Late‐onset mucopolysaccharidosis type IIIA mimicking Usher syndrome

Mucopolysaccharidosis type IIIA (MPS IIIA or Sanfilippo syndrome type A) is an autosomal recessive lysosomal storage disorder caused by pathogenic variants in the SGSH gene encoding N‐sulfoglucosamine sulfohydrolase, an enzyme involved in the degradation of heparan sulfate. MPS IIIA is typically characterized by neurocognitive decline and hepatosplenomegaly with childhood onset. Here, we report on a 53‐year‐old male subject initially diagnosed with Usher syndrome for the concurrence of retinitis pigmentosa and sensorineural hearing loss. Clinical exome sequencing identified biallelic missense variants in SGSH, and biochemical assays showed complete deficiency of sulfamidase activity and increased urinary glycosaminoglycan excretion. Reverse phenotyping revealed left ventricle pseudo‐hypertrophy, hepatosplenomegaly, bilateral deep white matter hyperintensities upon brain MRI, and decreased cortical metabolic activity by PET‐CT. On neuropsychological testing, the proband presented only partial and isolated verbal memory deficits. This case illustrates the power of unbiased, comprehensive genetic testing for the diagnosis of challenging mild or atypical forms of MPS IIIA.

lysosomal storage disorder caused by deficiency of N-sulfoglucosamine sulfohydrolase, an enzyme involved in the catabolism of heparan sulfate (HS) (Andrade et al., 2015).N-sulfoglucosamine sulfohydrolase is encoded by the SGSH gene (Scott et al., 1995).MPS IIIA is a neurodegenerative disease with typical onset in childhood, presenting with neurocognitive decline, hepatosplenomegaly, and variable systemic manifestations, including musculoskeletal abnormalities, cardiovascular problems, and hearing loss (Andrade et al., 2015;Shapiro et al., 2017).
The mildest end of the spectrum includes attenuated and adult-onset phenotypes with mild cognitive deficit.In only a few cases lack of neuronopathic features was reported (Nijmeijer et al., 2019;Van Hove et al., 2003).Here, we report on an atypical presentation of MPS IIIA in an individual presenting with retinitis pigmentosa (RP) and sensorineural hearing loss (SNHL) that led to the clinical diagnosis of Usher syndrome until his 50s when clinical exome sequencing revealed biallelic missense variants in SGSH.Post-genotyping phenotyping uncovered several subclinical abnormalities of MPS IIIA.

| METHODS
Ethics statement and detailed methods of exome sequencing can be found in Supporting Information.

