title: Triggering of Suicidal Erythrocyte Death by Ruxolitinib.
abstract: OBJECTIVE
  The JAK1/JAK2 tyrosine kinase inhibitor ruxolitinib is widely used for the treatment of myeloproliferative neoplasm-associated myelofibrosis and other malignancies. Most important side effects include anemia. A common cause of anemia is accelerated suicidal death of erythrocytes or eryptosis, which is characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Mechanisms contributing to the triggering of eryptosis include oxidative stress, Ca2+ entry with increase of cytosolic Ca2+ activity ([Ca2+]i), and activation of distinct kinases, such as p38 mitogen activated protein (MAP) kinase. The present study explored whether and how ruxolitinib induces eryptosis.
  
  
  METHODS
  Phosphatidylserine exposure at the cell surface was estimated from annexin-V-binding, cell volume from forward scatter, hemolysis from hemoglobin release, [Ca2+]i from Fluo3-fluorescence, and ROS formation from DCFDA dependent fluorescence.
  
  
  RESULTS
  A 48 hours exposure of human erythrocytes to ruxolitinib (25 µM) significantly increased the percentage of annexin-V-binding cells and significantly decreased forward scatter. Ruxolitinib did not significantly modify Fluo3-fluorescence and DCFDA fluorescence and the effect of ruxolitinib on annexin-V-binding was not significantly modified by removal of extracellular Ca2+. The effect of ruxolitinib on annexin-V-binding was, however, significantly blunted by the p38 MAP kinase inhibitor SB203580 and virtually abolished by the p38 MAP kinase inhibitor skepinone.
  
  
  CONCLUSIONS
  Ruxolitinib triggers cell shrinkage and phospholipid scrambling of the erythrocyte cell membrane, an effect in part requiring p38 MAP kinase activity.
authors.affiliation: Department of Physiology, University of Tuebingen, Tuebingen, Germany.
authors.forename: Marilena
authors.initials: M
authors.lastname: Briglia
authors.forename: Antonella
authors.initials: A
authors.lastname: Fazio
authors.forename: Caterina
authors.initials: C
authors.lastname: Faggio
authors.forename: Stefan
authors.initials: S
authors.lastname: Laufer
authors.forename: Kousi
authors.initials: K
authors.lastname: Alzoubi
authors.forename: Florian
authors.initials: F
authors.lastname: Lang
mesh_terms: D002118:Calcium / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D002462:Cell Membrane / Q000187:drug effects; D048429:Cell Size / Q000187:drug effects; D004912:Erythrocytes / Q000166:cytology / Q000187:drug effects*; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D009570:Nitriles; D018384:Oxidative Stress / Q000187:drug effects; D010718:Phosphatidylserines / Q000378:metabolism; D011720:Pyrazoles / Q000494:pharmacology*; D011725:Pyridines / Q000494:pharmacology; D011743:Pyrimidines
chemical_list: D007093:Imidazoles; D009570:Nitriles; D010718:Phosphatidylserines; D011720:Pyrazoles; D011725:Pyridines; D011743:Pyrimidines; C540383:ruxolitinib; C093642:SB 203580; D002118:Calcium