Multidrug resistance (MDR)-associated transporters contribute to the persistence of many types of cancers during chemotherapy. Three decades of research to inhibit MDR-associated transporters have, so far, produced only limited benefits for cancer treatment. This proposal aims to develop functional assays that will make it possible to characterize better, and hence target effectively, MDR-associated transporters from cancer cells. This research will proceed in three steps. First, it will develop chip-based assay platforms for functional biophysical characterization of multidrug resistance-associated transporters. These assays will be combined with electrophysiology and single molecule fluorescence detection. Second, it will investigate, under well-controlled conditions, the effect of parameters that affect the transport rate, such as the transmembrane potential, the lipid composition, the concentration of ATP and co-transported molecules (e.g. glutathione) to understand better the function of MDR-associated transporters and possible ways to modulate this function. And third, it will develop assays that can monitor the transport rate of individual reconstituted MDR-associated transport proteins in planar lipid bilayers by taking advantage of extremely small volumes (picoliters) in microfabricated recording chips and by combining electrophysiology with single molecule optical detection. We will use these assays to characterize MDR-associated transporters on the single-protein-level in analogy to single ion channel patch clamp recordings. Finally we aim to investigate the activation state and the effect of activation (e.g. by phosphorylation) on the single transporter efflux rates. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM081705-02
Application #
7470036
Study Section
Enabling Bioanalytical and Biophysical Technologies Study Section (EBT)
Program Officer
Chin, Jean
Project Start
2007-08-01
Project End
2012-07-31
Budget Start
2008-08-01
Budget End
2009-07-31
Support Year
2
Fiscal Year
2008
Total Cost
$275,931
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Biomedical Engineering
Type
Schools of Engineering
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Yusko, Erik C; Bruhn, Brandon R; Eggenberger, Olivia M et al. (2017) Real-time shape approximation and fingerprinting of single proteins using a nanopore. Nat Nanotechnol 12:360-367
Horger, Kim S; Liu, Haiyan; Rao, Divya K et al. (2015) Hydrogel-assisted functional reconstitution of human P-glycoprotein (ABCB1) in giant liposomes. Biochim Biophys Acta 1848:643-53
Rao, Divya K; Liu, Haiyan; Ambudkar, Suresh V et al. (2014) A combination of curcumin with either gramicidin or ouabain selectively kills cells that express the multidrug resistance-linked ABCG2 transporter. J Biol Chem 289:31397-410
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Majd, Sheereen; Yusko, Erik C; MacBriar, Alexander D et al. (2009) Gramicidin pores report the activity of membrane-active enzymes. J Am Chem Soc 131:16119-26

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