Durg Resistance Mediated by Overexpression of MRPI in an Inducible System
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Authors
Pelz, Kirsten
Issue Date
2000
Type
Thesis
Language
en_US
Keywords
Alternative Title
Abstract
The superfamily of A TP-binding cassette transport proteins known as multidrug
resistance proteins (MRPs) are important to cancer therapy due to their function in
conferring tumor cells with resistance to multiple drugs. MRP 1 is one of six members in
this family that has been identified previously and has been strongly implicated in the
reduction of tumor cell sensitivity to chemotherapeutic drugs. Because of its location in
the membranes of many tumor cell lines, the MRPI transport protein plays a crucial role
in the uptake and efflux of chemotherapuetic drugs. In this study, we have utilized K562 acute myeloid leukemia (AML) cells transfected with MRP 1 cDNA in an inducible expression system. By introducing a plasmid containing MRPI under control of a tetracycline-response promotor into a K562 , cell line, it is possible to induce the expression of MRP 1 through the addition of doxycycline (Dox), a derivative of tetracycline. The response protein functions as a control switch for MRP 1 expression by "turning on" transcription of the protein when Dox is added to the cell media. Drug resistance observed in the transfected cell line can then be attributed solely to MRP 1 overexpression due to the fact that the presence of Dox is necessary for MRPI expression to occur. Following demonstration of MRP 1 overexpression in transfected cells induced with Dox, both induced and uninduced cells were treated with several chemotherapeutic agent. Using cytotoxicity assays, we determined that transfectants overexpressing MRPI displayed a reduction in sensitivity to vincristine and daunorubicin during long-term exposure (72 hours) and to methotrexate during short-term (4 hour) exposure. Increased resistance to the antifolate drug, ZD1694, was not observed during either long- or shortterm exposure.
As expected, net uptake of radio labelled methotrexate (eH]MTX ) in the transfected K562 cell line was greater for uninduced cells while the efflux rate for eH]MTX was slightly faster in induced cells (i.e. cells overexpressing the MRPI transporter) due to the fact that MRPI has the ability to pump drug out of the cells. Thus, using a tightly controlled system of expression, we have strong evidence to support the role of MRP I-mediated drug resistance in tumor cell lines. This may be important in
regard to providing effective treatments for future cancer patients.
Description
iv, 28 p.
Citation
Publisher
Kalamazoo College
License
U.S. copyright laws protect this material. Commercial use or distribution of this material is not permitted without prior written permission of the copyright holder.