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(Circulation. 2004;109:26-29.)
© 2004 American Heart Association, Inc.

Brief Rapid Communications

Unusual Effects of a QT-Prolonging Drug, Arsenic Trioxide, on Cardiac Potassium Currents

Benoit Drolet, PhD; Chantale Simard, PhD; Dan M. Roden, MD

From the Division of Clinical Pharmacology, Vanderbilt University School of Medicine, Nashville, Tenn.

Correspondence to Dan M. Roden, MD, Professor of Medicine and Pharmacology, Director, Division of Clinical Pharmacology, 532 Robinson Research Building, Vanderbilt University School of Medicine, Nashville, TN 37232. E-mail dan.roden{at}vanderbilt.edu

Received October 8, 2003; revision received November 4, 2003; accepted November 14, 2003.

	Abstract

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Background— Cases of QT prolongation, torsades de pointes, and sudden death have been reported with arsenic trioxide (As₂O₃), a highly effective agent for acute promyelocytic leukemia. In this study, we evaluated the effects of As₂O₃ on repolarizing cardiac ion currents.

Methods and Results— In HERG- or KCNQ1+KCNE1-transfected CHO cells (n=32; total), As₂O₃ caused concentration-dependent block of both I_Kr and I_Ks, with an IC₅₀ for tail current block of 0.14±0.01 µmol/L for I_Kr and 1.13±0.06 µmol/L for I_Ks. In contrast to other QT-prolonging drugs, As₂O₃ also activated a time-independent current that additional experiments identified as I_K-ATP.

Conclusions— As₂O₃ blocks both I_Kr and I_Ks at clinically relevant concentrations. On the other hand, it also activates I_K-ATP, which maintains normal repolarization. We infer that variability in the extent of QT interval prolongation and onset of ventricular arrhythmias during arsenic therapy represents competing effects to block and activate multiple repolarizing potassium currents.

Key Words: torsades de pointes • drugs • ion channels

	Introduction

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Approximately 20% to 30% of patients with acute promyelocytic leukemia (APL) relapse with current standard all-trans retinoic acid and anthracycline-based chemotherapy regimen.¹ In the mid-1990s, studies from China reported that arsenic trioxide (As₂O₃) achieved complete remission in as many as 90% of patients with APL,^2,3 and additional trials have confirmed these results.^4,5 However, treatment has also been associated with QT prolongation, torsades de pointes, and sudden death.^4,6–8 Because most of the patients receiving As₂O₃ have been exposed to cardiotoxic chemotherapy, cardiac dysfunction is thought to be universal before As₂O₃ therapy begins.^6,9 In addition, hypokalemia and hypomagnesemia are among the most common As₂O₃-related side effects.¹ Thus, it has been proposed that QT prolongation and ventricular arrhythmias associated with arsenic could be exacerbated by concurrent electrolyte disturbances or previous chemotherapy-induced cardiac damage.^4,6–8

Recently, clinically relevant concentrations of As₂O₃ (1 to 10 µmol/L) have been reported to prolong the action potential duration in guinea pig papillary muscle.⁹ In another study using rabbit hearts, polymorphic ventricular tachycardia was observed with As₂O₃ 30 µmol/L.¹⁰ In this study, we therefore investigated the effects of As₂O₃ on cardiac repolarizing currents and identified an unexpectedly complex profile that may underlie variability in the arrhythmogenic potential of arsenic.

	Methods

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Experiments were performed in CHO cells transfected with 2 µg (each) of HERG, KCNQ1+KCNE1, or Kir6.2+SUR2A cDNAs. Kir6.2 and SUR2A cDNAs were kindly provided by Dr Joseph Bryan, Baylor College of Medicine, Houston, Tex. Cells were transfected using FuGENE 6 (Roche Applied Science). Green fluorescent protein (GFP) was coexpressed to identify transfected cells. All whole cell currents were recorded at 22°C to 23°C. Cells were held at -80 mV and pulsed to -40 to +60 mV for 1 second (HERG) or 5 seconds (KCNQ1+KCNE1), and tail currents were measured at -40 mV. The maximal tail current amplitudes were sampled after the capacitive transient in a 20-ms window. The composition of extracellular and internal pipette solutions for the HERG and KCNQ1+KCNE1 experiments were as described previously.¹¹ For the Kir6.2+SUR2A experiments, cells were held at 0 mV and pulsed to -100 to +50 mV for 100 ms. The composition of solutions for these experiments was as described previously.¹² As₂O₃ solutions of the extracellular buffer (30 nmol/L to 10 µmol/L) were prepared daily. Glibenclamide and pinacidil solutions in DMSO were prepared daily. The same concentration of DMSO (0.1% vol/vol) was also present in baseline and washout buffer solutions. As₂O₃, glibenclamide, and pinacidil were obtained from Sigma.

