HuR mediated post-transcriptional regulation as a new potential adjuvant therapeutic target in chemotherapy for pancreatic cancer

摘要

AIM: To investigate the expression of HuR in pancreatic ductal adenocarcinoma (PDA) and to assess the effects of HuR silencing on the expression of cyclooxygenase-2 (COX-2) and heme oxygenase-1 (HO-1) and the in vitro response to gemcitabine (GEM) treatment in pancreatic cell lines.METHODS: We compared the expression of HuR, COX-2, and HO-1 in PDA and normal pancreatic tissue using quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blot. In addition, the HuR, COX-2 and HO-1 were analyzed in four types of cancer cell lines (MiaPaca2, Su.86.86, Capan-1, and Capan-2) with and without GEM treatment. Immunocytofluorescence analysis was used to investigate HuR localization in cells. Cell viability and response to GEM after HuR silencing were determined with the 3-(4,5-dimethylthiazol-2-Yl)-2,5-diphenyltetrazolium bromide test and the crystal violet clonogenic assay, respectively. To measure apoptosis, activation of caspases 3/7 was evaluated using immunofluorescence.RESULTS: In PDA tissue obtained from patients not treated with GEM, HuR mRNA expression was 3.2 times lower (P < 0.05) and COX-2 and HO-1 mRNA expression was 2.3-fold and 7.2-fold higher (P < 0.05), respectively, than normal pancreatic tissue (from organ donor). qRT-PCR analysis showed that HuR, COX-2, and HO-1 mRNA were overexpressed in all cancer cell lines treated with the half maximal inhibitory concentration (IC50) dose of GEM compared with control cells (P < 0.05). Western blot analysis revealed that COX-2 and HO-1 levels were significantly decreased in cancer cells after HuR silencing. Furthermore, HuR silencing increased the response to GEM treatment and decreased cell viability by 11.6%-53.7% compared to control cell lines. Caspases 3 and 7 were activated after HuR silencing and GEM treatment in all pancreatic cancer cell lines. In comparison, treatment with GEM alone did not activate caspases 3 and 7 in the same cell lines.CONCLUSION: HuR mediated post-transcriptional upregulation of COX-2 and HO-1 expression after GEM treatment in pancreatic cancer cells. HuR silencing significantly increased the effectiveness of GEM treatment in vitro.