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Mass Spectrometric Analysis of Glycosphingolipid Antigens
Published on: April 16, 2013
Identification of glycan structure alterations on cell membrane proteins in desoxyepothilone B resistant leukemia
Miyako Nakano1, Rohit Saldanha, Anja Göbel
1Department of Chemistry and Biomolecular Sciences, Macquarie University, NSW 2109, Australia.
Abstract:
Resistance to tubulin-binding agents used in cancer is often multifactorial and can include changes in drug accumulation and modified expression of tubulin isotypes. Glycans on cell membrane proteins play important roles in many cellular processes such as recognition and apoptosis, and this study investigated whether changes to the glycan structures on cell membrane proteins occur when cells become resistant to drugs. Specifically, we investigated the alteration of glycan structures on the cell membrane proteins of human T-cell acute lymphoblastic leukemia (CEM) cells that were selected for resistance to desoxyepothilone B (CEM/dEpoB). The glycan profile of the cell membrane glycoproteins was obtained by sequential release of N- and O-glycans from cell membrane fraction dotted onto polyvinylidene difluoride membrane with PNGase F and β-elimination respectively. The released glycan alditols were analyzed by liquid chromatography (graphitized carbon)-electrospray ionization tandem MS. The major N-glycan on CEM cell was the core fucosylated α2-6 monosialo-biantennary structure. Resistant CEM/dEpoB cells had a significant decrease of α2-6 linked sialic acid on N-glycans. The lower α2-6 sialylation was caused by a decrease in activity of β-galactoside α2-6 sialyltransferase (ST6Gal), and decreased expression of the mRNA. It is clear that the membrane glycosylation of leukemia cells changes during acquired resistance to dEpoB drugs and that this change occurs globally on all cell membrane glycoproteins. This is the first identification of a specific glycan modification on the surface of drug resistant cells and the mechanism of this downstream effect on microtubule targeting drugs may offer a route to new interventions to overcome drug resistance.
Insights
Drug-resistant leukemia cells show altered cell surface glycans. This study identifies decreased sialic acid on N-glycans in resistant cells, linked to reduced ST6Gal enzyme activity, offering potential new strategies against cancer drug resistance.
Area of Science:
- Biochemistry
- Molecular Biology
- Cancer Research
Background:
- Cancer cells develop resistance to chemotherapy through various mechanisms.
- Cell surface glycans play critical roles in cellular functions and interactions.
- Understanding glycan alterations in drug-resistant cancer cells is crucial for developing new therapies.
Purpose of the Study:
- To investigate changes in cell membrane glycan structures in human T-cell acute lymphoblastic leukemia (CEM) cells selected for resistance to desoxyepothilone B (dEpoB).
- To identify specific glycan modifications associated with acquired drug resistance in leukemia.
Main Methods:
- Sequential release of N- and O-glycans from cell membrane fractions.
- Analysis of released glycan alditols using liquid chromatography (graphitized carbon)-electrospray ionization tandem mass spectrometry (MS).
- Quantification of sialylation and assessment of ST6Gal enzyme activity and mRNA expression.
Main Results:
- Resistant CEM/dEpoB cells exhibited a significant decrease in α2-6 linked sialic acid on N-glycans compared to sensitive CEM cells.
- This reduction in sialylation was attributed to decreased activity and mRNA expression of β-galactoside α2-6 sialyltransferase (ST6Gal).
- The observed glycan changes were global across all cell membrane glycoproteins.
Conclusions:
- Acquired resistance to dEpoB in leukemia cells is associated with global alterations in membrane glycosylation.
- Decreased α2-6 sialylation of N-glycans is a key feature of dEpoB resistance in these cells.
- These findings provide novel insights into drug resistance mechanisms and suggest potential therapeutic targets for overcoming resistance to microtubule-targeting drugs.
