DIPG-50. Bioinformatic evaluation of genes involved in sphingomyelin biosynthesis in Diffuse Midline Glioma H3K27 altered/DIPG: dysregulation of sphingosine 1-phosphate (SP1)

Abstract

Sphingosine 1-phosphate (S1P), a bioactive signalling lipid, interacts with a network of metabolic enzymes, receptors, transporters, and epigenetic partners. This network is well described in many cancers; however, little is known about its potential impact in DIPG. Expression of HDAC1 (binding target of S1P) and genes associated with the sphingomyelin (SM) pathway were examined in datasets identified in the National Centre for Biotechnology Information, Gene Expression Omnibus, and analysed using the R2: Genomics Analysis and Visualization Platform (http://r2.amc.nl). The Paugh-DIPG dataset (27 DIPG samples) and normal samples (20 years and younger - Berchtold dataset) were compared. To avoid issues related to batch effects, expression values for each gene of interest and controls were exported into separate files to determine differentially expressed genes. Internal genes include housekeeping; ACTB, GAPDH, B2M, TBP; downregulated in DIPG; GPR6, NGB, and upregulated in DIPG; MMP16, PDGFRA, TP53, CSPG4. Genes of interest; SPHK1, SPHK2, SGPL1, ACER1, ACER3, KDSR, SMPD1-4, CPTP, GLTP, DEGS1, CERK, CERS1-6, ASAH1, SGPP1, SGPP2 and HDAC1. To test for significance, each dataset was standardised using ACTB housekeeping gene. Values including Log-transformed fold change were analysed using the non-parametric, Mann-Whitney test. 7 of the 16 genes were dysregulated relative to expression in normal brain (p<0.0002). SPHK2 and SMPD3 were downregulated, and HDAC1, SGPL1, DEGS1, CERS4, and ASAH1 were upregulated in DIPG compared to normal. To identify genes more likely associated with DIPG (vs development), we evaluated gene expression in Brainspan dataset (brspv10rs). Validation of SPHK2 and SGPL1 protein expression (responsible for the synthesis and cleavage of SP1) is underway. Current work is focused on the intracellular processing and function (isoform specific inhibitors) of S1P in DIPG cells. Given its reported role in several cancer hallmarks, a better understanding of the sphingomyelin biosynthesis pathway in DMG/DIPG is merited and may lead to novel therapeutic targets.