Abstract A17: Silencing of Nuclear factor (erythroid-derived 2)-like 2 (NFE2L2) impairs growth and proliferation of hepatoblastoma cells

Abstract

Abstract Background: Hepatoblastoma (HB) is the most common primary, malignant liver tumor in children, occurring primarily in children under the age of 4 years. Although chemotherapy in conjunction with surgical resection has improved the prognosis for patients with HB, disease-free survival rates remain less than 50% for patients with advanced, metastatic or relapsed disease, highlighting the need for novel treatments. Additional subsets of tumors with a worse prognosis include those with histologic features of hepatocellular carcinoma (HCC) and tumors which develop in children older than 4 years. Nuclear factor erythroid-2–related factor 2 (NFE2L2, also known as Nrf2), a key transcriptional regulator for the cellular response to oxidative stress in normal cells, has been shown to contribute to malignant phenotypes of cancers. Recently, activating mutations in NFE2L2 have been reported in HCC and also in HB by our research team (Trevino et al1). These activating mutations of NFE2L2 and/or high expression of NFE2L2 have been observed notably in several pediatric liver tumors with high-risk features associated with a poor outcome. We hypothesize that NFE2L2 activation/overexpression may contribute to an aggressive tumor phenotype, and investigated its role in HB cell proliferation. Methods: Hep293TT cells, previously established using primary tumor tissues from a highly aggressive hepatoblastoma from a 5-year-old Caucasian female, were transfected with NFE2L2 small hairpin RNAs (shRNAs) to determine the effect of NFE2L2 suppression on cell proliferation and to identify potential NFE2L2-regulated targets. Analyses of Hep293TT cell proliferation and cell cycle kinetics were conducted using cell viability assays (CellTiter-Glo Luminescence), propidium iodide staining and FACS analysis. A comprehensive transcriptome analyses of Hep293TT cells was carried out by performing RNA sequencing (RNA-seq) using an Illumina HiSeq 2000. Results: RNA-seq analysis was performed on a set of pediatric cancer cell lines including Hep293TT, as well as reference liver cell lines HepG2 and Hep3B. After normalization, the analysis revealed high expression of NFE2L2 in Hep293TT cells with fold change of 2.01 relative to the mean expression level from the pediatric cancer cell line panel (14 cell lines). Compared to other liver cancer cell lines, fold change of 1.28 and 1.5 were measured from Hep3B and HepG2, respectively. Differential expression between NFE2L2 knockdown and parental cell will further demonstrate the molecular and functional influence of NFE2L2. Furthermore, knockdown of NFE2L2 inhibited cell proliferation in Hep293TT cells by inducing cycle arrest. Conclusions: Our results suggest that NFE2L2 may contribute to cell growth and proliferation in hepatoblastoma. Further studies are warranted to determine if NFE2L2 status can be used to stratify highly aggressive pediatric liver tumors and establish NFE2L2 as a potentially novel target for the treatment of HBs. 1Trevino et al, AACR Annual Meeting, 2012, Abstract #4592 This work was supported by grants from the Cancer Prevention and Research Institute of Texas RP101195 and NIH R21 CA122565. Citation Format: Hima Bansal, Tina Chen, Zhao Lai, Donald Parsons, Lisa Trevino, Dolores Lopez-Terrada, Milton Finegold, Dinesh Rakheja, Sarah Comerford, Robert Hammer, Bradley Pollock, Jaclyn Hung, Sanjay Bansal, Yidong Chen, Gail Tomlinson. Silencing of Nuclear factor (erythroid-derived 2)-like 2 (NFE2L2) impairs growth and proliferation of hepatoblastoma cells. [abstract]. In: Proceedings of the AACR Special Conference on Pediatric Cancer at the Crossroads: Translating Discovery into Improved Outcomes; Nov 3-6, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;74(20 Suppl):Abstract nr A17.