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Restitution of Tumor Suppressor miR-145 Using Magnetic Nanoparticles Inhibits Pancreatic Cancer
Saini Setua2, Stephen W. Behrman*1, Sheema Khan2, Murali M. Yallapu2, Meena Jaggi2, Subhash C. Chauhan2
1Surgery, Univ. of Tennessee, Memphis, TN; 2Pharmaceutical Sciences, University of Tennessee Health Science Center, Memphis, TN
Background: Pancreatic cancer (PC) is the fourth leading cause of cancer related death in the United States, with a 5-year survival rate of less than 5%. MicroRNAs have been identified as attractive targets for therapeutic intervention. The functional significance of lost microRNAs have been reported in several human malignancies, including PC. Therefore, restoring lost miRNA function can provide a potential therapeutic benefit. Prior work has identified microRNA-145 (miR-145) as a tumor suppressor miRNA in pancreatic cancer. The restoration of miR-145 downregulates a number of oncogenes including mucin MUC13 - a glycoprotein that is aberrantly expressed in PC, and efficiently inhibits tumor growth in mice. The main challenge for successful translation of microRNAs into clinical practice remains an effective in vivo delivery system. The focus of this study was to develop and assess the efficacy of a miRNA delivery method for PC treatment.
Methods: Magnetic nanoparticle (MNP) based nanoformulation of miR-145 (miR-145-MNPF) was developed for the intracellular delivery and sustained release of miR-145. The positively charged polyethyleneimine molecules were used to increase the loading efficiency of miR-145. MUC13 expressing PC cell lines (HPAF-II and Capan-1) were utilized. Following transient transfection of miR-145-MNPF, Western blotting and immunofluorescence techniques were used to investigate the effects of miR-145 restoration on a number of oncoproteins including MUC13. Additionally, functional studies of the effects of miR-145 restitution using miR-145-MNPF including cell proliferation, colony formation, cell migration, and cell invasion assays were analyzed.
Results: miR-145 expression was progressively suppressed over the course of development from PanIN I-III to late stage poorly differentiated PDAC. Treatment of cells with miR-145-MNPF led to efficient intracellular delivery and upregulation of miR-145 as observed through prussian blue staining and qRT-PCR respectively. miR-145 restitution resulted in significant downregulation of target oncogenes including MUC13, HER2, P-AKT and p53 as observed through Western blotting and immunofluorescence techniques. miR-145-MNPF inhibited cell proliferation, clonogenicity, migration, and invasion of PC cells. MNPF mediated restitution of miR-145 effectively sensitized PC cells to paclitaxel and TRAIL therapy.
Conclusions: 1) MNP based delivery systems can be efficiently used for microRNA replacement therapy in order to restore lost microRNAs in cancer. 2) miR-145-MNPF efficiently restores miR-145 in pancreatic cancer cells and inhibits growth and invasion of PC. 3) miR-145 restitution using miR-145-MNPF may offer a potential therapeutic strategy for pancreatic cancer treatment alone or in combination with other therapies.
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