The University of Hull
Development of microfluidic devices to study drugs effects and develop novel targets against circulating and bone-marrow derived Chronic Lymphocytic Leukaemia
Supervisors:
Dr John Greenman - Division of Cancer, Postgraduate Medical Institute
Dr David Allsup - Hull & East Yorkshire Hospital NHS Trust
Professor Steve Haswell - Dept of Chemistry, Faculty of Science
This project aims to fabricate and optimise microfluidic devices for studying circulating leukaemia cells and those found in the bone-marrow. The work will focus on Chronic Lymphocytic Leukaemia (CLL), the commonest form of leukaemia in the Western World. The devices will provide a way of comparing how circulating cells or small bone-marrow samples respond when treated with chemotherapy drugs. In addition, the device will be used to study the expression and functionality of plasma-membrane-associated adenosine triphosphate (ATP) to assess the potential of this molecule as a prognostic marker and/or target for therapeutic intervention using mass spectrometry. Ultimately it is anticipated that this device will aid in diagnosis and treatment of CLL; and is equally applicable to many other leukaemias and lymphomas.
Microfluidic-based cell handling systems. These have recently demonstrated the enormous advantage that such devices offer in mimicking the in vivo environment; offering considerable potential to study tissue ex vivo whilst retaining the true in vivo integrity. The principle aim of the current work is to develop microfluidic systems that can be used to test the responsiveness of human tissue, both normal and diseased, to the effects of drugs, nutrients or injury. This approach differs fundamentally from current techniques that use either isolated cells or whole animal models by allowing us to study the responses of human tissue, which represent more closely the natural, or pathological, state. We believe that this technology has the potential to revolutionise approaches for identifying new therapeutic drugs in diseases such as cancer, and in understanding the biology and regenerative capacity of the heart. Such insights will in turn offer new strategies for clinical management of disease states.
Plan of investigation One of the major problems with treating CLL is targeting the tumour “hidden away” in the bone marrow, rather than in the circulation. We propose to use bespoke microfluidic devices to compare drug effects against circulating CLL cells and CLL bone-marrow. Initial experiments will establish the unique parameters required to maintain CLL bone-marrow in the microfluidic chambers, using chips optimised for similar-sized biopsies from liver tissue. Conditions such as media flow, media supplementation, O2 and CO2 concentrations will all be optimised. Once these parameters are determined, parallel experiments will be undertaken using CLL bone marrow tissue biopsies and circulating CLL from the same patient to test the activity of a selection of commonly used chemotherapy drugs (fludarabine, dexamethasone, cyclophosphamide). The assays employed will include: simple live:dead staining, MTS proliferation, Annexin V-FITC binding and Lactate Dehydrogenase (LDH) release.
Dr John Greenman - Division of Cancer, Postgraduate Medical Institute
Dr David Allsup - Hull & East Yorkshire Hospital NHS Trust
Professor Steve Haswell - Dept of Chemistry, Faculty of Science
Purposes of proposed investigation
This project aims to fabricate and optimise microfluidic devices for studying circulating leukaemia cells and those found in the bone-marrow. The work will focus on Chronic Lymphocytic Leukaemia (CLL), the commonest form of leukaemia in the Western World. The devices will provide a way of comparing how circulating cells or small bone-marrow samples respond when treated with chemotherapy drugs. In addition, the device will be used to study the expression and functionality of plasma-membrane-associated adenosine triphosphate (ATP) to assess the potential of this molecule as a prognostic marker and/or target for therapeutic intervention using mass spectrometry. Ultimately it is anticipated that this device will aid in diagnosis and treatment of CLL; and is equally applicable to many other leukaemias and lymphomas.
Background CLL is the commonest leukaemia in the Western world, which in 50% of case is indolent and non-progressive, however the remaining patients suffer from a progressive disease that is ultimately fatal. It is characterised by a proliferation of malignant lymphocytes in the peripheral blood, marrow infiltration with resultant bone marrow failure, lymphadenopathy and hepatosplenomegaly. Patients usually die from infections consequential upon an immunosuppressed state induced by a combination of disease and the consequences of therapy. Numerous chemotherapeutic agents have activity in CLL with many inducing prolonged remissions, although none prolong overall survival. The majority of patients requiring treatment will eventually experience relapse of their disease and there is therefore a need for new and novel therapeutic agents.
Microfluidic-based cell handling systems. These have recently demonstrated the enormous advantage that such devices offer in mimicking the in vivo environment; offering considerable potential to study tissue ex vivo whilst retaining the true in vivo integrity. The principle aim of the current work is to develop microfluidic systems that can be used to test the responsiveness of human tissue, both normal and diseased, to the effects of drugs, nutrients or injury. This approach differs fundamentally from current techniques that use either isolated cells or whole animal models by allowing us to study the responses of human tissue, which represent more closely the natural, or pathological, state. We believe that this technology has the potential to revolutionise approaches for identifying new therapeutic drugs in diseases such as cancer, and in understanding the biology and regenerative capacity of the heart. Such insights will in turn offer new strategies for clinical management of disease states.
Plan of investigation One of the major problems with treating CLL is targeting the tumour “hidden away” in the bone marrow, rather than in the circulation. We propose to use bespoke microfluidic devices to compare drug effects against circulating CLL cells and CLL bone-marrow. Initial experiments will establish the unique parameters required to maintain CLL bone-marrow in the microfluidic chambers, using chips optimised for similar-sized biopsies from liver tissue. Conditions such as media flow, media supplementation, O2 and CO2 concentrations will all be optimised. Once these parameters are determined, parallel experiments will be undertaken using CLL bone marrow tissue biopsies and circulating CLL from the same patient to test the activity of a selection of commonly used chemotherapy drugs (fludarabine, dexamethasone, cyclophosphamide). The assays employed will include: simple live:dead staining, MTS proliferation, Annexin V-FITC binding and Lactate Dehydrogenase (LDH) release.