Breakthroughs in myeloproliferative neoplasms

Abstract

The discovery of the JAK2V617F mutation ushered the field of Philadelphia-negative myeloproliferative neoplasms (MPNs) into the era of targeted therapy. Currently, there are several JAK2 inhibitors in clinical trials for patients with MPNs, particularly for patients with myelofibrosis (MF). These drugs act by blocking the proliferation of neoplastic cells by disrupting the JAK2–STAT signaling and by abrogating inflammatory cytokine signaling which is dependent on JAK kinases. Therapy with JAK2 inhibitors can improve splenomegaly and debilitating constitutional symptoms in great majority of MF patients, improving greatly their quality of life. Long-term follow-up will reveal whether these drugs can also prolong survival by better controlling signs and symptoms of the MF. There are other compounds in clinical trials for MPNs, including the new immunomodulatory drug pomalidomide, and inhibitor of mammalian target of Rapamycin everolimus. In this article, we briefly review the latest therapeutic advances in the field of Philadelphia-negative MPNs.

Keywords:

• JAK2V617F
• JAK2 inhibitors
• Ruxolitinib
• Everolimus
• Pomalidomide

Introduction

The classic Philadelphia-negative myeloproliferative neoplasms (Ph-negative MPNs) include polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis. MF can also develop secondary to transformation of both PV and ET (post-PV/ET MF). Our understanding of the pathophysiology of these disorders changed in 2005 with the discovery of the JAK2V617F mutation in about 95% of patients with PV and 60% of patients with ET and MF.1 The JAK2V617F mutation causes cytokine hyper-responsiveness and constitutive activation of the JAK2 tyrosine kinase (TK) with subsequent increased signaling through receptors associated with JAK2, such as erythropoietin receptor (EPOR) and thrombopoietin receptor (MPL).1 The net result is increased activation of pro-survival and anti-apoptotic pathways, including the JAK–STAT and phosphatidylinositol 3-kinase/Akt/mammalian target of Rapamycin (PI3K–Akt–mTOR) pathways.1 Discovery of the JAK2V617F mutation ushered Ph-negative MPNs in the era of molecular biology and targeted therapy. In the years after the report of JAK2V617F, other mutations such as JAK2 exon 12 mutations and MPL W515K/L mutations were detected in patients with JAK2V617F-negative MPNs, all sharing the common theme of leading to increased signaling through receptors of hematopoietic cytokines. Other mutations in genes unrelated to cytokine signaling, such as TET2, ASXL1, RUNX1, IDH1, IDH2, and EZH2, were recently reported, revealing a level of complexity in the pathogenesis of these disorders that was previously unknown.1 More importantly, these mutations represent possible therapeutic targets for intervention, and after just 2 years from the discovery of JAK2V617F, clinical trials with JAK2 inhibitors in patients with MF started enrollment.2 It is now an exciting time in the field of Ph-negative MPNs, and we review here the more recent clinical results of novel agents for these disorders.

JAK2 Inhibitors

Introduction

The JAK2 enzyme belongs to the JAK2 family of TKs, including JAK1, JAK2, JAK3, and TYK2. JAK2 signaling is essential for erythropoiesis, thrombopoiesis, and granulocyte development, as it associates with the intracellular portion of EPOR, MPL, and receptors which utilize CD131 (e.g. receptors for GM-CSF and IL-3). The JAK2V617F mutation occurs in the JH2 (JAK homology 2) domain, also called the pseudokinase domain.1 The V617F mutation leads to increased JAK2 activity by disrupting the inhibitory activity the JH2 domain exerts over the JH1 domain.

