Ruxolitinib Impairs Chemokine-Directed Migration and Delays Activation of Human Cytotoxic T Lymphocytes by Modulating P-Glycoprotein Function

Cytotoxic T lymphocytes (CTLs) rely on tightly regulated activation, efflux capacity, and migration to maintain immune surveillance and establish memory pools. Modulation of these pathways can profoundly affect T cell differentiation and function. We investigated how ruxolitinib, a clinically approved JAK1/2 inhibitor, impacts P-glycoprotein (Pgp, MDR1, ABCB1) expression and function, T cell activation markers, and chemokine-driven transmigration in human CD8⁺ T cells. Ruxolitinib modulated Pgp activity at multiple levels. It induced and stabilized ABCB1 mRNA during acute TCR activation, directly inhibited Pgp efflux function, and stimulated basal ATPase activity in a manner similar to the known Pgp substrate verapamil. Functional assays classified ruxolitinib as a Pgp modulator and inducer. Additionally, ruxolitinib suppressed TCR-induced upregulation of PD-1 and CD8 surface markers, suggesting interference with activation-associated differentiation pathways. Finally, ruxolitinib significantly impaired CCL19-driven transmigration of CTLs across endothelial monolayers at clinically relevant concentrations, highlighting its effect on lymphoid homing mechanisms. Our findings reveal that ruxolitinib profoundly reshapes CTL function by altering efflux regulation, activation dynamics, and migratory behavior. These insights have implications for optimizing JAK-targeted therapies in immune modulation, vaccination, transplantation, and memory T cell-based immunotherapies.