Lee BH et al. (2011), Effects of the histamine H(1) receptor antagoni...

XB-ART-43812

Acta Pharmacol Sin 2011 Sep 01;329:1128-37. doi: 10.1038/aps.2011.66.

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Effects of the histamine H(1) receptor antagonist hydroxyzine on hERG K(+) channels and cardiac action potential duration.

Lee BH , Lee SH , Chu D , Hyun JW , Choe H , Choi BH , Jo SH .

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To investigate the effects of hydroxyzine on human ether-a-go-go-related gene (hERG) channels to determine the electrolphysiological basis for its proarrhythmic effects. hERG channels were expressed in Xenopus oocytes and HEK293 cells, and the effects of hydroxyzine on the channels were examined using two-microelectrode voltage-clamp and patch-clamp techniques, respectively. The effects of hydroxyzine on action potential duration were examined in guinea pig ventricular myocytes using current clamp. Hydroxyzine (0.2 and 2 μmol/L) significantly increased the action potential duration at 90% repolarization (APD(90)) in both concentration- and time-dependent manners. Hydroxyzine (0.03-3 μmol/L) blocked both the steady-state and tail hERG currents. The block was voltage-dependent, and the values of IC(50) for blocking the steady-state and tail currents at +20 mV was 0.18±0.02 μmol/L and 0.16±0.01 μmol/L, respectively, in HEK293 cells. Hydroxyzine (5 μmol/L) affected both the activated and the inactivated states of the channels, but not the closed state. The S6 domain mutation Y652A attenuated the blocking of hERG current by ~6-fold. The results suggest that hydroxyzine could block hERG channels and prolong APD. The tyrosine at position 652 in the channel may be responsible for the proarrhythmic effects of hydroxyzine.

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Species referenced: Xenopus laevis
Genes referenced: gnao1 kcnh1 kcnh2

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	Figure 1. The effect of hydroxyzine on action potentials in isolated guinea pig ventricular myocytes. (A) Superimposed action potentials recorded before and after exposure to various concentrations of hydroxyzine. (B) Concentration- and time-dependent prolongation of action potential duration at 90% (APD90). At each experimental condition (after exposure to 0.2 μmol/L for 5 min, 0.2 μmol/L for 10 min, 2 μmol/L for 5 min, and 2 μmol/L for 10 min), the APD90 values were normalized to control APD90 (obtained in the absence of drug). bP<0.05. n=4−9.
	Figure 2. The effect of hydroxyzine on human ether-a-go-go-related gene (hERG) currents (IHERG) elicited by depolarizing voltage pulses in Xenopus oocytes. Superimposed current traces elicited by depolarizing voltage pulses (4 s) in 10 mV increments from a holding potential of −70 mV (upper panel) in the absence of hydroxyzine (control, center panel) and presence of 5 μmol/L hydroxyzine (lower panel).
	Figure 3. The effect of hydroxyzine on hERG channels expressed in Xenopus oocytes. (A) Plot of the normalized hERG current measured at the end of the depolarizing pulse (IHERG) against pulse potential in control and hydroxyzine conditions. The maximal amplitude of the IHERG in control solution was normalized to 1. (B) Plot of the normalized tail current measured at its peak following repolarization. The peak amplitude of the tail current in control solution was normalized to 1. The control data were fitted with the following Boltzmann equation, y=1/{1+exp[(-V+V1/2)/dx]}, yielding a V1/2 of −19.2 mV. (C) Activation curves with the values normalized to their respective maximum for each concentration of hydroxyzine. n=3−5.
	Figure 4. The concentration dependence of hydroxyzine-induced inhibition of hERG channels stably expressed in HEK293 cells. (A) Superimposed IHERG traces were elicited with 4-s depolarizations to +20 mV from a holding potential of −80 mV, and the tail current was recorded at −60 mV for 6 s in the absence or presence of 0.03, 0.1, 0.3, and 1 μmol/L hydroxyzine, as indicated. The voltage protocol was applied every 15 s. The dotted line represents zero current. (B) Concentration dependence curve of inhibition by hydroxyzine for steady-state currents measured at the end of the depolarizing pulse to +20 mV or peak tail currents. The respective normalized currents were plotted against the various concentrations of hydroxyzine. The solid lines are fits of the data points using the Hill equation (see Methods).
	Figure 5. Voltage dependence of hydroxyzine-induced blockade of hERG channels expressed in Xenopus oocytes. (A) Current traces from a cell depolarized to −20 mV (left panel), +10 mV (middle panel), or +40 mV (right panel) before and after application of 5 μmol/L hydroxyzine, showing increased blockade of hERG current at more positive potentials. The protocol consisted of 4 s depolarizing steps to −20 mV, +10 mV, or +40 mV from a holding potential of −70 mV, followed by repolarization to −60 mV. (B) Hydroxyzine-induced hERG current inhibition at various voltages. At each depolarizing voltage step (-20, +10, or +40 mV), the tail currents in the presence of 5 μmol/L hydroxyzine were normalized to the tail current obtained in the absence of drug. n=4.
	Figure 6. Relative changes in sustained hERG currents in response to hydroxyzine in Xenopus oocytes. (A) An example recording of currents under control conditions (control) and after application of 5 μmol/L hydroxyzine (for 7 min without any intervening test pulses). (B) Relative current obtained by dividing the current in 5 μmol/L hydroxyzine by the control current shown in in panel A. Current inhibition increased with depolarization time to 56% at 2 s in this representative cell, indicating that primarily open and/or inactivated channels were blocked.
	Figure 7. Blocking of inactivated hERG channels by hydroxyzine in Xenopus oocytes. (A) Inhibition of inactivated channels by 5 μmol/L hydroxyzine. hERG channels were inactivated by an initial voltage-step to +80 mV, followed by channel opening at 0 mV. (B) The corresponding relative block during the 0 mV step is shown. Maximum inhibition was achieved in the inactivated state during the first step, and no additional time-dependent inhibition occurred upon channel opening during the second voltage step.
	Figure 8. Concentration-dependent inhibition of WT and Y652A mutant hERG channels expressed in Xenopus oocytes. (A) Representative traces of WT and mutant hERG currents in the presence and absence of the indicated concentrations of hydroxyzine. The effects of the drug on WT and Y652A tail currents were recorded at −140 mV (instead of −60 mV as above) following a 4 s activating pulse. (B) The concentration-response curves were fitted with a logistic dose-response equation to yield IC50 values of 5.9±1.5 μmol/L (n=3) and 35.1±50.5 μmol/L (n=4) for WT and Y652A hERG channels, respectively.
	Figure 9. Docking of hydroxyzine within the inner cavity of a HERG channel homology model. (A) The major microspecies of hydroxyzine protonation at pH 7.4. (B) Inner view of hydroxyzine docked to the ligand binding site in the HERG channel. Hydroxyzine is shown in stick form and the dotted line indicates a hydrogen bond between the blocker and channel. The HERG channel inhibitor hydroxyzine shows interactions with THR623 (subunit A) and TYR652 (subunit C). (C) PoseView analysis of protein-ligand interactions. Hydrogen bonding is depicted as a dotted line between the participating atoms. The green lines with residue names and numbers indicate hydrophobic interactions between the drug and channel. Note the hydrogen bond between the hydrogen atom of the protonated nitrogen (in the hydroxyzine molecule) and the carbonyl oxygen atom of THR623(I).

References [+] :

Choe, A novel hypothesis for the binding mode of HERG channel blockers. 2006, Pubmed