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Title: Driver genes in CTCL : a genomic approach for screening and a detailed analysis of a candidate gene
Author: Woollard, Wesley Jackson
ISNI:       0000 0004 6347 7340
Awarding Body: King's College London
Current Institution: King's College London (University of London)
Date of Award: 2017
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The cutaneous T-cell lymphomas (CTCLs) mycosis fungoides (MF) and Sézary syndrome (SS) are T-cell malignancies affecting the skin. The heterogeneous genomic landscape of CTCL has hindered the identification of driver genes. However, maturing sequencing technologies and candidate gene studies of chromosomal hotspots such as 9p21 can be used to identify putative driver events. The aims of this thesis were to investigate: (i) if MTAP (found on 9p21) undergoes selective loss; (ii) if MTAP loss affects dimethyl-arginine status; and (iii) identify putative driver genes of CTCL. Tumour samples from 280 CTCL patients were analysed by QPCR for copy number changes across 9p21. There is a high propensity for MTAP loss (34%) occurring independently of CDKN2A loss (12%) across all CTCL stages. Expression of MTAP mRNA was measured by QPCR in 20 tumours, reduced MTAP mRNA coincides with MTAP loss. However, reduced mRNA also occurs in tumours without MTAP loss indicating selective pressure to silence MTAP. Genomic screening indicates that mutations are rare in CDKN2A and MTAP in CTCL suggesting that epigenetic mechanisms may be repressing MTAP expression. Promoter methylation studies in CTCL cell lines suggest aberrant methylation inhibits MTAP expression. Knock-down of MTAP by RNAi in HEK293 cells suggested protein-arginine methylation maybe reduced in MTAP absence; though further confirmation is required. A genomic screen of 101 tumours and 32 healthy controls, including 10 germline matched whole exomes and 91 tumours undergoing targeted sequencing was used to probe for driver events. Novel putative drivers identified include: genes which maintain genome integrity; POT1 (14%) and BRCA2 (14%), master epigenetic regulators; ASXL3 (8%) and KDM6A (6%), and programmed cell death regulators; PDCD11 (35%) and TRPM3 (11%). In conclusion, putative driver events of CTCL have been identified using; (i) targeted analysis of a candidate region to identify selective pressure silencing MTAP; and (ii) a genomic sequencing approach to define a shortlist of genes.
Supervisor: Mitchell, Tracey ; Whittaker, Sean Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available