title: The anti-protozoal drug pentamidine blocks KIR2.x-mediated inward rectifier current by entering the cytoplasmic pore region of the channel.
abstract: OBJECTIVE
  Pentamidine is a drug used in treatment of protozoal infections. Pentamidine treatment may cause sudden cardiac death by provoking cardiac arrhythmias associated with QTc prolongation and U-wave alterations. This proarrhythmic effect was linked to inhibition of hERG trafficking, but not to acute block of ion channels contributing to the action potential. Because the U-wave has been linked to the cardiac inward rectifier current (I(K1)), we examined the action and mechanism of pentamidine-mediated I(K1) block.
  
  
  METHODS
  Patch clamp measurements of I(K1) were made on cultured adult canine ventricular cardiomyocytes, K(IR)2.1-HEK293 cells and K(IR)2.x inside-out patches. Pentamidine binding to cytoplasmic amino acid residues of K(IR)2.1 channels was studied by molecular modelling.
  
  
  RESULTS
  Pentamidine application (24 h) decreased I(K1) in cultured canine cardiomyocytes and K(IR)2.1-HEK293 cells under whole cell clamp conditions. Pentamidine inhibited I(K1) in K(IR)2.1-HEK293 cells 10 min after application. When applied to the cytoplasmic side under inside-out patch clamp conditions, pentamidine block of I(K1) was acute (IC(50)= 0.17 microM). Molecular modelling predicted pentamidine-channel interactions in the cytoplasmic pore region of K(IR)2.1 at amino acids E224, D259 and E299. Mutation of these conserved residues to alanine reduced pentamidine block of I(K1). Block was independent of the presence of spermine. K(IR)2.2, and K(IR)2.3 based I(K1) was also sensitive to pentamidine blockade.
  
  
  CONCLUSIONS
  Pentamidine inhibits cardiac I(K1) by interacting with three negatively charged amino acids in the cytoplasmic pore region. Our findings may provide new insights for development of specific I(K1) blocking compounds.
authors.affiliation: Department of Medical Physiology, Division Heart & Lungs, UMCU, Utrecht, The Netherlands.
authors.forename: T P
authors.initials: TP
authors.lastname: de Boer
authors.forename: L
authors.initials: L
authors.lastname: Nalos
authors.forename: A
authors.initials: A
authors.lastname: Stary
authors.forename: B
authors.initials: B
authors.lastname: Kok
authors.forename: M J C
authors.initials: MJ
authors.lastname: Houtman
authors.forename: G
authors.initials: G
authors.lastname: Antoons
authors.forename: T A B
authors.initials: TA
authors.lastname: van Veen
authors.forename: J D M
authors.initials: JD
authors.lastname: Beekman
authors.forename: B L
authors.initials: BL
authors.lastname: de Groot
authors.forename: T
authors.initials: T
authors.lastname: Opthof
authors.forename: M B
authors.initials: MB
authors.lastname: Rook
authors.forename: M A
authors.initials: MA
authors.lastname: Vos
authors.forename: M A G
authors.initials: MA
authors.lastname: van der Heyden
mesh_terms: D000818:Animals; D000981:Antiprotozoal Agents / Q000494:pharmacology*; D015153:Blotting, Western; D002460:Cell Line; D003593:Cytoplasm / Q000187:drug effects* / Q000378:metabolism; D004285:Dogs; D006801:Humans; D009154:Mutation; D018408:Patch-Clamp Techniques; D010419:Pentamidine / Q000494:pharmacology*; D024661:Potassium Channels, Inwardly Rectifying / Q000037:antagonists & inhibitors* / Q000235:genetics
chemical_list: D000981:Antiprotozoal Agents; C489461:Kir2.1 channel; D024661:Potassium Channels, Inwardly Rectifying; D010419:Pentamidine