FET fusion oncoproteins disrupt physiologic DNA repair networks and induce ATR synthetic lethality in cancer.

The genetic principle of synthetic lethality is clinically validated in cancers with loss of specific DNA damage response (DDR) pathway genes (i.e. BRCA1/2 tumor suppressor mutations). The broader question of whether and how oncogenes create tumor-specific vulnerabilities within DDR networks remains unanswered. Native FET protein family members are among the earliest proteins recruited to DNA double-strand breaks (DSBs) during the DDR, though the function of both native FET proteins and FET fusion oncoproteins in DSB repair remains poorly defined. Here we focus on Ewing sarcoma (ES), an EWS-FLI1 fusion oncoprotein-driven pediatric bone tumor, as a model for FET rearranged cancers. We discover that the EWS-FLI1 fusion oncoprotein is recruited to DNA DSBs and interferes with native EWS function in activating the DNA damage sensor ATM. Using preclinical mechanistic approaches and clinical datasets, we establish functional ATM deficiency as a principal DNA repair defect in ES and the compensatory ATR signaling axis as a collateral dependency and therapeutic target in FET rearranged cancers. Thus, aberrant recruitment of a fusion oncoprotein to sites of DNA damage can disrupt normal DSB repair, revealing a mechanism for how oncogenes can create cancer-specific synthetic lethality within DDR networks.