Malignant transformation of gastric cells is accompanied by the deregulated expression of
glycosyltransferases leading to the biosynthesis of
tumor-associated
glycans such as the
sialyl-Lewis X antigen (SLex). SLex presence on cell surface
glycoconjugates increases the invasive capacity of
gastric cancer cells and is associated with
tumor metastasis.
ST3Gal IV enzyme is involved in the synthesis of SLex
antigen and overexpressed in gastric
carcinomas. Herein, we identified the
glycoproteins carrying SLex in
gastric cancer cells overexpressing
ST3Gal IV enzyme and evaluated their
biomarker potential for gastric
carcinoma. Methods: SLex modified
glycoproteins were identified applying western blot and mass spectrometry. Immunoprecipitation, proximity
ligation assay (PLA),
E-selectin binding assay and CRISPR/cas9 knockout experiments were performed to characterize the presence of SLex on the identified
glycoprotein.
Protein N-
glycans of the SLex
protein carrier were in deep analyzed by porous-graphitized-
carbon liquid-chromatography and tandem mass spectrometry glycomics. In silico expression analysis of α2-3
sialyltransferase ST3Gal IV and SLex
protein carrier was performed and the conjoint expression of the SLex modified
glycoproteins evaluated by immunohistochemistry and PLA in a series of gastric
carcinomas. Results:
Carcinoembryonic antigen (CEA; CEACAM5) was identified and validated by different methodologies as a major carrier of SLex. N-glycomics of CEA revealed that complex N-
glycans are capped with α2-3 linked
sialic acid (Neu5Acα2-3Galβ1-4GlcNAc). Data set analysis of
ST3Gal IV and CEA showed that
ST3Gal IV expression was associated with patient´s poor survival, whereas CEA did not show any prognostic value. The co-expression of both CEA and SLeX was observed in 86,3% of gastric
carcinoma cases and 74,5% of the total cases displayed the conjoint CEA+SLexin situ PLA expression. This expression was associated with clinicopathological features of the
tumors, including infiltrative pattern of
tumor growth, presence of venous invasion and patient's poor survival. CEA immunoprecipitation from gastric
carcinoma tissues also confirmed the presence of SLex. Conclusion: CEA is the major
glycoprotein carrying SLex in gastric
carcinoma and the conjoint detection of CEA-SLex is associated with aggressive
tumor features highlighting its PLA detection as a
biomarker of
gastric cancer patient prognosis for
theranostic applications.