(±)-Forbesione

Ham-1 cholangiocarcinoma cells

0.5 µM; 1 µM; 2 µM; 4 µM; 8 µM

24 h

Induced concentration-dependent S-phase cell cycle arrest (P<0.01).
Reduced the proportion of cells in the G₀/G₁ (P<0.05) and G₂/M (P<0.05) phases.
Increased the percentage of cells in the sub-G₁ fraction (apoptotic cells) significantly in a concentration-dependent manner (P<0.01).

Ham-1 cholangiocarcinoma cells

2 µM; 4 µM; 8 µM; 16 µM

24 h

Induced bright green, condensed, and fragmented chromatin, which are hallmarks of apoptosis, in treated cells, while untreated cells showed uniformly green-stained, normal nuclei.

Ham-1 cholangiocarcinoma cells

2 µM; 4 µM; 8 µM; 16 µM; 32 μM

6, 24, 48 h

Increased the percentage of apoptotic cells in a concentration- and time-dependent manner.
Resulted in 50.75% (P<0.01), 55.57% (P<0.001), and 64.43% (P<0.001) apoptotic cells at 6, 24, and 48 h respectively when treated with 32 µM.

Ham-1 cholangiocarcinoma cells

2 µM; 4 µM; 8 µM

24 h

Significantly decreased protein expression of cyclin E (P<0.01), cyclin A (P<0.05), and cyclin-dependent kinase 2 (Cdk2) (P<0.01).
Significantly increased protein expression of p21 (P<0.05) and p27 (P<0.05) compared to control cells.\nIncreased protein expression of Fas (P<0.05), Fas-associated death domain (FADD) (P<0.05), and activated caspase-3 (P<0.05) in the death receptor pathway.
Decreased protein expression of procaspase-8 (P<0.05) and procaspase-3 (P<0.01) in the death receptor pathway.
Increased protein expression of B-cell lymphoma-2-like protein 4 (Bax) (P<0.05), activated caspase-9 (P<0.05), and activated caspase-3 (P<0.05) in the mitochondrial pathway.
Decreased protein expression of B-cell lymphoma-2 (Bcl-2) (P<0.05), procaspase-9 (P<0.05), and procaspase-3 (P<0.01) in the mitochondrial pathway.
Increased protein expression of activated caspase-12 (P<0.05), activated caspase-9 (P<0.05), and activated caspase-3 (P<0.05) in the endoplasmic reticulum pathway.
Decreased protein expression of procaspase-12 (P<0.05), procaspase-9 (P<0.05), and procaspase-3 (P<0.01) in the endoplasmic reticulum pathway.\nSignificantly decreased protein expression of NF-κB/p65 (P<0.01) compared to control cells.
Significantly increased protein expression of inhibitor of κB-α (IκB-α) (P<0.05) compared to control cells.\nSignificantly decreased protein expression of cytokeratin 19 (CK19) (P<0.01) compared to control cells.
Significantly decreased protein expression of proliferating cell nuclear antigen (PCNA) (P<0.05) compared to control cells.

human cholangiocarcinoma KKU-100 cells

0.25 µM; 0.5 µM; 1 µM; 2 μM; 1 μM (combined with 0.025 μM Doxorubicin)

48 h

Increased apoptosis in KKU-100 cells in a dose-dependent manner, with the highest single-agent dose (2 μM) inducing ~32% apoptotic cells.
Induced ~75% apoptotic cells when combined with 0.025 μM Doxorubicin, which was significantly higher than either single agent alone.

human cholangiocarcinoma KKU-100 cells

0.25 μM; 0.5 μM; 1 μM (single agent); 0.25 μM + 0.006 μM Doxorubicin (C1); 0.5 μM + 0.012 μM Doxorubicin (C2); 1 μM + 0.025 μM Doxorubicin (C3)

48 h

Decreased Bcl-2 expression (0.27-fold at 1 μM) and increased Bax expression (2.06-fold at 1 μM) as a single agent, leading to a Bax/Bcl-2 ratio of 1.63; the C3 combination further decreased Bcl-2 to 0.10-fold and increased Bax to 3.24-fold, resulting in a Bax/Bcl-2 ratio of 12.00.
Decreased survivin (0.35-fold at 1 μM), procaspase-9 (0.70-fold at 1 μM), and procaspase-3 (0.76-fold at 1 μM) while increasing activated caspase-9 (42.00-fold at 1 μM) and activated caspase-3 (42.00-fold at 1 μM) as a single agent; the C3 combination enhanced these effects, reducing procaspase-9 and procaspase-3 to 0-fold and increasing activated caspase-9 and caspase-3 to 64.00-fold and 54.00-fold, respectively.
Increased IκB-α (1.15-fold at 1 μM) and decreased pIκB-α (0.14-fold at 1 μM) and NF-κB/p65 (0.12-fold at 1 μM) as a single agent; the C3 combination further increased IκB-α to 1.21-fold and decreased pIκB-α and NF-κB/p65 to 0.01-fold each.
Decreased MRP1 expression (0.71-fold at 1 μM) as a single agent, while the C3 combination reduced MRP1 to 0.34-fold, a significantly greater effect than single-agent treatment.