Orthotopic Models of Human Gastric Carcinoma in Nude Mice: Applications for Study of Tumor Growth and Progression

Susan Jones‐Bolin1, Bruce Ruggeri1

1 Cephalon, Inc., West Chester, Pennsylvania
Publication Name:  Current Protocols in Pharmacology
Unit Number:  Unit 14.4
DOI:  10.1002/0471141755.ph1404s37
Online Posting Date:  June, 2007
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Abstract

Gastric carcinoma (GC) remains a leading cause of cancer related deaths worldwide with the 5‐year survival rate in the U.S. at ∼5% to 15% with existing therapies. This tumor is aggressive and has often metastasized to distant sites (liver, lung, and adjacent intestine) by the time of diagnosis. Treatment options (surgery, radiation, and chemotherapy) are limited and the disease carries a grave prognosis for most patients (50% 5‐year survival for distal GC; 10% 5‐year survival for proximal GC). An orthotopic model of human GC in nude mice provides an excellent way to evaluate the pathogenesis of tumor growth and metastasis in order to develop effective therapies, as well as to better understand the underlying biology of gastric tumor growth and metastasis. The protocol described in this unit details the development and characterization of an orthotopic model of human GC in athymic nude mice with diffuse lymphatic and hepatic metastatic spread. This model closely mimics the course of the human disease.

Keywords: Human gastric carcinoma; Athymic nude mice; Orthotopic; Metastasis

     
 
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Table of Contents

  • Basic Protocol 1: Orthotopic Model of Human Gastric Carcinoma in Athymic Nude Mice
  • Basic Protocol 2: Pharmacological Efficacy Studies Using the Athymic Nude Mouse Orthotopic Gastric Carcinoma Model
  • Alternate Protocol 1: Establishing Cell Lines from Gastric Carcinoma Tumor Tissue
  • Reagents and Solutions
  • Commentary
  • Literature Cited
  • Figures
  • Tables
     
 
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Materials

Basic Protocol 1: Orthotopic Model of Human Gastric Carcinoma in Athymic Nude Mice

  Materials
  • GTL‐16 human gastric carcinoma cell line or human gastric carcinoma cell lines with comparable phenotype (such as NCI‐N87, ATCC)
  • Gastric carcinoma growth medium (see recipe)
  • 1× trypsin/EDTA (Mediatech)
  • 1× PBS
  • Nude mice, female, 6‐ to 8‐weeks‐old (20 to 25 g; athymic nu/nu or comparable strain; Charles River Labs)
  • Matrigel synthetic basement membrane (Collaborative Research)
  • Ketamine/xylazine mixture
  • Isoflurane with nose cone apparatus
  • Tissue culture flasks
  • 1‐ml syringes and 27‐G needles
  • 50‐ml conical centrifuge tubes
  • Hemacytometer or a commercial cell counter
  • 70‐mm (or larger) petri dish, sterile
  • Surgical instruments including (two sets of instruments recommended):
    • 4.5‐in. iris scissors, straight, sharp ends
    • Disposable scalpels
    • 4.75‐in. Adson forceps or delicate dressing forceps, serrated
    • 4.5‐in. tissue forceps, 1 × 2 teeth
    • 5.5‐in. Mayo‐Hegar or similar needle holder
    • Suture: prolene 6/0, taper needle, for suturing tissue to stomach; one pack per 5 to 10 mice (Webster Veterinary Supply)
    • Vicryl 5/0 or 6/0 –or‐ PDS II 5/0, for closing peritoneal layer; one pack per 5 to 10 mice (Webster Veterinary Supply)
    • 9‐mm wound clips, with applier and remover (VWR)
  • Autoclave for sterilizing instruments, gauze sponges, and wound clips
  • Circulating water heating pad: Gaymar T‐pump or similar
  • Sterile towels
  • Distilled water to fill pump for heating pads
  • Sterile alcohol prep pads, medium
  • Sterile gauze sponges (2‐in. × 2‐in.)
  • Sharpie marker, ear punch, or tattooing device to identify individual mice
  • Additional reagents and equipment for euthanizing using CO 2 (Donovan and Brown, )

Basic Protocol 2: Pharmacological Efficacy Studies Using the Athymic Nude Mouse Orthotopic Gastric Carcinoma Model

  Materials
  • Nude mice exhibiting appropriate metastatic phenotype ( protocol 1)
  • Additional reagents and equipment for nude mice exhibiting appropriate metastatic phenotype ( protocol 1)

Alternate Protocol 1: Establishing Cell Lines from Gastric Carcinoma Tumor Tissue

  • Mice with tumors (from protocol 1)
  • DMEM containing 10% (v/v) FBS, ice‐cold
  • Gastric carcinoma growth medium (see recipe)
  • 10% (v/v) DMSO/90% (v/v) FBS
  • 70‐µm nylon cell strainer (BD Biosciences)
  • Plunger from 1‐ml syringe
  • 75‐cm2 tissue culture flask
  • Humidified 5% CO 2, 37°C incubator
  • 1.5‐ml cryotubes (e.g., Nunc)
  • Nalgene Cryo 1°C Freezing Container (Nalge Nunc International)
  • −80°C freezer
  • Liquid nitrogen freezer
  • 30‐G needle
  • Additional reagents and equipment for developing mice with tumors ( protocol 1)
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Figures

Videos

Literature Cited

Literature Cited
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