MicroRNAs and breast cancer stem cells: Potential role in breast cancer therapy

MicroRNAs (miRNAs) can control cancer and cancer stem cells (CSCs), and this topic has drawn immense attention recently. Stem cells are a tiny population of a bulk of tumor cells that have enormous potential in expansion and metastasis of the tumor. miRNA have a crucial role in the management of the function of stem cells. This role is to either promote or suppress the tumor. In this review, we investigated the function and different characteristics of CSCs and function of the miRNAs that are related to them. We also demonstrated the role and efficacy of these miRNAs in breast cancer and breast cancer stem cells (BCSC). Eventually, we revealed the metastasis, tumor formation, and their role in the apoptosis process. Also, the therapeutic potential of miRNA as an effective method for the treatment of BCSC was described. Extensive research is required to investigate the employment or suppression of these miRNAs for therapeutics approached in different cancers in the future.     

乳腺癌干细胞分选及相关信号通路研究进展

肿瘤干细胞是指存在于肿瘤组织中的具有干细胞特性,即能够多向分化和自我更新的一类细胞群。随着肿瘤干细胞概念的提出,乳腺癌干细胞成为当今科研领域的一个研究热点。因此,了解如何分选乳腺癌干细胞及如何维持其"干性"对治疗及预防乳腺癌具有至关重要的意义。主要从乳腺癌干细胞分选、相关信号通路、上皮-间充质转换(EMT)等方面进行综述。     

Modulatory roles of microRNAs in the regulation of different signalling pathways in large bowel cancer stem cells

There are emerging data to suggest that microRNAs (miRNAs) have significant roles in regulating the function of normal cells and cancer stem cells (CSCs). This review aims to analyse the roles of miRNAs in the regulation of colon CSCs through their interaction with various signalling pathways. Studies showed a large number of miRNAs that are reported to be deregulated in colon CSCs. However, few of the studies available were able to outline the function of miRNAs in colon CSCs and uncover their signalling pathways. From those miRNAs, which are better described, miR‐21 followed by miR‐34, miR‐200 and miR‐215 are the most reported miRNAs to have roles in colon CSC regulation. In particular, miRNAs have been reported to regulate the stemness features of colon CSCs mainly via Wnt/B‐catenin and Notch signalling pathways. Additionally, miRNAs have been reported to act on processes involving CSCs through cell cycle regulation genes and epithelial–mesenchymal transition. The relative paucity of data available on the significance of miRNAs in CSCs means that new studies will be of great importance to determine their roles and to identify the signalling pathways through which they operate. Such studies may in future guide further research to target these genes for more effective cancer treatment. miRNAs were shown to regulate the function of cancer stem cells in large bowel cancer by targeting a few key signalling pathways in cells.     

RUNX1 and RUNX2 transcription factors function in opposing roles to regulate breast cancer stem cells

Breast cancer stem cells (BCSCs) are competent to initiate tumor formation and growth and refractory to conventional therapies. Consequently BCSCs are implicated in tumor recurrence. Many signaling cascades associated with BCSCs are critical for epithelial-to-mesenchymal transition (EMT). We developed a model system to mechanistically examine BCSCs in basal-like breast cancer using MCF10AT1 FACS sorted for CD24 (negative/low in BCSCs) and CD44 (positive/high in BCSCs). Ingenuity Pathway Analysis comparing RNA-seq on the CD24^−/low versus CD24^+/high MCF10AT1 indicates that the top activated upstream regulators include TWIST1, TGFβ1, OCT4, and other factors known to be increased in BCSCs and during EMT. The top inhibited upstream regulators include ESR1, TP63, and FAS. Consistent with our results, many genes previously demonstrated to be regulated by RUNX factors are altered in BCSCs. The RUNX2 interaction network is the top significant pathway altered between CD24^−/low and CD24^+/high MCF10AT1. RUNX1 is higher in expression at the RNA level than RUNX2. RUNX3 is not expressed. While, human-specific quantitative polymerase chain reaction primers demonstrate that RUNX1 and CDH1 decrease in human MCF10CA1a cells that have grown tumors within the murine mammary fat pad microenvironment, RUNX2 and VIM increase. Treatment with an inhibitor of RUNX binding to CBFβ for 5 days followed by a 7-day recovery period results in EMT suggesting that loss of RUNX1, rather than increase in RUNX2, is a driver of EMT in early stage breast cancer. Increased understanding of RUNX regulation on BCSCs and EMT will provide novel insight into therapeutic strategies to prevent recurrence.     

