TY - JOUR T1 - Desensitization of the Human Motilin Receptor by Motilides JF - Journal of Pharmacology and Experimental Therapeutics JO - J Pharmacol Exp Ther SP - 1397 LP - 1405 DO - 10.1124/jpet.104.081497 VL - 313 IS - 3 AU - Leen Thielemans AU - Inge Depoortere AU - Jason Perret AU - Patrick Robberecht AU - Yaoquan Liu AU - Theo Thijs AU - Chris Carreras AU - Emmanuel Burgeon AU - Theo L. Peeters Y1 - 2005/06/01 UR - http://jpet.aspetjournals.org/content/313/3/1397.abstract N2 - Tachyphylaxis may have contributed to the failure of the motilide ABT-229 [N-ethyl, N-methyl 4′′ deoxy erythromycin (EM)-B enolether] in clinical trials. We compared the desensitizing potency of structurally related motilides [EM-A, EM-A enolether (ME4), N-ethyl, N-methyl EM-A (ME36), EM-B enolether (ME67), N-ethyl, N-methyl EM-A enolether (EM523), ABT-229 and 4′′ deoxy EM-A enolether (KOS1326)] in a Chinese hamster ovary (CHO)-K1 cell line expressing the human motilin receptor (MTLR) and in rabbit duodenal segments. CHO-MTLR cells were preincubated with motilides prior to stimulation with motilin. The negative logarithm of the preincubation concentration reducing the maximal motilin-induced Ca2+ flux to 50% was calculated (pDC50). Internalization was visualized in CHO-K1 cells containing an enhanced green fluorescent protein (EGFP)-tagged MTLR and quantified in binding experiments. The contractile response of repeated stimulations was measured in duodenal segments. In CHO-MTLR cells, the pDC50 was ABT-229 (8.78) > motilin (7.77) > EM-A (4.78), different from their order of potency to induce Ca2+ release (pEC50): motilin (9.39) > ABT-229 (8.46) > EM-A (7.11). In cells with the EGFP-tagged MTLR, ABT-229 decreased membrane fluorescence by 25 ± 2% compared with 16 ± 2% for motilin and 8 ± 2% for EM-A. Binding studies confirmed that EM-A did not induce MTLR internalization (residual binding 96 ± 4% compared with motilin, 31 ± 3% and ABT-229, 21 ± 1%). Comparison of the pDC50 and pEC50 values of the other motilides ME4 (5.90; 8.08), ME67 (6.03; 8.12), ME36 (3.32; 6.62), EM-523 (6.02; 8.22), and KOS1326 (7.32; 8.14) suggested that the strong desensitizing properties of ABT-229 are mostly related to the removal of the 4″-OH of the cladinose sugar. The decline of the contractile response in duodenal segments correlated with the pDC50. The ability to desensitize and internalize the MTLR is not only determined by potency. This may be an important criterion for the development of a clinically useful compound. The American Society for Pharmacology and Experimental Therapeutics ER -