The somatic activating janus kinase 2 mutation (JAK2)V617F is detectable in most patients with polycythemia vera (PV). Here we report that CP‐690,550 exerts greater antiproliferative and pro‐apoptotic activity against cells harboring JAK2V617F compared with JAK2WT. CP‐690,550 treatment of murine factor‐dependent cell Patersen–erythropoietin receptor (FDCP‐EpoR) cells harboring human wild‐type or V617F JAK2 resulted in inhibition of cell proliferation with a 50% inhibitory concentration (IC50) of 2.1 µM and 0.25 µM, respectively. Moreover, CP‐690,550 induced a significant pro‐apoptotic effect on murine FDCP‐EpoR cells carrying JAK2V617F, whereas a lesser effect was observed for cells carrying wild‐type JAK2. This activity was coupled with inhibition of phosphorylation of the key JAK2V617F‐dependent downstream signaling effectors signal transducer and activator of transcription (STAT)3, STAT5, and v‐akt murine thymoma viral oncogene homolog (AKT). Furthermore, CP‐690,550 treatment of ex‐vivo‐expanded erythroid progenitors from JAK2V617F‐positive PV patients resulted in specific, antiproliferative (IC50 = 0.2 µM) and pro‐apoptotic activity. In contrast, expanded progenitors from healthy controls were less sensitive to CP‐690,550 in proliferation (IC50 > 1.0 µM), and apoptosis assays. The antiproliferative effect on expanded patient progenitors was paralleled by a decrease in JAK2V617F mutant allele frequency, particularly in a patient homozygous for JAK2V617F. Flow cytometric analysis of expanded PV progenitor cells treated with CP‐690,550 suggests a possible transition towards a pattern of erythroid differentiation resembling expanded cells from normal healthy controls. (Cancer Sci 2008; 99: 1265–1273)