Statistical Analysis
Data are presented as mean±SEM. Concentration-dependent block of HERG or KCNQ1+KCNE1 tail current was tested by the Hotelling t² test. P<0.05 was considered statistically significant.

	Results

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Figure 1A shows currents elicited in a HERG-transfected cell under baseline conditions and after 20 minutes of As₂O₃ 100 nmol/L (Figure 1B). In this cell, As₂O₃ 100 nmol/L caused a 60% reduction of the tail current. Unexpectedly, however, there was also an increase in time-independent outward current (Figure 1B, arrow). Figures 1C and 1D show currents elicited before and during As₂O₃ 1 µmol/L. In this cell, As₂O₃ 1 µmol/L caused a 92% reduction of the tail current and clear activation of this time-independent outward current (Figure 1D). Figure 1E shows concentration dependence of the effect on I_Kr tail current, with an IC₅₀ of 0.14±0.01 µmol/L. Figures 1F through 1J show a similar concentration-dependent block of I_Ks (assessed by reduction of tail currents) and, again, activation of a time-independent outward current. I_Ks was 1 order of magnitude less sensitive to block, with an IC₅₀ of 1.13±0.06 µmol/L. Nearly all drugs that cause QT prolongation and torsades de pointes block I_Kr, and some (eg, quinidine and azimilide) also block I_Ks, often with somewhat higher IC₅₀ values.^13,14 However, activation of an outward current, as in Figure 1, has not been reported previously. Because this effect may reflect activation of a current endogenous to CHO cells, we next step-studied cells transfected with GFP only. Figure 2A shows currents elicited in a GFP-transfected cell under baseline conditions, and Figure 2B shows currents elicited after 20 minutes of As₂O₃ 1 µmol/L. This gating pattern is reminiscent of I_K-ATP, and Figure 2C shows that the current was blocked by subsequent exposure to the I_K-ATP blocker glibenclamide 10 µmol/L. Figures 2D through 2F show near-identical behaviors with the I_K-ATP activator pinacidil 10 µmol/L (Figure 2E) and 10 minutes after the addition of glibenclamide 10 µmol/L (Figure 2F). To additionally test the hypothesis that As₂O₃ activates I_K-ATP, we studied Kir6.2+SUR2A-transfected CHO cells. Figures 2G through 2I show results similar to those in Figures 2A through 2C, demonstrating activation of I_K-ATP by As₂O₃ 1 µmol/L. Figures 2J through 2L show that pinacidil 10 µmol/L activates a similar current and glibenclamide 10 µmol/L blocks this current, as expected.

View larger version (29K): [in this window] [in a new window]   Figure 1. A and B, Currents elicited by 1-second steps in a HERG-transfected CHO cell at baseline and after 20 minutes of As2O3 100 nmol/L. C and D, Currents elicited before and after 20 minutes of As2O3 1 µmol/L. Arrows in B and D indicate a time-independent outward current elicited by As2O3. E, HERG tail current amplitude, measured at +20 mV (n=16), normalized to control, plotted as a function of As2O3 concentration, and fitted to the Hill equation. F and G, Currents elicited by 5-second steps in a KCNQ1+KCNE1-transfected CHO cell at baseline and after 20 minutes of As2O3 1 µmol/L. H and I, Currents elicited before and after 20 minutes of As2O3 10 µmol/L. J, KCNQ1+KCNE1 tail current amplitude, measured at +60 mV (n=16), normalized to control, plotted as a function of As2O3 concentration, and fitted to the Hill equation.

