- 作者: Fe-Lin Lin Wu; Yu-Fen Liang; Yuan-Chen Chang; Hao-Hsin Yo; Ming-Feng Wei; Li-Jiuan Shen
- 作者服務機構: School of Pharmacy, College of Medicine, National Taiwan University, Taiwan, R.O.C.
- 中文摘要: --
- 英文摘要:
Background: Sensitivity of cancer cells to recombinant arginine deiminase (rADI) depends on expression of
argininosuccinate synthetase (AS), a rate-limiting enzyme in synthesis of arginine from
citrulline. To understand the efficiency of RNA interfering of AS in sensitizing the resistant
cancer cells to rADI, the down regulation of AS transiently and permanently were performed in
vitro, respectively.
Methods: We studied the use of down-regulation of this enzyme by RNA interference in three human
cancer cell lines (A375, HeLa, and MCF-7) as a way to restore sensitivity to rADI in resistant
cells. The expression of AS at levels of mRNA and protein was determined to understand the
effect of RNA interference. Cell viability, cell cycle, and possible mechanism of the restore
sensitivity of AS RNA interference in rADI treated cancer cells were evaluated.
Results: AS DNA was present in all cancer cell lines studied, however, the expression of this enzyme at
the mRNA and protein level was different. In two rADI-resistant cell lines, one with endogenous
AS expression (MCF-7 cells) and one with induced AS expression (HeLa cells), AS small interference RNA (siRNA) inhibited 37-46% of the expression of AS in MCF-7 cells. ASsiRNA
did not affect cell viability in MCF-7 which may be due to the certain amount of residual AS
protein. In contrast, ASsiRNA down-regulated almost all AS expression in HeLa cells and caused
cell death after rADI treatment. Permanently down-regulated AS expression by short hairpin
RNA (shRNA) made MCF-7 cells become sensitive to rADI via the inhibition of 4E-BP1-
regulated mTOR signaling pathway.
Conclusions: Our results demonstrate that rADI-resistance can be altered via AS RNA interference. Although
transient enzyme down-regulation (siRNA) did not affect cell viability in MCF-7 cells,
permanent down-regulation (shRNA) overcame the problem of rADI-resistance due to the more
efficiency in AS silencing. - 中文關鍵字: --
- 英文關鍵字: --