This study aims to investigate the antitumor effects of saikosaponin A (SSA) on acute myeloid leukemia (AML) and elucidate its underlying mechanisms, particularly focusing on the endoplasmic reticulum stress (ERS)-mediated MAPK-p-JNK signaling pathway.The inhibitory effects of SSA on the proliferation of AML cell lines K562 and HL60 were evaluated using CCK8 and EdU assays. Apoptotic effects induced by SSA were analyzed via flow cytometry. RNA sequencing was performed to identify differentially expressed genes and enriched signaling pathways. Western blot analysis was utilized to confirm the involvement of ERS and activation of the MAPK-p-JNK signaling pathway. Further validation of the potential mechanism of SSA-induced apoptosis was conducted using SP600125 and 4PBA. The in vivo anti-AML efficacy of SSA was assessed using a xenograft model.SSA exhibited significant inhibitory effects on the proliferation of AML cell lines K562 and HL60, with IC50 values at 12, 24, and 48 hours demonstrating time- and dose-dependency (19.84 μM, 17.86 μM, and 15.38 μM for K562; 22.73 μM, 17.02 μM, and 15.25 μM for HL60, respectively). Western blot analysis demonstrated that SSA induces apoptosis in AML cells through the mitochondrial apoptotic pathway. Transcriptomic profiling and Western blot analyses confirmed that SSA activates the ERS-mediated p-JNK signaling pathway to induce apoptosis in AML, a process that can be reversed by the addition of 4PBA or SP600125. Furthermore, SSA significantly reduced tumor volume and weight in a NOD-SCID mouse xenograft model without causing notable toxicity to the liver, kidneys, lungs, or heart, while also activating the ERS and p-JNK signaling pathways in vivo.SSA induces apoptosis in AML cells by activating the ERS-mediated p-JNK signaling pathway, exhibiting significant anti-AML effects both in vitro and in vivo, accompanied by a favorable safety profile.