Induction of fibroblast senescence generates a non-fibrogenic myofibroblast phenotype that differentially impacts on cancer prognosis

Cancer-associated fibroblasts (CAF) remain a poorly characterized, heterogeneous cell population. Here we characterized two previously described tumor-promoting CAF sub-types, smooth muscle actin (SMA)-positive myofibroblasts and senescent fibroblasts, identifying a novel link between the two

Hepatocyte growth factor, a key tumor-promoting factor in the tumor microenvironment

The tumor microenvironment plays a key role in tumor development and progression. Stromal cells secrete growth factors, cytokines and extracellular matrix proteins which promote growth, survival and metastatic spread of cancer cells. Fibroblasts are the predominant constituent of the tumor stroma and Hepatocyte Growth Factor (HGF), the specific ligand for the tyrosine kinase receptor c-MET, is a major component of their secretome. Indeed, cancer-associated fibroblasts have been shown to promote growth, survival and migration of cancer cells in an HGF-dependent manner. Fibroblasts also confer resistance to anti-cancer therapy through HGF-induced epithelial mesenchymal transition (EMT) and activation of pro-survival signaling pathways such as ERK and AKT in tumor cells. Constitutive HGF/MET signaling in cancer cells is associated with increased tumor aggressiveness and predicts poor outcome in cancer patients. Due to its role in tumor progression and therapeutic resistance, both HGF and MET have emerged as valid therapeutic targets. Several inhibitors of MET and HGF are currently being tested in clinical trials. Preclinical data provide a strong indication that inhibitors of HGF/MET signaling overcome both primary and acquired resistance to EGFR, HER2, and BRAF targeting agents. These findings support the notion that co-targeting of cancer cells and stromal cells is required to prevent therapeutic resistance and to increase the overall survival rate of cancer patients. HGF dependence has emerged as a hallmark of therapeutic resistance, suggesting that inhibitors of biological activity of HGF should be included into therapeutic regimens of cancer patients

The Interplay between Colon Cancer Cells and Tumour-Associated Stromal Cells Impacts the Biological Clock and Enhances Malignant Phenotypes

Cancer cells interrelate with the bordering host microenvironment that encompasses the extracellular matrix and a nontumour cellular component comprising fibroblasts and immune-competent cells. The tumour microenvironment modulates cancer onset and progression, but the molecular factors managing this interaction are not fully understood. Malignant transformation of a benign tumour is among the first crucial events in colorectal carcinogenesis. The role of tumour stroma fibroblasts is well-described in cancer, but less well-characterized in benign tumours. In the current work we utilized fibroblasts isolated from tubulovillous adenoma, which has high risk for malignant transformation, to study the interaction between benign tumour stroma and the circadian clock machinery. We explored the role of the biological clock in this interplay taking advantage of an experimental model, represented by the co-culture of colon cancer cells with normal fibroblasts or tumour-associated fibroblasts, isolated from human colorectal tumour specimens. When co-cultured with tumour-associated fibroblasts, colon cancer cells showed alterations in their circadian and metabolic parameters, with decreased apoptosis, increased colon cancer cell viability, and increased resistance to chemotherapeutic agents. In conclusion, the interactions among colon cancer cells and tumour-associated fibroblasts affect the molecular clockwork and seem to aggravate malignant cell phenotypes, suggesting a detrimental effect of this interplay on cancer dynamics

Expression, Tissue Distribution and Function of miR-21 in Esophageal Squamous Cell Carcinoma

