title: Targeting of CD34+CD38- cells using Gemtuzumab ozogamicin (Mylotarg) in combination with tipifarnib (Zarnestra) in Acute Myeloid Leukaemia.
abstract: BACKGROUND
  The CD34+CD38- subset of AML cells is enriched for resistance to current chemotherapeutic agents and considered to contribute to disease progression and relapse in Acute Myeloid Leukaemia (AML) patients following initial treatment.
  
  
  METHODS
  Chemosensitivity in phenotypically defined subsets from 34 primary AML samples was measured by flow cytometry following 48 hr in vitro treatment with gemtuzumab ozogamicin (GO, Mylotarg) and the farnesyltransferase inhibitor tipifarnib/zarnestra. The DNA damage response was measured using flow cytometry, immunofluorescence and immunohistochemistry.
  
  
  RESULTS
  Using a previously validated in vitro minimal residual disease model, we now show that the combination of GO (10 ng/ml) and tipifarnib (5 μM) targets the CD34+CD38- subset resulting in 65% median cell loss compared to 28% and 13% CD34+CD38- cell loss in GO-treated and tipifarnib-treated cells, respectively. Using phosphokinome profiling and immunofluorescence in the TF-1a cell line, we demonstrate that the drug combination is characterised by the activation of a DNA damage response (induction of γH2A.X and thr68 phosphorylation of chk2). Higher induction of γH2AX was found in CD34+CD38- than in CD34+CD38+ patient cells. In a model system, we show that dormancy impairs damage resolution, allowing accumulation of γH2AX foci.
  
  
  CONCLUSIONS
  The chemosensitivity of the CD34+CD38- subset, combined with enhanced damage indicators, suggest that this subset is primed to favour programmed cell death as opposed to repairing damage. This interaction between tipifarnib and GO suggests a potential role in the treatment of AML.
authors.affiliation: Division of Haematology, University of Nottingham, Nottingham, UK.
authors.forename: Mays
authors.initials: M
authors.lastname: Jawad
authors.forename: Ning
authors.initials: N
authors.lastname: Yu
authors.forename: Claire
authors.initials: C
authors.lastname: Seedhouse
authors.forename: Karuna
authors.initials: K
authors.lastname: Tandon
authors.forename: Nigel H
authors.initials: NH
authors.lastname: Russell
authors.forename: Monica
authors.initials: M
authors.lastname: Pallis
mesh_terms: D051997:ADP-ribosyl Cyclase 1 / Q000378:metabolism*; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D000617:Aminoglycosides / Q000494:pharmacology*; D061067:Antibodies, Monoclonal, Humanized / Q000494:pharmacology*; D018952:Antigens, CD34 / Q000378:metabolism*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004249:DNA Damage / Q000187:drug effects; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D000079982:Gemtuzumab; D006657:Histones / Q000378:metabolism; D006801:Humans; D053650:Interleukin-3 Receptor alpha Subunit / Q000378:metabolism; D015470:Leukemia, Myeloid, Acute / Q000188:drug therapy / Q000378:metabolism*; D014411:Neoplastic Stem Cells / Q000187:drug effects / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism; D000090243:Nucleophosmin; D015363:Quinolones / Q000494:pharmacology*; D063268:Sialic Acid Binding Ig-like Lectin 3 / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D051941:fms-Like Tyrosine Kinase 3 / Q000378:metabolism
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000617:Aminoglycosides; D061067:Antibodies, Monoclonal, Humanized; D018952:Antigens, CD34; D000970:Antineoplastic Agents; D006657:Histones; D053650:Interleukin-3 Receptor alpha Subunit; D009687:Nuclear Proteins; D015363:Quinolones; D063268:Sialic Acid Binding Ig-like Lectin 3; D000090243:Nucleophosmin; D000079982:Gemtuzumab; D051941:fms-Like Tyrosine Kinase 3; D051997:ADP-ribosyl Cyclase 1; C402769:tipifarnib