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. 2019 Sep 6;134(16):1323–1336. doi: 10.1182/blood.2019000015

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© 2019 by The American Society of Hematology

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graphic file with name bloodBLD2019000015f2-1.jpg — Suz12 loss cooperates with JAK3(M511I) in driving an aggressive T-ALL in vivo. (A) Scheme of bone marrow transplantation set-up. HSPCs were isolated from the bone marrow of Cas9 donor mice, followed by retroviral transduction with constructs overexpressing JAK3(M511I) and/or Suz12gRNA before injection into recipient mice. (B) Clonal evolution of different populations (WT, S, J, J+S) in the blood of a representative JAK3(M511I)+Suz12gRNA mouse (M1R15) over time, showing competitive advantage of J+S cells over other populations. Cell populations: nontransduced (wild-type; WT), Suz12gRNA (S; BFP only), JAK3(M511I) (J; GFP only), and JAK3(M511I)+Suz12gRNA (J+S, BFP+GFP double positive). (C) WBC counts of recipient mice over time for 4 different cohorts: JAK3(M511I)+Suz12gRNA, JAK3(M511I), Suz12gRNA, and control (GFP and BFP empty vectors, EV) mice. A WBC count of 30 000 was used as cutoff for DFS. (D) Survival curve showing DFS of mice in the same 3 cohorts as in C. Mice in which Suz12gRNA-TS5 instead of TS1 was used were designated with a box and darker color. P values were calculated with Gehan-Breslow-Wilcoxon test. (E-F) Survival curves of JAK3(M511I)+Suz12gRNA (E) and Suz12gRNA (F) leukemias that were serially transplanted. (G) Representative flow cytometry stainings for CD8 (APC-Cy7, y-axis) and CD4 (PerCP-Cy5.5, x-axis) of thymus and spleen of leukemia cells (pregated on GFP and/or BFP) from 3 different cohorts at time of sacrifice. For all figures except Fig. 2D, Suz12gRNA TS1 was used in Suz12gRNA and JAK3(M511I)+Suz12gRNA mice.

graphic file with name bloodBLD2019000015f2-2.jpg — Suz12 loss cooperates with JAK3(M511I) in driving an aggressive T-ALL in vivo. (A) Scheme of bone marrow transplantation set-up. HSPCs were isolated from the bone marrow of Cas9 donor mice, followed by retroviral transduction with constructs overexpressing JAK3(M511I) and/or Suz12gRNA before injection into recipient mice. (B) Clonal evolution of different populations (WT, S, J, J+S) in the blood of a representative JAK3(M511I)+Suz12gRNA mouse (M1R15) over time, showing competitive advantage of J+S cells over other populations. Cell populations: nontransduced (wild-type; WT), Suz12gRNA (S; BFP only), JAK3(M511I) (J; GFP only), and JAK3(M511I)+Suz12gRNA (J+S, BFP+GFP double positive). (C) WBC counts of recipient mice over time for 4 different cohorts: JAK3(M511I)+Suz12gRNA, JAK3(M511I), Suz12gRNA, and control (GFP and BFP empty vectors, EV) mice. A WBC count of 30 000 was used as cutoff for DFS. (D) Survival curve showing DFS of mice in the same 3 cohorts as in C. Mice in which Suz12gRNA-TS5 instead of TS1 was used were designated with a box and darker color. P values were calculated with Gehan-Breslow-Wilcoxon test. (E-F) Survival curves of JAK3(M511I)+Suz12gRNA (E) and Suz12gRNA (F) leukemias that were serially transplanted. (G) Representative flow cytometry stainings for CD8 (APC-Cy7, y-axis) and CD4 (PerCP-Cy5.5, x-axis) of thymus and spleen of leukemia cells (pregated on GFP and/or BFP) from 3 different cohorts at time of sacrifice. For all figures except Fig. 2D, Suz12gRNA TS1 was used in Suz12gRNA and JAK3(M511I)+Suz12gRNA mice.