Colorectal cancer organoid-stroma biobank allows subtype-specific assessment of individualized therapy responses
Description
2 Tab-delimited Text files contain the raw luminescence data of the drug treatment experiments: Drug_sensitivity_all_lines.txt: Sensitivity of patient-derived tumor organoids to 5-FU, Oxaliplatin, SN-38 and Gefitinib in mono- and coculture with auto- or heterologous CAFs. Chemogenomic_coculture_screen.txt: Sensitivity of patient-derived tumor organoids to chemogenomic drug library. Experiments were performed in presence of CAFs and in presence or absence of a sub-lethal concentration of Gefitinib.
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Organoid viability in co-cultures was measured using luciferase/GFP transduced organoids. Cells were either cultured for 3 days in full medium, followed by two rounds of washes and culture for 6 days in reduced medium. On day 3, drugs were added to the reduced medium with a D300e digital dispenser (Tecan). 0.1-5 nM SN-38 (Selleckchem), 0.5-60 μM 5-Fluorouracil (Sigma Aldrich), 0.5-60 μM Oxaliplatin (Sigma Aldrich) and 0.01-10 μM Gefitinib (Selleckchem) were administered in 7-point dilutions. Compounds were dissolved in DMSO and DMSO content in all wells was normalized, not exceeding 1% final volume. Medium was replenished after three days and cell viability was assessed in triplicates after 6 days of drug exposure using the One-Glo Ex assay. Luminescence was recorded on a SpectraMax iD3 Multi-Mode Microplate Reader (Molecular Devices). Individual values were excluded if cells were lost due to technical errors or if seeding was not uniform. For drug library screening in resistant co-cultures, luciferase expressing organoids (O14 or O23) and non-modified CAFs (F14) were seeded as described above in rich medium. The library contained 186 test compounds and 106 negative controls, that were assembled as part of the SGC Chemical Probe programs including the Donated Probe program and the EUbOPEN consortium. Master plates of compounds were dissolved in DMSO at 1 mM or 10 mM. 3 days after seeding the culture medium was changed to growth factor reduced medium (as above) following co-treatment. The library was administered with an automatic multichannel pipette (Integra Mini 96) at final concentration of 0.1 or 1 μM (10,000 fold dilution) to co- cultures in presence and absence of a sub-lethal dose of Gefitinib, (O14: 1 μM and O23: 0.2 μM) or SN-38 (O23: 1.36 nM). Each experiment was performed in two replicates. The medium was renewed after 3 days, and treatment was stopped after 6 days followed by measurement of organoid viability using ONE-Glo® EX (Promega).