Autocrine RTK driven proliferation and colony formation in BCa cell lines.

(A) PCR products showing siRNA suppression (upper and middle panels in each group) vs. 18S rRNA loading controls (lower panel in each group) resolved by agarose gel electrophoresis and visualized using ethidium bromide staining, from cell lines used for results shown in panels B–F. Upper left: J82 cells transfected with siRNA directed against GAS6 or AXL used for results shown in panel B. Upper middle: FL3 cells transfected with siRNA directed against MST1 or MST1R used for results shown in panel C. Upper right: FL3 cells transfected with siRNA directed against CSF1 or CSF1R used for results shown in panel D. Lower left: J82 cells transfected with siRNA directed against AXL used for results shown in panel E. Lower right: FL3 cells transfected with siRNA directed against MST1R or CSF1R used for results shown in panel F. (B) Proliferation (abs at 570 nm, 3 d, 37°C) of J82 cells in 1% FBS previously transfected with siRNA directed against GAS6 (siGAS6), AXL (siAXL) or control siRNA (siControl) that were left untreated or treated with Gas6 (5.7 nM) in the absence or presence of cabozantinib (cabo, 300 nM). (C) Proliferation of FL3 cells in 1% FBS previously transfected with siRNA directed against MST1 (siMST1), MST1R (siMST1R) or control siRNA (siControl) that were left untreated or treated with MST1 (5 nM) in the absence or presence of cabozantinib (cabo, 300 nM). (D) Proliferation of FL3 cells in 1% FBS previously transfected with siRNA directed against CSF1 (siCSF1), CSF1R (siCSFR) or control siRNA (siControl) that were left untreated or treated with CSF1 (10 nM) in the absence or presence of cabozantinib (cabo, 300 nM). (E) Anchorage independent growth (abs at 570 nm) of J82 cells in 1% FBS previously transfected with siRNA directed against AXL (siAXL) or control siRNA (siControl) that were left untreated or treated with Gas6 (5.7 nM) in the absence or presence of cabozantinib (cabo, 300 nM). (F) Anchorage independent growth of FL3 cells in 1% FBS previously transfected with siRNA directed against MST1R (siMST1R), CSF1R (siCSFR) or control siRNA (siControl) that were left untreated or treated with MST1 (5 nM) or CSF1 (10 nM) in the absence or presence of cabozantinib (cabo, 300 nM). In panels C—F, values are the mean +/- SD from triplicate samples; asterisks indicate significant difference from untreated control (p < 0.05, ns, not significant). In all panels, data are representative of 3 or more independent experiments.