Abstract 485: Chronic exposure to a low dose of interferon-β increases levels of DNA damage resistance signature genes through unphosphorylated ISGF3

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Ionizing radiation and DNA damaging chemotherapeutic agents are used widely to treat many cancers, but the presence of cells that resist DNA damage often leads to failure to cure. One of the specific features of therapy-resistance is high expression of Interferon-Related DNA damage resistance Signature (IRDS) genes. The IRDS genes are a subset of (about 30 genes) Interferon Stimulated Genes (ISGs) including IFI27, OAS2, Mx1, and IFIT1. However, pro-apoptotic and anti-proliferative ISGs, such as IRF1, caspases, and FASL, are not included in this signature. In this study, we elucidated the molecular mechanism of how the expression of IRDS genes are increased, with no increase of other ISGs, in therapy-resistant cancer cells. When cells expressed high levels of IRF9 and STATs 1 and 2 in the absence of phosphorylation of Tyr-701-STAT1 and Tyr-690-STAT2, the expression of IRDS genes was increased, but that of other ISGs was not. The un-phosphorylated STAT1, STAT2, and IRF9 proteins formed a complex resembling Interferon Stimulated Gene Factor 3 (ISGF3), a major transcription factor in type I interferon signaling. While ISGF3, consisting of phosphorylated STATs 1 and 2 and IRF9, mediates the expression of whole set of ISGs, the unphosphorylated ISGF3 (U-ISGF3) induces only IRDS genes. ChIP assay revealed that U-ISGF3 selectively bound to the promoters of IRDS genes (IFI27, OAS2, and Mx1), but not to those of other ISGs (MyD88, IFI16, and IRF1). Continuous exposure of cells to a low level of interferonβ, which might be produced by cancer cells or surrounding immune cells, lead to steady-state increases in the expression of only U-ISGF3 and IRDS genes with no increase of phosphorylated ISGF3 and non-IRDS ISGs. We examined the correlation between the U-ISGF3 levels and DNA damage resistance in various small cell lung carcinoma (SCLC) cell lines. The expression of IRF9 and STATs 1 and 2 proteins varied in different SCLC cell lines, and high levels of U-ISGF3 correlated with resistance to DNA damage. H82, H1048, and H146 cells, which express low levels of U-ISGF3, were sensitive to DNA damage induced by radiation, etoposide, and doxorubicin, while H196, H1688, and H2195 cells expressing high levels of U-ISGF3 were resistant to DNA damage. In response to radiation, γH2A.X levels were significantly increased and remained high up to 48 hrs in H1048 cells, but the increased γH2A.X levels returned to lower levels in H196 cells, suggesting that U-ISGF3-induced gene products may increase DNA damage repair. When STAT1 was knocked down to reduce U-ISGF3 levels in H196 cells, γH2A.X levels increased by radiation remained high. Knocking IRF9 down also decreased the resistance to DNA damage. In conclusion, U-ISGF3, which is increased by a low dose of interferons, plays an important role in inducing DNA damage resistance, and its targeting sensitizes cancer cells to DNA damage induced by various cancer therapies. Citation Format: HyeonJoo Cheon, Elise G. Holvey-Bates, George R. Stark. Chronic exposure to a low dose of interferon-β increases levels of DNA damage resistance signature genes through unphosphorylated ISGF3. [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 485. doi:10.1158/1538-7445.AM2014-485