Malignant cell-specific pro-tumorigenic role of type I interferon receptor in breast cancers

Abstract

Within the microenvironment of solid tumors, stress associated with deficit of nutrients and oxygen as well as tumor-derived factors triggers the phosphorylation-dependent degradation of the IFNAR1 chain of type I interferon (IFN1) receptor and ensuing suppression of the IFN1 pathway. Here we sought to examine the importance of these events in malignant mammary cells. Expression of non-degradable IFNAR1^S526A mutant in mouse mammary adenocarcinoma cells stimulated the IFN1 pathway yet did not affect growth of these cells in vitro or ability to form subcutaneous tumors in the syngeneic mice. Remarkably, these cells exhibited a notably accelerated growth when transplanted orthotopically into mammary glands. Importantly, in human patients with either ER+ or ER- breast cancers, high levels of IFNAR1 were associated with poor prognosis. We discuss the putative mechanisms underlying the pro-tumorigenic role of IFNAR1 in malignant breast cells.

Keywords: IFNAR1; PD-L1; breast cancer; immune therapy; interferon receptor; mammary adenocarcinoma; type I interferon.

Figure 1.

Expression of SA-GFP in breast cancer cells does not affect proliferation in vitro. (a, b) Representative histograms of IFNAR1 cell surface levels measured by flow cytometry on GFP and SA-GFP E0771 (a) and AT-3 cells (b). (c) Immunoblot analysis of activated (pSTAT1, pSTAT3) and total Stat1 and Stat3 in E0771 cells line expressing GFP or SA-GFP. Levels of GAPDH were used as a loading control. (d) qPCR analysis of the ISGs mRNA levels in E0771 cells expressing GFP or SA-GFP. The data are shown as mean ± SEM (n = 3). (e, f) Cell proliferation assay of GFP and SA-GFP E0771 (e) and AT-3 (f) cells. Cells (mean±SEM, n = 3) were counted for 3 days after seeding.

Figure 2.

Breast cancer cells expressing SA-GFP grow faster orthotopically but not subcutaneously in syngeneic mouse model. (a, b) Subcutaneous (a) and orthotopic (b) E0771-tdTomato-GFP (GFP) and E0771-tdTomato-Ifnar1SA-GFP (SA-GFP) tumor growth in WT mice. (c) Orthotopic AT-3-GFP (GFP) and AT-3-Ifnar1SA-GFP (SA-GFP) tumor growth in WT mice. (d) qPCR analysis of the expression of ISGs from GFP and SA-GFP E0771 tumors from (B). The data are shown as average fold change relative to E0771-GFP control ± SEM (n = 9 per group).

Figure 3.

Breast cancer cells with induced IFN1 signaling have higher PD-L1 levels and response to anti-PD-L1 treatment. (a,b) Flow cytometry analysis of PD-L1 cell surface levels on GFP and SA-GFP E0771 (a) and AT-3 (b) cells. A representative histogram (left) and quantification (right) are shown. (c,d) The effect of anti-PD-L1 treatment on SA-GFP E0771 tumor growth in WT syngeneic mice. Data are mean±SEM of tumor volume (c) and tumor weight (d) for 8 IgG- and 10 anti-PD-L1-treated mice per group.

Figure 4.

The high levels of IFNAR1 inversely correlate with breast cancer poor prognosis. (a) Representative immunohistochemistry staining of human breast cancer expressing high or low levels of IFNAR1 in cancer cells. Scale bars represent 50 µm. (be) Breast cancer patients with high cancer cell levels of IFNAR1 have poor progression-free survival in comparison with patients with low IFNAR1 expression, an effect observed in all patients (b), within ER-positive patients (c), within ER-negative patients (d) or within HER2-positive patients (e).