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Investigating the cellular and anti-cancer effects of REST and HDAC inhibition in Daoy medulloblastoma cells

Alshawli, Abdulelah Saleh (2019) Investigating the cellular and anti-cancer effects of REST and HDAC inhibition in Daoy medulloblastoma cells. PhD thesis, University of Leeds.

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Abstract

REST is a transcriptional repressor protein and has been suggested to maintain the self-renewal potential of several brain tumours including medulloblastoma. REST represses transcription by recruiting several chromatin modifiers, including histone deacetylase (HDAC) enzymes to gene promoters which have been associated with several cancers. Nevertheless, HDAC inhibitors (HDACis) have shown promising results in targeting specifically tumour cells. Interestingly, REST expressing cells showed higher sensitivity to HDACis yet, it is not known if they induce their action through the HDACs recruited in REST repression complexes. Also, it is not fully understood how HDACis specifically inhibit the growth of tumour cells, and in what way normal cells protect themselves from the hyperacetylation effect of these drugs. In this study, I examined the contribution of REST in the medulloblastoma Daoy cell line by modulating its expression using CRISPR/Cas9. To explore the HDACis anti-cancer molecular mechanism, I profiled the gene expression of HDACis-treated tumour and normal cells at a single cell level using the next generation sequencing analysis. The results showed that disrupting REST expression induced upregulation of neuronal genes, slightly (~6 %) reduced the cell growth, prolonged the cell cycle, and reduced the migration ability of the tumour cells. However, it did not stimulate apoptosis or reduce the sensitivity of the cell to the HDACis. Treating the tumour cells with HDACis resulted in activating the TNFα signalling via NFκB pathway and modulating the expression of the cell cycle and Myc pathways. Whereas, exposing the normal cells to HDACis caused considerable changes on the transcriptome level. Collectively, the data of this study showed that blocking REST expression did not lead to tumour cell death. It also showed the ability of HDACis to induce their action independent of HDACs recruited in REST complexes. Treating with HDACis caused global gene expression changes in tumour and normal cells which activate a defined signalling pathways however, this raises concerns regarding the toxic side effects of HDACis in normal cells. Hence, the use of HDACis should be revised carefully even with their minimal side-effect in clinical trials.

Item Type: Thesis (PhD)
Keywords: HDAC, HDAC Inhibitors, HDACis, REST, Medulloblastoma, CRISPR/Cas9, shRNA, scNGS, 10x Genomics, SAHA, MS-275, Neurons, Daoy
Academic Units: The University of Leeds > Faculty of Biological Sciences (Leeds) > Institute for Molecular and Cellular Biology (Leeds)
Depositing User: Mr Abdulelah Alshawli
Date Deposited: 08 Jan 2020 12:12
Last Modified: 01 Aug 2020 00:18
URI: http://etheses.whiterose.ac.uk/id/eprint/24412

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