A new anionic surfactant (RapiGest SF) was successfully used for site-specific analysis of glycosylation in human alpha-1-acid glycoprotein (AGP). By means of this analytical approach combined with capillary HPLC-mass spectrometry (and tandem mass spectrometry), the N-linked glycosylation pattern of AGP was explored. On the basis of mass matching and MS/MS experiments ca 80 different AGP-derived glycopeptides were identified. Glycosylation shows a markedly different pattern for the various glycosylation sites. At sites I and II, triantennary complex-type oligosaccharides predominate and at sites III, IV and V, tetra-antennary complex-type oligosaccharides predominate. Sites IV and V show the presence of additional N-acetyl lactosamine (Gal-GlcNAc) units (even higher degree of branching and/or longer antennae are also present).

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Cancer can be viewed as a disorder generating from an uncoupling of gene expression that controls cellular proliferation and differentiation. Therefore, much attention has been paid to cancer cell lines that can be induced to differentiate after in vitro exposure to chemical or chemotherapeutic agents (for reviews, see refs. 1-3). The K-562 human leukemia cell line provides a useful system for studying human erythroid differentiation because it expresses markers of erythroid lineages, such as hemoglobin.Of the variety of chemotherapeutic agents used in cancer treatment, some can induce cell differentiation (4). In our previous studies we used drugs directed against key enzymes of purine and pyrimidine metabolism (5-7). Activities of target enzymes of these drugs were those that were tightly linked with in vivo and in vitro proliferative activity (5,8,9). The activity of IMP dehydrogenase (IMP:NAD' oxidoreductase; EC 1.1.1.205), the rate-limiting enzyme of de novo GTP biosynthesis, markedly increased in various types of cancer cells; therefore, this enzyme was suggested as a sensitive target for anticancer chemotherapy (5, 10). Tiazofurin (2-i-D-ribofuranosyl-4-thiazolecarboxamide;NSC-286193) potently inhibits the proliferation ofa variety of experimental and human neoplasms and is now in phase I and II clinical trial (11, 12

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Human serum acid alpha-1-glycoprotein (AGP, orosomucoid) content of healthy individuals and cancer patients was measured, isolated and purified using a protocol of fast and biocompatible sample preparation, ion exchange and dye-ligand affinity chromatographic methods. In comparison to the healthy individuals significantly higher serum AGP levels were found in a wide spectrum of cancer patients, indicating its diagnostic value in the malignant disease. Oligosaccharide content of AGP samples was separated following PNGase F enzyme digestion and analysed by RP-HPLC and MALDI-TOF mass spectrometry. RP-HPLC and MALDI-TOF mass spectrometric analysis of sugar constituents of AGP specimen originated from selected cancer patients with high serum AGP levels indicated the appearance of anomal distribution of bi-, tri- and tetra-antennary oligosaccharide structures compared to the healthy controls.

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According to the high marker level in lung and liver metastases, the marker might be useful in monitoring both patients with disease free ocular melanomas, to detect liver metastasis and high-risk patients after surgical removal of the primary tumour to reveal lung metastases.

Genetic variants of human plasma alpha-1 acid glycoprotein (AGP) have been studied in cancer, compared with a group of healthy control. AGP has four genetic variants: AGP F1, F2, and S variants correspond to the ORM1 gene whereas AGP A corresponds to the ORM2 gene. The proportion of ORM1 and ORM2 variants were studied in plasma using a novel UPLC-MS method. Plasma total AGP level was 0.5 +/- 0.2 g L(-1) and the proportions of the ORM1 and ORM2 variants were 76.3 +/- 8.2% and 23.7 +/- 8.2%, respectively. In cancer plasma AGP levels increased fourfold and the proportion of ORM1 variants increased to 88.7 +/- 6.8%. Changes in the proportion of genetic variants due to cancer were clearly significant, as shown by statistical analysis. Three different cancer types have been studied, lymphoma, melanoma, and ovarian cancer. The results did not show any difference depending on cancer type. The results indicate that, in accordance with prior expectations, the ORM1 variant is predominantly responsible for the acute-phase property of AGP.