GLS in Philadelphia-negative and JAK2 V617F-driven myeloproliferative neoplasms (MPNs).

Abstract

e15092 Background: Philadelphia-negative myeloproliferative neoplasms (MPNs) are clonal hematopoietic malignancies. Nearly all MPNs are driven by somatic mutations in either: JAK2, CALR, or MPL. Although all three MPN driver mutations lead to constitutive activation of JAK/STAT signaling, JAK inhibitors are not curative and fail to alter disease progression and display unwanted side effects. Allogeneic stem cell transplantation remains the only curative therapy for MPNs, but is associated with substantial morbidity and mortality. Recently, glutaminolysis has been shown to play a critical role in cancer cell metabolism. During glutaminolysis, glutamine is metabolized through in a two-step reaction, the first of which involves the enzyme glutaminase (GLS) catalyzing the hydrolysis of glutamine to glutamate. Glutamate can then fuel for energy production into the TCA cycle, among several other possible metabolic fates. As many cancers have proven to be dependent on this pathway, targeting GLS has become an attractive therapeutic avenue. Because glutaminolysis has been understudied in MPNs, we sought to determine whether this pathway represents a novel, targetable vulnerability in MPNs. Methods: We evaluated the mRNA levels of GLS in peripheral blood mononuclear cells from 30 MPN patients and 5 healthy donors. We tested by qPCR for GLS and the protein levels of GLS by western blot in TF-1 cells, a human myeloid cell line, stably overexpressing either JAK2, MPL, or CALR mutated proteins. The latter also under treatment with the JAK inhibitor ruxolinitinib. We also evaluated the GLS enzyme activity through the use of a fluorimetric assay. Lastly, we tested the sensitivity of MPN cells to GLS inhibition with a GLS inhibitor, CB-839, which is currently in advanced phase clinical trials for other cancers including Myelodysplastic Syndromes. Results: GLS mRNA expression was increased in all MPN patients regardless of their driver mutation, where expression in JAK2V617F patients was higher in MF versus ET patients (p < 0.001). GLS protein expression and activity were increased in TF-1 cells expressing JAK2, MPL, and CALR mutations. We also found that GLS mRNA and protein expression was up-regulated in a JAK/STAT-dependent manner. Interestingly, despite increased expression of GLS across all MPN driver mutations, only JAK2 V617F cells demonstrated significant sensitivity to GLS inhibition with CB-839 in vitro and with preliminary data in vivo. We found that combination treatment with JAK inhibitor ruxolitinib further inhibited cell viability. Conclusions: GLS up-regulation is a common feature of all MPNs, and is JAK/STAT dependent. JAK2 V617F expressing cells show significant sensitivity to GLS inhibition. Combination treatment with ruxolitinib further enhances this effect. Treatment with CB-839 may thus represent a novel therapeutic avenue for JAK2 V617F+ MPNs.