Fig. 1

GSDMB is induced in response to anti-HER2 therapies, and its silencing increases the sensitive to lapatinib treatment. A Representative immunofluorescence images of GSDMB and HER2 expression in GSDMB/HER2-positive gastric carcinomas. Scale bar, 20 µm. Nuclei were counterstained with DAPI. Representative pie chart of GSDMB expression statistics in HER2 + gastric carcinomas (see Supplementary Table 1 for extra clinical features and statistics). B-C Relative mRNA (left) and protein levels (right) of GSDMB and HER2 in HCC1954 (B) and OE19 (C) cells treated with IC50 of lapatinib (2 µM and 0.7 µM, respectively) at indicated time points. D GSDMB and HER2 protein levels in lapatinib resistant HCC1954 and OE19 cells (LR) and their corresponding control cells (C) treated chronically with lapatinib and DMSO, respectively, and after ten days of drug removal. Study of the different cytotoxic effect of the chronic lapatinib treatment in these cells was analyzed by cell viability assays. E Relative mRNA levels of GSDMB and HER2 in trastuzumab (TRC1 and TRC2) and lapatinib (LRC1 and LRC1) resistant tumors, compared to the parental tumor (C) derived from a HER2 breast cancer PDXs. F GSDMB expression was reduced in HCC1954 LR (left) and OE19 LR cells (right) by two specific siRNAs (siGB1 and siGB2), in comparison with the control (siNTC). Cytotoxic effect of the presence of lapatinib (2 µM and 1.5 µM, respectively) in GSDMB-siRNAs-silenced cells HCC1954 LR (left) and OE19 LR (right) cells was assessed by cell viability assays. G Cytotoxic effect after 72 h treatment with IC50 of lapatinib (2 µM and 0.7 µM, respectively) in GSDMB-shRNAs-silenced cells HCC1954 and OE19 (right) cells was assessed by cell growth (viability assays, left panel) and death (Annexin V FITC and PI, right panel). Annexin V-FITC positive cells alone (A + /PI-) and Annexin V-FITC and PI doubled stained (A + /PI +) were defined as apoptotic cells. The number over the bars indicate the ratio of cell death relative to the shNTC condition. Statistical significance was determined by two-tailed unpaired t-test (*P < 0.05; **P < 0.01). Data are shown as the mean ± s.e.m. Three independent experiments with similar results were performed. In (B, C, E), gene expression was normalized to the mRNA levels of GAPDH. A, Annexin V; PI, propidium iodide. NTC, non-targeting control. DMSO, dimethyl sulfoxide. Lap, lapatinib