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Cardiovascular Research 2003 57(3):660-669; doi:10.1016/S0008-6363(02)00726-5
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Copyright © 2003, European Society of Cardiology

Effects of propafenone and its main metabolite, 5-hydroxypropafenone, on HERG channels

Cristina Arias*, Teresa González, Ignacio Moreno, Ricardo Caballero, Eva Delpón, Juan Tamargo and Carmen Valenzuela

Institute of Pharmacology and Toxicology, CSIC/UCM, School of Medicine, Universidad Complutense, 28040 Madrid, Spain

c.arias{at}ift.csic.es

* Corresponding author. Tel.: +34-91-394-1474; fax: +34-91-394-1470.

Received 26 August 2002; accepted 8 October 2002


   Abstract
Top
 Abstract
1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
 References
 
Objectives: Propafenone is a class Ic antiarrhythmic drug used to maintain sinus rhythm in patients with atrial fibrillation. During chronic therapy, it undergoes extensive first-pass hepatic metabolism to 5-hydroxypropafenone. In the present study we have analysed the effects of propafenone and 5-hydroxypropafenone on HERG current. Methods: The whole-cell configuration of the patch-clamp technique was used in CHO cells stably transfected with the gene encoding HERG channels. Results: Propafenone and 5-hydroxypropafenone (2 µM) inhibited HERG current by 78.7±2.3% (n=7) and 71.1±4.1% (n=7, P0.05) when measured at the end of 5-s depolarizing pulses to –10 mV. Block measured at the maximum peak of tail currents recorded at –60 mV was similar for propafenone (78.3±2.0%, n=7, P0.05) and higher for 5-hydroxypropafenone (79.3±1.5%, n=7, P<0.05). Propafenone and 5-hydroxypropafenone shifted the midpoint of the activation curve by –10.2±0.9 mV (n=7, P<0.01) and –7.4±1.1 mV (n=10, P<0.01), respectively. Both drugs accelerated the deactivation and the inactivation process of HERG current. Propafenone, but not 5-hydroxypropafenone, inhibited to a higher extent HERG current at the end of 5-s depolarizing pulses to 0 mV than after promoting the transition of HERG channels from the inactivated to the opened state. Conclusions: These results indicate that propafenone and its main active metabolite, 5-hydroxypropafenone, block HERG channels to a similar extent by binding predominantly to the open state of the channel.

KEYWORDS Antiarrhythmic agents; Arrhythmia (mechanisms); Ion channels; K-channel; Membrane currents


   1. Introduction
Top
 Abstract
 1. Introduction
2. Materials and methods
 3. Results
 4. Discussion
 References
 
Propafenone is a class Ic antiarrhythmic drug widely used for conversion of atrial fibrillation to sinus rhythm [1–7], to prevent recurrences in patients with atrial fibrillation [8] and, occasionally, in the treatment of certain ventricular tachycardias [9]. As such, it is able to depress intracardiac conduction velocity as a consequence of its binding to the open state of cardiac Na+ channels [10]. However, propafenone, at the same range of concentrations, inhibits several K+ currents: ITO, IKur, IKr, IKs and IK1 [11–15] and, at higher concentrations, the L-type Ca2+ current (ICa) [16,17]. Furthermore, propafenone also exhibits β-adrenergic receptor blocking effects [18,19]. During chronic therapy, propafenone undergoes extensive first-pass hepatic metabolism dependent of the CYP2D6 isoform of the cytochrome P450 [20–22] to 5-hydroxypropafenone [23], which accumulates in plasma arising similar concentrations than those of propafenone [9,21,24–27]. This active metabolite is more potent than propafenone as Na+ channels blocker and, like the parent compound, it has been classified as a class Ic antiarrhythmic drug [26–29], although it is less potent than propafenone to block L-type Ca2+ and hKv1.5 channels, as well as β-adrenergic receptors [1,14,26,29–32].

Propafenone and 5-hydroxypropafenone have been reported to prolong the cardiac action potential in guinea pig atrial fibres, rabbit sino-atrial node cells, as well as in atrial and ventricular muscle fibres from guinea pig chronically treated with propafenone [18,31,33–35]. These observations suggest that propafenone and/or its active metabolite block K+ currents involved in the cardiac repolarization, both at the atrial and the ventricular level. It has been previously described that propafenone blocks HERG channels [13,15,36]. However, the effects induced by 5-hydroxypropafenone on cardiac K+ currents other than atrial ITO and hKv1.5 [14,37] are unknown. Therefore, the purpose of the present study was to analyse and compare the effects of 5-hydroxypropafenone and propafenone on HERG channels, which determine the ventricular action potential duration [38,39]. Preliminary report of the present study has been published in abstract form [40].


