An activating mutation in the Janus kinase 2 gene (JAK2) (G1849T, which produces JAK2 V617F) occurs at a high frequency in Bcr-Abl-negative myeloproliferative neoplasms (MPNs). JAK2 V617F induces cytokine-independent growth in cell lines and, in murine models, recapitulates much of the pathobiology observed in MPN patients, suggesting that small-molecule inhibitors targeting JAK2 may be therapeutically useful. Some orally bioavailable inhibitors of JAK2 are already in clinical trials.
NS-018 is a novel JAK2 inhibitor that inhibits JAK2 enzyme activity with an IC50 value of less than 1 nM. NS-018 shows 30–50-fold selectivity for JAK2 over other JAK-family kinases such as JAK1, JAK3 and TYK2.
We tested NS-018 in a murine model of MPN induced by JAK2 V617F. Mice expressing JAK2 V617F controlled by the H2Kb promoter (V617F-TG mice) show an MPN phenotype: leukocytosis, thrombocytosis, progressive anemia, hepatosplenomegaly with extramedullary hematopoiesis, megakaryocyte hyperplasia and bone marrow fibrosis. They also exhibit body weight loss and high mortality compared to wild-type controls. Bone-marrow cells show constitutive activation of STAT5 and cytokine-independent growth of erythroid colony-forming units (CFU-E).
NS-018 inhibited cytokine-independent CFU-E growth and constitutive activation of STAT5 in V617F-TG cells in vitro. For in vivo experiments, V617F-TG mice were divided into treatment and vehicle control groups after disease was established at 12 weeks after birth. NS-018 was administered for 24 weeks by oral gavage at doses of 25 mg/kg or 50 mg/kg bid, and the control groups received vehicle only. Mice were monitored by blood counts, and a subset of mice was euthanized for detailed histopathology and fluorescence activated cell sorting analysis. During the study, 12 of 34 mice died in the vehicle group, whereas 1 of 36 mice died in the 50 mg/kg group. There was a statistically significant prolongation of survival in the 50 mg/kg group (p<0.01). Mice treated with NS-018 gained more weight than vehicle-treated mice, and were comparable to wild-type mice. V617F-TG at 12 weeks old showed severe leukocytosis with average white blood cell counts of 24 × 1010/L. After two weeks of NS-018 treatment, the leukocyte count was reduced to 59% in 25 mg/kg group and 39% in the 50 mg/kg group compared to vehicle group, and the effect was maintained until the end of the study. The inhibitory effect of NS-018 on T or B lymphocytes was much less than on myeloid cells. The 50 mg/kg group showed no progression of anemia. NS-018 treatment also improved hepatosplenomegaly in a dose-dependent manner. In the spleen, Mac-1/Gr-1+ myeloid cells associated with extramedullary hematopoiesis were significantly decreased, and B220+ B cells were increased by NS-018 treatment. In correlation with reduction of organ weights and infiltrating myeloid cells, there was also clear evidence of a dose-dependent reduction in the histopathology of extramedullary hematopoiesis in the spleen, liver, and lungs of NS-018-treated mice. In contrast to the improvement in the pathology of these organs, NS-018 had little impact on the progression of fibrosis and megakaryocyte hyperplasia in bone marrow. No significant toxicity was observed in treated mice.
In conclusion, NS-018 demonstrated therapeutic efficacy in a murine model of MPN induced by JAK2 V617F. In V617F-TG, which closely mimics human MPN, NS-018 significantly improved survival, body weight loss, hepatosplenomegaly, leukocytosis and anemia progression, thus confirming the viability of a targeted-therapy approach in managing JAK2 V617F positive MPNs. On the basis of these preclinical experiments, NS-018 appears to be an excellent candidate for phase I/II studies in patients with Bcr-Abl-negative MPNs.
Nakaya:Nippon Shinyaku Co., Ltd: Employment. Homan:Nippon Shinyaku Co., Ltd: Employment. Kotera:Nippon Shinyaku Co., Ltd: Employment. Shibayama:Nippon Shinyaku Co., Ltd: Employment. Naito:Nippon Shinyaku Co., Ltd: Employment. Shimoda:Nippon Shinyaku Co., Ltd: Research Funding.
Asterisk with author names denotes non-ASH members.