Effects Of Imatinib On Mucin O-glycosylation In K562Cells

Objective: To investigate the expression variation of mucins tumor-associatedcarbohydrate antigens (TACAs) and of mRNA of glycosyltransferases synthesizingthese TACAs in chronic myelogenous leukemia cell line K562after treated withimatinib,and to study the change of colocalization between glycosyltransferasesppGalNAc-Ts and endoplasmic reticulum in K562cell after treated with imatinib. Topreliminary study the expression difference of mucins TACAs and mRNA ofglycosyltransferases synthesizing these TACAs between K562cells and imatinibresistant K562cells.Methods:(1) After K562cells treated with imatinib, flow cytometry was used todetect the expression level of mucins TACAs in K562cell surface and immunofluoresc-ence was performed for testing the degree of colocalization between ppGalNAc-Ts andendoplasmic reticulum in K562cells.(2) RT-PCR was used to detect the expression ofmRNA level of glycosyltransferases synthesizing mucins TACAs.(2) Imatinib wasadded to K562cell cultures in stepwise increasing concentrations to develop a drugresistent cell subline, RT-PCR, apoptosis assay and MTT assay were used to identify theimatinib resistant K562cell subline; the expression difference of mucins TACAs andmRNA of glycosyltransferases synthesizing mucins TACA between K562cells andimatinib resistance K562cells was detected respectively by flow cytometry andRT-PCR.Results:(1) After K562cells treated with imatinib, the expression level of Tnantigen was increased,while T antigen and sialylated T antigen were decreased; and thedegree of colocalization of ppGalNAc-Ts and endoplasmic reticulum was decreased.(2)With the increasing concentration of imatinib,the expression of mRNA of glycosyltrans-ferases synthesizing mucins TACAs were all decreased in different degrees.(3) Theexpression of MDR1mRNA were significantly increased in imatinib-resistant K562cells, and the imatinib-resistant K562cells has passed the test by the cell apoptotic assay and cell proliferation assay. Then the imatinib-resistant K562cells was namedK562A. The expression level of Tn antigen and sialylated T antigen on K562A cellssurface were higher than on K562cells surface,while T antigen were lower by the flowcytometry. Compared to K562cell, the mRNA level of ppGalNAc-T2, ST3Gal1andST6GalNAc4in K562A cells were higher, while the mRNA level of C1Gal-T1andST6GalNAc1in K562A cell lines were lower.Conclusion:(1) Imatinib can affect the expression of mucin tumor-associatedcarbohydrate antigens (TACAs) on K562cell surface,such as Tn antigen, T antigen, andthe sialylated T antigen.(2) According to the reference, the tyrosine kinase activity ofthe BCR-ABL fusion protein can enhance Src kinase activity, Src kinase impelppGalNAc-Ts being transported from the Golgi to the endoplasmic reticulum by COP Ivesicles, and O-glycan synthesis mode changes, as a result, the expression of mucinsTACAs on K562cell surface maybe affected. So imatinib may affect the iniatiation ofO-glycosylation and the expression mucin TACAs on cell surface by inhibiting thetyrosine kinase activity of the BCR-ABL.(3) Imatinib affecting expression of TACAson K562cell surface may also be caused by inhibiting the expression of theglycosyltransferase gene related with TACAs.(4) High expression of P-glycoproteinmay be the mian mechanism of imatinib resistance in K562A cells. Tn antigen andsialylated T antigen levels on K562A cells surface are higher than that on K562cells,while T antigen is opposite. The different level of mucin TACAs between K562cellsand K562A cells may be due to the changed level of glycosyltransferases synthesizingTACAs in K562A cells.