Abstract
A decrease of the intracellular concentration of doxorubicin by activation of ABC-transporters, mainly P-glycoprotein, leads to a reduction of its chemotherapeutical efficacy. To overcome multidrug resistance, digitonin, steroidal saponin, was selected to enhance cell permeability, increase intracellular accumulation, and anticancer effect of doxorubicin. We investigated the cytotoxicity and P-glycoprotein modulatory effect of digitonin in combination with doxorubicin in resistant leukemia and colon cells. MTT assay was applied to evaluate the cell viability and reversal effect of this combination. Rhodamine123 and calcein efflux assays were used for investigate P-glycoprotein function by flow cytometry. At the molecular level, RT-PCR confirmed the data obtained. Digitonin exhibits a significant effect on viability of Caco-2 and CEM/ADR5000 cells with IC
50
values 15.17µM and 16.02µM, respectively. The co-incubation of doxorubicin with non-toxic concentration of digitonin (5µM) resulted in an enhance doxorubicin cytotoxicity in Caco-2 and CEM/ADR5000 cells by 1.9- and 1.2-fold, respectively. Digitonin increase Rho123 and calcein accumulation in Caco-2 and CEM/ADR5000 cells in dose dependent manner. Moreover, 5µM digitonin increases the accumulation of Rho123 and calcein 1.3- and 1.1-fold of verapamil activity in Caco-2 cells. RT-PCR data indicate that 5µM digitonin down-regulated P-gp/MDR1 mRNA to 80% of the control level. In conclusion, digitonin enhances the antitumor effect of doxorubicin and exhibits P-glycoprotein modulatory effect, so it considered as an efficient additive to the chemotherapeutic principle.
Keywords:
digitonin, anticancer, doxorubicin resistance