title: The protease inhibitor atazanavir blocks hERG K(+) channels expressed in HEK293 cells and obstructs hERG protein transport to cell membrane.
abstract: OBJECTIVE
  Atazanavir (ATV) is a HIV-1 protease inhibitor for the treatment of AIDS patients, which is recently reported to provoke excessive prolongation of the QT interval and torsades de pointes (TdP). In order to elucidate its arrhythmogenic mechanisms, we investigated the effects of ATV on the hERG K(+) channels expressed in HEK293 cells.
  
  
  METHODS
  hERG K(+) currents were detected using whole-cell patch clamp recording in HEK293 cells transfected with EGFP-hERG plasmids. The expression of hERG protein was measured with Western blotting. Two mutants (Y652A and F656C) were constructed in the S6 domain within the inner helices of hERG K(+) channels that were responsible for binding of various drugs. The trafficking of hERG protein was studied with confocal microscopy.
  
  
  RESULTS
  Application of ATV (0.01-30 μmol/L) concentration-dependently decreased hERG K(+) currents with an IC50 of 5.7±1.8 μmol/L. ATV (10 μmol/L) did not affect the activation and steady-state inactivation of hERG K(+) currents. Compared with the wild type hERG K(+) channels, both Y652A and F656C mutants significantly reduced the inhibition of ATV on hERG K(+) currents. Overnight treatment with ATV (0.1-30 μmol/L) concentration-dependently reduced the amount of fully glycosylated 155 kDa hERG protein without significantly affecting the core-glycosylated 135 kDa hERG protein in the cells expressing the WT-hERG protein. Confocal microscopy studies confirmed that overnight treatment with ATV obstructed the trafficking of hERG protein to the cell membrane.
  
  
  CONCLUSIONS
  ATV directly blocks hERG K(+) channels via binding to the residues Y652 and F656 in the S6 domain, and indirectly obstructs the transport of the hERG protein to the cell membrane.
authors.affiliation: Department of Pharmacology, School of Medicine, Zhengzhou University, Zhengzhou 450001, China.
authors.forename: Sheng-na
authors.initials: SN
authors.lastname: Han
authors.affiliation: Zhengzhou Central Hospital Affiliated to Zhengzhou University, Zhengzhou 450007, China.
authors.forename: Xiao-yan
authors.initials: XY
authors.lastname: Sun
authors.affiliation: Jiangsu Key Laboratory for Molecular & Medical Biotechnology, College of Life Science in Nanjing Normal University, Nanjing 210046, China.
authors.forename: Zhao
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Department of Pharmacology, School of Medicine, Zhengzhou University, Zhengzhou 450001, China.
authors.forename: Li-rong
authors.initials: LR
authors.lastname: Zhang
mesh_terms: D000069446:Atazanavir Sulfate; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D051638:Ether-A-Go-Go Potassium Channels / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D057809:HEK293 Cells / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D009842:Oligopeptides / Q000494:pharmacology*; D017354:Point Mutation; D011480:Protease Inhibitors / Q000494:pharmacology*; D021381:Protein Transport / Q000187:drug effects; D011725:Pyridines / Q000494:pharmacology*
chemical_list: D051638:Ether-A-Go-Go Potassium Channels; D009842:Oligopeptides; D011480:Protease Inhibitors; D011725:Pyridines; D000069446:Atazanavir Sulfate
references.citation: Br J Pharmacol. 2006 Nov;149(5):481-9
references.pmid: 16967046
references.citation: Proc Natl Acad Sci U S A. 2000 Oct 24;97(22):12329-33
references.pmid: 11005845
references.citation: J Cardiovasc Pharmacol. 2006 Jan;47(1):21-9
references.pmid: 16424781
references.citation: HIV Med. 2006 Jul;7(5):317-22
references.pmid: 16945077
references.citation: Mol Pharmacol. 2004 Aug;66(2):240-9
references.pmid: 15266014
references.citation: Pediatr Cardiol. 2000 May-Jun;21(3):189-96
references.pmid: 10818172
references.citation: Chest. 1997 Sep;112(3):646-53
references.pmid: 9315796
references.citation: Ther Clin Risk Manag. 2009 Feb;5(1):99-116
references.pmid: 19436623
references.citation: J Pharmacol Exp Ther. 2007 Feb;320(2):525-34
references.pmid: 17095614
references.citation: N Engl J Med. 2004 Mar 4;350(10):1013-22
references.pmid: 14999113
references.citation: Clin Infect Dis. 2008 Aug 1;47(3):e36-8
references.pmid: 18558878
references.citation: J Pharmacol Exp Ther. 2005 Jan;312(1):316-23
references.pmid: 15340016
references.citation: J Appl Toxicol. 2008 Mar;28(2):104-11
references.pmid: 17516459
references.citation: Br J Clin Pharmacol. 2009 Jan;67(1):76-82
references.pmid: 19076152
references.citation: Basic Clin Pharmacol Toxicol. 2010 Aug;107(2):643-9
references.pmid: 20662825
references.citation: J Biol Chem. 2006 Mar 3;281(9):5877-84
references.pmid: 16407206
references.citation: AIDS. 2011 Jan 28;25(3):367-77
references.pmid: 21150558
references.citation: Acta Pharmacol Sin. 2014 Dec;35(12):1473-84
references.pmid: 25418379
references.citation: J Med Assoc Thai. 2013 Apr;96(4):501-3
references.pmid: 23691707
references.citation: Ann Pharmacother. 2002 Jun;36(6):1006-8
references.pmid: 12022902
references.citation: Naunyn Schmiedebergs Arch Pharmacol. 2011 Feb;383(2):119-39
references.pmid: 21120454
references.citation: Naunyn Schmiedebergs Arch Pharmacol. 2014 Jan;387(1):23-32
references.pmid: 24045971
references.citation: N Engl J Med. 2003 Nov 20;349(21):1993-2003
references.pmid: 14627784
references.citation: Lancet. 2002 Nov 30;360(9347):1747-8
references.pmid: 12480430
references.citation: Mol Pharmacol. 2004 Nov;66(5):1201-12
references.pmid: 15308760
references.citation: Cardiovasc Res. 2003 Apr 1;58(1):32-45
references.pmid: 12667944
references.citation: Rev Esp Cardiol. 2002 Aug;55(8):878-81
references.pmid: 12199987
references.citation: Acta Pharmacol Sin. 2013 Oct;34(10):1359-66
references.pmid: 23933653
references.citation: Eur J Pharmacol. 2011 Jan 10;650(1):138-44
references.pmid: 20951697
references.citation: Lancet. 2005 Feb 19-25;365(9460):682-6
references.pmid: 15721475
references.citation: Clin Infect Dis. 2007 Mar 15;44(6):e67-8
references.pmid: 17304444