Imipramine Inhibits Osteosarcoma Invasion via Src Inactivation and Caspase‐Dependent Apoptosis

Osteosarcoma (OS) is an aggressive malignancy characterised by high metastatic potential and poor prognosis. Imipramine, a tricyclic antidepressant, has shown potential anticancer effects. This study evaluates the cytotoxic, pro‐apoptotic and anti‐invasion effects of imipramine on OS cells in vitro and in vivo, as well as its underlying mechanisms. Imipramine significantly reduced U‐2 OS and MG 63 cell viability in a time‐ and dose‐dependent manner, confirmed through MTT and colony formation assays. It induced apoptosis via caspase‐dependent pathways, as evidenced by increased cleaved caspase‐3, ‐8 and ‐9 levels and reduced expression of anti‐apoptotic proteins such as MCL‐1. Imipramine activated both extrinsic and intrinsic apoptosis pathways in vitro and in vivo, increasing Fas, Fas‐L, BAX and BAK while suppressing anti‐apoptotic factors like BCL‐2 and XIAP. Transwell assays showed dose‐dependent inhibition of cell migration and invasion, supported by suppressed Src phosphorylation and downregulation of EMT markers (Snail‐1 and Slug). In U‐2 OS xenograft‐bearing mice, imipramine significantly inhibited tumour growth in a dose‐dependent manner, with the 30 mg/kg group showing the smallest tumour volume and slowest growth rate ( p = 0.0098). Tumour weights were significantly reduced without impacting body weight or liver and kidney function markers (AST, ALT, γ‐GT and CREA). Histopathological analyses revealed no significant abnormalities in vital organs. Imipramine exerts potent anti‐OS effects by suppressing Src‐mediated invasion and enhancing caspase‐dependent apoptosis through extrinsic and intrinsic pathways. It inhibits tumour progression without inducing systemic toxicity, demonstrating its potential as a therapeutic candidate for OS.