Growth and Molecular Characteristics of Temozolomide-Resistant Human A172 and R1 Glioblastoma Cells

Glioblastoma recurrence is caused by tumor cells resistance which can be initial or acquired resu-lting from therapy. Studies searching the markers that would allow predicting the level of glioblastoma cell therapy resistance are in progress. Complexity of the problem is due to a high heterogeneity of individual tumors as well as due to cellular composition in each tumor. In the present work, comparative study on the influence of temozolomide (in Temodal® form) single exposure on the well-known glioblastoma cell line A172 as well as on new line R1 was performed. Treatment of A172 (highly temozolomide-sensitive cell line) with 0.1 mM temozolomide showed that only individual cells persisted and resumed the proliferation. In R1 glioblastoma, single cells survived and resumed the proliferation upon treatment with 1.0 mM temozolomide. Populations obtained from these proliferating cells were designated as resistant ones. The presence of MGMT enzyme and the expression of genes responsible for chemotherapy resistance and tumor progression (MGMT, ABCB1, ABCC1, ABCG2), and of growth factor genes (VEGF, HGF), as well as the presence of IL-6 and IL-8 cytokines (and their encoding genes) was examined in resistant A172 and R1 cells. In A172 cells, methylated status of MGMT gene promoter and the absence of MGMT gene expression were confirmed. It was firstly shown that R1 glioblastoma was heterogeneous in methylation status of MGMT gene promoter and MGMT enzyme expression. In A172 and R1 resistant cell populations, the level of MGMT gene promoter methylation was lower than in intact cells, while MGMT gene expression was enhanced that could be clue to a greater resistance of these cells to chemotherapy. Expression of most genes connected with chemotherapy resistance and more aggressive course of the disease, as well as of growth factors genes and interleukin genes in resistant A172 cells was higher than in intact cells. In contrast, in resistant R1 cells, expression of the same genes (excluding ABCC1 and VEGF, in which the expression level changed insignificantly) was lower than in intact cells. Our results confirmed the significance of MGMT in formation of glioblastoma cell resistance to temozolomide. Prognostic value of other studied parameters is still considered ambiguous.