title: Dynamics of ceramide generation and metabolism in response to fenretinide--Diversity within and among leukemia.
abstract: Fenretinide, N-(4-hydroxyphenyl)retinamide, (4-HPR), a synthetic retinoid, owes its cancer-toxic effects in part to the generation of ceramide, a potent tumor-suppressing sphingolipid. As such, 4-HPR has garnered considerable interest as a chemotherapeutic. Cancer cells, however, via various metabolic routes, inactivate ceramide, and this can limit 4-HPR efficacy. As relatively little is known regarding 4-HPR-induced ceramide management in acute myelogeneous leukemia (AML), we undertook the present study to evaluate the impact of 4-HPR on ceramide production, metabolism, and cytotoxicity. In KG-1, HL-60, and HL-60/VCR (multidrug resistant) human leukemia cells, 4-HPR induced 15-, 2-, and 20-fold increases in ceramide (measured using [3H]palmitic acid), respectively. By use of specific inhibitors we show that ceramide was produced by sphingomyelinase and de novo pathways in response to 4-HPR exposure. HL-60/VCR cells metabolized ceramide to glucosylceramide (GC). 4-HPR exposure (1.25-10 μM) reduced viability in all cell lines, with approximate IC50's ranging from 1 to 8.0 μM. Reactive oxygen species (ROS) were generated in response to 4-HPR treatment, and the concomitant cytotoxicity was reversed by addition of vitamin E. 4-HPR was not cytotoxic nor did it elicit ceramide formation in K562, a chronic myeloid leukemia cell line; however, K562 cells were sensitive to a cell-deliverable form of ceramide, C6-ceramide. Treatment of Molt-3, an acute lymphoblastic leukemia cell line, with 4-HPR revealed moderate ceramide production (5-fold over control), robust conversion of ceramide to GC and sphingomyelin, and resistance to 4-HPR and C6-ceramide. In conclusion, this work demonstrates diversity within and among leukemia in 4-HPR sensitivity and ceramide generation and subsequent metabolism. As such, knowledge of these metabolic pathways can provide guidance for enhancing ceramide-driven effects of 4-HPR in treatment of leukemia.
authors.affiliation: Department of Biochemistry and Molecular Biology, East Carolina University, Brody School of Medicine, East Carolina Diabetes and Obesity Institute, Greenville, NC 27834, USA; Affiliated with Department of Pharmacology, Faculty of Veterinary Medicine, South Valley University, Qena 83523, Egypt.
authors.forename: Samy A F
authors.initials: SA
authors.lastname: Morad
authors.affiliation: Department of Biochemistry and Molecular Biology, East Carolina University, Brody School of Medicine, East Carolina Diabetes and Obesity Institute, Greenville, NC 27834, USA.
authors.forename: Traci S
authors.initials: TS
authors.lastname: Davis
authors.affiliation: University of Virginia Cancer Center, Charlottesville, VA 22908-0716, USA.
authors.forename: Mark
authors.initials: M
authors.lastname: Kester
authors.affiliation: University of Virginia Cancer Center, Charlottesville, VA 22908-0716, USA; Department of Medicine, Hematology/Oncology, University of Virginia, Charlottesville, VA 22908-0716, USA.
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Loughran
authors.affiliation: Department of Biochemistry and Molecular Biology, East Carolina University, Brody School of Medicine, East Carolina Diabetes and Obesity Institute, Greenville, NC 27834, USA. Electronic address: cabotm@ecu.edu.
authors.forename: Myles C
authors.initials: MC
authors.lastname: Cabot
mesh_terms: D000970:Antineoplastic Agents / Q000378:metabolism / Q000494:pharmacology*; D002470:Cell Survival / Q000187:drug effects; D002518:Ceramides / Q000096:biosynthesis*; D002855:Chromatography, Thin Layer; D017313:Fenretinide / Q000378:metabolism / Q000494:pharmacology*; D018922:HL-60 Cells; D006801:Humans; D007938:Leukemia / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D000970:Antineoplastic Agents; D002518:Ceramides; D017382:Reactive Oxygen Species; D017313:Fenretinide
keywords: Acute myelogenous leukemia; Ceramide metabolism; Fenretinide/4-HPR; Leukemia; Sphingolipid metabolism