Fucosidosis—Clinical Manifestation, Long-Term Outcomes, and Genetic Profile—Review and Case Series

Fucosidosis is a neurodegenerative disorder which progresses inexorably. Clinical features include coarse facial features, growth retardation, recurrent upper respiratory infections, dysostosis multiplex, and angiokeratoma corporis diffusum. Fucosidosis is caused by mutations in the FUCA1 gene resulting in α-L-fucosidase deficiency. Only 36 pathogenic variants in the FUCA1 gene are related to fucosidosis. Most of them are missense/nonsense substitutions; six missense and 11 nonsense mutations. Among deletions there were eight small and five gross changes. So far, only three splice site variants have been described—one small deletion, one complete deletion and one stop-loss mutation. The disease has a significant clinical variability, the cause of which is not well understood. The genotype–phenotype correlation has not been well defined. This review describes the genetic profile and clinical manifestations of fucosidosis in pediatric and adult cases.


Introduction
Fucosidosis (OMIM# 230000) is a lysosomal storage disease (LSD) caused by biallelic pathogenic variants in FUCA1 gene localized on chromosome 1p36.11 [1][2][3]. Mutations result in α-L-fucosidase (EC 3.2.1.51) deficiency [4] in almost all patients [5]. The enzyme is a homotetramer composed of subunits of different masses (50 to 60 kDa), which result from variations in N-glycosylation and proteolytic processing [6,7]. As a result of the hydrolytic enzyme deficiency, incomplete catabolism of N-and O-glycosylproteins results in the accumulation of fucose-containing glycolipids and glycoproteins in various tissues and urine [7]. In particular, glycolipids expressing blood group antigens (H, X) accumulate in the liver.
Fucose (C 6 H 12 O 6 ), a deoxy sugar isolated both as free sugar and as a component of oligosaccharides in human and sheep milk, is found in most of the plasma glycoproteins in the mucopolysaccharides and mucolipids of various human and animal tissues, where it is present mainly as a terminal sugar [8]. The impaired hydrolysis of terminal α-L-fucoside linkages in glycosphingolipids and glycoproteins leads to accretion of fucose-containing oligosaccharides, glycoproteins, and glycolipids in the brain, liver and skin [1,2]. Oligosaccharides and glycoasparagines are deposited in tissues and excessively excreted in urine. This variant alters the acceptor splice site of exon 3 and the consequence of this change is not predictable, but a skip of exon 3 is very likely. [35]   Most of them are missense/nonsense substitutions, in which there are six missense and 11 nonsense mutations. Deletions occur frequently, including eight small and five gross changes. So far, only three splice site variants have been described-one small deletion, one complete deletion, and one stop-loss mutation [25].
All pathogenic variants result in barely detectable α-L-fucosidase activity and significantly deficient cross-reacting immune material (CRIM; below 6% of normal mean), implying that the mutant fucosidase proteins are unstable and degrade at a great rate [1,5,30].
A pseudogene FUCA1P was found on chromosome 2q31-q32. It shares 80% identity with α-L-fucosidase gene and contains no intron as well as open reading frame [22]. A locus on chromosome 6q24.2 (FUCA2) has been shown to influence plasma fucosidase activity (representing 10-20% of the total cellular fucosidase activity), but not its activity in leukocytes. This gene is not involved in fucosidosis [49].

Molecular Testing
Molecular genetic diagnostics include a mix of FUCA1-gene-targeted testing and chromosomal microarray analysis depending on the molecular background of the disease. FUCA1 sequence analysis detects most of the known pathogenic changes-intragenic small deletions/insertions (INDELs) and missense, nonsense, and splice sites variants. Gross deletions/duplications (exons or the whole gene) are detected by chromosomal microarray analysis (CMA). The sequence analysis should be performed first. If only one or no pathogenic variant is found, array comparative genomic hybridization (aCGH) analysis is recommended.
FUCA1-gene-targeted analysis requires that the clinician determine that fucosidosis is the most probable disease in patient, whereas multigene panel testing proves to be valuable in making differential diagnoses. Because the clinical manifestation of fucosidosis is broad ranging, individuals with the distinctive phenotype may successfully be diagnosed using single-gene testing, while those in whom the clinical diagnosis is unclear need verification using more comprehensive testing, including whole-exome sequencing.

