title: Organometallic nucleosides induce non-classical leukemic cell death that is mitochondrial-ROS dependent and facilitated by TCL1-oncogene burden.
abstract: BACKGROUND
  Redox stress is a hallmark of the rewired metabolic phenotype of cancer. The underlying dysregulation of reactive oxygen species (ROS) is interconnected with abnormal mitochondrial biogenesis and function. In chronic lymphocytic leukemia (CLL), elevated ROS are implicated in clonal outgrowth and drug resistance. The pro-survival oncogene T-cell leukemia 1 (TCL1) is causally linked to the high threshold towards classical apoptosis in CLL. We investigated how aberrant redox characteristics and bioenergetics of CLL are impacted by TCL1 and if this is therapeutically exploitable.
  
  
  METHODS
  Bio-organometallic chemistry provided compounds containing a cytosine nucleobase, a metal core (ferrocene, ruthenocene, Fe(CO)3), and a 5'-CH2O-TDS substituent. Four of these metal-containing nucleoside analogues (MCNA) were tested for their efficacy and mode of action in CLL patient samples, gene-targeted cell lines, and murine TCL1-transgenic splenocytes.
  
  
  RESULTS
  The MCNA showed a marked and selective cytotoxicity towards CLL cells. MCNA activity was equally observed in high-risk disease groups, including those of del11q/del17p cytogenetics and of clinical fludarabine resistance. They overcame protective stromal cell interactions. MCNA-evoked PARP-mediated cell death was non-autophagic and non-necrotic as well as caspase- and P53-independent. This unconventional apoptosis involved early increases of ROS, which proved indispensible based on mitigation of MCNA-triggered death by various scavengers. MCNA exposure reduced mitochondrial respiration (oxygen consumption rate; OCR) and induced a rapid membrane depolarization (∆ΨM). These characteristics distinguished the MCNA from the alkylator bendamustine and from fludarabine. Higher cellular ROS and increased MCNA sensitivity were linked to TCL1 expression. The presence of TCL1 promoted a mitochondrial release of in part caspase-independent apoptotic factors (AIF, Smac, Cytochrome-c) in response to MCNA. Although basal mitochondrial respiration (OCR) and maximal respiratory capacity were not affected by TCL1 overexpression, it mediated a reduced aerobic glycolysis (lactate production) and a higher fraction of oxygen consumption coupled to ATP-synthesis.
  
  
  CONCLUSIONS
  Redox-active substances such as organometallic nucleosides can confer specific cytotoxicity to ROS-stressed cancer cells. Their P53- and caspase-independent induction of non-classical apoptosis implicates that redox-based strategies can overcome resistance to conventional apoptotic triggers. The high TCL1-oncogenic burden of aggressive CLL cells instructs their particular dependence on mitochondrial energetic flux and renders them more susceptible towards agents interfering in mitochondrial homeostasis.
authors.affiliation: Laboratory of Lymphocyte Signaling and Oncoproteome, Department I of Internal Medicine, Center for Integrated Oncology (CIO) Köln-Bonn, and Excellence Cluster for Cellular Stress Response and Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany. christian_prinz@gmx.de.
authors.forename: Christian
authors.initials: C
authors.lastname: Prinz
authors.affiliation: Laboratory of Lymphocyte Signaling and Oncoproteome, Department I of Internal Medicine, Center for Integrated Oncology (CIO) Köln-Bonn, and Excellence Cluster for Cellular Stress Response and Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany. elena.vasyutina@uk-koeln.de.
authors.forename: Elena
authors.initials: E
authors.lastname: Vasyutina
authors.affiliation: Laboratory of Lymphocyte Signaling and Oncoproteome, Department I of Internal Medicine, Center for Integrated Oncology (CIO) Köln-Bonn, and Excellence Cluster for Cellular Stress Response and Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany. gregor.lohmann@uk-koeln.de.
authors.forename: Gregor
authors.initials: G
authors.lastname: Lohmann
authors.affiliation: Laboratory of Lymphocyte Signaling and Oncoproteome, Department I of Internal Medicine, Center for Integrated Oncology (CIO) Köln-Bonn, and Excellence Cluster for Cellular Stress Response and Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany. alexandra.schrader@uk-koeln.de.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Schrader
authors.affiliation: Division of Organic Chemistry, University of Cologne, Cologne, Germany. romansks@uni-koeln.de.
authors.forename: Steffen
authors.initials: S
authors.lastname: Romanski
authors.affiliation: Division of Organic Chemistry, University of Cologne, Cologne, Germany. pghirschhaeuser@gmx.de.
authors.forename: Christoph
authors.initials: C
authors.lastname: Hirschhäuser
authors.affiliation: Laboratory of Lymphocyte Signaling and Oncoproteome, Department I of Internal Medicine, Center for Integrated Oncology (CIO) Köln-Bonn, and Excellence Cluster for Cellular Stress Response and Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany. petra.mayer@uk-koeln.de.
authors.forename: Petra
authors.initials: P
authors.lastname: Mayer
authors.affiliation: Department of Pediatric Hematology/Oncology, Children's Hospital Cologne, Cologne, Germany. corazonfrias1@yahoo.com.
authors.forename: Corazon
authors.initials: C
authors.lastname: Frias
authors.affiliation: Department I of Internal Medicine, CIO Köln-Bonn, and CECAD, University of Cologne, Cologne, Germany. carmen.herling@uk-koeln.de.
authors.forename: Carmen D
authors.initials: CD
authors.lastname: Herling
authors.affiliation: Department I of Internal Medicine, CIO Köln-Bonn, and CECAD, University of Cologne, Cologne, Germany. michael.hallek@uk-koeln.de.
authors.forename: Michael
authors.initials: M
authors.lastname: Hallek
authors.affiliation: Division of Organic Chemistry, University of Cologne, Cologne, Germany. schmalz@uni-koeln.de.
authors.forename: Hans-Günther
authors.initials: HG
authors.lastname: Schmalz
authors.affiliation: Department of Pediatric Hematology/Oncology, Children's Hospital Cologne, Cologne, Germany. prokopa@kliniken-koeln.de.
authors.forename: Aram
authors.initials: A
authors.lastname: Prokop
authors.affiliation: Department of Internal Medicine, University of Erlangen, Erlangen, Germany. dimitrios.mougiakakos@uk-erlangen.de.
authors.forename: Dimitrios
authors.initials: D
authors.lastname: Mougiakakos
authors.affiliation: Laboratory of Lymphocyte Signaling and Oncoproteome, Department I of Internal Medicine, Center for Integrated Oncology (CIO) Köln-Bonn, and Excellence Cluster for Cellular Stress Response and Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany. marco.herling@uk-koeln.de.
authors.forename: Marco
authors.initials: M
authors.lastname: Herling
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D001343:Autophagy / Q000187:drug effects; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D004734:Energy Metabolism / Q000187:drug effects; D006706:Homeostasis / Q000187:drug effects; D006801:Humans; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000473:pathology*; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D009336:Necrosis; D009705:Nucleosides / Q000737:chemistry / Q000494:pharmacology*; D009857:Oncogenes; D009942:Organometallic Compounds / Q000737:chemistry / Q000494:pharmacology*; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D012307:Risk Factors; D017154:Stromal Cells / Q000187:drug effects / Q000473:pathology; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
chemical_list: D009705:Nucleosides; D009942:Organometallic Compounds; D011518:Proto-Oncogene Proteins; D017382:Reactive Oxygen Species; C090851:TCL1A protein, human; D016159:Tumor Suppressor Protein p53; D053148:Caspase 3