Synergistic effects of combining PARP inhibitor (AZD2281) and ATR inhibitor (AZD6738) in Ewing Sarcoma cell lines

Ewing Sarcoma (ES) is an aggressive pediatric solid tumor. Even though overall-survival for localized patients is approximately 70%, the overall-survival for high risk ES patients has not improved in the last 20 years. Therefore, there is a need for exploration of new therapeutic agents in ES. Recent evidence has demonstrated that ES cells behave like BRCA-deficient tumor types which renders them sensitive to PARP inhibitors in vitro and in vivo. However, a phase II study of the efficacy of single-agent PARP inhibition in patients with relapsed ES did not significantly improve outcome. As single-agent therapy is rarely expected to result in significant clinical responses, in this study, we plan to validate potential targeted combination therapies with PARP inhibitors in ES. Since ES appears to demonstrated BRCA-deficient biology with impaired homologous recombination, cells are expected to be sensitive to both PARP inhibitors and ATR inhibitors, drugs which have a role in regulating DNA damage and impairing homologous recombination. In breast cancer and ovarian cell lines with genetic BRCA-deficiency, PARP and ATR inhibitors have synergistic activity. We hypothesize that these inhibitors will also have synergistic anti-Ewing activity. Furthermore, we recognize that ES cells demonstrate remarkably quiet genomes suggesting that there is minimal ongoing DNA-damage when cells are growing unperturbed. Therefore, we also plan to test the effect of adding low-dose genotoxic chemotherapy to induce additional sensitivity to the combination of PARP and ATR inhibitors in ES. The specific aims of this study were to explore the possible anti-tumor effect of PARP inhibitors combined with ATR inhibitors in ES cell lines, and to explore whether low dose genotoxic chemotherapy with SN38 can potentiate the anti-tumor effect of combined PARP and ATR inhibition in ES cell lines. We studied the anti-Ewing Sarcoma effect of the combination of a PARP inhibitor, AZD2281, and an ATR inhibitor, AZD6738, across a range of doses with and without low doses of a DNA damaging agent, SN38 (irinotecan metabolite), in two ES cell lines. We analyzed synergy by determining the Combination Index (CI) and Fractional Inhibition (FA) of each combination. We found that the ATR inhibitor, AZD6738, was synergistic across large range of concentrations when combined with the PARP inhibitor, AZD2281, in ES cell lines. We also found that treatment of cells with low doses of SN38 increases ES cell sensitivity to treatment with the PARP inhibitor and ATR inhibitor combination. This study provides preclinical support for additional studies exploring these combinations in ES. Given the low number of pediatric patients with ES compared to adult cancer patients, there will be limited attempts in combining these agents in clinical trials. Therefore, the development of an in vivo trial testing the safety and efficacy of this combination in ES mouse models is proposed.2020-07-03T00:00:00