Tumors systemically initiate metastatic niches in distant target metastatic organs. These niches, composed of bone marrow-derived hematopoietic cells, provide permissive conditions for future metastases. However, the mechanisms by which these cells mediate outgrowth of metastatic tumor cells are not completely known. Using mouse models of spontaneous breast cancer, we show enhanced recruitment of bone marrow-derived CD11b

The progression of cancer to metastatic disease is a major cause of death. We identified miR-708 being transcriptionally repressed by polycomb repressor complex 2-induced H3K27 trimethylation in metastatic breast cancer. miR-708 targets the endoplasmic reticulum protein neuronatin to decrease intracellular calcium level, resulting in reduction of activation of ERK and FAK, decreased cell migration, and impaired metastases. Ectopic expression of neuronatin refractory to suppression by miR-708 rescued cell migration and metastasis defects. In patients with breast cancer, miR-708 expression was decreased in lymph node and distal metastases, suggesting a metastasis-suppressive role. Our findings uncover a mechanistic role for miR-708 in metastasis and provide a rationale for developing miR-708 as a therapeutic agent against metastatic breast cancer.

Increased compliance with an Enhanced Recovery After Surgery pathway is associated with improved clinical outcomes after resection for primary lung cancer. Several elements, including early mobilization, appear to be more influential than others.

Inflammation is inextricably associated with primary tumor progression. However, the contribution of inflammation to tumor outgrowth in metastatic organs has remained underexplored. Here, we show that extrinsic inflammation in the lungs leads to the recruitment of bone marrow-derived neutrophils, which degranulate azurophilic granules to release the Ser proteases, elastase and cathepsin G, resulting in the proteolytic destruction of the antitumorigenic factor thrombospondin-1 (Tsp-1). Genetic ablation of these neutrophil proteases protected Tsp-1 from degradation and suppressed lung metastasis. These results provide mechanistic insights into the contribution of inflammatory neutrophils to metastasis and highlight the unique neutrophil protease-Tsp-1 axis as a potential antimetastatic therapeutic target.T he contribution of inflammation to primary tumor progression is well documented (1); however, little is known about its role in metastatic outgrowth in distant organs. The lung, which is a frequent site of metastasis from extrapulmonary neoplasms, is susceptible to inflammatory insults. Bacterial infection-induced, metastasis-conducive environments in the lung (2, 3) and cigarette smoke-induced inflammation were associated with pulmonary metastasis from breast cancer (2, 4).Bacterial lipopolysaccharide (LPS) is a well-characterized inducer of inflammation because its binding to toll-like receptor 4 (TLR4) results in nuclear factor kappa B (NF-κB) activation and expression of proinflammatory cytokines, including interleukin-1 beta (IL-1β), tumor necrosis factor alpha (TNF-α), and IL-6 (5). LPS-induced acute lung injury is marked by increased neutrophil influx and up-regulation of proinflammatory cytokines. Similar phenotypes are observed in other lung inflammatory conditions, including asthma (6), chronic obstructive pulmonary disease (7), and pneumonia (8, 9). LPS-mediated lung inflammation is associated with breast and colon cancer metastasis to the lungs (10-12).The mechanisms by which inflammation contributes to metastatic outgrowth in distant organs have remained underexplored. From a clinical perspective, although blocking primary tumor invasion and blocking dissemination are considered effective approaches in suppressing metastasis, an important question is how best to treat patients whose tumor has already metastasized. Thus, approaches are required to block tumor outgrowth in secondary organs for effective treatment of metastatic cancers. In this study, using two independent models of lung inflammation, we show enhanced recruitment of neutrophils, which degranulate to release the Ser proteases, neutrophil elastase (NE) and cathepsin G (CG), to degrade thrombospondin-1 (Tsp-1) in the lung microenvironment, enhancing metastatic outgrowth. Protease deficiency protected Tsp-1 from proteolysis and suppressed metastasis, providing a previously unidentified mechanism of Tsp-1 regulation in the metastatic organ. Results Neutrophil-Mediated Lung Inflammation Enhances MetastaticOutgrowth. To determine the contribu...

