title: Triggering of Suicidal Erythrocyte Death by Gefitinib.
abstract: OBJECTIVE
  The epidermal growth factor receptor-tyrosine kinase inhibitor gefitinib is effective against several malignancies and is mainly utilized in the treatment of epidermal growth factor receptor mutation positive non-small cell lung cancer. The anti-cancer effect of the drug involves stimulation of apoptosis. Side effects of gefitinib include anemia. At least in theory, the development of anemia during gefitinib treatment could result from triggering of eryptosis, the suicidal erythrocyte death characterized by cell shrinkage and by cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Signaling potentially stimulating eryptosis include increase of cytosolic Ca2+ activity ([Ca2+]i) and generation of oxidative stress. The present study explored, whether gefitinib stimulates eryptosis and, if so, whether its effect involves Ca2+ entry and/or oxidative stress.
  
  
  METHODS
  Flow cytometry was employed to quantify cell volume from forward scatter, phosphatidylserine exposure at the cell surface from annexin-V-binding, [Ca2+]i from Fluo3-fluorescence, and reactive oxygen species (ROS) abundance from 2',7'-dichlorodihydrofluorescein diacetate (DCFDA) dependent fluorescence.
  
  
  RESULTS
  A 48 hours exposure of human erythrocytes to gefitinib (≥ 2 µg/ml) significantly decreased forward scatter and significantly increased the percentage of annexin-V-binding cells. Gefitinib did not significantly increase Fluo3-fluorescence but the effect of gefitinib on annexin-V-binding was significantly blunted by removal of extracellular Ca2+. Gefitinib further significantly increased DCFDA fluorescence.
  
  
  CONCLUSIONS
  Gefitinib triggers erythrocyte shrinkage and phospholipid scrambling of the erythrocyte cell membrane, an effect at least in part dependent on extracellular Ca2+ and paralleled by oxidative stress.
authors.affiliation: Department of Internal Medicine III, Eberhard-Karls-University of Tuebingen, Tuebingen, Germany.
authors.forename: Abdulla
authors.initials: A
authors.lastname: Al Mamun Bhuyan
authors.affiliation: Department of Internal Medicine III, Eberhard-Karls-University of Tuebingen, Tuebingen, Germany.
authors.forename: Teresa
authors.initials: T
authors.lastname: Wagner
authors.affiliation: Department of Internal Medicine III, Eberhard-Karls-University of Tuebingen, Tuebingen, Germany.
authors.forename: Hang
authors.initials: H
authors.lastname: Cao
authors.affiliation: Department of Internal Medicine III, Eberhard-Karls-University of Tuebingen, Tuebingen, Germany.|Department of Molecular Medicine II, Medical Faculty, Heinrich Heine University, Duesseldorf, Germany.
authors.forename: Florian
authors.initials: F
authors.lastname: Lang
mesh_terms: D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects*; D016923:Cell Death / Q000187:drug effects; D004910:Erythrocyte Membrane / Q000378:metabolism*; D005260:Female; D000077156:Gefitinib; D006801:Humans; D008297:Male; D018384:Oxidative Stress / Q000187:drug effects*; D010743:Phospholipids / Q000378:metabolism*; D011799:Quinazolines / Q000494:pharmacology*; D017382:Reactive Oxygen Species / Q000378:metabolism*
chemical_list: D010743:Phospholipids; D011799:Quinazolines; D017382:Reactive Oxygen Species; D000077156:Gefitinib; D002118:Calcium
keywords: Calcium; Eryptosis; Oxidative stress; Phosphatidylserine