Objective:MiR-21 is an oncomir expressed by malignant cells and/or tumor microenvironment components. In this study we focused on understanding the effects of stromal miR-21 on esophageal malignant cells.Design:MiR-21 expression was evaluated in formalin-fixed paraffin-embedded samples from patients with esophageal squamous-cell carcinoma (SCC) by quantitative RT-PCR. MiR-21 tissue distribution was visualized with in situ hybridization. A co-culture system of normal fibroblasts and esophageal cancer cells was used to determine the effects of fibroblasts on miR-21 expression levels, and on SCC cell migration and invasion.Results:MiR-21 was overexpressed in SCCs, when compared to the adjacent non-tumor tissues (P = 0.0007), and was mainly localized in the cytoplasm of stromal cells adjacent to malignant cells. Accordingly, miR-21 expression was increased in tumors with high versus low stromal content (P = 0.04). When co-cultured with normal fibroblasts, miR-21 expression was elevated in SCC cells (KYSE-30), while its expression was restricted to fibroblasts when co-cultured with adenocarcinoma cells (OE-33 and FLO-1). MiR-21 was detected in conditioned media of cancer cell lines, illustrating the release of this miRNA into the environment. Co-culturing with normal fibroblasts or addition of fibroblast conditioned media caused a significant increase in cell migration and invasion potency of KYSE-30 cells (P<0.0001). In addition, co-culturing cancer cells with fibroblasts and expression of miR-21 induced the expression of the cancer associated fibroblast (CAF) marker S100A4.Conclusions:MiR-21 expression is mostly confined to the SCC stroma and its release from fibroblasts influences the migration and invasion capacity of SCC cells. Moreover, miR-21 may be an important factor in "activating" fibroblasts to CAFs. These findings provide new insights into the role of CAFs and the extracellular matrix in tumor microenvironment formation and in tumor cell maintenance, and suggest miR-21 may contribute to cellular crosstalk in the tumor microenvironment. © 2013 Nouraee et al

α-Smooth muscle actin expression in cancerassociated fibroblasts in canine epithelial tumors

Tumor tissues contain not only cancer cells but also other cell types including, fibroblasts, immune cells, and endothelial cells, which interact with cancer cells. In human medicine, cancer-associated fibroblasts (CAFs) have been reported to promote tumor growth. CAFs are known to express α-smooth muscle actin (α-SMA), and this expression is correlated with poor prognosis in humans with cancer. However, the role of CAFs in canines and α-SMA expression in canine CAFs remains unknown. This study evaluated whether CAFs are present within the stroma of various types of canine epithelial tumors, for example, mammary gland tumors, squamous cell carcinoma, and anal sac adenocarcinoma, and assessed α-SMA expression in CAFs isolated from canine epithelial tumors. α-SMA analysis of tumor tissues revealed a cytoplasmic localization with variable levels of expression. α-SMA was detected in 60.9% (14/23) of epithelial tumor tissues and in 80% (8/10) of anal sac adenocarcinoma tissues. CAFs and normal fibroblasts (NFs) were isolated from tumor and skin tissues. The size of CAFs was variable, and most CAFs had large cell volume, in contrast to NFs. Most CAFs expressed α-SMA stress fibers and had higher α-SMA protein levels than NFs. Taken together, our findings provide evidence that canine CAFs express α-SMA in various canine epithelial tumors. Further studies are required to investigate the correlation between canine CAFs and clinical parameters and to elucidate the mechanisms underlying the effects of CAFs on cancer progression

The fibroblast Tiam1-osteopontin pathway modulates breast cancer invasion and metastasis

Background The tumor microenvironment has complex effects in cancer pathophysiology that are not fully understood. Most cancer therapies are directed against malignant cells specifically, leaving pro-malignant signals from the microenvironment unaddressed. Defining specific mechanisms by which the tumor microenvironment contributes to breast cancer metastasis may lead to new therapeutic approaches against advanced breast cancer. Methods We use a novel method for manipulating three-dimensional mixed cell co-cultures, along with studies in mouse xenograft models of human breast cancer and a histologic study of human breast cancer samples, to investigate how breast cancer-associated fibroblasts affect the malignant behaviors of breast cancer cells. Results Altering fibroblast Tiam1 expression induces changes in invasion, migration, epithelial-mesenchymal transition, and cancer stem cell characteristics in associated breast cancer cells. These changes are both dependent on fibroblast secretion of osteopontin and also long-lasting even after cancer cell dissociation from the fibroblasts, indicating a novel Tiam1-osteopontin pathway in breast cancer-associated fibroblasts. Notably, inhibition of fibroblast osteopontin with low doses of a novel small molecule prevents lung metastasis in a mouse model of human breast cancer metastasis. Moreover, fibroblast expression patterns of Tiam1 and osteopontin in human breast cancers show converse changes correlating with invasion, supporting the hypothesis that this pathway in tumor-associated fibroblasts regulates breast cancer invasiveness in human disease and is thus clinically relevant. Conclusions These findings suggest a new therapeutic paradigm for preventing breast cancer metastasis. Pro-malignant signals from the tumor microenvironment with long-lasting effects on associated cancer cells may perpetuate the metastatic potential of developing cancers. Inhibition of these microenvironment signals represents a new therapeutic strategy against cancer metastasis that enables targeting of stromal cells with less genetic plasticity than associated cancer cells and opens new avenues for investigation of novel therapeutic targets and agents.National Institute of General Medical Sciences (U.S.) (GM074825

Heterogeneous cancer-associated fibroblast population potentiates neuroendocrine differentiation and castrate resistance in a CD105-dependent manner.

