Antisense inhibition of microRNA-21 or -221 arrests cell cycle, induces apoptosis, and sensitizes the effects of gemcitabine in pancreatic adenocarcinoma

Pancreas. 2009 Oct;38(7):e190-9. doi: 10.1097/MPA.0b013e3181ba82e1.

Abstract

Objectives: The contribution of overexpressed microRNA-21 and -221 (miR-21 and miR-221) to the malignant phenotype was determined by inhibiting these miRNAs using antisense oligonucleotides.

Methods: The effects of antisense to miR-21 and miR-221 on cell proliferation, cell cycle arrest, induction of apoptosis, combinatorial effects with gemcitabine, and effects on target protein levels were studied.

Results: Low nanomolar concentrations of both antisense oligonucleotides reduced proliferation of pancreatic cancer cell lines. Reduced proliferation was less pronounced in the normal ductal epithelial cell line human pancreatic Nestin-expressing cell or in pancreatic cancer cell lines exposed to an irrelevant control oligonucleotide. Inhibition of miR-21 and miR-221 increased the amount of apoptosis in HS766T cells by 3- to 6-fold compared with the control oligonucleotide. HS766T cells exposed to miR-21 antisense resulted in cell cycle arrest (G1 phase). Protein levels of tumor suppressor targets of the miRNAs were increased by antisense to miR-21 (PTEN and RECK) and miR-221 (p27). Antisense to miR-21 and miR-221 sensitized the effects of gemcitabine, and the antisense-gemcitabine combinations were synergistic at high fraction affected.

Conclusions: We demonstrate that antisense to miR-21 and miR-221 results in significant cell killing under various conditions and that antisense oligonucleotides targeted to miRNA represents a potential new therapy for pancreatic cancer.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenocarcinoma / genetics
  • Adenocarcinoma / metabolism
  • Adenocarcinoma / pathology
  • Antimetabolites, Antineoplastic / pharmacology
  • Apoptosis / drug effects*
  • Apoptosis / genetics
  • Blotting, Western
  • Cell Cycle / drug effects*
  • Cell Cycle / genetics
  • Cell Line
  • Cell Line, Tumor
  • Cyclin-Dependent Kinase Inhibitor p21 / metabolism
  • Cyclin-Dependent Kinase Inhibitor p27 / metabolism
  • Deoxycytidine / analogs & derivatives*
  • Deoxycytidine / pharmacology
  • G1 Phase / drug effects
  • G1 Phase / genetics
  • Humans
  • MicroRNAs / genetics*
  • Oligonucleotides, Antisense / genetics*
  • PTEN Phosphohydrolase / metabolism
  • Pancreatic Neoplasms / genetics
  • Pancreatic Neoplasms / metabolism
  • Pancreatic Neoplasms / pathology
  • Reverse Transcriptase Polymerase Chain Reaction
  • Transfection

Substances

  • Antimetabolites, Antineoplastic
  • Cyclin-Dependent Kinase Inhibitor p21
  • MIRN21 microRNA, human
  • MIRN221 microRNA, human
  • MicroRNAs
  • Oligonucleotides, Antisense
  • Deoxycytidine
  • Cyclin-Dependent Kinase Inhibitor p27
  • gemcitabine
  • PTEN Phosphohydrolase
  • PTEN protein, human