Free sialic acid storage disorder: Progress and promise

Abstract

Lysosomal free sialic acid storage disorder (FSASD) is an extremely rare, autosomal recessive, neurodegenerative, multisystemic disorder caused by defects in the lysosomal sialic acid membrane exporter SLC17A5 (sialin). SLC17A5 defects cause free sialic acid and some other acidic hexoses to accumulate in lysosomes, resulting in enlarged lysosomes in some cell types and 10-100-fold increased urinary excretion of free sialic acid. Clinical features of FSASD include coarse facial features, organomegaly, and progressive neurodegenerative symptoms with cognitive impairment, cerebellar ataxia and muscular hypotonia. Central hypomyelination with cerebellar atrophy and thinning of the corpus callosum are also prominent disease features. Around 200 FSASD cases are reported worldwide, with the clinical spectrum ranging from a severe infantile onset form, often lethal in early childhood, to a mild, less severe form with subjects living into adulthood, also called Salla disease. The pathobiology of FSASD remains poorly understood and FSASD is likely underdiagnosed. Known patients have experienced a diagnostic delay due to the rarity of the disorder, absence of routine urine sialic acid testing, and non-specific clinical symptoms, including developmental delay, ataxia and infantile hypomyelination. There is no approved therapy for FSASD. We initiated a multidisciplinary collaborative effort involving worldwide academic clinical and scientific FSASD experts, the National Institutes of Health (USA), and the FSASD patient advocacy group (Salla Treatment and Research [S.T.A.R.] Foundation) to overcome the scientific, clinical and financial challenges facing the development of new treatments for FSASD. We aim to collect data that incentivize industry to further develop, obtain approval for, and commercialize FSASD treatments. This review summarizes current aspects of FSASD diagnosis, prevalence, etiology, and disease models, as well as challenges on the path to therapeutic approaches for FSASD.

Keywords: Hypomyelination; Infantile sialic acid storage disorder; Lysosomal membrane transporter; N-acetylneuraminic acid; SLC17A5; Salla disease; Sialic acid.

Figure 1:. Compilation of FSASD Features

(A) Electron micrograph of a skin biopsy from an intermediate FSASD subject. Dermis revealing blood vessels with endothelial cells (E) and pericytes, a nerve (N) bundle with Schwann cells (SC), and fibroblasts (F). The endothelial cells, fibroblasts, and Schwann cells have numerous enlarged, vacuolar shaped, lysosomes (3860×). Inset: Schwann cell containing enlarged lysosomes, most of which are electron lucent; some contain fine fibrillar material (17,550×). Image derived from [17], with permission from Elsevier Inc. (B) Brain MRI of the same intermediate FSASD subject as in (A) at 10 months of age (right images) compared to age-matched control images (left). Top: Axial T1-weighted, Bottom: Sagittal midline T1-weighted. Note widespread and profound hypomyelination throughout the cerebral and cerebellar hemispheres and small corpus callosum (red arrows). FSASD images derived from [17], with permission from Elsevier Inc. (C) Coarse facial features of FSASD include hypertelorism, flat-bridged nose, depressed nasal bridge, broad nasal tip, long philtrum, broad forehead/brachycephaly, depicted in a 4.5 year old girl [10] and a 30-month old girl [17], both presenting with intermediate FSASD. Images with permission from Elsevier Inc. (D) Ultrastructural images of control and Slc17A5^−/− (knock-out, FSASD) mice cervical spinal cord (top) and optic nerve (bottom) cut in cross section demonstrate a decrease in the number of myelinated axons in these tissues in FSASD mice. Scale bars, 2 μm. Image derived from [87] (Copyright 2009 Society for Neuroscience). (E) Fibroblasts from healthy individuals (Control) and an FSASD patient (FSASD) were metabolically labelled with either ManNAl or Neu5NAl for 8 hours and labeled with AzidoFluor 545 fluorescent probe (red) and the nuclear dye DAPI (blue). Cells were then examined using confocal microscopy (Scale bars: 50 μm). Top images: After incorporation of ManNAl, labeled sialylated glycoconjugates were mainly observed in the perinuclear Golgi-like region of both control and FSASD cells, indicating that FSASD cells have the capacity to transform ManNAl into CMP-Neu5NAl, which was then incorporated into the newly synthesized glycoconjugates. Bottom Images: The FSASD cells labeled with Neu5NAl displayed no staining. These results show the inability of Neu5NAl to reach the cytosol and be converted to CMP-Neu5NAl in FSASD cells, consistent with cellular Neu5Al import through the endocytic pathway [128], thus circumventing the absence of a plasma membrane sialic acid transporter. These results confirm not only the crucial role of SLC17A5 in Neu5NAl metabolism, but also the potential of this metabolic labeling methodology to decipher deficiencies in sialic acid pathways. Images derived from [98], with permission from The Royal Society of Chemistry.

