I. Cyclodextrin-based receptors for peptide recognition. II. Molecular determinants of flecainide action on the human cardiac sodium channel

Chapter 1. Cyclodextrin-based receptors for peptide recognition . A synthetic approach to cyclodextrin trimers based on semi-rigid scaffolds is described. The synthetic challenges involved in the preparation of these receptors are discussed. The binding properties of the receptors with different L-tryptophan and t-butyl phenylalanine based substrates are explored. Trimeric binding was demonstrated to be stronger than dimeric binding. The thermodynamics of complexation were explored. Cyclodextrin dimers were investigated for their ability to bind natural peptide hormones. A detailed study of the binding of a secondary face, ester linked dimer to α-melanocyte hormone was undertaken. Finally, versatile synthetic routes to cyclodextrin-peptide hybrids are presented. Both stepwise solid phase synthesis and post-synthetic covalent modification methods are discussed. The binding properties of cyclodextrin-peptide hybrids containing two and three β-cyclodextrins are described.; Chapter 2. Molecular determinants of flecainide action on the human cardiac sodium channel. A novel neutral analog of the antiarrhythmic drug flecainide was designed and synthesized. A quaternary analog of flecainide, previously reported but not fully characterized, was synthesized. The effects of these drugs in wild type and mutant human cardiac sodium channels were investigated and compared with flecainide, lidocaine, and quaternary lidocaine (QX-314). The neutral analog was found to shift the steady-state inactivation curve without conditioning pulses and shown to have a high affinity for the inactivated state.{09}The binding to the inactivated state by the neutral analog and lidocaine was disrupted by F1760A and Y1767A mutations. This represents the first direct evidence of flecainide binding to the inactivated state of the channel.; Blocking properties of quaternary flecainide were shown to be similar to quaternary lidocaine. Both drugs exert strong use-dependent block when applied internally but weak block when applied externally. This suggests an intracellular access route for flecainide to its binding site. The quaternary compound, like flecainide, but unlike the neutral analog, does not shift the inactivation curve without conditioning pulses.; These data provide further understanding of the molecular mechanism of flecainide action in cardiac sodium channels and demonstrate the utility of a new molecular tool for the investigation of flecainide action in mutant sodium channels associated with cardiac disease.