Functional genomic screening to unravel mechanisms underlying resistance to conventional induction therapy in T-cell acute lymphoblastic leukaemia

Abstract

Long term cure is achieved in more than 80% of children with T-cell acute lymphoblastic leukaemia (T-ALL), which can be attributed to the refinement of intensive, MRD driven, chemotherapy regimens. However, poor response to induction therapy is associated with unfavourable outcomes. Vincristine, Dexamethasone, Asparaginase and Daunorubicin (VXLD) form the cornerstone of T-ALL induction therapy. Identifying mechanism of resistance to induction chemotherapy could help us improve the efficacy of treatment and outcomes for patients with poor response to current therapy. This project aimed to develop a preclinical model which allowed the identification of pathways underlying drug resistance to T-ALL induction therapy and inform the development of a more targeted approach to reduce disease burden in refractory disease. This thesis describes the optimisation of a 4 drug treatment regimen in Rag2-/- gamma c-/- mice that mirrors the clinical induction protocol. This preclinical in vivo model has utility both as a model to explore drug resistance as well as a benchmark to assess efficacy and toxicity of new compounds or drug combinations. In this work the established in vivo model of induction therapy was combined with CRISPR screening to identify mechanisms of chemo-resistance. The MAGeCKFlute algorithm was used to identify differential representation of CRISPR guide RNAs under VXLD treatment pressure, and identified several interconnecting pathways in the generation of chemo-resistance. The role of the apoptotic regulator BCL-X was further investigated, as gene specific guides were depleted (suggestive of drug target). Targeted inhibition of BCL-X has already entered the clinical arena, and may provide a means to overcome induction resistance. The successful development of this preclinical model incorporating VXLD chemotherapy has increased our understanding of T-ALL biology and identified potential drug resistance mechanisms. Further work will focus on the validation of these putative drug targets which may revert induction failure in future therapeutic approaches.