| CASE REPORT
A white male subject was referred to the eye clinic at the age of 29 years for RP and SNHL that led to the clinical diagnosis of Usher syndrome.He was the third of five siblings born to consanguineous (first cousins) healthy parents.A younger sister died at 7 months of age due to sudden infant death syndrome.The other three siblings did not have any vision or hearing complaints (Figure 1a).The proband was born at term of uncomplicated pregnancy with normal perinatal history.He achieved developmental milestones on time and attended school until the age of 18 years without any learning disabilities.At 29 years because of night-blindness, he had an ophthalmological examination that showed a best-corrected visual acuity (BCVA) of 20/30 in the right eye (sphere À3) and 20/30 in the left eye (sphere À2.5 = cylinder À0.5 alpha 90 ).The Ishihara color vision test revealed achromatopsia.Intraocular pressure was 14 mmHg in both eyes, and lenses were clear under biomicroscopy.Fundus examination revealed dystrophy of the retinal pigment epithelium (RPE) with bone spicule-shaped pigment deposits within and beyond the vascular arcades, while the far-periphery had a normal appearance (Figure 2a).Fundus autofluorescence (FAF) imaging indicated reduced autofluorescence, more evident along the vascular arcades (Figure 2b).Optical coherence tomography (OCT) showed reduced  2c).The Goldmann visual field (GVF) test with a III4e target showed a constricted visual field with ring scotoma (Figure 2d).In full-field electroretinography (ERG), scotopic responses were below the noise level and the photopic ones were subnormal (data not shown).Taken together, these observations led to a diagnosis of pericentral RP according to Hamel's criteria (Hamel, 2006).On annual follow-up visits, the retinal degeneration progressively worsened, and over a 20-year-period led to a visual acuity of hand motion and non-recordable ERGs (data not shown).
F I G U R E 2 (a) Fundus color image revealed a waxy pallor of the optic disc with a circumpapillary atrophy, and bone spicule-shaped pigment deposits within and beyond the vascular arcades, while the far-periphery had a normal appearance.(e) Pure tone audiometry in the right ear (in red) and left ear (in blue) at the frequency range between 250 and 8000 Hz showing mild, bilateral, sensorineural hearing loss, which is slightly more evident at higher frequencies.AC, air conduction; BC, bone conduction.(f-h) Brain MRI of the proband in T2 FLAIR revealing bilateral para-trigonal deep white matter hyperintensity (white arrows) which extends to the posterior portion of the corona radiata and to the posterior portion of the left internal capsule (shown in axial (f), coronal (g), and sagittal (h) images).(i) 18 F-FDG brain PET scan showing symmetrical reduction of 18 F-FDG in occipital region (white arrows) and thalamic hypometabolism (red arrows).Asymmetrical distribution of the tracer was observed in parietal, frontotemporal, and temporal regions (orange arrows).
The proband also had complains of hearing difficulties and at the age of 29 years an audiometric test showed mild bilateral, symmetric SNHL, with a pure tone average (PTA) of 30 dB in the frequency range 500-4000 Hz (Figure 2e).Impedance testing produced normal Type "A" tympanogram bilaterally with ipsilateral and contralateral stapedial reflexes across the entire frequency range (data not shown).
Regular audiological follow-up visits (the most recent of which at the age of 54 years) showed that the mild hearing loss was nonprogressive.
He never reported any memory problems, cognitive or behavioral issues.At the most recent physical exam, when he was 53 years-old, his weight was 72.5 kg (À1.43 SD), his height was 150 cm (À3.6 SD) and his occipito-frontal circumference was 57 cm (+2.8 SD).He was non-dysmorphic and had no hepatosplenomegaly on physical exam.His neurological exam was normal except for minimal cerebellar features, including mild telekinetic tremor (left>right) and dysdiadochokinesia.
Clinical exome sequencing did not detect causative variants in Usher syndrome genes nor in other genes causing either RP or SNHL, but it revealed two missense variants in the SGSH gene, validated by Sanger sequencing.Each variant was detected in one of the parents (Figure 1b).Among the three living siblings, II:2 and II:5 were each heterozygous for one of the two variants (p.Leu146Pro), whereas II:1 did not carry either of them (Figure 1a).Additional variants considered after the filtering steps are shown in Table S1.The p.(Arg182His) variant (NM_001352921: c.545G > A) replaces a highly conserved arginine residue, had an allele frequency of 1.6 Â 10 À5 in gnomAD (i.e., found in 4 out of 244,336 alleles and only at the heterozygous state), and a Combined Annotation Dependent Depletion (CADD; http://cadd.gs.washington.edu)score of 34.This variant is annotated in ClinVar as pathogenic (Variation ID: VCV001071734.3)and had been previously reported in patients with mild or non-neuronopathic phenotypes (Nijmeijer et al., 2019), while an Arginine-to-Cysteine substitution of the same codon was found in several patients with typical MPS IIIA (Di Natale et al., 1998;Valstar et al., 2010;Zanetti et al., 2019).The p.Leu146Pro variant (NM_001352921: c.437 T > C) had an allele frequency of 4 Â 10 À6 in gnomAD (i.e.found in 1 out of 250,638 alleles, only in the heterozygous state), and displayed a CADD score of 23.It was annotated as VUS in ClinVar (Variation ID: VCV000552534.3)but can be classified as likely pathogenic based on the ACMG criteria (Richards et al., 2015) PM1, PM2, PM3, PP3, PP4, and PP5.This variant was previously identified in homozygous state in an Italian MPS IIIA patient with a severe phenotype (Di Natale et al., 1998).The p.Leu146Pro substitution appears to interfere with H-bonding to Ser144 and helix α5 stability of the protein (Sidhu et al., 2014).Both variants were absent in almost 4000 whole or clinical exomes of Italian patients of our internal database.The enzymatic assay on white blood cells showed complete absence of heparin sulfamidase activity (0 nmol/mg/17 h; normal values 4.3 ± 2.8), and the urinary glycosaminoglycans (uGAGs) were higher than normal (50.2 mg GAG/g creatinine; normal range for age: 3-30 mg GAG/g creatinine).
Clinical manifestations of MPS IIIA were then actively assessed.
An abdominal ultrasound showed hepatomegaly with increased Our case and the few reported subjects (10 cases from 5 families) with late-onset (age at diagnosis: >40 years) and attenuated MPS IIIA (Nijmeijer et al., 2019;Van Hove et al., 2003) all showed very lowto-absent sulfamidase activity in the same range of severe infantile forms (Table S2), supporting the lack of correlation between residual enzyme activity and clinical severity (Kubaski et al., 2020).In contrast, the increased concentrations of uGAGs were milder (average about 5.4-fold higher than the upper limit of normal) (Nijmeijer et al., 2019;Van Hove et al., 2003) (Table S2) compared to infantile forms (about 15-fold higher than the upper limit of normal) (Kubaski et al., 2020).
The first symptoms that prompted the diagnosis of mild MPS IIIA were vision problems.Patients with mucopolysaccharidoses often have ocular manifestations such as retinopathy, optic nerve abnormalities, corneal clouding, and glaucoma (Ashworth et al., 2006;Fahnehjelm et al., 2012;Ferrari et al., 2011).In MPS IIIA, the most frequent ocular feature is a progressive retinopathy (Muschol et al., 2022;Wilkin et al., 2016), that is also found in all patients with late-onset forms (Nijmeijer et al., 2019), suggesting that retinal homeostasis is particularly sensitive to SGSH deficiency.Photoreceptor dysfunction was indeed an early sign of neuronal symptoms also in a mouse model of MPS IIIA (Intartaglia et al., 2020).Although retinopathy was apparently isolated at diagnosis, post-genotyping evaluations revealed multi-system subclinical involvement in most individuals with late-onset MPS IIIA, including our case.Although common in all types of MPS, hearing loss was found only in our case (Buhrman et al., 2014;da Silveira et al., 2018).
The mechanisms underlying the milder manifestations in the face of complete deficiency of enzyme activity remain unclear, but it is possible that genetic modifiers, such as genes involved in synthesis or degradation of GAGs, may be implicated in attenuating disease severity.Larger cohorts of mild MPS IIIA patients are needed to investigate these hypotheses.

