Antiproliferative effect of new chalcone derivatives in human colorectal cancer HCT116 cells
Proceedings for Annual Meeting of The Japanese Pharmacological Society
Backgroud: Antiproliferative activity of chalcones against various cancer cells was documented. Aim of this work was to investigate mechanism of antiproliferative effect of synthetic chalcones on human colorectal cancer cells. Methods: Antiproliferative activity of chalcones on HCT116 colorectal cancer cell line was tested using MTT and BrdU assay. Flow cytometric analysis was used to determine changes in cell cycle, activity of caspase-3 and -9, poly (ADP ribose) polymerase (PARP), release of
... (PARP), release of cytochrome c, status of Bcl-2 family proteins as well as mitochondrial membrane potential (MMP). Immunoflorescence allowed us to detect microtubule dysregulation. Apoptosis of HCT116 cells induced by 1C was assessed by annexin V/PI double staining and acridine orange/propidium iodide (AO/PI) staining. Expression of selected genes regulating apoptosis and key signaling pathways was evaluated using quantitative real-time PCR. Flow cytometry and Western blot analysis were used to evaluate activity of trancription factors and signal molecules. Results: The most active compound (1C) inhibited proliferation of HCT 116 cells with IC50= 4,13 microM and was selected for further analysis. Cell cycle analysis showed block in G2/M phase as well as increase in the pro-apoptotic population of cells with sub-G1 DNA content. Apoptosis was confirmed by annexin V/PI double staining and AO/PI staining. The apoptosis was associated with the loss of MMP, PARP cleavage, caspase-3 and -9 activation, release of cytochrome c, as well as changes in the levels of Bcl-2 and p53 proteins. Immunoflorescence revealed that 1C induced disruption of microtubule structure. Moreover, 1C increases phosphorylation of stress kinases p38 and ERK 1/2 what is associated with induction of apoptosis and cell cycle arrest. Conclusion: New synthetic derivative 1C significantly inhibits proliferation of HCT116 cells by induction of apoptosis and cell cycle arrest via targeting important signaling pathways. The process was also associated with disruption of microtubule structure.