Adipose-derived stem cells and cancer cells fuse to generate cancer stem cell-like cells with increased tumorigenicity

Adipose-derived stem cells (ADSCs) are a type of mesenchymal stem cells isolated from adipose tissue and have the ability to differentiate into adipogenic, osteogenic, and chondrogenic lineages. Despite their great therapeutic potentials, previous studies showed that ADSCs could enhance the proliferation and metastatic potential of breast cancer cells (BCCs). In this study, we found that ADSCs fused with BCCs spontaneously, while breast cancer stem cell (CSC) markers CD44⁺CD24^-/lowEpCAM⁺ were enriched in this fusion population. We further assessed the fusion hybrid by multicolor DNA FISH and mouse xenograft assays. Only single nucleus was observed in the fusion hybrid, confirming that it was a synkaryon. In vivo mouse xenograft assay indicated that the tumorigenic potential of the fusion hybrid was significantly higher than that of the parent tumorigenic triple-negative BCC line MDA-MB-231. We had compared the fusion efficiency between two BCC lines, the CD44-rich MDA-MB-231 and the CD44-poor MCF-7, with ADSCs. Interestingly, we found that the fusion efficiency was much higher between MDA-MB-231 and ADSCs, suggesting that a potential mechanism of cell fusion may lie in the dissimilarity between these two cell lines. The cell fusion efficiency was hampered by knocking down the CD44. Altogether, our findings suggest that CD44-mediated cell fusion could be a potential mechanism for generating CSCs.     

Influence of adjuvant radiotherapy on circulating epithelial tumor cells and circulating cancer stem cells in primary non-metastatic breast cancer

Background: There is an unmet need to identify biomarkers that directly reflect response to adjuvant radiotherapy (RT). Circulating epithelial tumor cells (CETCs) represent the liquid component of solid tumors and are responsible for metastatic relapse. CETC subsets with cancer stem cell characteristics, circulating cancer stem cells (cCSCs), play a pivotal role in the metastatic cascade. Monitoring the most aggressive subpopulation of CETCs could reflect the aggressiveness of the remaining tumor burden. There is limited data on the detection and monitoring changes in CETC and cCSC numbers during RT in early breast cancer.Methods: CETC numbers were analyzed prior to, at midterm and at the end of RT in 52 primary non-metastatic breast cancer patients. Hormone receptor status was determined in CETCs prior to and at the end of RT. For the identification of cCSCs cell suspensions from the peripheral blood of patients were cultured in vitro under conditions favoring growth of tumorspheres.Results: Hormone receptor status in CETCs before RT was comparable to that in primary tumor tissue. Prior to RT numbers of CETCs correlated with aggressiveness of primary tumors. cCSCs could be successfully identified and monitored during RT. Prior to RT patients treated with neoadjuvant chemotherapy had significantly higher numbers of CETCs and tumorspheres compared to patients after adjuvant chemotherapy. During RT, the number of CETCs decreased continuously in patients after neoadjuvant chemotherapy but not after adjuvant chemotherapy.Conclusion: Monitoring the number of CETCs and the CETC subset with cancer stem cell properties during RT may provide additional clinically useful prognostic information.     

From milk to malignancy: the role of mammary stem cells in development, pregnancy and breast cancer

Adult stem cells of the mammary gland (MaSCs) are a highly dynamic population of cells that are responsible for the generation of the gland during puberty and its expansion during pregnancy. In recent years significant advances have been made in understanding how these cells are regulated during these developmentally important processes both in humans and in mice. Understanding how MaSCs are regulated is becoming a particularly important area of research, given that they may be particularly susceptible targets for transformation in breast cancer. Here, we summarize the identification of MaSCs, how they are regulated and the evidence for their serving as the origins of breast cancer. In particular, we focus on how changes in MaSC populations may explain both the increased risk of developing aggressive ER/PR(-) breast cancer shortly after pregnancy and the long-term decreased risk of developing ER/PR(+) tumors.     

