Search

Background and Purpose The 5-HT3 receptor antagonist palonosetron is an important treatment for emesis and nausea during cancer therapy. Its clinical efficacy may result from its unique binding and clearance characteristics and receptor down-regulation mechanisms. We investigated the mechanisms by which palonosetron exerts its long-term inhibition of 5-HT3 receptors for a better understanding of its clinical efficacy. Experimental Approach Cell surface receptors (recombinantly expressed 5HT3A or 5HT3AB in COS-7 cells) were monitored using [3H]granisetron binding and ELISA after exposure to palonosetron. Receptor endocytosis was investigated using immunofluorescence microscopy. Key Results Chronic exposure to palonosetron reduced the number of available cell surface [3H]granisetron binding sites. This down-regulation was not sensitive to either low temperature or pharmacological inhibitors of endocytosis (dynasore or nystatin) suggesting that internalization did not play a role. This was corroborated by our observation that there was no change in cell surface 5-HT3 receptor levels or increase in endocytic rate. Palonosetron exhibited slow dissociation from the receptor over many hours, with a significant proportion of binding sites being occupied for at least 4 days. Furthermore, our observations suggest that chronic receptor down-regulation involved interactions with an allosteric binding site. Conclusions and Implications Palonosetron acts as a pseudo-irreversible antagonist causing prolonged inhibition of 5-HT3 receptors due to its very slow dissociation. In addition, an irreversible binding mode persists for at least 4 days. Allosteric receptor interactions appear to play a role in this phenomenon.

Selective and potent antagonists for the β(2) -adrenoceptor are potentially interesting as experimental and clinical tools, and we sought to identify novel ligands with this pharmacology.A range of pharmacological assays was used to assess potency, affinity, selectivity (β(2) -adrenoceptor vs. β(1) -adrenoceptor) and efficacy.Ten novel compounds were identified but none had as high affinity as the prototypical β(2) -adrenoceptor blocker ICI-118,551, although one of the novel compounds was more selective for β(2) -adrenoceptors. Most of the ligands were inverse agonists for β(2) -adrenoceptor-cAMP signalling, although one (5217377) was a partial agonist and another a neutral antagonist (7929193). None of the ligands were efficacious with regard to β(2) -adrenoceptor-β-arrestin signalling. The (2S,3S) enantiomers were identified as the most active, although unusually the racemates were the most selective for the β(2) -adrenoceptors. This was taken as evidence for some unusual enantiospecific behaviour.In terms of improving on the pharmacology of the ligand ICI-118,551, one of the compounds was more selective (racemic JB-175), while one was a neutral antagonist (7929193), although none had as high an affinity. The results substantiate the notion that β-blockers do more than simply inhibit receptor activation, and differences between the ligands could provide useful tools to investigate receptor biology.