Identification of inhibitory potential of acamprosate, roxindole and L-ascorbic acid against tryptophan 2, 3 dioxygenase using experimental and computational approaches

Background: Tryptophan 2,3-dioxygenase (TDO) is a haem-containing enzyme of the kynurenine pathway, which is imperative for tryptophan metabolism. Primarily found in the liver, TDO facilitates the breakdown of TRP into N-formyl kynurenine. TDO is regarded as a promising target for antidepressant therapies. As an enzyme responsible for tryptophan degradation, its inhibition may enhance serotonin availability, which plays a key role in mood regulation. Objectives: This study identifies TDO as a potential target for central nervous system drugs (acamprosate, roxindole, and L-ascorbic acid) that may alter brain activity and potentially impact mood and behavior. Methods: The study involved male Albino Wistar rats, each weighing between 150 and 200 grams. The rats were decapitated, the livers were promptly excised within 10 seconds, and perfused in situ with ice-cold saline. The perfused livers were then immediately frozen at -80?°C for subsequent analysis. In vitro TDO enzyme activity was assessed in frozen liver homogenates. Enzymatic activity was measured for both the holoenzyme and total enzyme spectrophotometrically. Molecular docking of the selected compounds with TDO was conducted using AutoDock Vina. The crystal structure of TDO was retrieved from the Protein Data Bank (PDB), while ligand structures were obtained from PubChem. Results: In vitro experiments revealed that these drugs inhibited apoenzyme activity by 68-85%, while total enzyme activity was reduced by 34%, 38% and 37% for acamprosate, roxindole, and L-ascorbic acid, respectively. Further validation through molecular docking analysis confirmed their strong binding affinity to the TDO active site, with l-ascorbic acid showing the highest binding energy (-7.2 kcal/mol), followed by acamprosate (-6.7 kcal/mol) and roxindole (-6.4 kcal/mol). Conclusion: These findings suggest that acamprosate, roxindole, and L-ascorbic acid act as competitive TDO inhibitors, potentially enhancing serotonin synthesis and mitigating depressive symptoms.