Aberrantly Expressed Embryonic Protein NODAL Alters Breast Cancer Cell Susceptibility to γδ T Cell Cytotoxicity

Abstract

Gamma delta (γδ) T cells kill transformed cells, and increased circulating γδ T cells levels correlate with improved outcome in cancer patients; however, their function within the breast tumor microenvironment (TME) remains controversial. As tumors progress, they begin to express stem-cell associated proteins, concomitant with the emergence of therapy resistant metastatic disease. For example, invasive breast cancers often secrete the embryonic morphogen, NODAL. NODAL has been shown to promote angiogenesis, therapy resistance and metastasis in breast cancers. However, to date, little is known about how this secreted protein may interact with cells in the TME. Herein we explore how NODAL in the TME may influence γδ T cell function. We have assessed the proximity of γδ T cells to NODAL in a cohort of triple negative breast tumors. In all cases in which γδ T cells could be identified in these tumors, γδ T cells were found in close proximity to NODAL-expressing tumor cells. Migration of γδ and αβ T cells was similar toward MDA-MB-231 cells in which NODAL had been knocked down (shN) and MDA-MB-231 scrambled control cells (shC). Furthermore, Vδ1 γδ T cells did not migrate preferentially toward conditioned medium from these cell lines. While 24-h exposure to NODAL did not impact CD69, PD-1, or T cell antigen receptor (TCR) expression on γδ T cells, long term exposure resulted in decreased Vδ2 TCR expression. Maturation of γδ T cells was not significantly influenced by NODAL stimulation. While neither short-nor long-term NODAL stimulation impacted the ability of γδ T cells to kill MCF-7 breast cancer cells, the absence of NODAL resulted in greater sensitivity of targets to γδ T cell cytotoxicity, while overexpression of NODAL conferred resistance. This appeared to be at least in part due to an inverse correlation between NODAL and surface MICA/B expression on breast cancer target lines. As such, it appears that NODAL may play a role in strategies employed by breast cancer cells to evade γδ T cell targeting, and this should be considered in the development of safe and effective γδ T cell immunotherapies.

Clinicopathological Correlates of γδ T Cell Infiltration in Triple-Negative Breast Cancer

“…Several studies have brought evidence of the presence of γδ T cells in the tumor microenvironment of TNBC [ 25 , 32 , 33 , 34 ]. Ma et al previously evaluated the correlations between γδ T cell infiltration and BC characteristics and prognosis in 46 patients [ 50 ].…”

“…Indeed, they display a strong major histocompatibility complex-independent reactivity against many tumor cell types; they also have no alloreactivity and can be massively expanded from human samples [ 31 ]. Although evidence indicates that γδ T cells have a role in cancer, data on their frequency in cancer tissues and clinicopathological correlates remain scarce, particularly in TNBC, for which only a small number of samples have been analyzed [ 25 , 32 , 33 , 34 ]. Yet, the precise characterization of TNBC stromal components could allow refining the prognostic evaluation and identifying additional targets for immune modulation in this cancer subtype, for which treatments were lacking.…”

Embryonic protein NODAL regulates the breast tumor microenvironment by reprogramming cancer-derived secretomes

“…For example, NODAL seems to induce a breast cancer secretome that promotes angiogenesis through regulation of the angiogenic factors platelet-derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) [45] . Furthermore, NODAL expression is inversely correlated with susceptibility to gamma delta (γδ) T cell cytotoxicity, at least in part through decreased surface expression of the immune activating danger signal MHC class I polypeptide-related sequence A/B (MICA/B) [50] . CAFs from a gastric cancer mouse model have recently been shown to promote cancer cell proliferation and resistance to doxorubicin via NODAL secretion [51] and NODAL appears to induce CAF-like phenotypes in mouse and human fibroblast cell lines [52] .…”

Embryonic Protein NODAL Regulates the Breast Tumour Microenvironment by Reprogramming Cancer-Derived Secretomes

“…Recent efforts have shown that NODAL alters breast cancer cell susceptibility to γδ T cell killing by acting on cancer cells to decrease recognizable antigens on the cell surface. Furthermore, long-term NODAL stimulation reduced Vδ2 T cell antigen receptor expression, suggesting activation of an as-of-yet unidentified signaling pathway in primary γδ T cells50 . We build upon these studies by showing that NODAL may affect TME function and composition directly or indirectly by broadly regulating the breast cancer secretome.…”

“…For example, NODAL seems to induce a breast cancer secretome that promotes angiogenesis through regulation of the angiogenic factors platelet-derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) 45 . Furthermore, NODAL expression is inversely correlated with susceptibility to gamma delta (γδ) T cell cytotoxicity, at least in part through decreased surface expression of the immune activating danger signal MHC class I polypeptide-related sequence A/B (MICA/B) 50 . CAFs from a gastric cancer mouse model have recently been shown to promote cancer cell proliferation and resistance to doxorubicin via NODAL secretion 51 and NODAL appears to induce CAF-like phenotypes in mouse and human fibroblast cell lines 52 .…”