Competitive Binding of Ozanimod and Other Sphingosine 1-Phosphate Receptor Modulators at Receptor Subtypes 1 and 5

Ozanimod, a sphingosine 1-phosphate (S1P) receptor modulator, binds with high affinity selectively to S1P receptor subtypes 1 (S1P(1)) and 5 (S1P(5)), and is approved in multiple countries for treating adults with relapsing forms of multiple sclerosis (MS) or moderately to severely active ulcerative colitis (UC). Other S1P receptor modulators have been approved for the treatment of MS or are in clinical development for MS or UC, but it is unknown whether these compounds bind competitively with each other to S1P(1) or S1P(5). We developed a competitive radioligand binding assay using tritiated ozanimod and demonstrate full displacement of ozanimod by S1P (endogenous ligand), suggesting that ozanimod binds to the S1P(1) and S1P(5) orthosteric binding sites. S1P receptor modulators FTY720-p, siponimod, etrasimod, ponesimod, KRP-203-p, and amiselimod-p also completely displacing radiolabeled ozanimod; thus, on a macroscopic level, all bind to the same site. Molecular docking studies support these results and predict the binding of each molecule to the orthosteric site of the receptors, creating similar interactions within S1P(1) and S1P(5). The absolute free energy perturbation method further validated key proposed binding modes. Functional potency tightly aligned with binding affinities across S1P(1) and S1P(5) and all compounds elicited S1P(1)-mediated β-arrestin recruitment. Since all the S1P modulators included in this study display similar receptor pharmacology and compete for binding at the same site, they can be considered interchangeable with one another. The choice of any one particular agent should therefore be made on the basis of overall therapeutic profile, and patients can be offered the opportunity to switch S1P medications without the potential concern of additive S1P pharmacology.