title: Prolonged inhibition of 5-HT₃ receptors by palonosetron results from surface receptor inhibition rather than inducing receptor internalization.
abstract: OBJECTIVE
  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.
  
  
  METHODS
  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.
  
  
  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
  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.
authors.affiliation: Division of Neuroscience, Medical Research Institute, Ninewells Medical School, University of Dundee, Dundee, UK.
authors.forename: J Daniel
authors.initials: JD
authors.lastname: Hothersall
authors.forename: Christopher
authors.initials: C
authors.lastname: Moffat
authors.forename: Christopher N
authors.initials: CN
authors.lastname: Connolly
mesh_terms: D000494:Allosteric Regulation; D000818:Animals; D000932:Antiemetics / Q000378:metabolism / Q000494:pharmacology*; D019556:COS Cells; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism; D002522:Chlorocebus aethiops; D015536:Down-Regulation / Q000187:drug effects*; D004705:Endocytosis / Q000187:drug effects; D017829:Granisetron / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D006835:Hydrazones / Q000494:pharmacology; D007546:Isoquinolines / Q000378:metabolism / Q000494:pharmacology*; D007700:Kinetics; D009761:Nystatin / Q000494:pharmacology; D000077924:Palonosetron; D021381:Protein Transport / Q000187:drug effects; D011812:Quinuclidines / Q000378:metabolism / Q000494:pharmacology*; D011869:Radioligand Assay; D044406:Receptors, Serotonin, 5-HT3 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D058831:Serotonin 5-HT3 Receptor Antagonists / Q000378:metabolism / Q000494:pharmacology*
chemical_list: D000932:Antiemetics; D017829:Granisetron; D006835:Hydrazones; D007546:Isoquinolines; D009761:Nystatin; D000077924:Palonosetron; D011812:Quinuclidines; D044406:Receptors, Serotonin, 5-HT3; D011994:Recombinant Proteins; D058831:Serotonin 5-HT3 Receptor Antagonists; C509409:HTR3A protein, human; C509410:HTR3B protein, human; C511794:N'-(3,4-dihydroxybenzylidene)-3-hydroxy-2-naphthahydrazide