Screening potential P-glycoprotein inhibitors by combination of a detergent-free membrane protein extraction with surface plasmon resonance biosensor.
10.1016/j.apsb.2022.03.016
- Author:
Yuhong CAO
1
;
Jiahao FANG
1
;
Yiwei SHI
1
;
Hui WANG
1
;
Xiaofei CHEN
1
;
Yue LIU
1
;
Zhenyu ZHU
1
;
Yan CAO
1
;
Zhanying HONG
1
;
Yifeng CHAI
1
Author Information
1. School of Pharmacy, Second Military Medical University, Shanghai Key Laboratory for Pharmaceutical Metabolites Research, Shanghai 200433, China.
- Publication Type:Journal Article
- Keywords:
Affinity calculation;
Inhibitor screening;
Membrane proteins;
Multidrug resistance;
Natural products;
P-Glycoprotein;
Styrene maleic acid;
Surface plasmon resonance
- From:
Acta Pharmaceutica Sinica B
2022;12(7):3113-3123
- CountryChina
- Language:English
-
Abstract:
P-glycoprotein (P-gp) highly expressed in cancer cells can lead to multidrug resistance (MDR) and the combination of anti-cancer drugs with P-gp inhibitor has been a promising strategy to reverse MDR in cancer treatment. In this study, we established a label-free and detergent-free system combining surface plasmon resonance (SPR) biosensor with styrene maleic acid (SMA) polymer membrane proteins (MPs) stabilization technology to screen potential P-gp inhibitors. First, P-gp was extracted from MCF-7/ADR cells using SMA polymer to form SMA liposomes (SMALPs). Following that, SMALPs were immobilized on an SPR biosensor chip to establish a P-gp inhibitor screening system, and the affinity between P-gp and small molecule ligand was determined. The methodological investigation proved that the screening system had good specificity and stability. Nine P-gp ligands were screened out from 50 natural products, and their affinity constants with P-gp were also determined. The in vitro cell verification experiments demonstrated that tetrandrine, fangchinoline, praeruptorin B, neobaicalein, and icariin could significantly increase the sensitivity of MCF-7/ADR cells to Adriamycin (Adr). Moreover, tetrandrine, praeruptorin B, and neobaicalein could reverse MDR in MCF-7/ADR cells by inhibiting the function of P-gp. This is the first time that SMALPs-based stabilization strategy was applied to SPR analysis system. SMA polymer can retain P-gp in the environment of natural lipid bilayer and thus maintain the correct conformation and physiological functions of P-gp. The developed system can quickly and accurately screen small molecule ligands of complex MPs and obtain affinity between complex MPs and small molecule ligands without protein purification.
