Preliminary Data of the First Year of Newborn Screening for Metachromatic Leukodystrophy (MLD) in Lombardy

Metachromatic leukodystrophy (MLD) is a rare, autosomal recessive lysosomal storage disorder caused by a deficiency of the enzyme arylsulfatase A (ARSA, MIM #250100), which leads to progressive demyelination in the central and peripheral nervous systems, resulting in severe neurodegeneration and premature death. With the recent approval of the first ex vivo gene therapy for MLD, newborn screening (NBS) programs have begun to implement pilot studies for early detection, as identifying affected individuals during the pre-symptomatic therapeutic window is crucial. We present the analytical workflow and the NBS algorithm developed during the first year of MLD screening in Lombardy. The screening strategy comprised a first-tier test (1TT) using mass spectrometry to quantify sulfatide concentrations, a second-tier test (2TT) measuring ARSA enzymatic activity, and a third-tier test (3TT) based on whole-exome sequencing. A total of 16,130 newborns were screened for MLD. Of these, 6.41% required retesting (1TT), 1.53% underwent duplicate 2TT, and 0.36% proceeded to 3TT. No neonates were recalled, and no cases of MLD were identified through the screening program. The first-year implementation of MLD NBS in Lombardy demonstrates that a mass spectrometry-based sulfatide assay combined with a multi-tiered screening algorithm is feasible and reliable. Incorporating genetic analysis alongside expanded validation of additional sulfatide species may enhance specificity, reduce false positives, and facilitate timely identification of infants affected by MLD.