A functional SMAD1 locus is required for the therapeutic efficacy of DAC treatment. (A-B) SMAD1^WT and SMAD1^KO SU-DHL-4 cells were injected subcutaneously into the flanks of NSG mice. From day 12 after transplantation, mice were treated systemically with 0.25 mg/kg DAC for 2 cycles, 5 days per week. The tumor volume was measured continuously (A), and tumor weights were recorded at the end point (B). Data in panels A-B are pooled from 2 independent studies, and P values were calculated using the Mann-Whitney U test (for every time point relative to the red curve in panel A). (C-E) SMAD1^WT and SMAD1^KO SU-DHL-4 cells were subjected to lentiviral gene transfer of luciferase and injected IV into MISTRG mice. Mice were treated with 0 or 0.25 mg/kg DAC starting from day 15 after injection. IVIS images of all included mice (C) and the radiance over 5 weeks (D) are shown. (D) P values were calculated using the Mann-Whitney U test (for every time point relative to the red curve). (E) Survival curve showing the percentage of injected mice without symptoms of disease (paralysis, weight loss). The log-rank (Mantel-Cox) test was used for statistical comparisons. Data in panels C-E are pooled from 2 studies; the scale marked with a red asterisk (*) (far right panel in panel C) applies to the mice denoted by the asterisk (*) in panel C. Differences in panels D-E show trends only; 1 mouse (KO, on DAC) died from its disease before the study end point, and 1 mouse (WT, on DAC) died from nondisease-related causes. *P < .05, **P < .01, ***P < .001.