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GASTRIC Publication-derived

Patient-Derived Organoids from Metastatic Gastrointestinal Cancers: Establishment, Characterization, and Drug Response Assessment

Source Vlachogiannis et al. · The Institute of Cancer Research, Royal Marsden Hospital, London

👤 Georgios Vlachogiannis, Somaieh Hedayat, Alexandra Vatsiou, Yann Jamin, Nicola Valeri ⏱ 14 days 📋 5 phases 🧫 Patient-Derived Biopsies from Metastatic GI Cancers (Colorectal, Gastroesophageal, Cholangiocarcinoma)

Abstract

This protocol establishes patient-derived organoids (PDOs) from biopsies of metastatic gastrointestinal cancers and characterizes their genomic and transcriptomic profiles relative to parental tumors. PDOs are cultured in three-dimensional matrigel and assessed for ex vivo responses to a library of 55 clinically relevant anticancer compounds, with results directly compared to patient clinical outcomes to validate PDOs as a co-clinical trial platform for personalized medicine and drug discovery.

Cell source
Patient-Derived Biopsies from Metastatic GI Cancers (Colorectal, Gastroesophageal, Cholangiocarcinoma)
Application
Disease modeling and Drug screening

Protocol overview

33 steps across 5 phases

PDO Establishment from Patient Biopsies Day 0–3
  1. 1 Biopsy Collection and Transport
  2. 2 Tissue Mincing and Conditioning
  3. 3 Enzymatic Digestion
  4. 4 Mechanical Cell Release
  5. 5 Cell Collection and Centrifugation
  6. 6 Matrigel Suspension and Plating
  7. 7 Matrigel Solidification
  8. 8 Overlay with Complete Organoid Media
  9. 9 Media Refresh Schedule
PDO Passaging and Biobanking Day 7–14 (and ongoing)
  1. 1 Mechanical Harvesting
  2. 2 TrypLe Digestion
  3. 3 Mechanical Dissociation to Single Cells
  4. 4 Cell Washing and Centrifugation
  5. 5 Resuspension in Matrigel and Reseeding
  6. 6 Matrigel Solidification and Media Overlay (as in Phase 1, Steps 7–8)
  7. 7 Biobanking (Cryopreservation)
3D PDO Drug Assay Setup Day 0–14 (from cell dissociation)
  1. 1 PDO Harvesting and Dissociation
  2. 2 Cell Counting
  3. 3 Cell Dilution in Matrigel
  4. 4 Seeding in 96-Well Plates
  5. 5 Matrigel Solidification
  6. 6 Media Overlay
  7. 7 First Media Refresh (24 hours post-seeding)
Drug Addition and Treatment (3D Assay or Screen) Day 3–10
  1. 1 Drug Media Preparation (Day 3)
  2. 2 First Media/Drug Addition
  3. 3 Subsequent Drug-Containing Media Refreshes (Days 5, 7, 9 or 10)
  4. 4 Viability Assay Preparation (End of Treatment, typically Day 9–10)
  5. 5 Viability Reading
3D PDO Drug Screening (Automated Compound Library Approach) Day 0–14
  1. 1 Assay Plate Preparation Using ECHO Liquid Dispenser
  2. 2 Screen Execution (Parallel to Drug Assay Setup, Phase 3)
  3. 3 Compound Resuspension and Media Addition (Day 3)
  4. 4 Subsequent Media Refreshes and Viability Assay (Days 5–10)
  5. 5 Data Analysis and Hit Validation

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.

This wording is awaiting legal review.

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