Abstract 3400: Integrated functional RNAi screening and structural genomics identifies inverse co-modulators of TP53 family and NF-kB transitional activation as potential therapeutic targets in head and neck squamous cell carcinoma

Abstract

Abstract Head and neck squamous cell carcinoma (HNSCC) is the 6th most common cancer, with an annual incidence of over 600,000 new cases worldwide, and 50-60% mortality. Dysregulation of tumor suppressor TP53 in HNSCC occurs in over 90% of cases. We and others have observed that TP53 related family members p63 and p73 are also deregulated, together preventing transcription of growth arrest and apoptosis genes. Conversely, we have shown that members of the NF-κB/REL family of transcription factors are aberrantly activated, and drive expression of genes that promote cell proliferation, survival, inflammation, angiogenesis,and resistance in ∼70% of HNSCC cases. We have previously published that 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 and may serve as critical targets for therapy. 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, using antibiotic selection and fluorescence activated cell sorting. In collaboration with the RNAi screening facility at NIH, kinome and druggable target genes have been screened in our NF-κB reporter cell lines. Screening was performed by 48 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. These targets include genes participating in Toll/IKK such as IRAK1, PI3K/AKT signaling such as PIK3CA and PIK3R3, mitotic kinases such as BUB1B and AURKA, and calmodulin signaling such as CALM2 and CAMK1G. Candidate hits from RNAi screening data were first prioritized by integration with expression, mutation, and DNA copy number variation data from The Cancer Genome Atlas (TCGA) database. To further investigate the interactive signaling regulation between NF-κB and TP53 pathways, selected siRNAs that can inhibit NF-κB activation were cross screened in our TP53 reporter cell lines. Transcriptional activation of TP53 was induced with doxorubicin or cisplatin following 48 hr RNAi knockdown. We have shown that knockdown of targets in the PI3K/AKT pathway can inversely increase TP53 activation while inhibiting NF-κB activity. Selected targets were further validated by demonstration of anti-proliferative activity of single siRNAs in multiple HNSCC cell lines. Preclinical studies with PI3K-mTOR small molecule inhibitors indicate potential to co-modulate NF-κB and TP53 and inhibit HNSCC in vitro and in vivo. We predict that inhibition of targets that inversely modulate both TP53 and NF-κB signaling will be less prone to resistance and more effective in treatment of HNSCC. Supported by NIDCD intramural projects ZIADC000016, 73 and 74. Citation Format: Anthony D. Saleh, Shaleeka Cornelius, Hui Cheng, Scott Martin, Pinar Ormanoglu, Xinping Yang, Zhong Chen, Carter VanWaes. Integrated functional RNAi screening and structural genomics identifies inverse co-modulators of TP53 family and NF-kB transitional activation as potential therapeutic targets in head and neck squamous cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3400. doi:10.1158/1538-7445.AM2014-3400