MDM2 Inhibition in a Subset of Sarcoma Cell Lines Increases Susceptibility to Radiation Therapy by Inducing Senescence in the Polyploid Cells

PURPOSE: Inhibition of p53 by amplification of MDM2 is one of the key contributors in the oncogenesis of a subset of soft tissue sarcoma (STS) known as well-/dedifferentiated liposarcoma. A small molecule MDM2 antagonist, nutlin-3, induces the p53 pathway by disrupting the interaction between MDM2 and p53. Radiation therapy is an integral component for treating liposarcoma and induces p53. Based on wild-type TP53 status in liposarcoma, it was investigated whether MDM2 inhibition with irradiation led to enhanced reactivation of p53 and subsequent p53-mediated effects in liposarcoma. METHODS AND MATERIALS: Clonogenic assays, immunoblotting, flow cytometry/fluorescence-activated cell sorting, and senescence assays were employed in liposarcoma cell lines after co-treatment with nutlin-3 and radiation. RESULTS: Upon treatment with nutlin-3, 2 of the well-/dedifferentiated liposarcoma (MDM2(Amp)/TP53(WT)) cell lines displayed radiosensitivity with sensitization enhancement ratio values of >1. In contrast, the cell line with mutant TP53 showed sensitization enhancement ratio values of ∼1. Immunoblotting revealed induced reactivation of the p53-MDM2-p21 signaling axis in response to combination therapy in all cell lines with wild-type TP53. Removal of MDM2 inhibitor (with or without radiation therapy) led to the emergence of ploidy-based heterogeneous subpopulations (4N and >4N) in TP53 wild-type cells and not in TP53 mutant cells. Immunoblotting of cell cycle markers (G1, G2/M) revealed the generation of 4N G1 cells. Sorting and long-term fate analysis of different populations (2N, 4N, and >4N) by colony assay displayed attenuated colony-forming potential and augmented senescence of the 4N and >4N cells contributing to the radiosensitization effect. CONCLUSIONS: Nutlin-3 increases the vulnerability of liposarcoma cell lines to radiation by augmented activation of p53. The cells underwent senescence. Presence and activation of p53 are required for exertion of the radiosensitizing effect by nutlin-3, but this is not the sole determinant of the effect. This study opens avenues for the clinical translation in a stratified group of patients with liposarcoma.