Gastroenterology

Gastroenterology

Volume 139, Issue 3, September 2010, Pages 1030-1040
Gastroenterology

Basic—Liver, Pancreas, and Biliary Tract
CD39/ENTPD1 Expression by CD4+Foxp3+ Regulatory T Cells Promotes Hepatic Metastatic Tumor Growth in Mice

https://doi.org/10.1053/j.gastro.2010.05.007Get rights and content

Background & Aims

Adenosine mediates immune suppression and is generated by the ectonucleotidases CD39 (ENTPD1) and CD73 that are expressed on vascular endothelial cells and regulatory T cells (Tregs). Although tumor-infiltrating immune cells include Foxp3+ Tregs, it is not clear whether local adenosine generation by Tregs promotes tumor growth in a CD39-dependent manner. In this study, we have examined the effect of CD39 expression by Tregs on effector immune cell responses to hepatic metastases in vivo.

Methods

A model of hepatic metastatic cancer was developed with portal vein infusion of luciferase-expressing melanoma B16/F10 cells and MCA38 colon cancer cells in wild-type (wt) and mutant mice null for Cd39. Chimeric mice were generated by bone marrow transplantation (BMT) using Cd39 null or wt C57BL6 donors and irradiated recipient mice.

Results

We demonstrate that hepatic growth of melanoma metastatic tumors was strongly inhibited in mice with Cd39 null vasculature or in wt mice with circulating Cd39 null bone marrow–derived cells. We show functional CD39 expression on CD4+Foxp3+ Tregs suppressed antitumor immunity mediated by natural killer (NK) cells in vitro and in vivo. Finally, inhibition of CD39 activity by polyoxometalate-1, a pharmacologic inhibitor of nucleoside triphosphate diphosphohydrolase activity, significantly inhibited tumor growth (P < .001).

Conclusions

CD39 expression on Tregs inhibits NK activity and is permissive for metastatic growth. Pharmacologic or targeted inhibition of CD39 enzymatic activity may find utility as an adjunct therapy for secondary hepatic malignancies.

Section snippets

Animals

Six- to 14-week-old male C57BL6 Cd39 null mice were used.8 Age-, sex-, and strain-matched wild-type (wt) mice and (γc)/Rag2−/− mice were purchased from Taconic Farms (Germantown, NY). Rag1−/− mice were from The Jackson Laboratory (Bar Harbor, ME). Foxp3-GFP (green fluorescent protein) knock-in mice were generated as described.13

Animal Experimentation Protocols were reviewed and approved by the Institutional Animal Care and Use Committees of Beth Israel Deaconess Medical Center.

Antibodies and Reagents

All chemicals

CD39 Expression Facilitates Tumor Growth

Portal vein infusion of luciferase-expressing melanoma B16/F10 (luc-B16/F10) cells and MCA38 colon cancer cells resulted in cells being retained in liver sinusoids; no tumors were found in lungs or other sites within 14 days (data not shown). Melanoma-bearing Cd39 null mice had longer times to euthanasia and death than the wt mice (P < .0001; Figure 1A).

Tumor growth in liver was substantially inhibited in Cd39 null mice (melanoma data in Figure 1B and C and colonic tumor studies in Figure 2A

Discussion

We have already shown that CD39 expression on endothelium is important for angiogenesis and is required for tumor growth in the lung.11, 18 We show now that infiltrating Tregs inhibit NK cell–mediated tumor cytotoxicity in a CD39-dependent manner in the liver. Tregs exhibit immunosuppressive effects on effector T cells, operational by adenosine-mediated pathways modulated by CD39 and CD73.9 Our current findings strengthen the notion that the expression of CD39 and associated changes in

Acknowledgments

The authors thank members of the Longwood Small Animal Imaging Facility of BIDMC for assistance with the in vivo bioluminescence imaging.

X.S. and Y.W. contributed equally to this work, and the surnames are arranged alphabetically.

References (55)

  • M. Mandapathil et al.

    Generation and accumulation of immunosuppressive adenosine by human CD4+CD25highFOXP3+ regulatory T cells

    J Biol Chem

    (2010)
  • T. Ogawa et al.

    Anti-tumor angiogenesis therapy using soluble receptors: enhanced inhibition of tumor growth when soluble fibroblast growth factor receptor-1 is used with soluble vascular endothelial growth factor receptor

    Cancer Gene Ther

    (2002)
  • R.S. Chari et al.

    Chemotherapy and regional therapy of hepatic colorectal metastases: expert consensus statement by Bartlett et al

    Ann Surg Oncol

    (2006)
  • D.W. Hoskin et al.

    Adenosine as a possible inhibitor of killer T-cell activation in the microenvironment of solid tumours

    Int J Cancer

    (1994)
  • G. Burnstock

    Purinergic signaling and vascular cell proliferation and death

    Arterioscler Thromb Vasc Biol

    (2002)
  • A. Ohta et al.

    A2A adenosine receptor protects tumors from antitumor T cells

    Proc Natl Acad Sci U S A

    (2006)
  • K. Enjyoji et al.

    Targeted disruption of cd39/ATP diphosphohydrolase results in disordered hemostasis and thromboregulation

    Nat Med

    (1999)
  • S. Deaglio et al.

    Adenosine generation catalyzed by CD39 and CD73 expressed on regulatory T cells mediates immune suppression

    J Exp Med

    (2007)
  • E. Bettelli et al.

    Reciprocal developmental pathways for the generation of pathogenic effector TH17 and regulatory T cells

    Nature

    (2006)
  • A. Sato et al.

    Antitumor activity of IFN-lambda in murine tumor models

    J Immunol

    (2006)
  • X. Huang et al.

    Combined therapy of local and metastatic 4T1 breast tumor in mice using SU6668, an inhibitor of angiogenic receptor tyrosine kinases, and the immunostimulator B7.2-IgG fusion protein

    Cancer Res

    (2002)
  • Y.J. Day et al.

    Renal protection from ischemia mediated by A2A adenosine receptors on bone marrow-derived cells

    J Clin Invest

    (2003)
  • C. Goepfert et al.

    Disordered cellular migration and angiogenesis in cd39-null mice

    Circulation

    (2001)
  • G. Beldi et al.

    Natural killer T cell dysfunction in CD39-null mice protects against concanavalin A-induced hepatitis

    Hepatology

    (2008)
  • D.I. Gabrilovich et al.

    Myeloid-derived suppressor cells as regulators of the immune system

    Nat Rev Immunol

    (2009)
  • J.D. Fontenot et al.

    Foxp3 programs the development and function of CD4+CD25+ regulatory T cells

    Nat Immunol

    (2003)
  • M.J. Smyth et al.

    CD4+CD25+ T regulatory cells suppress NK cell-mediated immunotherapy of cancer

    J Immunol

    (2006)
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    Conflicts of interest The authors disclose no conflicts.

    Funding This study has been supported by the National Institutes of Health (NHLBI PO1-HL076540 and RO1-HL094400).

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