Fig. 1

TRAP IL-1 blocks cell cycle, proliferation, and clonogenicity in CTX-resistant SW48. SW48 and SW48 CXR cells (2 × 10³) were seeded in 96-well plate. The day after SW48 cells were treated with control medium, IL-1 (α and β, 10 ng/ml each), CTX (50 μg/ml) either alone or in combination with TRAP IL-1 (20 μg/ml). SW48 CXR cells were treated with control cetuximab (50 μg/ml), only medium (10% FBS), or TRAP IL-1 (TRAP 20 μg/ml). After 5 days from seeding, cell viability was assessed through AlamarBlue assay. Histograms show results obtained from SW48 (a) and SW48 CXR (b). Statistical analysis was carried out using one-way ANOVA and significance determined with respectively Dunnett’s and Tukey’s multiple comparisons test **p < 0.01, ***p < 0.001. To measure clonogenicity, 1 × 10⁴ of SW48 and SW48-CXR were seeded in 12-well plates in triplicate and treated after 24 h as described in a and b. After 10 days, cells were fixed in 4% PFA and stained with crystal violet for 30 min. The ability of cells to grow in a colony was determined by counting them through ImageJ software. Percentage of covered area is shown for SW48 (c) and SW48-CXR (d). Statistical analysis was carried out using one-way ANOVA and significance calculated with Dunnett’s multiple comparisons test *p < 0.05, **p < 0.01, ***p < 0.001. (e) Cell cycle of SW48 and SW48-CXR (5 × 10⁵) cells treated or not for 48 h with TRAP IL-1 (20 μg/ml) was determined by analyzing DNA content of cells stained with propidium iodide by FACS. Data were processed though Citoflex software