title: Schedule-dependent cytotoxic synergism of pemetrexed and erlotinib in human non-small cell lung cancer cells.
abstract: OBJECTIVE
  This study was undertaken to select the optimal combination schedule of erlotinib and pemetrexed for the treatment of relapsed non-small cell lung cancer (NSCLC) using a panel of human NSCLC lines.
  
  
  METHODS
  Human NSCLC cell lines, with variable expression of the known molecular determinants of erlotinib sensitivity, were exposed to pemetrexed and erlotinib using different schedules. Antitumor effect was measured by growth inhibition by cell count and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, cell cycle distribution and apoptosis by flow cytometry, and expression of cell cycle mediators by immunoblots. The cytotoxic interaction between pemetrexed and erlotinib (i.e., synergistic, additive, or antagonistic) was determined by median effect analysis.
  
  
  RESULTS
  When cells were exposed to concurrent pemetrexed and erlotinib or sequential pemetrexed followed by erlotinib, cytotoxic synergism was observed in both erlotinib-sensitive and erlotinib-resistant human NSCLC cell lines. This was independent of the mutation status of epidermal growth factor receptor or K-Ras genes. Synergism was associated with a combination of cell cycle effects from both agents. In contrast, exposure of cells to erlotinib followed by pemetrexed was mostly antagonistic in erlotinib-sensitive cells and additive at best in erlotinib-resistant cells. Antagonism was associated with erlotinib-induced G(1)-phase blockade of erlotinib-sensitive cells, which protects cells from pemetrexed cytotoxicity. Pemetrexed induced an epidermal growth factor receptor-mediated activation of the phosphatidylinositol 3-kinase/AKT pathway, which was inhibited by erlotinib and a specific phosphatidylinositol 3-kinase inhibitor, LY294002.
  
  
  CONCLUSIONS
  The combination of pemetrexed and erlotinib is synergistic in NSCLC in vitro if exposure to erlotinib before pemetrexed is avoided, particularly in tumors that are sensitive to erlotinib. Based on these findings, a randomized phase II study comparing the progression-free survival between an intermittent combination of erlotinib and pemetrexed (experimental arm) and pemetrexed alone (control arm) in patients with relapsing NSCLC has been initiated.
authors.affiliation: Department of Oncology, Montefiore Medical Center, Albert Einstein College of Medicine, Bronx, New York 10461, USA. tli@montefiore.org
authors.forename: Tianhong
authors.initials: T
authors.lastname: Li
authors.forename: Yi-He
authors.initials: YH
authors.lastname: Ling
authors.forename: I David
authors.initials: ID
authors.lastname: Goldman
authors.forename: Roman
authors.initials: R
authors.lastname: Perez-Soler
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000008:administration & dosage*; D000970:Antineoplastic Agents / Q000627:therapeutic use*; D017209:Apoptosis; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D004357:Drug Synergism; D066246:ErbB Receptors / Q000235:genetics; D000069347:Erlotinib Hydrochloride; D011905:Genes, ras; D005971:Glutamates / Q000008:administration & dosage*; D006147:Guanine / Q000008:administration & dosage / Q000031:analogs & derivatives*; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy*; D008954:Models, Biological; D000068437:Pemetrexed; D047428:Protein Kinase Inhibitors / Q000008:administration & dosage*; D011799:Quinazolines / Q000008:administration & dosage*
chemical_list: D000964:Antimetabolites, Antineoplastic; D000970:Antineoplastic Agents; D005971:Glutamates; D047428:Protein Kinase Inhibitors; D011799:Quinazolines; D000068437:Pemetrexed; D006147:Guanine; D000069347:Erlotinib Hydrochloride; D066246:ErbB Receptors