The Interaction of Bortezomib with P-Gp, MRP-1 and BCRP Drug Transporters: Implications for Therapeutic Applications of Bortezomib in Advanced Multiple Myeloma and Other Neoplasias.

Multidrug transporters are energy-dependent transmembrane proteins which can efflux a broad range of anticancer drugs and thereby play a role in resistance to the actions of substrate agents. Classically, three transporters, p-glycoprotein (Pgp; MDR-1; ABCB1), multidrug resistant protein-1 (MRP-1; ABCC1) and breast cancer resistance protein (BCRP; MXR; ABCG2), have been found to have the broadest substrate specificity and a strong correlation with drug resistance in vitro and in vivo in many models and forms of cancer. We have sought to characterize the interaction of bortezomib with these transporters and thereby explore the potential for these agents to play a role in resistance. Bortezomib is a novel proteosome inhibitor with significant activity in multiple myeloma, although subsets of patients remain refractory to the activity of the drug. Hence, better characterization of the interactions of this drug with classical resistance mechanisms may identify improved treatment applications.

We investigated the role of these transporters by using isogenic cell line models which are resistant due to overexpression of a particular transporter: DLKP lung cancer cell line that overexpresses MRP-1; DLKP-A which overexpresses Pgp; and DLKP-SQ-Mitox which overexpresses BCRP. DLKP-A cells exhibited a 4.6-fold decrease in responsiveness to bortezomib compared to parental DLKP cells. In DLKP-SQ-Mitox, bortezomib-induced cytotoxicity was comparable to DLKP. When bortezomib was combined with elacridar, a Pgp and BCRP inhibitor, significant synergy was evident in DLKP-A (100% viable cells with single agent treatment versus 11% with the combination), but not DLKP-SQ-Mitox. Sulindac, an MRP-1 inhibitor, combined with bortezomib failed to produce any synergy in MRP-1 positive DLKP cells. Conversely, combination assays of Pgp substrate cytotoxics such as doxorubicin with Bortezomib were largely additive in nature. This indicates that bortezomib has little, if any, direct Pgp inhibitory activity, as combinations of a traditional Pgp inhibitor (such as elacridar) and doxorubicin would show marked synergy rather than just an additive effect in Pgp positive cells. To further characterize the extent of this interaction with Pgp, we conducted cytotoxicity assays in cell lines with varying levels of Pgp overexpression. NCI/Adr-res (ovarian cancer, high Pgp overexpression), RPMI-Dox40 (multiple myeloma, moderate Pgp overexpression) and A549-taxol (lung cancer, low Pgp overexpression). The combination of bortezomib and elacridar that produced the most synergy was in cell lines expressing moderate to high levels of Pgp expression. Cell lines with lower Pgp expression produced an additive cytotoxicity. We next examined whether bortezomib had any direct effect on Pgp expression. In RPMI-Dox40 cells, Pgp expression is reduced in a time-dependent manner with bortezomib treatment.

Our studies therefore show that bortezomib is a substrate for Pgp but not the other drug efflux pumps. In tumor cells expressing high levels of Pgp, the efficacy of bortezomib is synergistically enhanced by combinations with a Pgp inhibitor, while bortezomib treatment itself can reduce the expression of Pgp. This study suggests that in the subset of patients with advanced multiple myeloma or solid tumors which express high levels of Pgp, inhibition of its function could contribute to enhanced responsiveness to bortezomib.

Richardson:millenium: Membership on an entity's Board of Directors or advisory committees, Research Funding; celgene: Membership on an entity's Board of Directors or advisory committees, speakers bureau up to 7/1/09; MLNM: speakers bureau up to 7/1/09. Mitsiades:Millennium Pharmaceuticals : Consultancy, Honoraria; Novartis Pharmaceuticals : Consultancy, Honoraria; Bristol-Myers Squibb : Consultancy, Honoraria; Merck &Co: Consultancy, Honoraria; Kosan Pharmaceuticals : Consultancy, Honoraria; Pharmion: Consultancy, Honoraria; PharmaMar: licensing royalties ; Amgen Pharmaceuticals: Research Funding; AVEO Pharma: Research Funding; EMD Serono : Research Funding; Sunesis Pharmaceuticals: Research Funding. Anderson:Celgene: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Millennium: Consultancy, Research Funding; Biotest AG: Consultancy, Research Funding.