UPLC-MS/MS High-Risk Screening for Sphingolipidoses Using Dried Urine Spots

Abstract

Background: Early detection of sphingolipidoses is crucial to prevent irreversible complications and improve patient outcomes. The use of urine samples dried on filter paper (DUS) is a non-invasive strategy that simplifies the collection, storage, and shipping of samples compared to using liquid urine specimens. Objectives: (1) Develop and validate a multiplex ultra-performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS) methodology using DUS to quantify twenty-one lysosphingolipids normalized to creatinine for eight different sphingolipidoses. (2) Establish normal reference values to evaluate the clinical utility of the methodology. Methods: Samples were eluted from a 5 cm filter paper disk (~1 mL of urine) and extracted on Oasis MCX solid-phase extraction cartridges prior to injection in the UPLC-MS/MS system. Results: Urinary lysosphingolipids were stable on DUS at −80 °C and −30 °C for 117 days, at 21.5 °C and 4 °C for at least 26 days, and at 35 °C for 3 days. Globotriaosylsphingosine, glucosylsphingosine, and their analogs were elevated in patients with Fabry disease and Gaucher disease, respectively, compared to controls (p-value < 0.0001). The analysis of related analog profiles suggests a better overall reliability in detecting patients early, especially for Fabry patients. Conclusions: This approach is feasible and might be useful for the early detection, monitoring, and follow-up of patients with sphingolipidoses.