Channel Openings Are Necessary but not Sufficient for Use-dependent Block of Cardiac Na Channels by Flecainide: Evidence from the Analysis of Disease-linked Mutations

Na channel blockers such as flecainide have found renewed usefulness in the diagnosis and treatment of two clinical syndromes arising from inherited mutations in SCN5A, the gene encoding the subunit of the cardiac voltage–gated Na channel. The Brugada syndrome (BrS) and the LQT-3 variant of the Long QT syndrome are caused by disease-linked SCN5A mutations that act to change functional and pharmacological properties of the channel. Here we have explored a set of SCN5A mutations linked both to BrS and LQT-3 to determine what diseasemodified channel properties underlie distinct responses to the Na channel blocker flecainide. We focused on flecainide block that develops with repetitive channel activity, so-called use-dependent block (UDB). Our results indicate that mutation-induced changes in the voltage-dependence of channel availability (inactivation) may act as determinants of flecainide block. The data further indicate that UDB by flecainide requires channel opening, but is not likely due to open channel block. Rather, flecainide appears to interact with inactivation states that follow depolarization-induced channel opening, and mutation-induced changes in channel inactivation will alter flecainide block independent of the disease to which the mutation is linked. Analysis of flecainide block of mutant channels linked to these rare disorders has provided novel insight into the molecular determinants of drug action. I N T R O D U C T I O N Local anesthetic molecules such as lidocaine and flecainide block Na channels and have been used therapeutically to manage cardiac arrhythmias (Rosen et al., 1975; Rosen and Wit, 1983; Wit and Rosen, 1983). Despite the prospective therapeutic value of the inherent voltageand use-dependent properties of channel block by these drugs in the treatment of tachyarrhythmias, their potential has been overshadowed by toxic side effects (Rosen and Wit, 1987; Weissenburger et al., 1991). Recently however, Na channel blockers have again proven useful for diagnosis and treatment of Brugada and LQT-3 syndromes, two inherited diseases linked to mutations in SCN5A, the gene that encodes the subunit of the cardiac voltage–gated Na channel (Brugada et al., 1999). Na channel blockade by the flecainide is of particular interest as it had been shown to reduce QT prolongation in carriers of some LQT-3 mutations (Brugada et al., 1999), and (Benhorin et al., 2000; Windle et al., 2001)and to evoke ST segment elevation, a hallmark of the Brugada syndrome (BrS),* in patients with a predisposition to the disease (Brugada et al., 2000). Thus, in the case of LQT-3, flecainide has potential therapeutic application, whereas for BrS it has proven useful as a diagnostic tool. However, in some cases, flecainide has been reported to provoke BrS symptoms (ST segment elevation) in patients carrying LQT-3 mutations (Priori et al., 2000). Furthermore, flecainide preferentially blocks some LQT-3 or BrS-linked mutant Na channels (Abriel et al., 2000; Grant et al., 2000; Nagatomo et al., 2000; Viswanathan et al., 2001). Investigation of the drug interaction with these and other LQT-3– and BrSlinked mutations may indicate the usefulness of flecainide in the detection and management of these disorders and in determining whether or not it is reasonable to use this drug to identify potential disease-specific mutations. Moreover, exploration of the relationship between altered channel structure and drug efficacy can provide new insight into molecular determinants of flecainide block of the human cardiac sodium channel. Local anesthetic compounds, such as lidocaine and flecainide, block Na channels in a useand voltagedependent manner. These actions have been widely interpreted within the framework of the modulated receptor hypothesis that ion channel states can alter drug affinity and that charged and neutral drugs interact with a common receptor but gain access to it via distinct hydrophobic and hydrophilic pathways (Hille, 1977a; Hondeghem and Katzung, 1977). Use-dependent block by flecainide, but not lidocaine, depends critically on channel openings (Ragsdale et al., 1994; Qu et al., Corresponding author: Robert S. Kass, Department of Pharmacology, College of Physicians and Surgeons of Columbia University, 630 W. 168th St. New York, NY 10032. Fax: 212-342-2703; E-mail: