Bicinchoninic Acid Protein Assay in the Determination of Adriamycin Cytotoxicity Modulated by the MDR Glycoprotein

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Abstract
The development of simultaneous resistance to structurally unrelated drugs in cancer cells is a major obstacle to effective cancer chemotherapy. This multidrug-resistance (MDR) phenomenon is largely attributed to overexpression of a 170 kD glycoprotein, which serves as a transmembrane efflux pump in extruding a variety of natural anticancer drugs such as vinblastine, doxorubicin, and taxol from cancer cells. It is desirable, therefore, to discover compounds that can block the efflux mechanism and thus reverse drug resistance. The bicinchoninic acid protein assay has been adapted for use in a microtiter plate, into an easy, indirect method for screening MDR efflux blockers in plant extracts. This spectrophotometric assay is used to determine the enhancement of adriamycin cytotoxicity against resistant cancer cells by plant extracts or pure compounds indirectly. We have shown that the optical density measured (amount of cellular protein present) correlates with the number of viable cells and that fluorescence of Adriamycin associated with the cell correlates with the concentrations of Adriamycin added to the media. In addition, the relative efficacy of MDR reversal by various alkaloids has been determined.