Medical Biology - Theses
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ItemStudies of the role of Mcl-1 in haemopoiesis and leukaemia( 2015)Cell death by apoptosis plays a critical role during embryonic development and in maintaining tissue homeostasis. Consequently, defective apoptosis can lead to degenerative diseases, autoimmunity and tumour development. In mammals, there are two converging apoptosis pathways: the ‘extrinsic’ pathway, which is triggered by engagement of cell surface ‘death receptors’ such as Fas; and the ‘intrinsic’ pathway, which is triggered by diverse cellular stresses, and is regulated by pro- and anti-apoptotic members of the Bcl-2 family of proteins. The principal focus of my studies is Mcl-1, an inhibitor of the intrinsic apoptosis pathway. Mcl-1 is overexpressed in a variety of cancers, including acute myeloid leukaemia (AML) where high levels of Mcl-1 are associated with poor prognosis and drug resistance. Using mouse genetic models, I have investigated the consequences of overexpression of Mcl-1 for haemopoiesis and autoimmunity (Part I) and for the development and treatment of AML (Part II). I. To determine the impact of simultaneously inhibiting the intrinsic apoptosis pathway via overexpression of Mcl-1 and the extrinsic apoptosis pathway via a non-functional Fas receptor, mcl-1 transgenic mice were crossed with faslpr/lpr mice. The combined mutations had little impact on myelopoiesis apart from an increase in macrophages, mainly in the spleen. All major lymphoid subsets were elevated, however, including the “unusual” T cells characteristic of faslpr/lpr mice. Furthermore, the onset of autoimmune disease was markedly accelerated. Thus, consistent with other genetic studies, the intrinsic and extrinsic apoptosis pathways synergise to control autoimmunity. II. To determine the impact of Mcl-1 in AML, I used a mouse model induced by retroviral expression of MLL-AF9, the fusion oncoprotein created by the t(9;11) translocation often found in childhood and treatment-induced adult AML. Overexpression of Mcl-1 or its pro-survival relative, BCL-2, increased the leukaemic burden in the spleen and blood of sick mice although it did not accelerate morbidity. AMLs overexpressing Mcl-1 or BCL-2 tended to have a higher proportion of mature cells compared to ‘wild type’ MLL-AF9 leukaemias. Unlike ‘wild type’ MLL-AF9 leukaemias, which were readily transplantable in non-irradiated recipients, most MLL-AF9 leukaemias overexpressing Mcl-1 and many overexpressing BCL-2 would only transplant if injected into lightly-irradiated recipients. Possible reasons for this unexpected result are discussed. In vitro experiments using short-term lines derived from primary tumours demonstrated that overexpression of Mcl-1 or BCL-2 in MLL-AF9 tumours increased resistance to standard drugs used to treat AML in the clinic. However, even those overexpressing Mcl-1 or BCL-2 were sensitive to the proteasome inhibitor, bortezomib, and to various CDK inhibitors as single agents. The addition of the BH3-mimetic ABT-737 enhanced the response of MLL-AF9 AMLs of all genotypes to standard therapeutics. In contrast, when added to bortezomib or CDK inhibitors, ABT-737 only enhanced the sensitivity of the AMLs that overexpressed BCL-2. Future studies will compare the efficacy of these drug regimens in vivo in transplanted syngeneic immuno-competent mice.