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. 2022 Dec 13;141(10):1105–1118. doi: 10.1182/blood.2022017619

Figure 2.

Figure 2.

Increased RUNX1A:RUNX1C ratio induces malignant ML-DS phenotype. (A-C) Neonatal (top) and fetal (bottom) CD34+ HSPCs were lentivirally transduced with RUNX1A, RUNX1C, or EV control. (A) Colonies after plating CD34+ HSPCs from 2 independent donors in methylcellulose-based colony-forming unit (CFU) assays (mean ± standard deviation [SD], n = 2, 1-way ANOVA). Percentage of immature (CD41+CD61+/CD42−) and mature megakaryocytes (CD41+CD61+/CD42+) (B) and immature CD34+ cells after 7 days in media promoting megakaryocytic differentiation (mean ± SD, n = 5, 2-way ANOVA) (C). (D-F) Cells derived from patients with ML-DS were lentivirally transduced with RUNX1A, RUNX1C, or EV control. (D) Percentage of GFP+ transduced cells normalized to day 0 after transduction (mean ± SD, n = 3, 1-way ANOVA, ∗∗P < .01). Bar graphs show the percentage of CD41+CD117+ and CD41+CD117− megakaryocytic cells (E) and CD33+CD117+ and CD33+CD117− myeloid cells (F) 5 days after transduction (mean ± SD, n = 3, 2-way ANOVA). (G) Experimental setup for evaluating RUNX1A:RUNX1C restoration in vivo. ML-DS blasts from 2 patients were transduced with RUNX1A (GFP+), RUNX1C (GFP+), or EV control (GFP+) and mixed 1:1 with EV control–transduced blasts (dTomato+), before transplantation into sublethally irradiated recipient mice. (H) Ratio of GFP+ to dTomato+ cells in the bone marrow of mice euthanized 4 to 8 weeks after transplantation (n = 5, Kruskal-Wallis test). dT, dTomato; ns, not significant.