Figure 4
Combination treatment sensitizes melanoma cells but not primary cells to TRAIL. (a) SBCL2 (RGP), WM-115 (VGP) and 451-LU (MM) were stimulated with TRAIL (100 ng/ml), cisplatin (20 μM) or UVB (200 J/m2) alone, or the combination treatment. After 16 h, apoptosis was determined using a Cell Death Detection ELISA. [R]=resistant; [s]=slightly sensitive; [S]=sensitive. *P<0.01 comparing co-treated with TRAIL-only-treated cells of the same line. (b) 451-LU cells (MM) were stimulated as in (a). After 16 h, protein status of XIAP and cleavage pattern of caspase-3 were determined by western blot analysis. β-Actin served as loading control. One out of three experiments is shown. (c) Primary fibroblasts, keratinocytes and melanocytes as well as 451-LU melanoma cells were treated and analyzed as in (a). Results in (a) and (c) show the mean±S.D. of three independent experiments. (d) Skin equivalents containing 451-LU nests were subjected to either short-term treatment: TRAIL (250 ng/ml), cisplatin (20 μM), UVB (400 J/m2) or in combination applied twice at days 13 and 15; or long-term treatment: TRAIL (250 ng/ml), cisplatin (2.5 μM), UVB (200 J/m2) or in combination applied five times over 10 days. At day 17, tumor volumes determined using ImageJ software. (e) Statistical analysis of tumor nest sizes counting the 10 largest melanoma nests, and overall tumor nest numbers (mean±S.D.) of three sections of skin equivalents derived from five different donors. *P<0.05, **P<0.01 comparing co-treated with TRAIL-only-treated tumor nests
