Abstract 2946: AY4, an agonistic anti-death receptor 4 monoclonal antibody, induces death receptor 4-mediated apoptotic cell death in anaplastic thyroid cancer cells via downregulation of Bcl-xL with reactive oxygen species generation.

Abstract

Abstract Background: Anaplastic thyroid carcinoma (ATC) is an aggressive human tumor with a median survival of 6 months. We previously developed an agonistic anti-death receptor 4 monoclonal antibody, AY4, and demonstrated the anti-tumor effects of AY4 in head and neck cancer cells. Presently, we show that ATC cells are sensitive to AY4 and that the sensitivity correlates with reduced expression level of Bcl-xL and reactive oxygen species (ROS) generation. Methods: The effects of AY4 in Thyroid cell lines (C-643, U-HTH7, HTH83 and SW1736) was studied on apoptosis using MTT assy, Annexin V assay and Western blot analysis and was examed on the production of reactive oxygen species with DCFDA staining. HTH83 and C-643 cells were transfected with Bcl-xL siRNA using Lipofectamine 2000. Results: AY4 induced cell death of C-643, U-HTH 7, HTH83, and SW1736 cells. To elucidate the role of ROS generation in AY4-induced apoptosis of ATC cells, U-HTH 7 and SW1736 cells were pretreated with an anti-oxidant (N-acetyl-cysteine, NAC) followed by AY4 treatment. The cell death was blocked by NAC. AY4-induced cell death was accompanied by the down-regulation of the anti-apoptotic protein, Bcl-xL. To examine the link between the apoptotic response and Bcl-xL protein expression, U-HTH 7 cells were transfected with Bcl-xL plasmid. The consequence of the over-expression of Bcl-xL appeared to decrease AY4-mediated cell death by blocking ROS generation in U-HTH 7 cells. In contrast, Bcl-xL knock-down using small interfering RNA of Bcl-xL enhanced AY4 sensitivity in HTH83 and C-643 cells, and rendered the cells sensitive to AY4-induced cell death.. Conclusion: The results support the conclusion that the expression level of Bcl-xL is important in the AY4-induced apoptosis of ATC cells through ROS generation. AY4 may be a promising tool for ATC therapy. Keywords: Death receptor 4, agonistic antibody, apoptosis, ROS, anaplastic thyroid carcinoma (ATC) Citation Format: Bok-Soon Lee, Hyun-Young Cha, Yoo Seob Shin, Sung Un Kang, Hye Sook Hwang, Sam Kang, Jae Won Chang, Yong- Sung Kim, Chul-Ho Kim. AY4, an agonistic anti-death receptor 4 monoclonal antibody, induces death receptor 4-mediated apoptotic cell death in anaplastic thyroid cancer cells via downregulation of Bcl-xL with reactive oxygen species generation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2946. doi:10.1158/1538-7445.AM2013-2946