Generation of biologically responsive colon-like intestinal tissue patches from human induced pluripotent stem cells using a rapid co-differentiation platform

• Developed a rapid (8-day) serum-free co-differentiation protocol for hiPSCs to generate multiple intestinal cell lineages (epithelial, mesenchymal, endothelial) simultaneously. • Derived cells exhibited intestinal identity with early colonic polarization, confirmed by bulk and single-cell RNA sequencing, and were reproducible across multiple hiPSC lines and independent centers. • Colon-like intestinal patches (CL-IPs) formed on collagen hydrogels and, after transplantation into mice, developed organized tissue structures including crypts, stroma, muscle, and human vasculature that connected to the host circulation. • No evidence of residual pluripotency was observed, supporting the safety of the approach for potential therapeutic applications.