Abstract 3104: PAX3-FOXO1 increases fibroblast reprogramming efficiency and drives self-renewal in alveolar rhabdomyosarcoma

Abstract Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children. Most alveolar (ARMS) cases bear the chimeric transcription factor PAX3/7-FOXO1 and have a worse prognosis compared to the embryonal subtype (ERMS). Previous studies identified a CSC subpopulation in ERMS but no study looked at the translocation positive RMS so far. This knowledge would be important to guide future treatment strategies. Here, we show that no CSC subpopulation can be detected in ARMS by a range of common methods. Furthermore, in vivo limiting dilution experiments confirmed that even 100 ARMS cells are sufficient to generate tumors in NOD/SCID mice. Thus, we hypothesized that ARMS follows a clonal evolution model. In line with this hypothesis, the stem cell factors Nanog, Oct4, and Sox2 were homogeneously expressed in ARMS cell lines and genetic loss- or gain-of-function experiments did not change cellular physiology unlike silencing of PAX3-FOXO1 which caused cell cycle arrest and apoptosis. Based on these observations we then investigated whether PAX3-FOXO1 itself can act as stem cell gene. For this we reprogrammed human fibroblast with the known factors Oct4, Sox2, Klf4, and c-Myc in presence or absence of PAX3-FOXO1. Interestingly, the presence of the fusion protein significantly increased the reprogramming efficiency leading to the generation of almost double the number of colonies of induced pluripotent stem cells (iPSC) than the control cells. PAX3-FOXO1 iPSC expressed the core stem cell genes as well as the PAX3-FOXO1 target gene AP2beta. Strikingly, PAX3-FOXO1 iPSC showed reduced ability to differentiate in vitro into all three germ layers. Ongoing experiments aim at confirming these results also in vivo by generation of teratomas. In conclusion and in contrary to ERMS, fusion protein positive ARMS seem to follow the clonal evolution model in which each clone has tumorigenic potential. Moreover, PAX3-FOXO1 is the critical factor for ARMS self-renewal and a general inhibitor of differentiation. Citation Format: Beat W. Schafer, Elisa Casanova, Sampoorna Satheesha, Melanie Müller, Paolo Cinelli, Paolo Cinelli. PAX3-FOXO1 increases fibroblast reprogramming efficiency and drives self-renewal in alveolar rhabdomyosarcoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3104. doi:10.1158/1538-7445.AM2014-3104