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. 2010 Sep 28;103(9):1380–1390. doi: 10.1038/sj.bjc.6605896

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Copyright © 2010 Cancer Research UK

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Fenretinide sensitises ESFT cells to the DR apoptotic pathway to enhance cell death. (A) TC-32 cells were treated with fenretinide (Fen; 1.5 μM, 16 h) or vehicle control (Veh; ethanol treated cells), media were changed and cells were subsequently treated with the DR ligands TRAIL, FasL or NGF (0–40 ng/ml, 24 h). Viable cell number was determined by the Trypan blue exclusion assay. Results are presented as the mean of the viable cell number calculated as a percentage of untreated control cells±s.e.m. (n=9). Statistics indicate significant interactions between fenretinide and DR ligands, which is indicative of enhanced cell death; ^*P⩽0.001. TC-32 cells were pretreated with fenretinide (1.5 μM, 16 h) before DR ligands (40 ng/ml, 0-24 h) and caspase-8 activity was detected by flow cytometry. (Bi) Effect of fenretinide and (Bi) TRAIL, (Bii) FasL or NGF time course of caspase-8 cleavage in TC-32 cells. Results are presented as the mean of the percentage of cells with cleaved caspase-8, as detected within the M1 region±s.e.m. (n=9), ^*P⩽0.001. Untreated and etoposide (30 μM, 24 h)-treated Jurkat cells served as negative and positive control samples, respectively. (C) Total protein lysates from ESFT cells pretreated with fenretinide (Fen, 1.5 μM, 16 h) before TRAIL treatment (40 ng/ml, 24 h) were examined for both full length (FL, 22 kDa) and truncated (cleaved) bid (tBid, 15 kDa) by immunoblotting. Equal protein loading was confirmed by hybridisation to tubulin. M=molecular weight markers. Negative (Neg) control=untreated Jurkat cells, positive (Pos) control=etoposide-treated (25 μM, 16 h) Jurkat cells. Immunoblots are representative from three independent experiments.