Bladder cancer (BCa) is an aggressive malignancy with limited effective treatment options, and its poor outcomes largely result from delayed detection and therapeutic resistance. Triosephosphate isomerase 1 (TPI1) has been associated with tumor progression in various cancers, but its specific function in BCa remains poorly characterized. This study evaluated cancer-related markers and identified glycolysis as a key factor negatively impacting survival in BCa. Additionally, TPI1 was recognized as a potential prognostic marker, with its expression significantly elevated in BCa tissues compared to normal counterparts. Higher TPI1 levels were strongly linked to unfavorable clinical outcomes. Functional assays demonstrated that TPI1 overexpression significantly promoted BCa cell growth, migration, and invasive capabilities in vitro and in vivo. Mechanistically, TPI1 interacted with serine/threonine kinase B (AKT) and murine double minute 2 (MDM2) to form a protein complex, which enhanced the AKT-driven phosphorylation of MDM2 at serine 166 site, thereby promoting tumor protein p53 (p53) ubiquitination degradation. Furthermore, the truncated MDM2-F2 mutant (spanning 181-360) bound to TPI1, with amino acid 317 playing a critical role in this interaction. Notably, reducing AKT expression counteracted the p53 ubiquitination triggered by elevated TPI1.