title: OSU-03012 sensitizes breast cancers to lapatinib-induced cell killing: a role for Nck1 but not Nck2.
abstract: BACKGROUND
  Lapatinib is characterized as an ErbB1/ErbB2 dual inhibitor and has recently been approved for the treatment of metastatic breast cancer. In this study, we examined mechanisms associated with enhancing the activity of lapatinib via combination with other therapies.
  
  
  METHODS
  In the present studies, estrogen receptor (ER) positive and ER negative breast cancer cells were genetically manipulated to up- or downregulate eIF2-alpha, its phospho-mutant, Nck1, or Nck2, then treated with OSU-03012, lapatinib or the combination and assayed for cytotoxicity/cytostaticity using clonogenic assays.
  
  
  RESULTS
  Treatment of breast cancer cell lines with lapatinib and OSU-03012 (a small molecule derivative of the Cox-2 inhibitor celecoxib) induced synergistic cytotoxic/cytostatic effects. This combination therapy corresponded to an increase in the phosphorylation of eIF2-α at serine⁵¹ and a decrease in Nck1 expression. Ectopic expression of phospho-mutant eIF2-α (Ser⁵¹Ala) or downregulation of eIF2-α in addition to downregulation of the eIF2-α kinase PERK inhibited the synergistic and cytotoxic effects. Furthermore, ectopic expression of Nck1, but not Nck2 abolished the decrease in cell viability observed in combination-treated cells. Downregulation of Nck1 failed to "rescue" the ablation of the cytotoxic/cytostatic effects by the phospho-mutant of eIF2-α (Ser⁵¹Ala) demonstrating that Nck1 downregulation is upstream of eIF2-α phosphorylation in the anti-survival pathway activated by lapatinib and OSU-03012 treatment. Finally, co-immunoprecipitation assays indicated that eIF2-α dissociates from the Nck1/PP1 complex after OSU-03012 and lapatinib co-treatment.
  
  
  CONCLUSIONS
  These data indicate that OSU-03012 and lapatinib co-treatment is an effective combination therapy, which functions to enhance cell killing through the Nck1/eIF2 complex. Hence, this complex is a novel target for the treatment of metastatic breast cancer.
authors.affiliation: University of Richmond, 28 Westhampton Way, Richmond, VA 23173, USA.
authors.forename: N Winston
authors.initials: NW
authors.lastname: West
authors.forename: Aileen
authors.initials: A
authors.lastname: Garcia-Vargas
authors.forename: Charles E
authors.initials: CE
authors.lastname: Chalfant
authors.forename: Margaret A
authors.initials: MA
authors.lastname: Park
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000378:metabolism*; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D004357:Drug Synergism; D005260:Female; D006801:Humans; D047468:Immunoprecipitation; D000077341:Lapatinib; D015513:Oncogene Proteins / Q000378:metabolism*; D011720:Pyrazoles / Q000008:administration & dosage; D011799:Quinazolines / Q000008:administration & dosage; D015398:Signal Transduction / Q000187:drug effects* / Q000502:physiology; D013449:Sulfonamides / Q000008:administration & dosage; D014162:Transfection
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C485544:NCK2 protein, human; C078332:Nck protein; C500894:OSU 03012; D015513:Oncogene Proteins; D011720:Pyrazoles; D011799:Quinazolines; D013449:Sulfonamides; D000077341:Lapatinib