Intraperitoneal and Subcutaneous Tumor Models for Assessing Anti‐Neoplastic Agents in Rodents

Melinda Hollingshead1

1 National Cancer Institute, Frederick, Maryland
Publication Name:  Current Protocols in Pharmacology
Unit Number:  Unit 5.28
DOI:  10.1002/0471141755.ph0528s18
Online Posting Date:  November, 2002
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Abstract

Evaluation of compounds for in vivo antineoplastic activity can be achieved in a variety of animal models. Two models commonly employed for these studies are the intraperitoneal challenge survival model and the subcutaneous tumor implant model. The challenge survival model involves intraperitoneal tumor inoculation, test compound treatment and monitoring for survival. This assay can be modulated for stringency by altering the treatment dose, route and schedule. The second model, subcutaneous tumor implants, is more strenuous as the test compound must cross physiologic barriers to reach the target tumor. Additionally, the subcutaneous model is more labor intensive to setup, monitor and evaluate.

     
 
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Table of Contents

  • Basic Protocol 1: Intraperitoneal Tumor Implantation
  • Basic Protocol 2: Subcutaneous Tumor Implantation
  • Support Protocol 1: Preparation of Drug Solution
  • Support Protocol 2: Tumor Staging
  • Support Protocol 3: Subcutaneous Tumor Growth Calculations
  • Commentary
  • Literature Cited
  • Figures
  • Tables
     
 
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Materials

Basic Protocol 1: Intraperitoneal Tumor Implantation

  Materials
  • HL‐60 leukemia cells (DCTD Tumor Repository; http://www.dtp.nci.nih.gov)
  • RPMI‐1640 medium with 2 mM glutamine
  • Fetal bovine serum (FBS)
  • 0.4% trypan blue solution, tissue‐culture grade
  • SCID female mice, 6 weeks old (Charles River Laboratories or Taconic Animals)
  • Test compound (see protocol 3)
  • 75‐, 162‐, or 175‐cm2 tissue‐culture flasks
  • 50‐ml centrifuge tubes
  • Hemacytometer
  • 1‐ml syringes with 25‐G needles
NOTE: All reagents and equipment coming into contact with live cells must be sterile and sterile technique must be employed throughout the cell preparation procedure.NOTE: SCID mice are immunocompromised and must be housed and maintained under barrier conditions to prevent infection.NOTE: All cell culture incubations are performed in a humidified 37°C, 5% CO 2 incubator.

Basic Protocol 2: Subcutaneous Tumor Implantation

  Materials
  • LOX‐IMVI human melanoma cells (DCTD Tumor Repository; http://www.dtp.nci.nih.gov)
  • RPMI‐1640 medium containing 2 mM glutamine
  • Fetal bovine serum (FBS)
  • Endotoxin‐free 0.9% saline (Quality Biological), sterile
  • 0.05% trypsin/0.53 mM sodium EDTA solution
  • Athymic nude female mice, 6 weeks old (Charles River Laboratories or Taconic Animals)
  • Test compound (see protocol 3)
  • 162‐cm2 tissue‐culture flasks
  • 50‐ml centrifuge tubes
  • Hemacytometer
  • 1‐ml syringes with 25‐G needles
NOTE: All reagents and equipment contacting live cells must be sterile and sterile technique must be used appropriately.NOTE: Athymic nude mice are immunocompromised and must be housed and maintained under barrier conditions to prevent infection.NOTE: All cell culture incubations are performed in a humidified 37°C, 5% CO 2 incubator.

Support Protocol 1: Preparation of Drug Solution

  Materials
  • Test compound
  • 100% dimethyl sulfoxide (DMSO; Burdick and Jackson)
  • Endotoxin‐free saline, sterile
  • Polyoxyethylenesorbitan monooleate (Tween 80)
  • Analytical balance
  • 2‐, 5‐, 10‐, or 30‐ml Wheaton serum vials, amber
  • Vial stoppers
  • Parafilm
  • 37°C water bath

Support Protocol 2: Tumor Staging

  Materials
  • Tumor‐bearing mice
  • Millimeter calipers
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Figures

Videos

Literature Cited

Literature Cited
   Andriole, G.L., Mule, J.J., Hansen, C.T., Linehan, W.M., and Rosenberg, S.A. 1985. Evidence that lymphokine‐activated killer cells and natural killer cells are distinct based on an analysis of congenitally immunodeficient mice. J. Immunol. 135:2911‐2913.
   Auerbach, R. and Auerbach, W. 1981. Regional differences in the growth of normal and neoplastic cells. Science 215:127‐134.
   Dorshkind, E., Keller, G.M., Phillips, R.A., Miller, R.G., Bosma, G.C., O'Toole, M., and Bosma, M.J. 1984. Functional status of cells from lymphoid and myeloid tissues in mice with severe combined immunodeficiency disease. J. Immunol 132:1804‐1808.
   Goldin, A. and Wolpert‐Defilippes, M.K. 1979. Nude mouse models as predictors of chemotherapy in man: Thymidine and pyrimidines. Bull Cancer 66:61‐66.
   Iversen, P.O., Lewis, I.D., Turczynowicz, S., Hasle, H., Niemeyer, C., Schmiegelow, K., Bastiras, S., Biondi, A., Hughes, T.P., and Lopez, A.F. 1997. Inhibition of granulocyte‐macrophage colony‐stimulating factor prevents dissemination and induces remission of juvenile myelomonocytic leukemia in engrafted immunodeficient mice. Blood 90:4910‐4917.
   Mosier, D.E., Stell, K.L., Gulizia, R.J., Torbett, B.E., and Gilmore, G.L. 1993. Homozygous scid/scid; beige/beige mice have low levels of spontaneous or neonatal T cell–induced B cell generation. J. Exp. Med. 177:191‐194.
   Plowman, J., Dykes, D.J., Hollingshead, M., Simpson‐Herren, L., and Alley, M.C. 1997. Human tumor xenograft models in NCI drug development. In Anticancer Drug Development Guide Preclinical Screening, Clinical Trials, and Approval (B.A. Teicher, ed.) pp. 101‐125. Humana Press, Totowa, New Jersey.
   Tomayko, M. and Reynolds, C.P. 1989. Determination of subcutaneous tumor size in athymic (nude) mice. Cancer Chemother. Pharmacol. 24:148‐154.
   Venditti, J.M. 1971. Treatment schedule dependency of experimentally active antileukemic (L1210) drugs. Cancer Chemother. Reports Part 3 2:35‐59.
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