Abstract 1946: Activating transcription factor 5 (ATF5) in highly expressed in Stage 4, MYCN-amplified neuroblastoma

Abstract

Abstract Background: Activating transcription factor 5 (ATF5), a member of the ATF/CREB family transcription factor, has been shown to regulate the differentiation of neuroprogenitor cells into mature neurons. ATF5 has also been implicated in progression of glioblastoma multiforme. We have recently identified ATF5 as a potential synthetic lethal gene to MYCN-amplified neuroblastoma (NB) through a whole genome shRNA screen. In the present study we investigate the role of ATF5 in NB tumorigenesis. Methods: We analyzed the human NB patient data GEO (GSE45480, Kocak et al.) to investigate and correlate the expression of ATF5 with different stages of NB. Immunohistochemical staining for ATF5 was performed on primary human NB tumors. Quantitative RT-PCR and western blot for ATF5 were conducted on a panel of NB cell lines. Immunocytochemistry was performed to determine the intercellular localization of ATF5. ATF5 was silenced by a tetracycline inducible (tet-on) shRNA construct in BE(2)-C and SK-N-DZ cells. The cells were subjected to cell viability assay and soft agar colony formation assay to assess the function of ATF5. Results: Analysis of NB patient data GEO (GSE45480) from Kocak et al., demonstrated that ATF5 is highly expressed in Stage 4 MYCN-amplified tumors compared to Stage 4, MYCN-non-amplified tumors (P = 2.56e-10) or Stage 1 tumors (P = 7.30e-18). ATF5 is also highly expressed in Stage 4, MYCN-non-amplified tumors vs. Stage 1 tumors (P = 2.8e-7). High expression of ATF5 strongly correlated with poor survival of NB patients (P = 2.88e-6, NCI Oncogenomics, Obertheur data set). Quantitative PCR detected ATF5 transcript in all eight NB cell lines tested. Western blot analyses demonstrated high expression of ATF5 in the MYCN-amplified neuroblastoma cell lines BE(2)-C, SK-N-DZ, and IMR32. Nuclear ATF5 staining was detected in all three cell lines as well as in Stage 4 NB tumors. Silencing of ATF5 reduced BE(2)-C cell viability to 30%, 48 hr after the addition of doxycycline. Silencing of ATF5 significantly reduced the colony formation of BE(2)-C and SK-N-DZ cells in soft agar (p<0.05) Conclusions: Our study demonstrates that ATF5 is highly expressed in Stage 4 MYCN-amplified tumors, and that silencing of ATF5 inhibits proliferation. These results suggest that ATF5 has a role in NB tumorigenesis and it represents a potential therapeutic target in MYCN-amplified neuroblastoma. Citation Format: Debarshi Banerjee, Shuobo Zhang, Gonzalo Lopez, Andrea Califano, Angela Kadenhe-Chiweshe, Darrell Yamashiro. Activating transcription factor 5 (ATF5) in highly expressed in Stage 4, MYCN-amplified neuroblastoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1946. doi:10.1158/1538-7445.AM2015-1946