Abstract
Background: rac-Fenoterol is a β2-adrenoceptor agonist (β2-AR) used in the treatment of asthma. It has two chiral centers and is marketed as a racemic mixture of R,R′- and S,S′-fenoterol (R-F and S-F). Here we report the separation of the R-F and S-F enantiomers and the evaluation of their binding to and activation of the β2-AR. Methods: R-F and S-F were separated from the enantiomeric mixture by chiral chromatography and absolute configuration determined by circular dichroism. β2-AR binding was evaluated using frontal affinity chromatography with a stationary phase containing immobilized membranes from HEK-293 cells that express human β2-AR and standard membrane binding studies using the same membranes. The effect of R-F and SF on cardiomyocyte contractility was also investigated using freshly isolated adult rat cardiomyocytes. Results: Chiral chromatography of rac-fenoterol yielded separated peaks with an enantioselectivity factor of 1.21. The less retained peak was assigned the absolute configuration of S-F and the more retained peak R-F. Frontal chromatography using membrane-bound β2-AR as the stationary phase and rac-3H-fenoterol as a marker ligand showed that addition of increasing concentrations of R-F to the mobile phase produced concentration-dependent decreases in rac-3H-fenoterol retention, while similar addition of S-F produced no change in rac-3H-fenoterol retention. The calculated dissociation constant of R-F was 472 nM and the number of available binding sites 176 pmol/column, which was consistent with the results from the membrane binding study 460 ± 55 nM (R-F) and 109,000 ± 10,400 nM (S-F). In the cardiomyocytes, R-F increased maximum contractile response from (265 ± 11.6)% to (306 ± 11.8)% of resting cell length (P < 0.05) and reduced EC50 from -7.0 ± 0.270 to -7.1 ± 0.2 log[M] (P < 0.05), while S-F had no significant effect. Discussion: Previous studies have shown that rac-fenoterol acts as an apparent β2-AR/Gs selective agonist and fully restores diminished β2-AR contractile response in cardiomyocytes from failing hearts of spontaneously hypertensive rats (SHR). Here we report the separation of the enantiomers of rac-fenoterol and that R-F is the active component of rac-fenoterol. Further evaluation of R-F will determine if it has enhanced selectivity and specificity for β2-AR/Gs activation and if it can be used in the treatment of congestive heart failure.
Original language | English (US) |
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Pages (from-to) | 822-827 |
Number of pages | 6 |
Journal | Chirality |
Volume | 18 |
Issue number | 10 |
DOIs | |
State | Published - 2006 |
Keywords
- Enantiomers
- Fenoterol
- β-adrenergic agonist
ASJC Scopus subject areas
- Analytical Chemistry
- Catalysis
- Pharmacology
- Drug Discovery
- Spectroscopy
- Organic Chemistry