Role of thymidine phosphorylase and Rad51 in HSP90 inhibition induced cytotoxic effect in non-small-cell lung cancer cells

• Main Authors: Sheng-Chieh Tseng, 曾聖捷
• Other Authors: Yun-Wei Lin
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/13092432424382210910

碩士 === 國立嘉義大學 === 生化科技學系研究所 === 101 === Elevated thymidine phosphorylase (TP) levels, a key enzyme in the pyrimidine nucleoside salvage pathway, are associated with an aggressive disease phenotype and poor prognoses. Furthermore, Repair protein Rad51 is involved in protecting non-small lung cancer (NSCLC) cell lines against chemotherapeutic agent-induced cytotoxicity. Moreover, Heat shock protein 90 (HSP90) is an exciting new target in cancer therapy. This study investigated the role of Rad51 and TP expression in HSP90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG)-induced cytotoxicity in NSCLC cell line, A549. The 17-AAG treatment decreased cellular Rad51 and TP protein and mRNA levels and phosphorylated MKK1/2-ERK1/2 and AKT protein levels. Specific inhibition of Rad51 and TP expression by siRNA further enhanced 17-AAG-induced cytotoxicity and cell death and growth inhibition that had been induced by 17-AAG. Furthermore, enhancement of ERK1/2 or AKT activation by transfecting the cancer cells with constitutively active MKK1/2 or AKT expression vectors significantly restored the 17-AAG-reduced TP protein levels as well as cell viability. The 17-AAG treatment disrupted the interaction between HSP90 and TP or HSP90 and Rad51 triggered two proteins degradation through the ubiquitin-26S proteasome pathway. The U0126 (MKK1/2 inhibitors) or LY294002 (PI3K inhibitor) treatment decreased the TP expression and cell viability induced by 17-AAG. our results suggest that the down-modulation of TP and Rad51 protein induced by 17-AAG represents a key factor in enhancing the cytotoxic effects in NSCLC cells. Key words: HSP90; Rad51; ERK1/2; thymidine phosphorylase; cytotoxicity; non-small cell lung cancer