Sialylation of EGFR by the ST6Gal-I sialyltransferase promotes EGFR activation and resistance to gefitinib-mediated cell death.

Abstract

BACKGROUND:

The ST6Gal-I sialyltransferase is upregulated in numerous cancers, and high expression of this enzyme correlates with poor patient prognosis in various malignancies, including ovarian cancer. Through its sialylation of a select cohort of cell surface receptors, ST6Gal-I modulates cell signaling to promote tumor cell survival. The goal of the present study was to investigate the influence of ST6Gal-I on another important receptor that controls cancer cell behavior, EGFR. Additionally, the effect of ST6Gal-I on cancer cells treated with the common EGFR inhibitor, gefitinib, was evaluated.

RESULTS:

Using the OV4 ovarian cancer cell line, which lacks endogenous ST6Gal-I expression, a kinomics assay revealed that cells with forced overexpression of ST6Gal-I exhibited increased global tyrosine kinase activity, a finding confirmed by immunoblotting whole cell lysates with an anti-phosphotyrosine antibody. Interestingly, the kinomics assay suggested that one of the most highly activated tyrosine kinases in ST6Gal-I-overexpressing OV4 cells was EGFR. Based on these findings, additional analyses were performed to investigate the effect of ST6Gal-I on EGFR activation. To this end, we utilized, in addition to OV4 cells, the SKOV3 ovarian cancer cell line, engineered with both ST6Gal-I overexpression and knockdown, as well as the BxPC3 pancreatic cancer cell line with knockdown of ST6Gal-I. In all three cell lines, we determined that EGFR is a substrate of ST6Gal-I, and that the sialylation status of EGFR directly correlates with ST6Gal-I expression. Cells with differential ST6Gal-I expression were subsequently evaluated for EGFR tyrosine phosphorylation. Cells with high ST6Gal-I expression were found to have elevated levels of basal and EGF-induced EGFR activation. Conversely, knockdown of ST6Gal-I greatly attenuated EGFR activation, both basally and post EGF treatment. Finally, to illustrate the functional importance of ST6Gal-I in regulating EGFR-dependent survival, cells were treated with gefitinib, an EGFR inhibitor widely used for cancer therapy. These studies showed that ST6Gal-I promotes resistance to gefitinib-mediated apoptosis, as measured by caspase activity assays.

CONCLUSION:

Results herein indicate that ST6Gal-I promotes EGFR activation and protects against gefitinib-mediated cell death. Establishing the tumor-associated ST6Gal-I sialyltransferase as a regulator of EGFR provides novel insight into the role of glycosylation in growth factor signaling and chemoresistance.

• Authors: Colleen M Britain, Andrew T Holdbrooks, Joshua C Anderson, Christopher D Willey, Susan L Bellis
• Journal: Journal of ovarian research (J Ovarian Res) Vol. 11 Issue 1 Pg. 12 (Feb 05 2018) ISSN: 1757-2215 [Electronic] England
• PMID: 29402301 (Publication Type: Journal Article)

Chemical References

• Antineoplastic Agents
• Protein Kinase Inhibitors
• Proteome
• Quinazolines
• Epidermal Growth Factor
• Sialyltransferases
• ErbB Receptors
• Protein-Tyrosine Kinases
• Gefitinib
• beta-D-Galactoside alpha 2-6-Sialyltransferase

Topics

• Antineoplastic Agents (pharmacology)
• Apoptosis (drug effects)
• Cell Line, Tumor
• Drug Resistance, Neoplasm
• Epidermal Growth Factor (metabolism, pharmacology)
• ErbB Receptors (metabolism)
• Gefitinib
• Gene Expression
• Gene Knockdown Techniques
• Humans
• Protein Kinase Inhibitors (pharmacology)
• Protein-Tyrosine Kinases (metabolism)
• Proteome
• Proteomics (methods)
• Quinazolines (pharmacology)
• Sialyltransferases (genetics, metabolism)
• Signal Transduction (drug effects)
• beta-D-Galactoside alpha 2-6-Sialyltransferase