Abstract 1172: JAK2V617F-mutant megakaryocytes contribute to stem cell aging in myeloproliferative neoplasms

The myeloproliferative neoplasms (MPNs) are stem cell disorders characterized by hematopoietic stem cell (HSC) expansion and an increased propensity to develop hematologic malignancies. The acquired kinase mutation JAK2V617F plays a central role in MPNs. In this work we tested the hypothesis that the JAK2V617F-bearing megakaryocytes (MKs), an important component of the hematopoietic niche, promotes HSC dysfunction in MPNs. Methods JAK2V617F Flip-Flop (FF1) mice and Pf4-Cre mice (Radek Skoda, Switzerland) were crossed to generate MK lineage-specific human JAK2V617F knock-in mouse line (Pf4/FF1). Animal experiments were performed in accordance with the Institutional Animal Care and Use Committee guideline. Results We previously reported that, at 28 wks of age, the Pf4/FF1 mice developed a murine myeloproliferative syndrome with moderate thrombocytosis, splenomegaly, increased marrow MKs, and a 3-fold increase of CD45+CD201+CD150+CD48- HSCs with increased donor engraftment in a competitive repopulation assay compared to Pf4-cre control mice. (Zhan Leukemia 2016; Zhang Stem Cells 2018) Rigorous, extremely sensitive RT-PCR assays eliminated the possibility that HSCs in Pf4/FF1 mice express the mutant kinase. To study the effect of JAK2V617F-bearing MKs on HSC function during aging, we followed the Pf4/FF1 mice up to 2 years of age. At 2 yr, there were significant neutrophilia in Pf4/FF1 mice but no change was observed in hemoglobin, platelet count, or marrow HSC cell numbers compared to age-matched control mice. Indeed, while HSC numbers significantly increased in old (2-yr) control mice compared to young (6-mo) control mice (3.3-fold, p=0.0002, n=4-8), there was no significant increase of HSCs between old and young Pf4/FF1 mice (1.6-fold, p=0.158, n=6-8). A competitive marrow transplantation experiment was performed in which CD45.2 donor marrow cells from 2-yr old Pf4/FF1 or control mice were injected intravenously together with competitor CD45.1 wild-type marrow cells intro lethally irradiated wild-type recipient (CD45.1) (n=8-9 in each group). During a 4-month follow up, old HSCs from the Pf4/FF1 mice displayed hallmarks of murine hematopoietic aging including a lower peripheral blood donor chimerism, increased myeloid blood cell production, and decreased lymphoid cells compared to old HSCs from control mice. To begin to understand how JAK2V617F-bearing MKs promote the aging phenotype of HSCs in old Pf4/FF1 mice, qRT-PCR of purified HSCs identified up-regulation of p21 (2-fold, p = 0.019) in Pf4/FF1 mice compared to controls. Conclusion The Pf4/FF1 mice, in which the JAK2V617F expression is restricted to MKs, developed a myeloproliferative phenotype at young age and accelerated stem cell aging at old age. The precise mechanism by which JAK2V617F-bearing MKs affect the MPN marrow niche and stem cell dysfunction during disease progression deserve further investigation. Citation Format: Helen Wong, Melissa Castiglione, Molly Quan, Huichun Zhan. JAK2V617F-mutant megakaryocytes contribute to stem cell aging in myeloproliferative neoplasms [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1172.