Spontaneous Breast Cancer Bone Metastasis is Associated with Mesenchymal Phenotype and Immune Suppression

Introduction:

Mortality from breast cancer is almost exclusively due to metastasis. Bone metastasis occur in up to 85% of patients with advanced breast cancer, and are associated with morbidity and mortality. Breast cancer-induced bone metastases are characterized by changes in the bone microenvironment and frequently osteolytic lesions. These alterations disrupt the homeostasis of the immune cell populations within the bone marrow, presumably affecting the entire immune balance of the body.

Materials and methods:

To characterize the changes at the metastatic microenvironment we generated bone-seeking variants of 4T1 cells by two rounds of in vivo selection (4T1.1 and 4T1.2). We calibrated the timeline for the formation of a pre-metastatic niche, before the formation of macro-metastases, and mapped the early changes of various immune cell populations in multiple organs, as well as the transcriptome of bone-tropic cells compared with the parental cells.

Results and discussion:

We found that the bone-metastasizing 4T1.2 cells had a more mesenchymal phenotype compared to the parental 4T1, suggesting that bone metastasizing cells undergo epithelial to mesenchymal transition in order to invade the bone. In addition, the bone seeking cells exhibit "osteomimicry" behavior, reflected by increased expression of bone-specific genes (ALP, BSP, OCN), suggesting that adapting to the specific metastatic microenvironment is required for metastatic seeding. Moreover, analysis of cell populations in BM of bone metastases-bearing mice revealed an immune-suppressed microenvironment, including an increase in the population of granulocytic BMDCs and a decrease in T cells. Importantly, the immune cell changes in bone, blood, spleen and lungs preceded the formation of spontaneous bone metastases, suggesting that formation of an immune suppressed niche facilitates organ colonization, in agreement with the notion that breast cancer is a systemic disease.

Conclusions:

In this work, we established and characterized a spontaneous model of breast cancer bone metastasis, and demonstrated that alterations in the bone microenvironment occur prior to metastases detection. Our findings may provide the basis for identifying new targets that could prevent bone metastatic relapse.