Abstract 4819: MicroRNA-21 (miR-21) in pancreatic ductal adenocarcinoma (PDAC): Correlation with clinical outcome and pharmacological aspects underlying its role in the modulation of gemcitabine activity

Abstract

Abstract MiR-21 was reported to be overexpressed in PDAC and contributed to tumor invasion and resistance to gemcitabine. Further studies are needed to test whether miR-21 could be used to stratify patients for treatment and provide mechanistic insights for new therapeutic targets. The aim of this study was to evaluate whether miR-21 expression was associated with the overall survival (OS) of PDAC patients treated with gemcitabine, and investigate its role in PDAC invasive behaviour and chemoresistance. MiR-21 expression was evaluated in cells (including 7 PDAC cell lines, 7 primary PDAC cultures, fibroblasts and a normal pancreatic ductal cell line) and tissues (neoplastic specimens from 77 PDAC patients (32 in metastatic/inoperable and 45 in adjuvant setting) and normal ductal samples) isolated by laser microdissection. The role of miR-21 on the pharmacological effects of gemcitabine was studied in cells transfected with a specific miR-21 precursor (pre-mir). Modulation of apoptosis, Akt activation and VEGF secretion was analyzed by Annexin and ELISA assays, respectively. Quantitative PCR evaluated the correlation of miR-21 with invasion-related genes and VEGF. Inhibitors of PI3K and mTOR (LY294002 and rapamycin) were used to test whether modulation of these signalling pathways affected molecular mechanisms activated by miR-21 and gemcitabine. Patients with high miR-21 expression had a significantly shorter OS both in adjuvant and inoperable/metastatic setting, while a trend toward correlation was found between low expression and clinical benefit. Multivariate analysis in all patients with adequate follow-up (N=58), confirmed the negative prognostic significance of high miR-21 expression (HR:2.6, 95%CI:1.7-3.4; P:0.001). MiR-21 expression was correlated with the expression in their respective tissues in the primary cultures and with gemcitabine resistance in all the PDAC cells. Treatment with gemcitabine resulted in a significant increase of miR-21 expression, ranging from 2 to 19-fold, in 13 PDAC cell cultures. Pre-mir transfection significantly decreased apoptosis induction by gemcitabine and up-regulated metalloproteinase-2/−9 and VEGF mRNA expression in 2 selected PDAC cultures. In contrast, addition of LY294002 and rapamycin resulted in reduction of phospho-Akt and prevented pre-mir induced resistance to gemcitabine. In conclusion, miR-21 expression was correlated with OS in PDAC gemcitabine-treated patients. In addition, miR-21 expression is associated with reduced apoptosis, increased Akt phosphorylation and expression of genes involved in invasive behaviour resulting in gemcitabine chemoresistance, which was prevented by Akt inactivation. Our findings indicate that inactivation of PI3K/mTOR pathway may increase gemcitabine sensitivity in PDAC and warrants clinical studies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4819.