Glycoengineered anti-CD39 promotes anticancer responses by depleting suppressive cells and inhibiting angiogenesis in tumor models
Haohai Zhang, Lili Feng, Paola de Andrade Mello, Changchuin Mao, Richard Near, Eva Csizmadia, Leo Li-Ying Chan, Keiichi Enjyoji, Wenda Gao, Haitao Zhao, Simon C. Robson
Haohai Zhang, Lili Feng, Paola de Andrade Mello, Changchuin Mao, Richard Near, Eva Csizmadia, Leo Li-Ying Chan, Keiichi Enjyoji, Wenda Gao, Haitao Zhao, Simon C. Robson
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Research Article Angiogenesis Immunology

Glycoengineered anti-CD39 promotes anticancer responses by depleting suppressive cells and inhibiting angiogenesis in tumor models

Abstract

Immunosuppressive cells accumulating in the tumor microenvironment constitute a formidable barrier that interferes with current immunotherapeutic approaches. A unifying feature of these tumor-associated immune and vascular endothelial cells appears to be the elevated expression of ectonucleotidase CD39, which in tandem with ecto-5′-nucleotidase CD73, catalyzes the conversion of extracellular ATP into adenosine. We glycoengineered an afucosylated anti-CD39 IgG2c and tested this reagent in mouse melanoma and colorectal tumor models. We identified major biological effects of this approach on cancer growth, associated with depletion of immunosuppressive cells, mediated through enhanced Fcγ receptor–directed (FcγR-directed), antibody-dependent cellular cytotoxicity (ADCC). Furthermore, regulatory/exhausted T cells lost CD39 expression, as a consequence of antibody-mediated trogocytosis. Most strikingly, tumor-associated macrophages and endothelial cells with high CD39 expression were effectively depleted following antibody treatment, thereby blocking angiogenesis. Tumor site–specific cellular modulation and lack of angiogenesis synergized with chemotherapy and anti–PD-L1 immunotherapy in experimental tumor models. We conclude that depleting suppressive cells and targeting tumor vasculature, through administration of afucosylated anti-CD39 antibody and the activation of ADCC, comprises an improved, purinergic system–modulating strategy for cancer therapy.

Authors

Haohai Zhang, Lili Feng, Paola de Andrade Mello, Changchuin Mao, Richard Near, Eva Csizmadia, Leo Li-Ying Chan, Keiichi Enjyoji, Wenda Gao, Haitao Zhao, Simon C. Robson

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Figure 4

αCD39 mAb induces trogocytosis in vitro and in vivo.

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αCD39 mAb induces trogocytosis in vitro and in vivo. CHO cells overexpre...
CHO cells overexpressing mCD39 tagged with EGFP were used as donor cells, while murine macrophage J774A.1 cells were used as acceptor cells in the trogocytosis assay. (A) CHO-mCD39-EGFP cells were cultured alone or in combination with J774A.1 cells. When CHO-mCD39 EGFP cells were cultured in Transwells, J774A.1 cells were positioned in the bottom chamber. The cocultures were treated with the indicated mAb for 48 hours. Flow cytometry analyses of signals on CHO-mCD39-EGFP cells in those cultures are shown. (B) Representative confocal microscopy images of cocultured CHO-mCD39-EGFP and J774A.1 cells treated with the indicated mAb. The red arrow marks the engulfed CD39-EGFP in J774A.1 cells. Scale bars: 20 μm. (C) Flow cytometry analysis of EGFP signal on CHO-mCD39-EGFP cells of cocultures treated with indicated concentrations of αCD39 mAb. (D) CHO-mCD39-EGFP and J774A.1 cells were cocultured at indicated ratios and treated with 2 μg/mL αCD39 mAb. Flow cytometry analysis of the EGFP signal on CHO-mCD39-EGFP cells is shown. (E and F) MC38 tumor–bearing mice were treated with 2 doses of 5 mg/kg control (CTRL) or αCD39 mAb on days 8 and 11 after tumor implantation. B16F10 tumor–bearing mice were treated with 2 doses of 10 mg/kg CTRL or αCD39 mAb on days 11 and 14 after tumor implantation. Tissues were harvested 24 hours after the second dose of treatment and processed for CD39 IHC staining (E) or FACS analysis (F). Scale bars: 100 μm. Data are shown as mean ± SEM. The Mann-Whitney test was used for the statistical analysis. Data in A–F are representative of at least 2 independent experiments. **P < 0.01, ***P < 0.001.