Comparative molecular field analysis of fenoterol derivatives interacting with an agonist-stabilized form of the β2-adrenergic receptor

Abstract

The β2-adrenergic receptor (β2-AR) agonist [3H]-(R,R′)-methoxyfenoterol was employed as the marker ligand in displacement studies measuring the binding affinities (Ki values) of the stereoisomers of a series of 4′-methoxyfenoterol analogs in which the length of the alkyl substituent at α′ position was varied from 0 to 3 carbon atoms. The binding affinities of the compounds were additionally determined using the inverse agonist [3H]-CGP-12177 as the marker ligand and the ability of the compounds to stimulate cAMP accumulation, measured as EC50 values, were determined in HEK293 cells expressing the β2-AR. The data indicate that the highest binding affinities and functional activities were produced by methyl and ethyl substituents at the α′ position. The results also indicate that the Ki values obtained using [3H]-(R,R′)-methoxyfenoterol as the marker ligand modeled the EC50 values obtained from cAMP stimulation better than the data obtained using [3H]-CGP-12177 as the marker ligand. The data from this study was combined with data from previous studies and processed using the Comparative Molecular Field Analysis approach to produce a CoMFA model reflecting the binding to the β2-AR conformation probed by [3H]-(R,R′)-4′-methoxyfenoterol. The CoMFA model of the agonist-stabilized β2-AR suggests that the binding of the fenoterol analogs to an agonist-stabilized conformation of the β2-AR is governed to a greater extend by steric effects than binding to the [3H]-CGP-12177-stabilized conformation(s) in which electrostatic interactions play a more predominate role.

Recruitment ofβ-Arrestin 1 and 2 to theβ2-Adrenoceptor: Analysis of 65 Ligands

“…Many agonists analyzed in this study are full agonists with respect to G-protein activation and cAMP production (Toll et al, 2011;Brunskole Hummel et al, 2013); however, they turned out to be only (weak) partial agonists regarding barr recruitment, indicating a strong bias toward G a smediated cAMP formation. Compounds showing low efficacies for barr recruitment were ALB, FOR, SAL, zinterol, and in particular the FEN derivatives (Baker, 2010;Brunskole Hummel et al, 2013;Plazinska et al, 2014). Reduced efficacies may result from modification of the two hydroxyl groups in the m and p positions of the catechol moiety of the endogenous agonists and ISO.…”

Interaction of fenoterol stereoisomers with β2-adrenoceptor-Gsα fusion proteins: antagonist and agonist competition binding

“…Observation of increased affinities for the ligands modified at the aminoalkyl tail confirm the rationale for the synthesis of compounds 2 and 3 (Jozwiak et al 2007): First, an oxygen at 4′-position of the phenyl-(compounds 1 and 2) or naphthyl-ring (compound 3) enables hydrogen bond interactions. Second, introduction of a more extensive aromatic naphthyl moiety causes increased affinity of 3-stereoisomers by favorable bulk interactions (Plazinska et al 2014a).…”

“…Furthermore, molecular models based on recent β 2 AR crystal structures revealed two possible orientations of the aminoalkyl tail of fenoterol-like ligands into the extended orthosteric binding site as well (Plazinska et al 2013). Plazinska et al (2014a) used [ 3 H](R,R′)-2 binding data to generate a comparative molecular field analysis (CoMFA) model with an optimized reflection of agonist binding to β 2 AR. Data obtained at β 2 AR-G s α fusion proteins with [ 3 H](R,R′)-2 as marker match the predicted affinities of the refined CoMFA model with only minor deviations (Table 3) (Plazinska et al 2014a).…”

“…Plazinska et al (2014a) used [ 3 H](R,R′)-2 binding data to generate a comparative molecular field analysis (CoMFA) model with an optimized reflection of agonist binding to β 2 AR. Data obtained at β 2 AR-G s α fusion proteins with [ 3 H](R,R′)-2 as marker match the predicted affinities of the refined CoMFA model with only minor deviations (Table 3) (Plazinska et al 2014a). The analyses emphasize the importance of a proper stereo-configuration at both chiral C-atoms for an optimal interaction with the receptor (Jozwiak et al 2011;Plazinska et al 2014a, b).…”

“…As a fenoterol analog itself, this high-affinity radioactive marker (K D of 4.88±0.41 nM in HEK-β 2 AR) is particularly suited to analyze binding of fenoterol-like ligands at the β 2 AR (Jozwiak et al 2010a). Therefore, combination of pharmacological analyses using this novel radioligand at HEK-β 2 AR with updated crystal structures of the active and inactive β 2 AR already allowed the improvement of in silico models describing the stereoselective interaction of fenoterol derivatives with the receptor (Plazinska et al 2013(Plazinska et al , 2014a.…”