Figure 4

Effects of Crizotinib in Pre-clinical Models of ALK + IBC. A-B. Treatment of mice bearing FC-IBC01 xenografts with DMSO vehicle control had no detectable apoptosis as determined by detection of TUNEL staining as a marker of programmed cell death. Figure 4 A shows the lack of green fluorescence associated with TUNEL staining and Figure 4 B shows a lack of green fluorescence which detects TUNEL staining, with detection of blue fluorescence which is associated with nuclear DNA based on Topro-3 staining. C and D. Treatment of mice bearing FC-IBC01 xenografts with 83 mg/kg Crizotinib resulted in significant apoptosis of FC-IBC01 tumor cells, as assessed by detection of TUNEL as a marker for programmed cell death, as denoted by the green fluorescence (Figure 4 C) and double label of green fluorescence which detects TUNEL staining and blue fluorescence detects nuclear DNA based on Topro-3 staining (Figure 4 D). E and F. Positive control for TUNEL staining using DNAse I treatment of xenograft tissues. G. Comparative quantitative analysis of Crizotinib-induced apoptosis in FC-IBC01 tumor xenografts demonstrates a significant increase in detection of TUNEL positive cells (P ≤ 0.0001). H and I. Comparative analysis of inhibitory effects of Crizotinib on phospho-ALK-Y-1604 signaling activation in FC-IBC01 and Mary-X pre-clinical models of IBC. Treatment of mice bearing FC-IBC01 (H) or Mary-X (I) with 83 mg/kg Crizotinib significantly inhibited phospho-ALK- Y-1604 (P ≤ 0.0001).