The novel quaternary ammonium salt, CHF5407, showed subnanomolar affinities for human muscarinic M1, M2 and M3 receptors, and dissociated very slowly (t1/2 = 166 min from hM3 receptors (t1/2 = 166 min) with a large part of the receptorial complex (54%) remaining undissociated at 32h from radioligand washout. In contrast, [3H]-CHF5407 dissociated quickly from hM2 receptors (t1/2=31 min), whereas [3H]-tiotropium dissociated slowly from both hM3 (t1/2=163 min) and hM2 receptor (t1/2=297 min). In the guinea-pig isolated trachea and human isolated bronchus, CHF5407 produced a potent (pIC50=9.0-9.6) and long-lasting (up to 24h) inhibition of M3-receptor mediated contractile responses to carbachol. In the guinea-pig electrically-driven left atrium, the M2 receptor-mediated inhibitory response to carbachol was recovered more quickly in CHF5407-pretreated, than in tiotropium-pretreated preparations. CHF5407, administered intratracheally (i.t.) to anaesthetized guinea pigs, potently inhibited acetylcholine (Ach)-induced bronchoconstriction with an ED50 value of 0.15 nmoles/kg. The effect was substained over a period of 24 h, with a residual 57% inhibition 48 h after antagonist administration at 1 nmoles/kg. In conscious guinea pigs, inhaled CHF5407 inhibited Ach-induced bronchoconstriction for at least 24 h as did tiotropium at similar dosages. Cardiovascular parameters in anaesthetised guinea pigs were not significantly changed by CHF5407, up to 100 nmoles/kg i.v. and up to 1000 nmoles/kg i.t. In conclusion, CHF5407 shows a prolonged antibronchospastic activity both in vitro and in vivo, due to a very slow dissociation from M3 receptors. In contrast, CHF5407 is markedly short-acting at M2 receptors, a behaviour not shared by tiotropium.
Bronchodilator Activity of (3R)-3[[[ (3-fluorophenyl) [(3,4,5trifluorophenyl)methyl]amino] oxy]carbonyl]-1-[2-oxo-2-(2-thienyl)ethyl]-1-azoniabicyclo[2.2.2]octane bromide (CHF5407) a Potent, Long-Acting and Selective Muscarinic M3 Receptor Antagonist / Villetti G; Pastore F; Bergamaschi M; Bassani F; Bolzoni PT; Battipaglia L; Amari G; Rizzi A; Delcanale M; Volta R; Cenacchi V; Cacciani F; Zaniboni M; Berti F; Rossoni G; Harrison S; Petrillo P; Santoro E; Scudellaro R; Mannini F; Geppetti P; Razzetti R; Patacchini R; Civelli M.. - In: JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS. - ISSN 0022-3565. - 335(2010), pp. 622-635. [10.1124/jpet.110.170035]