Xuefen Le Bourhis

CD44⁺/CD24^– phenotype has been used to identify human and canine mammary cancer stem-like cells. In canine mammary tumors, CD44⁺/CD24^– phenotype has been associated with high grade and lymph node infiltration. However, several studies have reported opposing results regarding the clinical significance of phenotypic groups formed by the combination of CD44 and CD24 in both human and canine mammary tumors. So far, no study has investigated the correlation between these phenotypes and survival in dogs. The aim of this study was to investigate the expression and distribution of CD44 and CD24 in canine mammary carcinomas and to correlate them with histological diagnosis and survival in a well-characterized cohort. Immunohistochemistry was performed in 96 mammary carcinomas with antibodies against CD44 and CD24. Expression of CD44⁺ and CD44⁺/CD24^– phenotype was detected in 75 of 96 (78%) and 63 of 96 (65.6%) carcinomas, respectively. Their expression was associated with tumor type, occurring more often in tubular complex carcinomas than in solid carcinomas. CD44⁺/CD24^– phenotype was associated with a better overall survival ( P = .001). CD24⁺ expression was detected in 52 of 96 tumors (54%) and CD44^–/CD24⁺ phenotype in 39 of 96 tumors (40.6%). Both were associated with poor clinicopathological parameters (high grade, and emboli). No correlation with overall survival was observed. CD44⁺/CD24^– expression was associated with a better prognosis and occurred at high frequency and high level, indicating that this phenotype is not suitable to detect cancer stem cells in canine mammary carcinomas. Although further studies are needed, our results suggest that CD24 may constitute a valuable marker of poor prognosis for canine mammary carcinomas.

Cancer stem cells (CSCs), are thought to be at the origin of tumor development and resistance to therapies. Thus, a better understanding of the molecular mechanisms involved in the control of CSC stemness is essential to the design of more effective therapies for cancer patients. Cancer cell stemness and the subsequent expansion of CSCs are regulated by micro-environmental signals including neurotrophins. Over the years, the roles of neurotrophins in tumor development have been well established and regularly reviewed. Especially, nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) are reported to stimulate tumor cell proliferation, survival, migration and/or invasion, and favors tumor angiogenesis. More recently, neurotrophins have been reported to regulate CSCs. This review briefly presents neurotrophins and their receptors, summarizes their roles in different cancers, and discusses the emerging evidence of neurotrophins-induced enrichment of CSCs as well as the involved signaling pathways.

The discovery of cancer stem cells (CSCs) fundamentally advanced our understanding of the mechanisms governing breast cancer development. However, the stimuli that control breast CSC self‐renewal and differentiation have still not been fully detailed. We previously showed that nerve growth factor (NGF) and its precursor proNGF can stimulate breast cancer cell growth and invasion in an autocrine manner. In this study, we investigated the effects of NGF and proNGF on the breast CSC compartment and found that NGF or proNGF enrich for CSCs in several breast cancer cell lines. This enrichment appeared to be achieved by increasing the number of symmetric divisions of quiescent/slow‐proliferating CSCs. Interestingly, in vitro NGF pretreatment of MCF‐7 luminal breast cancer cells promoted epithelial to mesenchymal transition in tumors of severe combined immunodeficient mice. Furthermore, p75^NTR, the common receptor for both neurotrophins and proneurotrophins, mediated breast CSC self‐renewal by regulating the expression of pluripotency transcription factors. Our data indicate, for the first time, that the NGF/proNGF/p75^NTR axis plays a critical role in regulating breast CSC self‐renewal and plasticity. Stem Cells 2015;33:342–353

