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. Author manuscript; available in PMC: 2021 Dec 15.
Published in final edited form as: Cancer Res. 2021 Apr 5;81(12):3347–3357. doi: 10.1158/0008-5472.CAN-20-3611

Figure 2. NADI-351 inhibits recruitment of Notch1 to the NTC and to the HES1 promoter.

Figure 2.

(A) CSL-dependent affinity pulldown indicates NADI-351 (10 μM) selectively inhibits Notch1ICD and MAML1 NTC assembly in MDA-MB-231 in a time-dependent manner (red boxes). (B) NADI-351 (10 μM) inhibits Notch target genes (HES1, HES5 and HEY1) expression in MDA-MB-231 cells through RT-qPCR at 6 h of treatment (n = 3). NADI-351 (10 μM) selectively inhibits recruitment of Notch1ICD (C,E) and MAML1 (D) to the NTC and to the HES1 promoter in MDA-MB-231 cells by ChIP at 6 h of treatment (n = 6). (F) CSL-dependent affinity pulldown indicates that high concentrations of NADI-351 selectively inhibits the recruitment of Notch1ICD and MAML1 to DNA-bound CSL at 48h in MDA-MB-231. (G) Effect of NADI-351 against individual inducible NotchICD-driven luciferase reporters (Notch1-3). Error bars are representative of at least three independent experiments, and values indicate mean ± SD [(B), (C), (D) and (E)]. p values ≤ 0.05 are considered statistically significant and indicated by an asterisk. *P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001; ns (no significant difference).