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Theses Doctoral

Mechanisms of Mutation-Specific Inhibition of Late Na+ Current in Long QT Syndrome Type 3

Robey, Seth Hamilton

The mechanical contraction of the heart is tightly coupled to rapid and concerted electrical excitation of the cardiac muscle. This electrical activity is facilitated by a highly synchronized conduction system consisting of channels, pumps, and transporters that facilitate the flow of charged ions between cellular compartments, the cytoplasm, and the interstitial fluid between cells. The biophysical properties of these membrane proteins have been studied for many years, but their role in the generation of potentially lethal cardiac arrhythmias and their interactions with drugs remains an important field of research. The cardiac isoform of the voltage-gated Na+-channel, Nav1.5, has garnered widespread interest because of its role in the generation of electrical impulses in the cardiac myocyte, its association with congenital conduction disorders and acquired cardiac arrhythmias, and its unique pharmacological properties.
The Congenital Long QT Syndrome Type 3 (LQT3) arises from heritable mutations in SCN5A - the gene encoding Nav1.5 - that disrupt the inactivation process responsible for imparting a refractory period and that often cause a sustained depolarizing late current (INaL). The gain of function depolarizing currents arising from LQT3 mutant channels cause a prolongation of the ventricular action potential and leave patients susceptible to asynchronous electrical activity, ventricular arrhythmias, and sudden cardiac death. The disruption of channel inactivation can arise through a wide range of modalities, including changes in inactivation voltage-dependence and kinetics, and has been shown to occur with varying degrees of severity. Because of this range of phenotypes there is heterogeneity in the risk factors for arrhythmia and sudden cardiac death and
in the utility of Na+-channel blocking antiarrhythmic drugs. Moreover, INaL has been implicated as a proarrhythmic and potentiating factor in several acquired cardiac ailments including heart failure, ischemia, and hypertrophy. There is therefore a large unmet need for improved understanding of INaL and mechanisms of its selective inhibition, and LQT3 mutant channels provide a reliable experimental model for this class of cardiac arrhythmias.
This study will employ a combination of electrophysiological and computational methods to unravel mechanisms by which mutant Nav1.5 produces pro-arrhythmic currents and the interactions of different disease-causing mutant channels with a set of clinically relevant antiarrhythmic drugs. Chapter 1 of this study presents a functional characterization of one LQT3 mutation, F1473C, that was discovered in a patient with severe QT prolongation, frequent ventricular arrhythmias, and a poor response to pharmacological intervention. This mutation gives rise to INaL by a mechanism that is functionally distinct from the mechanism discovered previously in the canonical LQT3 mutation, ΔKPQ (1505-1507del), and causes a unique response to channel inhibitors. In order to better understand the mechanisms of this divergent pharmacology, Chapter 2 presents the development of a series of computational models which explore the gating dysfunctions that cause INaL and how these pathological changes can influence the predicted safety and efficacy of pharmacological intervention. These models predict that the majority of mutation- specific drug effects can be attributed to differential mutant channel gating, but raise the possibility that mutations may directly alter the physical chemical interaction between drugs and channels. Finally, Chapter 3 presents an attempt to explore this possibility using an innovative chemical biology technique - the site-specific incorporation of unnatural amino acids - that allows for the measurement of precise chemical interactions hypothesized to vary in a mutation-dependent manner. The findings presented in this work promote the need for patient-specific screening of
antiarrhythmic agents and lay the groundwork for the use of in silico systems analysis of cardiovascular pharmacology.


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More About This Work

Academic Units
Pharmacology and Molecular Signaling
Thesis Advisors
Kass, Robert S.
Ph.D., Columbia University
Published Here
January 20, 2017