Quantitative Proteomic Analysis of Bromotetrandrine and Tetrandrine in K562 Cell Line Using 18O-labeling Method
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Abstract:
Objective To compare quantitative proteomic analysis of bromotetrandrine (W198) which was a Class I new antitumor drug in China and tetrandrine (Tet) in K562 cell line using 18O-labeling method. Methods To illustrate its mechanism, a shotgun quantitative proteomic strategy employing 2D LC-MS-MS and trypsin catalyzed 18O-labeling quantification was carried out in this study. Compared to normal chronic leukemia cell line K562 and K562 induced by Tet, the proteomic changes of K562 induced by W198 were investigated. In order to validate the quantitation by the 18O-labeling, the analysis was done on an equivalent sample composed of the same amount of labeled and unlabeled proteins from normally cultured cells to act as a reference to the comparative sample. Results A threshold of ± 2-fold change for deciding whether a protein concentration was changed was settled for the following experiments. Comparing the 105 identified soluble proteins’ expression levels of the apoptosis starting up K562 cells after W198 induction with the normally cultured cells, 16 proteins were found with significantly altered expression levels after W198 treatment. Eight proteins were up-expressed including HMGB2, peroxiredoxin-2, and eIF4A-I, etc. Eight proteins were down-expressed including TCP-1, GRP94, GST-π, and SFGHs, etc. Compared to K562 induced by Tet, eight proteins of K562 were found with significantly altered expression levels after W198 treatment. Five proteins were up-expressed including HSP 90-β and 40S ribosomal protein S15a, etc. Three proteins were down-expressed including phosphoglycerate kinase 1, isoform 5 of interleukin enhancer-binding factor 3, etc. Conclusion The 18O-labeling MS-MS-based method is ideal as a discovery tool, but it is not suitable for validation using a large number of samples. Other more effective methods, such as Western blotting should be used for further validation of candidate cancer proteins discovered from 18O-labeling samples. In total, 105 soluble proteins were discovered, and 16 proteins were found with significantly altered expression levels after W198 treatment. These repressed or activated proteins are the potential drug targets of W198, which may provide novel targets for future development of biomarkers for cancer therapy.
TAN Ying, GE Zhi-qiang, LIU Chang-xiao. Quantitative Proteomic Analysis of Bromotetrandrine and Tetrandrine in K562 Cell Line Using 18O-labeling Method[J]. Chinese Herbal Medicines (CHM),2012,4(1):43-52
