Figure 4.

TWIST1 regulates ABCB1. (A) Membranous ABCB1 IHC staining of DKFZ TMA and Nottingham TMA was scored and correlated with metastatic status (M0/M+) in a total of 259 patients (Pearson’s chi-squared exact test P = .04). (B) Schematic of the E-box binding site in the Exon 3 promoter region of ABCB1. (C) ChIP qPCR analysis of the promoter regions in GAPDH, SNAI2, and ABCB1, showing enrichment with TWIST1 antibody for GAPDH and ABCB1 compared with IgG antibody control. Data are expressed as a percentage of the input (paired Student’s t-test *P ≤ .05; n ≥ 3). (D and E) MED6 and MED6-TWIST1 (2.6 × 10⁴ cells/well) cells were treated with vardenafil (10 µM) or vehicle (H₂0) at days 0–4 in the 3D-BME model. (D) Time-lapse images were taken at ×10 magnification (scale bars represent 100 µm). (E) The mean aggregate area was quantified from time-lapse images taken between days 0 and 4 (unpaired t-test with Sidak–Bonferroni correction *P ≤ .05; n ≥ 2).