FTY720 phosphorylation is dispensable for its antileukemic activity on Jak2^V617F cells. (A) Schematic representation of FTY720 conversion into FTY720-P and FTY720/PP2A interplay; inhibitors are indicated in black. (B) Effect of FTY720 and its non-immunosuppressive derivatives (OSU-2S and FTY720-S-regioisomer) on the clonogenic potential, expressed as percentage of CFCs, of Ba/F3, Ba/F3-Jak2^V617F, and primary CD34⁺ PV cells (left). Jak2 expression in vehicle- and FTY720-treated CD34⁺ progenitors derived from the BM or PB of PV patients (top right). Jak2 expression in Ba/F3-Jak2^V617F cells treated with the PP2A activator FTY720 alone or in combination with okadaic acid used at concentration (0.25 nM) that inhibits PP2A activity only (bottom right). (C) PP2A activity in Ba/F3-Jak2^V617F, HEL, primary CD34⁺ NBM, and PV cells untreated or treated with FTY720 (2.5 µM) alone or in combination with the sphingosine kinase inhibitor DMS (2.5 µM), the immunosuppressive and S1PR1 agonist FTY720-P (2.5 µM), or with the nonimmunosuppressive FTY720 derivatives (FTY720-S-regioisomer and OSU-2S) (left). Jak2 protein levels in HEL cells untreated and treated with FTY720 (2.5 µM) alone or in combination with DMS (2.5 µM) or treated with FTY720-P (2.5 µM) (top right). Proliferation of Ba/F3-Jak2^V617F cells (or Ba/F3-MigR1 used as a control) untreated or treated with FTY720 (500 nM) alone or in combination with DMS (500 nM) (bottom right).