Abstract 5005: Combined targeting of IKKα and β effectively suppresses NF-κB activation, cell survival and migration in head and neck squamous cell cancers
Authors: Carol Yan, Liesl Nottingham, Clint Allen, Tsu-Fan Cheng, Arun Pattatheyil, Xinping Yang, David Gius, Lenny Dang, Zhong Chen, Carter Van Waes
Journal: Cancer Research (2011)

Abstract Activation of the transcription factor NF-kappaB (NF-kB) has been found to be a key promoter of tumorigenesis and a contributor of chemotherapy resistance in many cancers, including head and neck squamous cell carcinomas (HNSCC). The NK-kB pathway is regulated through both canonical and noncanonical pathways involving the IkB kinases (IKK) α and β which together, have been implicated in cancer pathogenesis. However, the individual roles of the IKK subunits have not been previously elucidated and potential therapeutic targets still largely remain to be characterized. Endogenous IKKα and β expression and phosphorylation were analyzed by IHC staining and western blot in HNSCC tumor specimens and cell lines (UM-SCC 11A and 11B). Individual and dual overexpression and knockdown of IKKα and β were assessed for their effects on NF-kB activation by a luciferase reporter assay, and on tumor cell proliferation and migration in vitro via MTT and wound healing assays, respectively. HNSCC tissue and cell lines exhibited aberrant expression with increased nuclear localization and phosphorylation of IKKα and β when compared to normal controls. In a panel of 9 HNSCC cell lines, endogenous IKKα was more abundantly expressed than IKKβ. While individual overexpression of wt or constitutively activated IKKβ was more potent than IKKα, dual overexpression of IKKα and β exhibited the strongest induction of NF-kB activation. siRNA knockdown of either kinase resulted in decreased cell proliferation and migration and NF-kB activation, and a combinatorial effect was seen with double IKKα and β knockdown. Additionally, chemical inhibitors targeting IKKα and β were used to evaluate their potential for suppressing NF-kB activation and targeting the HNSCC malignant phenotype in vitro. Four small molecule inhibitors were studied that preferentially targeted IKKα&amp;gt;β (17-DMAG), IKKβ (MLN120b and SC514), or both IKKα and β (wedelolactone). Compared to the IKKβ specific inhibitor, MLN120b, 17-DMAG more potently inhibited NF-kB DNA binding in electromobility shift assays (EMSA) and NF-kB reporter gene activation. MTT proliferation assays and flow cytometric DNA cell cycle analysis demonstrated more potent effects of 17-DMAG and wedelolactone in inhibiting cell proliferation. We conclude that overexpression and signaling mediated by both IKKα and β contribute to NF-kB activation, and pathogenesis of proliferation and migration of HNSCC cells in vitro. Furthermore, dual siRNA silencing or chemical inhibition of IKKα and β kinases resulted in combinatorial inhibitory effects on NF-kB activation and the malignant phenotype. In the face of intrinsic and acquired chemoresistance limiting treatment options, IKKα/β represent promising novel therapeutic targets for fighting HNSCC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5005. doi:10.1158/1538-7445.AM2011-5005