Similar to other TK inhibitors in current use in hematology, JAK2 inhibitors are adenosine triphosphate (ATP)-mimetic drugs which compete with ATP for the ATP-binding site in the TK, since ATP is the major source of phosphate for the phosphorylation reaction. The V617F mutation is located in the JH2 domain, and most currently available JAK2 inhibitors do not distinguish between V617F-mutated and non-mutated cases. This might explain the observed fact that these drugs have clinical activity in MF independently of the JAK2V617F mutational status.2 Recent evidence suggests that all patients with MPNs have common underlying pathophysiological abnormality in the deregulated JAK–STAT signaling, regardless of the presence or absence of JAK2V617F mutation, or any other known mutations.1

Besides abrogating neoplastic signaling induced by abnormally active JAK2–STAT pathway, there is evidence to suggest that some JAK2 inhibitors (compounds that are dual JAK1/JAK2 inhibitors) also work by normalizing pro-inflammatory cytokine levels and inhibiting cytokine signaling.2 MF is considered a pro-inflammatory disorder, and elevated levels of many cytokines (e.g. IL-6, IL-8, tumor necrosis factor-alpha) are present and contribute to the bone marrow (BM) fibrosis, osteosclerosis, angiogenesis, and development of constitutional symptoms seen in this disease. In studies with dual JAK1/JAK2 inhibitors cytokine levels normalize, and this was associated with improvement in systemic symptoms, weight gain, and functional capacity.2

Clinical results with selected JAK2 inhibitors (Table 1)

Table 1. JAK2 inhibitors in clinical development at the present time

Drug	Current phase	Target(s)	Spleen response rate (by IWG-MRT)	Other responses	Toxicities
Ruxolitinib	III	JAK1, JAK2	44%	Symptoms, pruritus, exercise ability	Diarrhea, anemia, thrombocytopenia
TG101348	I/II; III planned	JAK2	39%	Symptoms, pruritus, exercise ability	Diarrhea, anemia
CYT387	I/II	JAK1, JAK2, TYK2	46%	Symptoms, pruritus, exercise ability, erythroid response rate	Peripheral neuropathy, ‘first dose’ effect
LY2784544	I	JAK2 V617F	NA	NA	NA
NS-018	I	JAK2	NA	NA	NA
BMS911543	I	JAK2	NA	NA	NA

Ruxolitinib is an orally available JAK1/JAK2 inhibitor that had activity in animal models of MPNs. A phase I/II study evaluated ruxolitinib in 153 patients with MF.2 The maximum tolerated dose (MTD) was 25 mg twice daily, but a dose adjusted schedule starting at 15 mg twice daily with subsequent increases in case of no response represented the best balance between safety and efficacy. The response rate (RR) by the International Working Group on Myelofibrosis Research and Treatment (IWG-MRT) criteria was 44% for the whole cohort, and 52% in the cohort who started at 15 mg twice daily. Importantly, RRs were not affected by JAK2 mutational status: 51% (V617F) versus 45% (wild-type). There was also significant improvement in exercise ability (as measured by the 6-minute walking test), myelofibrosis-associated symptoms and weight gain. Improvement in symptoms was correlated with a decrease in the levels of pro-inflammatory cytokines. No improvement in BM fibrosis or JAK2 allele burden was seen. Grade 3–4 toxicities included thrombocytopenia and new onset of transfusion dependent anemia. Patients who started in the 15 mg twice daily cohort had lower rates of grade 3–4 thrombocytopenia (3%) and anemia (8%).

Two phase III studies of ruxolitinib were recently presented in abstract form.3,4 These studies compared ruxolitinib against placebo (COMFORT-I) or best available therapy (COMFORT-II). The primary endpoint of both trials was the proportion of patients with a ⩾35% reduction in spleen volume as assessed by magnetic resonance imaging (corresponding to a ⩾50% reduction in spleen length by palpation). Both studies confirmed the superiority of ruxolitinib: RR was 42% versus 1% (COMFORT-I; P<0·0001) and 28·5% versus 0% (COMFORT-II).3,4 Secondary endpoints included improvements in quality of life and symptoms and also showed better results with ruxolitinib. Similar to the phase I/II trial, most common toxicities with ruxolitinib were grade 3–4 anemia and thrombocytopenia, but rarely led to discontinuation of therapy (one patient for each toxicity).

