Table 2.

Summary characteristics of included publications (n = 21) and clinical studies (n = 5).

Reference or NCT number	Study type	Source & type of experimental model	Limitions	Main findings
(42)	Analysis of the mutational landscape using high-throughput sequencing technologies	Excised from mice with RC patient tumor xenograft Ex vivo	• Lack of data from post treatment human rectal cancer specimens. • Did not show accordance exists between organoid and biopsy data • Small sample size	Assessed for ST6GAL-1 protein with and without chemoradiation treatment on patient-derived xenograft and organoid models and identified ST6GAL-1 protein as a mediator for resistance to clinical chemoradiation therapy through restraining apoptosis.
(43)	Personalized medicine based on the testing of individual PDOs; Studying the tumor microenvironment with PDOs; Cancer modeling by genetic engineering of organoids	Biopsies from pre-CRT tumor and normal In vivo and ex vivo	• Lack of data from post treatment human rectal cancer specimens. • Did not mention about the type and number of cells seeded for organoid culture. • Lacked the success rate and cell composition of the established organoid.	Developed RC PDOs and primary stroma cells and identified that interleukin-1α (IL-1α) after irradiation polarizes cancer-associated fibroblasts toward the inflammatory phenotype together with triggering oxidative DNA damage; Displayed the impact factor in chemoradiotherapy resistance and disease progression.
(44)	Reviewing biomarkers and models used in RC	–	–	Reviewed published findings associated with biomarkers discovery and pre-clinical models (included RC PDOs) in RC.
(45)	Personalized medicine based on the testing of individual PDOs	Surgically or endoscopically resected tumor tissues of patients undergoing neoadjuvant therapy Ex vivo	• Organoid culture lacked microenvironmental regulation of tumor response. • Did not mention about the type and number of cells seeded for organoid culture. • Lacked the success rate and cell composition of the established organoid.	Analyzed radiosensitivity of PDOs and provided a readout predictive of neoadjuvant therapy for selecting patients who need pre-treatment.
(46)	Reviewing PDOs models for precision medicine	–	–	Evaluated the potential of PDO models (included RC PDOs and distinguished RC research) in predictive translational research.
(47)	Conducting clinical trial for translational research from bench to bedside; Personalized medicine based on the testing of individual PDOs	–	–	Started ACO/ARO/AIO-21 phase I trial to test the IL-1 receptor antagonist (IL-1 RA) anakinra combining with CRT therapy for RC based on previous achievement (43), which set up a great example for translational application from bench to bedside.
(38)	Personalized medicine based on the testing of individual PDOs	0.5×0.5×0.5 cm for surgically resected specimens and 1.5×0.2×0.2 cm for ultrasound-guided core-needle biopsy tissue Ex vivo	• Results need further validation in the prospective, randomized controlled study. • Organoids culture was in the absence of tumor microenvironment. • Lacked the purity and cell composition report of the established organoid.	The sensitivity, specificity, and accuracy of the RC PDOs for predicting chemotherapy regimens response were 63.33%, 94.12%, and 79.69%.
(48)	Analysis of the mutational landscape using high-throughput sequencing technologies; Personalized medicine based on the testing of individual PDOs	Biopsy samples Ex vivo	• Small sample size • Did not mention about the type and number of cells seeded for organoid culture. • Organoids culture was in the absence of tumor microenvironment. • Did not perform a drug sensitivity test.	Established a prediction model through a machine learning algorithm combining clinical and experimental radio response data; Radiation responses in clinic were positively correlated with the paired cultures.
(41)	Analysis of the mutational landscape using high-throughput sequencing technologies	Colon-endoscopic biopsy from participants accepted preoperative chemoradiotherapy (pCRT) Ex vivo	• Did not show accordance exists between organoid and biopsy data	High expression of VSTM2L reduced γ-H2AX expression in RC PDOs treated with CRT.
(49)	Analysis of the mutational landscape using high-throughput sequencing technologies	Biopsy samples Ex vivo	• Small sample size • Did not mention about the type and number of cells seeded for organoid culture. • Lacked the success rate and cell composition of the established organoid.	Developed RC PDOs to detect genes and pathways that participate in the radio-resistance of LARC by biological and bioinformatic analysis approaches; Identified cathepsin E (CTSE) that was negatively correlated with the radio-resistance in PDOs.
(50)	Reviewing PDOs models for precision medicine	–	–	Described CRT prediction value of organoids (included RC PDOs) for GI cancers.
(51)	Drug screening to develop novel treatment strategies; Personalized medicine based on the testing of individual PDOs	Resected specimens Ex vivo	• Small sample size • Did not mention about the type and number of cells seeded for organoid culture. • Lacked the purity and cell composition report of the established organoid.	Screened PDOs with a customized medium-throughput drug library consist of 33 single agents and three 5-FU-based drug combinations with Leucovorin (FLV), Oxaliplatin (FLOX), and SN-38 (FLIRI).
