Protein Kinase Inhibitor Alpha Drives Vincristine Resistance in Ewing Sarcoma via cAMP-EPAC Signaling Reprogramming.

Ewing sarcoma (ES) is an aggressive bone malignancy with poor outcomes for chemotherapy-resistant patients, yet the mechanisms underlying vincristine resistance remain unclear. Here, we identify protein kinase inhibitor alpha (PKIA) as a critical driver of chemoresistance through cAMP-EPAC signaling reprogramming. Transcriptomic analysis of vincristine-resistant ES cells revealed PKIA upregulation, which correlated with poor survival in clinical cohorts (HR = 2.14, p < 0.001). Mechanistically, PKIA overexpression elevated intracellular cAMP levels but suppressed PKA activity, instead activating the noncanonical EPAC-Rap1-ERK pathway to promote drug efflux and survival. Pharmacological inhibition of EPAC with ESI-09 reversed resistance (IC~50~ reduction: 52%, p < 0.01), while PKIA knockdown restored vincristine sensitivity in xenografts. Strikingly, PKIA exhibited a dual role, with low expression in primary ES (potentially tumor-suppressive) and high expression in resistant/metastatic tumors (prosurvival), mirroring observations in prostate and hepatocellular cancers. Our work establishes PKIA as a therapeutic vulnerability and supports targeting the cAMP-EPAC axis to overcome chemoresistance in high-risk ES.