title: Dissociation of E-4031 from the HERG channel caused by mutations of an amino acid results in greater block at high stimulation frequency.
abstract: OBJECTIVE
  We have reported identification of the amino acid whose mutation reduces effects of quinidine on the HERG channel. Although the residue (isoleucine at 647) is not in the recently reported methanesulfonanilide binding site, a single concentration of E-4031 (10 microM) was less effective to I647 mutant channels than wild type HERG channel. We designed the present experiment to further investigate influence of mutations at 647 on the effects of methanesulfonanilides.
  
  
  RESULTS
  HERG channels were expressed in Xenopus oocytes and their currents were measured by a two-microelectrode voltage clamp method. Of the two mutations initially studied (I647A and I647F), the I647F had a greater influence and differentially affected the effects of dofetilide and E-4031. The IC(50) for dofetilide of the two mutant channels (I647A and I647F) was increased only 2-fold, but the IC(50) for E-4031 was increased 6-fold (I647A) and 14-fold (I647F). Aromatic residues other than phenylalanine were then substituted for I647, and found to reduce the effects of E-4031. Whereas E-4031 dissociated from the mutant channels during rested state, dofetilide little dissociated. The mutant channels that showed recovery from E-4031 block were inhibited greater at 1 Hz than at 0.1 Hz.
  
  
  CONCLUSIONS
  The present results indicate that dissociation of a drug from the HERG channel results in greater block at high frequency. Although the mechanism by which the mutations cause the dissociation of E-4031 is uncertain, it is noteworthy that one methanesulfonanilide dissociates from the channel more easily than another.
authors.affiliation: Department of Pharmacology, Yamagata University School of Medicine, Yamagata, Japan. kuishii@med.id.yamagata-u.ac.jp
authors.forename: Kuniaki
authors.initials: K
authors.lastname: Ishii
authors.forename: Mirei
authors.initials: M
authors.lastname: Nagai
authors.forename: Masahiro
authors.initials: M
authors.lastname: Takahashi
authors.forename: Masao
authors.initials: M
authors.lastname: Endoh
mesh_terms: D000818:Animals; D000889:Anti-Arrhythmia Agents / Q000378:metabolism / Q000494:pharmacology*; D027682:Cation Transport Proteins; D004305:Dose-Response Relationship, Drug; D051638:Ether-A-Go-Go Potassium Channels; D007532:Isoleucine / Q000235:genetics; D008133:Long QT Syndrome; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D010627:Phenethylamines / Q000494:pharmacology; D010880:Piperidines / Q000378:metabolism / Q000494:pharmacology*; D017354:Point Mutation; D026902:Potassium Channel Blockers / Q000494:pharmacology; D015221:Potassium Channels / Q000187:drug effects / Q000235:genetics*; D024642:Potassium Channels, Voltage-Gated; D011725:Pyridines / Q000378:metabolism / Q000494:pharmacology*; D013449:Sulfonamides / Q000494:pharmacology; D014981:Xenopus
chemical_list: D000889:Anti-Arrhythmia Agents; D027682:Cation Transport Proteins; D051638:Ether-A-Go-Go Potassium Channels; C092509:KCNH6 protein, human; D010627:Phenethylamines; D010880:Piperidines; D026902:Potassium Channel Blockers; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D011725:Pyridines; D013449:Sulfonamides; D007532:Isoleucine; C063968:E 4031; C063533:dofetilide