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An orthotopic xenograft model with survival hindlimb amputation allows investigation of the effect of tumor microenvironment on sarcoma metastasis

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Abstract

Overall survival rates for pediatric high-grade sarcoma have improved greatly in the past few decades, but prevention and treatment of distant metastasis remain the most compelling problems facing these patients. Traditional preclinical mouse models have not proven adequate to study the biology and treatment of spontaneous distant sarcoma metastasis. To address this deficit, we developed an orthotopic implantation/amputation model in which patient-derived sarcoma xenografts are surgically implanted into mouse hindlimbs, allowed to grow, then subsequently amputated and the animals observed for development of metastases. NOD/SCID/IL-2Rγ-null mice were implanted with either histologically intact high grade sarcoma patient-derived xenografts or cell lines in the pretibial space and affected limbs were amputated after tumor growth. In contrast to subcutaneous flank tumors, we were able to consistently detect spontaneous distant spread of the tumors using our model. Metastases were seen in 27–90 % of animals, depending on the xenograft, and were repeatable and predictable. We also demonstrate the utility of this model for studying the biology of metastasis and present preliminary new insights suggesting the role of arginine metabolism and macrophage phenotype polarization in creating a tumor microenvironment that facilitates metastasis. Subcutaneous tumors express more arginase than inducible nitric oxide synthase and demonstrate significant macrophage infiltration, whereas orthotopic tumors express similar amounts of inducible nitric oxide synthase and arginase and have only a scant macrophage infiltrate. Thus, we present a model of spontaneous distant sarcoma metastasis that mimics the clinical situation and is amenable to studying the biology of the entire metastatic cascade.

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Abbreviations

NSG:

NOD/SCID/IL-2Rγ-null

PBS:

Phosphate-buffered saline

BSA:

Bovine serum albumin

iNOS:

Inducible nitric oxide synthase

TBST:

1×TBS and 0.05 % Tween 20

DAB:

3,39-diaminobenzidine

NO:

Nitric oxide

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Acknowledgments

This work was supported by grants from the National Institutes of Health (1R01CA138212-01) and the Liddy Shriver Sarcoma Initiative (to DML), as well as from the Pablove Foundation (to MH). The authors also wish to acknowledge the support of the Giant Food Children’s Cancer Research Fund, the Heather Brooke Foundation, and the Love for Luca Foundation.

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Authors and Affiliations

  1. Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University, Bunting-Blaustein Cancer Research Building, Room 2M51, 1650 Orleans St., Baltimore, MD, 21231, USA

    Seth D. Goldstein, Masanori Hayashi, Catherine M. Albert, Kyle W. Jackson & David M. Loeb

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  1. Seth D. Goldstein
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  2. Masanori Hayashi
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  3. Catherine M. Albert
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Correspondence to David M. Loeb.

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The authors declare that they have no actual or potential conflicts of interest.

Research involving human and animal rights

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. All procedures performed in studies involving animals were in accordance with the ethical standard of the institution at which the studies were conducted. This article does not contain any studies with human participants performed by any of the authors.

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Goldstein, S.D., Hayashi, M., Albert, C.M. et al. An orthotopic xenograft model with survival hindlimb amputation allows investigation of the effect of tumor microenvironment on sarcoma metastasis. Clin Exp Metastasis 32, 703–715 (2015). https://doi.org/10.1007/s10585-015-9738-x

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  • DOI: https://doi.org/10.1007/s10585-015-9738-x

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