Crosstalk between EZH2 and BCL6 in regulating chronic myeloid leukaemia (CML) stem cell survival: from epigenomics to novel therapeutic approaches

Bugler, Jane (2019) Crosstalk between EZH2 and BCL6 in regulating chronic myeloid leukaemia (CML) stem cell survival: from epigenomics to novel therapeutic approaches. PhD thesis, University of Glasgow.

Abstract

Chronic Myeloid Leukaemia (CML) is a haematopoietic stem cell disorder characterised by the presence of the BCR-ABL1 fusion gene, which produces a constitutively active tyrosine kinase responsible for disease progression. Despite the introduction of tyrosine kinase inhibitors (TKIs) to target this overactive tyrosine kinase, the low cure rate and risk of disease recurrence and progression remains a major clinical issue. This is thought to occur due to a pool of TKIpersistent leukaemia stem cells (LSCs). Thus, understanding how LSCs evade TKI therapy and maintain their survival is important to identify novel drug targets and eradicate residual disease. Evidence suggests that the epigenetic regulator EZH2, is required for LSC survival. A combination of EZH2 inhibitors and TKIs targeted LSCs in vitro and in vivo, and resulted in the re-activation of several targets involved in cell cycle and apoptosis, including those normally regulated by the transcription factor BCL6. BCL6 is a survival factor in CML cells upregulated following TKI treatment, driving an anti-apoptotic phenotype. The hypothesis of this project was that BCL6 and EZH2 work through similar survival pathways that underpin an important network in CML which is therapeutically targetable. Genetic knockdowns of BCL6 and EZH2 in CML cell lines ± TKI significantly decreased cell expansion and increased apoptosis, and dual targeting of BCL6 and EZH2 resulted in additional effects on cell expansion. CML and non-CML CD34+ primary cells were treated with varying concentrations of a BCL6 inhibitor ± TKI or EZH2i, significantly decreasing colony forming cell (CFC) and long-term cultureinitiating cell (LTC-IC) outputs, suggesting targeting of more primitive CML cells. To further examine this relationship, chromatin immunoprecipitation (ChIP) was performed in CML cell lines and CD34+ cells. Whilst BCL6 and EZH2 targets were predominantly mutually exclusive, sets of their targets were enriched in many of the same downstream pathways. Moreover, a coordinated pattern of occupancy was observed when BCL6 and EZH2 shared targets. RNA-sequencing revealed strong enrichment for c-MYC and differentiation gene signatures following targeting of BCL6 and EZH2. Integrating these pathways with ChIP-seq data generated a network map highlighting the importance of c-MYC in the BCL6 and EZH2 network, as well as identifying putative targets mediating LSC survival that require further investigation as potential novel therapies in CML.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Keywords: chronic myeloid leukaemia, CML, leukaemia stem cells, epigenetics.
• Subjects: R Medicine > RZ Other systems of medicine
• Colleges/Schools: College of Medical Veterinary and Life Sciences > School of Cancer Sciences
• Supervisor's Name: Vetrie, Prof. David
• Date of Award: 2019
• Embargo Date: 6 November 2023
• Depositing User: Ms Jane Bugler
• Unique ID: glathesis:2019-75174
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 08 Nov 2019 13:16
• Last Modified: 06 Apr 2023 10:32
• Thesis DOI: 10.5525/gla.thesis.75174
• URI: https://theses.gla.ac.uk/id/eprint/75174