This study aimed to investigate the effect of esculetin (Esc) on the viability and apoptosis of triple negative breast cancer (TNBC) cells and its mechanism. In this study, TNBC cell lines MDA-MB-231 and MDA-MB-468 were utilized as experimental models. The anti-proliferative effects of Esc on TNBC cells were evaluated through MTT assay, morphological observation, and colony formation assay. Transwell migration and invasion assays were employed to assess Esc's impact on TNBC cell motility and invasiveness. Hoechst 33258 staining and flow cytometry were performed to examine apoptosis induction. Western blot was used to quantify protein expression levels of Janus kinase 2 (JAK2)/Signal transducer and activator of transcription 3 (STAT3) pathway (JAK2, p-JAK2, STAT3, p-STAT3) and apoptosis-related proteins including cysteinyl aspartate specific proteinase 9 (Caspase-9), Caspase-3, poly ADP-ribosepolymerase (PARP), Cleaved Caspase-9, Cleaved Caspase-3 and Cleaved PARP. The results showed that compared with the control group (Con), Esc could significantly inhibit the proliferation and metastasis of TNBC cells (P<0.001). Western blot analysis revealed that, compared to Con, Esc significantly downregulated the protein expression of JAK2, p-JAK2, STAT3, and p-STAT3 (P<0.05), while upregulating the expression of Cleaved Caspase-9, Cleaved Caspase-3, and Cleaved PARP (P<0.05). Hoechst staining and flow cytometry demonstrated that Esc markedly induced apoptosis in TNBC cells compared to Con. Furthermore, co-treatment with Esc and the JAK2/STAT3 inhibitor WP1066 synergistically enhanced TNBC cell apoptosis (P<0.01). In summary, Esc likely induces apoptosis in TNBC cells by suppressing the JAK2/STAT3 signaling pathway, thereby activating the caspase cascade.