Repurposing FDA-approved allosteric drugs as non-competitive inhibitors of human UGTs: An integrated computational study and biochemical validation

Abstract

UDP-glucuronosyltransferases (UGTs) catalyze a major detoxification route for xenobiotics, yet chemical tools that modulate their activity are virtually absent and the possibility of allosteric regulation is largely unexplored. We combined ligand-based target prediction, structure-based docking, long-timescale molecular dynamics and MM/PBSA free-energy calculations to search the pharmacopeia for hidden UGT modulators. Six FDA-approved allosteric drugs were screened against the catalytic domains of UGT1A1, 1A9, 2B7 and 2B15. Docking and 200-ns simulations revealed a previously unrecognized cryptic pocket ∼10 Å from the catalytic histidine that stably accommodated all six ligands; ivacaftor and cinacalcet produced the deepest free-energy basins and the most favorable MM/PBSA binding energies (−35.5 and − 31.2 kcal mol⁻¹, respectively). FEL analysis showed that ligand binding funnels the enzyme's conformational ensemble into a single deep basin, providing a dynamic explanation for the V_max depression observed in vitro. In recombinant enzymes and pooled human-liver microsomes these two compounds inhibited glucuronidation with low-micromolar potency (IC₅₀ ≈ 3–4 μM) while the remaining drugs were an order of magnitude weaker, mirroring the computational ranking. Lineweaver–Burk and Michaelis–Menten analyses showed pure non-competitive behavior, confirming that pocket occupancy depresses V_max without altering K_m. The strong correlation between calculated ΔG_bind and experimental pIC₅₀ (ρ = 0.89) validates the predictive pipeline. Pharmacokinetic modelling suggests that clinically relevant concentrations of ivacaftor and cinacalcet may modulate UGT1A9- and UGT1A1-mediated clearance, revealing a previously unrecognized drug–drug-interaction risk. Altogether, the work delivers the small-molecule allosteric probes for UGTs, establishes a general workflow for mining existing drugs as phase-II enzyme modulators, and provides a structural framework for developing isoform-selective UGT therapeutics.