Cytokine stimulation induces proliferation and growth of
acute myeloid leukemia (AML) blasts and high levels of
cytokines have been associated with poor prognosis in AML. The Jak-Stat pathway constitutes a major mediator of
cytokine activity. We investigated whether
WP-1034, a novel Jak-Stat inhibitor, is active against AML blasts. OCIM2 and fresh AML cells were incubated with 1 to 6 microM
WP-1034 to determine its effect on proliferation.
WP-1034 effectively inhibited proliferation of OCIM2 cells and fresh AML samples. We then analyzed the expressions of Stat 1, 3, and 5, as well as Phospho-Stat 1, 3, and 5 by Western immunoblotting after incubation of OCIM2 cells without and with 1 to 10 microM
WP-1034 for 2 hours, and at 5 microM from 20 minutes up to 4 hours and found that
WP-1034 blocked Stat 3 and 5 activation. Analysis of cell cycle status by PI staining and flow cytometry showed that
WP-1034 caused cell cycle arrest of OCIM2 cells in sub-Go phase. We then evaluated the induction of apoptosis of OCIM2 cells following incubation with
WP-1034 at 3 to 6 microM by
annexin V-CY5 assay and analyzed
caspase 3 and PARP cleavage using Western immunoblotting. We found that
WP-1034 induced apoptosis of OCIM2 cells and that induction of apoptosis involved cleavage of
caspase 3 and the
DNA repair enzyme poly (
adenosine diphosphate [
ADP]-ribose)
polymerase (PARP). Taken together, our data suggest that
WP-1034 is a potent inhibitor of AML cell proliferation by inhibition of Stat 3 and 5 and induction of caspase-dependent apoptosis.