title: Capillary electrophoresis for screening of 20S proteasome inhibitors.
abstract: A method for studying 20S proteasome inhibitors by capillary electrophoresis (CE) has been developed. Proteasome plays a fundamental role in degrading key regulatory proteins. The 20S proteasome can degrade intrinsically disordered proteins in an ATP-independent manner without additional "helper" molecules. The discovery of new proteasome inhibitors with little or no toxicity is highly desirable in anticancer therapy. In this study, the inhibitory effects of MG132 and MG115 on the 20S proteasome were evaluated by CE for the first time. The optimized CE conditions were as follows: fused-silica capillary of 30 cm effective length and 75 microm internal diameter, pressure injection of 0.5 psi for 5 s, 50 mM Hepes buffer (pH 7.6) with 2% dimethyl sulfoxide, constant voltage of 20 kV, and detection wavelength at 340 nm. Also, the new method was used to study the inhibitory effects of 30 novel peptidyl vinyl ester derivatives of MG132. The 50% inhibition concentrations (IC(50) values) of MG132 and MG115 were 40.0 and 84.7 nM, respectively. Two new compounds, XP32 and XP35, showed considerable inhibitory effects on the 20S proteasome. When the concentrations of them were fixed at 172 nM, their inhibition rates were 36.2% and 29.1%, respectively. The results showed that the CE method was powerful, sensitive, and fast and required little sample. It could be employed as one of the reliable drug screening methods for 20S proteasome inhibitors.
authors.affiliation: Department of Pharmaceutical Analysis, School of Pharmaceutical Science, Peking University, Beijing 100191, People's Republic of China.
authors.forename: Wenjing
authors.initials: W
authors.lastname: Chen
authors.forename: Ke
authors.initials: K
authors.lastname: Mou
authors.forename: Bo
authors.initials: B
authors.lastname: Xu
authors.forename: Xiaomei
authors.initials: X
authors.lastname: Ling
authors.forename: Jingrong
authors.initials: J
authors.lastname: Cui
authors.forename: Ping
authors.initials: P
authors.lastname: Xu
mesh_terms: D000818:Animals; D003374:Coumarins / Q000378:metabolism; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D019075:Electrophoresis, Capillary / Q000379:methods*; D004952:Esters / Q000737:chemistry; D006868:Hydrolysis; D020128:Inhibitory Concentration 50; D007976:Leupeptins / Q000737:chemistry / Q000494:pharmacology; D016014:Linear Models; D011480:Protease Inhibitors / Q000032:analysis* / Q000737:chemistry / Q000494:pharmacology*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D061988:Proteasome Inhibitors; D011817:Rabbits; D015203:Reproducibility of Results
chemical_list: D003374:Coumarins; D004952:Esters; D007976:Leupeptins; D011480:Protease Inhibitors; D061988:Proteasome Inhibitors; C089486:carbobenzoxy-leucyl-leucyl-norvalinal; D046988:Proteasome Endopeptidase Complex; C028743:7-amino-4-methylcoumarin; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde