Defining cAMP regulated cell death in Glioblastoma

Abstract

Glioblastoma (GBM) is the most aggressive type of malignant brain tumour in adults. Despite treatment, GBM patients have a poor median survival rate of approximately 15 months. This poor survival is related to apoptosis resistance in GBM tumour cells. The cyclic adenosine monophosphate (cAMP) signalling pathway is a well-established second messenger system that regulates both cell survival and death, but its role in GBM is still not fully understood. As such, this thesis investigated if the cAMP pathway affects the expression of genes regulating apoptosis and cell viability in GBM cells. It also examined whether expression differences in these genes correlate with patient survival and treatment response, alongside the effect of forskolin-induced cAMP activation on tumour behaviour. To achieve this, cAMP gene expression signature analysis was performed using different public datasets, followed by literature validation of 30 selected genes. TCGA-GBM data were used for gene expression and survival analyses, while selected cAMP genes were used to analyse pathway enrichment across glioma grades and sex differences. An in vivo GBM mouse model was treated with forskolin, TMZ or vehicle control, followed by H&E staining, IHC for pCREB, BIM and cleaved caspase-3, and digital image analysis. Furthermore, the 30-gene cAMP signature showed different expression levels across GBM samples. Survival analyses did not show a significant survival effect for the selected cAMP genes. cAMP enrichment was lower in GBM than in LGG, and male and female GBM samples had similar enrichment scores. In the histological analysis, the forskolin group showed a less dense tumour region on H&E staining. IHC staining showed BIM, cleaved caspase 3 and pCREB signals in tumour regions, with stronger staining in the forskolin group. These results show that cAMP activation affects apoptosis in GBM tissue and cell viability