B cell receptor induced autophagy and endocytosis in Chronic Lymphocytic Leukaemia

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Skills Priority Alignment: Advanced TherapeuticsChronic Lymphocytic Leukaemia (CLL) is the most prevalent adult leukaemia in Europe and the United States and is currently incurable. CLL is largely split into two subgroups, as indolent (M-CLL) and an aggressive (U-CLL) disease characterised by their IGHV mutational status. The disease is driven through activation of the B cell receptor (BCR) and inhibitors that target this pathway are revolutionising this disease, but these agents are taken daily, not curative and resistance to these drugs is already emerging. Therefore novel strategies or salvage therapies to treat this disease are urgently required.Consequently, a better understanding of the BCR signalling pathways is essential if we are to identify novel therapeutic targets. The Steele group has demonstrated that basal expression of markers such as LC3BII and GABARAPL2 which are associated with autophagy, a process known to promote tumorigenesis, is elevated in CLL samples compared to healthy donor (HD) cells (p=.009 & p=.013 respectively). Moreover, LC3BII expression is greater still in U-CLL samples compared to M-CLL (p=.02). It has been demonstrated the novel finding that activating CLL cells using bead immobilised (BI) or soluble anti-IgM could further augment LC3BII expression in a time dependent manner and this correlated with the cells ability to signal via the BCR. Moreover, the BCR-induced increase in LC3BII expression was greater in U-CLL samples compared to M-CLL (p=.004) or HD (p=.023) cells. CLL cells incubated with BI anti-IgM for 18-24h were also able to take up the beads (2.8 micrometre) coated with anti- IgM, but not the isotype control, by an unknown process. The size of these beads suggests that clathrin-mediated endocytosis was unlikely. However, a mechanism called LC3-associated phagocytosis (LAP) is known to plays a role in the uptake of extracellular pathogens that engage receptor mediated signalling pathway such as FcR but has not been described for sIgM. The LAP process is distinct from autophagy but still requires LC3BII to occur. Both autophagy and LAP are associated with antigen processing and presentation by the MHCII receptor, whereas autophagy alone is also involved in antigen processing for presentation by the MHCI receptor. Therefore, the aims of this project are to 1.) Determine which signalling pathways are involved in BCR-induced autophagy and LAP following treatment with soluble and BI-anti-IgM, 2.) Determine how autophagy and LAP are involved in antigen presentation to MHCII in CLL and normal B cells and 3.) Determine how these pathways differ between HD and CLL cells so that we can target these pathways therapeutically.This PhD studentship will bring together expertise from several key academics and link established research strengths in B-cell malignancies, cell signalling, novel therapeutics and imaging technologies between medicine (Dr Steele, Prof Cragg and Dr James), biosciences (Dr Tumbarello) and Defence science and technology laboratories (DSTL, Porton Salisbury, Dr Caroline Rowland) and the Crick (Dr Tooze). This project will improved our understanding of how autophagy and LC3 associated phagocytosis (LAP) pathways regulate CLL pathogenesis, leading to the identification of novel therapies or treatment strategies for patients.