Establishment of Patient‐Derived Xenograft (PDX) Models of Human Breast Cancer

Xiaomei Zhang1, Michael T. Lewis2

1 Lester and Sue Smith Breast Center, Baylor College of Medicine, Houston, Texas, 2 Department of Molecular and Cellular Biology and Department of Radiology, Baylor College of Medicine, Houston, Texas
Publication Name:  Current Protocols in Mouse Biology
Unit Number:   
DOI:  10.1002/9780470942390.mo120140
Online Posting Date:  March, 2013
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Abstract

Patient‐derived xenograft (PDX) models of human breast cancer are proving useful for preclinical evaluation of experimental therapeutics. However, until recently, generation of PDX models reflecting the full spectrum of human breast cancers has been an elusive goal. We recently developed a method for establishing serially transplantable, phenotypically stable, human breast cancer xenograft models in immunocompromised mice with comparatively high efficiency (overall ∼25%). These xenografts represent the major clinically defined subtypes of breast cancer [e.g. estrogen receptor positive (ER+), HER2 positive (HER2+), and “triple negative” (TN) breast cancers]. This method, and methods being developed in other laboratories, may soon allow for conducting “animal clinical trials” once sufficient numbers of clinically relevant models are generated. Curr. Protoc. Mouse Biol. 3:21‐29 © 2013 by John Wiley & Sons, Inc.

Keywords: SCID/Beige immunocompromised mice; NOD‐SCID IL2Rgammanull immunocompromised mice (NSG); preclinical breast cancer models; transplantation methods; mammary gland

     
 
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Table of Contents

  • Introduction
  • Basic Protocol 1: Transplantation of Patient‐Derived Specimens and Established Xenograft Tissue into Immunodeficient Mice
  • Basic Protocol 2: Harvest and Serial Passage of Xenograft Tumor Tissues
  • Commentary
  • Literature Cited
  • Figures
     
 
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Materials

Basic Protocol 1: Transplantation of Patient‐Derived Specimens and Established Xenograft Tissue into Immunodeficient Mice

  Materials
  • Primary human tumor tissue: Whenever possible, fresh tumor specimens are delivered to our research laboratory within one hour of removal from the patient [if immediate transplantation is not feasible, we store tissue in tissue culture medium (e.g. RPMI medium 1640; Invitrogen Life Technologies, cat. no.11875‐093) temporarily at 4°C for up to 24 hr]
  • Hank′s Balanced Salt Solution (HBSS; Invitrogen Life Technologies, cat. no. 14025092)
  • Isoflurane or injectable solution: e.g., Rodent Combo Anesthetic III (ketamine 37.6 mg/ml; xylazine 1.92 mg/ml; acepromazine 0.38 mg/ml)
  • CB17.Cg‐PrkdcscidLystbg/Crl (SCID/Beige) female mice, 3 weeks old (10 to 15 grams), Charles River Laboratories International or NOD.Cg‐Prkdcscid Il2rgtm1Wjl/SzJ(NOD‐SCID IL2Rgammanull; NSG) female mice, 3 weeks old (10 to 15 grams), The Jackson Laboratory
  • 10% Povidone‐Iodine solution (Butler Schein Animal Health. cat. no. 003660)
  • 70% (v/v) ethanol
  • 17β‐Estradiol pellet, 60‐day release, 0.36 mg/pellet (Innovative research of America, cat. no. SE‐121)
  • 100‐mm petri dishes (BD Biosciences, cat. no. 351005), sterile
  • Sterile stainless steel disposable scalpels, size 10 (EXEL International Medical Products, cat. no. 29550)
  • Micro dissecting scissors 4.5‐in. straight sharp (Roboz, cat. no. RS‐5916)
  • Castroviejo micro dissecting spring scissors (Roboz, cat. no. RS‐5658SC)
  • Mouse ear tags (Fisher Scientific, cat. no. INS1005‐1X)
  • Animal hair clippers
  • Sterile stainless steel reusable precision trochar (Innovative Research of America, cat. no. MP‐182)
  • Micro dissecting forceps, serrated, full curve (Roboz, cat. no. RS‐5138)
  • Micro dissecting forceps, straight, fine sharp tips (Roboz, cat. no. RS‐5090)
  • Wound clip applier, 9 mm (Roboz, cat. no. RS‐9260)
  • Wound clips, 9 mm (Roboz, cat. no. RS‐9265)
  • Wound clip remover (Roboz, cat. no. RS‐9263)
  • Biosafety hood for sterile removal of mammary glands and tumor transplantation
  • Sterile cotton‐tipped applicators (Fisher Scientific, cat. no. 8884541300)
  • Surgical board
  • High temperature fine‐tipped cautery (SELECT Cautery Tips; PSS World Medical, cat. no. 243)
  • Sterile paper towels or gauze pads
  • Clean, autoclaved cages
  • Slide drying bench (bench warmer) (Fisher Scientific, cat. no. 11‐474‐470)

Basic Protocol 2: Harvest and Serial Passage of Xenograft Tumor Tissues

  Materials
  • Tumor‐bearing mice (see protocol 1)
  • 10% buffered formalin
  • Tumor stock medium for fresh tumor transport: RPMI 1640 with 10% heat‐inactivated newborn calf serum (HINCS; Invitrogen Life Technologies, cat. no. 26010074)
  • Liquid nitrogen
  • Freezing medium for long‐term storage of tumor fragments: Dulbecco's modified Eagles Medium (DMEM; Invitrogen Life Technologies, cat. no. 11965‐092) with 10% fetal bovine serum (FBS; Invitrogen Life Technologies, cat. no.12662‐029) and 10% sterile‐filtered dimethyl sulfoxide (DMSO; Sigma‐Aldrich, cat. no. D2438) and 1% penicillin‐streptomycin
  • Hank′s Balanced Salt Solution (HBSS; Invitrogen Life Technologies, cat. no. 14025092)
  • 50‐ml conical tubes
  • Cryovials
  • −80°C freezer
  • 37°C water bath
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Figures

Videos

Literature Cited

Literature Cited
   Clarke, R. 1996. Human breast cancer cell line xenografts as models of breast cancer. The immunobiologies of recipient mice and the characteristics of several tumorigenic cell lines. Breast Cancer Res. Treat. 39:69‐86.
   DeOme, K.B., Faulkin, L.J.J., and Bern, H. 1959. Development of mammary tumors from hyperplastic alveolar nodules transplanted into gland‐free mammary fat pads of female C3H mice. Cancer Res. 19:515‐520.
   Gupta, P.B. and Kuperwasser, C. 2006. Contributions of estrogen to ER‐negative breast tumor growth. J. Steroid Biochem. Mol. Biol. 102:71‐78.
   Gupta, P.B., Proia, D., Cingoz, O., Weremowicz, J., Naber, S.P., Weinberg, R.A., and Kuperwasser, C. 2007. Systemic stromal effects of estrogen promote the growth of estrogen receptor‐negative cancers. Cancer Res. 67:2062‐2071.
   Iyer, V., Klebba, I., McCready, J., Arendt, L.M., Betancur‐Boissel, M., Wu, M.F., Zhang, X., Lewis, M.T., and Kuperwasser, C. 2012. Estrogen promotes ER‐negative tumor growth and angiogenesis through mobilization of bone marrow‐derived monocytes. Cancer Res. 2:2705‐2713.
   Xia, Z., Taylor, P.R., Locklin, R.M., Gordon, S., Cui, Z., and Triffitt, J.T. 2006. Innate immune response to human bone marrow fibroblastic cell implantation in CB17 scid/beige mice. J. Cell Biochem. 98:966‐980.
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