Genotype-Phenotype Correlation
Since most affected individuals are homozygotes for different mutations, strict genotypephenotype correlation is not well defined. Only few of them are repeated in unrelated patients. As a rule, patients are homozygous for two null alleles, such as the gross deletions, including G33 or G34 genotype (Tables 1 and 2), have a severe phenotype [1,16].      The clinical heterogeneity is less commonly associated with different non-allelic mutations, as complementation studies between type I and type II fucosidosis (see below) did not restore α-L-fucosidase activity [51]. The possible existence of different co-allelic mutations in the fucosidase gene, responsible for different clinical phenotypes, has not yet been investigated [5]. This hypothesis however disagrees with the existence of different clinical phenotypes within the same family [52]. The clinical variability of fucosidosis is broad and it was proposed that other genetic or non-genetic factors may take part in it [7].

Prenatal Diagnosis
Fucosidosis is inherited in an autosomal recessive manner, which means that each sibling of the patient from the same biological parents has a 25% risk of being affected, a 50% chance of being an heterozygous healthy carrier, and a 25% chance of being healthy and not a carrier [1].
Once the FUCA1 pathogenic variants have been identified in an affected family member, parents may decide on prenatal testing in the subsequent pregnancy or preimplantation genetic testing (PGT-M).
Only molecular genetic testing can be used. Biochemical testing is not useful because the expression level is nearly 50% lower in mutation carriers than in the normal control [36]. The Western blot analysis showed that mutation caused the decrease of protein FUCA1 expression level and mutation carriers still had partial protein expression, but lower than normal controls [36].

Clinical Manifestation
Fucosidosis is a neurodegenerative disorder which progresses inexorably. Its features, such as coarse facial features, growth retardation, recurrent upper respiratory infections, dysostosis multiplex, and angiokeratoma corporis diffusum are common in other LSDs. The mean age at presentation of the first symptom in 60 patients was 1.2 +/− 0.8 years (range: 0.0-5.5 years) [1].

Type I and II
Although the first case of fucosidosis was described in 1966 [12], the enzyme deficiency was only characterized in 1968 [53].
In view of the significant variability in clinical manifestations in fucosidosis, two main types have been defined [54,55].

•
Type I: a rapidly-progressing neurodegenerative course, leading to decerebration and death typically before the age of 10 years [13]; • Type II: a milder course, with slower neurological symptoms progression, possible survival into adulthood, and most patients develop angiokeratoma corporis diffusum [14].

Skin
Dermatological abnormalities were documented in 60% of patient and included telangiectasiae on the skin or conjunctivae and angiokeratoma [1]. Angiokeratoma corporis diffusum was present in most (51%) patients [1], particularly in those living ten years and longer, although it is not pathognomonic for fucosidosis, it raises a suspicion of the diagnosis [56]. The older the patient is when examined, the more likely that angiokeratomas are found [57].
Red, purple pinhead-sized raised skin lesions usually develop on the lower abdomen and genitalia, while tiny scattered angiomas cover most of the body [1]. Angiokeratomas, within the papillary dermis, are proliferative ectatic blood vessels limited by a flattened endothelium containing erythrocytes [58]. Some patients have only telangiectasia without angiokeratoma [18].
Valero-Rubio et al. [60] showed that FUCA1 expression disturbances, resulting from its down-regulation, affect mainly genes related to keratinocyte differentiation/epidermal development and immune responses. This might have been instigated by abnormal transcription factor expression (FOXN1). The authors thus postulated that skin lesions in fucosidosis might be caused by dysfunctions in common aetiological overlapping molecular cascades [60].

