Regulation Of Yap1 During Hypoxia And Its Novel Role In Vascular Mimicry And Angiogenesis

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related mortality. As in other solid tumors, angiogenesis is necessary for the growth and progression of NSCLC. Vascular mimicry is a phenomenon in which non-endothelial tumor cells form angiogenic vascular structures. We have shown in the past that NSCLC cancer stem-like cells (CSCs) have unique ability to undergo vascular mimicry. The present study shows that transcriptional co-activator YAP1, the oncogenic component of the Hippo pathway, enables the NSCLC CSCs to form angiogenic tubule-like structures in matrigel. Inhibition of YAP1 or depletion of YAP1 suppressed vascular mimicry of CSCs as well as angiogenic tubule formation by HUVECs in matrigel as well as in a fibrin gel bead assay (FIBA) assay. The NSCLC CSCs were found to have higher mRNA expression of VEGF receptor II (KDR) and Angiopoietin-2 (AngPT-2) and depletion of YAP1 reduced the expression of VEGF, KDR and AngPT-2 mRNA; these promoters were induced by YAP1 in transient transfection assays, suggesting a possible molecular mechanism by which YAP1 regulates angiogenesis. Hypoxia is a strong inducer of tumor angiogenesis, cancer progression and metastasis. Our experiments showed that NSCLC cells grown in hypoxic conditions or treated with hypoxia mimetic DMOG have higher YAP1 mRNA and protein expression. However, such a change was not observed in the expression of YAP1 orthologue, TAZ, nor in the canonical Hippo pathway proteins like LATS1u00262, Mst1u00262, Sav or Mob. We identified a novel regulation of YAP1 by prolyl hydroxylase PHD2, which is mainly known to regulate HIF1α. PHD2 was found to hydroxylate proline residue(s) in YAP1 in a region between aa 284 to aa 289 as seen by mutational analysis. YAP1 was found to directly bind to PHD2 and depletion of PHD2 or treatment with DMOG which is an inhibitor of prolyl hydroxylases, elevated YAP1 protein levels in the nucleus. Further, YAP1 was found to associate with HIF1α as detected by co-immunoprecipitation experiments and could enhance HIF1α-mediated induction of the VEGF promoter. Proximity ligation assays performed on TMA showed enhanced YAP1 and HIF1α interaction in lung tumor tissues compared to normal cells. Our data suggest a novel regulation of YAP1 in hypoxic environment that supports angiogenesis and tumor growth. Citation Format: Namrata Bora Singhal, Srikumar Chellappan. Regulation of YAP1 during hypoxia and its novel role in vascular mimicry and angiogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2145. doi:10.1158/1538-7445.AM2017-2145