In silico assessment of neuromuscular blocking agents and fluoroquinolones as ligands of the Mas-related G protein-coupled receptor X2.

BACKGROUND: Neuromuscular blocking agents (NMBAs) may trigger severe perioperative hypersensitivity reactions. A recently proposed mechanism involves off-target activity of NMBAs. They are thought to directly activate the Mas-related G protein-coupled receptor X2 (MRGPRX2), leading to mast cell degranulation and drug-induced hypersensitivity reactions. This study investigated which NMBAs exert this effect, whether in silico findings are consistent with cell-based experiments, and how the affinity of NMBAs for MRGPRX2 compares with that of fluoroquinolones, known MRGPRX2 agonists. We also sought to predict MRGPRX2 binding-site mutations that might enhance interactions with these drugs. METHODS: Molecular docking, molecular dynamics (MD) simulations, and molecular mechanics Poisson-Boltzmann surface area binding free-energy calculations were combined with a β-hexosaminidase release assay in MRGPRX2-expressing RBL-2H3 stable cell line. Computational alanine scanning was performed to predict the impact of receptor mutations. RESULTS: We demonstrated that atracurium-induced mast cell degranulation can be attributed to its immediate metabolite, laudanosine, which binds strongly to MRGPRX2. Pipecuronium, but not rocuronium, suxamethonium, or vecuronium, exhibited high affinity for MRGPRX2 in MD simulations. Complexes of ciprofloxacin, levofloxacin, and moxifloxacin with MRGPRX2 demonstrated excellent stability in MD simulations. RBL-MX2 responses to NMBAs and fluoroquinolones were highly consistent with our MD findings. Alanine substitutions reduced the affinity of ligands for MRGPRX2. CONCLUSIONS: In contrast to fluoroquinolones, NMBAs displayed different affinity for MRGPRX2. Cellular responses in bench experiments closely reflected the MD predictions. Alanine scanning revealed that the MRGPRX2 binding pocket exhibits low susceptibility to single-site mutations that enhance receptor responsiveness.