(52)	Reviewing pre-clinical models used in RC	–	–	Described different pre-clinical model (included PDOs) used in RC research.
(53)	Reviewing biomarkers and models used in RC	–	–	Reviewed published paper associated with potential biomarkers and cell-based models (included RC PDOs) to predict treatment response in RC.
(16)	Analysis of the mutational landscape using high-throughput sequencing technologies; Drug screening to develop novel treatment strategies; Personalized medicine based on the testing of individual PDOs	Tissue biopsies from patients with newly diagnosed LARC who were treatment-naive in a phase III clinical trial NCT02605265 Ex vivo	• Lacked the purity and cell composition report of the established organoid.	Established an organoid biobank with PDOs obtaining similar histological and genetic features of original tumors; identify the role of predicting LARC patient Chemoradiation responses in the clinic.
(54)	Drug screening to develop novel treatment strategies	7 rectal endoscopic biopsy and 1 colon cancer sample from low anterior resection Ex vivo	• Lacked the success rate and cell composition of the established organoid.	Butyrate could enhance the curative effect of radiotherapy while protecting the normal mucosa; Identified FOXO3A as a factor with non-responsive cases to butyrate in PDOs.
(27)	Personalized medicine based on the testing of individual PDOs	Endoscopic biopsies from 26 Stages 2 and 3 rectal cancer patients prior to receiving 5FU/RT In vivo and ex vivo	• Small sample size	Identified the ability of cetuximab to enhance RT effectiveness; Used PDOs to improve patient selection based on mutational profile. Success rate:90%
(55)	Analysis of the mutational landscape using high-throughput sequencing technologies	Endoscopic Biopsies from therapy-naïve rectal cancer patients Ex vivo	• Did not mention about the type and number of cells seeded for organoid culture. • Lacked the success rate and cell composition of the established organoid.	Compared the gene profiling of organoids derived from a normal rectum and rectal tumors and their responses to calcitriol; Identified rectal tumor organoid-specific genes associated with biosynthetic machinery, including those encoding the RNA polymerase II subunits POLR2H and POLR2J.
(56)	Personalized medicine based on the testing of individual PDOs	Did not mention Ex vivo	• Did not mention about the type and number of cells seeded for organoid culture. • Lacked the success rate and cell composition of the established organoid. • Lack details in culture methods • Small sample size	Similarly, two patient-derived organoid models containing relatively low AC expression were found to be comparatively more radiosensitive than three other models containing higher levels of AC.
(39)	Analysis of the mutational landscape using high-throughput sequencing technologies; Drug screening to develop novel treatment strategies; Personalized medicine based on the testing of individual PDOs; Investigation of intratumoral heterogeneity and tumor evolution	Endoscopic biopsies from pre- and post-treatment patient samples In vivo and ex vivo	• Need studies with larger populations to investigate the prediction value. • Did not mention about the type and number of cells seeded for organoid culture.	RC PDO cultures reserved architecture and molecular features of the original tumors and their in vitro responses to clinical treatment correlated with the outcomes of individual patients’ tumors; PDOs from patients with RC under multimodal therapy engraft into the rectal mucosa of mice, which indicating a success in vivo RC PDO model.
(40)	Protocols for RC PDO establishment	Surgery or biopsy Ex vivo	• Lack details in culture methods • Lacked the success rate and cell composition of the established organoid.	Developed protocols for establishing RC cancer organoids; performed high-throughput drug sensitivity testing.
NCT03577808	Personalized medicine based on the testing of individual PDOs;	Pre-treatment biopsies Ex vivo	–	Validation of Organoids Potential Use as a Companion Diagnostic in Predicting Neoadjuvant Chemoradiation Sensitivity in Locally Advanced Rectal Cancer
NCT05352165	Personalized medicine based on the testing of individual PDOs; Drug screening to develop novel treatment strategies;	Not mentioned	–	A Prospective Multicenter Randomized Controlled Trial of the Clinical Efficacy of Neoadjuvant Therapy Based on Organoids Drug Sensitivity Versus Empirical Neoadjuvant Therapy in the Treatment of Advanced Rectal Cancer.
NCT04371198	Determine the feasibility of establishing patient-derived organoids.	Pre-treatment rectal adenocarcinoma biopsies. Ex vivo	–	Accessing the feasibility of the Biospecimen Collection Protocol for establishing Patient-Derived Organoids for Rectal Cancer
NCT05401318	Personalized medicine based on the testing of individual PDOs; Drug screening to develop novel treatment strategies;	Fresh tumor samples from colon and rectal cancer patients Ex vivo	–	Accessing the prediction value of the PDOs and investigating the effect of Pre-treatment with cytotoxic agents which can induce cellular immunotherapy efficacy against solid tumors in PDOs
NCT04842006	Personalized medicine based on the testing of individual PDOs;	Not mentioned Ex vivo	–	Population distribution of PDO treatment response is compared to their corresponding clinical response by response MRI and pathological response.