Growth inhibition and its mechanism of isochaihulactone and n-butylidenephthalide on human prostate cancer cell lines

• Main Authors: Sheng-Chun Chiu, 邱勝軍
• Other Authors: Cheng-Yoong Pang
• Format: Others
• Language: zh-TW
• Published: 2011
• Online Access: http://ndltd.ncl.edu.tw/handle/16516222907179072900

博士 === 慈濟大學 === 醫學科學研究所 === 99 === Abstract: Prostate cancer (PCa) is the most common malignancy in American men and the second leading cause of deaths from cancer. It is the fifth top malignanacy and seventh top cancer mortality in Taiwan men. Moreover, the Complementary Alternative Medicine (CAM) has been proven to be helpful in cancer therapy attracted us to investigate the anti-tumor potential of several herbal extracts. Nan-Chai-Hu (Bupleurum scorzonerifolium, BS) is an important Chinese herb in the treatment of influenza, fever, malaria, cancer, and the menstrual disorders in China, Japan, and many otherparts of Asia. Our laboratory have a novel lignan from the extract fraction of BS, isochaihulactone, that has antitumoral activity against lung cancer A549 cells in vitro and in vivo. Danggui (Angelica sinensis, AS), a traditional Chinese medicine for menopausal symptoms and is recommended as a tonic, hemopoetic, spasmolytic and analgesic drug in clinical practice. In our previous study, n-butylidenephthalide (BP), a compound derived from AS chloroform extract, showed a dramatic antitumoral effect against malignant brain tumors in vitro and in vivo. Isochaihulactone caused cell cycle arrest at G2/M phase in LNCaP cells, which was correlated with an increase of p53 and p21 levels and downregulation of the checkpoint proteins cdc25c, cyclin B1 and cdc2. Bcl-2 phosphorylation and caspase activation were also observed. Isochaihulactone induced phosphorylation of c-Jun-N-terminal kinase (JNK), and JNK inhibitor partially reduced isochaihulactone-induced cell death. Isochaihulactone also induced the expressions of EGR-1 and NAG-1. Expression of NAG-1 was reduced by JNK inhibitor, and knocking down of NAG-1 inhibited isochaihulactone-induced cell death. Two human prostate cancer cell lines, LNCaP and PC-3, were treated with BP, and evaluated for their viability and cell cycle profiles. BP caused cell cycle arrest at G0/G1 phase, which was correlated with an increase of p21 and p27 levels and downregulation of the checkpoint proteins cyclin D1 and cdk2. To determine the mechanism of BP-induced growth arrest and cell death in prostate cancer cell lines, we performed a microarray study to identify alterations in gene expression induced by treatment with the BP in the LNCaP cells. Several BP-inducible genes, including the GADD153/CHOP, an endoplasmic reticulum stress (ER stress)-regulated gene, were identified. Treatment with BP induced ER stress, as evidenced by increased expression of the downstream molecules GRP78/BiP, IRE1-? and GADD153/CHOP in both LNCaP and PC-3 cell lines. We then tested the contribution of ER stress related genes to protect against BP-induced cell death using RNA interference. Blockage of IRE1-? or GADD153/CHOP expression by siRNA significantly reduced BP-induced cell death in LNCaP cells. Western blot showed the involvement of JNK1/2 signaling pathway. Blockage of JNK1/2 signaling by JNK siRNA resulted in decreased the expression of IRE1 and GADD153/CHOP genes, implicating that BP-induced ER stress may be elicited via JNK1/2 signaling in prostate cancer cells. The anti-proliferative effect of BP was further demonstrated in LNCaP xenograft animal model. Taken together, our study suggests that isochaihulactone can significantly inhibit LNCaP cell growth. In the other aspect, our results suggested that the anti-proliferative effect of BP is facilitated by ER stress in prostate cancer cells.