Supplementary Figure 1 from Critical Role of STAT3 in IL-6–Mediated Drug Resistance in Human Neuroblastoma

Supplementary Figure 1 - PDF file 231K, Inhibition of STAT3 activation in CHLA-255 cells by anti-IL-6R blocking antibody. A, Expression of pSTAT3 and STAT3 in CHLA-255 cells was examined by Western blot analysis as in Fig. 1A. When indicated, cells were treated with tocilizumab (2g/mL) for 12 hours before treatment with IL-6 with or without sIL-6R. The data are representative of 2 separate experiments showing similar results. B, Nuclear extracts from CHLA-255 treated with IL-6 and sIL-6R in the presence and absence of tocilizumab as above described were examined for STAT3 DNA-binding activity by EMSA as indicated in Materials and Methods. Lane 8 is same as lane 5, except for the addition of a supershifting anti-STAT3 antibody also added to lane 2 (DU145 control). Top band (labeled STAT3) represents STAT3/STAT3 homodimers bound to DNA. C, CHLA-255 transfected with a STAT3 Firefly luciferase vector and a Renilla luciferase control vector were treated with IL-6 (10ng/mL) alone or in combination with sIL-6R (25ng/mL) as shown in Fig. 1 in the absence or presence of tocilizumab (4g/mL). After 24 hours, the activity of the STAT3 reporter was examined by Dual Luciferase assay as indicated in Materials and Methods. The data represent the mean (SD) ratio Firefly/Renilla luciferase activity from five samples for each experimental condition