No abstract

Immunosuppressive elements within the tumor microenvironment, such as tumor-associated macrophages (TAM), can present a barrier to successful anti-tumor responses by cytolytic T cells. Here we employed preclinical syngeneic p53 null mouse models of triple-negative breast cancer (TNBC) to develop a treatment regimen that harnessed the immunostimulatory effects of low-dose cyclophosphamide coupled with the pharmacologic inhibition of TAMs using either a small molecule CSF1R inhibitor or an anti-CSF1R antibody. This therapeutic combination was effective in treating several highly aggressive TNBC murine mammary tumor and lung metastasis models. Single cell RNA sequencing characterized tumor-infiltrating lymphocytes (TIL) including helper T cells and antigen-presenting B cells that were highly enriched in responders to combination therapy. In one model that exhibited long-term post-treatment tumor regression, high dimensional imaging techniques identified the close spatial localization of B220+/CD86+-activated B cells and CD4+ T cells in tertiary lymphoid structures that were present up to 6 weeks post-treatment. The transcriptional and metabolic heterogeneity of TAMs was also characterized in two closely related claudin-low/mesenchymal subtype tumor models with differential treatment responses. A murine TAM signature derived from the T12 model was highly conserved in human claudin-low breast cancers, and high expression of the TAM signature correlated with reduced overall survival in breast cancer patients. This TAM signature may help identify human claudin-low breast cancer patients that will benefit from the combination of cyclophosphamide and anti-CSF1R therapy. These studies illustrate the complexity of the tumor immune microenvironment and highlight different immune responses that result from rational immunotherapy combinations.

niches may dictate the cellular fates and therapeutic responses of DTCs and microscopic metastases. Cancer cells may exploit the niches' normal roles, including protection from aberrant immune activation and remodeling/repair of bones, to facilitate metastatic progression. Therefore, identification of the cancer-niche crosstalk pathways, especially those involved in immunosurveillance and tissue repair, may lead to novel mechanistic insights and therapeutic targets.There are challenges and opportunities in our further investigations of BM niches in metastatic colonization. Bone-resident cells apparently of the same type may actually be heterogeneous with regard to the location, lineage, and functions. This is exemplified by pericytes/MSCs (195,196) and endothelial cells (197), which may reconcile the seemingly contradictory findings (e.g., refs. 81, 87). Future studies are needed for more precise definition of various niches. To this end, single-cell transcriptomic or proteomic analyses that preserve the spatial information may be highly valu-

The bone microenvironment is dynamic and undergoes remodeling in normal and pathological conditions. Whether such remodeling impacts disseminated tumor cells and bone metastasis remains poorly understood. Here, we demonstrated that pathological fractures increase metastatic colonization around the injury. NG2+ cells are a common participant of bone metastasis initiation and bone remodeling in both homeostatic and fractured conditions. NG2+ bone mesenchymal stromal cells (BMSCs) often co-localize with DTCs in the perivascular niche. Both DTCs and NG2+ BMSCs are recruited to remodeling sites. Ablation of NG2+ lineage impaired bone remodeling and concurrently diminished metastatic colonization. In co-cultures, NG2+ BMSCs, especially when undergoing osteo-differentiation, enhanced cancer cell proliferation and migration. Knockout of N-cadherin in NG2+ cells abolished these effects in vitro, and phenocopied NG2+ lineage depletion in vivo. These findings uncover dual roles of NG2+ cells in metastasis and remodeling, and indicate that osteo-differentiation of BMSCs promotes metastasis initiation via N-cadherin-mediated cell-cell interaction.