Mode of Action Analyses of Neferine, a Bisbenzylisoquinoline Alkaloid of Lotus (Nelumbo nucifera) against Multidrug-Resistant Tumor Cells

Abstract

Neferine, a bisbenzylisoquinoline alkaloid isolated from the green seed embryos of Lotus (<i>Nelumbo nucifera</i> Gaertn), has been previously shown to have various anti-cancer effects. In the present study, we evaluated the effect of neferine in terms of P-glycoprotein (P-gp) inhibition via <i>in vitro</i> cytotoxicity assays, R123 uptake assays in drug-resistant cancer cells, <i>in silico</i> molecular docking analysis on human P-gp and <i>in silico</i> absorption, distribution, metabolism, and excretion (ADME), quantitative structure activity relationships (QSAR) and toxicity analyses. Lipinski rule of five were mainly considered for the ADME evaluation and the preset descriptors including number of hydrogen bond donor, acceptor, hERG IC₅₀, logp, logD were considered for the QSAR analyses. Neferine revealed higher toxicity toward paclitaxel- and doxorubicin-resistant breast, lung or colon cancer cells, implying collateral sensitivity of these cells toward neferine. Increased R123 uptake was observed in a comparable manner to the control P-gp inhibitor, verapamil. Molecular docking analyses revealed that neferine still interacts with P-gp, even if R123 was pre-bound. Bioinformatical ADME and toxicity analyses revealed that neferine possesses the druggability parameters with no predicted toxicity. In conclusion, neferine may allocate the P-gp drug-binding pocket and prevent R123 binding in agreement with P-gp inhibition experiments, where neferine increased R123 uptake.