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Title: The genetics and kinetics of BCL2 driven lymphoid malignancies
Author: Webster, Philip
ISNI:       0000 0004 6348 0558
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2015
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Introduction: Non-Hodgkin Lymphoma (NHL) is rising in incidence. Treatment of this genetically heterogeneous disease has toxic side effects and significant numbers of relapsers / non-responders. BCL2, an anti-apoptotic protein, is commonly overexpressed in NHL as a result of the t(14;18) translocation. A number of BCL2 inhibitors have shown success in clinical trials but variable efficacy has meant that none have been licenced for use. Methods: Retroviral insertional mutagenesis (RIM), using Moloney Murine Leukaemia Virus (MoMuLV) in transgenic mice overexpressing BCL2, was used to identify putative target genes deregulated alongside BCL2 in lymphomagenesis. This project aimed to update MoMuLV integration site identification and sequencing, allowing quantification of integration site clonal abundance. Cohorts of mice were sacrificed at time points prior to disease onset in order to interrogate integration site kinetics. To test the oncogenic potential of candidate genes, C57BL/6 Vav-BCL2 p53+/- mouse B cells were retrovirally transduced with genes of interest and transplanted into mice to study the speed of lymphoma onset. Results & Conclusions: A novel, high throughput, quantitative library preparation and sequencing protocol compatible with an Illumina platform was validated. RIM screening in BCL2 transgenic and wild-type mice identified different insertion sites profiles, detecting known oncogenes and tumour suppressor genes as well as novel candidate genes involved in pathways of lymphoid organ development, B-cell activation and differentiation. Study of insertion kinetics over time showed three patterns of clonal abundance and also allowed the study of specific gene deregulation prior to disease onset. Overexpression of Cd86 slowed disease onset whilst Ildr1 expedited disease onset suggesting the former is a tumour suppressor gene and the latter an oncogene. Discovering genes mutated with BCL2 in lymphoma may help to explain the lack of efficacy of BCL2 inhibitors and also identify novel therapeutic targets.
Supervisor: Uren, Anthony Sponsor: Medical Research Council
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral