Development of trastuzumab-resistant human gastric carcinoma cell lines and mechanisms of drug resistance

Sci Rep. 2015 Jun 25;5:11634. doi: 10.1038/srep11634.

Abstract

Trastuzumab has been successfully employed for the treatment of Her-2-positive gastric cancer. However, there are problems with both primary and secondary resistance to trastuzumab. In this study, we employed the human gastric carcinoma cell line NCI-N87 with high Her-2 expression to create trastuzumab-resistant NCI-N87/TR cells by stepwise exposure to increasing doses of trastuzumab. Western blotting and Real-time PCR were conducted to detect protein and gene levels. Compared with NCI-N87 cells, the expression of P-IGF-1R and P-AKT proteins was significantly increased in NCI-N87/TR cells (both P = 0.000), while PTEN gene and protein expression showed a significant decrease (both P = 0.000). In addition, mutations of the PTEN gene were detected at exons 5, 7, and 8. The sensitivity of NCI-N87/TR cells to trastuzumab was increased by transfection with the PTEN gene, or by incubation with a PI3K inhibitor (LY294002) or an IGF-IR inhibitor (AG1024), as well as siRNA targeting PI3K p110 or IGF-1R. Taken together, our findings showed that activation of the PI3K-AKT signaling pathway was one of the major mechanisms leading to resistance of NCI-N87/TR gastric cancer cells to trastuzumab, which was probably associated with PTEN gene down-regulation and mutation, as well as with over-activity of the IGF-1R signaling pathway.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antineoplastic Agents / pharmacology
  • Blotting, Western
  • Cell Line, Tumor
  • Cell Survival / drug effects
  • Cell Survival / genetics
  • Chromones / pharmacology
  • Class Ia Phosphatidylinositol 3-Kinase / genetics
  • Class Ia Phosphatidylinositol 3-Kinase / metabolism
  • Dose-Response Relationship, Drug
  • Drug Resistance, Neoplasm / drug effects*
  • Drug Resistance, Neoplasm / genetics*
  • Enzyme Inhibitors / pharmacology
  • Gene Expression Regulation, Neoplastic / drug effects*
  • Humans
  • Morpholines / pharmacology
  • PTEN Phosphohydrolase / genetics
  • PTEN Phosphohydrolase / metabolism
  • Phosphoinositide-3 Kinase Inhibitors
  • Proto-Oncogene Proteins c-akt / genetics
  • Proto-Oncogene Proteins c-akt / metabolism
  • RNA Interference
  • Receptor, IGF Type 1 / antagonists & inhibitors
  • Receptor, IGF Type 1 / genetics
  • Receptor, IGF Type 1 / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Signal Transduction / drug effects
  • Signal Transduction / genetics
  • Stomach Neoplasms / genetics
  • Stomach Neoplasms / metabolism
  • Stomach Neoplasms / pathology
  • Trastuzumab / pharmacology*
  • Tyrphostins / pharmacology

Substances

  • Antineoplastic Agents
  • Chromones
  • Enzyme Inhibitors
  • Morpholines
  • Phosphoinositide-3 Kinase Inhibitors
  • Tyrphostins
  • tyrphostin AG 1024
  • 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
  • Class Ia Phosphatidylinositol 3-Kinase
  • Receptor, IGF Type 1
  • Proto-Oncogene Proteins c-akt
  • PTEN Phosphohydrolase
  • Trastuzumab