Fig. 6

Combined ILEI KD and crizotinib treatment reduces tumor growth by inhibiting proliferation and MMP expression. a, j Fold growth ± SEM of NCI-H441 (a) and NCI-H1993 (j) control (shCont) and ILEI KD (shILEI) tumors upon vehicle or crizotinib (Crizo) treatment normalized to size at treatment start. Crizotinib-treated NCI-H1993 tumors were allowed to grow for an additional 11 days after treatment termination. b, k Tumor masses ± SEM of NCI-H441 (b) and of NCI-H1993 (k) shCont and shILEI tumors of vehicle or Crizo-treated mice. c, l Percentage of Ki67 positive tumor cells ± SEM of NCI-H441 (c) and NCI-H1993 (l) shCont and shILEI tumors of vehicle or Crizo-treated mice. d, m Percentage of activated caspase 3 (actCasp3) positive tumor cells ± SEM of NCI-H441 (d) and NCI-H1993 (m) shCont and shILEI tumors of vehicle or crizotinib-treated mice. e, n Gelatin zymography of protein extracts of NCI-H441 (e) and NCI-H1993 (n) shCont and shILEI tumors of vehicle- or crizotinib-treated mice (n = 3 pro group). Recombinant pro-MMP9, assay control. f, o Quantification of gels of panel e (f) and n (o). Pro- (filled color) and activated (patterned color) MMP-9 and MMP-2 levels ± SEM were normalized to respective total MMP levels of vehicle-treated control tumors. Statistics compares total MMP-9 and MMP-2 levels. g, p mRNA expression levels of MMP9 in NCI-H441 (g) and of MMP2 in NCI-H1993 (p) shCont and shILEI tumors of vehicle- or crizotinib-treated mice (n = 3 pro group). Expression was normalized to GAPDH and shown as fold change ± SEM over vehicle treated control tumors. h Representative images of MMP-9 IHC on NCI-H441 shCont and shILEI tumor sections of vehicle or crizotinib-treated mice. Arrowheads mark intracellular granular MMP-9 localization. Scale bar, 100 μm. i Percentage of MMP-9 positive tumor cells ± SEM in NCI-H441 shCont and shILEI tumors of vehicle or crizotinib-treated mice. Statistical significance was determined by two-way ANOVA (a, j), Student’s t-test (a, j) and one-way ANOVA (b, c, d, f, g, i, k, l, m, o, p) and is marked with asterisks (*p < 0.05; **p < 0.01; ***p < 0.001)