Colorectal cancer cell line-derived organoid model with stem cell properties captures the regrowing state of residual cancer cells after neoadjuvant chemotherapy.

The effectiveness of colorectal cancer (CRC) therapy is limited owing to the absence of treatments targeting drug-tolerant residual cancer cells. Although neoadjuvant therapy is effective, pathological examination of residual tumors has revealed the presence of small clusters of LGR5-positive cancer cells in the fibrous tissue. Here, we established a colorectal cancer cell line-derived organoid (CCD-organoid) regrowth model using a patient-derived cell line with cancer stem cell properties and demonstrated that it displayed the morphological characteristics of small clusters in clinical tissues. Time course analysis of single-cell RNA sequencing of the CCD-organoid regrowth model revealed various states and dynamic alterations within non-cycling cells. We identified subpopulations highly expressing protein translation-related genes RPL17 and EEF1G. To identify key signals for the transition of residual cancer cells to regrowth, we evaluated inhibitors targeting pathways such as the Wnt pathway, reactive oxygen species pathway, and RNA polymerase I pathway, highlighted in the single-cell RNA sequencing analysis. Only the polymerase I-inhibitor BMH-21 significantly reduced tumor growth both in vitro and in vivo, indicating the critical cell subpopulation driving recurrence. Our results demonstrate the possibility of a unique therapeutic target for CRC treatment targeting drug-tolerant residual cancer cells.