Abstract A026: Targeting PAX3-FOXO1 expression through translational and transcriptional control

Abstract FOXO1 gene fusions including PAX3-FOXO1 (P3F) and PAX7-FOXO1 (P7F) drive poor outcomes in pediatric rhabdomyosarcoma (RMS), yet direct targeting of these oncogenic drivers remains a challenge. We sought to define therapeutic vulnerabilities of FOXO1 fusion positive RMS by exploiting mechanisms of translational and transcriptional control. We previously identified the GATOR2 complex, a mediator of amino acid-dependent mTORC1 activation, as a selective dependency in fusion positive (FP) RMS. We now show that P3F and P7F require mTORC1-driven cap dependent translation for their own expression, explaining this genetic dependency. We used measures of cap-dependent translation including phosphorylation of 4-EBP1, polysome shift assays of mRNA, and pulse labeling of newly synthesized proteins after treatment with allosteric and bi-steric mTORC1 inhibitors. The effects of these agents, alone or in combination with BET/bromodomain inhibition, were evaluated in cell line and patient-derived xenograft (PDX) models. We found that the bi-steric mTORC1 inhibitor RMC-6272, but not the allosteric inhibitor rapamycin, suppressed p-4EBP1, eliminated P3F from polysomes, and decreased newly synthesized P3F protein in vitro for up to 3 days. In FP PDX models, RMC-6272 reduced P3F and P7F protein within 24 hours, though they rebounded by day 8 of treatment. One month of RMC-6272 treatment could induce remissions up to 8 weeks, though tumors that recurred demonstrated acquired resistance when reimplanted into secondary hosts. These tumors no longer showed reductions in P3F when re-challenged with RMC-6272, consistent with restoration of oncoprotein translation. Given the established sensitivity of P3F transcription to BET inhibitors, the BET inhibitor BMS-986158 was combined with RMC-6272. Combination therapy further reduced P3F protein levels compared to RMC-6272 alone, and prolonged suppression at day 8, suggesting more durable suppression of the fusion can be achieved by concurrently blocking transcription and translation. From the above, we conclude that mTORC1-driven cap-dependent translation is a critical vulnerability in FP RMS. Bi-steric mTORC1 inhibition reduces P3F protein in vitro and in vivo, but is limited by adaptive resistance. Combination therapy with BET inhibition sustains suppression of P3F, providing a rationale for dual targeting of transcription and translation in pre-clinical development of therapies for FP RMS. Ongoing studies aim to define the contribution of the PAX3 and PAX7 5′ untranslated region (UTR) in conferring cap dependence, and the preclinical activity of mTORC1/BET inhibitor combinations. Citation Format: Sinead Carse, Yolanda Pan, Rishika Mehta, Pushpendra K. Sahu, Matthew Watson, Amit J. Sabnis. Targeting PAX3-FOXO1 expression through translational and transcriptional control [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Fusion-Positive Cancer: From Discovery to Therapy; 2026 Jan 13-15; Philadelphia PA. Philadelphia (PA): AACR; Cancer Res 2026;86(1_Suppl):Abstract nr A026.