Prolonged inhibition of 5-HT(3) receptors by palonosetron results from surface receptor inhibition rather than inducing receptor internalization

BACKGROUND AND PURPOSE: The 5-HT(3) 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-HT(3) receptors for a better understanding of its clinical efficacy. EXPERIMENTAL APPROACH: Cell surface receptors (recombinantly expressed 5HT(3)A or 5HT(3)AB in COS-7 cells) were monitored using [(3)H]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 [(3)H]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-HT(3) 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-HT(3) 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.