Abstract B02: Elevated YAP expression associates with EMT, stemness, and angiogenic properties of TNBC cell lines and recurrence in TNBC patients

Abstract

Abstract Triple-negative breast cancer (TNBC) is considered one of the aggressive breast cancer subtypes with higher risk of recurrence and mortality compared to other subtypes. There is a need to identify molecular markers and drivers that could distinguish metastatic subset of TNBC. Based on our previous study (Kulkarni et al., Oncotarget 2018), we investigated for association of elevated YAP expression specifically in TNBC tissue samples and cell lines. Six TNBC cell lines with different degrees of YAP expression were selected for the study. Cell lines were grouped based on high, intermediate, or low nuclear YAP expression. After stable knockdown of YAP expression using retrovirus, these three groups of TNBC cell lines were assessed for migration, mammosphere formation, and angiogenic properties. Further, detailed characterization of media supernatants was carried out to identify angiogenic markers altered after YAP knockdown. Metastatic potential of the cell lines after YAP knockdown was assessed using mouse tail vein injection model for lung metastasis nodules and lung Mets were analyzed for angiogenic marker. With appropriate ethical approvals, frozen TNBC tumors, along with adjacent normal tissue, were analyzed for YAP and its angiogenic target expression. The expressions were co-related with clinical parameters such as age at diagnosis, grade, lymph node positivity, and recurrence. TNBC cell lines with high YAP and intermediate YAP expression showed significantly reduced migration and mammosphere formation after YAP knockdown. Similarly, media supernatants from high YAP and medium-YAP cell lines showed severely affected angiogenic potential after YAP knockdown. Fluidigm analysis for YAP-signature (Kulkarni et al., Oncotarget 2018) gene expression showed significantly reduced expression of YAP-target genes involved in EMT and stemness in these cell lines. Proteomic analysis of media supernatants revealed expression of two angiogenic factors to be downregulated after YAP knockdown in all 4 cell lines. Metastatic potential of high-YAP expressing cell line in mouse tail vein injection model was severely affected after YAP knockdown. The two TNBC cell lines with low YAP expression did not show any effect on mammosphere formation or angiogenic properties after YAP knockdown; neither had any effect on expression of the angiogenic targets. Patient samples with elevated YAP expression in the tumor compared to adjacent normal tissue showed strong association with younger age at diagnosis, high grade, and lymph node positivity. Our analysis of six TNBC cell lines has uncovered YAP to be significantly associated with increased mesenchymal properties of TNBC cell lines. Analysis of TNBC tissue samples strengthened this association of elevated YAP expression with aggressive clinical parameters within TNBC patients. With further validation, YAP expression can be employed as a prognostic marker to predict aggressive and recurrent TNBC for close follow-up and aggressive treatment planning. Citation Format: Madhura Kulkarni, Nurfarhanah Bte Syed Sulaiman, Supriya Srivastava, Tuan Zea Tan, Thomas Choudary Putti, Xiaomeng Wang, Wanjin Hong, Soo Chin Lee, Yoshiaki Ito. Elevated YAP expression associates with EMT, stemness, and angiogenic properties of TNBC cell lines and recurrence in TNBC patients [abstract]. In: Proceedings of the AACR Special Conference on the Hippo Pathway: Signaling, Cancer, and Beyond; 2019 May 8-11; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(8_Suppl):Abstract nr B02.