1
Centre of Glycomics, Institute of Chemistry, Slovak Academy of Sciences, 84538 Bratislava, Slovakia
2
Centre for Inherited Metabolic Disorders, National Institute of Children's Diseases, 83340 Bratislava, Slovakia
3
Department of Paediatrics, Faculty of Medicine, Comenius University in Bratislava, and National Institute of Children's Diseases, 83340 Bratislava, Slovakia
Background: Alpha-mannosidosis is a rare lysosomal storage disorder,
caused by decreased activity of -D-mannosidase. This enzyme is involved
in the hydrolysis of mannosidic linkages in N-linked oligosaccharides. Due to the
mannosidase defect, undigested mannose-rich oligosaccharides (Man2GlcNAc -
Man9GlcNAc) accumulating in cells are excreted in large quantities in urine.
Methods: In this work, we determined the levels of urinary mannose-rich
oligosaccharides in a patient subjected to novel enzyme replacement therapy.
Urinary oligosaccharides were extracted using solid phase extraction (SPE),
labeled by fluorescent tag 2-aminobenzamide, and quantified by high-performance
liquid chromatography (HPLC) with fluorescence detector (FLD). The identity of
peaks was determined by matrix-assisted laser desorption/ionization time-of-flight/time-of-flight (MALDI-TOF/TOF) mass spectrometry. In addition, the levels of urinary
mannose-rich oligosaccharides were also quantified by H nuclear magnetic
resonance (NMR) spectroscopy. The data were analyzed using one-tailed paired
t-test and Pearson’s correlation tests. Results: Compared to
levels before the administration of therapy, an approximately two-folds decrease
in total mannose-rich oligosaccharides after one month of treatment was observed
by NMR and HPLC. After four months, an approximately ten-folds significant
decrease in total urinary mannose-rich oligosaccharides was detected, suggesting
therapy effectiveness. A significant decrease in the levels of oligosaccharides
with 7–9 mannose units was detected by HPLC. Conclusions: The
application of both HPLC-FLD and NMR in quantification of oligosaccharide
biomarkers is a suitable approach for monitoring of therapy efficacy in
alpha-mannosidosis patients.
Keywords
alpha-mannosidosis
HPLC
mass spectrometry
NMR
velmanase alpha
Funding
2019/7-CHÚSAV-4/Ministry of Health of Slovak Republic
VEGA 2/0060/21/Ministry of Health of Slovak Republic
Maroš Krchňák,
Rebeka Kodríková,
Mária Matulová,
Marek Nemčovič,
Iveta Uhliariková,
Jaroslav Katrlík,
Anna Šalingová,
Anna Hlavatá,
Katarína Juríčková,
Peter Baráth,
Ján Mucha,
Zuzana Pakanová. The Application of HPLC-FLD and NMR in the Monitoring of Therapy Efficacy in Alpha-Mannosidosis. Front. Biosci. (Landmark Ed)2023, 28(2), 39.
https://doi.org/10.31083/j.fbl2802039
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Model Title
Fig. 1.
Selected region of H NMR spectra in which anomeric H1
signals of saccharides appear. The diagnostic signal of 1,3,6-linked mannose
units in -configuration (1,3,6-Man) in mannose-rich
oligosaccharides, accumulated in the urine of alpha-mannosidosis patients, is
highlighted by red color. GlcNAc and GlcNAc –
reducing end glucosamine in - and -configuration; Man,
mannose; HOD, residual signal of water.
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