Discovery of β-arrestin-biased β2-adrenoceptor agonists from 2-amino-2-phenylethanol derivatives

Abstract

β-Arrestins are a small family of proteins important for signal transduction at G protein-coupled receptors (GPCRs). β-Arrestins are involved in the desensitization of GPCRs. Recently, biased ligands possessing different efficacies in activating the G protein- versus the β-arrestin-dependent signals downstream of a single GPCR have emerged, which can be used to selectively modulate GPCR signal transduction in such a way that desirable signals are enhanced to produce therapeutic effects while undesirable signals of the same GPCR are suppressed to avoid side effects. In the present study, we evaluated agonist bias for compounds developed along a drug discovery project of β₂-adrenoceptor agonists. About 150 compounds, including derivatives of fenoterol, 2-amino-1-phenylethanol and 2-amino-2-phenylethanol, were obtained or synthesized, and initially screened for their β-adrenoceptor-mediated activities in the guinea pig tracheal smooth muscle relaxation assay or the cardiomyocyte contractility assay. Nineteen bioactive compounds were further assessed using both the HTRF cAMP assay and the PathHunter β-arrestin assay. Their concentration-response data in stimulating cAMP synthesis and β-arrestin recruitment were applied to the Black-Leff operational model for ligand bias quantitation. As a result, three compounds (L-2, L-4, and L-12) with the core structure of 5-(1-amino-2-hydroxyethyl)-8-hydroxyquinolin-2(1H)-one were identified as a new series of β-arrestin-biased β₂-adrenoceptor agonists, whereas salmeterol was found to be G_s-biased. These findings would facilitate the development of novel drugs for the treatment of both heart failure and asthma.

Keywords: asthma; biased agonism; cAMP; fenoterol; heart failure; salmeterol; β-arrestin; β2-adrenoceptor agonists.

Scheme 1

Synthesis of 2-amino-1-phenylethanol derivatives. Reagents and conditions: (i) amines, ethanol, 5 °C to room temperature (rt), 5 h; (ii) NaBH₄, ethanol and water, rt

Scheme 2

Synthesis of B-24 and B-30. Reagents and conditions: (i) isopropylamine, ethanol, reflux, 13 h

Scheme 3

Synthesis of L-2, L-4, L-6, and L-12. Reagents and conditions: (i) mCPBA, DCM, 0 °C to rt, 30 min; (ii) Ac₂O, 100 °C, 3 h; (iii) CH₃COCl, AlCl₃, DCE, 0 °C to reflux; (iv) C₆H₅CH₂Br, K₂CO₃, in N,N-dimethylformamide, 40 °C, 3 h; (v) Br₂, BF₃Et₂O, DCM, reflux, 15 min; (vi) NaBH₄, CH₃OH, DCM, 0 °C, 30 min; (vii) KOH, CH₃OH, DCM, 0 °C, 30 min; (viii) isopropylamine, N-propylamine, tert-butylamine or N-hexylamine, ZnCl₂, CH₃CN, reflux, 12 h; (ix) H₂, 10% Pd/C, CH₃OH, rt, 2 h

Fig. 1

ΔΔLog(τ/K_A) values of the compounds for β-arrestin signaling over G_s signaling. The significance of the ligand bias was determined by comparing each value with that of (R)-ISO using Bonferroni’s t-test. *P < 0.05, ***P < 0.001 (Error bars are standard errors, n = 5)