Loss of DUSP1 Is Synthetic Lethal to JAK2^V617F

Introduction.

Myeloproliferative neoplasm (MPNs) are clonal stem cell disorders characterized by increased production of mature blood cells. Ph-negative MPNs (Ph- MPNs) encompass polycythemia vera (PV), essential thrombocythemia (ET), primary myelofibrosis (PMF) and chronic neutrophilic leukemia (CNL), amongst others. The discovery of Jak2 kinase activating mutations in MPNs provided a rationale for the targeting of Jak2 by small molecule kinase inhibitor. Interestingly, MPNs induced by MPL (MPL^W515L) or Calreticulin (CALR^fs-frame-shift mutations) were also found to be dependent on JAK2 signaling. Thus, provided a rationale for JAK2 targeting in this group of MPNs. However, unlike imatinib (BCR-ABL inhibitor) treatment in chronic myeloid leukemia (CML), ruxolitinib (Jak2 inhibitor) treatment does not eliminate the mutant clone. Besides, inhibition or genetic deletion of Jak2 kinases causes substantial hematologic toxicity that includes moderate to severe thrombocytopenia and anemia as well as non-hematological adverse events, including neurological toxicity. The latter led to the discontinuation of clinical development of several JAK2 inhibitors. Furthermore, mouse knock-out studies suggest that prolonged inhibition of Jak2 will be detrimental for normal hematopoietic and adult tissue homeostasis. These observations warrant identifying additional therapeutic targets that must be safe and able to eliminate the mutant clone.

Any therapy with the stated goal to successfully treat and possibly eradicate mutant cells and eventually cure must show differential toxicity toward tumor cells relative to normal cells. In this regard, CML represents a valuable paradigm to understand the mechanisms driving therapeutic response to TKI treatment. The cytotoxic response and clonal selectivity of BCR-ABL inhibitor are due to the dependence of leukemic cells on driver oncogene (BCR-ABL) kinase, where acute inhibition of BCR-ABL by imatinib results in oncogenic shock causing cell death. A recent study demonstrated that the AP1 transcription factor c-Fos and dual specificity phosphatase, Dusp1, mediate dependence to oncogenic kinase signaling. Perhaps more interestingly, these studies provided evidence that expression levels of c-Fos and Dusp1 regulate apoptotic thresh-hold in cancer cells such that lower levels confer sensitivity, while higher levels drive resistance to drug treatments. Thus, these two genes not only crucial for oncogenic transformation but also regulate the therapeutic efficacy of kinase inhibitors (Kesarwani M, et. al. Nature Medicine, 2017).

Results.

We reasoned that MPN induced by activated Jak2 signaling, similar to LSCs in CML, might have elevated expression of c-Fos and Dusp1 resulting to a higher apoptotic threshold, and thus, abrogating the cytotoxic effect of Jak2 inhibitors. As envisioned, a gene expression analysis of c-Fos and Dusp1 revealed higher expression of Dusp1 (4-10 fold in comparison to normal cells), but not the c-fos, in both mouse (induced by expression of Jak2^V617F, Mpl^W515L, and Calr^fs in mouse bonemarrow cells) and human primary MPN patient samples (data not presented). Interestingly, mice transplanted with Jak2^V617F expressing bonemarrow cells show dependence on Dusp1. Genetic deletion of Dusp1 is synthetic lethal to Jak2^V617F expressing hematopoietic cells (Fig. 1). Mice transplanted with Jak2^V617F expressing cells lacking Dusp1 show gradual depletion and completely cleared from the bonemarrow within five weeks of post-transplantation (Fig. 1E). In contrast, mice transplanted with cells expressing pMSCV-Ires-GFP (vector) and MPL^W515L show stable engraftment. These data provide evidence that Jak2^V617F is uniquely addicted to Dusp1 and crucial for Jak2^V617F mediate pathogenesis. Therefore, targeting of Dusp1 may exert curative response in Jak2 mutated MPNs, which constitute ~90% MPN cases.

Conclusion.

We show that both mouse and human MPNs have higher Dusp1 expression irrespective of MPN drivers (JAK2, MPL, and CALR). Jak2^V617F is uniquely addicted to the Dusp1 expression, where genetic deletion of Dusp1 is lethal to Jak2^V617F expressing cells, but not to the other MPN drivers, such as, MPL^W515L. Thus, providing a rationale for targeting the Dusp1 in Jak2 mutated MPNs to eliminate the mutant clones and cure the disease.

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