You are already at the latest version Animal models are standard for safety evaluation, yet physiological differences limit human translation. Doxorubicin (DOX), a chemotherapeutic, can cause off-target gastrointestinal (GI) toxicity and treatment discontinuation. This study evaluated human colonoids as a potential human-relevant epithelial model for DOX-induced GI toxicity by comparing transcriptomic responses across human colonoids, mouse colonoids, and male C57BL/6J mouse colon tissue.