Glioblastoma multiform (GBM) tumors are very heterogeneous, organized in a hierarchical pattern, including cancer stem cells (CSC), and are responsible for development, maintenance, and cancer relapse. Therefore, it is relevant to establish new GBM cell lines with CSC characteristics to develop new treatments. A new human GBM cell line, named R2J, was established from the cerebro-spinal fluid (CSF) of a patient affected by GBM with leptomeningeal metastasis. R2J cells exhibits an abnormal karyotype and form self-renewable spheres in a serum-free medium. Original tumor, R2J, cultured in monolayer (2D) and in spheres showed a persistence expression of CD44, CD56 (except in monolayer), EGFR, Ki67, Nestin, and vimentin. The R2J cell line is tumorigenic and possesses CSC properties. We tested in vitro the anticancer effects of sodium selenite (SS) compared to temozolomide TMZ. SS was absorbed by R2J cells, was cytotoxic, induced an oxidative stress, and arrested cell growth in G2M before inducing both necrosis and apoptosis via caspase-3. SS also modified dimethyl-histone-3-lysine-9 (H3K9m2) levels and decreased histone deacetylase (HDAC) activity, suggesting anti-invasiveness potential. This study highlights the value of this new GBM cell line for preclinical modeling of clinically relevant, patient specific GBM and opens a therapeutic window to test SS to target resistant and recurrent GBM.