Retrovirus-Mediated Gene Transfer of the Multidrug Resistance-Associated Protein (MRP) cDNA Protects Cells from Chemotherapeutic Agents

Abstract

Transduction of hematopoietic progenitors with a multidrug resistance gene like mdr-1 or mrp aims to protect bone marrow from toxicity of chemotherapeutic agents. The interest in the use of mrp as an alternative to mdr-1 gene transfer for bone marrow protection lies in its different modulation. Indeed, classical P-gp reversal agents, tested in the clinic to decrease mdr-1 tumor resistance, have little or no effect on MRP function. This would allow, in the same patient, the use of reversal agents to decrease P-gp tumor resistance without reversing bone marrow protection of the transduced hematopoietic cells provided by multidrug resistance-associated protein (MRP). As a first step, we have constructed and tested two different mrp-containing vectors with either the Harvey retroviral long terminal repeat (LTR) or PGK as promoters and generated ecotropic producer cells. We have shown by Southern blot analysis that retroviral supernatant from these producer cells can efficiently transmit the mrp gene to target cells. Mrp expression could be detected by fluorescence-activated cell sorting (FACS) analysis in the producer cells. The transduced cells have increased resistance to doxorubicin, vincristine, and etoposide. Furthermore, chemoprotection of the transduced cells was increased after selection with chemotherapeutic agents in the presence of glutathione, a co-factor for MRP function. These data indicate that mrp retroviral vectors may be useful for chemoprotection and selection. Retroviral vectors containing the mrp cDNA have been constructed with long terminal repeat (LTR) and PGK promoters. They allow for a stable and efficient gene transfer in mrp-transduced fibroblasts, express the Mrp protein, and are chemoresistant to vincristine, doxorubicin, and etoposide. Selection of the transduced cells in chemotherapeutic media is possible. Those vectors are interesting for chemoprotection and for selection.