Abstract 1325: PCI-5002 leads to enhanced radiosensitization via induction of necrotic cell death by promoting autophagy in non-small cell lung tumor xenograft model

Abstract

Abstract Purpose: We previously showed that Bax/Bak inhibition induces autophagy and promotes radiosensitivity in vitro and in vivo. Therefore, we further investigated the specific mechanism by which radiation triggers necrotic cell death via autophagy in H460 cells treated with PCI-5002, water-soluble zinc ionophores, in vitro or in vivo. Methods: Clonogenic assay was used to determine radiosensitivity. H460 lung cancer xenografts was generated and immunohistochemistry (IHC) was performed on tumor sections for tumor cell proliferation (Ki67 staining), angiogenesis (von Willebrand Factor staining), and apoptosis (cleaved caspase-3 antibody staining). Autophagy was determined by p62 protein, which is named sequestosome 1(SQSTM1) and links polyubiquitinated proteins to the autophagic machinery in vivo and punctuate localization of GFP-LC3 fusion protein, which observed under a confocal fluorescence microscope in vitro. Necrosis was determined with HMGB1 release and Propidium lodide (PI) staining. Results: The PCI-5002 markedly enhanced sensitivity of MEF WT cells to radiation (DER=1.43, p=0.003) in clonogenic assay. This enhanced radiosensitization was associated with an increase in characteristic punctuate localization of GFP-LC3 when was blocked a cleaved caspase-3 in MEF WT cells treated with PCI-5002, suggesting that autophagy was induced simultaneously. We found that PCI-5002 with radiation showed a dramatically enhanced sensitivity of H460 cells to radiation (DER=1.92, p=0.001) in clonogenic assay, and tumor growth delay in a mouse hind limb xenograft model in vivo, and was well tolerated compared to control. In addition, the concurrent administration of PCI-5002 and radiation in vivo significantly decreased cell proliferation and vascular density compared to radiation alone. Consistent with the in vitro results, treatment both PCI-5002 and radiation showed a dramatically increase in HMGB1 as a marker for Necrosis and a significant decrease in p62 protein level for detecting autophagic flux in the lung tumor xenograft model. Conclusions: These results suggest that induction of necrosis via promotion of autophagy may be utilized as a novel strategy to enhance radiation therapy in lung cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1325.