Understanding the role and regulation of PTEN in Adenovirus 5 infection

Cancer is caused by random, accumulated gene mutations (for example, to the tumour suppressor p53 and PTEN genes) that lead to dysregulated cell growth and proliferation and to invasion and metastasis. Current treatments include chemotherapy, radiotherapy and surgery, but these can be insufficient. Novel therapies are needed, including the designing of adenoviruses (Ads) to target and kill cancer cells.
Loss of the p53 gene product in cancer cells has been previously exploited to confer conditional replication of certain mutant Ads in cancer cells without affecting normal cells. p53 activates PTEN transcription and the PTEN protein is stabilised by p53. PTEN could present another promising target to enable cancer-cell specific targeting of Ad-based therapies.
Previous work in the Blair lab that led up to this project showed that both PTEN mRNA and protein were reduced by approx. 50% in Ad5-infected human cells. In this project, the activity of the PTEN promoter was analysed during Ad5 infection of cells in which the normal p53 genes were either present or absent (mediated by targeted deletion). Using dual luciferase assays that measure the activity of a transfected PTEN promoter, it was established that there was no significant difference in PTEN promoter activity between Ad5-infected or mock-infected cells and that the presence or absence of p53 did not affect PTEN promoter activity in Ad5-infected cells. This suggests that any reduction in PTEN mRNA that occurs during Ad5 infection is mediated at a post-transcriptional level. In addition, Ad5 was previously shown in the Blair lab to be restricted for growth in a glioblastoma cell line, U87MG, which is PTEN-null. Here, PTEN-expressing derivatives of U87MG were generated by retroviral transduction. These cell lines were characterised in preparation for detailed comparative studies on Ad5 replication and killing of isogenic PTEN-null and PTEN-positive cells.