Targeting the endoplasmic reticulum stress pathway with tunicamycin and melatonin in the aggressive GBM T98G cell line

Glioblastoma (GBM) defined as degree 4 astrocytoma and is the most agressive primary brain tumor in the central nervous system. Tumor's heterogenous structure, invasiveness and inability of various therapeutics to pass through the blood brain barrier challange to find an effective and targeted therapy viort he GBM. Cancer researches indicated the importance of targeting endoplasmic reticulum stress and combining different types of therapeutics increases the effectiveness of the therapy. Tunicamycin is a natural antibiotic that causes endoplasmic reticulum stress via inhibiting N-linked glycosylation. On the other hand, melatonin is a biogenic amine that known for anticancer, antiviral antiinflammation effects. The aim of this thesis project is to determine molecular mechanisms of a combination therapy with the help of melatonin's anticancer effect in T98G GBM cell line that targets endoplasmic reticulum stress via investigating PERK/eIF2α/ATF4/CHOP axis. Therapeutic doses that are used for the thesis were determined by MTT test results and references from different research articles. In this context, qRT-PCR and western blot analysis were used to determine the effect of single and combined usage of therapeutics in PERK/eIF2α/ATF4/CHOP axis. In the analysis of qRT-PCR it is indicated that the application of single and combined doses of tunicamycin activated eIF2α/ATF4 axis and induces endoplasmic reticulum stress. Similarly in the western blot analysis, values of eIF2α and p-eIF2α found higher in single and combined tunicamycin groups. In addition to that, values of ATF4 found highest in the combined group. Molecular flourescent staining was used in order to determine single and combined usage of bioactive natural product effect on the apoptosis. Molecular imaging results showed that an effective apoptosis signal was observed in the combined group. All experiment results indicated that melatonin and tunicamycin combination could be a potential strategy for the treatment of GBM.