Abstract 2483: Estrogen receptor beta enhances chemotherapy response in GBM
Authors: Zhou Mei , Sareddy Gangadhara R. , Liu Jinyou , Li Mengxing , Viswanadhapalli Suryavathi , Li Xiaonan , Tekmal Rajeshwar R , Brenner Andrew , Vadlamudi Ratna K. 
Published: August 2018

Background: The incidence of developing glioblastoma (GBM) is greater in men than in women, and women of reproductive age have a survival advantage over men and postmenopausal women. These correlative findings suggest that estrogens play a significant role in suppression of GBM, but how they might do so is poorly understood. GBM cells express the estrogen receptor beta (ERβ) that functions as a tumor suppressor. However, the in vivo significance of endogenous ERβ and its role in GBM progression are incompletely understood. The objective of this study is to examine the role of endogenous ERβ in the progression and chemotherapy response of GBM.

Methods: To study the functions of endogenous ERβ in GBM cells, we have generated ERβ knockout (ERβKO) GBM cells using CRISPR/Cas9 system and lentivital-ERβshRNA transfected primary GBM cells (ERβKD). As a second model, we generated multiple ERβ overexpressing GBM model cells (both established and primary GBM cells) using lentiviral transduction. Effect of ERβ-KO, -KD or -overexpression was studied using cell proliferation, invasion and apoptosis using established in vitro assays. Mechanistic studies were conducted using RNA-seq, RT-qPCR and signaling analysis. In vivo role of ERβ was studied using orthotopic models of GBM and mouse survival was determined using Kaplan-Meier survival curves.

Results: ERβKO or ERβKD increased, while ERβ overexpression reduced the proliferation of GBM cells. To determine the effect of ERβKO on mice survival, U251-WT and U251-ERβKO cells were implanted intracranially into immunocompromised mice. Compared to control mice, ERβKO mice had significant reduction in survival. Further, IHC analysis of tumor sections from these mice revealed that ERβKO tumors had more expression of the proliferation marker Ki-67 than the control tumors. RNA-seq analyses using U87WT and U87ERβKO cells revealed that the ERβKO-modulated genes were related to DNA damage response, p53 pathway, NF-κB signaling, JAK-STAT, and mTOR pathways. Further, RNA-seq studies using U87 and U87ERβ overexpression models revealed downregulation of a number of genes involved in DNA repair, DNA damage response (DDR), ATM signaling and cell cycle checkpoint control. Mechanistic studies showed that ERβ modulates expression of genes involved in apoptosis and DDR, thus enhancing the response to chemotherapy drugs. Compared to control, ERβKO cells exhibited resistance to DNA-damaging agents including cisplatin and temozolomide, while ERβ-overexpressing cells were highly sensitive to these DNA-damaging agents. We also validated ERβ sensitization to temozolomide using orthotopic GBM models.

Conclusion: Using ERβ-KO and -KD GBM model cells, we have provided strong evidence demonstrating tumor suppressor and therapy sensitization potential of endogenous ERβ. Our results suggest that upregulation of ERβ expression/functions is an attractive therapy for treating GBM.

Citation Format: Mei Zhou, Gangadhara R. Sareddy, Jinyou Liu, Mengxing Li, Suryavathi Viswanadhapalli, Xiaonan Li, Rajeshwar R Tekmal, Andrew Brenner, Ratna K. Vadlamudi. Estrogen receptor beta enhances chemotherapy response in GBM [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2483.