Introduction

Mucopolysaccharidoses (MPSs) are a group of lysosomal storage disorders (LSDs) caused by deficiency in the activity of each of 11 lysosomal enzymes required for degradation of glycosaminoglycans (GAGs). The 11 genes coding for the lysosomal enzymes are IDUA, IDS, SGSH, NAGLU, HGSNAT, GNS, GALNS, GLB1, ARSB, GUSB , and HYAL1 . Deficiency in these enzymes results in abnormal accumulation of GAGs in lysosomes of most cells leading to progressive cellular damage and multiple organ failure, which is the phenotypic hallmark of most MPS types. There is a high degree of phenotypic overlap between the seven types of MPSs which are inherited in either autosomal recessive (MPS types I, III, IV, VI, VII, and IX) or X-linked (MPS type II) (Brusius-Facchin et al. 2019).
MPSs are diagnosed through a combination of clinical phenotype and biochemical testing, such as quantitation of urinary GAGs and measurement of enzyme activity in leukocytes or fibroblasts. Measurement of GAGs using tandem mass spectrometry is a plausible way to screen for MPS patients, but it cannot provide a definitive differential diagnosis for the disease type. Biochemical characterization of the GAGs using two-dimensional electrophoresis can help with achieving a more specific differential diagnosis, but it requires enzymatic confirmation. However, performing several enzymatic assays in screen-positive patients could be time-consuming and not cost-effective. Molecular characterization of MPS genes using Sanger or next-generation sequencing is another diagnostic method that is considered as accurate as of the enzymatic assay, especially when the assignment of patients to the costly enzyme replacement therapies is taken into account. Such molecular approaches could also be beneficial for predicting the prognosis and providing genetic counseling to the family and their at-risk relatives about carrier screening and prenatal or preimplantation genetic diagnosis (Brusius-Facchin et al. 2019; Filocamo et al. 2018; Kadali et al. 2019).
Sanger sequencing of the MPS genes has been widely used for molecular diagnosis in clinically and/or biochemically diagnosed patients over the last two decades. This individualized approach, which requires exon-by-exon sequencing of the corresponding genes, might be time-consuming and less cost-effective. Furthermore, when it comes to the differential diagnoses of each MPS type, which encompass a wide range of other types of MPSs and the other LSDs, Sanger sequencing cannot be considered an effective tool. In a handful of recent research articles, next-generation sequencing (NGS) has therefore been suggested as a quick, efficient, and reliable alternative to Sanger sequencing. Next-Generation
Sequencing allows achieving a comprehensive molecular investigation of all MPS types as well as their differential diagnosis in a single test, which could be of paramount importance, particularly in patients with non-specific overlapping phenotypes. Whole exome sequencing (WES) and targeted capture NGS are two NGS-based methods that have been applied for molecular diagnosis of genetic disorders, and each has its own capabilities and limitations. While the low sensitivity of the Whole-Exome Sequencing (WES) resulting from its limited coverage could be considered a drawback, employing targeted capture NGS approach makes the technology much less error-prone due to the higher coverage and mean depth achievable in targeted regions. Additionally, the application of target panels with a larger number of genes encompassing MPS genes and their differential diagnosis can facilitate reaching a conclusive diagnosis (Cobos et al. 2015; Costa-Motta et al. 2014; Filocamo et al. 2018; Kadali et al. 2019; Wood et al. 2013).
Here, we report on the genetic investigation of a cohort of 302 patients from 289 unrelated families with the clinical and/or biochemical diagnosis of various types of MPS which is the largest cohort of Iranian patients reported so far. This report is part of a national project on “Iran MucoPolysaccharidosis REdiagnosis Study (IMPRESsion)”, which has been carried out to determine the genetic profile of MPSs in Iran. The recruited patients have been investigated using either Sanger sequencing or NGS panel, targeting 594 genes associated with inborn errors of metabolism. Moreover, in search of possible founder mutations and/or region-specific pathogenic variants, the geographical origin of the studied patients has also been collected. Such data can facilitate mutation detection of patients originating from the designated areas, and propose a stepwise diagnostic approach starting from testing region-specific pathogenic variants, followed by more comprehensive testing by NGS-based methods.