Reduced expression of small GTPases and hypermethylation of the folate binding protein gene in cisplatin-resistant cells

Br J Cancer. 2004 Jul 19;91(2):270-6. doi: 10.1038/sj.bjc.6601956.


Reduced accumulation of cisplatin is the most consistent feature seen in cisplatin-resistant (CP-r) cells that are cross-resistant to other cytotoxic compounds, such as methotrexate. In this report, defective uptake of a broad range of compounds, including [(14)C]-carboplatin, [(3)H]MTX, [(3)H]folic acid (FA), [(125)I]epidermal growth factor, (59)Fe, [(3)H]glucose, and [(3)H]proline, as well as (73)As(5+) and (73)As(3+), was detected in CP-r human hepatoma and epidermal carcinoma cells that we have previously shown are defective in fluid-phase endocytosis. Downregulation of several small GTPases, such as rab5, rac1, and rhoA, which regulate endocytosis, was found in CP-r cells. However, expression of an early endosomal protein and clathrin heavy chain was not changed, suggesting that the defective endocytic pathway is clathrin independent. Reduced expression of the cell surface protein, folate-binding protein (FBP), which is a carrier for the uptake of MTX, was also observed in the CP-r cells by confocal immunofluorescence microscopy and Real-Time PCR. Reactivation of the silenced FBP gene in the CP-r cells by a DNA demethylation agent, 2-deoxy-5-aza-cytidine (DAC) demonstrates that hypermethylation occurred in the CP-r cells. The uptake of [(14)C]carboplatin, [(3)H]FA, and [(3)H]MTX increased in an early stage CP-r cell line (KB-CP1) after treatment with DAC. Both a defective endocytic pathway and DNA hypermethylation resulting in the downregulation of small regulatory GTPases and cell surface receptors contribute to the reduced accumulation of a broad range of compounds in CP-r cells.

MeSH terms

  • Antineoplastic Agents / pharmacology*
  • Azacitidine / analogs & derivatives*
  • Azacitidine / pharmacology
  • Carboplatin / pharmacology
  • Carrier Proteins / genetics*
  • Carrier Proteins / metabolism
  • Cell Survival / drug effects
  • Cisplatin / pharmacology*
  • DNA Methylation*
  • Decitabine
  • Down-Regulation
  • Drug Resistance, Neoplasm / genetics*
  • Endocytosis / physiology
  • Folate Receptors, GPI-Anchored
  • Folic Acid / pharmacology
  • Gene Expression Regulation, Neoplastic / physiology*
  • Humans
  • Immunoblotting
  • Methotrexate / pharmacology
  • Monomeric GTP-Binding Proteins / genetics*
  • Monomeric GTP-Binding Proteins / metabolism
  • RNA, Messenger / metabolism
  • Receptors, Cell Surface / genetics*
  • Receptors, Cell Surface / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Tumor Cells, Cultured / drug effects


  • Antineoplastic Agents
  • Carrier Proteins
  • Folate Receptors, GPI-Anchored
  • RNA, Messenger
  • Receptors, Cell Surface
  • Decitabine
  • Folic Acid
  • Carboplatin
  • Monomeric GTP-Binding Proteins
  • Azacitidine
  • Cisplatin
  • Methotrexate