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Translational pharmacology of selected compounds at the human and canine α1a adrenergic receptors The pharmacological modulation of adrenergic receptor (AR) subtypes is widely used for a variety of medical conditions, including hypertension, heart failure, prostatic hypertrophy and depression (Rossier et al. 1999). Whilst no significant species differences in agonist pharmacology have been identified for the α1a receptor to date, certain antagonists show unexpectedly low potency in isolated tissue assay (Hieble, 2000). The aim of the present study was to investigate the translational pharmacology of compounds at the human and previously undefined canine α1a AR. A radioligand filter binding method was used to compare the affinity of compounds, selected from different chemical series, with previously demonstrated α1 AR affinity (Michelotti et al. 2000). The binding assay used membranes prepared from Chinese Hamster Ovary cell lines stably expressing either the human or canine a 1a receptor. The affinity of test compounds for the a 1a receptor were assessed by their ability to compete with specific binding of [3H]-prazosin, with non-specific binding determined using 10μM phentolamine. Assay buffer contained 20mM HEPES, 5mM MgCl2, 0.5mM EDTA and 0.02% bacitracin at pH 7.4 (25°C). Briefly, solubilised compound, [3H]-prazosin and diluted membrane preparation (at predetermined optimal protein concentration) were mixed and incubated for 1hour at 25°C. Binding was terminated by harvesting with a Brandel cell harvester and ice-cold buffer and plates read using an NXT Topcount. An IC50 value (concentration of drug which reduced specific [3H]-prazosin binding by 50%), was determined for each test compound from a 10 point concentration effect curve and converted to a Ki value using the method described by Cheng and Prussoff (1973). Resulting data are shown in Table 1. Statistical analysis was conducted utilising a Student’s t-test, with p<0.05 considered significant.
Table 1. Comparison of known a 1 adrenergic receptor (AR) agonist and antagonist binding affinity to the human and canine a 1a AR. Results are given as arithmetic mean pKi ± S.D. (n). Compounds with a statistically significant difference in binding affinity are marked with*. These data demonstrate that certain compounds from distinct chemical series bind with significantly different affinities to the human and canine a 1a adrenoreceptors.
Cheng Y.C. & Prussoff W.H. (1973) Biochem. Pharmacol, 22: 3099-3108 |
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