The phytochemical study of Euphorbia pedroi led to the isolation of a new tetracyclic triterpenoid with an unusual spiro scaffold, spiropedroxodiol (1), along with seven known terpenoids (2–8). Aiming at obtaining compounds with improved multidrug-resistance (MDR) reversal activity, compound 8, an ent-abietane diterpene, was derivatized by introducing nitrogen-containing and aromatic moieties, yielding compounds 9–14. The structures of compounds were characterized by detailed spectroscopic analysis, including 2D NMR experiments (COSY, HMQC/HSQC, HMBC, and NOESY). Compounds 1–14 were evaluated for their MDR-reversing activity on human ABCB1 gene transfected mouse lymphoma cells (L5178Y-MDR) through a combination of functional and chemosensitivity assays. The natural compounds 1–8 were further evaluated on resistant human colon adenocarcinoma cells (Colo320), and, additionally, their cytotoxicity was assessed on noncancerous mouse (NIH/3T3) and human (MRC-5) embryonic fibroblast cell lines. While spiropedroxodiol (1) was found to be a very strong MDR reversal agent in both L5178Y-MDR and Colo320 cells, the chemical modifications of helioscopinolide E (8) at C-3 positively contributed to increase the MDR reversal activity of compounds 10, 12, and 13. Furthermore, in combination assays, compounds 1 and 7–14 enhanced synergistically the cytotoxicity of doxorubicin. Finally, by means of molecular docking, the key residues and binding modes by which compounds 1–14 may interact with a murine P-glycoprotein model were identified, allowing additional insights on the efflux modulation mechanism of these compounds.
Ricardo J. Ferreira 1, Annamária Kincses 2, Márió Gajdács 2, Gabriella Spengler 2, Daniel J. V. A. dos Santos 1,3, Joseph Molnár 2, and Maria-José U. Ferreira 1*
1 iMed.ULisboa (Research Institute for Medicines), Faculty of Pharmacy, Universidade de Lisboa, Avenida Prof. Gama Pinto, 1649-003 Lisboa, Portugal.
2 Department of Medical Microbiology and Immunobiology, Faculty of Medicine, University of Szeged, Dóm tér 10, H-6720 Szeged, Hungary.
3 LAQV@REQUIMTE/Department of Chemistry and Biochemistry, Faculty of Sciences, University of Porto, Rua do Campo Alegre, 4169-007 Porto, Portugal.