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RUNX1 loss renders hematopoietic and leukemic cells dependent on IL-3 and sensitive to JAK inhibition
Amy C. Fan, … , Purvesh Khatri, Ravindra Majeti
Amy C. Fan, … , Purvesh Khatri, Ravindra Majeti
Published August 15, 2023
Citation Information: J Clin Invest. 2023;133(19):e167053. https://doi.org/10.1172/JCI167053.
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Research Article Hematology Inflammation

RUNX1 loss renders hematopoietic and leukemic cells dependent on IL-3 and sensitive to JAK inhibition

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Abstract

Disease-initiating mutations in the transcription factor RUNX1 occur as germline and somatic events that cause leukemias with particularly poor prognosis. However, the role of RUNX1 in leukemogenesis is not fully understood, and effective therapies for RUNX1-mutant leukemias remain elusive. Here, we used primary patient samples and a RUNX1-KO model in primary human hematopoietic cells to investigate how RUNX1 loss contributes to leukemic progression and to identify targetable vulnerabilities. Surprisingly, we found that RUNX1 loss decreased proliferative capacity and stem cell function. However, RUNX1-deficient cells selectively upregulated the IL-3 receptor. Exposure to IL-3, but not other JAK/STAT cytokines, rescued RUNX1-KO proliferative and competitive defects. Further, we demonstrated that RUNX1 loss repressed JAK/STAT signaling and rendered RUNX1-deficient cells sensitive to JAK inhibitors. Our study identifies a dependency of RUNX1-mutant leukemias on IL-3/JAK/STAT signaling, which may enable targeting of these aggressive blood cancers with existing agents.

Authors

Amy C. Fan, Yusuke Nakauchi, Lawrence Bai, Armon Azizi, Kevin A. Nuno, Feifei Zhao, Thomas Köhnke, Daiki Karigane, David Cruz-Hernandez, Andreas Reinisch, Purvesh Khatri, Ravindra Majeti

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Figure 2

RUNX1 loss in human HSPCs causes hematopoietic and stem cell defects.

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RUNX1 loss in human HSPCs causes hematopoietic and stem cell defects.
(...
(A) CD34+ HDR HSPCs were plated in methylcellulose-based colony-forming assays and assessed for colony formation at 14 days. n = 4 CB. Two-Way ANOVA, Šidák’s multiple-comparison test: **P < 0.01, ***P < 0.001, ****P < 0.0001. (B) CD34+ HDR HSPCs were plated in collagen-based megakaryocyte colony-forming assays, and colonies were quantified after 10 days. n = 4 CB. Paired t test: ***P < 0.001. (C) CD34+ HDR HSPCs were plated in stem retention media (serum-free media with SCF, TPO, and FLT3L) and analyzed by flow cytometry for cell count at days 6. n = 8 CB donors. Paired t test: ****P < 0.0001. BFU, burst-forming units. GEMM, granulocyte, erythrocyte, monocyte, megakaryocyte. (D) CD34+ HDR HSPCs were incubated with EdU for 2 hours, stained with DAPI, and evaluated for cell-cycle status. n = 3 CB. Two-way ANOVA, Šidák’s multiple-comparison test: **P < 0.01. (E) CD34+ HDR HSPCs were injected intrafemorally into sublethally irradiated NSG mice, and human CD45+ HDR+ engraftment was evaluated upon sacrifice (at 24–26 weeks after transplantation). Unpaired t test NS (not significant). n = 5 CB donors, 9–18 mice. (F) BFP+ AAVS1 and mCherry+ RUNX1-KO cells were injected intrafemorally at a 1:1 ratio into NSG mice, and relative engraftment within the human CD45+ compartment was evaluated 18 weeks after transplantation. The FACS plot indicates representative ratio upon injection. The graph shows relative engraftment at 18 weeks; paired t test: **P < 0.01. n = 3 CB donors, 6 mice.

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