Abstract 2889: The Yes-Associated Protein plays a role in tumor formation, stemness, and therapy response in high risk neuroblastoma

Abstract

Neuroblastoma (NB) is the most common extra-cranial solid tumor of childhood. Despite intensive multimodal therapy, >50% of high risk (HR) NB patients relapse with chemotherapy-resistant disease. RAS/RAF/MAPK pathway mutations and increased Yes-Associated Protein (YAP) transcriptional activity are more prevalent in relapsed NB compared to diagnostic tumors. YAP is a transcriptional co-activator, regulated by the Hippo pathway that binds with TEAD in the nucleus to activate genes promoting cell growth, self-renewal, and survival. YAP aberrant transcription has been implicated in many tumor types, yet the role for YAP in NB is unknown. YAP protein is heterogeneously expressed in HR NB cell lines. We used human derived NB cell lines stably transduced to express (NGPYAP) or inhibit (SK-N-ASshYAP, NLFshYAP) YAP to further characterize YAP’s role in NB. While TEAD expression increases with YAP genetic inhibition, expression of YAP paralog TAZ did not change. SK-N-ASshYAP xenografts in NSG mice showed significantly delayed time to tumor formation and slower growth compared to empty vector transduced xenografts. Unmodified NB cells grown as neurospheres in neural basal media showed increased YAP expression and stemness markers, OCT4 and SOX2, by RT-qPCR. OCT4 and SOX2 were dependent on YAP expression as they did not increase in NLFshYAP neurospheres. Furthermore, NLFshYAP had decreased number and size of neurospheres compared to control. Given YAP’s role in tumor stemness, we treated unmodified NB cells with 13-cis-retinoic acid (RA) to see if YAP-induced stemness would overcome RA-induced differentiation. Interestingly, YAP expression and downstream targets, OCT4 and SOX2, increased in response to RA treatment. This suggests that RA paradoxically induces YAP transcriptional activity and may be selecting for a more chemoresistant tumor initiating phenotype. Lastly, YAP knockdown sensitized NRAS- and NF-1-mutated NBs to the MEK inhibitor, Trametinib, and to chemotherapy in vitro. In adult cancers, YAP’s role in therapy resistance is due to its regulation of anti-apoptotic Bcl-2 family proteins. However, YAP knockdown or induced expression in NBs did not change Bcl-2 family protein levels, nor did it affect Bim binding patterns to Mcl-1 or Bcl-2. Thus, investigations are ongoing to understand how YAP negatively regulates Trametinib response. We treated NB cells with Verteporfin (VP), an FDA-approved drug used to treat macular degeneration that inhibits YAP-TEAD interactions in other cancers. While VP inhibited YAP expression in NBs, it also had significant off target effects, potently killing YAP null NBs via light-activated generation of reactive oxygen species. Given the relevance of YAP in relapsed NBs and its effects on NB tumor formation, stemness, and therapy resistance, the need for targeted and specific YAP inhibitors is critical to reinstating therapy sensitivity at relapse. Citation Format: Jenny Shim, Yoo Joo Lee, Kaitlyn Janssen, Alexa Fritz, Kelly C. Goldsmith. The Yes-Associated Protein plays a role in tumor formation, stemness, and therapy response in high risk neuroblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2889.