title: The aromatase inhibitor letrozole and inhibitors of insulin-like growth factor I receptor synergistically induce apoptosis in in vitro models of estrogen-dependent breast cancer.
abstract: BACKGROUND
  Endocrine-dependent, estrogen receptor positive breast cancer cells proliferate in response to estrogens, synthesized by the cytochrome p450 aromatase enzyme. Letrozole is a potent nonsteroidal aromatase inhibitor that is registered for the treatment of postmenopausal women with advanced metastatic breast cancers and in the neoadjuvant, early, and extended adjuvant indications. Because crosstalk exists between estrogen receptor and insulin-like growth factor I receptor (IGF-IR), the effect of combining a selective IGF-IR inhibitor (NVP-AEW541) with letrozole was assessed in two independent in vitro models of estrogen-dependent breast cancer.
  
  
  METHODS
  MCF7 and T47D cells stably expressing aromatase (MCF7/Aro and T47D/Aro) were used as in vitro models of aromatase-driven breast cancer. The role of the IGF-IR pathway in breast cancer cells stimulated only by 17beta-estradiol or androstenedione was assessed by proliferation assays. The combination of letrozole and NVP-AEW541 was assessed for synergy in inhibiting cell proliferation using Chou-Talalay derived equations. Finally, combination or single agent effects on proliferation and apoptosis were assessed using proliferation assays, flow cytometry, and immunoblotting.
  
  
  RESULTS
  Both MCF7 and T47D cells, as well as MCF7/Aro and T47D/Aro, exhibited sensitivity to inhibition of 17beta-estradiol dependent proliferation by NVP-AEW541. Letrozole combined with NVP-AEW541 synergistically inhibited androstenedione-dependent proliferation in aromatase-expressing cells with combination index values of 0.6 or less. Synergistic combination effects correlated with higher levels of apoptosis as compared with cells treated with the single agent alone. Treatment with either agent also appeared to inhibit IGF-IR signalling via phosphoinositide 3-kinase. Notably, IGF-IR inhibition had limited effect on estrogen-dependent proliferation in the cell lines, but was clearly required for survival, suggesting that the combination of letrozole and IGF-IR inhibition sensitizes cells to apoptosis.
  
  
  CONCLUSIONS
  Inhibition of the IGF-IR pathway and aromatase was synergistic in two independent estrogen-dependent in vitro models of breast cancer. Moreover, synergism of NVP-AEW541 and letrozole correlated with induction of apoptosis, but not cell cycle arrest, in the cell lines tested. Combination of IGF-IR inhibitors and letrozole may hold promise for the treatment of patients with estrogen-dependent breast cancers.
authors.affiliation: Novartis Institutes of BioMedical Research Basel, Oncology Research, Klybeckstrasse 141, CH-4057, Basel, Switzerland. joanna.lisztwan@novartis.com
authors.forename: Joanna
authors.initials: J
authors.lastname: Lisztwan
authors.forename: Astrid
authors.initials: A
authors.lastname: Pornon
authors.forename: Bin
authors.initials: B
authors.lastname: Chen
authors.forename: Shiuan
authors.initials: S
authors.lastname: Chen
authors.forename: Dean B
authors.initials: DB
authors.lastname: Evans
mesh_terms: D017209:Apoptosis; D047072:Aromatase Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D004967:Estrogens / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D000077289:Letrozole; D009362:Neoplasm Metastasis; D009570:Nitriles / Q000494:pharmacology*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D017526:Receptor, IGF Type 1 / Q000378:metabolism*; D015398:Signal Transduction; D014230:Triazoles / Q000494:pharmacology*
chemical_list: D047072:Aromatase Inhibitors; D004967:Estrogens; D009570:Nitriles; D014230:Triazoles; D000077289:Letrozole; D019869:Phosphatidylinositol 3-Kinases; D017526:Receptor, IGF Type 1; D051057:Proto-Oncogene Proteins c-akt