Targeting dormancy in acute myeloid leukaemia

Doolan, Lara (2023) Targeting dormancy in acute myeloid leukaemia. PhD thesis, University of Nottingham.

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Acute myeloid leukaemia (AML) represents the poorest survival rate of all leukaemias, and the majority of patients who do achieve remission relapse within 3 years. Relapse is attributed to the presence of quiescent leukaemia initiating cells (LICs) which evade standard-of-care chemotherapy that targets the proliferative bulk of the disease. Novel therapeutics which are able to target dormant LICs are therefore required to improve patient outcomes in AML. Previous work in our group generated a model for dormancy in AML using the CD34+CD38- cell line TF1a, treated with TGFβ1 and rapamycin, to enrich for dormancy. Microarray analysis and gene expression connectivity mapping were used to identify a list of FDA-approved drugs predicted to target dormancy enriched TF1a cells (TF1a-DE). In this thesis, the top 8 hits from this analysis were screened to assess their efficacy in targeting TF1a-DE cells. This identified pantoprazole, a proton pump inhibitor, as able to inhibit colony formation and chemo-sensitise TF1a-DE to AraC.

RNA-sequencing was utilised to expand our molecular signature for dormancy. Firstly, TF1a-DE cells were subjected to RNA-sequencing which identified 795 upregulated genes and 444 genes downregulated genes. Kaplan-Meier analysis and Cox multivariate survival analysis of differentially expressed genes identified in this model identified PLXNC1, SPATS2L and ELN as being associated with overall survival, independent of known risk factors age, cytogenetic risk group, and white blood cell count (WBC).

Finally, our group has previously developed a 12-day AML co-culture in which primary AML cells are stained with permanent fluorescent dye and co-cultured with MS-5 murine stromal cell line and sorted based on fluorescence intensity into dormant and cycled fractions. RNA-sequencing and differential expression analysis was performed on dormant and cycled primary AML cells to generate a gene signature of 523 upregulated genes, and 162 down regulated genes. Kaplan-Meier analysis and Cox multivariate analysis which included risk group, age and WBC identified MMP7 and CLNK as being independently associated with overall survival. 17 genes were commonly upregulated, and 5 genes downregulated in both TF1a-DE and dormant AML cells. ELN was identified as an upregulated gene in both models for dormancy. In summary, this thesis has characterised dormant AML cells through gene expression analysis of TF1a-DE cells and dormant primary AML cells, and shown the ability of pantoprazole to chemo-sensitise TF1a-DE cells to AraC and inhibit colony formation.

Item Type: Thesis (University of Nottingham only) (PhD)
Supervisors: Seedhouse, Claire
Thompson, Alex
Keywords: Acute myeloid leukaemia; Leukaemia initiating cells; TF1a-DE cells; RNA-sequencing; AraC
Subjects: W Medicine and related subjects (NLM Classification) > WH Hemic and lymphatic system
Faculties/Schools: UK Campuses > Faculty of Medicine and Health Sciences > School of Medicine
Item ID: 76442
Depositing User: Doolan, Lara
Date Deposited: 13 Dec 2023 04:40
Last Modified: 13 Dec 2023 04:40

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