Phenotypic changes on central nervous system (CNS) tumor cell lines cultured in vitro 2D and 3D models and treated with cisplatin.

Despite aggressive therapy, most patients with brain tumors present disease relapse due to the cellular and molecular nature of these tumors. One of the models that best explains the heterogeneity observed in CNS tumors is the presence of cancer stem cells (CSCs). In this paper, we evaluated platinum-based response in brain tumor U-87 MG, LN-18, and KELLY cell lines cultured in monolayer (2D) and neurosphere (CSC enrichment- 3D) models. We evaluated mRNA expression of heat shock proteins (HSPA1A, HSPB1, HSPA1AL, TRAP1, and HSPD1), and DNA repair gene ERCC1. Changes in cell cycle and glycosylation profile were assessed by flow cytometry. After treatment with cisplatin, we found that the mRNA expression of HSPs markedly increased in the U-87 MG and LN-18 neurosphere cells. In KELLY monolayer cells, cisplatin induced upregulation of all genes. In KELLY neurosphere cells, only the HSPA1A, HSPB1, TRAP1, and HSPD1 genes were upregulated. The proportion of cells in the G0/G1 phase was significantly higher in U-87 MG neurosphere cisplatin-treated cells. A trend towards a greater proportion of cells in the S phase of U-87 MG monolayer cisplatin-treated cells was also observed. On the other hand, a significant decrease in the number of cells in the S phase and an increase in G2/M was observed in LN-18 monolayer cisplatin-treated cells. Glycosylation analysis using lectins showed a higher surface binding for PNA in the U-87 MG treated monolayer and a lower binding for Concanavalin A in the treated neurospheres. The binding of Isolectin GS-IB4, GSII, and SBA in KELLY monolayer cisplatin-treated cells was lower whereas the proportion of cells labeled with Concanavalin A was higher. In the KELLY neurosphere cisplatin-treated cells, the binding of Concanavalin A was lower than nontreated cells. Thus, our findings strongly supported the idea that definitions of phenotypic characteristics may help to establish better therapeutic strategies for brain tumors.

Central nervous system (CNS) tumor cell heterogeneity contributes to differential platinum-based response in an in vitro 2D and 3D cell culture approach.

One of the models that best explains the cellular heterogeneity observed in central nervous system (CNS) tumors is the presence of cancer stem cells (CSCs). CSCs can originate from differentiated adult cells that return to an undifferentiated stage through the mechanism known as epithelial-mesenchymal transition (EMT). In this paper, we evaluated cellular and molecular heterogeneity and the participation of the epithelial-mesenchymal transition (EMT) in glioblastoma (U-87 MG and LN-18) and neuroblastoma (KELLY and IMR-32) cell lines cultured in monolayer (2D) and neurosphere (CSC enrichment- 3D) models. For this, after treatment with cisplatin, we studied different cell subpopulations by immunophenotyping using neural stem cell/progenitor markers (ALDH, CD24, CD56, and CD133), mesenchymal stem cell markers (CD73, CD90, CD105, and CD146) and hematopoietic markers (CD14, CD19, CD34, CD45, and HLA-DR) and mRNA expression profiles of genes related to EMT, such as ZEB1, TWIST1, TGFB1, STAT3, and lncRNA HOTAIR. In addition, we evaluated the growth capacity of residual cells when treated with cisplatin using the chorioallantoic membrane (CAM) model to study disease relapse. After treatment with cisplatin, we found that the expression of STAT3 and TGFB1 genes markedly increased in the neurosphere of the IMR-32 cell line, and TWIST1 was upregulated in the neurosphere of LN-18. Only the nontreated monolayer of LN-18, KELLY, and IMR-32 amplified the lncRNA HOTAIR. The IMR-32 cell line exhibited an enrichment of CD24+/ALDH+ and this cell subset decreased after cisplatin treatment. We observed the loss of CD146+/CD73+ cell subpopulations in U-87 MG monolayer and neurosphere models, after cisplatin treatment, while in LN-18 monolayer cisplatin-treated cells, CD73+/CD90+ cell subpopulations increased. Neuroblastoma cell lines showed CD14+/HLA-DR- cell subpopulations representative of myeloid-derived suppressor cells (MDSCs). Tumors generated from residual cells, after exposure to cisplatin, grafted on CAM showed patterns of organization different from those of the controls. Thus, our findings strongly supported the idea that definitions of tumor phenotypic characteristics may help to establish better therapeutic strategies for the development of new drug targets.