title: Enhancement of fludarabine sensitivity by all-trans-retinoic acid in chronic lymphocytic leukemia cells.
abstract: BACKGROUND
  A subset of patients with fludarabine-resistant chronic lymphocytic leukemia has previously been shown to express elevated intracellular levels of the concentrative high-affinity fludarabine transporter hCNT3, without any detectable related activity. We have recently shown that all-trans-retinoic acid is capable of inducing hCNT3 trafficking to plasma membrane in the MEC1 cell line. We, therefore, evaluated the effect of all-trans-retinoic acid on hCNT3 in primary chronic lymphocytic leukemia cells as a suitable mechanism to improve fludarabine-based therapy of chronic lymphocytic leukemia.
  
  
  METHODS
  Cells from 23 chronic lymphocytic leukemia patients wild-type for P53 were analyzed for ex vivo sensitivity to fludarabine. hCNT3 activity in chronic lymphocytic leukemia cell samples was evaluated by measuring the uptake of [8-(3)H]-fludarabine. The amounts of transforming growth factor-β1 and hCNT3 messenger RNA were analyzed by real-time polymerase chain reaction. The effect of all-trans-retinoic acid on hCNT3 subcellular localization was analyzed by confocal microscopy and its effect on fludarabine-induced apoptosis was evaluated by flow cytometry analysis using annexin V staining.
  
  
  RESULTS
  Chronic lymphocytic leukemia cases showing higher ex vivo basal sensitivity to fludarabine also had a greater basal hCNT3-associated fludarabine uptake capacity compared to the subset of patients showing ex vivo resistance to the drug. hCNT3 transporter activity in chronic lymphocytic leukemia cells from the latter patients was either negligible or absent. Treatment of the fludarabine-resistant subset of chronic lymphocytic leukemia cells with all-trans-retinoic acid induced increased fludarabine transport via hCNT3 which was associated with a significant increase in fludarabine sensitivity.
  
  
  CONCLUSIONS
  Improvement of ex vivo fludarabine sensitivity in chronic lymphocytic leukemia cells is associated with increased hCNT3 activity after all-trans-retinoic acid treatment.
authors.affiliation: Departament de Bioquímica i Biologia Molecular, Universitat de Barcelona, IBUB & CIBER EHD Diagonal 645, 08028 Barcelona, Spain. pfernandezcalotti@ub.edu
authors.forename: Paula X
authors.initials: PX
authors.lastname: Fernández-Calotti
authors.forename: Mónica
authors.initials: M
authors.lastname: Lopez-Guerra
authors.forename: Dolors
authors.initials: D
authors.lastname: Colomer
authors.forename: Marçal
authors.initials: M
authors.lastname: Pastor-Anglada
mesh_terms: D000368:Aged; D000369:Aged, 80 and over; D017304:Annexin A5; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D001692:Biological Transport / Q000187:drug effects; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000188:drug therapy* / Q000473:pathology; D008297:Male; D026901:Membrane Transport Proteins / Q000819:agonists* / Q000096:biosynthesis; D008875:Middle Aged; D061251:Primary Cell Culture; D012333:RNA, Messenger / Q000096:biosynthesis; D060888:Real-Time Polymerase Chain Reaction; D014212:Tretinoin / Q000494:pharmacology*; D014316:Tritium; D014740:Vidarabine / Q000031:analogs & derivatives* / Q000494:pharmacology
chemical_list: D017304:Annexin A5; D000970:Antineoplastic Agents; D026901:Membrane Transport Proteins; D012333:RNA, Messenger; C065171:cif nucleoside transporter; D014316:Tritium; D014212:Tretinoin; D014740:Vidarabine; C024352:fludarabine