Abstract
BackgroundDysregulated signaling of the JAK/STAT pathway is a common feature of chronic myeloproliferative neoplasms (MPN), usually associated with JAK2V617F mutation. Recent clinical trials with JAK2 inhibitors showed significant improvements in splenomegaly and constitutional symptoms in patients with myelofibrosis but meaningful molecular responses were not documented. Accordingly, there remains a need for exploring new treatment strategies of MPN. A potential additional target for treatment is represented by the PI3K/AKT/mammalian target of rapamycin (mTOR) pathway that has been found constitutively activated in MPN cells; proof-of-evidence of efficacy of the mTOR inhibitor RAD001 has been obtained recently in a Phase I/II trial in patients with myelofibrosis. The aim of the study was to characterize the effects in vitro of mTOR inhibitors, used alone and in combination with JAK2 inhibitors, against MPN cells.FindingsMouse and human JAK2V617F mutated cell lines and primary hematopoietic progenitors from MPN patients were challenged with an allosteric (RAD001) and an ATP-competitive (PP242) mTOR inhibitor and two JAK2 inhibitors (AZD1480 and ruxolitinib). mTOR inhibitors effectively reduced proliferation and colony formation of cell lines through a slowed cell division mediated by changes in cell cycle transition to the S-phase. mTOR inhibitors also impaired the proliferation and prevented colony formation from MPN hematopoietic progenitors at doses significantly lower than healthy controls. JAK2 inhibitors produced similar antiproliferative effects in MPN cell lines and primary cells but were more potent inducers of apoptosis, as also supported by differential effects on cyclinD1, PIM1 and BcLxL expression levels. Co-treatment of mTOR inhibitor with JAK2 inhibitor resulted in synergistic activity against the proliferation of JAK2V617F mutated cell lines and significantly reduced erythropoietin-independent colony growth in patients with polycythemia vera.Conclusions/SignificanceThese findings support mTOR inhibitors as novel potential drugs for the treatment of MPN and advocate for clinical trials exploiting the combination of mTOR and JAK2 inhibitor.
Highlights
The chronic myeloproliferative neoplasms (MPN), which include polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (PMF) [1], are characterized by a V617F point mutation in the exon 14 of Janus Kinase 2 (JAK2) that occurs in 95% of PV and 60% of ET or PMF patients [2,3]
Dysregulation of the JAK2/STAT pathway represents a central mechanism in the pathogenesis of MPNs: (i) the JAK2V617F gain-of-function mutation occurs in the majority of patients with PV and 60% of PMF and essential thrombocythemia [2,3], (ii) other mutations (MPL, LNK, CBL) found in 5–10% can activate the JAK/STAT pathway [5,6,7,8,50], (iii) mouse models indicate that those mutations are able to induce a myeloproliferative disorder [11,12,13,14,15,16,17,18,19], (iv) the JAK/STAT pathway is involved in the dysregulated cytokine expression that accompanies MPNs and underlies some tract of the clinical phenotype [51], and (v) targeting activated JAK2 with ATP-competitive JAK2 inhibitors resulted in measurable clinical improvements in patients with myelofibrosis [25,26]
In this study we have focused on the mammalian target of rapamycin, a key downstream target of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway, with the objective to characterize the efficacy of mTOR inhibitors in different cellular models of MPNs, including primary cells
Summary
The chronic myeloproliferative neoplasms (MPN), which include polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (PMF) [1], are characterized by a V617F point mutation in the exon 14 of Janus Kinase 2 (JAK2) that occurs in 95% of PV and 60% of ET or PMF patients [2,3]. JAK2 exon 12 mutations are detected in 2–3% of PV [4] and mutations in MPL, the receptor for thrombopoietin, have been reported in 5–10% of ET and PMF patients [5,6,7,8] These molecular abnormalities result in a constitutive activation of the JAK/signal transducer and activator of transcription (STAT) signaling pathway and contribute to cytokine hypersensitivity and cytokine independent growth of the mutant cells, as exemplified by the erythropoietin-independent erythroid colonies (EEC) typically found in most PV patients [9,10]. It seems unlikely that eradication of the MPN clone can be achieved with (available) JAK2 inhibitors; novel drugs and more effective therapeutic strategies need to be sought In this regard, it has been shown that cotreatment of the HDACi panobinostat and the JAK2 inhibitor TG101209 resulted in greater attenuation of JAK/STAT signaling in human and mouse JAK2V617F-mutated cells and increased cytotoxicity against MPN CD34+ cells compared to individual drugs [28]. The aim of the study was to characterize the effects in vitro of mTOR inhibitors, used alone and in combination with JAK2 inhibitors, against MPN cells
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