   2. Materials and methods
Top
 Abstract
 1. Introduction
 2. Materials and methods
3. Results
 4. Discussion
 References
 
2.1 Cell culture
CHO cells stably transfected with the gene encoding HERG channels were cultured at 37 °C in Ham's-F12 medium supplemented with geneticine (600 µg/ml), penicillin–streptomycin (800 UI and 200 µg/ml, respectively) and bovine serum 10%, in a 5% CO2 atmosphere. Cultures were passaged every 3–5 days by use of a brief trypsin treatment. Before experimental use, the cells were removed from the dish with a rubber policeman, a procedure that left the majority of the cells intact. The cell suspension was stored at room temperature (21–23 °C) and used within the next 12 h in all the experiments reported.

2.2 Electrophysiological recording
The intracellular pipette-filling solution contained (in mM): K–aspartate 80, KCl 50, phosphocreatine 3, KH2PO4 10, MgATP 3, HEPES–K 10, and EGTA 5 and was adjusted to pH 7.25 with KOH. The bath solution contained (in mM): NaCl 130, KCl 4, CaCl2 1.8, MgCl2 1, HEPES–Na 10, and glucose 10, and was adjusted to pH 7.4 with NaOH. Propafenone (Sigma Chemicals, St. Louis, MO, USA) and 5-hydroxypropafenone (a gift from Knoll AG, Ludwigshafen, Germany) were dissolved in distilled deionized water to yield stock solutions of 10 mM from which further dilutions were made. HERG currents were recorded at room temperature (21–23 °C) using the whole-cell patch-clamp technique [41] with an Axopatch 200A patch-clamp amplifier (Axon Instruments, Foster City, CA, USA). Micropipettes were pulled from borosilicate glass capillary tubes (GD-1; Narishige, Tokyo, Japan) on a programmable horizontal puller (Sutter Instrument Co., San Rafael, CA, USA) and heat-polished with a microforge (Narishige). Micropipettes resistance was 1–3 M{Omega}. Mean uncompensated access resistance was 3.7±0.3 M{Omega} (n=13), and cell capacitance was 14.3±1.1 pF (n=13). Since the mean maximum current was 0.8±0.1 nA (n=13), the averaged voltage error of 2.8±0.4 mV (n=13). In all experiments reported, voltage error was <5 mV.

HERG currents were filtered at 1 kHz and sampled at 2 kHz. Cells were held at –80 mV. After control data were obtained, bath perfusion was switched to drug-containing solution. The effects of drug infusion were monitored with test pulses to –10 mV applied every 30 s until steady state was obtained. Steady-state current–voltage relationships (IV) were obtained by averaging the current over a small window (2–5 ms) at the end of 5-s depolarizing pulses from –80 mV up to +50 mV in 10-mV steps. Between –80 and –50 mV only passive linear leak was observed and least squares fit to these data were used for passive leak correction. Deactivating tail currents were recorded at –60 mV. The activation curves were obtained from the tail current amplitude measured just after the capacitive transient. Other voltage-clamp pulse protocols are described in the Results section. Command potentials, data acquisition and measurements were done using the Clampfit program of pClamp 6.0.1, Origin 6.0.1 (Microcal Software, Northampton, MA, USA) and by custom-made analysis programs.

Deactivation was fitted to a biexponential process:

Formula (1)
where {tau}1 and {tau}2 are the system time constants, A1 and A2 are the amplitudes of each component of the exponential, and C is the baseline value. Half-maximal voltages (Eh) and slope factors (s) of activation were determined by fitting data with a Boltzmann equation: y=1/[1+ exp(–(E–Eh)/s)]. The curve-fitting procedure used a non-linear least-squares (Gauss–Newton) algorithm; results were displayed in linear and semilogarithmic format, together with the difference plot. Goodness of fit and the required number of exponential components were judged by comparing {chi}2 values statistically (F test) and by inspection for systematic non-random trends in the difference plot. Drug-induced block was measured at the end of depolarizing pulses of 5 s in duration from –80 to –10 mV, unless indicated otherwise.

