Figure 2
XIAP overexpression inhibits ADCC response in SUM149 cells through caspase inhibition. (a) Percent cell lysis of SUM149 FG9 and wtXIAP cells incubated with cetuximab alone, PBMC alone or cetuximab and IL-2 activated PBMCs for 4 h, n=2–3. (b) Schematic of XIAP mutants used in this study. D148A mutation is known to disrupt binding of caspases 3 and 7, whereas W310A disrupts caspase-9 binding. The H467A point mutation abolishes E3 ubiquitin ligase activity. (c) Western immunoblot of XIAP expression in cell lines transduced as indicated. FG9 is an empty vector as described in Materials and Methods. (d) Caspase activity and (e) viability of XIAP variant cell lines treated as indicated. Bars represent fold change in luminescence (d) or mean±S.E.M. % cell death (e), n=2–3. (f) Percent cell lysis of wtXIAP, +DW/AA and +H/A cells incubated with cetuximab alone, PBMC alone or cetuximab and IL-2 activated PBMCs for 4 h. Bars represent mean±S.E.M. calculated percent lysis, n=2–3. *P<0.05, **P<0.005, #P<0.001 (compared with FG9 unless otherwise indicated)
