title: Role of human longevity assurance gene 1 and C18-ceramide in chemotherapy-induced cell death in human head and neck squamous cell carcinomas.
abstract: In this study, quantitative isobologram studies showed that treatment with gemcitabine and doxorubicin, known inducers of ceramide generation, in combination, supra-additively inhibited the growth of human UM-SCC-22A cells in situ. Then, possible involvement of the human homologue of yeast longevity assurance gene 1 (LASS1)/C(18)-ceramide in chemotherapy-induced cell death in these cells was examined. Gemcitabine/doxorubicin combination treatment resulted in the elevation of mRNA and protein levels of LASS1 and not LASS2-6, which was consistent with a 3.5-fold increase in the endogenous (dihydro)ceramide synthase activity of LASS1 for the generation of C(18)-ceramide. Importantly, the overexpression of LASS1 (both human and mouse homologues) enhanced the growth-inhibitory effects of gemcitabine/doxorubicin with a concomitant induction of caspase-3 activation. In reciprocal experiments, partial inhibition of human LASS1 expression using small interfering RNA (siRNA) prevented cell death by about 50% in response to gemcitabine/doxorubicin. In addition, LASS1, and not LASS5, siRNA modulated the activation of caspase-3 and caspase-9, but not caspase-8, in response to this combination. Treatment with gemcitabine/doxorubicin in combination also resulted in a significant suppression of the head and neck squamous cell carcinoma (HNSCC) tumor growth in severe combined immunodeficiency mice bearing the UM-SCC-22A xenografts. More interestingly, analysis of endogenous ceramide levels in these tumors by liquid chromatography/mass spectroscopy showed that only the levels of C(18)-ceramide, the main product of LASS1, were elevated significantly (about 7-fold) in response to gemcitabine/doxorubicin when compared with controls. In conclusion, these data suggest an important role for LASS1/C(18)-ceramide in gemcitabine/doxorubicin-induced cell death via the activation of caspase-9/3 in HNSCC.
authors.affiliation: Department of Biochemistry and Molecular Biology, Medical University of South Carolina, 173 Ashley Avenue, Charleston, SC 29425, USA.
authors.forename: Can E
authors.initials: CE
authors.lastname: Senkal
authors.forename: Suriyan
authors.initials: S
authors.lastname: Ponnusamy
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Rossi
authors.forename: Jacek
authors.initials: J
authors.lastname: Bialewski
authors.forename: Debijyati
authors.initials: D
authors.lastname: Sinha
authors.forename: James C
authors.initials: JC
authors.lastname: Jiang
authors.forename: S Michal
authors.initials: SM
authors.lastname: Jazwinski
authors.forename: Yusuf A
authors.initials: YA
authors.lastname: Hannun
authors.forename: Besim
authors.initials: B
authors.lastname: Ogretmen
mesh_terms: D000818:Animals; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D015153:Blotting, Western; D002294:Carcinoma, Squamous Cell / Q000188:drug therapy*; D020169:Caspases / Q000187:drug effects / Q000378:metabolism; D045744:Cell Line, Tumor; D002518:Ceramides / Q000737:chemistry / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D002853:Chromatography, Liquid; D003841:Deoxycytidine / Q000008:administration & dosage / Q000031:analogs & derivatives; D004317:Doxorubicin / Q000008:administration & dosage; D005455:Fluorescent Antibody Technique; D006258:Head and Neck Neoplasms / Q000188:drug therapy*; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D016513:Mice, SCID; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism; D010957:Plasmids; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051049:Sphingosine N-Acyltransferase; D015996:Survival Rate; D000093542:Gemcitabine
chemical_list: D002518:Ceramides; D008565:Membrane Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D003841:Deoxycytidine; D004317:Doxorubicin; D010088:Oxidoreductases; C473786:dihydroceramide synthase; C493790:CERS1 protein, human; C505722:CERS5 protein, human; D051049:Sphingosine N-Acyltransferase; D020169:Caspases; D000093542:Gemcitabine