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INTESTINAL Publication-derived

Patient-Derived CRC Organoids Predict Drug Response

Source Vlachogiannis et al. · The Institute of Cancer Research and Royal Marsden NHS Trust, London, UK · 10.1126/science.aao2774

👤 Vlachogiannis G, Hedayat S, Vatsiou A, Jamin Y, Fernandez-Mateos J, Khan K, Lampis A, Ber K, Romanber J, Pietrber P, Bber JD, Sadanandam A, Cunningham D, Valeri N ⏱ 70 days 📋 10 phases 🧫 Patient-Derived (Metastatic CRC Biopsies from Clinical Trials)

Abstract

First prospective clinical study demonstrating that patient-derived tumor organoids from metastatic GI cancer biopsies can predict clinical drug response. Organoid sensitivity testing achieved 100% sensitivity and 93% specificity compared to actual patient outcomes, establishing organoids as viable tools for precision oncology.

Cell source
Patient-Derived (Metastatic CRC Biopsies from Clinical Trials)
Application
Precision Oncology

Protocol overview

43 steps across 10 phases

Tumor Tissue Acquisition and Processing Day 0
  1. 1 Patient Enrollment and Biopsy Collection
  2. 2 Histological Assessment and Sample Preparation
Patient-Derived Organoid (PDO) Establishment Day 1–14
  1. 1 Tissue Dissociation and Cell Isolation
  2. 2 Seeding in 3D Culture System
  3. 3 PDO Culture Maintenance
  4. 4 Passaging and Expansion
Histopathological and Immunohistochemical Characterization Day 7–21
  1. 1 Specimen Preparation and H&E Staining
  2. 2 Immunohistochemistry (IHC) for Diagnostic Markers
  3. 3 Chromogenic In Situ Hybridization (CISH) for Gene Amplifications
  4. 4 Immunostaining for Biomarker Expression
Molecular Profiling (NGS and Gene Expression) Day 7–28
  1. 1 DNA Extraction and Targeted NGS Panel
  2. 2 Mutational Analysis and Copy Number Variation (CNA) Detection
  3. 3 Whole-Genome Sequencing (WGS) Validation
  4. 4 Digital-Droplet PCR (ddPCR) Validation of Key CNAs
  5. 5 RNA Extraction and Gene Expression Profiling (RNA-Seq)
High-Throughput 3D Drug Screening Day 21–35
  1. 1 Organoid Preparation for Drug Screening
  2. 2 Seeding in 3D Drug Screening Format
  3. 3 Drug Library Preparation
  4. 4 Drug Dosing and Treatment
  5. 5 Cell Viability Assessment (GI50 Determination)
  6. 6 Hit Validation at Lower Drug Concentrations
Signaling Pathway Analysis and Mechanism of Action Day 7–14
  1. 1 Organoid Treatment with Pathway Inhibitors
  2. 2 Protein Extraction from Treated Organoids
  3. 3 Western Blot Analysis of Pathway Activation
  4. 4 Cell Cycle and Apoptosis Assessment
  5. 5 DNA Damage Assessment
Establishment of PDO-Derived Orthotopic Xenografts (PDO-Xenografts) Day 28–35
  1. 1 PDO Preparation for Xenograft Implantation
  2. 2 Mouse Strain Selection and Xenograft Site
  3. 3 Surgical Implantation
  4. 4 Tumor Engraftment Monitoring
PDO-Xenograft In Vivo Drug Response Studies Day 42–70
  1. 1 Randomization and Treatment Initiation
  2. 2 Serial Tumor Monitoring During Treatment
  3. 3 Functional Imaging (DCE-MRI and Susceptibility-Contrast MRI)
  4. 4 Survival Monitoring (Long-Term Studies)
Histopathological Analysis of PDO-Xenografts Day 70 (at endpoint)
  1. 1 Tumor Harvest and Fixation
  2. 2 H&E Staining and Morphological Assessment
  3. 3 Immunohistochemical Assessment of Vasculature
  4. 4 Assessment of Histopathological Growth Patterns (HGPs)
  5. 5 Necrosis Quantification
Correlation of PDO and Clinical Response Data Day 70+
  1. 1 Compilation of Clinical Response Data
  2. 2 Matching PDO/Xenograft Responses to Patient Clinical Responses
  3. 3 Sensitivity, Specificity, and Predictive Value Calculation
  4. 4 Mechanistic Investigation of Discordant Cases

Full SOP

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Attribution

This SOP was authored by Organthis based on the published method in Vlachogiannis et al.. The originating laboratory holds no rights in this SOP and has not endorsed it unless marked Verified.

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