Skip to main content
. 2022 Dec 1;15(1):e15631. doi: 10.15252/emmm.202115631

Figure 3. The therapeutic effect of IMPDH inhibition on AML is p53‐p21 independent.

Figure 3

  1. Immunoblotting of the cord blood cells and those expressing RUNX1‐ETO or MLL‐AF9. The cells were incubated with/without 1 μM MPA (M) and 100 μM Guanosine (G) for 24 h.
  2. CB‐MLL‐AF9#2 cells were transduced with a Non‐targeting (NT) control and an shRNA targeting p21 (sh‐p21). Cells were incubated with/without 1 μM MPA (M) and 100 μM guanosine (G) for 24 h. Total cell lysates were analyzed by western blotting using the antibodies for p21 and Tubulin.
  3. CB‐MLL‐AF9#2 cells transduced with NT or sh‐p21 shRNAs were incubated with MPA at the indicated concentration for 72 h. Cell viability assays were performed using WST‐1 in three technical replicates.
  4. CB‐MLL‐AF9#2 cells were transduced by vector control or p53DD (dominant‐negative) and were incubated with/without 1 μM MPA (M) and 100 μM guanosine (G) for 24 h. Total cell lysates were analyzed by western blotting using the antibodies for p53 and Tubulin. Note the elevated levels of endogenous p53 protein (upper arrow) in p53DD (lower arrow)‐expressing cells.
  5. Cells were incubated with MPA at the indicated concentration with/without 100 μM guanosine for 72 h then measured with WST‐1 in three technical replicates.
  6. Mouse bone marrow c‐Kit+ cells derived from Trp53−/− mice were transduced with MLL‐AF9‐GFP and were transplanted into mice to generate p53‐deficient (p53−/−) leukemia cells. GFP+ MLL‐AF9 leukemia cells were collected from bone marrows of vehicle‐ or FF‐10501‐01‐treated wild‐type and p53−/− leukemic mice. Expression levels of p53 and Tubulin were assessed by western blotting 24 h after the treatment.
  7. Experimental scheme used in (H). The p53‐deficient MLL‐AF9‐bearing mice were treated with vehicle, FF‐10501‐01 or DS‐5272 every other day from day 1.
  8. Kaplan–Meier survival curves of p53−/− MLL‐AF9 leukemia mice treated with vehicle or FF‐10501‐01 or DS‐5272 (n = 6 per group). Statistical significance was evaluated by the log‐rank test (left). Frequency of GFP+ leukemia cells in peripheral blood at day 19 (right, n = 6 per group). A two‐tailed unpaired t‐test was used for the comparison.

Data information: All data are shown as mean ± SEM.

Source data are available online for this figure.