Essential role for Stat5a/b in myeloproliferative neoplasms induced by BCR-ABL1 and JAK2(V617F) in mice.

STAT5 proteins are constitutively activated in malignant cells from many patients with leukemia, including the myeloproliferative neoplasms (MPNs) chronic myeloid leukemia (CML) and polycythemia vera (PV), but whether STAT5 is essential for the pathogenesis of these diseases is not known. In the present study, we used mice with a conditional null mutation in the Stat5a/b gene locus to determine the requirement for STAT5 in MPNs induced by BCR-ABL1 and JAK2(V617F) in retroviral transplantation models of CML and PV. Loss of one Stat5a/b allele resulted in a decrease in BCR-ABL1-induced CML-like MPN and the appearance of B-cell acute lymphoblastic leukemia, whereas complete deletion of Stat5a/b prevented the development of leukemia in primary recipients. However, BCR-ABL1 was expressed and active in Stat5-null leukemic stem cells, and Stat5 deletion did not prevent progression to lymphoid blast crisis or abolish established B-cell acute lymphoblastic leukemia. JAK2(V617F) failed to induce polycythemia in recipients after deletion of Stat5a/b, although the loss of STAT5 did not prevent the development of myelofibrosis. These results demonstrate that STAT5a/b is essential for the induction of CML-like leukemia by BCR-ABL1 and of polycythemia by JAK2(V617F), and validate STAT5a/b and the genes they regulate as targets for therapy in these MPNs.

Conformational control inhibition of the BCR-ABL1 tyrosine kinase, including the gatekeeper T315I mutant, by the switch-control inhibitor DCC-2036.

Acquired resistance to ABL1 tyrosine kinase inhibitors (TKIs) through ABL1 kinase domain mutations, particularly the gatekeeper mutant T315I, is a significant problem for patients with chronic myeloid leukemia (CML). Using structure-based drug design, we developed compounds that bind to residues (Arg386/Glu282) ABL1 uses to switch between inactive and active conformations. The lead "switch-control" inhibitor, DCC-2036, potently inhibits both unphosphorylated and phosphorylated ABL1 by inducing a type II inactive conformation, and retains efficacy against the majority of clinically relevant CML-resistance mutants, including T315I. DCC-2036 inhibits BCR-ABL1(T315I)-expressing cell lines, prolongs survival in mouse models of T315I mutant CML and B-lymphoblastic leukemia, and inhibits primary patient leukemia cells expressing T315I in vitro and in vivo, supporting its clinical development in TKI-resistant Ph(+) leukemia.

Molecular pathogenesis and therapy of polycythemia induced in mice by JAK2 V617F.

BACKGROUND: A somatic activating mutation (V617F) in the JAK2 tyrosine kinase was recently discovered in the majority of patients with polycythemia vera (PV), and some with essential thrombocythemia (ET) and chronic idiopathic myelofibrosis. However, the role of mutant JAK2 in disease pathogenesis is unclear. METHODS AND FINDINGS: We expressed murine JAK2 WT or V617F via retroviral bone marrow transduction/transplantation in the hematopoietic system of two different inbred mouse strains, Balb/c and C57Bl/6 (B6). In both strains, JAK2 V617F, but not JAK2 WT, induced non-fatal polycythemia characterized by increased hematocrit and hemoglobin, reticulocytosis, splenomegaly, low plasma erythropoietin (Epo), and Epo-independent erythroid colonies. JAK2 V617F also induced leukocytosis and neutrophilia that was much more prominent in Balb/c mice than in B6. Platelet counts were not affected in either strain despite expression of JAK2 V617F in megakaryocytes and markedly prolonged tail bleeding times. The polycythemia tended to resolve after several months, coincident with increased spleen and marrow fibrosis, but was resurrected by transplantation to secondary recipients. Using donor mice with mutations in Lyn, Hck, and Fgr, we demonstrated that the polycythemia was independent of Src kinases. Polycythemia and reticulocytosis responded to treatment with imatinib or a JAK2 inhibitor, but were unresponsive to the Src inhibitor dasatinib. CONCLUSIONS: These findings demonstrate that JAK2 V617F induces Epo-independent expansion of the erythroid lineage in vivo. The fact that the central erythroid features of PV are recapitulated by expression of JAK2 V617F argues that it is the primary and direct cause of human PV. The lack of thrombocytosis suggests that additional events may be required for JAK2 V617F to cause ET, but qualitative platelet abnormalities induced by JAK2 V617F may contribute to the hemostatic complications of PV. Despite the role of Src kinases in Epo signaling, our studies predict that Src inhibitors will be ineffective for therapy of PV. However, we provide proof-of-principle that a JAK2 inhibitor should have therapeutic effects on the polycythemia, and perhaps myelofibrosis and hemostatic abnormalities, suffered by MPD patients carrying the JAK2 V617F mutation.