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Determination of specific glycobiomarkers in alpha-mannosidosis patients by NMR and mass spectrometry

Zuzana Pakanová 1 Mária Matulová 1 Marek Nemčovič 1 Filip Pančík 1 Filip Květoň 1 Iveta Uhliariková 1 Anna Šalingová 2 Anna Hlavatá 3 Ján Mucha 1 Peter Baráth 1

1Institute of Chemistry, Slovak Academy of Sciences, Bratislava
2National Institute of Children´s Diseases, Department of Laboratory Medicine, Bratislava
3Comenius University, Faculty of Medicine and National Institute of Children´s Diseases, Department of Paediatrics, Bratislava
zuzana.pakanova@savba.sk

Alpha-mannosidosis is a progressive lysosomal storage disorder, firstly described in 1967[1]. It is caused by a partial or complete deficiency of alpha-mannosidose, enzyme playing an essential role in oligosaccharide and glycoprotein degradation. Mannose-rich oligosaccharides are accumulated in lysosomes of all tissues, resulting in impaired cellular function and apoptosis [2]. Due to the variability of clinical manifestation and multisystematic involvement, its definitive diagnostics may be challenging. MALDI TOF/TOF mass spectrometric analyses of permethylated oligosaccharides from urine of alpha-mannosidosis patients revealed characteristic set of specific mannose-based glycobiomarkers, compiled from Man2GlcNAc1 (m/z 722.3) up to Man8GlcNAc1 (m/z 1947.0). In 1H NMR spectra, characteristic signals of mannose units, originated from mannose in various linkages and locations in these oligosaccharides, as well as α,βGlcNAc from reducing ends were identified. Despite of non-quantitative nature of MALDI TOF/TOF analysis, 1H NMR was used for an overall quantification of mannose oligosaccharides. It is based on the integration of the signal, representing the core 1,3,6-linked βMan unit bound to the reducing end GlcNAc unit. Significantly increased intensities of 1,3-linked αMan signals in 1H NMR spectra confirmed results from mass spectrometry, suggesting Man2GlcNAc1 as the most abundant structure found in urine of alpha-mannosidosis patients. Presented methods were developed to become the basis for a precise monitoring of therapy efficacy in the future.

This work was supported by Ministry of Health of the Slovak Republic under the project registration number 2019/7-CHÚSAV-4, VEGA 2/0130/18, and APVV-18-0336. This contribution is the result of the implementation of the “Technical infrastructure for biomedical research” project, ITMS 26230120008, which was supported by the Research & Development Operational Programme and funded by the ERDF.
[1] Malm D, Nilssen O (2008). Alpha-mannosidosis. Orphanet J Rare Dis.3(1):21.
[2] Beck M, Olsen KJ, Wraith JE et al. (2013). Natural history of alpha-mannosidosis: a longitudinal study. Orphanet J Rare Dis.8:88.