Mnt deletion enhances the survival of mice transplanted with primary MLL::AF9 AML cells. (A) Kaplan-Meier survival analysis of mice transplanted with primary (T0) MLL::AF9 AMLs of the indicated genotypes. Eight mice were transplanted IV with 5 ×105 BM cells from each primary (T0) AML mouse on day 0 and treated by oral gavage with TAM (200 mg/kg body weight) or vehicle on days 3, 4, and 5. TAM-treated mice transplanted with Mntfl/flCreERT2/MLL::AF9 AML cells showed significantly delayed morbidity (pink curve; median survival, 57 days) when compared with the TAM-treated control mice transplanted with CreERT2/MLL::AF9 AML cells (blue curve; median survival, 24 days; supplemental Table 3). The arrows indicate the lifespan of mice with AMLs 9843, 9842, 9841, 9844. n = number of recipient mice; number in brackets indicates the number of independently derived MLL::AF9 AMLs transplanted. Statistical significance was determined using a log-rank (Mantle-Cox) test with Bonferroni correction (∗∗∗∗P < .0001). (B-C) Mnt deletion was assessed by polymerase chain reaction (B) and western blot (C) analysis of the BM cells obtained from individual transplant recipients at autopsy. Results are shown for 2 representative primary AMLs, CreERT2/MLL::AF9 2244 (blue) and Mntfl/flCreERT2/MLL::AF9 2205 (pink); individual transplant recipient mice are identified. The DNA ladder (base pair) and marker protein sizes (kilodalton) are indicated. Of note, in panel B, no AML cells were detectable in the BM of cured mice (9841 and 9844), as shown by the absence of the 450 bp CreERT2-specific DNA fragment. Panel C showing the MNT and MYC protein levels with actin as the loading control. Both MYC and MNT are undetectable in the BM of cured’ mice (9841 and 9844), as is the case for normal BM cells (not shown). Asterisks indicate a nonspecific band; AH15 is a control Eμ-Myc lymphoma CL in which Mnt was deleted by CRISPR/Cas9. KO, knockout; TAM, tamoxifen; WT, wild type.
Figure 3.

Mnt deletion enhances the survival of mice transplanted with primary MLL::AF9 AML cells. (A) Kaplan-Meier survival analysis of mice transplanted with primary (T0) MLL::AF9 AMLs of the indicated genotypes. Eight mice were transplanted IV with 5 ×105 BM cells from each primary (T0) AML mouse on day 0 and treated by oral gavage with TAM (200 mg/kg body weight) or vehicle on days 3, 4, and 5. TAM-treated mice transplanted with Mntfl/flCreERT2/MLL::AF9 AML cells showed significantly delayed morbidity (pink curve; median survival, 57 days) when compared with the TAM-treated control mice transplanted with CreERT2/MLL::AF9 AML cells (blue curve; median survival, 24 days; supplemental Table 3). The arrows indicate the lifespan of mice with AMLs 9843, 9842, 9841, 9844. n = number of recipient mice; number in brackets indicates the number of independently derived MLL::AF9 AMLs transplanted. Statistical significance was determined using a log-rank (Mantle-Cox) test with Bonferroni correction (∗∗∗∗P < .0001). (B-C) Mnt deletion was assessed by polymerase chain reaction (B) and western blot (C) analysis of the BM cells obtained from individual transplant recipients at autopsy. Results are shown for 2 representative primary AMLs, CreERT2/MLL::AF9 2244 (blue) and Mntfl/flCreERT2/MLL::AF9 2205 (pink); individual transplant recipient mice are identified. The DNA ladder (base pair) and marker protein sizes (kilodalton) are indicated. Of note, in panel B, no AML cells were detectable in the BM of cured mice (9841 and 9844), as shown by the absence of the 450 bp CreERT2-specific DNA fragment. Panel C showing the MNT and MYC protein levels with actin as the loading control. Both MYC and MNT are undetectable in the BM of cured’ mice (9841 and 9844), as is the case for normal BM cells (not shown). Asterisks indicate a nonspecific band; AH15 is a control Eμ-Myc lymphoma CL in which Mnt was deleted by CRISPR/Cas9. KO, knockout; TAM, tamoxifen; WT, wild type.

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