Cancer cells develop resistance to a number of cytotoxic drugs. The project will utilise novel approaches to examine how the drug efflux pump P-glycoprotein confers resistance
Chemotherapy is used in several settings for the therapy or management of cancer. Unfortunately, the success of chemotherapy is limited in many forms of cancer; which display an inherent resistance. However, many cancer types display an initial response to chemotherapy, only to acquire a resistant phenotype during treatment. Resistance to multiple anti-cancer drugs is a multi-factorial cellular response and thought to be responsible for treatment failure. The presence of a drug efflux pump (i.e. P-glycoprotein) with broad substrate specificity on cancer cells is one of the key mediators of drug resistance. P-glycoprotein (P-gp) prevents sufficient accumulation of cytotoxic drugs within cancer cells; thereby preventing cell death. P-gp has been the subject of intensive investigation for over 30 years and to date there are no successful strategies to combat its ability to confer drug resistance in cancer.
P-gp is an ATP driven drug efflux pump and the ability to interact with over 200 known chemicals remains a biological enigma. Similarly, many aspects of its mechanism of action remain elusive. One of the main research goals of the Callaghan Laboratory is to describe the molecular mechanism of this complex drug resistance mediator.
A key aspect of delivering the above objective is to utilise molecular based techniques using purified P-gp. Purified P-gp must be reconstituted into lipid vesicles, which may be used to measure generate transport activity. The drug transport activity of P-gp may also be dissected into the components of drug binding and ATP hydrolysis. Each of the assays (transport, binding and hydrolysis) provides distinct information about the overall process. The project will have two major objectives:
- To reconstitute purified P-gp into nano-disc structures to enable measurement of binding properties of anti-cancer drugs. The nano-disc structures do not have a vesicular lumen and are “non-sided”; thereby facilitating direct measurement of drug binding and ATP hydrolysis.
- To reconstitute P-gp into giant unilamellar vesicles (GUVs) to enable measurement of drug transport activity. Conventional liposomes (d~100nm) are notoriously difficult to utilise in the measurement of transport for hydrophobic drugs. The use of GUVs (d~1µm) alleviates the problems of bilayer tension and high drug accumulation that interfere with transport studies in conventional vesicles.
The project will suit students with an interest in cancer drug resistance, membrane transport processes and/or molecular studies with proteins. Training will be provided in membrane protein purification, chromatography and reconstitution into a variety of lipid systems. Assay systems will involve fluorescence spectroscopy, radioligand binding assays and advanced fluorescence microscopy. Further details may be obtained from Associate Professor Callaghan.