Figure 2.
Mutant Idh2 does not cooperate with KMT2A-MLLT3 in vitro or in vivo. (A) Schematic diagram of experimental workflow. LSK cells or GMPs from WT or Vav1-Cre;Idh2R140Q/WT mice were transduced with KMT2A-MLLT3-GFP, sorted, and transplanted into sublethally irradiated WT recipients or plated in methylcellulose for colony-formation assays. (B-C) Colony formation and serial replating of LSK cells from Vav1-Cre;Idh2R140Q/WT or WT control mice transduced with KMT2A-MLLT3. Number of colonies (B) and number of cells (C) at each plating, per 500 cells plated in methyl cellulose of KMT2A-MLLT3–transformed LSK cells from 2 individual experiments. Cells were replated every 5 days. Error bars represent SD. (D) Survival of sublethally irradiated mice transplanted with 10 000 Vav1-Cre;Idh2R140Q/WT LSK cells transduced with KMT2A-MLLT3 (n = 5) or transplanted with 10 000 WT LSK cells transduced with KMT2A-MLLT3 (n = 5). (E) Growth of WT + KMT2A-MLLT3 and Vav1-Cre;Idh2R140Q/WT + KMT2A-MLLT3 leukemic cells [derived from leukemic mice in (D)] were treated with increasing concentrations of DOT1L inhibitor (EPZ5676). Cells were counted every 3 days using trypan blue, and 10 000 cells were replated and treated with fresh compound. Error bars represent SD. (F) Growth of WT + KMT2A-MLLT3 and Vav1-Cre;Idh2R140Q/WT + KMT2A-MLLT3 leukemic cells [from (D)] treated with increasing concentrations of mutant IDH2 inhibitor (AGI-6780). Cells were counted every 3 days using trypan blue, and 10 000 cells were replated and treated with fresh compound. Error bars represent the SD of technical duplicates. (G-H) Colony formation and serial replating of GMPs from Vav1-Cre;Idh2R140Q/WT or WT control mice transduced with KMT2A-MLLT3. Number of colonies (G) and number of cells (H) at each plating, per 500 cells plated in methyl cellulose of KMT2A-MLLT3–transformed GMPs from 2 individual transplants. Cells were replated every 5 days. Error bars represent SD. (I) Survival of sublethally irradiated mice transplanted with 10 000 Vav1-Cre;Idh2R140Q/WT GMPs transduced with KMT2A-MLLT3 (n = 5) or with 10 000 WT GMPs transduced with KMT2A-MLLT3 (n = 5). *P < .01, **P < .001; Student t test. DMSO, dimethyl sulfoxide; ns, not significant.

Mutant Idh2 does not cooperate with KMT2A-MLLT3 in vitro or in vivo. (A) Schematic diagram of experimental workflow. LSK cells or GMPs from WT or Vav1-Cre;Idh2R140Q/WT mice were transduced with KMT2A-MLLT3-GFP, sorted, and transplanted into sublethally irradiated WT recipients or plated in methylcellulose for colony-formation assays. (B-C) Colony formation and serial replating of LSK cells from Vav1-Cre;Idh2R140Q/WT or WT control mice transduced with KMT2A-MLLT3. Number of colonies (B) and number of cells (C) at each plating, per 500 cells plated in methyl cellulose of KMT2A-MLLT3–transformed LSK cells from 2 individual experiments. Cells were replated every 5 days. Error bars represent SD. (D) Survival of sublethally irradiated mice transplanted with 10 000 Vav1-Cre;Idh2R140Q/WT LSK cells transduced with KMT2A-MLLT3 (n = 5) or transplanted with 10 000 WT LSK cells transduced with KMT2A-MLLT3 (n = 5). (E) Growth of WT + KMT2A-MLLT3 and Vav1-Cre;Idh2R140Q/WT + KMT2A-MLLT3 leukemic cells [derived from leukemic mice in (D)] were treated with increasing concentrations of DOT1L inhibitor (EPZ5676). Cells were counted every 3 days using trypan blue, and 10 000 cells were replated and treated with fresh compound. Error bars represent SD. (F) Growth of WT + KMT2A-MLLT3 and Vav1-Cre;Idh2R140Q/WT + KMT2A-MLLT3 leukemic cells [from (D)] treated with increasing concentrations of mutant IDH2 inhibitor (AGI-6780). Cells were counted every 3 days using trypan blue, and 10 000 cells were replated and treated with fresh compound. Error bars represent the SD of technical duplicates. (G-H) Colony formation and serial replating of GMPs from Vav1-Cre;Idh2R140Q/WT or WT control mice transduced with KMT2A-MLLT3. Number of colonies (G) and number of cells (H) at each plating, per 500 cells plated in methyl cellulose of KMT2A-MLLT3–transformed GMPs from 2 individual transplants. Cells were replated every 5 days. Error bars represent SD. (I) Survival of sublethally irradiated mice transplanted with 10 000 Vav1-Cre;Idh2R140Q/WT GMPs transduced with KMT2A-MLLT3 (n = 5) or with 10 000 WT GMPs transduced with KMT2A-MLLT3 (n = 5). *P < .01, **P < .001; Student t test. DMSO, dimethyl sulfoxide; ns, not significant.

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