Abstract 1163: Estrogen receptor beta (ESR2)-mediated tumor suppression of glioblastoma involves modulation of cell cycle and DNA damage response pathways
Authors: Vadlamudi Ratna K. , Sareddy Gangadhara Reddy , Li Xiaonan , Liu Jinyou , Garcia Lauren , Brenner Andrew 
Published: July 2016

Abstract
Purpose: Glioblastomas (GBM) are deadly brain tumors, have greater incidence in males than females. Epidemiological evidence supports a role of estrogen in tumor suppression. Estrogen signaling is mediated by estrogen receptor alpha (ESR1) and beta (ESR2). Our earlier studies showed that GBM preferentially express ESR2. However, mechanisms by which E2-ESR2 signaling promote suppression of GBM is poorly understood. The objective of this study is to determine the mechanisms by which ESR2 promote tumor suppression.
Experimental design: Established and patient derived primary GBM cells overexpressing or under expressing ESR2 were generated using lentiviral vectors. Three different ESR2 agonists (LY500307, Liquiritigenin, and Erb041) were used in the study and transcription changes elicited by ESR2 were profiled using RNA-seq. Cell cycle was analyzed using FACS analysis. In vivo efficacy of ESR2 agonists was tested in an orthotopic model using syngeneic murine GBM model (GL26), tumor growth was measured using bioluminescence, and survival was recorded. The synergy experiments using 119 FDA approved drugs was done using cell viability assay.
Results: ESR2 agonist treatment or ESR2 overexpression significantly reduced the viability and colony formation and induced apoptosis of various GBM cell lines with minimal effect on normal astrocytes. Further, ESR2 agonist (LY500307 5mg/kg/day) treatment significantly improved survival of GL26 tumor-bearing mice. RNA-seq studies using ESR2 overexpression or ligands treatment revealed downregulation of a number of genes involved in DNA repair, DNA damage response and cell cycle. The canonical pathways modulated by ESR2 included cell cycle, DNA damage, p53 signaling, and checkpoint regulation. In addition, pathways related to molecular mechanism of cancer, ILK signaling, Wnt signaling and glioma invasion were also altered. ESR2 agonist treatment significantly increased the percentage of cells in G2/M phase in U87, U251 and LN229 GBM cells when compared to vehicle further supporting the RNA-seq pathway analysis of cell cycle and G2/M DNA damage checkpoint regulation. Using combinational screening of 119 FDA-approved drugs combined with a low dose of ESR2 agonists (below the IC50 dose), we discovered synergism of ESR2 agonist treatment with several DNA-damaging agents. We independently validated synergy using additional ESR2 agonists with currently used chemotherapy drugs in GBM including temozolomide.
Conclusions: Our results demonstrate that ESR2 as tumor suppressor for GBM and ESR2 agonists have potential as a therapeutic agent for GBM. ERβ agonists’ ability to suppress pathways involved in DNA repair can be exploited to promote apoptosis of GBM cells.
Citation Format: Ratna K. Vadlamudi, Gangadhara Reddy Sareddy, Xiaonan Li, Jinyou Liu, Lauren Garcia, Andrew Brenner. Estrogen receptor beta (ESR2)-mediated tumor suppression of glioblastoma involves modulation of cell cycle and DNA damage response pathways. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1163.