Multidrug resistance (MDR) and existing cancer stem cells (CSCs) are two major problems in successful treatment of cancers. Expression of breast cancer resistance protein (BCRP) is known to associate with both MDR and CSCs, suggesting that BCRP is an interesting and ideal target for development of chemosensitizing agents for better treatment of drug resistant cancers and elimination of CSCs. Recently, we identified a novel BCRP inhibitor PZ39 and its analogue compound PZ39C8 which are capable of inhibiting BCRP function and inducing BCRP degradation in lysosome. In the present study, we (1) investigated the mechanism of inhibitor-induced BCRP degradation using a stable cell line expressing GFP-BCRP in combination with pharmacological agents of endocytosis and trafficking and (2) determined the inhibitor binding site in BCRP using compound-conjugated Sepharose 4B beads and pull-down assay. We found that (1) the lysosomal inhibitor Bafilomycin A but not the proteosome inhibitor MG132 abolished PZ39C8-induced BCRP degradation; (2) dynasore, a potent endocytosis inhibitor, and 3-Methyladenine, a autophagy inhibitor, both efficiently blocked the PZ39C8-induced BCRP degradation from plasma and internal membranes; (3) following co-treatment with PZ-39C8 and endocytosis, trafficking, and lysosome inhibitors, BCRP co-localized with the LAMP-1, LC3II and Clathrin, EEA1, the markers of Lysosome, autophagy and endocytic pathways, respectively; and (4) PZ39C8 directly binds BCRP and the binding site/s appears to locate in the domain including TM5-loop-TM6. Taken together, we conclude that PZ39C8 binds to TM5-ECL3-TM6 of BCRP and induces BCRP degradation in lysosome via endocytic and autophagic dependent pathways. Thus, PZ39C8 is potentially a valuable probe for structure-function studies of BCRP and a lead compound for developing therapeutics targeting BCRP-mediated MDR in combinational cancer chemotherapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 739. doi:10.1158/1538-7445.AM2011-739