Abstract A16: Fusion-negative rhabdomyosarcoma originating from endothelial progenitors

Rhabdomyosarcoma (RMS) is the most common soft-tissue sarcoma in childhood, and despite rigorous clinical trials the survival for children with high-risk RMS has not changed for three decades. RMS is subdivided into two major classes, fusion-positive (FP) and fusion-negative RMS (FN-RMS), based on the presence or absence of the PAX3-FOXO1 or PAX7-FOXO1 gene fusions. RMS occurs at locations throughout the body with nearly 40% of tumors occurring in the head and neck. Tumor location and fusion status are key prognostic factors. RMS resembles developing skeletal muscle and has been speculated to originate from genetically compromised skeletal muscle progenitors. However, the genes that control RMS development and specify location remain elusive. RMS also occurs in tissues devoid of skeletal muscle such as the urinary bladder, prostate, and biliary tree, suggesting the possibility of origins outside of the skeletal muscle lineage. Currently, the cell of origin and the factors that specify RMS location and thus prognosis are unknown. Previously, we showed that activation of Sonic Hedgehog signaling through expression of a conditional, constitutively active Smoothened allele, SmoM2, under control of an adipocyte-restricted adipose protein 2 (aP2)-Cre recombinase transgene in mice gives rise to aggressive skeletal muscle tumors that display the histologic and molecular characteristics of human FN-RMS. In this model, tumorigenesis occurs with high penetrance (~80%), is early onset (by 2 months of age), and is anatomically restricted to the head and neck. Also, unlike previous RMS models, this model requires no additional background mutations, such as inactivation of p53, and drives only FN-RMS neoplasia. We illustrated that the transcriptome of the aP2-Cre;SmoM2 tumors recapitulates both other mouse FN-RMS models as well as human FN-RMS. With the short latency and anatomic restricted tumor location, we sought to leverage this model to explore the cell of origin of FN-RMS. Here we use genetic fate mapping with fluorescent reporter mice to interrogate the cell of origin of FN-RMS in the aP2-Cre;SmoM2 model. Tracing the aP2-Cre labeled cells with reporter mice illustrated labeled cells in both brown and white adipose tissue as well as a discrete population of cells lying between skeletal muscle fibers but not beneath the laminin sheath. These aP2-Cre labeled cells are distinct from Pax7-positive skeletal muscle stem cells or satellite cells and do not contribute to myotube formation in vitro or in vivo. Gene profiling of tomato positive cells isolated by FACS from the sternocleidomastoid (SCM) of aP2-Cre;R26-Tom and aP2-Cre;R26-Tom;SmoM2 revealed that these aP2-Cre labeled cells in muscle interstitium are endothelial cells. In SCM sections, aP2-Cre labeled cells colocalize with Pecam1. When compared to aP2-Cre;R26-Tom mice, the addition of oncogenic SmoM2 (aP2-Cre;R26-Tom;SmoM2) results in embryonic expansion of the aP2-labeled muscle interstitial cells and formation of FN-RMS. These expansions became positive for the myogenic regulatory factor MyoD1 and did not express Pecam1 at embryonic day 17.5. Subsequent gene expression analysis revealed that both myogenic regulatory factors and endothelial genes are upregulated in tumor cells. Skeletal muscle and endothelial cells share a common progenitor, and together these results suggest that aberrant Sonic Hedgehog signaling promotes a myogenic fate switch in aP2-Cre expressing endothelial progenitor cells that results in FN-RMS formation. Citation Format: Catherine J. Drummond, Jason A. Hanna, Matthew R. Garcia, Daniel Devine, Jennifer Peters, Victoria Frohlich, David Finkelstein, Mark E. Hatley. Fusion-negative rhabdomyosarcoma originating from endothelial progenitors [abstract]. In: Proceedings of the AACR Conference on Advances in Sarcomas: From Basic Science to Clinical Translation; May 16-19, 2017; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(2_Suppl):Abstract nr A16.