CD8+ T cells from experimental in situ breast carcinoma interfere with bone homeostasis

Abstract

Before bone colonization, immune cells primed by breast primary tumor cells actively modify the bone microenvironment, disturbing the complex and tightly homeostatic signaling network regulated by osteoblasts and osteoclasts. Indeed, we have shown that RANKL⁺ CD4⁺ T cells specific for the 4T1 mammary carcinoma cell line, arrive at the bone marrow (BM) before metastatic cells and set the pre-metastatic niche. In the absence of RANKL expressed by T cells, there is no pre-metastatic osteolytic disease and bone metastases are blocked. Adding to the role of T cells, we have recently demonstrated that dendritic cells (DCs) provide a positive feedback loop to the osteolytic profile induced by the metastatic tumor. In this setting, DCs are able to differentiate into potent bone resorbing osteoclast-like cells keeping their antigen-presenting cell (APC) properties to maintain RANKL⁺ CD4⁺ Th17 T cells activities, via IL-23 expression. Here we show that 67NR non-metastatic tumor cells, a sibling of 4T1 tumor cell line, induce an increase in trabecular bone mass on day 11 post-tumor implant. This observation was associated with an expansion of the osteoblastic lineage cells accompanied by a reduction of osteoclasts numbers. Moreover, BM derived CD8⁺ T cells from 67NR tumor-bearing mice, express an anti-osteoclastogenic cytokine milieu enriched by IFN-γ, IL-10 and producing low levels of RANKL. The frequency of BM derived CD8⁺ FoxP3⁺ regulatory T cells, known as potent suppressors of osteoclastogenesis both in vitro and in vivo, was also increased in such animals. This milieu was capable to suppress 4T1 tumor-specific CD4⁺ T cells phenotype in vivo and in vitro and strongly inhibited bone metastases establishment, restoring trabecular bone mass volume. We concluded that the 67NR⁺ tumor derived CD8⁺ T cells phenotypes, either contributing to bone homeostasis and/or control of 4T1 breast tumor pre-metastatic disease, interfere with osteoclasts and osteoblasts activities inside BM. Our study highlights the opposing roles of subverted tumor CD4⁺ and CD8⁺ T cell subtypes in directing breast cancer progression and bone metastases establishment. For non-metastatic tumors, the role of T cells regarding bone remodeling has never been addressed before. As far as we know, this is the first description that an in situ carcinoma can modify distant sites. In the case showed here, modification of the distant bone site disfavors pre-metastatic bone niche formation.

Keywords: Bone metastasis; Breast cancer and pre-metastatic niche; Breast tumor; CD8(+) T cells; CD8(+) T regulatory cells; Osteoblasts; Osteoclasts.