Benjamin Ganne
email: benjamin.ganne@chu-limoges.fr

Benjamin Dauriat

Laurence Richard

Foudil Lamari

Karima Ghorab

Laurent Magy

Mehdi Benkirane

Alexandre Perani

Valentine Marquet

Patrick Calvas

Benjamin Ganne1
email: benjamin.ganne.92@gmail.com

Benjamin Dauriat1

Laurence Richard2

Foudil Lamari3

Karima Ghorab2

Laurent Magy2

Mehdi Benkirane4

Alexandre Perani1

Valentine Marquet1

Patrick Calvas5

Catherine Yardin1

Sylvie Bourthoumieu1

GM2 gangliosidosis AB variant: first case of late onset and review of the literature

ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Introduction

Gangliosides are amphiphilic molecules, discovered by Ernst Klenk in the 1930s [START_REF] Sandhoff | Gangliosides and gangliosidoses: principles of molecular and metabolic pathogenesis[END_REF], and more than 180 different types have been described in vertebrates [START_REF] Leal | GM2 Gangliosidoses: clinical features, pathophysiological aspects, and current therapies[END_REF].

These glycosphingolipids are metabolized through the trans-Golgi network from a disaccharide lactosylceramide core (LacCer), on which N-acetylgalactosamine (GalNac) and galactose (Gal) can be added. They differ by the length of the fatty acid and saccharide chain as well as the number and position of the N-acetylneuraminic substrate (NANA or Neu5Ac).

Their degradation into ceramide requires lysosomal hydrolases and lipid-binding protein, the "sphingolipid activator proteins" (SAPs).

In humans, gangliosides are mainly found in the nervous system, and concentrated in the plasma membrane of neurons, in domains called lipid rafts [3]. They are involved in different cellular functions, such as intercellular recognition, signal transduction, myelin stabilization, or even cell adhesion [START_REF] Yu | Structures, biosynthesis, and functions of gangliosides-an overview[END_REF][START_REF] Schnaar | The biology of gangliosides[END_REF].

GM2 represents only 4-5% of brain gangliosides, and their degradation into GM3 requires hexosaminidase A (HexA), a heterodimer composed of an α subunit, encoded by HEXA (15q23, OMIM*606,869#272,800), and a β subunit, encoded by HEXB (5q13.3, OMIM*606,873#268,800). Dimerization of α subunits (αα) or β subunits (ββ) forms respectively hexosaminidase S (HexS) and hexosaminidase B (HexB). These homodimeric hexosaminidases cannot substitute for hexosaminidase A in GM2 ganglioside degradation.

While HexA can cleave both neutral and anionic substrates, HexB can only degrade neutral substrates and catabolizes globosides into globotriaosylceramides [START_REF] Tropak | Construction of a hybrid β-hexosaminidase subunit capable of forming stable homodimers that hydrolyze GM2 ganglioside in vivo[END_REF]. HexS, however, does not participate in GM2 degradation in vivo [7,[START_REF] Mahuran | Biochemical consequences of mutations causing the GM2 gangliosidoses[END_REF].

The hydrolysis of GM2 also requires a non-enzymatic protein, the GM2 activator protein (GM2-AP), encoded by GM2A (5q33.1, OMIM*613,109#272,750). GM2-AP forms a complex with GM2 ganglioside allowing interaction between hexosaminidase A and GM2 ganglioside, its membrane-embedded substrate. Pathogenic mutations of HEXA, HEXB, and GM2A are responsible for autosomal recessive diseases known as GM2 gangliosidosis, caused by excessive intralysosomal accumulation of GM2 gangliosides, especially in neuronal cells.

Mutations in HEXA, encoding the α chain of HexA, lead to Tay-Sachs disease or B variant, while mutations in HEXB, encoding the β chain, lead to deficiency of both HexA and HexB causing Sandhoff disease or O variant. Mutations of GM2A, encoding GM2-AP, lead to AB variant, which is rarely encountered and indistinguishable from Tay-Sachs disease.

Different clinical GM2 gangliosidosis subtypes have been described according to the age of onset. The infantile form is mainly characterized by hypotonia and seizures that appear in the first year, with premature death around 4-5 years [START_REF] Bley | Natural history of infantile G(M2) gangliosidosis[END_REF]. Juvenile forms have an onset between 2 and 10 years old, with ataxia, psychomotor deterioration, spasticity, and seizures, leading to death before adulthood [START_REF] King | The juvenile gangliosidoses: a timeline of clinical change[END_REF]. Late-onset forms (or adult forms) present with a heterogeneous phenotype and a better prognosis than the previously described forms, with inferior motoneuron deficiency, spinocerebellar ataxia, muscular atrophy, and psychiatric disorders [START_REF] Masingue | Natural history of adult patients with GM2 gangliosidosis[END_REF].

