Katelyn R. Ludwig, Victor Collins, Natasha J. Caplen, Christine Heske
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Submitter: Katelyn Ludwig (Postdoc or fellow)
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email: katelyn.ludwig@nih.gov
Ewing sarcoma (EWS) is an aggressive bone and soft tissue malignancy predominantly affecting children and adolescents in the second decade of life. The current first-line chemotherapeutic regimen for treatment of EWS involves cyclical administration of vincristine, doxorubicin, and cyclophosphamide (VDC) alternating with ifosfamide and etoposide (IE). Although outcomes in patients who present with localized disease have improved, long-term outcomes in patients with metastatic, relapsed, or refractory EWS are dismal, with survival rates of less than 20%. In this study, we have investigated rational drug combinations of the newest generation of DNA-PK inhibitors (M3814 or AZD-7648) and DNA damaging agents relevant to the treatment of EWS. Rational combinations of drugs were assessed for their effect on the growth of a panel of EWS cell lines, including TC32, TC71, SK-N-MC, RDES, TC106, EW8, and CHLA-258. We analyzed the effect of drug treatment (single agent and in combination) on cell growth using IncuCyte-based imaging and quantitation of cell proliferation and a metabolic readout of cell viability. We assessed markers of apoptosis and DNA damage using western blotting, flow cytometry, and micronuclei counts. We further assessed non-Ewing sarcoma cell lines to demonstrate the specific efficacy of this combination. Results of in vivo studies are ongoing and will be reported. We have demonstrated the synergy of M3814 or AZD-7648 with multiple chemotherapeutic agents that act as a TOP2 poisons, including the two clinically relevant agents, etoposide and doxorubicin. Furthermore, we observed that the combination of M3814 or AZD-7648 and etoposide activates markers of DNA damage and apoptosis. This study has identified a rational combination for EWS that may allow for lower doses of standard chemotherapeutic agents to be used while retaining or enhancing clinical efficacy.