Abstract
Many anticancer agents are currently available for clinical therapy, but very few agents are effective against some types of cancer. Fortunately, we have obtained a potential anticancer agent THKM76 from a plant product. In this purpose, we studied the effects of THKM76 on the morphology, cytotoxicity, and cell cycle progression in a human colon adenocarcinoma cell line (RKO) and a human fibroblast cell line (HF). For RKO cells, characteristic morphological changes and drug-induced apoptosis, related to the administration of the cytotoxic agent, were interpreted as degenerative in nature, but drug-treated HF cells revealed less characteristic features of damage. Cytotoxicity was evaluated by clonogenic survival assay showed that RKO was more sensitive to THKM76 than HF. In addition, by MTT assay, we found that treating the cells with THKM76 resulted in a decrease of cell viability in a dose- and a time-dependent manner. However, there were differences in survival rate between using clonogenic survival assay (assayed after release 7 days) and MTT assay (assayed immediately after exposure). According to the results and some evidences indicated that the cell growth delayed after release from treatment, we demonstrated that there were two major cytotoxic effects for inducing cell killing. (i) Direct effect: Cells were killed first during THKM76 treatment. (ii) Persisted effect: After removal of THKM76, drug-treated viable cells lost reproductive ability further and even died. Finally, flow cytometry analysis of the DNA content revealed G1 block, slight G2/M arrest, and the presence of a ‘sub-G2’ region in RKO cells after THKM76 treatment with indicated concentrations. Besides, after release from treatment, there was a significant increase of RKO cells arrested in G2/M phase. The results of combining the arrest status with irradiation supported a role for THKM76 as a radiosensitizer.