A particular feature of the infantile form is the presence of a macular cherry-red spot, which was the first characteristic sign described in Tay-Sachs disease, previously called "familial amaurotic idiocy" [START_REF] Tay | Symmetrical changes in the region of the yellow spot in each eye of an infant[END_REF].

AB variant is the rarest form, with less than 30 cases described in the literature, with no reported adult forms [START_REF] Sandhoff | Gangliosides and gangliosidoses: principles of molecular and metabolic pathogenesis[END_REF].

We describe a 22-year-old patient presenting a phenotype close to the late-onset form of GM2 gangliosidosis, in whom were detected two mutations in the GM2A gene, one predicted to be likely pathogenic, whereas the other was of unknown significance. We therefore studied GM2 accumulation in his plasma and fibroblasts as well as GM2A gene expression and cDNA Sanger sequencing, in order to conclude on the pathogenicity of these variants.

Patient

Proband is a young man born in 1998 from non-consanguineous parents in good health, as well as his younger brother.

At the age of 10 years, he consulted for gait disorders, length inequality of the lower limbs, calf hypotrophy, and claw toes. After a left foot arthrodesis at 16 years old, his gait worsened, leading to a clinical presentation of lower motor neuronopathy with motor deficit of the lower limb, and a spinocerebellar ataxia progressively settled.

No psychomotor development delay was noted during his childhood. He walked at 13 months, but had poor motor skills. He had no dysphagia, extra-pyramidal symptom, or sensory disorder.

At clinical examination, Achilles and patellar osteotendinous reflexes were absent, walking required assistance and was impossible on tiptoes or heels. In addition to the calf amyotrophy, there was interosseous amyotrophy, associated with slowness of writing without dyspraxia and without other motor deficit of the upper limbs. No fasciculation was noted.

A neuropsychological workup was carried out at the age of 22 and showed a minimal global alteration of cognitive functions, and slight autistic traits. The deficits were predominant in memory and executive functioning. An electromyogram was performed at the same time, showing pure motor involvement and chronic neurogenic pattern, mainly of the lower limbs, with both motor amplitude and conduction speed reduced.

A brain MRI performed at the age of 19 showed discrete atrophy of the cerebellar vermis, slight hyperintensity of posterior horn of the lateral ventricles, without cortical atrophy.

Hexosaminidase A and B enzymatic activities in blood were normal.

A NGS panel for hereditary ataxia showed two mutations in GM2A gene (NM_000405):

c.79A > T:p.Lys27*; nonsense variant inherited from his father, not reported in healthy population databases and predicted as likely pathogenic; c.415C > T:p.Pro139 Ser; missense variant, inherited from his mother, reported in the databases (rs767374072) and considered as a variant of undetermined significance (VOUS).

To establish the pathogenicity of this last variant with conflicting interpretations, we studied (1) accumulation of GM2 in our patient's fibroblasts using electron microscopy (EM) and immunofluorescence (IF), and in our patient's plasma using liquid chromatography coupled to a tandem mass spectrometer (LC-MS/MS) and (2) GM2A gene expression by RT-qPCR, completed with cDNA Sanger sequencing.

Material and methods

NGS analysis

Clinical exome sequencing was performed prior to our study using the TruSight One Expanded kit (TSOex, www.illum ina. com/ trusi ghtone) that targets approximately 6700 genes, most of them involved in human inherited diseases. Sequencing was performed using Illumina MiSeq or Next-Seq technology with 150-bp paired-end reads (Montpellier CHRU NGS platform).

Sequence alignment and variant calling were performed following the Broad Institute's Genome Analysis Toolkit (GATK) best practices, using the in-house pipeline Nenufaar (https:// github. com/ beboc he/ nenuf aar). Sequence reads were aligned against the reference hg19 human genome.

Variants were annotated with Annovar (http:// annov ar.openb ioinf ormat ics. org/) and Variant Studio (www. illum ina.com/ variantstu dio).

Copy number variation (CNV) analysis was performed with in-house algorithms that compare exon coverage ratios between patients of the same run (https:// github. com/ Mobid ic/ MobiC NV).

