Fig 1.
Combined treatments of CBP-93872 with oxaliplatin, cisplatin, 5-FU, or gemcitabine effectively suppresses cell growth.
(A) HT29 cells were treated with the indicated concentrations of CBP-93872 for 72 hrs, followed by WST-1 assay. Data are presented as means ± SD (n = 3). (B) Panc-1 cells were treated with the indicated concentrations of CBP-93872 for 72 hrs, followed by WST-1 assay. Data are presented as means ± SD (n = 3). (C) HT29 cells were treated with CBP-93872 (50 μM) in combination with indicated concentrations of oxaliplatin, cisplatin or 5-FU for 72 hrs, followed by WST-1 assay. (D) Panc-1 cells were treated with CBP-93872 (200 μM) in combination with indicated concentrations of oxaliplatin, cisplatin or gemcitabine for 72 hrs, followed by WST-1 assay. The black bars show individual treatments; while the white bars show combined treatment with CBP-93872. Data are presented as means ± SD (n = 3). Statistical significance was calculated using Student’s t-test (*, p < 0.01) (C, D).
Fig 2.
CBP-93872 enhances oxaliplatin-, cisplatin- and gemcitabine-induced apoptosis in HT29 cells or Panc-1 cells.
(A, B) HT29 cells were treated with oxaliplatin (30 μM) (A) or cisplatin (30 μM) (B) in the presence or absence of CBP-93872 (50 μM). Cells were harvested at 72 hrs, fixed and subjected to FACS analysis (left panels). The percentages of cells in sub-G1 phase are shown in the panels on the right. Data are presented as means ± SD (n = 3). Statistical significance was calculated using Student’s t-test (*, p < 0.01). (C) Panc-1 cells were treated with gemcitabine (0.1 μM) in the presence or absence of CBP-93872 (200 μM). (D-F) HT29 cells or Panc-1 cells, were collected at the times indicated. Total cell extracts were subjected to immunoblotting, using the indicated antibodies.
Fig 3.
CBP-93872 inhibits maintenance of G2 and S-phase checkpoints.
(A, B) HT29 cells were treated with oxaliplatin (30 μM) (A), or cisplatin (30 μM) (B) in the presence or absence of CBP-93872 (50 μM). Nocodazole (500 nM) was added to inhibit the exit of cells from mitosis. Cells were harvested at 48 hrs, fixed, and stained with anti-H3-pS10 antibodies to determine the mitotic index. Mitotic indices (left) and typical examples (right) are shown. Data are presented as means ± SD (n = 3). Statistical significance was calculated using Student’s t-test (*, p < 0.01). ((C) Panc-1 cells were treated with gemcitabine (0.1 μM) in the presence or absence of CBP-93872 (200 μM) with nocodazole (500 nM).
Fig 4.
CBP-93872 reduces the levels of oxaliplatin-, cisplatin- and gemcitabine-induced phosphorylations of ATR and Chk1.
(A, B) HT29 cells were treated with oxaliplatin (30 μM) (A), or cisplatin (30 μM) (B) in the presence of absence of CBP-93872 (50 μM). Cells were harvested at the times indicated, and total cell extracts were subjected to immmunoblotting using indicated antibodies. (C) Panc-1 cells were treated with gemcitabine (0.1 μM), in the presence or absence of CBP-93872 (200 μM).
