Thesis (Ph.D.)--Chulalongkorn University, 2013

Glycosmis parva Craib (Rutaceae) was reported to have cytotoxicity and anti-inflammatory activities with the reduction of COX-2 expression, in vitro. This study aimed to investigate the effect of ethyl acetate extract from leaves of G. parva (GPE) on human colorectal cancer expressing COX-2, HT-29, and not express COX-2, Colo-205 and its underlying mechanisms of action. Cytotoxicity of GPE against HT-29 and Colo-205 were exhibited in both dose- and time- dependent manner. The IC50 values of GPE against HT-29 were 69.49±2.04, 55.89±1.86 and 48.94±2.99 µg/ml at 24, 48 and 72 h, respectively. In Colo-205, the IC50 were 59.92±4.34, 28.85±1.44 and 25.42±1.65 µg/ml at 24, 48 and 72 h, respectively. GPE significantly induced apoptosis in both cell lines as evidenced by AnnexinV/FITC and PI staining. The effect of GPE on cytotoxicity and induction of apoptosis were higher in Colo-205 than HT-29. GPE at 25-100 µg/ml significantly inhibited cell proliferation in both cells. The effect was greater in HT-29 cells. Cell cycle analysis demonstrated that GPE caused a decrease in the cells in S phase which was associated with G0/G1 and G2/M accumulation. The changes in cell cycle pattern following GPE treatment were similar in both cell lines. Mechanistic studies suggested that the effects of GPE may be mediated through both COX-2 dependent and COX-2 independent pathway. The COX-2 dependent involved with reduction of COX-2 expression which affected downstream COX-2 pathway. The COX-2 independent pathway included the changes in the expression of cell cycle control genes and the alteration in the balance of Bcl-2 family gene expression. In HT-29, GPE down-regulated cyclin A and cyclin E expression and increased the expression of pro-apoptotic Bak while decreased the anti-apoptotic Bcl-2. It also significantly decreased COX-2 expression. In Colo-205, GPE decreased cyclin A and up-regulated p21 expression. It also decreased the expression of anti-apoptotic Bcl-2 and Bcl-XL. Taken together, GPE exhibited cytotoxic activity, induction of apoptosis, inhibition of cell proliferation and arrest cell cycle in both cells. The underlying mechanisms involved both COX-2 dependent and COX-2 independent pathway. These findings provide the fundamental knowledge of the anti-cancer effect of GPE which could be a potential compounds for the treatment of colorectal cancer.