Enhancement of tannic acid, esculetin, or berberine on the anticancer effects and mechanisms of arsenic trioxide

• Main Authors: Tseng-Hsi, 林增熙
• Other Authors: 周芬碧
• Format: Others
• Language: zh-TW
• Published: 2009
• Online Access: http://ndltd.ncl.edu.tw/handle/42001546493316978819

博士 === 中山醫學大學 === 生化暨生物科技研究所 === 97 === Cancer metastasis and drug resistance are major problems in modern cancer treatment. The dual effects of phytochemicals to enhance chemotherapy and protect normal cells are applicable. Arsenic trioxide (As2O3) is widely used in treating human acute promyelocytic leukemia (APL). However, solid tumors and other leukemia cells such as U937 promonocytic leukemia cells are insensitive to As2O3. Tannic acid (TA), a glucoside of gallic acid polymer, or esculetin, a coumarin derivative, has previously induced cell cycle arrest and apoptosis of cancer cells by displaying anticancer potential. We find that tannic acid treatment decreased the viability of HL-60 cells in a dose- and time-dependent manner. Flow cytometry analysis revealed that tannic acid significantly induced apoptosis of HL-60 cells as confirmed by a dose-dependent increase of sub-G1 fraction. The further analysis of DAPI stain and DNA integrity also evidenced that tannic acid caused chromosome condensation and DNA fragmentation. The molecular analysis showed that tannic acid-induced apoptosis of HL-60 cells was associated with disruption of mitochondrial membrane potential, release of cytochrome c, activation of caspase-9 and caspase-3, and cleavage of poly (ADP-ribose) polymerase (PARP). These effects were associated with the increase of intracellular superoxide in mitochondrial signal pathway which attributed to the down-regulation of superoxide dismutase (SOD). Esculetin inhibited proliferation and mitogen activated protein kinases (MAPKs) activation in human leukemia U937 cells. The study showed that esculetin, PD98059 (MEK/ERK inhibitor), and SP600125 (JNK inhibitor) similarly enhanced the As2O3-induced GSH depletion. In addition, esculetin enhanced the effect of As2O3 on caspase activation in U937 cells. The combined esculetin and As2O3 treatment increased Bid cleavage, Bax conformation change and cytochrome C release. The study also indicated that esculetin enhanced the As2O3-induced lysosomal leakage and apoptosis. Furthermore, pretreatment with N-acetylcysteine (NAC) reduced these enhanced effects. Based on these studies, esculetin enhances the As2O3-provoked apoptosis by modulating the MEK/ERK and JNK pathways and reducing intracellular GSH levels. GSH depletion led to higher oxidative stress which activated lysosomal–mitochondrial pathway of apoptosis. The wound scratch and Boyden chamber assays were applied to determine the effect of As2O3 and berberine on the migration capacity and invasiveness of glioma cancer cells. Zymography and Western blot analyses provided information on the effect of As2O3 and berberine on the intracellular translocation and activation of protein kinase C (PKC), and some PKC related downstream factors. The As2O3-mediated reduction in motility and invasion of glioma cells was enhanced upon co-treatment with berberine. Furthermore, it has been reported that PKC isoforms influence the morphology of the actin cytoskeleton, as well as the activation of metalloproteases MT1-MMP and MMP-2, reported to be involved in cancer cell migration. Treatment of glioma cells with As2O3 and berberine significantly decreased the activation of PKC α and ε and led to actin cytoskeleton rearrangements. The levels of two downstream transcription factors, myc and jun, and MT1-MMP and MMP-2 were also significantly reduced. Upon co-treatment of glioma cells with As2O3 and berberine, cancer cell metastasis can be significantly inhibited, most likely by blocking the PKC-mediated signaling pathway involved in cancer cell migration. This study is potentially interesting for the development of novel chemotherapeutic approaches in the treatment of malignant gliomas and cancer development in general.