In silico filtering was performed on a panel of 559 genes, corresponding to the majority of known ataxia and related disorder genes, using American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) guidelines for clinical sequence interpretation [START_REF] Richards | Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology[END_REF].

Fibroblast cultures

Fibroblast cultures were performed from our patient's cryopreserved fibroblasts and from three healthy negative controls, using Chang medium (CliniSciences, Nanterre, France, Ref T105-100ML) with antibiotics (penicillin-streptomycin mixture; Sigma-Aldrich, Darmstadt, Germany, Ref P0781-20ML) in 25 cm2 flasks, under 5% CO2 at 37 °C in an incubator.

Positive controls for GM2 gangliosidosis were obtained by exposing fibroblast cultures of healthy negative controls to imipramine, a tricyclic antidepressant, causing HexA inhibition and GM2 accumulation [START_REF] Navon | Depletion of cellular beta-hexosaminidase by imipramine is prevented by dexamethasone; implications for treating psychotic hexosaminidase-A deficient patients[END_REF][START_REF] Palmeri | Imipramine induced lipidosis and dexamethasone effect: morphological and biochemical study in normal and chronic GM2 gangliosidosis fibroblasts[END_REF]. Imipramine (Sigma-Aldrich, Darmstadt, Germany, Ref I7379) was reconstituted in PBS at 37°C to obtain a final concentration of 100 μM in the culture medium.

Gene expression by RT-qPCR

RNA extraction of fibroblast cultures was performed with the miRNeasy Mini kit (Qiagen, Hilden, Germany, Ref 217,004), and reverse transcription with the QuantiTect Reverse Transcription kit (Qiagen, Hilden, Germany, Ref 205,311), according to the supplier's recommendations. cDNA of negative controls (n = 6 fibroblast cultures, 2 for each negative control) and proband (n = 4 fibroblast cultures) were amplified by real-time PCR using the Rotor-Gene SYBR Green PCR Kit (Qiagen, Hilden, Germany, Ref 204,074) and the Corbett Research RG-6000 thermal cycler (Corbett Research, Australia). Primers were designed using the Primer Blast software (National Center for Biotechnology Information, Rockville Pike, Bethesda, USA): 3 primer pairs for GM2A (one for exons 1-2, and two for exons 2-3), and one primer pair for each reference gene, all supplied by Sigma-Aldrich (sequences available on request).

All qPCRs were performed in duplicate for GM2A and in triplicate for reference genes (HPRT and GUSB). GM2A gene expression (Qn) was determined using the ΔCt method and the patient/control ratio by ΔΔCt method [START_REF] Bustin | The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments[END_REF][START_REF] Vandesompele | Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes[END_REF].

Sanger sequencing

Previous cDNA were amplified using primers targeting exons 1 and 3 of GM2A and Taq'Ozyme Purple Mix 2 (Ozyme, Paris, France, Ref OZYA007-200), and then purified with ExoSAP-IT (Ozyme, Paris, France, Ref 78,200-200-UL). The purified PCR products were sequenced bi-directionally using forward and reverse primers and BigDye Terminator v1.1 Cycle Sequencing Kit (Thermo Fisher Scientific, Waltham, USA, Ref 4,337,452). Sequenced products were purified using DyeEx 2.0 Spin Kit (Qiagen, Hilden, Germany, Ref 63,206) and then subjected to capillary electrophoresis using a 3130xl Genetic Analyzer. Sequences were analyzed with Unipro UGENE V.40.1 software (Unipro, Novosibirsk, Russia).

Ultrastructure of fibroblasts by electron microscopy (EM)

Fibroblast cultures of our patient, negative controls, and positive controls were fixed in 2.5% glutaraldehyde, stained with 2% osmium tetroxide (OsO4), and embedded in epoxy resin. The microscope was a JEOL JEM-1400 Flash coupled to a Morada G3 camera (Electron Optic Instrument, West Orange, NJ).

Immunofluorescence (IF)

Fibroblast cultures of our patient, negative controls, and positive controls were washed twice with PBS for 5 min, then fixed with 1 mL of 4% paraformaldehyde for 15 min, and rinsed twice with PBS for 5,min, and once for 10 min. Fibroblasts were permeabilized with 0.1% Triton X-100 for 2 min and then saturated with 1% BSA (bovine serum albumin) for 45,im. They were then stained overnight using a GMB28 mouse monoclonal anti-GM2 (0.5 mg/mL, EMD Milipore Corporation, Temecula, USA, Ref MABN2550-100UL) as primary antibody, at 1:100 and 1:200. The secondary antibody was an Alexa Fluor 488 Goat anti-mouse IgM μ chain (2 mg/mL, Thermo Fisher Scientific, OR, USA, Ref A21042) used at 1:1000.

