High-grade gliomas are intrinsic malignant brain tumors and are resistant to the established treatment modalities. Epithelial to mesenchymal transition caused by hypoxic zone of tumor is responsible for chemoresistance. Standard chemotherapy (Temozolomide) is not much effective as a single agent and so there is a necessity of combining Temozolomide (TMZ) with other known glioma drugs for more effective cytotoxicity capable of abrogating chemoresistance. Cisplatin (CP), which is used as a second line of drug for glioma patients and acts as DNA damaging agent, could be a better choice for the combination strategy. In this study we have investigated the efficacy of the combination of TMZ and CP on glioma cell lines under normoxia (20% O2) and hypoxia (0.2% O2) and also compared its effect on epithelial to mesenchymal transition and cell migration.
The cell lines (A172 and LN229) with different p53 status were cultured under 20% (normoxia) and 0.2% (hypoxia) oxygen concentrations for the study. Cell viability analysis was done by MTT assay for the combination of TMZ and CP as compared to single agent and combination index was calculated. MTT assay was further correlated with caspase3/7 assay. Expression of EMT markers were quantified using Real Time PCR and wound healing assay was done to check the migration.
The combination of TMZ and CP showed a synergistic effect under both normoxia and hypoxia in A172 and LN229 cell lines with combination index less than 1. Caspase3/7 activity was found to be increased by the combination under both oxygen concentrations. Expression of mesenchymal markers was markedly reduced by the combination as compared to single agent whereas the expression of the epithelial markers was increased. The combination of TMZ and CP also reduced the migration of glioma cells as compared to single agent.
The synergistic combination of TMZ and CP markedly reduced epithelial to mesenchymal transition in glioma cell lines as compared to single agent which in turn reduced chemoresistance under hypoxia.
Clinical trial identification
All authors have declared no conflicts of interest.