dihydrosphingosine-1-phosphate and dihydroceramide

Sphingolipidoses are monogenic lipid storage diseases caused by variants in enzymes of lipid synthesis and metabolism. We describe an autosomal recessive complex neurological disorder affecting consanguineous kindred. All four affected individuals, born at term following normal pregnancies, had mild to severe intellectual disability, spastic quadriplegia, scoliosis and epilepsy in most, with no dysmorphic features. Brain MRI findings were suggestive of leukodystrophy, with abnormal hyperintense signal in the periventricular perioccipital region and thinning of the body of corpus callosum. Notably, all affected individuals were asymptomatic at early infancy and developed normally until the age of 8-18 months, when deterioration ensued. Homozygosity mapping identified a single 8.7 Mb disease-associated locus on chromosome 1q41-1q42.13 between rs1511695 and rs537250 (two-point LOD score 2.1). Whole exome sequencing, validated through Sanger sequencing, identified within this locus a single disease-associated homozygous variant in DEGS1, encoding C4-dihydroceramide desaturase, an enzyme of the ceramide synthesis pathway. The missense variant, segregating within the family as expected for recessive heredity, affects an evolutionary-conserved amino acid of all isoforms of DEGS1 (c.656A>G, c.764A>G; p.(N219S), p.(N255S)) and was not found in a homozygous state in ExAC and gnomAD databases or in 300 ethnically matched individuals. Lipidomcs analysis of whole blood of affected individuals demonstrated augmented levels of dihydroceramides, dihydrosphingosine, dihydrosphingosine-1-phosphate and dihydrosphingomyelins with reduced levels of ceramide, sphingosine, sphingosine-1-phosphate and monohexosylceramides, as expected in malfunction of C4-dihydroceramide desaturase. Thus, we describe a sphingolipidosis causing a severe regressive neurological disease.

Topics: Adolescent; Adult; Brain; Ceramides; Cerebrosides; Child; Child, Preschool; Exome Sequencing; Fatty Acid Desaturases; Female; Genetic Predisposition to Disease; Genetic Variation; Homozygote; Humans; Infant; Intellectual Disability; Lysophospholipids; Male; Mutation, Missense; Nervous System Diseases; Pedigree; Phenotype; Sequence Analysis, DNA; Sphingosine; Young Adult

Novel immunomodulatory molecule FTY720 is a synthetic analog of myriocin, but unlike myriocin FTY720 does not inhibit serine palmitoyltransferase. Although many of the effects of FTY720 are ascribed to its phosphorylation and subsequent sphingosine 1-phosphate (S1P)-like action through S1P(1,3-5) receptors, studies on modulation of intracellular balance of signaling sphingolipids by FTY720 are limited. In this study, we used stable isotope pulse labeling of human pulmonary artery endothelial cells with l-[U-(13)C, (15)N]serine as well as in vitro enzymatic assays and liquid chromatography-tandem mass spectrometry methodology to characterize FTY720 interference with sphingolipid de novo biosynthesis. In human pulmonary artery endothelial cells, FTY720 inhibited ceramide synthases, resulting in decreased cellular levels of dihydroceramides, ceramides, sphingosine, and S1P but increased levels of dihydrosphingosine and dihydrosphingosine 1-phosphate (DHS1P). The FTY720-induced modulation of sphingolipid de novo biosynthesis was similar to that of fumonisin B1, a classical inhibitor of ceramide synthases, but differed in the efficiency to inhibit biosynthesis of short-chain versus long-chain ceramides. In vitro kinetic studies revealed that FTY720 is a competitive inhibitor of ceramide synthase 2 toward dihydrosphingosine with an apparent K(i) of 2.15 microm. FTY720-induced up-regulation of DHS1P level was mediated by sphingosine kinase (SphK) 1, but not SphK2, as confirmed by experiments using SphK1/2 silencing with small interfering RNA. Our data demonstrate for the first time the ability of FTY720 to inhibit ceramide synthases and modulate the intracellular balance of signaling sphingolipids. These findings open a novel direction for therapeutic applications of FTY720 that focuses on inhibition of ceramide biosynthesis, ceramide-dependent signaling, and the up-regulation of DHS1P generation in cells.

Topics: Cells, Cultured; Ceramides; Chromatography, Liquid; Endothelium, Vascular; Fingolimod Hydrochloride; Humans; Immunosuppressive Agents; Lung; Lysophospholipids; Oxidoreductases; Phosphorylation; Phosphotransferases (Alcohol Group Acceptor); Propylene Glycols; Pulmonary Artery; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; RNA, Small Interfering; Serine C-Palmitoyltransferase; Sphingosine; Tandem Mass Spectrometry; Up-Regulation