IL-1 Signaling Promotes Clonal Expansion and Progression of Bone Marrow Fibrosis in JAK2V617F-Induced Myeloproliferative Neoplasm

Myeloproliferative neoplasms (MPN) are a group of clonal hematopoietic stem cell derived myeloid malignancies characterized by aberrant production of myeloid, erythroid or megakaryocytic lineage cells. JAK2V617F is the most common somatic driver mutation associated with MPN. Interestingly, JAK2V617F mutation can also be detected in healthy individuals with clonal hematopoiesis of indeterminate potential (CHIP) who do not exhibit overt changes in blood counts. This suggests that other factors might be involved in association with JAK2 mutation in clonal expansion and initiation/progression of MPN. Chronic inflammation is frequently associated with MPN. Interleukin 1 (IL-1) is a major regulator of inflammation. IL-1 consists of two related cytokines IL-1α and IL-1β. Both IL-1α and IL-1β bind to the IL-1 receptor 1 (IL-1R1) to initiate downstream signaling. Although elevated expression of IL-1α and IL-1β has been observed in MPN, their role in the pathogenesis of MPN has remained elusive.

In this study, we investigated the role of IL-1 signaling in JAK2V617F-induced MPN using a Jak2V617F knock-in mouse model. We observed elevated levels of IL-1α and IL-1β in mice expressing heterozygous (Jak2 ^VF/+) and homozygous Jak2V617F (Jak2 ^VF/VF) compared with WT control animals. Notably, IL-1α and IL-1β expression was significantly higher in Jak2 ^VF/VF mice exhibiting extensive bone marrow (BM) fibrosis compared with Jak2 ^VF/+ mice exhibiting polycythemia vera (PV), consistent with elevated levels of IL-1 in patients with myelofibrosis (MF).

Since both IL-1α and IL-1β levels were elevated in Jak2 ^VF/VF mice exhibiting MF, we utilized conditional IL-1R1 knockout (IL-1R1cKO) and Jak2 ^VF/VF mice to assess the role of IL-1 signaling in the initiation/progression of MF. As expected, Jak2 ^VF/VF mice exhibited a significant increase in WBC, neutrophil and platelet counts compared to WT control mice. Deletion of IL-1R1in Jak2 ^VF/VF mice (IL-1R1cKO; Jak2 ^VF/VF) significantly reduced the WBC, neutrophil and platelet counts to almost control levels. Flow cytometric analysis also showed a significant reduction of myeloid (Gr-1 ⁺) and megakaryocytic (CD41 ⁺) precursors in the BM and spleens of IL-1R1cKO; Jak2 ^VF/VF mice compared to Jak2 ^VF/VF mice. Moreover, deletion of IL-1R1 significantly reduced hematopoietic stem and progenitor cells (HSPC) in the BM of IL-1R1cKO; Jak2 ^VF/VF mice compared to Jak2 ^VF/VF mice. Spleen weight was significantly reduced in IL-1R1cKO; Jak2 ^VF/VF mice compared with Jak2 ^VF/VF mice and they were comparable to control WT mice. More importantly, deletion of IL-1R1 markedly reduced BM fibrosis in Jak2 ^VF/VF mice. These data suggest an important role of IL-1 signaling in the progression of BM fibrosis in Jak2V617F-induced MPN.

To test whether IL-1 signaling contributes to clonal expansion of JAK2 mutant HSPC, we performed competitive transplantation assays by mixing Mx1Cre; Jak2 ^VF/+ and Mx1Cre; IL-1R1 ^F/F; Jak2 ^VF/+ mice BM cells with CD45.1 ⁺ WT mice BM cells at a ratio of 1:1 and transplanted into lethally irradiated CD45.1 ⁺ recipient animals. At 4 weeks after BMT, the recipient animals were injected with pI-pC to induce Jak2V617F expression and IL-1R1 deletion. We observed significantly higher percentages of total CD45.2 ⁺ cells as well as CD45.2 ⁺ myeloid (Gr-1 ⁺), B- and T-cells in the peripheral blood of chimeric mice receiving Jak2 ^VF/+ BM compared with chimeric mice receiving IL-1R1cKO; Jak2 ^VF/+ BM. We also observed significantly reduced percentages of CD45.2 ⁺ LSK, LK, Gr-1 ⁺ and CD41 ⁺ cells in the BM of chimeric recipient animals receiving IL-1R1cKO; Jak2 ^VF/+ BM compared with Jak2 ^VF/+ BM. These results suggest a role of IL-1 signaling in clonal expansion of Jak2V617F mutant HSPC.

Additionally, we tested the effects of blocking IL-1R1 using an anti-IL-1R1 antibody in the homozygous Jak2V617F knock-in mouse model of MF. We observed that anti-IL-1R1 antibody treatment significantly reduced peripheral blood WBC and neutrophil counts and decreased HSPC and myeloid precursors in the BM of Jak2 ^VF/VF mice. Furthermore, anti-IL-1R1 antibody treatment significantly reduced splenomegaly and markedly reduced BM fibrosis in Jak2 ^VF/VF mice, suggesting that therapies targeting IL-1R1 could be useful for the treatment of myelofibrosis. Overall, our results suggest that IL-1 signaling contributes to clonal expansion of Jak2V617F mutant HSPC and progression of bone marrow fibrosis in MPN.