Metastatic tumors have been shown to establish permissive microenvironments for metastases via recruitment of bone marrow (BM)- derived cells. Here, we show that metastasis-incompetent tumors are also capable of generating such microenvironments. However, in these situations the otherwise pro-metastatic Gr1+ myeloid cells create a metastasis-refractory microenvironment via the induction of thrombospondin-1 (Tsp-1) by tumor-secreted prosaposin. (BM)-specific genetic deletion of Tsp-1 abolished the inhibition of metastasis, which was restored by BM transplant from Tsp-1+ donors. We also developed a 5-amino acid peptide from prosaposin as a pharmacological inducer of Tsp-1 in Gr1+ BM cells, which dramatically suppresses metastasis. These results provide mechanistic insights into why certain tumors are deficient in metastatic potential and implicate recruited Gr1+ myeloid cells as the main source of Tsp-1. The results underscore the plasticity of Gr1+ cells, which, depending on the context, promote or inhibit metastasis, and suggest that the peptide could be a potential therapeutic agent against metastatic cancer.

MicroRNAs (miRNAs) are key regulators of gene expression and contribute to a variety of biological processes. Abnormal miRNA expression has been reported in various diseases including pathophysiology of breast cancer, where they regulate protumorigenic processes including vascular invasiveness, estrogen receptor status, chemotherapy resistance, invasion and metastasis. The miRBase sequence database, a public repository for newly discovered miRNAs, has grown rapidly with approximately >10,000 entries to date. Despite this rapid growth, many miRNAs have not yet been validated, and several others are yet to be identified. A lack of a full complement of miRNAs has imposed limitations on recognizing their important roles in cancer, including breast cancer. Using deep sequencing technology, we have identified 189 candidate novel microRNAs in human breast cancer cell lines with diverse tumorigenic potential. We further show that analysis of 500-nucleotide pri-microRNA secondary structure constitutes a reliable method to predict bona fide miRNAs as judged by experimental validation. Candidate novel breast cancer miRNAs with stem lengths of greater than 30 bp resulted in the generation of precursor and mature sequences in vivo. On the other hand, candidates with stem length less than 30 bp were less efficient in producing mature miRNA. This approach may be used to predict which candidate novel miRNA would qualify as bona fide miRNAs from deep sequencing data with approximately 90% accuracy.

Wound healing involves the concerted action of various lymphoid and in particular myeloid cell populations. To characterize and quantitate different types of myeloid cells and to obtain information on their kinetics during wound healing, we performed multiparametric flow cytometry analysis. In healthy mice, neutrophil numbers increased early after injury and returned to near basal levels after completion of healing. Macrophages, monocyte‐derived dendritic cells (DCs), and eosinophils were abundant throughout the healing phase, in particular in early wounds, and Langerhans cells increased after wounding and remained elevated after epithelial closure. Major differences in healing‐impaired diabetic mice were a much higher percentage of immune cells in late wounds, mainly as a result of neutrophil, macrophage, and monocyte persistence; reduced numbers and percentages of macrophages and monocyte‐derived DCs in early wounds; and of Langerhans cells, conventional DCs, and eosinophils throughout the healing process. Finally, unbiased cluster analysis (PhenoGraph) identified a large number of different clusters of myeloid cells in skin wounds. These results provide insight into myeloid cell diversity and dynamics during wound repair and highlight the abnormal inflammatory response associated with impaired healing.

Healing of skin wounds is orchestrated by various types of immune cells, but little is known about the role of FoxP3 regulatory T cells (Tregs) in this process. Here, we determined if Tregs are important for wound healing in normal mice and if they contribute to the accelerated healing of mice overexpressing the growth and differentiation factor activin. Diphtheria toxin induced Treg depletion prior to injury caused impaired healing characterized by delayed reepithelialization, reduced wound contraction, and impaired vessel maturation. The accelerated wound repair of activin-transgenic mice was also abrogated. Mechanistically, we found a strong increase in IL-4 levels combined with overrepresentation of T-bet and GATA-3 αβ T cells in Treg-depleted 7-day wounds. In addition, numbers of IFN-γ- or IL-17A-producing CD4 and CD4 T cells were elevated. These results demonstrate that Treg depletion in wounds facilitates the expansion of an αβ T-cell population with features of Th1 and Th2 cells, and suggest that concomitant changes in the cytokine milieu disturb the healing process.