Voltage dependence of block was determined as follows: leak-corrected tail current in the presence of drug was normalized to matching control to yield the fractional block at each voltage [f=1–(IDrug/IControl)]. The voltage dependence of block was fitted to:

Formula (2)
where z, F, R and T have their usual meaning, {delta} represents the fractional electrical distance [42], i.e., the fraction of the transmembrane electrical field sensed by a single charge at the receptor site and KD* represents the apparent dissociation constant at the reference potential (0 mV).

2.3 Statistical methods
Results are expressed as mean±S.E.M. Direct comparisons between mean values in control conditions and in the presence of drug for a single variable were performed by a paired Student's t-test. Differences were considered significant if the p value was less than 0.05.


   3. Results
Top
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
4. Discussion
 References
 
3.1 HERG block induced by propafenone and 5-hydroxypropafenone
Clinically effective free plasma concentrations of propafenone and 5-hydroxypropafenone correspond to in vitro concentrations of 0.2–0.6 µM and 0.08–1.44 µM, respectively [9,21]. However, after chronic treatment both drugs accumulates in the heart, so that cardiac concentrations were 10- and 20-fold higher than those reported in plasma (equivalent to in vitro concentrations of 2–6 µM and 1.6–28.8 µM for propafenone and 5-hydroxypropafenone, respectively) [25]. In the present study, we have analysed the effects of the lowest drug concentration (2 µM) of propafenone and 5-hydroxypropafenone found in the human myocardium.

Fig. 1 shows original HERG current records obtained after applying 5-s depolarizing pulses from a holding potential of –80 to +50 mV in 10-mV steps in the absence and in the presence of 2 µM propafenone (Fig. 1A) and 5-hydroxypropafenone (Fig. 1B). Tail currents were elicited upon repolarization of the membrane potential to –60 mV after each voltage step. Depolarizations to values more positive than –50 mV elicited an outward K+ current followed by a hooked deactivating tail current. The maximum outward current was observed at {approx}–10 mV and it decreased at more positive membrane potentials due to the fast C-type inactivation of HERG channels [43]. Thus, steady-state drug-induced block was measured at the end of 5-s depolarizing pulses to –10 mV. Propafenone (2 µM) similarly inhibited HERG current both measured at the end of 5-s depolarizing pulses to –10 mV and at the maximum tail currents recorded at –60 mV (78.7±2.3% vs. 78.3±2.0%, n=7, P0.05). In contrast, 5-hydroxypropafenone inhibited HERG currents to a higher extent when measured at the maximum peak tail currents than at the end of depolarizing pulses from a holding potential to –10 mV (71.1±4.1% vs. 79.3±1.5%, n=7, P<0.05). From the calculated values of inhibition of the current at the end of 5-s pulses to –10 mV and assuming a Hill coefficient (nH) of 1, EC50 values averaged 0.55±0.07 µM (n=7) and 0.88±0.20 µM (n=7) for propafenone and 5-hydroxypropafenone, respectively. The EC50 value calculated for propafenone from the experiments reported here is in agreement to that previously reported [13,15,36]. All these results may suggest that both drugs block K+ efflux by binding to the open state of the channel. Moreover, the higher degree of block induced by 5-hydroxypropafenone measured at the maximum peak tail currents than at the end of 5-s depolarizing steps to –10 mV, indicate that the metabolite exhibits a higher affinity for the open state than the parent compound.


Figure 1
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  		Fig. 1 Original records obtained upon depolarization from a holding potential of –80 to +50 mV in 10-mV steps and upon repolarization to –60 mV. Current records obtained in the absence and in the presence of 2 µM propafenone (A) and 5-hydroxypropafenone (B).