Eye
Dilated and tortuous retinal veins were observed in 54%, dilated and tortuous conjunctival vessels in 53%, microaneurysms of conjunctival vessels in 41%, corneal opacities in 11%, and pigmentary retinopathy in 7% of patients and vision loss in 6% of patients [1]. Severe visual impairment is uncommon.
It was shown that storage material accumulates in conjunctival, retinal, and skin vessels. Histological assessment of the conjunctiva's endothelial cells has shown two distinct types of vacuoles-clear ones with a reticular structure similar to those seen in the mucopolysaccharidoses and dark inclusions with a dense granular material [61,62].
The anterior chambers are of normal depth with clear cornea and lenses. The fundi show macular changes similar to bull's eye retinopathy; the foveal area has fine brown pigmentation ringed by a zone of hypopigmentation. The rest of the macula has fine brownish and/or, slatey-blue pigmentary lesion with occasional clumps of dark brown discoloration. No undue tortuosity of the vessels are present, the discs and vessels are normal.
Snodgrass et al. [63] and Gatti, et al. [54] reported 'slight cloudiness of corneae' in some cases [64]. Borrone et al. [64] reported two affected siblings of whom one presented with corneal clouding, strabismus, papilledema with congestion, and tortuosity of retinal vessels, and the other with thin and tortuous vessels. Another 14-year-old female patient presented with a slight limitation in right abduction, an 18-prism diopter exophoria in primary gaze at distance with a latent component fine jerky vertical nystagmus. She had 100 s of arc stereopsis. On slit lamp examination telangiectatic vessels in the inferior conjunctiva and mild corneal stromal haze bilaterally proportional to her slightly decreased best corrected visual acuity were noted [65].
The height of 91% of patients was reported to be below the 5th centile [1], while the weight of 77% was below the fifth centile.

Cardiac Manifestations
Cardiomyopathy is not a common feature [18] but mild mitral regurgitation was found in 50% of patients investigated [69]. An enlarged heart with left ventricular hypertrophy and dilatation of right cavity can be observed on an X-ray and the electrocardiogram may show incomplete right bundle branch block [8]. Cloudy degeneration of myocardium and stasis in coronary vessels was also previously observed [8].

Respiratory/Recurrent Infections
Up to 78% of patients experience respiratory tract infections [1]. Intermittent otitis media, upper respiratory tract infections, and breathing difficulties during sleep as a result of partial upper airway obstruction were documented [70]. Moreover, a recurrent respiratory infection every 1 or 2 months, despite normal immunological tests [18] and diffuse emphysema with little areas of atelectasis of the lungs was previously observed [8].
Peripheral blood smears confirmed the presence of cytoplasmic vacuoles in many lymphocytes which were only slightly periodic acid-Schiff (PAS) positive [8].
Possible links between fucosidosis and cystic fibrosis were postulated [71]. High sweat electrolytes and recurrent infections of the respiratory tract are features of both disorders. Fucosidosis patients often have recurrent infections confined to areas of mucus-secreting ciliated epithelia but their immune system was shown to be intact and therefore it was suspected that there could be a local defect in mucus clearance. [71].
It was proposed that the terminal sugars fucose and sialic acid play a major role in defining the viscoelasticity of mucus. Thus, alterations in the enzymatic cleavage of these sugars affect mucus cross-linking and its viscoelasticity. Without cross-linking, cilia would flail about ineffectively in watery secretions [71].
Liver biopsy may reveal hepatic cells containing abundant stored ceramide tetra-and pentahexoside, producing a clear, swollen appearance of balloon or pseudogargoyle cells which are only weakly PAS positive [8,53].

Dysostosis Multiplex
Some patients also have non-specific features of dysostosis multiplex [74], which may affect up to 58% of patients [1]. An X-ray of the spine shows small thoracolumbar vertebrae with anterior tonguing, short odontoid pegs, cervical platyspondyly, and broad ribs [75]. Moreover, prognathism, ribs deformities, scoliosis with partial fusion of lower thoracic vertebrae, and gibbous deformity of lumbar vertebrae have been described, as well as absent scaphoid, trapezium, and trapezoid bones in the wrists [63]. Joint contractures have also been a common feature [30]. Willems et al. (1991) documented that up to 95% of patients have progressive mental health deterioration and up to 87% have motor deterioration, while seizures may occur in 38% of patients [1]. Forty-one percent of the patients lost the ability to sit, 38% lost the ability to stand, 50% became unable to walk alone, and 67% lost the ability to speak. Twenty-eight percent of the patients had a rapid neurologic deterioration with complete loss of the ability to walk, stand, sit, or talk before the age 5 years. Slower neurologic deterioration and maintenance of the ability to sit, stand, walk, or talk after the age of 10 was documented in 53% of the patients. In the remaining 19%, the course of neurologic deterioration was intermediate.

Neurology/Neurocognitive Function
Psychomotor decline was noted in 60% of patients at a median age of 18 ± 4.5 months [69]. Progressive neurological degeneration results in flexion contractures of legs and arms [63]. Spastic quadraparesis with increased deep tendon reflexes was previously described [19,68,73] and estimated to affect up to 40% of all fucosidosis cases [47]. Gradually-increasing spasticity with bilaterally-increased deep tendon reflexes and unsustained ankle clonus may result in unsteady gait, associated with excessive femoral anteversion, tibial torsion, mild metatarsus adductus, and intoeing [70].
Seizure types may vary with the secondary generalized epilepsy reported before [70]. Seizure was a feature in several cases [67] and documented in up to 38% of all fucosidosis cases [47].

