Chemosensitivity profiling of colorectal cancer patient-derived organoids reveals heterogeneous responses to 5-fluorouracil

To establish patient-derived organoid (PDO) models from colorectal cancer (CRC) patients and evaluate their fidelity and utility in assessing 5-fluorouracil (5-FU) chemosensitivity.Tumour samples from 30 untreated CRC patients were cultured into PDOs.Successful models were validated by hematoxylin and eosin (HE) and immunohistochemical (IHC) staining for morphology and biomarker expression.RNA sequencing (RNA-seq) was performed to identify transcriptional changes.5-FU sensitivity was tested using an ATP-based viability assay across early and late passages.Twenty-six PDO lines were successfully established (86.67% success rate) and retained the histopathological and molecular features of primary tumours.RNA-seq revealed significant dysregulation of extracellular matrix and immune-related pathways, consistent with the absence of a tumour microenvironment.5-FU sensitivity varied widely among PDOs, with IC 50 values ranging from 0.085 to 3.934 g/ml in first-generation cultures and narrowing to 0.078-2.134g/ml by passage 4, implying enhanced drug efficacy under microenvironment-deficient conditions.CRC PDOs effectively recapitulate tumour characteristics and exhibit heterogeneous 5-FU responses, supporting their potential as personalised drug-testing platforms.However, the absence of tumour microenvironment components may influence sensitivity outcomes and should be considered in future applications.