 
3.2 Voltage dependence of HERG channels block induced by propafenone and 5-hydroxypropafenone
Fig. 2A shows the IV relationships of HERG current obtained in the absence and in the presence of 2 µM propafenone or 5-hydroxypropafenone. In the absence of drug, the IV relationship exhibits the characteristic bell-shape that increases from –40 mV to {approx}–10 mV and, due to the fast C-type inactivation of HERG channels, it decreased with further depolarization [43,44]. Propafenone and 5-hydroxypropafenone decreased HERG current at all membrane potentials tested. Fig. 2B shows the activation curves obtained under control conditions and in the presence of 2 µM of either drug. The activation curves obtained in the presence of drug were normalized to match those obtained under control conditions (dashed lines). Propafenone and 5-hydroxypropafenone shifted the Eh values toward more negative membrane potentials by –10.2±0.9 mV (n=7, P<0.01) and –7.4±1.1 mV (n=10, P<0.01), respectively (Fig. 2B), suggesting an open channel block mechanism. Moreover, as it is shown in Fig. 2C, block steeply increased in the range of membrane potentials coinciding with the activation of HERG channels. At membrane potentials positive to 0 mV, block increased in a shallower manner. It is unlikely that this shallow voltage dependence was due to channel gating, since HERG activation had reached saturation over this voltage range (Fig. 2B). Propafenone and 5-hydroxypropafenone are bases and have a tertiary amine group with pKa=9. Therefore, at the intracellular pH of 7.25 propafenone and 5-hydroxypropafenone are mainly present in their charged form and the observed voltage dependence of block could be due to the effect of the transmembrane electrical field on the interaction between the propafenone ion and the channel receptor. If these drugs reach their receptor site from the inside, according to a Woodhull formalism [42], channel blockade is expected to increase in a voltage-dependent manner. Indeed, fitting the data to a Boltzmann function based on the Woodhull model, we obtained averaged fractional electrical distances of 0.09±0.01 (n=6) and 0.06±0.01 (n=6, P0.05) for propafenone and 5-hydroxypropafenone, respectively.


Figure 2
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  		Fig. 2 Voltage-dependent block of HERG channels induced by propafenone and 5-hydroxypropafenone. (A) IV relationships (5 s isochronal) of HERG channels obtained in the absence and in the presence of 2 µM of each drug. (B) Activation curves of HERG channels obtained under control conditions and in the presence of propafenone and 5-hydroxypropafenone (2 µM). Dotted lines reflect the normalized activation curves obtained in the presence of drug and matching control values. Note that both drugs shifted the activation curve in the hyperpolarizing direction. (C) Relative current vs. membrane potential. Block increases in the range of membrane potentials that coincides with the activation of HERG channels. At membrane potentials positive to 0 mV block slightly increased consistent with * values of {approx}0.09 for both drugs. Dashed lines represent the activation curves obtained under control conditions. Each point represents the mean±S.E.M. of six or seven experiments. ***, P<0.001 vs. control.

 
3.3 Time-dependent HERG channels block induced by propafenone and 5-hydroxypropafenone
Fig. 3A shows original records obtained after depolarizing to –10 mV under control conditions and in the presence of 2 µM propafenone or 5-hydroxypropafenone. In both cases, block induced at the end of the depolarizing pulse was higher than that observed at the beginning. In order to quantify this time dependence of block, the relative current (IDrug/IControl) was plotted and fitted by an exponential function (Fig. 3B). It was observed that at the beginning of the depolarizing pulse to –10 mV there was an instantaneous block that increased during depolarization pulse with an exponential kinetics. The time constant of onset of block during depolarizing pulses averaged 1486±183 ms (n=6) and 804±59 ms (n=5, P<0.05) for propafenone and 5-hydroxypropafenone, respectively. In the presence of propafenone and 5-hydroxypropafenone, an initial ‘instantaneous’ component of block was observed at the beginning of the depolarizing pulse (t=0 ms), which averaged 53.8±7.8% (n=6) and 46.7±8.7% (n=5), respectively, which was followed by the previously described time-dependent increase in block.


Figure 3
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  		Fig. 3 Time-dependent block induced by propafenone and 5-hydroxypropafenone during depolarization. (A) Current traces obtained after depolarizing the cell membrane from –80 to –10 mV in the absence and in the presence of propafenone or 5-hydroxypropafenone (2 µM). (B) Plot of the relative current (IDrug/IControl) vs. time of depolarization. Current exponentially decreased during depolarization.

 
Time dependence of block was also visible in the deactivation tail currents. Fig. 4A shows tail currents recorded in the absence and in the presence of propafenone or 5-hydroxypropafenone (2 µM). Both drugs decreased the maximum tail current and accelerated the deactivation process (Fig. 4B). This faster deactivation in the presence of drug may suggest an interaction with an activated or open state of the channel. If these drugs would bind to the activated state of the channel, we would expect a delay in the activation kinetics. Therefore, we analysed this process after applying a double pulse protocol consisting in a pulse to +40 mV of variable duration (between 50 and 1100 ms) followed by a pulse to –60 mV to record the tail currents. The HERG current kinetics was measured by plotting the maximum tail current against the duration of the prepulse to +40 mV. The kinetics of the activation process was not modified by propafenone (123.0±24.3 ms vs. 135.1±13.4 ms, n=4, P0.05) or 5-hydroxypropafenone (139.5±25.0 ms vs. 122.8±14.6 ms, n=5, P0.05).


