Xyloside-primed Chondroitin Sulfate/Dermatan Sulfate from Breast Carcinoma Cells with a Defined Disaccharide Composition Has Cytotoxic Effects in Vitro

Abstract

We previously reported that the xyloside 2-(6-hydroxynaphthyl) ␤-D-xylopyranoside (XylNapOH), in contrast to 2-naphthyl ␤-D-xylopyranoside (XylNap), specifically reduces tumor growth both in vitro and in vivo. Although there are indications that this could be mediated by the xyloside-primed glycosaminoglycans (GAGs) and that these differ in composition depending on xyloside and cell type, detailed knowledge regarding a structure-function relationship is lacking. In this study we isolated XylNapOH-and XylNap-primed GAGs from a breast carcinoma cell line, HCC70, and a breast fibroblast cell line, CCD-1095Sk, and demonstrated that both XylNapOH-and XylNap-primed chondroitin sulfate/dermatan sulfate GAGs derived from HCC70 cells had a cytotoxic effect on HCC70 cells and CCD-1095Sk cells. The cytotoxic effect appeared to be mediated by induction of apoptosis and was inhibited in a concentration-dependent manner by the XylNap-primed heparan sulfate GAGs. In contrast, neither the chondroitin sulfate/ dermatan sulfate nor the heparan sulfate derived from CCD1095Sk cells primed on XylNapOH or XylNap had any effect on the growth of HCC70 cells or CCD-105Sk cells. These observations were related to the disaccharide composition of the XylNapOH-and XylNap-primed GAGs, which differed between the two cell lines but was similar when the GAGs were derived from the same cell line. To our knowledge this is the first report on cytotoxic effects mediated by chondroitin sulfate/dermatan sulfate.

The dual role of the glycosaminoglycan chondroitin‐6‐sulfate in the development, progression and metastasis of cancer

“…Evidence supporting the suggestion that the 6‐ O ‐sulfation of CS can strongly influence the behavior of a tumor cells (Fig. ) has come from the observation that xyloside‐primed CS/DS that was produced by a breast carcinoma cell line demonstrated a strong cytotoxic activity toward both normal and tumor cells . This effect, which involved the induction of apoptosis, was neutralized by cancer‐cell‐synthesized heparan sulfate .…”

“…Moreover, the cytotoxic CS/DS also differed from the GAG that was produced by normal cells in respect to a significantly lower content of the IdoA residues (~10% versus ~40%, respectively) and to the occurrence of 4,6‐ O ‐disulfated disaccharides (E units) . However, despite their known biological potential, the presence of E units cannot be the only structural determinant that is responsible for the cytotoxic effect of the xyloside‐primed CS/DS because another CS/DS that was used in the research and that had a similar level of these disaccharides did not demonstrate any cytotoxic activity . Thus, the sequential context in which E units are presented such, as the 6‐ O ‐ or 4‐ O ‐sulfation of adjacent disaccharides, may be crucial for the biological properties of CS/DS.…”

“…Interestingly, CS/DS with these cytotoxic properties had a very high proportion of the 6-O-sulfated disaccharides in relation to the 4-O-sulfated ones (82-63% versus 15-30%) compared to the cytotoxically inactive GAG from normal breast fibroblasts, which contained~60% of the 4-O-sulfated disaccharides and~34% of the 6-O-sulfated ones [167,168]. Moreover, the cytotoxic CS/DS also differed from the GAG that was produced by normal cells in respect to a significantly lower content of the IdoA residues (~10% versus~40%, respectively) and to the occurrence of 4,6-O-disulfated disaccharides (E units) [167,168]. However, despite their known biological potential, the presence of E units cannot be the only structural determinant that is responsible for the cytotoxic effect of the xyloside-primed CS/DS because another CS/DS that was used in the research and that had a similar level of these disaccharides did not demonstrate any cytotoxic activity [167].…”

Glycosaminoglycan Domain Mapping of Cellular Chondroitin/Dermatan Sulfates

“…For preparation of the CS/DS used for GAGDoMa, we treated human breast fibroblasts, CCD-1095Sk, and human breast carcinoma cells, HCC70, with 100 μM of the xyloside 2-naphthyl β-D-xylopyranoside and the xyloside-primed GAGs were isolated from the media and depolymerized using the different bacterial enzymes (Fig. 1a) 22 . Thereafter, we subjected the samples to nanoflow reversed-phase ion-pairing (RPIP) chromatography using an in-house packed C18 column and dibutylamine (DBA) as an ion-pairing agent.…”

“…The MS analysis was performed by nanoflow reversed-phase dibutylamine ion-pairing chromatography with negative mode higher-energy collision dissociation (HCD) MS/MS, enabling fragment ion characterization, compositional profiling and semi-quantification of precursor ions. To challenge GAGDoMa, CS/DS primed on xylosides from two types of human cell lines were used as models as they were predicted to cover a wide range of structures 21,22 . Using this technology, we have identified over 200 structures and characterized more than 60 of these in depth.…”