Cancer-Type Expression of Tn Epitopes and LacdiNAc Structures: Human Cancer Cells Exhibit Distinctly Varying Levels of Heterogenous Ser/Thr Bearing Polypeptides, a Neutral β Galactosidase Converting T-Hapten to Tn, Ser/Thr: αGalNAc- and GlcNAc: β1-3/β1-4 GalNAc Transferase Activities

Abstract

Terminal sugar-alteration in carbohydrate chains such as GalNAc replacing Gal in prostate and pancreatic cancers and Gal3-O-sulfation in breast, colon and gastric tumors could play a crucial role in cancer pathogenesis. We found the activities of cancer cell GalNAc transferases (GalNAc-Ts) as 0%, 0% <20%, 20-50% and 20-120% respectively towards Galβ1-3GalNAcα-OBn, 4-FGlcNAcβ1-6 (Galβ1-3)GalNAcα-O-Bn, LacNAcβ-O-Bn, GlcNAcβ1-4 GlcNAcβ-O-Bn and GlcNAcβ1-6GalNAcα-O-Bn as compared to GlcNAcβ-OBn. α-[6-³H]GalNAc-ylated endogenous cancer cells Ser/Thr polypeptides by the corresponding cancer cell αGalNAc-T were at variable levels, heterogenous, and exhibited complete binding to VVL-agarose and non-binding to WGA-, WFL- and ConA- agarose. PNA-agarose binding and non-binding radioactive products from [6-³H] GalNAc-ylated exogenous acceptor GlcNAcβ1-6(Galβ1-3) GalNAcα-O-Al indicated cancer type variable β1-3Galactosidase activities at neutral pH. TLC analysis identified two radioactive products by confirming PNA-agarose data. WGA-agarose tight binding and VVL-agarose weak binding respectively of the products [6-³H] GalNAcβ1-4 and β1-3GlcNAcβ-OBn isolated from the exogenous acceptor GlcNAc-β-O-Bn by Sep-Pak C18 method were used to quantitate β1-4 and β1- 3GalNAc-T activities in cancer cells. DU4475, MDA-MB-435S, PA-1, LNCaP, PC3, DU145, EG7 and GL261- OVA over-expressed β1-3GalNAc-T activity. Tumorigenic MDA-MB-435/LLC6 as compared to non-tumorigenic MDA-MB-435S contained ~2-fold each of αGalNAc-T and β1-4GalNAc-T. The breast cancer DU4475 uniquely expressed 10-fold β1-3GalNAc-T with respect to β1-4GalNAc-T. HPLC identified negligible β1-6GalNAc-T in cancer cells and high-level β1-3GalNAc-T in pancreatic and gastric tumors. It is known that Tn epitopes correlate with cancer progression and metastasis and β-galactosidase is a senescence-biomarker and moleculartarget for ovarian cancer. It is apparent that βGalNAc-T, Tn polypeptides, αGalNAc-T and neutral β1-3galactosidase could play a crucial role in cancer pathogenesis.