Promyelocytic leukemia protein (PML) modulates the p53 tumor suppressor through its interaction with p53 and MDM2. We found that activated big MAP kinase 1 (BMK1) preferentially associates with PML isoform IV and disrupts PML-MDM2 interaction. Doxorubicin, a common chemotherapeutic agent, is known to promote PML-mediated p53 activation in part by promoting PML-dependent MDM2 nucleolar sequestration. We discovered that BMK1 deactivation coupled with doxorubicin synergistically enhanced MDM2 nucleolar sequestration and, consequently, promoted PML-mediated p53 upregulation leading to tumor cell apoptosis in vitro and tumor regression in vivo. Collectively, these results not only suggest that BMK1 activity has a role in suppressing p53 by blocking the interaction between PML and MDM2, but also implicate that pharmacological BMK1 inhibitor should significantly enhance the anticancer capacity of doxorubicin-based chemotherapy.