Pinoresinol isolated from Melia azedarach reverts multidrug resistance mediated by P-glycoprotein in chronic myeloid leukemia cells.

GONZÁLEZ M; VERA M; JORAY M; MACCIONI M; PALACIOS S; RUMJANEK V; CARPINELLA M

Simposio; VIII Simpósio de Oncobiologia.; 2014

Introduction and Aims: Multidrug resistance (MDR) is one of the major causes of chemotherapy failure in cancer treatment. An important MDR mechanism consists in the overexpression of the efflux pump P-glycoprotein (P-gp) which extrudes out of cells a variety of anticancer drugs by an ATP-dependent mechanism. This results in a decreased intracellular concentration of these agents and hence in their cytotoxicity. The development of safe and effective P-gp inhibitors is thus of great concern to researchers. There are few reports about the use of compounds isolated from plants of Argentina as MDR modulators. After performing an evaluation of the reversal effect over P-gp mediated MDR of 15 compounds isolated from native and naturalized plants from central Argentina in chronic myeloid leukemia (CML) cells, pinoresinol (1), isolated from M. azedarach, showed to be the most effective. This fact encouraged further studies related to the activity of this promising compound. Materials and Methods: The CML cell line K562 and its MDR variant due to P-gp overexpression, Lucena 1, were used. The effect of 1 on doxorubicin (DOX) cytotoxicity was evaluated by MTT assay. DOX accumulation and efflux assays and detection of P-gp expression were performed by flow cytometry. Molecular modelling studies were performed using a human P-gp model. P-gp ATPase activity was evaluated using the PREDEASY ATPase kit (SOLVO Biotechnology). The cytotoxicity of 1 on peripheral blood mononuclear cells was measured by MTT assay. Results: Compound 1 caused a decrease of 9.4 times in the IC50 of DOX in Lucena 1 cells at 40 µg/mL, showing the same level of activity as that of the commercial modulator verapamil. This result was confirmed by a significant increase in the intracellular DOX accumulation in MDR cells. This increase was noticed in the first hour of the experiment and remained over 13 hs of treatment. Once compound 1 was removed from the medium, it did not inhibited DOX efflux. These effects were not observed in K562. Docking studies revealed that 1 binds to aromatic residues of P-gp involved in a "v" vortex formed by the trans-membrane -helices 4, 5 and 6. On the other hand, P-gp ATPase activity was proved to be inhibited by 1 with an IC50 of 7.5 µg/mL. No effect on P-gp expression was observed. In addition, 1 did not affect cell viability of fresh human lymphocytes up to 160 µg/mL. Conclusions: These results suggest that compound 1 has great potential to be further developed as a P-gp inhibitor. This agent could be coadministered with an antineoplastic drug in order to improve cancer chemotherapy and patient?s prognosis.