Figure 2:. Intracellular Free Neu5Ac Metabolism and Associated Genetic Disorders

Intracellular free Neu5Ac metabolism comprises three processes: (A) Cytoplasmic free Neu5Ac biosynthesis is initiated with the conversion of UDP-N-acetyl glucosamine (UDP-GlcNAc) in a few enzymatic steps to Neu5Ac, which is activated in the nucleus to CMP-Neu5Ac and then transported back to the cytosol [31, 32, 40]. Cytosolic CMP-Neu5Ac is transported into the Golgi by SLC35A1 [129] where it serves as a substrate for sialyltransferases that sialylate nascent glycans [130]. Cytosolic CMP-Neu5Ac also strongly feedback-inhibits the first committed enzyme of sialic acid biosynthesis, UDP-GlcNAc 2 epimerase, providing negative feedback regulation of de novo cytoplasmic Neu5Ac synthesis [33, 34]. (B) Intralysosomal free Neu5Ac salvage occurs through recycling of glycans (glycoproteins, gangliosides) through endocytosis by the endo-lysosomal system, where lysosomal enzymes degrade the glycans into their individual building block molecules, including individual monosaccharides. Free Neu5Ac is released from glycans by neuraminidase enzymes [84, 86]. Neu5Ac is then transported from the lysosomal lumen into the cytosol by SLC17A5 [1]. (C) The fate of salvaged free Neu5Ac in the cytoplasm is unclear. A portion may be excreted from the cell, recycled in the Neu5Ac biosynthesis pathway for direct synthesis of CMP-Neu5Ac, or degraded/catabolized by N-acetylneuraminate pyruvate lyase (NPL) [38] into ManNAc and pyruvate. The ManNAc generated in the cytoplasm can either directly re-enter the Neu5Ac biosynthesis pathway or can be converted to N-acetylglycosamine (GlcNAc) for entry in the hexosamine pathway [38]. Several rare genetic disorders are associated with these pathways: (1) GNE myopathy (MIM#605820; ~950 reported cases [131]); (2) N-acetylneuraminic acid phosphate synthase (NANS) deficiency (MIM#605202; ~9 cases [40]); and (3) deficiency of SLC35A1, CDGIIf (MIM#603585; ~3 cases [129]) are characterized by decreased sialylation of glycans; (4) Sialidosis (MIM#256550; >100 cases [132]) is characterized by lysosomal accumulation of sialylated glycans. Three disorders are associated with increased urinary excretion of free Neu5Ac: (5) Sialuria (MIM#269921; ~ 11 cases [33]); (6) FSASD (MIM#269920, #604369; ~200 cases [1]); and (7) NPL deficiency (2 cases [38]).

Figure 3:. Topology model of SLC17A5

Simplified model of SLC17A5 (not to scale). SLC17A5 consists of 495 amino acids, 12 transmembrane domains and a N-terminal dileucine sorting motif (DRTPLL). Three frequent FSASD mutations are indicated (*). Transmembrane domain 4 (striped) lines a large aqueous cavity that is part of the substrate permeation pathway [4, 5].