NK cell heparanase controls tumor invasion and immune surveillance
Eva M. Putz, … , Mark D. Hulett, Mark J. Smyth
Eva M. Putz, … , Mark D. Hulett, Mark J. Smyth
Published June 5, 2017
Citation Information: J Clin Invest. 2017;127(7):2777-2788. https://doi.org/10.1172/JCI92958.
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Research Article Immunology Oncology

NK cell heparanase controls tumor invasion and immune surveillance

Abstract

NK cells are highly efficient at preventing cancer metastasis but are infrequently found in the core of primary tumors. Here, have we demonstrated that freshly isolated mouse and human NK cells express low levels of the endo-β-D-glucuronidase heparanase that increase upon NK cell activation. Heparanase deficiency did not affect development, differentiation, or tissue localization of NK cells under steady-state conditions. However, mice lacking heparanase specifically in NK cells (Hpsefl/fl NKp46-iCre mice) were highly tumor prone when challenged with the carcinogen methylcholanthrene (MCA). Hpsefl/fl NKp46-iCre mice were also more susceptible to tumor growth than were their littermate controls when challenged with the established mouse lymphoma cell line RMA-S-RAE-1β, which overexpresses the NK cell group 2D (NKG2D) ligand RAE-1β, or when inoculated with metastatic melanoma, prostate carcinoma, or mammary carcinoma cell lines. NK cell invasion of primary tumors and recruitment to the site of metastasis were strictly dependent on the presence of heparanase. Cytokine and immune checkpoint blockade immunotherapy for metastases was compromised when NK cells lacked heparanase. Our data suggest that heparanase plays a critical role in NK cell invasion into tumors and thereby tumor progression and metastases. This should be considered when systemically treating cancer patients with heparanase inhibitors, since the potential adverse effect on NK cell infiltration might limit the antitumor activity of the inhibitors.

Authors

Eva M. Putz, Alyce J. Mayfosh, Kevin Kos, Deborah S. Barkauskas, Kyohei Nakamura, Liam Town, Katharine J. Goodall, Dean Y. Yee, Ivan K.H. Poon, Nikola Baschuk, Fernando Souza-Fonseca-Guimaraes, Mark D. Hulett, Mark J. Smyth

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Figure 4

NK cell proliferation and function are unchanged by loss of heparanase.

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NK cell proliferation and function are unchanged by loss of heparanase. ...
(A and B) Purified BM NK cells from Hpsefl/fl NKp46-WT or Hpsefl/fl NKp46-iCre mice were labeled with CTV and cultured for 3 days in IL-15 as indicated (mean ± SD; n = 2 biological replicates). (A) Apoptosis was determined by annexin V and propidium iodide staining. (B) Proliferation was assessed by CTV dilution. (C) Purified splenic CFSE-labeled NK cells (2 × 105) were injected i.v. into B6.Rag2–/– Il2rg–/– mice. After 3 days, the proliferation of CD45+TCRβ–NK1.1+DX5+ NK cells in the indicated organs was determined by flow cytometry. Flow cytometric plot shows a representative proliferation profile. Data in the bar graph were pooled from 2 independent experiments (mean ± SEM; n = 8 per group). (D) The cytotoxicity of freshly isolated splenocytes or IL-2–activated NK cells (1,000 U/ml for 5 days) against YAC-1 and B16F10 target cells was tested at the indicated E/T ratios after 4 hours (mean ± SD; n = 3 biological replicates; 1 representative experiment of 2 experiments). (E) Splenocytes (5 × 106) were stimulated for 4 hours with 1 ng/ml IL-12, 100 ng/ml IL-15, and 10 ng/ml IL-18, and the expression of CD107a was assessed on TCRβ–NK1.1+DX5+ NK cells (mean ± SD; n = 4 mice per group). (F) Lung cells were stimulated for 4 hours in 1 ng/ml IL-12, 100 ng/ml IL-15, and 10 ng/ml IL-18, and the production of IFN-γ was measured by intracellular staining (mean ± SEM; n = 10; data were pooled from 3 independent experiments). (G) Purified splenic NK cells were stimulated in 50 ng/ml IL-15, 100 ng/ml IL-21, 1 ng/ml IL-12, 10 ng/ml IL-18, or anti-NK1.1 precoated wells. The release of IFN-γ was measured after 24 hours by CBA (mean ± SD; n = 2 biological replicates; 1 representative experiment of 2 experiments).