Structure-Based Discovery of Novel Glycinergic Modulators

Abstract

Marijuana's analgesic effects can be attributed to the allosteric modulation of glycine receptors (GlyRs) by Δ9-tetrahydrocannabinol (THC). GlyRs are pentameric ligand-gated chloride channels that control inhibitory neurotransmission in the brainstem and spinal cord. The glycinergic mechanism of cannabis-induced analgesia is independent of the other psychoactive effects of THC. Compounds specifically targeting the well-defined THC-binding site in GlyRs are likely to provide pain relief with fewer unwanted side effects. Here, we screened over 2 million drug-like molecules from the ZINC database on an ensemble of α3GlyR structures obtained from molecular dynamics simulations of the closed-state α3GlyR crystal structure (PDB ID: 5CFB) and a homology model derived from the open-state α1GlyR NMR structure (PDB ID: 2M6I). Computational dockings were specifically targeted to the known THC-binding site in the α3GlyR transmembrane domain. Each screened compound was ranked based on its total predicted binding affinity across the ensemble of α3GlyR structures. Top ranked compounds were selected for functional measurements in Xenopus laevis oocytes expressing human α3GlyR. Several lead candidates have been identified as strong modulators of α3GlyR, exhibiting positive and/or negative allosteric effects at micromolar concentrations. Further molecular dynamics simulations of α3GlyR in the presence of modulators revealed that in the closed-state α3GlyR, potentiating compounds showed an increased probability of contacting with residue S296 compared to inhibitors occupying the same THC-binding site. S296 on the third transmembrane helix has been previously shown to be critical for THC potentiation of GlyRs. In contrast, inhibitors showed a higher probability of close contact with residue S241 in the first transmembrane helix. Interestingly, the adjacent residue I240 has been previously reported as an inhibitory site. This study suggests that different interaction residue partners, even within the same binding site, may lead to distinctly different allosteric modulations in α3GlyR. Research supported by NIH grants.