Comparison of responsiveness to anticancer drugs between human ALI tumor colorectal organoids and 2D cultured colorectal cancer cell lines. Schematic experimental design of an anticancer drug treatment for human ALI tumor colorectal organoids from three patients and colorectal cancer cell lines (a). After tumor ALI organoids were seeded into Matrigel, they were treated with 5-fluorouracil (5-FU) ((b), (c) n = 5 each for three patients (T1, T2, and T3); 10–300 μg/mL) or Irinotecan ((d), (e) n = 5 each for three patients (T1, T2, and T3); 10–300 μM) for 6 days. Representative phase contrast images of tumor organoids treated with 5-FU or Irinotecan are shown (b, d). Scale bar: 200 μm. Cell viability was determined using Alamar Blue assay and 100% represents cell viability of each control (c, e). Effects of treatment with anticancer drugs on colorectal cancer cell lines, SW480, SW620, and HCT116. After cells were seeded into Matrigel, they were treated with 5-FU ((f) n = 5, each cell line, 0.1–100 μg/mL) or Irinotecan ((g) n = 5, each cell line, 0.1–100 μM) for 3 days. Cell viability was determined using an Alamar Blue assay and 100% represents cell viability of each control (f, g). Half maximal inhibitory concentration (IC50) values for 5-FU or Irinotecan in tumor organoids from each patient and colorectal cancer cell (h). Results were expressed as mean ± SEM.