Changes in serum IgG oligosaccharide chains with prostate cancer progression.

BACKGROUND: Changes of serum IgG oligosaccharide chain structure have been found in B cell lineage tumors and autoimmune diseases. Currently, the cancer-associated carbohydrate epitopes CA72-4 and CA15-3 are used as serum tumor markers. In the present study, we analyzed the structure of serum IgG oligosaccharide chains in prostate cancer (PCa) patients using the simple new method of fluorophore-associated carbohydrate electrophoresis (FACE). We also evaluated the relationship between changes of serum IgG oligosaccharide chain structure and serum concentration of prostate-specific antigen (PSA). MATERIALS AND METHODS: The structure of serum IgG oligosaccharide chains from 12 PCa patients (6 localized cancer, 6 metastatic cancer) and 10 healthy controls was evaluated by FACE. PSA levels in serum were determined by enzyme immunoassay. RESULTS: Fr 1 (monogalactosyl oligosaccharide) and Fr 2 (digalactosyl oligosaccharide) decreased significantly (p<0.05), while Fr 4 (agalactosyl IgG oligosaccharide) increased with PCa tumorprogression. The Fr 4/Fr 1+2 ratio in metastatic PCa patients was significantly higher than in healthy controls (p<0.05), and there was a significant correlation (r=0.84, p<0.05) between serum PSA levels and the Fr 4/Fr 1+2 ratio in all patients with PCa. CONCLUSION: The changes of serum IgG oligosaccharide chain structure with PCa progression are based on the abnormality of glycosylation in PCa metastasis. Therefore, the analysis of serum IgG oligosaccharide chain structure by FACE may be an auxiliary indicator of PSA for monitoring PCa progression.

Changes in oligosaccharide chains of autoantibodies to GRP78 expressed during progression of malignant melanoma stimulate melanoma cell growth and survival.

A correlation between expression of the glucose-regulated protein of 78 kDa (GRP78) in malignant melanoma tumors and poor patient survival is well established. In this study, in addition to demonstrating the expression of GRP78 in tumor tissue, we investigated the immune response against GRP78 in a group of patients with different progression stages of malignant melanoma. Furthermore, we analyzed the glycosylation status of GRP78 immunoglobulin (Ig) G autoantibodies at these stages and evaluated their capacities to affect the protein B-dependent protein kinase signaling pathway and unfolded protein response signaling mechanisms, all known to promote malignant melanoma cell proliferation and survival. We found that progression of disease correlates not only with enhanced expression of GRP78 in the tumor but also with an increase in GRP78 autoantibody serum titers in these patients. We also found that the glycosylation status of anti-GRP78 IgG changes as the disease progresses. The anti-GRP78 IgG is abnormally glycosylated in the Fc region and asymmetrically glycosylated in the Fab region. We demonstrate that hyperglycosylated anti-GRP78 IgGs stimulate cell proliferation through protein B-dependent protein kinase signaling pathways. They also mimic the effects of α2-macroglobulin on the upregulation of GRP78 and X-box binding protein 1, activating transcription factor 6 α, and serine/threonine-protein kinase/endoribonuclease precursor α as endoplasmic reticulum stress biomarkers and show no effect on expression or activation of caspases 3, 9, or 12. In conclusion, the anti-GRP78 IgG autoantibodies downregulate apoptosis and activate unfolded protein response mechanisms, which are essential to promote melanoma cell growth and survival.

Mass spectrometric approach for screening modifications of total serum N-glycome in human diseases: application to cirrhosis.

Congenital and acquired modifications of glycosylation in diseases are a rapidly growing field that demonstrates the importance of glycosylation in human biology. Unfortunately, in clinical biochemistry, very few tests are available to explore oligosaccharide metabolism on a large scale. Such an assay needs to be of high throughput, rapid, and preferentially noninvasive. In the present study, we describe a method to analyze qualitative variations of N-glycosylation of human serum proteins. The method is based on direct release of N-linked oligosaccharides from patient serum samples, a single-step purification, and a matrix-assisted laser desorption ionization time of flight mass spectrometric analysis. A complementary structural study of the released oligosaccharides was achieved by enzymatic digestions, linkage analysis, and electrospray ionization ion trap mass spectrometry (ESI-IT-MS) of the permethylated N-glycome. A total of 26 oligosaccharide structures were individualized, their presence in human serum being the result of the combination of the biosynthesis and catabolic pathways. Application of the protocol to the serum of patients with cirrhosis demonstrates the ability of this assay to identify acquired modifications of glycosylation. Furthermore, we have analyzed the N-glycans and showed the increase in bisecting N-acetylglucosamine residue, core fucosylation, and the presence of an important population of neutral oligosaccharides. The study of total serum N-glycome modifications is a preliminary for the discovery of new noninvasive diagnostic or prognostic biomarkers resulting from the variations of the N-glycan metabolism during diseases.