The Low-Potency, Voltage-Dependent HERG Blocker Propafenone—Molecular Determinants and Drug Trapping

Abstract

The molecular determinants of high-affinity human ether-a-gogo-related gene (HERG) potassium channel blockade by methanesulfonanilides include two aromatic residues (Phe656 and Tyr652) on the inner helices (S6) and residues on the pore helices that face into the inner cavity, but determinants for lower-affinity HERG blockers may be different. In this study, alanine-substituted HERG channel mutants of inner cavity residues were expressed in Xenopus laevis oocytes and were used to characterize the HERG channel binding site of the antiarrhythmic propafenone. Propafenone's blockade of HERG was strongly dependent on residue Phe656 but was insensitive or weakly sensitive to mutation of Tyr652, Thr623, Ser624, Val625, Gly648, or Val659 and did not require functional inactivation. Homology models of HERG based on KcsA and MthK crystal structures, representing the closed and open forms of the channel, respectively, suggest propafenone is trapped in the inner cavity and is unable to interact exclusively with Phe656 in the closed state (whereas exclusive interactions between propafenone and Phe656 are found in the open-channel model). These findings are supported by very slow recovery of wild-type HERG channels from block at Ϫ120 mV, but extremely rapid recovery of D540K channels that reopen at this potential. The experiments and modeling suggest that the open-state propafenone binding-site may be formed by the Phe656 residues alone. The binding site for propafenone (which may involve -stacking interactions with two or more Phe656 side-chains) is either perturbed or becomes less accessible because of closed-channel gating. This provides further evidence for the existence of gating-induced changes in the spatial location of Phe656 side chains.Pharmacological blockade of the cardiac 'rapid' delayed rectifier potassium (K ϩ ) current (I Kr ) is commonly associated with a drug's propensity to cause acquired long QT syndrome (Roden et al., 1996). The ␣-subunit of the I Kr channel is encoded by HERG (Sanguinetti et al., 1995). Molecular determinants of this channel's blockade by the archetypal highpotency HERG blockers, the methanesulfonanilides, reside in the inner cavity of the channel pore and are composed of amino acid residues in the H5 loop (adjacent to the selectivity filter) and the S6 transmembrane domain (Lees-Miller et al., 2000;Mitcheson et al., 2000a). For all drugs tested, the molecular determinants include the amino acid residues Phe656 and Tyr652 [except for vesnarinone, which does not require Tyr652 (Kamiya et al., 2001), and fluvoxamine, which can block HERG when Phe656 or Tyr652 are mutated ], and in many cases the molecular determinants additionally include a combination of Thr623, Ser624, Val625, Gly648, and Val659. Most drugs with a high-affinity for HERG have been shown to have an open-state-dependent blockade mechanism (Spector et al., 1996). Open-state blockade of HERG may involve drug trapping [e.g., MK-499 (Mitcheson et al., 2000b)], a 'foot in the door' type mechanism [e.g., chloroquine (Sanch...

The protease inhibitor atazanavir blocks hERG K+ channels expressed in HEK293 cells and obstructs hERG protein transport to cell membrane

“…However, not all hERG blocking drugs interact with these residues. For example, the blockade of hERG by propafenone [24] is substantially reduced by F656 but not by Y652. Our study showed that Y652A and F656C mutations markedly weakened the blockade of the hERG current by ATV (3-fold increase in the IC 50 for Y652A and F656C) compared with the WT-hERG.…”

Molecular Determinants of hERG Channel Block

“…A critical role for Phe656 was also proposed for binding of the antiarrhythmic agent dofetilide and quinidine (Lees-Miller et al, 2000), and the importance of both Tyr652 and Phe656 has been confirmed for several other drugs, including chloroquine (Sanchez-Chapula et al, 2002), quinidine (Sanchez-Chapula et al, 2003), halofantrine (Sanchez-Chapula et al, 2004), terfenadine and cisapride (Fernandez et al, 2004), lidoflazine (Ridley et al, 2004a), clofilium, and ibutilide . Some drugs, such as fluvoxamine (Milnes et al, 2003), propafenone (Witchel et al, 2004) and dronedarone (Ridley et al, 2004b), seem not to interact strongly with Tyr652 and/or Phe656; however, it is also not known whether these drugs interact with the other residues identified previously as part of the drug binding site of hERG. In this study, we extend our alanine-scan analysis of the Ser6 domain and pore helix region of hERG channels expressed in oocytes to four additional drugs (Fig.…”

Kinetics of Drug Interaction with the Kv11.1 Potassium Channel

“…It is well established that drug block of K v 11.1 can be voltage- (Paul et al, 2002;Witchel et al, 2004;Kamiya et al, 2008) and state-dependent (Perrin et al, 2008), and these properties may vary substantially from drug to drug (Perrin et al, 2008). Consequently, the apparent IC 50 can vary considerably depending on the voltage protocol used to measure block (Kirsch et al, 2004;Milnes et al, 2010).…”