Sphingosine-1-Phosphate Lyase Inhibition Alters the S1P Gradient and Ameliorates Crohn’s-Like Ileitis by Suppressing Thymocyte Maturation

Sphingosine-1-phosphate (S1P) receptor agonists are a promising therapeutic alternative to anti-integrin antibodies in inflammatory bowel disease. Here, we report that modulation of tissue S1P levels via inhibition of the S1P lyase might be an effective alternative, acting by interference with thymocyte maturation. Abstract Background Lymphocytes recirculate from tissues to blood following the sphingosine-1-phosphate (S1P) gradient (low in tissues, high in blood), maintained by synthetic and degradative enzymes, among which the S1P lyase (SPL) irreversibly degrades S1P. The role of SPL in the intestine, both during homeostasis and IBD, is poorly understood. We hypothesized that modulation of tissue S1P levels might be advantageous over S1P receptor (S1PR) agonists (eg, fingolimod, ozanimod, etrasimod), as without S1PR engagement there might be less likelihood of potential off-target effects. Methods First we examined SPL mRNA transcripts and SPL localization in tissues by quantitative reverse transcription polymerase chain reaction and immunohistochemistry. The in vivo effects of the SPL inhibitors 4-deoxypyridoxine hydrochloride (30 mg/L) and 2-acetyl-4 (tetrahydroxybutyl)imidazole (50 mg/L) were assessed through their oral administration to adult TNF∆ARE mice, which spontaneously develop Crohn’s-like chronic ileitis. The effect of SPL inhibition on circulating and tissue lymphocytes, transcriptional regulation of proinflammatory cytokines, and on the histological severity of ileitis was additionally examined. Tissue S1P levels were determined by liquid chromatography–mass spectrometry. Mechanistically, the potential effects of high S1P tissue levels on intestinal leukocyte apoptosis were assessed via terminal deoxynucleotidyl transferase dUTP nick end-labeling assay and annexin 5 staining. Finally, we examined the ability of T cells to home to the intestine, along with the effects of SPL inhibition on cellular subsets within immune compartments via flow and mass cytometry. Results S1P lyase was ubiquitously expressed. In the gut, immunohistochemistry predominantly localized it to small intestinal epithelia, although the lamina propria leukocyte fraction had higher mRNA transcripts. Inhibition of SPL markedly increased local intestinal S1P levels, induced peripheral lymphopenia, downregulated proinflammatory cytokines, and attenuated chronic ileitis in mice. SPL inhibition reduced T and myeloid cells in secondary lymphoid tissues and the intestine and dec