Figure 4
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  		Fig. 4 Time-dependent block induced by propafenone and 5-hydroxypropafenone on deactivation. (A) Tail currents recorded upon repolarization to –60 mV after a 5-s depolarizing pulse to –10 mV in the absence and in the presence of 2 µM propafenone or 5-hydroxypropafenone. (B) This panel shows both tail current traces normalized to match control values. Note that both propafenone and 5-hydroxypropafenone accelerated the deactivation kinetics.

 
In order to explore the effects of propafenone and 5-hydroxypropafenone on the inactivation kinetics of HERG channels, the membrane potential was held at –80 mV and a three pulse protocol was applied, consisting in a 2-s depolarizing prepulse to +40 mV followed by a 20-ms hyperpolarizing pulse to –120 mV before a 20-ms test pulse to different voltages between 0 and +50 mV (Fig. 5). Propafenone and 5-hydroxypropafenone accelerated the inactivation at all membrane potentials tested, with the exception of +50 mV in the case of 5-hydroxypropafenone (Fig. 5A). Block induced by both drugs was measured at the maximum peak current at 0 mV after a 20-ms hyperpolarizing pulse to –120 mV, which promotes the I->O transition and compared with that induced by propafenone and 5-hydroxypropafenone at the end of 5-s depolarizing pulses at the same potential (Fig. 5B). Propafenone produced a higher degree of block at the end of 5-s depolarizing pulses to 0 mV than at the maximum peak current at 0 mV after a 20-ms hyperpolarizing pulse to –120 mV; whereas 5-hydroxypropafenone induced a similar degree of block under both experimental conditions.


Figure 5
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  		Fig. 5 Effects of 2 µM propafenone or 5-hydroxypropafenone on HERG inactivation kinetics. Current records were obtained by using the protocol shown in the top of the figure. (A) Top panels show current records of test pulses obtained in the absence and in the presence of 2 µM of each drug. Bottom panels show plots of the time constant of inactivation at different membrane potentials in the absence and in the presence of each drug. (B) Graph showing the degree of block induced by propafenone or 5-hydroxypropafenone measured at the end of 5-s depolarizing pulses to 0 mV (End 5 s) or at the maximum peak current recorded at a test pulse to 0 mV after a depolarizing test pulse to +40 mV during 2 s and followed by a 20-ms hyperpolarizing pulse to –120 mV (Maximum) indicated by an arrow in the pulse protocol. Each point represents the mean±S.E.M. of four or five experiments. *: P<0.05 vs. control conditions.

 

   4. Discussion
Top
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
References
 
In the present paper the effects of propafenone and 5-hydroxypropafenone on HERG channels have been studied. We found that, at relevant clinical concentrations, propafenone and its main metabolite, 5-hydroxypropafenone, block to a similar extent HERG channels.

4.1 Effects of propafenone on HERG channels
Propafenone blocks HERG channels in a voltage- and time-dependent manner, consistent with drug binding preferentially to the open state of these channels. There are several pieces of evidence that support an open channel block mechanism: (a) propafenone induced block was voltage-dependent, in such a way that block steeply increased in the activation range of the channel; (b) it shifted the activation curve towards more negative membrane potentials; (c) propafenone-induced block was time-dependent, exponentially increasing during depolarization of the cell membrane; (d) time-dependent block was also evident in the tail currents, whose kinetics were accelerated in the presence of the drug; and (e) propafenone did not modify the activation kinetics of HERG current, suggesting that it does not bind to an activated state of the channel. In addition, propafenone accelerates the inactivation process of the current and the block measured at the end of 5-s depolarizing pulses was higher than that measured at the maximum peak current after the application of a 20-ms pulse to –120 mV (that promotes the transition from the inactivated to the opened state of HERG channels). Whereas the acceleration of the inactivation kinetics could be interpreted as another piece of evidence of open channel block, the higher degree of block observed at the end of 5-s depolarizing pulses to 0 mV in comparison with the degree of block measured at the same potential after a 20-ms hyperpolarizing pulse to –120 mV seems to indicate a higher affinity of propafenone for the inactivated state. Under this framework, the acceleration of the inactivation kinetics could be the consequence of the drug binding to the inactivated state. All these results suggest that propafenone inhibits HERG current after binding to the open and the inactivated states of the channel. These results are broadly similar to those previously reported for HERG channels expressed in Xenopus oocytes [15], with the exception that in the present study clinical propafenone concentrations resulted to inhibit these channels. This difference is likely due to the different expression system used in both studies. In fact, it has been reported that when studying the effects of lipophilic drugs (like propafenone or its metabolite) that act from the inside of the cell membrane, the apparent drug potency is approximately 10–100 times lower when using whole oocyte recordings than cell-free patch recordings (free of yolk) [45,46]. Also, in the presence of propafenone, an instantaneous block was observed at t=0 ms. This initial block, which appears before channel opening could be attributed to the drug interaction with a non-conducting state of the channel.

