Detection of glucosylsphingosine in dried blood spots for diagnosis of Gaucher disease by LC-MS/MS.

Abstract

INTRODUCTION:

Gaucher disease (GD) is caused by a deficiency of β-glucosidase (GCase), leading to accumulation of glucosylceramide (GlcC) and glucosylsphingosine (Lyso-Gb1). Lyso-Gb1 is a reliable biomarker for GD.

OBJECTIVES:

This study aims to develop a simple, effective and accurate method for the screening and diagnosis of GD using dried blood spot (DBS) samples.

METHODS:

Lyso-Gb1 in DBS was extracted by 50% acetonitrile aqueous solution containing isotope-labeled internal standard and analyzed using liquid chromatography tandem mass spectrometry (LC-MS/MS). A reference interval was established by analyzing samples from 277 healthy controls. Lyso-Gb1 was detected in the residual DBS samples from 142 high-risk patients with splenomegaly and/or thrombocytopenia. Based on GCase activity in DBS, samples were classified into four groups: confirmed GD patients (n = 52), GD carriers (n = 5), false positive (n = 36) and negative (n = 49).

RESULTS:

The optimized Lyso-Gb1 assay showed intra- and inter-assay variations ranged between 2.0%-8.2% and 3.8%-10.2%, respectively. Accuracies ranged from 93.5% to 112.6%. The lowest limit of quantification was 1 ng/mL. The normal reference interval of Lyso-Gb1 in DBS ranged from 2.1 to 9.9 ng/mL. Among the 142 subjects, except for one GD patient (Lyso-Gb1 > 2500 ng/mL), the Lyso-Gb1 concentrations in 51 GD patients ranged from 190.5 to 2380.6 ng/mL (the median 614.8 ng/mL). Also, one negative patient was found to have an elevated Lyso-Gb1 level (684.5 ng/mL), while the other patients were normal. The negative case was then confirmed to be an atypical GD patient with a c.1091A > G (p.Y364C) homozygous variant in PSAP gene by next generation sequencing.

CONCLUSIONS:

The optimized method to determine Lyso-Gb1 in DBS was demonstrated as a useful tool for the screening and diagnosis of GD.

• Authors: Chengfang Tang, Xuefang Jia, Fang Tang, Sichi Liu, Xiang Jiang, Xiaoyuan Zhao, Huiying Sheng, Minzhi Peng, Li Liu, Yonglan Huang
• Journal: Clinical biochemistry (Clin Biochem) Vol. 87 Pg. 79-84 (Jan 2021) ISSN: 1873-2933 [Electronic] United States
• PMID: 33188770 (Publication Type: Journal Article)
• Copyright: Copyright © 2020 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Chemical References

• Biomarkers
• Psychosine
• sphingosyl beta-glucoside
• beta-Glucosidase

Topics

• Adolescent
• Adult
• Biological Assay
• Biomarkers (blood)
• Case-Control Studies
• Child
• Child, Preschool
• Chromatography, Liquid (methods)
• Dried Blood Spot Testing (methods)
• Female
• Gaucher Disease (blood, diagnosis)
• Humans
• Infant
• Infant, Newborn
• Male
• Middle Aged
• Psychosine (analogs & derivatives, blood)
• Reference Values
• Tandem Mass Spectrometry (methods)
• Young Adult
• beta-Glucosidase (metabolism)