Abstract 3586: Silencing of GLSiso2 (GAC) decreases cell proliferation and induces cell death in glioblastoma cell line
Authors: Franco Yollanda Moreira , Soares Roseli Silva , Oba-Shinjo Sueli M. , Marie Suely Kazue Nagahashi 
Published: August 2019

Abstract
The metabolic reprogramming is currently recognized as one of the hallmarks of cancer. Tumor cells have unusual metabolic activity in comparison to normal cells, and ability to reprogram their metabolic machinery to meet their biosynthetic and bioenergetic needs. Factors such as hypoxia, mutation in oncogenes and changes in signaling pathways may induce an upregulation of anabolic processes and suppression of catabolic pathways that leads to the maintenance of bioenergetics, redox status, cell signaling, and biosynthesis, which support rapid cell proliferation. Tumor cells can use a variety of alternative energy production pathways depending on the availability of nutrients. Glutaminolysis, the process where glutamine is transported into the cells and converted into α-ketoglutarate (α-KG) to enter into the tricarboxylic acid (TCA) cycle, is regulated positively in cancer. Glutaminase (GLS) is the main regulator of this pathway and presents two forms in humans: kidney-type glutaminase (KGA or GAC) and liver-type GLS2 (LGA or GAB), both with distinct tissue distribution, regulation and functions. The GLS presents two isoforms: GLSiso1 and GLSiso2, and their regulation are not yet well understood. The aim of this study was to analyze the expression of GLS in astrocytoma of different malignant grades. Additionally, in vitro assays were performed in a glioblastoma cell line (U87MG) to access GLSiso2 functions in astrocytoma. Differential expression levels of both isoforms were detected in 173 astrocytomas of our group cohort. A gradual increase of both isoforms expression levels was observed in parallel to the increase of the tumor malignancy. However, high expression level of GLSiso1 was detected in normal brain samples, while very low expression of GLSiso2. Hence, the isoform 2 is more eligible and relevant as a therapeutic target. To analyze the role of GLSiso2 in gliomagenesis, the isoform-specific transcript expression was inhibited by the interfering RNA method in U87MG cells. The efficiency of silencing was confirmed at gene and protein levels by quantitative real time PCR and western blotting, respectively. Additionally, GLSiso2 silenced cells presented significant decrease of cell proliferation, a temozolamide-senzitizing effect and an increase of cell death, when compared to control cells. Total GLS protein distribution was analyzed in different grades of astrocytoma through immunohistochemistry. The higher the malignant grade of astrocytomas, the higher the expression of GLS observed in tumor cell cytoplasm. Interestingly GLS was also expressed in endothelium, with higher expression in GBM cases compared to lower grades of astrocytoma and non-neoplastic brain tissues. Further analysis of mechanistic role of GLS isoforms in the endothelium cell compartment might better clarify a possible complementary effect in the metabolic reprogramming over the tumor growth.
Citation Format: Yollanda Moreira Franco, Roseli Silva Soares, Sueli M. Oba-Shinjo, Suely Kazue Nagahashi Marie. Silencing of GLSiso2 (GAC) decreases cell proliferation and induces cell death in glioblastoma cell line [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3586.