Mesenchymal traits are selected along with stem features in breast cancer cells grown as mammospheres

Increasing evidence indicates that invasive properties of breast cancers rely on gain of mesenchymal and stem features, which has suggested that the dual targeting of these phenotypes may represent an appealing therapeutic strategy. It is known that the fraction of stem cells can be enriched by culturing breast cancer cells as mammospheres (MS), but whether these pro-stem conditions favor also the expansion of cells provided of mesenchymal features is still undefined. In the attempt to shed light on this issue, we compared the phenotypes of a panel of 10 breast cancer cell lines representative of distinct subtypes (luminal, HER2-positive, basal-like and claudin-low), grown in adherent conditions and as mammospheres. Under MS-proficient conditions, the increment in the fraction of stem-like cells was associated to upregulation of the mesenchymal marker Vimentin and downregulation of the epithelial markers expressed by luminal cells (E-cadherin, KRT18, KRT19, ESR1). Luminal cells tended also to upregulate the myoepithelial marker CD10. Taken together, our data indicate that MS-proficient conditions do favor mesenchymal/myoepithelial features, and indicate that the use of mammospheres as an in vitro tumor model may efficiently allow the exploitation of therapeutic approaches aimed at targeting aggressive tumors that have undergone epithelial-to-mesenchymal transition.     

Mesenchymal traits are selected along with stem features in breast cancer cells grown as mammospheres

Increasing evidence indicates that invasive properties of breast cancers rely on gain of mesenchymal and stem features, which has suggested that the dual targeting of these phenotypes may represent an appealing therapeutic strategy. It is known that the fraction of stem cells can be enriched by culturing breast cancer cells as mammospheres (MS), but whether these pro-stem conditions favor also the expansion of cells provided of mesenchymal features is still undefined.

In the attempt to shed light on this issue, we compared the phenotypes of a panel of 10 breast cancer cell lines representative of distinct subtypes (luminal, HER2-positive, basal-like and claudin-low), grown in adherent conditions and as mammospheres. Under MS-proficient conditions, the increment in the fraction of stem-like cells was associated to upregulation of the mesenchymal marker Vimentin and downregulation of the epithelial markers expressed by luminal cells (E-cadherin, KRT18, KRT19, ESR1). Luminal cells tended also to upregulate the myoepithelial marker CD10. Taken together, our data indicate that MS-proficient conditions do favor mesenchymal/myoepithelial features, and indicate that the use of mammospheres as an in vitro tumor model may efficiently allow the exploitation of therapeutic approaches aimed at targeting aggressive tumors that have undergone epithelial-to-mesenchymal transition.     

microRNAs in breast cancer: regulatory roles governing the hallmarks of cancer

A large number of etiological factors and the complexity of breast cancers present challenges for prevention and treatment. Recently, the emergence of microRNAs (miRNAs) as cancer biomarkers has added an extra dimension to the ‘molecular signatures’ of breast cancer. Bioinformatic analyses indicate that each miRNA can regulate hundreds of target genes and could serve functionally as ‘oncogenes’ or ‘tumour suppressor’ genes, and co‐ordinate multiple cellular processes relevant to cancer progression. A number of studies have shown that miRNAs play important roles in breast tumorigenesis, metastasis, proliferation and differentiation of breast cancer cells. This review provides a comprehensive overview of miRNAs with established functional relevance in breast cancer, their established target genes and resulting cellular phenotype. The role and application of circulating miRNAs in breast cancer is also discussed. Furthermore, we summarize the role of miRNAs in the hallmarks of breast cancer, as well as the possibility of using miRNAs as potential biomarkers for detection of breast cancer.     

Doxycycline inhibits the cancer stem cell phenotype and epithelial-to-mesenchymal transition in breast cancer

