Analysis of Glycosaminoglycans in Cerebrospinal Fluid from Patients with Mucopolysaccharidoses by Isotope-Dilution Ultra-Performance Liquid Chromatography–Tandem Mass Spectrometry

• Published in: Clinical chemistry (Baltimore, Md.) Vol. 57; no. 7; pp. 1005 - 1012
• Main Authors: Zhang, Haoyue, Young, Sarah P, Auray-Blais, Christiane, Orchard, Paul J, Tolar, Jakub, Millington, David S
• Format: Journal Article
• Language: English
• Published: Washington, DC American Association for Clinical Chemistry 01.07.2011; Oxford University Press
• Subjects: Analytical, structural and metabolic biochemistry; Aqueous solutions; Biological and medical sciences; Biomarkers - cerebrospinal fluid; Calibration; Child; Chondroitin Sulfates - cerebrospinal fluid; Chondroitin Sulfates - standards; Chromatography; Chromatography, High Pressure Liquid; Dermatan Sulfate - cerebrospinal fluid; Dermatan Sulfate - standards; Deuterium; Dimerization; Disease; Enzyme Replacement Therapy; Enzymes; Fundamental and applied biological sciences. Psychology; Hematopoietic Stem Cell Transplantation; Heparitin Sulfate - cerebrospinal fluid; Heparitin Sulfate - standards; Hexosamines - cerebrospinal fluid; Humans; Indicator Dilution Techniques; Injections, Intravenous; Injections, Spinal; Investigative techniques, diagnostic techniques (general aspects); Ionization; Liquid chromatography; Mass spectrometry; Medical sciences; Molecular biophysics; Mucopolysaccharidosis I - cerebrospinal fluid; Mucopolysaccharidosis I - therapy; Reference Standards; Reference Values; Tandem Mass Spectrometry; Uronic Acids - cerebrospinal fluid; Performance evaluation; Human; Diseases of the osteoarticular system; Isotope dilution; Metabolic diseases; Patient; Biochemistry; Liquid chromatography; Mucopolysaccharidosis; Cerebrospinal fluid; Enzymopathy; Genetic disease; Mass spectrometry MS/MS; Clinical biology; Glycosaminoglycan; Carbohydrate; Molecular biology; Performance
• ISSN: 0009-9147; 1530-8561; 1530-8561
• DOI: 10.1373/clinchem.2010.161141

New therapies for the treatment of mucopolysaccharidoses that target the brain, including intrathecal enzyme replacement, are being explored. Quantitative analysis of the glycosaminoglycans (GAGs) that accumulate in these disorders is required to assess the disease burden and monitor the effect of therapy in affected patients. Because current methods lack the required limit of quantification and specificity to analyze GAGs in small volumes of cerebrospinal fluid (CSF), we developed a method based on ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Samples of CSF (25 μL) were evaporated to dryness and subjected to methanolysis. The GAGs were degraded to uronic acid-N-acetylhexosamine dimers and mixed with internal standards derived from deuteriomethanolysis of GAG standards. Specific dimers derived from heparan, dermatan and chondroitin sulfates (HS, DS and CS) were separated by UPLC and analyzed by electrospray ionization MS/MS using selected reaction monitoring for each targeted GAG product and its corresponding internal standard. CSF from control pediatric subjects (n = 22) contained <0.38 mg/L HS, 0.26 mg/L DS, and 2.8 mg/L CS, whereas CSF from patients with Hurler syndrome (n = 7) contained concentrations of DS and HS that were at least 6-fold greater than the upper control limits. These concentrations were reduced by 17.5% to 82.5% after allogeneic transplantation and treatment with intrathecal and intravenous enzyme replacement therapy. The method described here has potential value in monitoring patients with mucopolysaccharidoses receiving treatment targeted to the brain.