title: Combination of EGFR/HER2 tyrosine kinase inhibition by BIBW 2992 and BIBW 2669 with irradiation in FaDu human squamous cell carcinoma.
abstract: OBJECTIVE
  To investigate the effect of the dual EGFR/HER2 (ErbB2) tyrosine kinase inhibitors BIBW 2992 and BIBW 2669 on proliferation and clonogenic cell survival of FaDu human squamous cell carcinoma in vitro, and on tumor growth after single-dose irradiation in nude mice.
  
  
  METHODS
  Cell proliferation, cell-cycle distribution and clonogenic cell survival after irradiation were assayed with and without BIBW 2992 or BIBW 2669 (3, 30, and 300 nM) in vitro. Tumor volume and tumor growth delay (GD(V2)) were determined in tumors growing in NMRI (nu/nu) nude mice, treated with (a) BIBW 2992 (20 mg kg(-1) body weight orally), BIBW 2669 (3-4 mg kg(-1) body weight orally) or carrier until a final tumor diameter of 15 mm, or, (b) 3 days before a 20-Gy single-dose irradiation or, (c) after a 20-Gy single-dose irradiation until reaching the final tumor diameter.
  
  
  RESULTS
  BIBW 2992 and BIBW 2669 significantly increased the doubling time of FaDu cells in vitro. A marked dose-dependent antiproliferative effect with blockade of the cells in G0/G1-phase of the cell cycle was found. Incubation with BIBW 2669 or BIBW 2992 for 3 days marginally increased radiosensitivity of FaDu cells in vitro. For BIBW 2992, this effect was statistically significant (p = 0.006). Daily oral application of BIBW 2669 or BIBW 2992 in mice bearing unirradiated FaDu tumors showed a marked antiproliferative effect with a significant prolongation of tumor growth delay (p < 0.0001). After drug application for 3 days, followed by 20-Gy single-dose irradiation, a slight effect of both drugs on tumor growth delay was seen. For BIBW 2669, this effect was statistically significant (p = 0.007). However, this effect disappeared when tumor volumes were normalized to the time point of irradiation suggesting that both drugs showed no or only a slight radiosensitizing effect in vivo. Daily application of BIBW 2669 or BIBW 2992 after a single-dose irradiation showed a clear inhibition of tumor growth with a significantly longer tumor growth delay after drug treatment compared to control tumors (p < 0.002). Enhancement ratios were smaller for irradiated than for unirradiated tumors, suggesting an additive effect for combinations with radiotherapy. In all treatment arms, the effects of BIBW 2669 were not significantly different from BIBW 2992.
  
  
  CONCLUSIONS
  BIBW 2669 and BIBW 2992 showed a clear antiproliferative effect in vitro, whereas radiosensitization was only marginal. The present data are the first to show an effect of combined irradiation and dual EGFR/ErbB2 inhibition on tumor growth delay in vivo. Further preclinical investigations using fractionated irradiation schedules and local tumor control as experimental endpoint are needed to evaluate a possible curative potential for the combination treatment.
authors.affiliation: Department of Radiation Oncology, Medical Faculty Carl Gustav Carus, University of Technology, Dresden, Germany.
authors.forename: Christina
authors.initials: C
authors.lastname: Schütze
authors.forename: Annegret
authors.initials: A
authors.lastname: Dörfler
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Eicheler
authors.forename: Daniel
authors.initials: D
authors.lastname: Zips
authors.forename: Sandra
authors.initials: S
authors.lastname: Hering
authors.forename: Flavio
authors.initials: F
authors.lastname: Solca
authors.forename: Michael
authors.initials: M
authors.lastname: Baumann
authors.forename: Mechthild
authors.initials: M
authors.lastname: Krause
mesh_terms: D000284:Administration, Oral; D000077716:Afatinib; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D002294:Carcinoma, Squamous Cell / Q000473:pathology / Q000532:radiotherapy*; D002455:Cell Division / Q000187:drug effects / Q000528:radiation effects*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000528:radiation effects*; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D066246:ErbB Receptors / Q000037:antagonists & inhibitors*; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D007012:Hypopharyngeal Neoplasms / Q000473:pathology / Q000532:radiotherapy*; D008297:Male; D051379:Mice; D008819:Mice, Nude; D009368:Neoplasm Transplantation; D011799:Quinazolines / Q000494:pharmacology*; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000037:antagonists & inhibitors*; D014183:Transplantation, Heterologous; D014410:Tumor Stem Cell Assay
chemical_list: D000077716:Afatinib; D000970:Antineoplastic Agents; D066246:ErbB Receptors; D011799:Quinazolines; D011838:Radiation-Sensitizing Agents; D018719:Erb-b2 Receptor Tyrosine Kinases; C522925:BIBW 2669