Counterstaining of nuclei was performed with DAPI, and we observed our slides with a Metafer epifluorescence microscope (MetaSystems), coupled with Isis software (MetaSystems), to calculate a percentage of fluorescent cells on each.

Liquid-phase chromatography-tandem mass spectrometry assay (LC-MS/MS)

GM2 concentration in our patient's plasma was determined by LC-MS/MS, carried out by Dr Lamari's team at the Pitie-Salpetriere Hospital in Paris, France. Plasma was mixed with a mixture of deuterated gangliosides (Cd3-GM2 and Cd3-GM3) as internal standards and proteins precipitated by ethanol. After mixing and centrifugation, the supernatant was injected to an Acquity UPLC-TQD system (Waters). Gangliosides were detected and quantified in a positive mode with multiple reaction monitoring. This method presents a sensitivity and specificity of 100% for GM2 gangliosidosis diagnosis, using as cutoff 58.9 nM for GM2 34:1, and 0.032 for the GM2 34:1/GM3 34:1 ratio.

Results

Analysis by RT-qPCR did not reveal a decrease in GM2A gene expression in our patient in comparison to our negative controls, with a mean Qn at 0.459 (0.452-0.467) for our patient against 0.474 (0.461-0.483) for the negative controls. The patient/control ratio at 0.97 corresponds to a normal GM2A gene expression. Moreover, the Sanger sequencing showed that both mutated alleles were expressed, even if the nonsense mutation seemed to be at a slightly lower level on the chromatograms (Fig. 1).

EM performed on positive controls revealed numerous concentric lamellar inclusions within lysosomes, characteristic of lysosomal storage diseases [START_REF] Ferreira | Lysosomal storage diseases[END_REF]. In contrast, ultrastructure of patient's fibroblasts did not show a significant increase in these structures compared to negative controls, not allowing them to be reliably distinguished.

The average number of cells labelled by the anti-GM2 antibody (1:100 and 1:200) is 26.1% (n = 4 fibroblast cultures) for our patient, 26.8% (n = 2 fibroblast cultures) for the negative control, and 42.6% (n = 2 fibroblast cultures) for the positive control. There was no difference between our patient and negative controls (Fig. 2).

Discussion

We have described the case of a 22-year-old young man suspected to have a compound heterozygous form of GM2 gangliosidosis AB variant, with lower motor neuronopathy and spinocerebellar ataxia.

Electron microscopy and immunofluorescence did not reveal a lysosomal accumulation of GM2, compared to negative and positive controls. There was also no difference in GM2A gene expression using RT-qPCR, and both alleles were expressed. Only a liquid-phase chromatography-tandem mass spectrometry assay showed an accumulation of GM2 in our patient's plasma, corresponding to an adult form of Tay-Sachs or Sandhoff disease.

Only a few cases of AB variant gangliosidosis have been described in the literature, and almost all of them are infantile forms where the symptoms appear before the first year of life with a macular cherry-red spot, psychomotor retardation, hyperacusis, seizures, and axial hypotonia.

A review of the literature of patients with AB variant GM2 gangliosidosis with available genomic data is presented in Table 1.

The only patient who presented symptoms at 12 months of life without macular cherry-red spot, and with axial hypertonia and ataxia in addition to the above "classic" clinical signs, was compound heterozygous for a nonsense variant (p.Glu87*) and a missense variant (p.Pro55Leu) [START_REF] Martins | Atypical juvenile presentation of GM2 gangliosidosis AB in a patient compoundheterozygote for c.259G > T and c.164C > T mutations in the GM2A gene[END_REF]. The latter was previously described as pathogenic in the literature, in an Indian family, for which three homozygous relatives developed chorea in childhood (7-8 years), but without cherry-red spot [START_REF] Salih | Mutation in GM2A leads to a progressive chorea-dementia syndrome. Tremor and Other Hyperkinetic Movements[END_REF].

