탠덤 질량분석기를 이용한 헌터 증후군과 연관된 iduronate-2-sulfatase 효소 활성도 직접 검사법 개발

Abstract

Introduction: Mucopolysaccharidosis type II (MPS II), also known as Hunter syndrome, is caused by a deficiency in iduronate-2-sulfatase. MPS II is a chronic, progressive lysosomal storage disorder that affects multiple organ systems but is challenging to diagnose during the early stages. Here we developed and evaluated the performance of ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) with a commercially available substrate for the detection of MPS II.

Methods: 4-Methylumbelliferyl α-L-idopyranosiduronic acid 2-sulfate (IDS-S) was used as a substrate for IDS, and its enzymatic product, 4-methylumbelliferyl α-L-idopyranosiduronic acid (IDS-P), was directly measured by UPLC-MS/MS. We determined the precision of our enzyme assay and the effects of sample amounts and incubation time on the results. Dried blood spots (DBSs) of 26 normal newborns and two patients with MPS II were analyzed.

Results: The intra- and inter-assay precisions were 7.9–10.5% and 4.8–10.2%, respectively. The amount of product obtained was proportional to the number of DBSs

however, a slight flattening was observed in the product versus DBS curve for more than one DBS. For our enzyme assay, the amount of product obtained increased linearly with the incubation period from 0 to 15 h. The enzyme activities measured in the DBSs were consistently lower in patients with MPS II than in normal newborns.

Conclusions: The performance of our enzyme assay was generally acceptable. In addition, it may be possible to simultaneously test multiple enzymes related to lysosomal storage diseases. To the best of our knowledge, this is the first report describing the use of MS/MS for the diagnosis of MPS II with a commercially available substrate. Our method provides a rapid, inexpensive, effective screening tool for the diagnosis of MPS II worldwide.