Neuroimaging
Neuroimaging features are commonly described. There is evidence of hypomyelination, with extensive, confluent, progressive, and symmetric signal anomaly in the periventricular and subcortical white matter [28,66,70,78,79]. Apart from the white matter changes, the grey matter changes were described in fucosidosis and are characterized by signal changes in the globi pallidi [15,28,66,[78][79][80]. This unique-for-fucosidosis feature manifests as marked hypointensity on T2/FLAIR sequences, and hyperintensity on T1 sequences [28,66,76,79]. A hypointense area on T2-weighted imaging in the bilateral globus pallidus may indicate the presence of high levels of iron in echo-gradient MRI [73], and, in combination with curvilinear T2-hyperintense areas within lentiform nuclei, creates a sign sometimes called "eye of the tiger" [18].
Generalized cerebral and cerebellar atrophy was also observed on CT scan with the disease progress [19,28,66,67,78] and was more commonly documented in patients with the type II phenotype [78]. Among patients who survived beyond the age of 30, cranial CT abnormalities included infra-and supratentorial volume loss, in particular in the frontal lobes [81].
Ventricular dilatation and focal areas of hypodensity were also observed [82]. Kau et al. [83] noted an increased cerebellar volume in the early stage of fucosidosis. The prominent white matter abnormalities and low signal of the globus pallidus may help distinguish it from other neurometabolic disorders [39,78].
Another modality, MS spectroscopy, typically shows a decreased N-acetyl aspartate (NAA)/choline ratio and a characteristic abnormal peak at 3.8 ppm [28,66] and a double peak at 1.2 ppm [80]. The level of N-acetylaspartate and creatinine ratio was shown be decreased with an unusual lactic acid peak at 1.33 ppm in MRI spectra in the bilateral basal ganglia and posterior limb of internal capsule [67]. In patients who had undergone hematopoietic stem cell transplantation (HSCT), magnetic resonance spectroscopy (MRS) confirmed the disappearance of an unusual lactic acid peak at 1.33 ppm, and the rise of the of NAA and creatinine ratio to normal [67].

Mortality
Death before age 10 years occurs in less than half patients (43%) and after age 20 similarly it occurs in 41% [1]. Patients with early symptoms tend to show faster neurologic deterioration leading to mortality at young age [1]. It was documented that around 60% of patients die secondary to respiratory infections and neurological deteriorations [69].

Animal Model
A Fuca1-deficient mouse model was generated by gene-targeting techniques [84]. The authors demonstrated behavioral alterations in Fuca1-deficient mice that coincided with early signs of neuropathology (at 3 months of age); subtle abnormalities in sensorimotor and cognitive abilities were identified. The animals displayed lysosomal dysregulation (increased Lamp1 expression) and evidence of neuroinflammation and secondary storage of GM2 ganglioside, as early manifestations of brain pathology [85]. These abnormalities preceded neuron loss but imply an early stage of neuropathology encompassing an extended endosomal-lysosomal network, secondary lipid storage, and emerging microgliosis and astrogliosis [86].

Supportive and MDT
The cornerstone of management is generally supportive with physiotherapy and other allied health input. The multidisciplinary team usually involves pediatric and adult metabolic physicians, metabolic physiotherapist, and ophthalmology, orthopedic, and cardiology specialists. In case of any neurological/neuropsychiatric complications, input from a neurology, neuropsychiatry teams should be sought, with a full neuropsychology assessment.
HSCT was applied in a small number of patients with fucosidosis with symptoms stabilization in some cases. It was first attempted in an asymptomatic 7-month-old child who was diagnosed with fucosidosis after the diagnosis of the symptomatic older brother [88]. After it was reported as an effective treatment, it was performed very uncommonly, and only after careful patient selection [3,89]. Earlier transplant appeared to be more effective in fucosidosis than transplant after symptoms fully manifest. After successful transplantation, a gradual increase in α-L fucosidase levels was observed in plasma, white blood cells, and cerebrospinal fluid, as well as an improvement in myelination on MRI scan for up to 4 years after transplantation [88,89].
In a case described by Milano et al. [89] at four years post-HSCT, there were major improvements in MRI findings, psychomotor development, swallowing, and number of respiratory tract infections. In a case of a symptomatic 3-year-old girl who underwent an umbilical cord blood (UCB) HSCT, a remarkable regression of neurological symptoms after HSCT was detected, cranial MRI results at two years post-HCT improved, a prior lactic acid peak disappeared on MRS, but no impact on dysostosis multiplex was observed [67]. Two out of three other patients showed decelerating of neurologic progress and a dramatic reduction in respiratory infections after HSCT [90,91]. Other HSCT-related trials were completed in 2008 [92].

