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Caveolin-1-/- null mammary stromal fibroblasts share characteristics with human breast cancer-associated fibroblasts.

ABSTRACT: Recently, we reported that human breast cancer-associated fibroblasts show functional inactivation of the retinoblastoma (RB) tumor suppressor and down-regulation of caveolin-1 (Cav-1) protein expression. However, it remains unknown whether loss of Cav-1 is sufficient to confer functional RB inactivation in mammary fibroblasts. To establish a direct cause-and-effect relationship, mammary stromal fibroblasts (MSFs) were prepared from Cav-1(-/-) null mice and subjected to phenotypic analysis. Here, we provide evidence that Cav-1(-/-) MSFs share many characteristics with human cancer-associated fibroblasts. The Cav-1(-/-) MSF transcriptome significantly overlaps with human cancer-associated fibroblasts; both show a nearly identical profile of RB/E2F-regulated genes that are up-regulated, which is consistent with RB inactivation. This Cav-1(-/-) MSF gene signature is predictive of poor clinical outcome in breast cancer patients treated with tamoxifen. Consistent with these findings, Cav-1(-/-) MSFs show RB hyperphosphorylation and the up-regulation of estrogen receptor co-activator genes. We also evaluated the paracrine effects of "conditioned media" prepared from Cav-1(-/-) MSFs on wild-type mammary epithelia. Our results indicate that Cav-1(-/-) MSF "conditioned media" is sufficient to induce an epithelial-mesenchymal transition, indicative of an invasive phenotype. Proteomic analysis of this "conditioned media" reveals increased levels of proliferative/angiogenic growth factors. Consistent with these findings, Cav-1(-/-) MSFs are able to undergo endothelial-like transdifferentiation. Thus, these results have important implications for understanding the role of cancer-associated fibroblasts and RB inactivation in promoting tumor angiogenesis.

SUBMITTER: Sotgia F

PROVIDER: S-EPMC2665737 | biostudies-literature | 2009 Mar

REPOSITORIES: biostudies-literature

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Caveolin-1-/- null mammary stromal fibroblasts share characteristics with human breast cancer-associated fibroblasts.

Sotgia Federica F Del Galdo Francesco F Casimiro Mathew C MC Bonuccelli Gloria G Mercier Isabelle I Whitaker-Menezes Diana D Daumer Kristin M KM Zhou Jie J Wang Chenguang C Katiyar Sanjay S Xu Huan H Bosco Emily E Quong Andrew A AA Aronow Bruce B Witkiewicz Agnieszka K AK Minetti Carlo C Frank Philippe G PG Jimenez Sergio A SA Knudsen Erik S ES Pestell Richard G RG Lisanti Michael P MP

The American journal of pathology 20090301 3

Recently, we reported that human breast cancer-associated fibroblasts show functional inactivation of the retinoblastoma (RB) tumor suppressor and down-regulation of caveolin-1 (Cav-1) protein expression. However, it remains unknown whether loss of Cav-1 is sufficient to confer functional RB inactivation in mammary fibroblasts. To establish a direct cause-and-effect relationship, mammary stromal fibroblasts (MSFs) were prepared from Cav-1(-/-) null mice and subjected to phenotypic analysis. Here ...[more]

PMID: 19234134

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Project description:BackgroundThe extensional signals in cross-talk between stromal cells and tumor cells generated from extracellular matrix molecules, soluble factor, and cell-cell adhesion complexes cooperate at the extra- and intracellular level in the tumor microenvironment. CAFs are the primary type of stromal cells in the tumor microenvironment and play a pivotal role in tumorigenesis and development. Hitherto, there is hardly any systematic analysis of the intrinsic relationship between CAFs function and its abnormal signaling pathway. The extreme complexity of CAFs' features and their role in tumor development are needed to be further investigated.Methodology/principal findingsWe primary cultured CAFs and NFs from early stages of breast cancer tissue and identified them using their biomarker by immunohistochemistry for Fibronectin, ?-SMA and FAP. Microarray was applied to analyze gene expression profiles of human breast CAFs and the paired NFs. The Up-regulated genes classified by Gene Ontology, signal pathways enriched by DAVID pathway analysis. Abnormal signaling pathways in breast cancer CAFs are involved in cell cycle, cell adhesion, signal transduction and protein transport being reported in CAFs derived from other tumors. Significantly, the altered ATM signaling pathway, a set of cell cycle regulated signaling, and immune associated signaling are identified to be changed in CAFs.Conclusions/significanceCAFs have the vigorous ability of proliferation and potential of invasion and migration comparing with NFs. CAFs could promote breast cancer cell invasion under co-culture conditions through up-regulated CCL18 and CXCL12. Consistently with its biologic behavior, the gene expression profiling analyzed by microarray shows that some of key signaling pathways, such as cell cycle, cell adhesion, and secreting factors play an important role in CAFs. The altered ATM signaling pathway is abnormally active in the early stage of breast cancer. The set of immune associated signaling may be involved in tumor cell immune evasion.

Project description:We have recently proposed a new model for understanding tumor metabolism, termed: "The Autophagic Tumor Stroma Model of Cancer Metabolism". In this new paradigm, catabolism (autophagy) in the tumor stroma fuels the anabolic growth of aggressive cancer cells. Mechanistically, tumor cells induce autophagy in adjacent cancer-associated fibroblasts via the loss of caveolin-1 (Cav-1), which is sufficient to promote oxidative stress in stromal fibroblasts. To further test this hypothesis, here we created human Cav-1 deficient immortalized fibroblasts using a targeted sh-RNA knock-down approach. Relative to control fibroblasts, Cav-1 deficient fibroblasts dramatically promoted tumor growth in xenograft assays employing an aggressive human breast cancer cell line, namely MDA-MB-231 cells. Co-injection of Cav-1 deficient fibroblasts, with MDA-MB-231 cells, increased both tumor mass and tumor volume by ~4-fold. Immuno-staining with CD31 indicated that this paracrine tumor promoting effect was clearly independent of angiogenesis. Mechanistically, proteomic analysis of these human Cav-1 deficient fibroblasts identified > 40 protein biomarkers that were upregulated, most of which were associated with i) myofibroblast differentiation, or ii) oxidative stress/hypoxia. In direct support of these findings, the tumor promoting effects of Cav-1 deficient fibroblasts could be functionally suppressed (nearly 2-fold) by the recombinant over-expression of SOD2 (superoxide dismutase 2), a known mitochondrial enzyme that de-activates superoxide, thereby reducing mitochondrial oxidative stress. In contrast, cytoplasmic soluble SOD1 had no effect, further highlighting a specific role for mitochondrial oxidative stress in this process. In summary, here we provide new evidence directly supporting a key role for a loss of stromal Cav-1 expression and oxidative stress in cancer-associated fibroblasts, in promoting tumor growth, which is consistent with "The Autophagic Tumor Stroma Model of Cancer". The human Cav-1 deficient fibroblasts that we have generated are a new genetically tractable model system for identifying other suppressors of the cancer-associated fibroblast phenotype, via a genetic "complementation" approach. This has important implications for understanding the pathogenesis of triple negative and basal breasts cancers, as well as tamoxifen-resistance in ER+ breast cancers, which are all associated with a Cav-1 deficient "lethal" tumor micro-environment, driving poor clinical outcome.

Dataset Information

Caveolin-1-/- null mammary stromal fibroblasts share characteristics with human breast cancer-associated fibroblasts.

Publications

Caveolin-1-/- null mammary stromal fibroblasts share characteristics with human breast cancer-associated fibroblasts.

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