An assay for quantitative analysis of polysialic acid expression in cancer cells

• Published in: Carbohydrate polymers Vol. 259; p. 117741
• Main Authors: Guo, Xiaoxiao, Elkashef, Sara M., Patel, Anjana, Ribeiro Morais, Goreti, Shnyder, Steven D., Loadman, Paul M., Patterson, Laurence H., Falconer, Robert A.
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.05.2021
• Subjects: acid hydrolysis; Animals; Cell Line, Tumor; Cell-based method; Chromatography, High Pressure Liquid; Chromatography, Ion Exchange; Chromatography, Reverse-Phase; drugs; Enzyme Inhibitors - chemistry; Enzyme Inhibitors - metabolism; Enzyme-Linked Immunosorbent Assay; fluorescence; glycomics; glycoproteins; Glycoside Hydrolases - metabolism; Inhibitor screening; polysaccharides; Polysialic acid; prognosis; Quantitative analysis; Rats; Sialic acid; Sialic Acids - analysis; Sialic Acids - metabolism; Sialyltransferases - antagonists & inhibitors; Sialyltransferases - metabolism; Inhibitor screening; Sialic acid; Cell-based method; Polysialic acid; Quantitative analysis
• ISSN: 0144-8617; 1879-1344; 1879-1344
• DOI: 10.1016/j.carbpol.2021.117741

[Display omitted] •Polysialic acid is of significant interest due its roles in biology and disease.•A cell-based assay for quantitative analysis of polySia expression in cancer cells.•A method applicable for evaluating small molecule polyST inhibitors.•A potentially vital tool with applications in drug discovery and in glycobiology. Polysialic acid (polySia) is a linear polysaccharide comprised of N-acetylneuraminic acid residues and its over-expression in cancer cells has been correlated with poor clinical prognosis. An assay has been developed for quantitative analysis of cellular polySia expression. This was achieved by extracting and purifying released polySia from glycoproteins by mild acid hydrolysis and optimised organic extraction. The polySia was further hydrolysed into Sia monomers, followed by fluorescent labelling and quantitative analysis. The assay was qualified utilising endoneuraminidase-NF to remove polySia from the surface of C6-ST8SiaII cancer cells (EC50 = 2.13 ng/mL). The result was comparable to that obtained in a polySia-specific cellular ELISA assay. Furthermore, the assay proved suitable for evaluation of changes in polySia expression following treatment with a small molecule inhibitor of polysialylation. Given the importance of polySia in multiple disease states, notably cancer, this is a potentially vital tool with applications in the fields of drug discovery and glycobiology.