Patients with PV and ET who are refractory or intolerant to conventional therapies (e.g. hydroxyurea, anagrelide) may also benefit from JAK2 inhibitors. A phase II study recruited 73 patients (PV = 34; ET = 39) who were refractory or intolerant to hydroxyurea.5 The MTD of ruxolitinib was 10 mg twice daily (PV) and 25 mg twice daily (ET). In PV, ruxolitinib decreased hematocrit to <45% and led to transfusion independence in 97% of patients. There was improvement in systemic symptoms such as pruritus and night sweats, and splenomegaly. In ET, platelets decreased to below upper limit of normal in 49% of patients after a median of 15 days, and most patients also had improvement in symptoms and splenomegaly.

TG101348 (also known as SAR302503) is a potent, orally available, selective JAK2 inhibitor. A phase I/II trial of TG101348 recruited 59 patients with high- or intermediate-risk MF (primary or post-PV/ET).6 The phase I portion of the trial determined the MTD to be 680 mg, and dose-limiting toxicities were grade 3–4 elevations in amylase and lipase. The RR in spleen size by the IWG-MRT criteria was 39% after six cycles of therapy, and 45% in MTD dose cohort. There was improvement in systemic symptoms, including early satiety, fatigue, night sweats, cough, and pruritus. However, no reduction in inflammatory cytokines was observed. Non-hematological toxicities with TG101348 include nausea (all grades: 69·5%; grades 3–4: 3·4%), vomiting (all grades: 57·6%; grades 3–4: 3·4%), and diarrhea (all grades: 64·4%; grades 3–4: 10·2%). New-onset of grade 3–4 transfusion-dependent anemia occurred in 35% of cases, while grade 3–4 thrombocytopenia was detected in 23·7%. Patients enrolled in the phase I/II trial have been transferred to an extension study to evaluate the long-term effects of TG101348 and a phase III trial is in the planning.

CYT387 is a JAK1/JAK2/TYK2 inhibitor that is currently in phase I/II clinical trials for patients with MF (primary or post-PV/ET). In pre-clinical studies, CYT387 normalized hematopoiesis and increased erythroid progenitors in the bone marrow of mice with JAK2V617F positive MPN. A phase I/II clinical trial was presented recently.7 So far, 60 patients have been recruited. The MTD was 300 mg once daily, and doses of 150 and 300 mg were deemed to be clinically effective. The splenomegaly RR was 46%, and median time to response was 0·4 months. Similar to other JAK2 inhibitors, there was also striking reductions in constitutional symptoms such as fever (CR: 100%), pruritus (CR: 75%), and night sweats (CR: 86%). Among 42 patients evaluable for anemia response (including 33 who were transfusion-dependent at baseline), the erythroid RR was 50%, and 58% in transfusion-dependent patients according to IWG-MRT criteria. Median duration of transfusion independency was 7·5 months. Unfortunately, application of IWG-MRT criteria has also been shown to result in responses in patients not on any therapy (e.g. placebo-treated patients in COMFORT-I blinded phase 3 ruxolitinib versus placebo trial, responded in 47% of cases)3 and improved, clinically significant criteria are urgently needed. Peripheral neuropathy (grade 1 only, 28%) and a first dose effect (defined as lightheadedness some time after taking the first dose of the drug; seen in 43% of subjects, grade 1) were the most common non-hematological side effects. Grade 3–4 thrombocytopenia developed in 27% of subjects, and grade 4 neutropenia in 5%. The clinical trial is still ongoing and more results are expected in major medical meetings.

Overall, the results of these clinical trials have demonstrated that JAK2 inhibitors have activity in most patients with MF and can be of great clinical value in treating such patients. The decrease in splenomegaly and improvement in debilitating constitutional symptoms can ameliorate these patients quality of life to a great extent. Future studies will continue to evaluate the role of JAK2 inhibitors for therapy of MF and other MPN, either alone or in combination with other drugs.

