Novel anxiolytic-antidepressant combinations TF-1 and TF-2 and their intervention mechanisms in a mouse model of comorbid anxiety and depression (CAD).

Background and Objective: To address the high mortality, slow onset, and suboptimal efficacy of current treatments for comorbid anxiety and depression (CAD), this study developed and systematically evaluated a novel combination formulation. Methods: The anxiolytic tandospirone (TDS) was combined with fluoxetine (FLU), escitalopram, or milnacipran for cell-based screening. The optimal combination was selected for pharmacokinetic studies in rats, followed by a 49-day pharmacodynamic evaluation and mechanistic investigation in a CAD mouse model. Results: Two optimal TDS-FLU combinations, TF-1 (TDS:FLU = 1.2:1,w/w) and TF-2 (TDS:FLU = 2.4:1,w/w), were obtained, both exhibiting good neuroprotective effects. Pharmacokinetic results showed that TF-1 and TF-2 significantly increased drug exposure and prolonged half-life compared with monotherapies of TDS and FLU. Pharmacodynamically, TF-1 and TF-2 exhibited superior anxiolytic-antidepressant efficacy compared with monotherapies after 28-day and 49-day interventions. At day 28, both formulations significantly increased open-field central-zone activity and sucrose preference, and reduced immobility time (all P < 0.05). Their therapeutic benefits were further augmented at day 49, with sustained and improved behavioral improvements upon long-term treatment. Mechanistic studies showed that both combinations elevated plasma and hippocampal 5-HT and BDNF levels, upregulated hippocampal 5-HT1AR and TrKB expression, and enhanced neuronal plasticity. Conclusion: TF-1 and TF-2 were superior to single agents in pharmacokinetics and long-term CAD efficacy, acting via 5-HT modulation and BDNF-TrKB activation, supporting their development as CAD combination therapies.