Development of a Single-Cell Technique to Increase Yield and Use of Gastrointestinal Cancer Organoids for Personalized Medicine Application.

Abstract Background Organoids are excellent three-dimensional in vitro models of gastrointestinal cancers. However, patient derived organoids (PDOs) remain inconsistent and unreliable for rapid actionable drug sensitivity testing due to size variation and limited material. Methods On day10/passage2 after standard creation of organoids, ½ PDOs were dissociated into single-cells with TrypLETM Express Enzyme/DNase I and mechanical dissociation; and ½ PDOs were expanded by standard technique. HE and N=6, pancreatic ductal adenocarcinoma, PDAC), which formed epithelialized cystic structures and by IHC exhibited CK7(high)/CK20(low) expression patterns mirroring parent tissues. Compared to paired standard PDOs, single-cells (N=2, PDAC; N=2, GC) showed similar architecture, albeit smaller and more uniform. Importantly, single-cells demonstrated similar sensitivity to cytotoxic drugs to matched PDOs. Conclusions PDO single-cells are accurate for rapid clinical drug testing in gastrointestinal cancers. Utilizing early passage PDO single-cells facilitates high-volume drug testing, decreasing time from tumor sampling to actionable clinical decisions and provides a personalized medicine platform to optimally select drugs for gastrointestinal cancer patients.