Heterogeneous prostatic carcinoma-associated fibroblasts (CAF) contribute to tumor progression and resistance to androgen signaling deprivation therapy (ADT). CAF subjected to extended passaging, compared to low passage CAF, were found to lose tumor expansion potential and heterogeneity. Cell surface endoglin (CD105), known to be expressed on proliferative endothelia and mesenchymal stem cells, was diminished in high passage CAF. RNA-sequencing revealed SFRP1 to be distinctly expressed by tumor-inductive CAF, which was further demonstrated to occur in a CD105-dependent manner. Moreover, ADT resulted in further expansion of the CD105+ fibroblastic population and downstream SFRP1 in 3-dimensional cultures and patient-derived xenograft tissues. In patients, CD105+ fibroblasts were found to circumscribe epithelia with neuroendocrine differentiation. CAF-derived SFRP1, driven by CD105 signaling, was necessary and sufficient to induce prostate cancer neuroendocrine differentiation in a paracrine manner. A partially humanized CD105 neutralizing antibody, TRC105, inhibited fibroblastic SFRP1 expression and epithelial neuroendocrine differentiation. In a novel synthetic lethality paradigm, we found that simultaneously targeting the epithelia and its microenvironment with ADT and TRC105, respectively, reduced castrate-resistant tumor progression, in a model where either ADT or TRC105 alone had little effect

Remodeling of extracellular matrix by normal and tumor-associated fibroblasts promotes cervical cancer progression

Background: Comparison of tissue microarray results of 29 cervical cancer and 27 normal cervix tissue samples using immunohistochemistry revealed considerable reorganization of the fibrillar stroma of these tumors. Preliminary densitometry analysis of laminin-1, α -smooth muscle actin (SMA) and fibronectin immunostaining demonstrated 3.8-fold upregulation of laminin-1 and 5.2-fold increase of SMA in the interstitial stroma, indicating that these proteins and the activated fibroblasts play important role in the pathogenesis of cervical cancer. In the present work we investigated the role of normal and tumor-associated fibroblasts. Methods: In vitro models were used to throw light on the multifactorial process of tumor-stroma interaction, by means of studying the cooperation between tumor cells and fibroblasts. Fibroblasts from normal cervix and cervical cancers were grown either separately or in co-culture with CSCC7 cervical cancer cell line. Changes manifest in secreted glycoproteins, integrins and matrix metallo-proteases (MMPs) were explored. Results: While normal fibroblasts produced components of interstitial matrix and TGF- β 1 that promoted cell proliferation, cancer-associated fibroblasts (CAFs) synthesized ample amounts of laminin-1. The following results support the significance of laminin-1 in the invasion of CSCC7 cells: 1.) Tumor-associated fibroblasts produced more laminin-1 and less components of fibrillar ECM than normal cells; 2.) The production of laminin chains was further increased when CSCC7 cells were grown in co-culture with fibroblasts; 3.) CSCC7 cells were capable of increasing their laminin production; 4.) Tumor cells predominantly expressed integrin α 6 β 4 laminin receptors and migrated towards laminin. The integrin profile of both normal and tumor-associated fibroblasts was similar, expressing receptors for fibronectin, vitronectin and osteopontin. MMP-7 secreted by CSCC7 cells was upregulated by the presence of normal fibroblasts, whereas MMP-2 produced mainly by fibroblasts was activated in the presence of CSCC7 cells. Conclusions: Our results indicate that in addition to degradation of the basement membrane, invasion of cervical cancer is accomplished by the remodeling of the interstitial stroma, which process includes decrease and partial replacement of fibronectin and collagens by a laminin-rich matrix