AUTHOR CONTRIBUTIONS
Alessandro De Falco and Marianthi Karali equally contributed to the design of the work, drafted the manuscript.Sandro Banfi and Nicola Brunetti-Pierri supervised the study, critically revised the manuscript.
Alessandro De Falco, Marianthi Karali, Chiara Criscuolo, Francesco Testa, Maria Rosaria Barillari, Margherita Scarpato, Valeria Gaudieri, Alberto Cuocolo, Anna Russo, Vincenzo Nigro, and Francesca Simonelli contributed to collection and analysis of clinical data.All authors have read and agreed to the published version of the manuscript.
(b) Fundus autofluorescence imaging of the boxed area in (a) showing hypoautofluorescence at the macular region extending beyond the vascular arcades.(c) Horizontal spectral-domain optical coherence tomography (SD-OCT) showing reduced macular thickness and RPE dystrophy.(d) Visual field test showing constriction of the III4e isopter.
105 Â 50 mm).The gallbladder was normal in shape but with two stones.The echocardiogram revealed pseudo-hypertrophy of the left ventricle and thickening of the right ventricle.Brain MRI showed bilateral paratrigonal deep white matter hyperintensity extending to the posterior portion of the corona radiata and the left internal capsule (Figure 2f-h).Brain PET-CT detected decreased cortical metabolic activity bilaterally in the occipital, parietal, fronto-temporal, and left temporal areas (Figure 2i).Within the context of normal cognitive function, neuropsychological tests showed only partial and isolated verbal memory deficits, that might be related at least in part to the visual impairment.4 | DISCUSSION We report on an adult individual presenting with RP and SNHL that initially suggested a clinical diagnosis of Usher syndrome but who was later recognized to be affected by MPS IIIA.Clinical phenotyping revealed previously unrecognized systemic and asymptomatic manifestations, including hepatosplenomegaly, pseudo-hypertrophy of the left ventricle, brain white matter hyperintensities, and diffuse metabolic cerebral abnormalities.This case illustrates the challenge of diagnosing mild adult-onset forms of MPS IIIA and the power of unbiased genome sequencing for the diagnosis of atypical disease presentations.Although enzyme replacement therapy (ERT) is not yet available for MPS IIIA (Suarez-Guerrero et al., 2016), early diagnosis of MPS IIIA is important to reduce disease-associated risks and to allow prompt initiation of ERT once it will become available.