Newborn Screening for Pompe Disease by Measuring Acid α-Glucosidase Activity Using Tandem Mass Spectrometry

Abstract

background: Pompe disease, caused by the deficiency of acid α-glucosidase (GAA), is a lysosomal storage disorder that manifests itself in its most severe form within the first months of life. Early detection by newborn screening is warranted, since prompt initiation of enzyme replacement therapy may improve morbidity and mortality. We evaluated a tandem mass spectrometry (MS/MS) method to measure GAA activity for newborn screening for Pompe disease. methods: We incubated 3.2-mm punches from dried blood spots (DBS) for 22 h with the substrate [7-benzoylamino-heptyl)-{2-[4-(3,4,5-trihydroxy-6-hydroxymethyl-tetrahydro-pyran-2-yloxy)-phenylcarbamoyl]- ethyl}-carbamic acid tert-butyl ester] and internal standard [7-d5-benzoylamino-heptyl)-[2-(4-hydroxy-phenylcarbamoyl)-ethyl]-carbamic acid tertbutyl ester]. We quantified the resulting product and internal standard using MS/MS. We assessed inter- and intrarun imprecision, carryover, stability, and correlation between enzyme activities and hematocrit and punch location and generated a Pompe disease–specific cutoff value using routine newborn screening samples. results: GAA activities in DBS from 29 known Pompe patients were <2 μmol/h/L. GAA activities in routine newborn screening samples were [mean (SD)] 14.7 (7.2) μmol/h/L (n = 10 279, median 13.3, 95% CI 14.46–14.74 μmol/h/L) and in normal adult samples 9.3 (3.3) μmol/h/L (n = 229, median 9, 95% CI 8.88–9.72 μmol/h/L). GAA activity was stable for 28 days between 37 °C and −80 °C. Carryover could not be observed, whereas intrarun and interrun imprecision were <10%. The limit of detection was 0.26 μmol/h/L and limit of quantification 0.35 μmol/h/L. conclusions: The measurement of GAA activities in dry blood spots using MS/MS is suitable for high-throughput analysis and newborn screening for Pompe disease.

Newborn Screening for Lysosomal Storage Disorders in Hungary

“…Assuming all four patients were affected, a frequency of 1:2,570 can be derived. In our study, we found similar mean enzyme activity and standard deviation in whole sample cohort as was found in the Austrian study (Dajnoki et al 2008). However, applying a cutoff value of 2 mmol/l/h would have missed three affected Pompe patients in our study.…”

“…Our frequency for Pompe of 1:4,447 at best to 1:20,012 is higher compared to the Taiwanese population. In an anonymous Austrian pilot study for NBS of Pompe disease only 4 (0.039%) of 10,279 patients would have been recalled for retesting using a cutoff of 2 mmol/l/h (Dajnoki et al 2008). Assuming all four patients were affected, a frequency of 1:2,570 can be derived.…”

“…Several pilot studies for single diseases or multiple LSDs with fluorometric and mass spectrometric methods were carried out in recent years (Dajnoki et al 2008;Orsini et al 2009;Spada et al 2006). Results suggest that some LSDs are quite frequent.…”

Newborn Screening for Hunter Disease: A Small-Scale Feasibility Study

“…Based on this result, the recall rate would be 0.07%. Although this number remains to be confirmed by larger studies, it is comparable to the results obtained in studies for other lysosomal storage disorders: Pompe disease (0.039%) (Dajnoki et al 2008) and Krabbe disease (0.06%) (Orsini et al 2009). A higher recall rate (0.82%) was reported by Chien et al for Pompe NBS in Taiwan, but this likely results from a common a-glucosidase variant with low enzyme activity present in the Asian population (Chien et al 2008).…”

“…Early intervention requires identification of patients at a very early age, before clinical features have become evident, i.e., by newborn screening (NBS) (Marsden and Levy 2010;Zhou et al 2011). NBS has been reported for a number of lysosomal storage disorders including Pompe disease (Chien et al 2008;Dajnoki et al 2008), Fabry disease (Dajnoki et al 2010), and Krabbe disease (Orsini et al 2009), but not for MPS II. Enzymatic assays for measuring the IDS activity in dried blood spots have been described in fluorometric (Oemardien et al 2011;Tolun et al 2012) and in tandem mass spectrometric (Wolfe et al 2011) platforms.…”