Abstract A016: Overcoming Promiscuous Tumor Formation: A Controlled Genetically Engineered Mouse Model of CIC::DUX4 Sarcoma

Abstract Objective: CIC::DUX4 sarcoma (CDS) is a rare and aggressive sub-type of sarcoma with no effective treatment options currently available. A significant barrier to identifying alternative therapeutic approaches is the lack of robust pre-clinical models. We previously generated 3 genetically engineered mouse models of CDS, however, all 3 models spontaneously formed tumors in the absence of Cre-recombinase. Accordingly, we developed a novel dural-recombinase FLEx-switch model (dFLEx CDS). In this study we show the dFLEx CDS model is leak-proof, spatially and temporally controllable, and recapitulates human CDS. Methods: The dFLEx CDS allele features a stop cassette proceeded by an inverted exon 1, and exon 2. Cre-mediated recombination inverts the first exon into frame and removes the upstream stop cassette while FLP-mediated recombination inverts exon 2 into frame. Both Cre and FLP-mediated recombination are required for CIC::DUX4 expression. To induce recombination, two approaches were used: (1) injection of Adeno-Cre-2A-FLP and (2) electroporation of pCAG-Cre and pCAG-FLPE plasmids. Additionally, dFLEx CDS mice were crossed with R26-FlpO deleter or R26-Cre mice to remove 1 of the 2 FLEx switches (sFLEx CDS). Results: Injection of AdCre-2A-FLP resulted in tumors within 60 days with 100% penetrance; however, tumor formation was not restricted to the injected hindlimb. Conversely, electroporation of separate pCAG-Cre and pCAG-FLPE plasmids resulted in spatially and temporally restricted tumors that express the CIC::DUX4 fusion and known CIC::DUX4 target genes. Interestingly, sFLEx CDS mice were less viable and suffered from spontaneous tumor formation, similar to our previous CDS GEMMs. Conclusion: dFLEx CDS is the first temporally and spatially controllable mouse model that recapitulates human CDS tumors. sFLEx CDS mice develop spontaneous tumors, further highlighting the potency of the CIC::DUX4 oncogene and need for multiple layers of recombination protection. The use of two FLEx systems minimizes unintended tumor development, providing a reliable platform for mechanistic insight and robust preclinical drug testing for CDS. Citation Format: MaKenna R. Browne, Peter G. Hendrickson, David G. Kirsch. Overcoming Promiscuous Tumor Formation: A Controlled Genetically Engineered Mouse Model of CIC::DUX4 Sarcoma [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 A016.