title: Evaluation of bioactive sphingolipids in 4-HPR-resistant leukemia cells.
abstract: BACKGROUND
  N-(4-hydroxyphenyl)retinamide (4-HPR, fenretinide) is a synthetic retinoid with potent pro-apoptotic activity against several types of cancer, but little is known regarding mechanisms leading to chemoresistance. Ceramide and, more recently, other sphingolipid species (e.g., dihydroceramide and dihydrosphingosine) have been implicated in 4-HPR-mediated tumor cell death. Because sphingolipid metabolism has been reported to be altered in drug-resistant tumor cells, we studied the implication of sphingolipids in acquired resistance to 4-HPR based on an acute lymphoblastic leukemia model.
  
  
  METHODS
  CCRF-CEM cell lines resistant to 4-HPR were obtained by gradual selection. Endogenous sphingolipid profiles and in situ enzymatic activities were determined by LC/MS, and resistance to 4-HPR or to alternative treatments was measured using the XTT viability assay and annexin V-FITC/propidium iodide labeling.
  
  
  RESULTS
  No major crossresistance was observed against other antitumoral compounds (i.e. paclitaxel, cisplatin, doxorubicin hydrochloride) or agents (i.e. ultra violet C, hydrogen peroxide) also described as sphingolipid modulators. CCRF-CEM cell lines resistant to 4-HPR exhibited a distinctive endogenous sphingolipid profile that correlated with inhibition of dihydroceramide desaturase. Cells maintained acquired resistance to 4-HPR after the removal of 4-HPR though the sphingolipid profile returned to control levels. On the other hand, combined treatment with sphingosine kinase inhibitors (unnatural (dihydro)sphingosines ((dh)Sph)) and glucosylceramide synthase inhibitor (PPMP) in the presence or absence of 4-HPR increased cellular (dh)Sph (but not ceramide) levels and were highly toxic for both parental and resistant cells.
  
  
  CONCLUSIONS
  In the leukemia model, acquired resistance to 4-HPR is selective and persists in the absence of sphingolipid profile alteration. Therapeutically, the data demonstrate that alternative sphingolipid-modulating antitumoral strategies are suitable for both 4-HPR-resistant and sensitive leukemia cells. Thus, whereas sphingolipids may not be critical for maintaining resistance to 4-HPR, manipulation of cytotoxic sphingolipids should be considered a viable approach for overcoming resistance.
authors: Department of Cell Biology and Histology, School of Medicine and Dentistry, University of the Basque Country, Barrio Sarriena s/n, 48940 Leioa (Bizkaia), Spain.
authors: Aintzane
authors: A
authors: Apraiz
authors: Jolanta K
authors: JK
authors: Idkowiak-Baldys
authors: María Dolores
authors: MD
authors: Boyano
authors: Gorka
authors: G
authors: Pérez-Yarza
authors: Yusuf A
authors: YA
authors: Hannun
authors: Aintzane
authors: A
authors: Asumendi
mesh_terms: D000704:Analysis of Variance; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D017313:Fenretinide / Q000494:pharmacology*; D006801:Humans; D007938:Leukemia / Q000188:drug therapy / Q000378:metabolism*; D010088:Oxidoreductases / Q000378:metabolism; D013107:Sphingolipids / Q000378:metabolism*; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000378:metabolism*; D014407:Tumor Cells, Cultured
chemical_list: D000970:Antineoplastic Agents; D013107:Sphingolipids; D017313:Fenretinide; D010088:Oxidoreductases; C108138:dihydroceramide desaturase; D013110:Sphingosine; C005682:safingol