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Fast and facile analysis of glycosylation and phosphorylation of fibrinogen from human plasma—correlation with liver cancer and liver cirrhosis

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Abstract

Hepatocellular carcinoma (HCC) is one of the deadliest cancers due to its late diagnosis with the main risk factor being liver cirrhosis (LC). Glycan structures from glycoproteins are usually altered in cancer. Blood plasma from 111 healthy and sick donors was analyzed to determine the post-translational modifications (PTM) of intact Aα-, Bβ-, and γ-subunits of fibrinogen, a glycoprotein predominantly produced in liver cells. Glycosylation and phosphorylation of the protein species were quantified by liquid chromatography coupled to mass spectrometry to correlate PTMs to pathological cases. Quantities of the PTMs were used for statistical classification by principal component analysis (PCA) and multivariate analysis of variance (MANOVA). As relevant clinical finding, patients with liver disease (HCC and/or LC) were distinguished from individuals without relevant chronic liver disease with 91% sensitivity and 100% specificity. Within the group of patients with liver disease, a robust separation between LC and HCC was not possible. In more detail, the phosphorylation of Aα-subunit is decreased in HCC patients, whereas the monophosphorylated state is significantly increased in LC patients. In terms of glycosylation, the amount of O-glycans in the Aα-subunit is decreased in LC patients, while sialylation and fucosylation of N-type glycans of Bβ- and γ-subunits are increased in LC and HCC. Based on PTM of fibrinogen, starting from plasma we can assign the status of an individual as healthy or as liver disease in less than 3 h.

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Acknowledgments

We sincerely thank Dr. M. Braun and Dr. L. Schmidt, Zentralinstitut für Transfusionsmedizin GmbH, Hamburg, for providing the plasma samples of healthy individuals. We also like to thank Dr. Maria Riedner, Department of Chemistry, University of Hamburg, for assistance with the mass spectrometry.

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  1. Florentine Klaus

    Present address: University of Münster, Institute for Hygiene, Robert Koch Str. 41, 48149, Münster, Germany

Authors and Affiliations

  1. Department of Chemistry, University of Hamburg, Martin-Luther-King-Platz 6, 20146, Hamburg, Germany

    Tim Nagel, Florentine Klaus & Bernd Meyer

  2. Department of Internal Medicine, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, 20246, Hamburg, Germany

    Ines Gil Ibanez, Henning Wege & Ansgar Lohse

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  1. Tim Nagel
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Nagel, T., Klaus, F., Ibanez, I.G. et al. Fast and facile analysis of glycosylation and phosphorylation of fibrinogen from human plasma—correlation with liver cancer and liver cirrhosis. Anal Bioanal Chem 410, 7965–7977 (2018). https://doi.org/10.1007/s00216-018-1418-7

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