Enzyme Replacement Therapy
Enzyme replacement therapy (ERT) for fucosidosis is currently being tested in preclinical studies. It is the first glycoprotein disorder to use intracisternal ERT to deliver a recombinant enzyme directly to the central nervous system (CNS) in a canine [93]. It was confirmed that the enzyme was higher near the injection site (39-73% of normal) and lower in deep brain structures (2.6-5.5% of normal), but it reached all areas of the CNS. The direct CNS delivery of the enzyme was shown to be safe and in a follow-up study (in the same experiment) partial improvement in neuropathology following intracisternal ERT has been shown, implying that perhaps a more intensive ERT protocol is required to improve outcomes [91,93].
Gene therapy may be an alternative treatment [94,95] but no clinical trials are available yet.

Discussion
Atypical presentation of fucosidosis may lead to delay in diagnosis, in particular in absence of coarse features or hepatosplenomegaly or joint contractures. As an example, a female patient aged 4 years developed swollen, painful knees, hips, wrists, hands and feet. While the inflammatory response was suggestive of polyartricular juvenile inflammatory rheumatoid arthritis [75], her symptoms were associated with the underlying metabolic disease. Diagnostic delay could be avoided with remembering the wide spectrum of clinical and radiological findings of fucosidosis with atypical presentations of disease.
Telangiectasia or angiokeratoma, despite it being inconspicuous in the early stage of the disease, might be its early indicator. Careful review of the classic symptoms, including visceromegaly, neurological deterioration, dysostosis multiplex, and neuroimaging, is important and allows for accurate diagnosis [18].
It is worth emphasizing that urinary screening of glycosaminoglycans (GAGs) does not detect this rare LSD, which requires specific enzyme analysis of plasma and leucocytes samples, followed by genetic analysis of FUCA1 mutations. In addition, a negative urinary oligosaccharide screen, seen in attenuated phenotype, does not exclude a diagnosis of LSD [75].
Neuroimaging is nearly always abnormal when performed in patients with fucosidosis. In terms of diagnostic investigations, biochemical testing is the first-line test and the preferred method for confirming the diagnosis. However, with easier access to next generation sequencing (NGS), more cases of fucosidosis are expected to be recognized, and-in such circumstances-the traditional pathway of biochemical testing will serve for verification. Given a potential benefit from transplantation in the early stages of disease, this is a potentially-treatable disorder [3,89]. Unfortunately, individuals are often diagnosed with advanced organ involvement and thus considered unsuitable for treatment with transplantation.
Genetic testing may be the only way to diagnose fucosidosis before the full spectrum of clinical manifestations will disclose. The genotype-phenotype correlation, as for other autosomal recessive disorders is, however, unclear and not straightforward. Given the genotypes summarized in Table 1, deletions, causing loss of enzyme activity (G7.2, G33, and G34) and variants leading to protein truncation, usually result in decreasing enzymatic activity (G2-G7.1, G9.1, G10.2, G11-G15, G17-G19, G22, G25-G29.2, G31, and G32.2) are expected to cause more severe fucosidosis manifestation. Unfortunately, the phenotypic descriptions are often lacking (Table 2). For variants altering the acceptor splice sites and available clinical records, we can conclude that G10.1 and G30 may result in later-onset disease and normal development in early childhood, with rapid progression from the age of three and cerebral atrophy in G10.1 [35] and regression from the age of two with very low, residual activity of α-L-fucosidase in G30 [47]. A severe course seems be the characteristic for G16, G20, G21, and G24, with observed psychomotor delayed from infancy in G24 [42], no activity of α-L-fucosidase in the leucocytes in G20 [40], and leukodystrophy described for G16 [37]. The conclusions need verification and further explanations of their pathomechanisms.