4.2 Effects of 5-hydroxypropafenone on HERG channels
Potency of 5-hydroxypropafenone and propafenone to block HERG channels were similar. Moreover, most HERG channel block characteristics observed for 5-hydroxypropafenone were similar to those found with the parent compound: (a) block induced by 5-hydroxypropafenone was voltage- and time-dependent, increasing in the range of membrane potentials that coincides with the activation of the channel; (b) 5-hydroxypropafenone shifted the activation curve towards more negative membrane potentials; (c) Block during depolarization to –10 mV exponentially increased with time; (d) The deactivation kinetics of HERG currents was accelerated in the presence of the drug; and finally (e) block measured at the end of 5-s depolarizing pulses to 0 mV was similar to that measured at the maximum peak current recorded at 0 mV after the application of a hyperpolarizing pulse to –120 mV. 5-Hydroxypropafenone, like propafenone, accelerated the inactivation kinetics of HERG current. However, in contrast to the parent compound, 5-hydroxypropafenone induced a higher degree of block when it was measured at the maximum peak of the tail currents recorded at –60 mV than when it was measured at the end of 5-s depolarizing pulses to –10 mV, thus suggesting a higher affinity for the open state of the channel. Moreover, block measured at the end of 5-s depolarizing pulses to 0 mV and at the maximum peak current recorded at 0 mV after applying a 20-ms hyperpolarizing pulse to –120 mV was similar, also suggesting a higher affinity for the open than for the inactivated state of HERG channels. Similarly to propafenone, an instantaneous block that can be attributed to a drug interaction with a closed state of the channel was observed at t=0 ms.

4.3 Clinical implications
Propafenone is extensively biotransformed to its main metabolite, 5-hydroxypropafenone, by a specific hepatic isoenzyme, cytochrome P-450 CYP2D6, whose activity is polymorphically distributed in humans [9,21]. Approximately 10% of patients have a deficiency in CYP2D6 (poor metabolizers), which results in plasma concentrations of 5-hydroxypropafenone much lower than the remaining population (extensive metabolizers) [32]. This explains why during long-term therapy total 5-hydroxypropafenone concentration at the steady-state varied from 6 to 73% [24].

The mean effective therapeutic plasma concentrations of propafenone varied from 3.14 to 1812 ng/ml (equivalent to 0.9–5.3 µM) [9,21,47]. Because propafenone is about 90% protein bound [9], in vitro concentrations of 0.2–0.6 µM probably correspond in action to clinically effective free drug concentrations. On the other hand, plasma concentrations of 5-hydroxypropafenone varied from 30 to 513 ng/ml (equivalent to 0.08–1.44 µM) [9,21,32]. In the present study, the EC50 values for the blockade of HERG channels of propafenone and its main metabolite were 0.6 and 0.9 µM, respectively. Thus, both drugs block HERG channels at concentrations within the therapeutic plasma levels. Moreover, 5-hydroxypropafenone is less protein bound, indicating that the free plasma concentration (pharmacologically active) of the metabolite may be high enough to contribute to the clinical effects of propafenone therapy [9].

4.4 Conclusions
This is the first study in which the effects of 5-hydroxypropafenone on HERG channels have been studied. The present results demonstrate that both the parent and the hydroxy-metabolite inhibit in a concentration-, voltage- and time-dependent manner the HERG current, mainly after binding to the open state of HERG channels. The efficacy of 5-hydroxypropafenone to block these channels at therapeutic concentrations could contribute to the previously reported efficacy of propafenone.

Time for primary review 17 days.


   Acknowledgements
 
This study has been supported by CICYT SAF99-0069 and FIS 01/1130 Grants. The authors want to thank Drs. S. Nattel, T.E. Hébert and M. Weerapura (University of Montreal, Montreal Heart Institute and McGill University) for providing us with the cell line expressing HERG channels and to Ms. Guadalupe Pablo for her excellent technical assistance.


   References
Top
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
 References
 

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