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 5

NK cell invasion is impaired in the absence of heparanase.

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NK cell invasion is impaired in the absence of heparanase. (A) The expre...
(A) The expression of CD62L and CXCR3 on TCRβ–NK1.1+DX5+ NK cells was analyzed by flow cytometry in the indicated organs of Hpsefl/fl NKp46-WT and Hpsefl/fl NKp46-iCre mice (mean ± SEM; n = 3–10). (B) Preactivated splenic NK cells (7.5 × 104) were seeded in the upper chamber of a Transwell insert. The number of migrating cells in response to 10% FBS and 20 ng/ml CXCL10 was assessed after 17 hours (mean ± SEM; n = 5; data were pooled from 2 independent experiments). (C) Heparanase enzymatic activity of isolated Hpse+/+ and Hpse–/– splenic NK cells was determined by a TR-FRET–based HS degradation assay (mean ± SD; n = 3). (D) Mice were injected s.c. with 100 μl Matrigel. Leukocyte infiltration into the Matrigel plugs was determined by flow cytometry after 3 days. Depicted are TCRβ–NK1.1+NKp46+ NK, TCRβ+CD4+, and TCRβ+CD8+ T cells, respectively (mean ± SEM; n = 9–10 mice per group; data were pooled from 3 independent experiments). (E) Mice were injected i.v. with 5 × 105 B16F10 melanoma cells. Lungs were harvested after 24 hours, and NK cell proportions and numbers were determined by flow cytometry (mean ± SEM; n = 9–12 mice per group; data were pooled from 3 independent experiments; direct comparison of NK cell proportions between B16F10 challenged NKp46-WT and NKp46-iCre mice P = 0.0118, Mann-Whitney U test, and NK cell numbers between B16F10 challenged NKp46-WT and NKp46-iCre mice: P = 0.0278, Mann-Whitney U test). (F–H) Mice were injected s.c. with 5 × 106 RMA-S-RAE-1β cells. Tumors were harvested on day 5 and analyzed by immunofluorescence (mean ± SEM; n = 4–6 per group). (F) Representative images of sections stained for NKp46+ NK cells (magenta) and DAPI (blue). Scale bars: 500 μm. Original magnification: ×20, tiled scan of whole tumor. (G) The distance of individual NK cells from the edge was calculated. (H) The number of NKp46+ cells per section was quantified by Imaris. Statistically significant differences were determined by Student’s t test (C), Mann-Whitney U test (D and H), or 1-way ANOVA with Tukey’s post test (E). *P < 0.05 and **P < 0.01.