Abstract 1724: Integrated functional RNAi screening and structural genomics identify inverse co-modulators of TP53 family and NF-κB transitional activation as potential therapeutic targets in head and neck squamous cell carcinoma
Authors: Anthony Saleh, Shaleeka Cornelius, Scott Martin, Pinar Ormanoglu, Hui Cheng, Rita Das, Xinping Yang, Zhong Chen, Carter Van Waes
Journal: Cancer Research (2015)

Abstract Head and neck squamous cell carcinoma (HNSCC) is the 6th most common cancer worldwide with a 50-60% mortality rate. Deregulation of p53 family members in HNSCC occurs in over 90% of cases, preventing transcription of growth arrest and apoptosis genes. Conversely, members of the NF-κB/REL family are aberrantly activated in about ∼70% of cases, and drive expression of pro-proliferation, inflammation, angiogenesis, and therapeutic resistance genes. The function of different TP53 and NF-κB family members are inversely modulated within two major subsets of HNSCC, suggesting that common molecules and pathways coordinate this modulation. To test this hypothesis, we have developed HNSCC stable cell lines that report transcriptional activation of TP53 or NF-κB individually through a β-lactamase reporter. Whole genome siRNA screening in our NF-κB reporter cell line has been performed. Screening was performed by 56-hour knockdown with siRNAs, followed by stimulation with TNF-α. The results have revealed known and novel targets that maintain oncogenic NF-κB signaling in HNSCC. Validating the assay, most structural components of the TNF receptor complex and downstream NF-κB pathway genes, such as IKKs, scored highly in the screen. We integrated this functional genomics data with structural mutation and expression data from TCGA to prioritize genes that are frequently deregulated in HNSCC specimens from patients. Several co-receptors that may support NF-κB signaling were identified such as WNT9/FZD6, NOTCH2, and TGFβR2. Our primary novel finding was the identification of genes that regulate mitosis, such as CDC2, CDC7, CHEK1, WEE1 and CALM2. Several components of the kinetochore were also identified including NDC80, PLK1,TTK and AURKA. To validate our screening data, individual assays were performed with two siRNAs for a panel of 27 genes from the pathways mentioned above. &amp;gt;95% of siRNAs tested validated our screening data. Genes scoring well in our analysis were also assayed in our p53 reporter cell line using cisplatin and TNF-α as stimuli. Genes that inversely activate p53 were identified. To further investigate the relationship between kinetochore function and NF-κB activation, we treated cells with various G2/M inhibitors such as docetaxel and showed decreased NF-κB signaling. Future experiments are underway to determine the signaling mechanism that connects the kinetochore to NF-κB regulators. Experiments are also ongoing to determine if aberrant activation of NF-κB can serve as a biomarker for sensitivity to cell cycle inhibitors. Citation Format: Anthony Saleh, Shaleeka Cornelius, Scott Martin, Pinar Ormanoglu, Hui Cheng, Rita Das, Xinping Yang, Zhong Chen, Carter Van Waes. Integrated functional RNAi screening and structural genomics identify inverse co-modulators of TP53 family and NF-κB transitional activation as potential therapeutic targets in head and neck squamous cell carcinoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1724. doi:10.1158/1538-7445.AM2015-1724