Abstract
α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid(AMPA)-type glutamate receptors (AMPARs) are the predominant excitatory neurotransmitter receptors in the brain, where they mediate synaptic transmission and plasticity. Excessive AMPAR activation leads to diseases such as epilepsy. AMPAR properties are modulated by auxiliary proteins and foremost by the transmembrane AMPAR regulatory proteins (TARPs). These distribute in unique expression patterns across the brain, rendering AMPAR/TARP complexes promising targets for region-specific therapeutic intervention. TARP γ8 is predominantly expressed in the forebrain and is enriched in the hippocampus, a region associated with temporal lobe epilepsy. Recent high-throughput medicinal chemistry screens have identified multiple promising compounds that selectively target AMPARs associated with γ8 and hold promise for epilepsy treatment. However, how these modulators target the receptor complex is currently unknown. Here, we use a combination of ligand docking, molecular dynamics simulations, and electrophysiology to address this question. We identify a conserved oxindole isostere, shared between three structurally diverse modulators (LY-3130481, JNJ-55511118, and JNJ-61432059) as the major module engaging γ8 by an H-bond to Asn-172 (γ8). The remaining variable region of each molecule likely targets the receptor complex in ligand-selective modes. Functional data reveal parallels in the underlying modulatory action of two prominent compounds. This work will aid development of refined AMPAR epilepsy therapeutics and facilitate to uncover the mechanisms by which TARPs modulate the receptor.
Highlights
transmembrane AMPAR regulatory protein (TARP) g8 predominates in the forebrain and is the major TARP in the hippocampus [21, 24], where AMPAtype glutamate receptors (AMPAR) are predominantly associated with g8 and another auxiliary subunit, cornichon-homologue 2 [25,26,27]
LY481 and JNJ-118 are the best characterized, blunting some of the positive modulatory action conferred by TARPs [10, 11], whereas JNJ-059 is a more recently reported compound, with substantially greater potency and of promise in rodent seizure models [13]
Because radiolabeled LY-481 derivatives can associate with free g8 in the absence of the AMPAR [36], we hypothesized that the oxindole isostere could directly engage with g8 to mediate the compounds’ effects
Summary
All three ligands share an oxindole isostere moiety (Fig. 2A, red substructure), which is critical for their potency, as determined by lead optimization campaigns [12, 13, 35] (see Fig. S1). The variable region (Fig. 2A, blue) differs substantially between ligands, and contrary to the oxindole isostere, alteration of this region never causes compound inactivation, suggesting that it is not a critical region for protein interaction [12, 13, 35] (Fig. S1). We considered the possibility that the oxindole isosteres exist as enol tautomers (in addition to the published keto isomers), which is apparent from 1H-NMR spectra of related structures in the Spectral Database for Organic Compounds (SDBS). Whereas the oxindole group of JNJ-059 favors the keto tautomer (SBDS 13584) (Fig. 2B, top row), the benzimidazolone of JNJ-118 could exist
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