GM2A gene expression was reported in five of the presented patients. Four had normal GM2A expression [START_REF] Schepers | Molecular analysis of a GM2-activator deficiency in two patients with GM2-gangliosidosis AB variant[END_REF][START_REF] Schroder | Molecular genetics of GM2-gangliosidosis AB variant: a novel mutation and expression in BHK cells[END_REF][START_REF] Xie | A Cys138-to-Arg substitution in the GM2 activator protein is associated with the AB variant form of GM2 gangliosidosis[END_REF], and only the patient from Chen had no mRNA on Northern blot [START_REF] Chen | Structure of the GM2A gene: identification of an exon 2 nonsense mutation and a naturally occurring transcript with an in-frame deletion of exon 2[END_REF]. This last patient was homozygous for the nonsense mutation p.Glu54*, thus considerably shortening the final protein, compared to other homozygous patients for a missense mutation (p.Cys138Arg or p.Arg169Pro), an inframe deletion mutation (p.Lys88Del), or a frameshift mutation (p.His137Profs*34). The absence of mRNA may be explained by the presence of this premature stop codon, leading to mRNA degradation through the "Nonsense-Mediated mRNA Decay" (NMD) system [START_REF] Garcia-Moreno | Perspective in alternative splicing coupled to nonsense-mediated mRNA decay[END_REF]. However, p.Lys27* mutation in our patient's cDNA might escape the NMD system, and only homozygous nonsense mutations would lead to a complete absence of mRNA. Gene expression studies in patients with a homozygous nonsense mutation would have allowed us to verify this hypothesis, but was not performed [START_REF] Renaud | GM2-gangliosidosis, AB variant: clinical, ophthalmological, MRI, and molecular findings[END_REF].

The visible lesions on brain MRI were extremely similar between patients. Among those for whom clinical signs started at or before 12 months, two showed periventricular and basal ganglia T2 hyper-signals [START_REF] Chen | Structure of the GM2A gene: identification of an exon 2 nonsense mutation and a naturally occurring transcript with an in-frame deletion of exon 2[END_REF][START_REF] Brackmann | Rare variant of GM2 gangliosidosis through activator-protein deficiency[END_REF], three had basal ganglia hypersignal with delayed myelination [START_REF] Renaud | GM2-gangliosidosis, AB variant: clinical, ophthalmological, MRI, and molecular findings[END_REF][START_REF] Brackmann | Rare variant of GM2 gangliosidosis through activator-protein deficiency[END_REF][START_REF] Sheth | GM2 gangliosidosis AB variant: novel mutation from Indiaa case report with a review[END_REF], and only one did not show any abnormality [START_REF] İnci | Two patients from Turkey with a novel variant in the GM2A gene and review of the literature[END_REF]. Only patients carrying the p.Pro55Leu mutation, in the heterozygous [START_REF] Martins | Atypical juvenile presentation of GM2 gangliosidosis AB in a patient compoundheterozygote for c.259G > T and c.164C > T mutations in the GM2A gene[END_REF] or homozygous state, had brain atrophy without signal alteration [START_REF] Salih | Mutation in GM2A leads to a progressive chorea-dementia syndrome. Tremor and Other Hyperkinetic Movements[END_REF]. Our patient's MRI showed no brain atrophy, but presented a localized atrophy of the cerebellar vermis comparable to that seen in adult forms of Tay-Sachs disease [START_REF] Masingue | Natural history of adult patients with GM2 gangliosidosis[END_REF][START_REF] Neudorfer | Lateonset Tay-Sachs disease: phenotypic characterization and genotypic correlations in 21 affected patients[END_REF], and a slight aspecific hyperintensity of posterior horn of the lateral ventricles.

A review of the literature on late-onset forms of GM2 gangliosidosis had clarified their clinical profile and evolution [START_REF] Masingue | Natural history of adult patients with GM2 gangliosidosis[END_REF]. Almost half of the patients had small signs during childhood, such as clumsiness or poor motor skills, which remained stable over time. The most frequent and earliest neurological manifestation was lower limb weakness in 81% of patients, with preferential of the psoas and quadriceps due to lower motor neuronopathy.

Half of the patients had to use a wheelchair after 20 years of disease progression, with a very heterogeneous pattern of gait degradation. Upper limb involvement has never been reported as the first sign of the disease and affects 53% of the patients, especially the triceps and interosseous muscles.

More than half of the patients had a cerebellar syndrome, which was not proportional to the cerebellar atrophy, present in 73% of the patients, and only 15% had cerebral atrophy.