P75^NTR, the common receptor for both neurotrophins and proneurotrophins, has been widely studied because of its role in many tissues, including the nervous system. More recently, a close relationship between p75^NTR expression and pluripotency has been described. p75^NTR was shown to be expressed in various types of stem cells and has been used to prospectively isolate stem cells with different degrees of potency. Here, we give an overview of the current knowledge on p75^NTR in stem cells, ranging from embryonic to adult stem cells, and cancer stem cells. In an attempt to address its potential role in the control of stem cell biology, the molecular mechanisms underlying p75^NTR signaling in different models are also highlighted. p75^NTR-mediated functions include survival, apoptosis, migration, and differentiation, and depend on cell type, (pro)neurotrophin binding, interacting transmembrane co-receptors expression, intracellular adaptor molecule availability, and post-translational modifications, such as regulated proteolytic processing. It is therefore conceivable that p75^NTR can modulate cell-fate decisions through its highly ramified signaling pathways. Thus, elucidating the potential implications of p75^NTR activity as well as the underlying molecular mechanisms of p75^NTR will shed new light on the biology of both normal and cancer stem cells.

The p75 neurotrophin receptor (p75^NTR) undergoes sequential proteolytic cleavages leading to the generation of a carboxyl‐terminal fragment (p75^NTR‐CTF) and an intracellular domain (p75^NTR‐ICD) in many cellular models. We have previously shown that p75^NTR is involved in the survival of breast cancer cells. Here, we demonstrated that p75^NTR cleavage occurs also in these cells. Surprisingly, p75^NTR‐CTF increased cell survival, whereas p75^NTR‐ICD had no effect. The pro‐survival effect of p75^NTR‐CTF was associated with a decrease of TNF‐related apoptosis‐inducing ligand (TRAIL)‐induced PARP and caspase 3 cleavages. Finally, our findings indicate that p75^NTR could favor cell survival via its carboxyl‐terminal fragment, independently of PI3‐kinase, NF‐κB, or MAP kinase signaling pathways.

Background: Although several anti-angiogenic therapies have been approved in the treatment of cancer, the survival benefits of such therapies are relatively modest. Discovering new molecules and/or better understating signaling pathways of angiogenesis is therefore essential for therapeutic improvements. The objective of the present study was to determine the involvement of nerve growth factor (NGF) in breast cancer angiogenesis and the underlying molecular mechanisms. Results: We showed that both recombinant NGF and NGF produced by breast cancer cells stimulated angiogenesis in Matrigel plugs in immunodeficient mice. NGF strongly increased invasion, cord formation and the monolayer permeability of endothelial cells. Moreover, NGF-stimulated invasion was under the control of its tyrosine kinase receptor (TrkA) and downstream signaling pathways such as PI3K and ERK, leading to the activation of matrix metalloprotease 2 and nitric oxide synthase. Interestingly, NGF increased the secretion of VEGF in both endothelial and breast cancer cells. Inhibition of VEGF, with a neutralizing antibody, reduced about half of NGF-induced endothelial cell invasion and angiogenesis in vivo. Conclusions: Our findings provided direct evidence that NGF could be an important stimulator for breast cancer angiogenesis. Thus, NGF, as well as the activated signaling pathways, should be regarded as potential new targets for anti-angiogenic therapy against breast cancer.

Blood vessel formation (neovascularization) in tumors can occur through two mechanisms: angiogenesis and vasculogenesis. Angiogenesis results from proliferation and sprouting of existing blood vessels close to the tumor, while vasculogenesis is believed to arise from recruitment of circulating cells, largely derived from the bone marrow, and de novo clonal formation of blood vessels from these cells. Increasing evidence in animal models indicate that bone marrow-derived endothelial precursor cells (EPC) can contribute to tumor angiogenesis. This review aims to collate existing literature and provide an overview on the current knowledge of EPC involvement in breast cancer angiogenesis. We also discuss recent attempts to use EPC as biomarker and therapeutic target in clinical trials.

