Guthridge, Mark A.Lopez, Angel FranciscoThomas, Daniel2015-10-292015-10-292012http://hdl.handle.net/2440/95873Acute myeloid leukaemia (AML) is a clonal disease of myelopoiesis characterised by poor survival due to frequent relapse. The cytokines interleukin-3 (IL-3) and granulocyte macrophage-colony stimulating factor (GM-CSF) are soluble glycoproteins that bind cell surface receptors on haemopoietic cells to stimulate their biological effects including cell survival and proliferation. Cancers cells usurp cytokine signalling pathways via various mechanisms to evade apoptosis and deregulate proliferation, two fundamental hallmarks of the cancer phenotype. Within a given patient’s leukaemia there are smaller sub-populations of cells that possess the unique ability to selfrenew and initiate tumour formation, termed “leukaemia stem cells” (LSCs). LSCs possess robust survival properties, are resistant to chemotherapy and over-express the IL-3 receptor α chain, however therapies directed specifically against LSCs are currently lacking. This body of work examines the cytokine receptor signalling events transduced by the IL-3/GM-CSF receptor active in acute myeloid leukaemia blasts and LSCs with the aim of discovering novel avenues for therapy. In this thesis four targets for AML therapy are investigated and developed:- (1) Firstly, we demonstrate feasibility of LSC targeting as a therapeutic strategy by demonstrating efficacy in vivo and in vitro of a blocking monoclonal antibody 7G3 directed to the extracellular domain of the IL-3 receptor CD123 which is over-expressed on AML stem cells. (2) Secondly, we show that in the majority of AML patients, a distinct serine in the IL-3 receptor beta chain is constitutively phosphorylated and is required for cytokine-mediated survival. In a search for druggable kinases that may be responsible for this phosphorylation we isolated phosphatidyl inositol 3-kinase with novel activity as a protein kinase and show that it is linked to cell survival. (3) We demonstrate that osteopontin (OPN) is a functionally relevant gene regulated by the serine-survival pathway that mediates survival of CD34⁺CD38⁻CD123⁺ leukemic stem/progenitors. Furthermore, increased expression of OPN is associated with poor prognosis in normal karyotype AML. (4) Finally, we identified a small molecule kinase inhibitor PIK-75 that is capable of the dual targetting of both PI3K and CDK9 in AML in vivo and in vitro. It is suggested that the simultaneous targeting of different molecules converging on a cell survival pathway represents a novel therapeutic avenue.leukaemia; cytokine receptor; targeted therapy; survival; signal transduction; kinaseThesis20120619173722