Psychiatric disorders (depression, maniac disorder, psychosis) affected about 30% of patients in Masingue's study, 94% of whom had Tay-Sachs disease, and could either correspond to the initial disorder of the disease or be isolated.

Neudorfer's study found the same proportion of clinical manifestations in their population of adult Tay-Sachs patients, except for psychiatric disorders, which were much more frequent (85.7% versus 30%) [START_REF] Neudorfer | Lateonset Tay-Sachs disease: phenotypic characterization and genotypic correlations in 21 affected patients[END_REF]. None of these studies reported ophtalmological disorders in late onset forms.

Masingue's study identified four different initial presentations of GM2 gangliosidosis in adults [START_REF] Masingue | Natural history of adult patients with GM2 gangliosidosis[END_REF]:

-Lower motoneuron disorder responsible for proximal lower limb weakness, with progression to the upper limbs; -Cerebellar ataxia (more frequently isolated in Sandhoff patients); -Psychiatric disorder (Tay-Sachs disease only); -Complex clinical presentation mixing the three above phenotypes.

The clinical course of our patient resembles that of an adult form of Tay-Sachs: initial involvement of the lower limbs with discrete signs in childhood of poor motor skills, progression to the upper limbs with involvement of the ulnar nerve and interosseous amyotrophy, cerebellar atrophy and slight cognitive impairment. The long-term evolution of the disease is mainly marked by the need to use a wheelchair to move around, the risk of developing psychiatric symptoms, and the occurrence of progressive cognitive decline.

Although all three clinical forms (infantile, juvenile, adult) have been described for Tay-Sachs and Sandhoff diseases, this was not the case for the AB variant, of which we report here the first late-onset case.

Neither EM nor IF allowed us to demonstrate a lysosomal accumulation of GM2 by functional deficiency of GM2-AP. RT-qPCR did not show a reduction in gene expression compared to controls, and cDNA Sanger sequencing revealed the presence of both alleles, with the p.Lys27* mutation potentially being slightly less expressed on chromatogram. On the other hand, the newly developed LC-MS/MS technique allowed us to show a slight increase of GM2 in our patient's plasma, with a concentration close to adult forms of Tay-Sachs and Sandhoff disease, allowing us to classify our GM2A:c.79A > T:p.Lys27* variant, not previously reported in healthy individual databases, as pathogenic. On the other hand, the GM2A:c.415C > T:p.Pro139Ser variant was only reported once in the Latin American population, with no known homozygous subject in the gnomAD V2.1.1, ExAC, or TOPMED databases. Alignment of GM2A gene across different species confirmed the conservation of Proline 139 through Metazoa, and interestingly, most of previously reported pathogenic missenses occurred on a well-conserved amino acid (Fig. 4). This study allows us to validate the PS3 ACMG criteria for this variant too and reclassify it from VOUS to be likely pathogenic (PM1/PM2/PP3/PS3). This situation highlights the necessity of functionnal analysis in unusual clinical presentations, allowing for confirmation of both mutations pathogenicity and diagnosis. Some pathogenic missenses appear to be hypomorphic and are much more represented in adult forms of Tay-Sachs disease, as HEXA:c.805G > A:p.Gly269Ser (rs121907954) [31,[START_REF] Paw | Molecular basis of adult-onset and chronic GM2 gangliosidoses in patients of Ashkenazi Jewish origin: substitution of serine for glycine at position 269 of the alpha-subunit of beta-hexosaminidase[END_REF]. Moreover, it has been described that the phenotype is often determined by the second allele in GM2 gangliosidosis, when a premature stop codon is present [START_REF] Mahuran | Biochemical consequences of mutations causing the GM2 gangliosidoses[END_REF].

Similarly, the GM2A:c.415C > T:p.Pro139Ser variant seems to be hypomorphic and may result in an adult or delayed form of AB variant when coupled to other nonsense mutation, or if present in the homozygous state. However, further mildly affected patients are required to confirm this hypothesis.

Conclusion

Although late-onset AB variant was not described to date in the literature, our case demonstrates that this pathology indeed follows the same rules as other GM2 gangliosidosis, with the existence of a clinical adult form, more likely to be linked to our GM2A:c.415C > T:p.Pro139Ser variant, which we were able to reclassify as likely pathogenic. Finally, the clinical course of our patient would be similar to that of the adult forms of Tay-Sachs, with foremost a progression of the lower limb deficiency and the necessity to use a wheelchair to move around.

Declarations

Conflict of interest The authors declare no competing interests.