The Trk family of neurotrophin tyrosine kinase receptors is emerging as an important player in carcinogenic progression in non-neuronal tissues. Here, we show that breast tumors present high levels of TrkA and phospho-TrkA compared to normal breast tissues. To further evaluate the precise functions of TrkA overexpression in breast cancer development, we have performed a series of biological tests using breast cancer cells that stably overexpress TrkA. We show that (1) TrkA overexpression promoted cell growth, migration and invasion in vitro; (2) overexpression of TrkA per se conferred constitutive activation of its tyrosine kinase activity; (3) signal pathways including PI3K-Akt and ERK/p38 MAP kinases were activated by TrkA overexpression and were required for the maintenance of a more aggressive cellular phenotype; and (4) TrkA overexpression enhanced tumor growth, angiogenesis and metastasis of xenografted breast cancer cells in immunodeficient mice. Moreover, recovered metastatic cells from the lungs exhibited enhanced anoikis resistance that was abolished by the pharmacological inhibitor K252a, suggesting that TrkA-promoted breast tumor metastasis could be mediated at least in part by enhancing anoikis resistance. Together, these results provide the first direct evidence that TrkA overexpression enhances the tumorigenic properties of breast cancer cells and point to TrkA as a potential target in breast cancer therapy.

Tamoxifen (TAM), is widely used as a single agent in adjuvant treatment of breast cancer. Here, we investigated the effects of TAM in combination with tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in estrogen receptor-α (ER-α)-positive and -negative breast cancer cells. We showed that cotreatment with TAM and TRAIL synergistically induced apoptosis regardless of ER-α status. By contrast, cotreatment did not affect the viability of normal breast epithelial cells. Cotreatment with TAM and TRAIL in breast cancer cells decreased the levels of antiapoptotic proteins including FLIPs and Bcl-2, and enhanced the levels of proapoptotic proteins such as FADD, caspase 8, tBid, Bax and caspase 9. Furthermore, cotreatment-induced apoptosis was efficiently reduced by FADD- or Bid-siRNA, indicating the implication of both extrinsic and intrinsic pathways in synergistic apoptosis induction. Importantly, cotreatment totally arrested tumor growth in an ER-α-negative MDA-MB-231 tumor xenograft model. The abrogation of tumor growth correlated with enhanced apoptosis in tumor tissues. Our findings raise the possibility to use TAM in combination with TRAIL for breast cancers, regardless of ER-α status.

This study was performed to determine the effect and action mechanisms of sodium butyrate (NaB) on the growth of breast cancer cells. Butyrate inhibited the growth of all breast cancer cell lines analysed. It induced cell cycle arrest in G1 and apoptosis in MCF‐7, MCF‐7ras, T47‐D, and BT‐20 cells, as well as arrest in G2/M in MDA‐MB‐231 cells. Transient transfection of MCF‐7 and T47‐D cells with wild‐type and antisense p53 did not modify butyrate‐induced apoptosis. Pifithrin‐α, which inhibits the transcriptional activity of P53, did not modify cell growth or apoptosis of MCF‐7 and T47‐D cells treated with butyrate. These results indicate that P53 was not involved in butyrate‐induced growth inhibition of breast cancer cells. Treatment of MCF‐7 cells with anti‐Fas agonist antibody induced cell death, indicating that Fas was functional in these cells. Moreover, butyrate potentiated Fas‐induced apoptosis, as massive apoptosis was observed rapidly when MCF‐7 cells were treated with butyrate and anti‐Fas agonist antibody. In addition, butyrate‐induced apoptosis in MCF‐7 cells was considerably reduced by anti‐Fas antagonist antibody. Western blot analysis showed that butyrate increased Fas and Fas ligand levels (Fas L), indicating that butyrate‐induced apoptosis may be mediated by Fas signalling. These results demonstrate that butyrate inhibited the growth of breast cancer cells in a P53‐independent manner. Moreover, it induced apoptosis via the Fas/Fas L system and potentiated Fas‐triggered apoptosis in MCF‐7 cells. These findings may open interesting perspectives in human breast cancer treatment strategy. British Journal of Pharmacology (2002) 135, 79–86; doi:10.1038/sj.bjp.0704456