indicate that UMEC reverses cholinergic inhibition of VI-induced cAMP production, and is a more potent muscarinic receptor antagonist when in combination with VI versus either alone.