Immunomodulatory Drugs

As mentioned, Ph-negative MPNs, in particular MF, are characterized by a deregulation of the immunological microenvironment and an increase in the levels of pro-inflammatory, pro-angiogenic, and fibrogenic cytokines, which contribute to disease pathogenesis by inducing osteosclerosis, BM fibrosis, mobilization of hematopoietic stem cells into the peripheral blood, and development of extramedullary hematopoiesis. Immunomodulatory drugs have anti-cytokine and anti-angiogenic properties. There are three immunomodulatory drugs in clinical use in MF: thalidomide, lenalidomide, and pomalidomide. We will focus on the results of the most recent compound, pomalidomide.

Pomalidomide is a thalidomide analogue that has 10 000-fold greater potency. A randomized phase II trial evaluated pomalidomide in 84 patients with MF.8 Patients could receive one of four therapies: pomalidomide 2 mg daily plus placebo (N = 22), pomalidomide 2 mg daily plus prednisone (N = 19), pomalidomide 0·5 mg daily plus prednisone (N = 22), and prednisone alone (N = 21).8 The best RR was seen in the pomalidomide 0·5 mg plus prednisone arm, with a 36% improvement in Hb and transfusion dependency. In another, non-randomized phase II study, 58 patients were treated with low-dose (0·5 mg) pomalidomide.9 Patients had to be transfusion dependent or have an Hb <10 g/dl. The overall RR was 17%, and was higher in JAK2V617F mutated patients (24% versus 0%). Other features associated with response include development of basophilia in the first month of therapy (38% versus 6%) and not having marked splenomegaly (38% versus 11%). Currently, a phase III study is randomizing transfusion-dependent MF patients to pomalidomide or placebo. Results are eagerly awaited.

Inhibitors of mTOR

The PI3K–Akt–mTOR pathway regulates diverse cellular processes such as proliferation, apoptosis, and cell growth and plays an important role in several types of tumors. PI3K can be activated directly by JAK2, with subsequent activation of Akt and subsequently mTOR. mTOR is a serine/threonine kinase which regulates protein synthesis, transcription, cell growth, proliferation, and apoptosis. Cells from patients with JAK2V617F-positive neoplasms show increased Akt phosphorylation, indicating constitutive activation of this pathway.

The immunosuppressant rapamycin (sirolimus) was the first mTOR inhibitor to be developed. Next-generation mTOR inhibitors include sirolimus analogues everolimus and temsirolimus. These drugs are currently approved for therapy of mantle cell lymphoma and renal cell carcinoma, among other indications. Based on pre-clinical data showing activation of Akt in JAK2V617F-positive MPNs, a phase I/II study evaluated the efficacy of everolimus as a single agent for therapy of patients with MF.10 No dose-limiting toxicity was observed with doses up to 10 mg/day, the highest dose was evaluated. A spleen response by IWG-MRT criteria was documented in 20% of patients. Patients also reported complete improvement of systemic symptoms (69%) and pruritus (80%). The overall RR by IWG-MRT criteria was 23%. Reduced phosphorylation of p70S6K (a known target of mTOR) was observed in responders while non-responders maintained phosphorylation status of this enzyme. Another biomarker of response was mRNA level of CCND1, a gene upregulated by the PI3K–Akt–mTOR pathway, with responders having lower mRNA level than non-responders. This study provides proof that targeting mTOR can be a valuable strategy in the therapy of Ph-negative MPNs.

Conclusion

The past 6 years have seen a great deal of advances in the field of Ph-negative MPNs, with increased understanding of the pathogenesis of these entities and the arrival of new therapeutic agents. However, we still have little understanding of how the various genetic lesions cooperate to produce the MPN phenotype. We also do not know precisely the mechanism of action of new agents, and we need to identify biomarkers for response so that therapy can be better tailored for the individual patient. Most studies still rely on these drugs as single agents, and future clinical trials will evaluate combination therapy in order to improve responses. With greater collaboration between basic and translational investigators and with help from patients, we can hope to surpass these challenges and provide better therapies for patients with Ph-negative MPNs.