Primary Human Non‐Small Cell Lung and Pancreatic Tumorgraft Models—Utility and Applications in Drug Discovery and Tumor Biology

Antonio Calles1, Belén Rubio‐Viqueira1, Manuel Hidalgo1

1 Clinical Research Programme, Spanish National Cancer Research Centre‐CNIO, Madrid, Spain
Publication Name:  Current Protocols in Pharmacology
Unit Number:  Unit 14.26
DOI:  10.1002/0471141755.ph1426s61
Online Posting Date:  June, 2013
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The number of therapeutic options for lung and pancreatic cancer is increasing because of the identification of new druggable molecular targets and development of new drug combinations. Reproducible, biologically relevant in vivo pre‐clinical models are critical for this effort. The generation of patient‐derived tumor xenografts has proven useful for integrating drug screening with biomarker discovery, discovering fundamental information in tumor biology, prioritizing drugs for clinical investigation, and personalizing treatments for these tumors. The protocol described in this unit details how to establish a direct in vivo subcutaneous primary tumorgraft and maintenance passages. The predictive value of a tumorgraft platform to guide personalized medicine is illustrated with the case of a patient with refractory advanced non‐small cell lung cancer (NSCLC). The outcome of a patient for whom their own pancreatic tumorgraft revealed a remarkable sensitivity to mitomycin C based on a PALB2 mutation is also detailed. Curr. Protoc. Pharmacol. 61:14.26.1‐14.26.21 © 2013 by John Wiley & Sons, Inc.

Keywords: lung cancer; pancreatic cancer; xenograft; tumorgraft; animal model; drug development

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Table of Contents

  • Introduction
  • Basic Protocol 1: Generation and Characterization of Primary Human Tumorgrafts of Lung and Pancreatic Carcinomas
  • Commentary
  • Literature Cited
  • Figures
  • Tables
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Basic Protocol 1: Generation and Characterization of Primary Human Tumorgrafts of Lung and Pancreatic Carcinomas

  • Human‐derived tumors (first implantation)/bank‐cryopreserved human tumorgrafts
  • 20% RPMI 1640 medium: RPMI 1640 medium (Gibco, cat. no. 11875‐093), 20% FBS (Gemini, cat. no. 100‐06) and 1% penicillin/streptomycin (Gibco, cat. no. 15140‐122), cold
  • 20% bleach spray bottle
  • 70% ethanol spray bottle
  • Matrigel basement membrane matrix, phenol‐red‐free (BD Biosciences, cat. no. 356237)
  • 4‐ to 6‐week‐old female athymic (nu/nu) mice or NOD‐SCID (Harlan Laboratories)
  • Isoflurane (forane liquid for inhalation, 100 ml; Baxter, cat. no. 1001936040)
  • Buprenorphine (Buprenex, 0.3 mg/ml; Reckitt Benckiser Healthcare, cat. no. NDC 12496‐0757‐1)
  • Individually wrapped sterile alcohol wipes
  • Freezing medium: RPMI 1640/20% FBS/10% DMSO (Sigma, cat. no. D5879), chilled
  • 15‐ and 50‐ml conical tubes (Falcon)
  • Personal protective equipment: sterile surgical gloves, sterile gown, face shield/mask, safety glasses, goggles, shoe covers
  • Biological safety level 2 cabinet
  • Sterile 60‐mm Petri dishes (BD Biosciences, cat. no. 351007)
  • Sterile surgical instruments:
    • 4‐in. microsurgery scissors, sharp‐edged
    • 4.5‐in. Iris scissors, straight
    • 4.5‐in. curved forceps, serrated
    • 4‐in. straight forceps, serrated
    • No. 10 scalpel handle
    • Individually‐wrapped sterile no. 10 scalpel blades
    • 9‐mm metallic staples, vet stapler, and staple remover
  • 6‐well tissue culture plates (BD Biosciences, cat. no. 353046)
  • Sterile 2‐ml microcentrifuge tubes
  • Mobile anesthesia unit composed of vaporizer and absorber for isoflurane, oxygen flask, and gas flow‐meter for oxygen; equipped with induction chamber for mice and face mask for maintenance of anesthesia (Parkland Scientific, cat. nos. V3000PK, 598254, COAX‐3T)
  • Individually wrapped sterile gauze pads
  • Instruments for mouse identification (ear punch tag, tattooing equipment, or microchip equipment)
  • Digital scale
  • Digital caliper
  • Equipment for euthanasia by CO 2 asphyxiation
  • 2‐ml cryovials, chilled
  • Isopropyl alcohol slow‐freezing container (freezing chamber) (Nalgene, cat. no. 5100‐0001)
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