Abstract A29: Respective role of TEAD and Smad signaling in YAP-mediated osteosarcoma tumor growth and lung metastatic development
Authors: Morice Sarah , Danieau Geoffroy , Mullard Mathilde , Amiaud Jérôme , Brion Régis , Renault Sarah , Tesfaye Robel , Dupuy Maryne , Talbot Julie , Ory Benjamin , Brounais-Le Royer Bénédicte , Corre Isabelle , Brouchet-Gomez Anne , Rédini Françoise , Verrecchia Franck 
Published: August 2020

Abstract
Osteosarcoma (OS) is the main malignant primary bone tumor in children and adolescents for whom the prognosis remains poor, especially when metastases are present at diagnosis. Because a) gene set enrichment analysis (GSEA) of expression data from a cohort of OS patients reveals a Hippo conserved signature in OS and b) proximity ligation (PLA) and immunoprecipitation (IP) assays have shown an interaction between YAP and TEAD, the main effector of Hippo signaling pathway, in OS cell lines, we investigated its implication in OS development. To elucidate the role of TEAD in YAP-driven OS development, we probed the consequences of YAP activation using overexpression of YAPS127A (constitutively active YAP) and YAPS94A (TEAD-binding deficient YAP protein). Firstly, in vivo and in vitro experiments demonstrate a) the crucial role of TEAD in YAP-driven cell proliferation and in vivo tumor growth and b) a correlation between the expression of TEAD1 and the survival outcomes of OS patients. In this context RNAseq analysis and GSEA identified more than 300 genes related to cell proliferation that are significantly overexpressed in YAPS127A cells compared to control cells and YAPS94A cells. Secondly, we study the effects of YAP/TEAD interactions on the main biologic functions’ links to metastasis dissemination. We show an increase of cell migration using KHOS-S94A and KHOS-S127A, suggesting that YAP/TEAD interaction is not necessary to increase migration in OS. GSEA identifies genes related to epithelial-mesenchymal-transition (EMT), cell migration and TGF-β that are significantly overexpressed in YAPS127A and YAPS94A cells, strongly suggesting that TEAD is not required in YAP-driven expression of genes related to EMT, cell migration, and TGF-β signaling. Thirdly, using PLA and IP, we show an interaction of YAP/SMAD3 in OS cell line independently of YAP/TEAD interaction. In this context, using Tβ²R1 inhibitor, SD-208, we demonstrate the role of TGF-β/smad signaling in YAP-driven cell migration and metastatic process in OS. Finally, we investigated the effect of verteporfin, identified as an inhibitor of the Hippo signaling pathway, on OS progression. In vivo experiments showed that verteporfin reduces primary tumor growth and lung metastases development. In vitro experiments demonstrated that verteporfin decreases cell viability and inhibits the TGF-β/Smad3 cascade and the response of TGF-β key targets involved in the metastasis dissemination process. Together, these results defined the specific role of TEAD and SMAD3 in YAP-driven OS tumor and lung metastatic process. In addition, we demonstrated that YAP could be considered as a therapeutic target in OS.
Citation Format: Sarah Morice, Geoffroy Danieau, Mathilde Mullard, Jérôme Amiaud, Régis Brion, Sarah Renault, Robel Tesfaye, Maryne Dupuy, Julie Talbot, Benjamin Ory, Bénédicte Brounais-Le Royer, Isabelle Corre, Anne Brouchet-Gomez, Françoise Rédini, Franck Verrecchia. Respective role of TEAD and Smad signaling in YAP-mediated osteosarcoma tumor growth and lung metastatic development [abstract]. In: Proceedings of the AACR Special Conference on the Advances in Pediatric Cancer Research; 2019 Sep 17-20; Montreal, QC, Canada. Philadelphia (PA): AACR; Cancer Res 2020;80(14 Suppl):Abstract nr A29.