View larger version (43K): [in this window] [in a new window]   Figure 2. A, B, and C, Currents elicited by 1-second steps in a GFP-transfected CHO cell under baseline conditions, after 20 minutes of As2O3 1 µmol/L, and 20 minutes after the addition of glibenclamide 10 µmol/L, respectively. D, E, and F, Currents elicited by 1-second steps in a GFP-transfected CHO cell under baseline conditions, after 20 minutes of pinacidil 10 µmol/L, and 20 minutes after the addition of glibenclamide 10 µmol/L. G, H, and I, Currents elicited by 100-ms steps in a Kir6.2+SUR2A-transfected CHO cell under baseline conditions, after 20 minutes of As2O3 1 µmol/L, and 20 minutes after the addition of glibenclamide 10 µmol/L. J, K, and L, Currents elicited by 100-ms steps in a Kir6.2+SUR2A-transfected CHO cell under baseline conditions, after 20 minutes of pinacidil 10 µmol/L, and 20 minutes after the addition of glibenclamide 10 µmol/L.

	Discussion

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Our results show that As₂O₃ is a potent blocker of I_Kr (IC₅₀, 0.14±0.01 µmol/L) and I_Ks (IC₅₀, 1.13±0.06 µmol/L). It has been shown that I_Kr block may lead to triggered tachyarrhythmias and sudden death.¹⁵ Moreover, adding the effect of an I_Ks blocker on an already-compromised I_Kr has been shown to additionally decrease the repolarization reserve, potentiating the action potential–prolonging effect of the I_Kr blocker.¹⁶ These concentrations are well within the therapeutically relevant range. In one study of 8 patients with relapsed APL, mean peak plasma arsenic concentration was 6.85 µmol/L (range, 5.54 to 7.30 µmol/L), and after 10 hours, it was 1 µmol/L.²

Unexpectedly, our data also showed that exposure of HERG- or KCNQ1+KCNE1-transfected CHO cells to As₂O₃ activates a time-independent outward current. This I_K-ATP-like current was also activated to a similar extent in GFP-transfected CHO cells exposed to As₂O₃ and could be reversed by adding glibenclamide 10 µmol/L, an I_K-ATP blocker.¹⁷ Indeed, the effects of As₂O₃ in GFP-transfected CHO cells were comparable to those of the I_K-ATP activator pinacidil.¹⁷ Interestingly, one of the most common non–life-threatening side effects of As₂O₃ is hyperglycemia, observed in up to 45% of patients.⁴ This effect has also been associated with other I_K-ATP activators, such as diazoxide.¹⁷ Moreover, depletion of intracellular ATP, as seen for example during cardiac ischemia, has been shown to activate I_K-ATP.¹⁷ Interestingly, arsenic is also recognized to uncouple cardiac mitochondrial oxidative phosphorylation.¹⁸ The mechanism is thought to be related to competitive substitution of arsenic for inorganic phosphate in the formation of ATP.¹⁸ As a result, arsenic-induced reduction of cardiac phosphorylation likely causes depletion of intracellular ATP and thus activation of cardiac I_K-ATP.

Therefore, while blocking both I_Kr and I_Ks at therapeutic concentrations, thereby producing a severe lesion in repolarization reserve, As₂O₃ also activates I_K-ATP, which may partially restore repolarization reserve and thus contribute to variability in the extent of QT-interval prolongation and onset of ventricular arrhythmias during arsenic therapy.

Conclusions
As₂O₃ is a potent blocker of both I_Kr and I_Ks. On the other hand, it also activates cardiac I_K-ATP, which may blunt QT prolongation and arrhythmia risk by restoring repolarization reserve. The risk of torsades de pointes can be reduced by adherence to guidelines for safe use of the drug. In addition, variability in the extent of QT effects among patients may reflect this unusual combination of potassium channel actions.

	Acknowledgments

 
Supported by the United States Public Health Service (HL46681, HL49989). Dr Roden holds the William Stokes Chair in Experimental Therapeutics, a gift of the Dai-ichi Corporation. Benoit Drolet is the recipient of a postdoctoral fellowship award from the Canadian Institutes of Health Research and the Heart and Stroke Foundation of Canada. The authors also thank Donna Choate for technical assistance.

	References

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