Cancer-associated fibroblasts (CAFs) are highly prominent in breast tumors, but their functional heterogeneity and origin are still largely unresolved. We report that bone marrow (BM)–derived mesenchymal stromal cells (MSCs) are recruited to primary breast tumors and to lung metastases and differentiate to a distinct subpopulation of CAFs. We show that BM-derived CAFs are functionally important for tumor growth and enhance angiogenesis via up-regulation of Clusterin. Using newly generated transgenic mice and adoptive BM transplantations, we demonstrate that BM-derived fibroblasts are a substantial source of CAFs in the tumor microenvironment. Unlike resident CAFs, BM-derived CAFs do not express PDGFRα, and their recruitment resulted in a decrease in the percentage of PDGFRα-expressing CAFs. Strikingly, decrease in PDGFRα in breast cancer patients was associated with worse prognosis, suggesting that BM-derived CAFs may have deleterious effects on survival. Therefore, PDGFRα expression distinguishes two functionally unique CAF populations in breast tumors and metastases and may have important implications for patient stratification and precision therapeutics.

Cancer-Associated Fibroblasts (CAFs) are the most prominent stromal cell type in breast tumors. CAFs promote tumor growth and metastasis by multiple mechanisms, including by mediating tumor-promoting inflammation. Immune modulation in the tumor microenvironment plays a central role in determining disease outcome. However, the functional interactions of CAFs with immune cells are largely unknown. Here we report a novel signaling axis between fibroblasts, cancer cells and immune cells in breast tumors that drives an immunosuppressive microenvironment, mediated by CAF-derived Chi3L1. We demonstrate that Chi3L1 is highly upregulated in CAFs isolated from mammary tumors and pulmonary metastases of transgenic mice, and in the stroma of human breast carcinomas. Genetic ablation of Chi3L1 in fibroblasts in vivo attenuated tumor growth, macrophage recruitment and reprogramming to an M2-like phenotype, enhanced tumor infiltration by CD8+ and CD4+ T cells and promoted a Th1 phenotype. These results indicate that CAF-derived Chi3L1 promotes tumor growth and shifts the balance of the immune milieu towards type 2 immunity. Taken together, our findings implicate fibroblast-derived Chi3L1 as a novel key player in the complex reciprocal interactions of stromal cells that facilitate tumor progression and metastasis, and suggest that targeting Chi3L1 may be clinically beneficial in breast cancer.

Cancer-Associated Fibroblasts (CAFs) were shown to orchestrate tumour-promoting inflammation in multiple malignancies, including breast cancer. However, the molecular pathways that govern the inflammatory role of CAFs are poorly characterised. In this study we found that fibroblasts sense damage-associated molecular patterns (DAMPs), and in response activate the NLRP3 inflammasome pathway, resulting in instigation of pro-inflammatory signalling and secretion of IL-1β. This upregulation was evident in CAFs in mouse and in human breast carcinomas. Moreover, CAF-derived inflammasome signalling facilitated tumour growth and metastasis, which was attenuated when NLRP3 or IL-1β were specifically ablated. Functionally, CAF-derived inflammasome promoted tumour progression and metastasis by modulating the tumour microenvironment towards an immune suppressive milieu and by upregulating the expression of adhesion molecules on endothelial cells. Our findings elucidate a mechanism by which CAFs promote breast cancer progression and metastasis, by linking the physiological tissue damage response of fibroblasts with tumour-promoting inflammation.

The mechanism of insulin dysregulation in children with hyperinsulinism associated with inactivating mutations of short-chain 3-hydroxyacyl-CoA dehydrogenase (SCHAD) was examined in mice with a knock-out of the hadh gene (hadh ؊/؊ ). Congenital hyperinsulinism is the most common cause of persistent hypoglycemia in infants and children (1). Six genetic loci have been associated with the disorder. The most common of these disorders are due to inactivating mutations of the sulfonylurea receptor 1 (SUR1) 2 and Kir6.2 subunits of the ␤-cell ATP-dependent potassium (K ATP ) channel or to activating mutations of glutamate dehydrogenase (GDH) and glucokinase. Recently, several children have been described with a recessively inherited form of hyperinsulinism that is associated with deficiency of a mitochondrial fatty acid ␤-oxidation enzyme, short-chain 3-hydroxyacyl-CoA dehydrogenase (SCHAD) encoded by the HADH gene on 4q (2-4). SCHAD catalyzes the third step in the ␤-oxidation cycle for medium and short-chain 3-hydroxy fatty acyl-CoAs. Children afflicted with SCHAD deficiency have recurrent episodes of hypoglycemia that can be prevented by treatment with diazoxide (2, 3) and also have characteristic accumulations of fatty acid metabolites, including plasma 3-hydroxybutyrylcarnitine and urinary 3-hydroxyglutaric acid (2, 3). This form of abnormal insulin regulation is unique, because other genetic disorders of mitochondrial fatty acid oxidation do not cause hyperinsulinism (5). In addition, the genetic defect in SCHAD deficiency is expected to impair, rather than increase, the production of ATP, which normally serves as the triggering signal for insulin release. An important clue to the mechanism of insulin dysregulation in SCHAD deficiency has been recently provided by the report * This work was supported, in whole or in part, by National Institutes of Health Grants DK53012 (to C. A. S.), DK22122 (to F. M. M.), DK 53761 (to I. N.), HL075421 (to A. W. S.). This work was also supported by a fellowship award from Society for Inherited Metabolic Disorders (to A. P.

Breast tumors are characterized by an extensive desmoplastic stroma, abundantly populated by fibroblasts. Cancer-associated fibroblasts (CAF) support tumorigenesis by stimulating angiogenesis, cancer cell proliferation, and invasion. CAF also orchestrate tumor-promoting inflammation in multiple tumor types, including breast cancer. However, the mechanisms through which normal tissue fibroblasts are reprogrammed to tumor-promoting CAFs are mainly obscure. Here, we show that mammary fibroblasts can be educated by breast cancer cells to become activated to a proinflammatory state that supports malignant progression. Proteomic analysis of breast cancer cell-secreted factors identified the secreted proinflammatory mediator osteopontin, which has been implicated in inflammation, tumor progression, and metastasis. Osteopontin was highly secreted by mouse and human breast cancer cells, and tumor cell-secreted osteopontin activated a CAF phenotypes in normal mammary fibroblasts in vitro and in vivo. Osteopontin was sufficient to induce fibroblast reprogramming and neutralizing antibodies against osteopontinblocked fibroblast activation induced by tumor cells. The ability of secreted osteopontin to activate mammary fibroblasts relied upon its known receptors CD44 and a V b 3 integrin. Strikingly, osteopontin silencing in tumor cells in vivo attenuated stromal activation and inhibited tumor growth. Our findings establish a critical functional role for paracrine signaling by tumor-derived osteopontin in reprograming normal fibroblasts into tumor-promoting CAFs. Cancer Res; 75(6); 963-73. Ó2015 AACR.