Sphingosine and Its Analog, the Immunosuppressant 2-Amino-2-(2-[4-octylphenyl]ethyl)-1,3-propanediol, Interact with the CB sub(1) Cannabinoid Receptor

Sphingosine-1-phosphate (S1P) and cannabinoid receptors are G-protein-coupled receptors that mediate the effects of S1P and endocannabinoids, respectively. Cannabinoid receptors also mediate the effects of Delta super(9)-tetrahydrocannabinol, the primary psychoactive ingredient in marijuana, whereas S1P receptors contribute to the immunosuppressant effects of 2-amino-2-(2-[4-octylphenyl]ethyl)-1,3-propanediol (FTY720). FTY720 is a sphingosine analog that can prevent renal graft rejections and suppress a variety of autoimmune disorders in animal models and clinical trials. We now report that both FTY720 and sphingosine interact with CB sub(1) but not CB sub(2) cannabinoid receptors. FTY720 and sphingosine inhibited the binding of the CB sub(1)-selective antagonist [ super(3)H]N-(piperidinyl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl )-4 -methyl-1H-pyrazole-3-carboxamide ([ super(3)H]SR141716A) and the cannabinoid agonist [ super(3)H](-)-cis-3-[2-hydroxy-4-(1,1-dimethylheptyl)phenyl]-trans -4- (3-hydroxypropyl)cyclohexanol ([ super(3)H]CP55,940) in a concentration-dependent manner in both CB sub(1)-expressing cell lines and mouse cerebellum. However, these compounds did not significantly alter [ super(3)H]CP55,940 binding to CB sub(2) receptors. In G-protein activation assays, FTY720 and sphingosine inhibited the maximal stimulation of guanosine 5'-O-(3-[ super(35)S]thio)triphosphate binding by the cannabinoid agonist R-(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]pyrrolo[1,2,3-d e ]-1,4-benzoxazinyl]-(1-naphthalenyl)methanone mesylate (WIN55,212-2) in a concentration-dependent manner, and this antagonist effect was not mimicked by S1P. FTY720 and sphingosine also inhibited activation of extracellular signal-regulated kinases 1 and 2 and Akt by WIN55,212-2 in intact Chinese hamster ovary (CHO) cells expressing CB sub(1) receptors and attenuated WIN55,212-2-stimulated internalization of a fluorescence-tagged CB sub(1) receptor in CHO cells. Moreover, both FTY720 and sphingosine produced rightward shifts in the concentration-effect curves of cannabinoid agonists for G-protein activation, indicating that they act as competitive CB sub(1) antagonists. These results suggest that the CB sub(1) receptor could be a novel target of FTY720 and that sphingosine could be an endogenous CB sub(1) antagonist.