Liver X receptors induce antiproliferative effects in basal-like breast cancer.

Liver X receptors (LXRs) are nuclear transcription factors important in the regulation of cholesterol transport, and glucose and fatty acid metabolism. The antiproliferative role of LXRs has been studied in a variety of malignancies and may represent a therapeutic opportunity in cancers lacking targeted therapies, such as triple-negative breast cancer. In this study, we investigated the impact of LXR agonists alone and in combination with carboplatin in preclinical models of breast cancer. In vitro experiments revealed a dose-dependent decrease in tumor cell proliferation in estrogen receptor-positive breast cancer cells, whereas LXR activation in vivo resulted in an increased growth inhibitory effect in a basal-like breast cancer model (in combination with carboplatin). Functional proteomic analysis identified differences in protein expression between responding and nonresponding models related to Akt activity, cell-cycle progression, and DNA repair. Furthermore, pathway analysis suggested that the LXR agonist in combination with carboplatin inhibits the activity of targets of E2F transcription factors and affects cholesterol homeostasis in basal-like breast cancer.

Basal-like breast cancer engages tumor-supportive macrophages via secreted factors induced by extracellular S100A4.

The tumor microenvironment (TME) may influence both cancer progression and therapeutic response. In breast cancer, particularly in the aggressive triple-negative/basal-like subgroup, patient outcome is strongly associated with the tumor's inflammatory profile. Tumor-associated macrophages (TAMs) are among the most abundant immune cells in the TME, shown to be linked to poor prognosis and therapeutic resistance. In this study, we investigated the effect of the metastasis- and inflammation-associated microenvironmental factor S100A4 on breast cancer cells (BCCs) of different subtypes and explored their further interactions with myeloid cells. We demonstrated that extracellular S100A4 activates BCCs, particularly the basal-like subtype, to elevate secretion of pro-inflammatory cytokines. The secreted factors promoted conversion of monocytes to TAM-like cells that exhibited protumorigenic activities: stimulated epithelial-mesenchymal transition, proliferation, chemoresistance, and motility in cancer cells. In conclusion, we have shown that extracellular S100A4 instigates a tumor-supportive microenvironment, involving a network of cytokines and TAM-like cells, which was particularly characteristic for basal-like BCCs and potentiated their aggressive properties. The S100A4-BCC-TAM interaction cascade could be an important contributor to the aggressive behavior of this subtype and should be further explored for therapeutic targeting.

Wnt expression and canonical Wnt signaling in human bone marrow B lymphopoiesis.

BACKGROUND: The early B lymphopoiesis in mammals is regulated through close interactions with stromal cells and components of the intracellular matrix in the bone marrow (BM) microenvironment. Although B lymphopoiesis has been studied for decades, the factors that are implicated in this process, both autocrine and paracrine, are inadequately explored. Wnt signaling is known to be involved in embryonic development and growth regulation of tissues and cancer. Wnt molecules are produced in the BM, and we here ask whether canonical Wnt signaling has a role in regulating human BM B lymphopoiesis. RESULTS: Examination of the mRNA expression pattern of Wnt ligands, Fzd receptors and Wnt antagonists revealed that BM B progenitor cells and stromal cells express a set of ligands and receptors available for induction of Wnt signaling as well as antagonists for fine tuning of this signaling. Furthermore, different B progenitor maturation stages showed differential expression of Wnt receptors and co-receptors, beta-catenin, plakoglobin, LEF-1 and TCF-4 mRNAs, suggesting canonical Wnt signaling as a regulator of early B lymphopoiesis. Exogenous Wnt3A induced stabilization and nuclear accumulation of beta-catenin in primary lineage restricted B progenitor cells. Also, Wnt3A inhibited B lymphopoiesis of CD133+CD10- hematopoietic progenitor cells and CD10+ B progenitor cells in coculture assays using a supportive layer of stromal cells. This effect was blocked by the Wnt antagonists sFRP1 or Dkk1. Examination of early events in the coculture showed that Wnt3A inhibits cell division of B progenitor cells. CONCLUSION: These results indicate that canonical Wnt signaling is involved in human BM B lymphopoiesis where it acts as a negative regulator of cell proliferation in a direct or stroma dependent manner.

From autonomy to community; new perspectives on tumorigenicity and therapy resistance.

Subclones of cancer cells evading treatment represent the major challenge in oncology. Despite recent advances, tumors not responding to treatments are still a severe risk to cancer patients, and oncologists have, as of now, little effective therapy to offer patients with systemic cancer disease. The widely discussed cancer stem cell (CSC) paradigm was originally launched as an explanation to the existence of small cell populations resistant to therapy within the heterogeneous tumor, but has so far unfortunately, offered little concrete improvement in cancer treatment regimes. The launch of the CSC hypothesis did, however, highlight the significance of therapy targeting specific tumor-driving processes, and even more importantly, an increased awareness of a phenomenon well known to stem cell researchers; non-genetic phenotypic heterogeneity of cells with common origin. Here, the scientific background of the CSC theory is revisited and the evidence for CSCs is discussed, along with the importance of considering CSC's dependency of their habitat for survival and growth. Furthermore, recent advances in cancer cell heterogeneity and new possibilities for studying therapy responses in cell clones within the natural tumor environment using patient derived xenograft (PDX) models, are reviewed.

Metabolic reprogramming supports the invasive phenotype in malignant melanoma.

