Abstract 358: LTB and LTBR mediates alternative Nf-kB activation through NIk and RELB/NF-kB2 to promote cell migration of HNSCC

Abstract

Abstract Nuclear factor-κB (NF-κB) includes a family of NF-κB/REL transcription factors that regulate key inflammatory and cancer genes, and which are aberrantly activated in head and neck squamous cell carcinomas (HNSCC). Lymphotoxin beta/Lymphotoxin beta receptor (LTβ/LTβR) can mediate activation of the “alternative” NF-κB pathway that includes NF-κB-inducing kinase (NIK), Inhibitor-κB kinase α (IKKα), and transcription factors RELB/NF-κB2. While LTβ is expressed in lymph nodes where HNSCC spread, the role of this pathway in pathogenesis of HNSCC is not well studied. Analysis of The Cancer Genome Atlas (TCGA) data reveals that LTβR is amplified or overexpressed in larynx or oral cavity HNSCC, while LTα/β, NIK, (MAP3K14) and RELB are overexpressed in oropharynx and tonsil cancers. Similarly, we observed overexpression of LTβR, NIK, and RELB proteins in subsets of HNSCC cell lines. In UM-SCC46, LTβ enhanced nuclear translocation of RELB/NF-κB2 (p52) subunit proteins, while LTβR siRNA knockdown decreased the expression of its target kinase NIK and RELB/NF-κB2 (p52). NIK knockdown also decreased RELB/NF-κB2 (p52) protein expression, whereas LTβ treatment stabilized NIK, and RELB/NF-κB2 (p52). Functional LTβ-mediated NF-κB activation was examined using a stably transfected NF-κB reporter cell line for β-lactamase/Fret activity. Knockdown of LTβR and NIK decreased NF-κB activity, whereas treatment with LTβ partially restored it. Treatment with a NIK inhibitor (1, 3[2H, 4H]-Iso-Quinoline Dione) reduced the protein expression of NIK, IKKα, and RELB/NF-κB2 (p52) in the cytoplasm and/or in the nucleus, and blocked LTβ induced translocation of RELB to nucleus. NIK and RELB knockdown by siRNA or using NIK inhibitor slowed HNSCC migration. Knockdown of NIK inhibited the LTβ-induced expression of MET, and SERPINE1 genes involved in migration, and knockdown of MET by siRNA inhibited the migration of HNSCC cell lines. Our findings show that LTβ/LTβ receptor promotes activation of the alternative NIK-IKKa-RELB/NF-κB2 pathway, to enhance MET-mediated cell migration in HNSCC. This may help explain why HNSCC exhibit migration and metastasis to local lymph nodes, where LTβ is expressed. The constitutive and LTβ induced activation of NIK and downstream NF-κB alternate pathway proteins, RELB and NF-κB2/p52 could be therapeutic targets in HNSCC. (Supported by NIDCD intramural projects ZIA-DC-000016, 74) Citation Format: Rita Das, Jamie Coupar, Anthony saleh, Paul E. Clavijo, Zhong Chen, Carter VanWaes. LTB and LTBR mediates alternative Nf-kB activation through NIk and RELB/NF-kB2 to promote cell migration of HNSCC [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 358. doi:10.1158/1538-7445.AM2017-358