Proteomic Discovery and Development of a Multiplexed Targeted MRM-LC-MS/MS Assay for Urine Biomarkers of Extracellular Matrix Disruption in Mucopolysaccharidoses I, II, and VI

Abstract

The mucopolysaccharidoses (MPS) are lysosomal storage disorders that result from defects in the catabolism of glycosaminoglycans. Impaired muscle, bone, and connective tissue are typical clinical features of MPS due to disruption of the extracellular matrix. Markers of MPS disease pathology are needed to determine disease severity and monitor effects of existing and emerging new treatments on disease mechanisms. Urine samples from a small cohort of MPS-I, -II, and -VI patients (n = 12) were analyzed using label-free quantative proteomics. Fifty-three proteins including many associated with extracellular matrix organization were differently expressed. A targeted multiplexed peptide MRM LC-MS/MS assay was used on a larger validation cohort of patient samples (MPS-I n = 18, MPS-II n = 12, MPS-VI n = 6, control n = 20). MPS-I and -II groups were further subdivided according to disease severity. None of the markers assessed were altered significantly in the mild disease groups compared to controls. β-galactosidase, a lysosomal protein, was elevated 3.6-5.7-fold significantly (p < 0.05) in all disease groups apart from mild MPS-I and -II. Collagen type Iα, fatty-acid-binding-protein 5, nidogen-1, cartilage oligomeric matrix protein, and insulin-like growth factor binding protein 7 concentrations were elevated in severe MPS I and II groups. Cartilage oligomeric matrix protein, insulin-like growth factor binding protein 7, and β-galactosidase were able to distinguish the severe neurological form of MPS-II from the milder non-neurological form. Protein Heg1 was significantly raised only in MPS-VI. This work describes the discovery of new biomarkers of MPS that represent disease pathology and allows the stratification of MPS-II patients according to disease severity.

Mucopolysaccharidosis Type I Newborn Screening: Best Practices for Diagnosis and Management

“…Kingma et al 27 developed an algorithm to predict clinical severity in MPS I based on genotyping and an optimized IDUA assay, however, this assay is not routinely available. In a preliminary proteomic study, the lysosomal protein b-galactosidase was elevated 3.6-to 5.7-fold in severe but not attenuated MPS I, 28 and 1 study has reported that serum heparin-cofactor IIthrombin complex levels can differentiate patients with severe MPS I from patients with attenuated MPS I. 29 Neither of these studies evaluated the utility of these biomarkers in the neonate or early infant, and, thus, their utility for use in NBS is limited.…”

Mucopolysaccharidosis Type II: One Hundred Years of Research, Diagnosis, and Treatment

“…As for MPSs, ideal biochemical biomarkers would be specific to a particular type or group of MPSs, help to discriminate more severe from less severe phenotypes, correlate with neurological involvement, respond to therapy, and be easily quantified [244]. In MPSs, primary biochemical biomarkers are represented by the primary storage material; for MPS II, they are partially degraded HS and DS fragments that accumulate in the lysosomes and extracellular matrices (ECM) and are secreted into the bloodstream and then excreted in the urine [245].…”

Free urinary glycosylated hydroxylysine as an indicator of altered collagen degradation in the mucopolysaccharidoses

“…One key early pathological feature is the disruption of the ECM. [13][14][15] The resulting consequence of this disruption needs to be further understood.…”