Modeling chemotherapy persistence in colorectal cancer using patient-derived organoids

Therapy resistance in colorectal cancer remains a major clinical challenge. Despite standard-of-care chemotherapies such as FOLFOX and FOLFIRI, tumors frequently develop resistance, leading to treatment failure. A growing body of research implicates drug-tolerant persister (DTP) cells-a reversible, slow-growing population that survives initial therapy-as a critical step in resistance development. These persister cells create a window for cancer to acquire additional adaptations-such as mutations and alternative signaling pathway activation-ultimately leading to irreversible resistance. Understanding this transition from persistence to full resistance is essential for developing therapies that can prevent relapse. To investigate these mechanisms in a clinically relevant context, we developed patient-derived organoid (PDO) models of colorectal cancer capable of surviving FOLFOX or FOLFIRI treatment. These organoids retain the heterogeneity and architecture of the original tumors, providing a physiologically faithful system to study persistence. Because chemotherapy impact varies by mechanism of action, standard assays may not fully capture response diversity. We therefore optimized viability readouts to measure treatment effects across the entire organoid population, capturing both cytotoxicity and partial survival. Using these persistent PDOs, we performed whole-exome sequencing alongside bulk and single-cell RNA sequencing to map the molecular features associated with the DTP state and subsequent resistance. This approach revealed pathways and cellular programs that enable tumor cells to tolerate chemotherapy, highlighting potential targets for overcoming resistance. By modeling clinically relevant persistence, these PDOs offer a platform to test interventions that eliminate persister populations before they evolve into fully resistant tumors.