Invasiveness is a hallmark of aggressive cancer like malignant melanoma, and factors involved in acquisition or maintenance of an invasive phenotype are attractive targets for therapy. We investigated melanoma phenotype modulation induced by the metastasis-promoting microenvironmental protein S100A4, focusing on the relationship between enhanced cellular motility, dedifferentiation and metabolic changes. In poorly motile, well-differentiated Melmet 5 cells, S100A4 stimulated migration, invasion and simultaneously down-regulated differentiation genes and modulated expression of metabolism genes. Metabolic studies confirmed suppressed mitochondrial respiration and activated glycolytic flux in the S100A4 stimulated cells, indicating a metabolic switch toward aerobic glycolysis, known as the Warburg effect. Reversal of the glycolytic switch by dichloracetate induced apoptosis and reduced cell growth, particularly in the S100A4 stimulated cells. This implies that cells with stimulated invasiveness get survival benefit from the glycolytic switch and, therefore, become more vulnerable to glycolysis inhibition. In conclusion, our data indicate that transition to the invasive phenotype in melanoma involves dedifferentiation and metabolic reprogramming from mitochondrial oxidation to glycolysis, which facilitates survival of the invasive cancer cells. Therapeutic strategies targeting the metabolic reprogramming may therefore be effective against the invasive phenotype.

Metastasis-associated protein S100A4 induces a network of inflammatory cytokines that activate stromal cells to acquire pro-tumorigenic properties.

Tumor cells have the ability to exploit stromal cells to facilitate metastasis. By using malignant melanoma as a model, we show that the stroma adjacent to metastatic lesions is enriched in the known metastasis-promoting protein S100A4. S100A4 stimulates cancer cells to secrete paracrine factors, such as inflammatory cytokines IL8, CCL2 and SAA, which activate stromal cells (endothelial cells and monocytes) so that they acquire tumor-supportive properties. Our data establishes S100A4 as an inducer of a cytokine network enabling tumor cells to engage angiogenic and inflammatory stromal cells, which might contribute to pro-metastatic activity of S100A4.

Extensive adipogenic and osteogenic differentiation of patterned human mesenchymal stem cells in a microfluidic device.

Microtechnology offers great prospects for cellular research by enabling controlled experimental conditions that cannot be achieved by traditional methods. This study demonstrates the use of a microfluidic platform for long-term cultivation (3 weeks) of human mesenchymal stem-like cells (MSCs), a cell population of high interest for tissue engineering. The typical high motility of the MSCs required a strategy for preventing cells from inhabiting the feeding channels and thus interfere with a steady perfusion of medium to the cell cultivation chamber. Hence, a straightforward and long-term patterning method was developed and implemented for reliable cell positioning within the device. This method was based on the modification of a polystyrene substrate into cell supportive and non-supportive regions by the use of selective oxygen plasma treatment and the triblock copolymer Pluronic. Also, a novel and size-effective "flip-chip" set-up for operating the devices was invented. Successful and reproducible adipogenic and osteogenic differentiation of MSCs in the device was demonstrated, verifying that an adequate long-term microfluidic cultivation environment was obtained. Strengths of the experimental protocol include ease of fabrication and maintenance (gravity driven), good cell performance (viability/differentiation), as well as the possibility of exposing the culture to heterogeneous laminar flow for experimental purposes.

CD10+ stromal cells form B-lymphocyte maturation niches in the human bone marrow.

In adult mammals, early B-lymphopoiesis takes place in the bone marrow in close association with stromal cells. Both the phenotype of the stromal cells and the molecules involved in this essential interaction are as yet inadequately described. In this study, all benign, differentiating B-cells (Pax-5+ lymphoid cells) are shown, by using two-colour immunohistochemistry on biopsies from human bone marrow, to be in close contact with scant dendritic CD10+ stromal cells until they leave via the sinusoids. This CD10+ stromal cell population does not fully overlap with the VCAM-1+ stromal cell population. Furthermore, using a set of B-cell differentiation markers (TdT, Pax-5, and CD20), B-cell development is shown to be spatially oriented, with maturation progressing towards bone marrow sinusoids. In conclusion, CD10+ stromal cells form distinct B-lymphocyte maturation niches in the human bone marrow.

Characterization of the novel human transmembrane protein 9 (TMEM9) that localizes to lysosomes and late endosomes.

We have identified and characterized the novel human transmembrane protein 9 (TMEM9). TMEM9 encodes a 183 amino-acid protein that contains an N-terminal signal peptide, a single transmembrane region, three potential N-glycosylation sites, and three conserved cys-rich domains in the N-terminus, but no hitherto known functional domains. The protein is highly conserved between species from Caenorhabditis elegans to man and belongs to a novel family of transmembrane proteins. The TMEM9 gene consists of at least 6 exons and is localized to chromosome 1q41. TMEM9 mRNA is expressed in a wide range of tissues and cells. COS-1 cells transfected with a TMEM9 expression plasmid gave three bands of about 28, 31, and 33kDa representing glycosylated forms of TMEM9 with a protein backbone of about 26kDa. In COS-1 cells transfected with a TMEM9-GFP expression construct,TMEM9-GFP is co-expressed with LAMP1 on late endosomes and lysosomes as well as on ER. Thus, TMEM9 is a phylogenetically conserved, widely expressed transmembrane protein with a potential, but unknown function in intracellular transport.