GATOR2-dependent mTORC1 activity is a therapeutic vulnerability in FOXO1 fusion positive rhabdomyosarcoma.

Oncogenic FOXO1 gene fusions drive a subset of rhabdomyosarcoma (RMS) with poor survival and to date these cancer drivers are therapeutically intractable. To identify new therapies for this disease, we undertook an isogenic CRISPR-interference screen to define PAX3-FOXO1 specific genetic dependencies and identified genes in the GATOR2 complex. GATOR2 loss in RMS abrogated amino acid-induced lysosomal localization of mTORC1 and consequent downstream signaling, slowing G1-S cell cycle transition. In vivo suppression of GATOR2 impaired the growth of tumor xenografts and favored the outgrowth of cells lacking PAX3-FOXO1. Loss of a subset of GATOR2 members can be compensated by direct genetic activation of mTORC1. RAS mutations are also sufficient to decouple mTORC1 activation from GATOR2, and indeed fusion negative RMS harboring such mutations exhibit amino acid-independent mTORC1 activity. A bi-steric, mTORC1-selective small molecule induced tumor regressions in fusion positive patient-derived tumor xenografts. These findings highlight a vulnerability in FOXO1 fusion positive RMS and provide rationale for the clinical evaluation of bi-steric mTORC1 inhibitors, currently in phase 1 testing, to treat this disease. Isogenic genetic screens can thus identify potentially exploitable vulnerabilities in fusion driven pediatric cancers which otherwise remain mostly undruggable.