A chemical biology approach to discover the biological targets of the antiepileptic drug lacosamide.

摘要

Lacosamide (VimpatRTM) is a potent antiepileptic drug that received market approval for the adjunctive treatment of partial-onset seizures in adults in Europe and the United States. The pharmacological studies document that lacosamide has a unique profile of activity that differentiates it from existing antiepileptic agents. Based on these findings, we hypothesized that lacosamide binds to different proteins, with low-to-modest affinity. This research project aims to identify the lacosamide biological targets that modulate function and toxicity. We propose a novel target identification approach where an affinity bait (AB) and a chemical reporter (CR) group are strategically placed within the lacosamide framework. These compounds are termed lacosamide AB&CR agents. The AB moiety leads to permanent capture of the binding protein while the bioorthogonal CR unit is used for either detection or isolation of the complex upon reaction with a probe. The understanding of lacosamide's mechanism(s) of action will help increase our understanding of epileptic disorders, and permit the rational development of new clinical agents.;In the first part of our study, we explored the structure-activity relationship (SAR) for the 3-oxy site in lacosamide. We showed that incorporation of non-bulky, hydrophobic groups at this site provided lacosamide derivatives with excellent activities in animal seizure models. This information was used to design and stereospecifically synthesize a series of lacosamide AB&CR agents where either the AB or the CR group was installed at the 3-oxy site. Most lacosamide AB&CR agents were evaluated for anticonvulsant activity in animal models.;In the second part of our study, the lacosamide AB&CR agents were utilized to interrogate the rat brain soluble and membrane-bound proteome for potential binding partners of lacosamide. We used several subcellular fractionation methods to deconvolute the rat brain proteome. Within each subcellular fraction, different protein purification methods were employed to partition the lysate and aid the identification process. Several potential proteins were selectively targeted by the lacosamide AB&CR agents. Further analysis did not confirm that these proteins were directly linked to lacosamide function. These studies documented the strengths and limitations of the AB&CR strategy for receptor identification and are discussed.