Background: Ewing sarcoma (EWS) is an aggressive malignancy of childhood with a dismal prognosis in patients with metastatic or recurrent disease. Previous research demonstrated EWS cell lines with the EWS-FLI1 translocation be sensitive to Poly (ADP ribose) polymerase inhibitors (PARPi). Although phase 1 clinical trials have failed to demonstrate significant single agent efficacy for EWS, preclinical studies suggest increased efficacy with the addition of DNA damaging agents to PARPi. We examined the PARPi BMN 673, olaparib, and veliparib in combination with irinotecan and temozolomide (TMZ) in a clinically relevant EWS orthotopic xenograft model. Methods: EWS orthotopic xenografts were created by injecting EW-8 luciferase labeled cells into the femur of CD-1 nude mice. Mice were randomized to the following treatment groups (see Table.) Allometric doses were administered on a clinically relevant schedule for two cycles. Complete response (CR), stable disease (SD), and progressive disease (PD) were determined using bioluminescence. Results: The addition of TMZ and irinotecan to BMN 673 and olaparib (Groups 1, 2) showed significant disease response in comparison to irinotecan and TMZ alone (Group 4). Mice that received BMN 673 had the best response with 100% achieving CR. Mice that received olaparib (Group 2) had a 50% CR and 50% SD. Groups 3, 4, and 5 all had 80% PD and 20% SD. Conclusions: The addition of the PARP inhibitors BMN 673 and olaparib to irinotecan and TMZ provides significant disease response and stabilization in an orthotopic xenograft model of EWS. These drug combinations may be effective in the treatment of pediatric patients with EWS. Ongoing pharmacokinetic and preclinical studies will be performed to determine if efficacy is likely to be achieved within the therapeutic window of this patient population.