Mitchell, Rachel; (2021) IKZF1 lesions in adult acute lymphoblastic leukaemia - the chicken or the egg? Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

The IKZF1 gene encodes the transcription factor IKAROS, a master regulator of lymphocyte differentiation. IKZF1 is commonly deleted in acute lymphoblastic leukaemia (ALL), particularly in BCR-ABL1+ ALL. Lesions in IKZF1 carry the hallmarks of RAG mediated deletion, the expression of which is controlled by IKAROS. The title of this thesis, “IKZF1 lesions… the chicken or the egg?”, refers to the unknown sequence of events which leads up to the deletion of IKZF1 since RAG expression dependent on IKAROS and therefore could not be deleted without RAG activity. The most common intragenic deletion involves the removal of the DNA binding domain in exons four to seven (Δ4-7), and produces a dominant negative (DN) which inhibits wild-type IKAROS function via dimerisation. Other focal deletions involving exons two or eight cause loss of function (LOF). Firstly, to ascertain if IKZF1 deletions (ΔIKZF1) had an impact on outcome, I screened 498 patients with B-ALL by end-point PCR for the four most common deletions: Δ4-7, Δ2-7 Δ4-8 and Δ2-8. IKZF1 lesions had no impact on outcome, despite minimal residual disease (MRD) positivity being more likely after induction therapy. A comparison of PCR and MLPA revealed a large number of discrepancies, highlighting that neither technique alone is sufficient for ΔIKZF1 detection. Next, I compared Ig/TCR and BCR-ABL1 based MRD to see if one technique was better at predicting survival. Several specimens had significantly different MRD levels, though neither of technique was proven better at predicting outcome. I also show that the type and number of Ig/TCR rearrangements differ significantly based on IKZF1 status, and that IKZF1 breakpoint sequences differ significantly based on BCR-ABL1 status, suggesting altered RAG activity. Finally, I established several cell line models in order to observe the effects of DN and LOF IKZF1 lesions on drug sensitivity. I showed that NALM-6 cells with LOF or DN IKZF1 lesions have increased resistance to dexamethasone, and that the presence of mesenchymal stromal cells in co-culture enhanced survival of ALL cells regardless of IKZF1 and BCR-ABL1 status. The data in this thesis has shown that while there is no impact of ΔIKZF1 on outcome in patients with ALL, it is likely that its impact is dependent on other cooperating lesions when these results are considered within the wider context of other trial data. Despite no impact seen in the clinical data, the cell line experiments have confirmed that ΔIKZF1 contributes to a more aggressive phenotype although the cause of this remains unclear.

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