Experimental evidence suggest that breast tumors originate from breast cancer stem cells (BCSCs), and that mitochondrial biogenesis is essential for the anchorage-independent clonal expansion and survival of CSCs, thus rendering mitochondria a significant target for novel treatment approaches. One of the recognized side effects of the FDA-approved drug, doxycycline is the inhibition of mitochondrial biogenesis. Here we investigate the mechanism by which doxycycline exerts its inhibitory effects on the properties of breast cancer cells and BCSCs, such as mammosphere forming efficiency, invasion, migration, apoptosis, the expression of stem cell markers and epithelial-to-mesenchymal transition (EMT) related markers of breast cancer cells. In addition, we explored whether autophagy plays a role in the inhibitory effect of doxycycline on breast cancer cells. We find that doxycyline can inhibit the viability and proliferation of breast cancer cells and BCSCs, decrease mammosphere forming efficiency, migration and invasion, and EMT of breast cancer cells. Expression of stem cell factors Oct4, Sox2, Nanog and CD44 were also significantly downregulated after doxycycline treatment. Moreover, doxycycline could down-regulate the expression of the autophagy marker LC-3BI and LC-3BII, suggesting that inhibiting autophagy may be responsible in part for the observed effects on proliferation, EMT and stem cell markers. The potent inhibition of EMT and cancer stem-like characteristics in breast cancer cells by doxycycline treatment suggests that this drug can be repurposed as an anti-cancer drug in the treatment of breast cancer patients in the clinic.     

Regulation of angiogenesis via Notch signaling in breast cancer and cancer stem cells

Breast cancer angiogenesis is elicited and regulated by a number of factors including the Notch signaling. Notch receptors and ligands are expressed in breast cancer cells as well as in the stromal compartment and have been implicated in carcinogenesis. Signals exchanged between neighboring cells through the Notch pathway can amplify and consolidate molecular differences, which eventually dictate cell fates. Notch signaling and its crosstalk with many signaling pathways play an important role in breast cancer cell growth, migration, invasion, metastasis and angiogenesis, as well as cancer stem cell (CSC) self-renewal. Therefore, significant attention has been paid in recent years toward the development of clinically useful antagonists of Notch signaling. Better understanding of the structure, function and regulation of Notch intracellular signaling pathways, as well as its complex crosstalk with other oncogenic signals in breast cancer cells will be essential to ensure rational design and application of new combinatory therapeutic strategies. Novel opportunities have emerged from the discovery of Notch crosstalk with inflammatory and angiogenic cytokines and their links to CSCs. Combinatory treatments with drugs designed to prevent Notch oncogenic signal crosstalk may be advantageous over λ secretase inhibitors (GSIs) alone. In this review, we focus on the more recent advancements in our knowledge of aberrant Notch signaling contributing to breast cancer angiogenesis, as well as its crosstalk with other factors contributing to angiogenesis and CSCs.     

乳腺癌干细胞分子特性及其治疗新策略

乳腺癌是一种严重影响女性健康甚至危及生命的疾病。环境、饮食习惯、疾病等均有可能诱发人类乳腺细胞癌基因活化或抑癌基因失活,其诱发过程可通过细胞、动物模型所证实。最新研究指出,乳腺癌肿瘤干细胞的分子特性研究为乳腺癌发生、发展的认识提供了重要的佐证。另外,MicroRNAs在乳腺癌中表现促进或抑制瘤细胞生长与分化。这些研究的深入开展将为乳腺癌的防治提供新思路。     

Identification of single chain antibodies to breast cancer stem cells using phage display

Recent evidence suggests that most malignancies are driven by “cancer stem cells” sharing the signature characteristics of adult stem cells: the ability to self renew and to differentiate. Furthermore these cells are thought to be quiescent, infrequently dividing cells with a natural resistance to chemotherapeutic agents. These studies theorize that therapies, which effectively treat the majority of tumor cells but ‘miss’ the stem cell population, will fail, while therapies directed at stern cells can potentially eradicate tumors. In breast cancer, researchers have isolated ‘breast cancer stem cells’ capable of recreating the tumor in vivo and in vitro. Generated new tumors contained both additional numbers of cancer stem cells and diverse mixed populations of cells present in the initial tumor, supporting the intriguing self‐renewal and differentiation characteristics. In the present study, an antibody phage library has been used to search for phage displayed‐single chain antibodies (scFv) with selective affinity to specific targets on breast cancer stem cells. We demonstrate evidence of two clones binding specifically to a cancer stem cell population isolated from the SUMl59 breast cancer cell line. These clones had selective affinity for cancer stem cells and they were able to select cancer stem cells among a large population of non‐stem cancer cells in paraffin‐embedded sections. The applicability of these clones to paraffin sections and frozen tissue specimens made them good candidates to be used as diagnostic and prognostic markers in breast cancer patient samples taking into consideration the cancer stern cell concept in tumor biology. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009     

Role of platelets and breast cancer stem cells in metastasis

The high mortality rate of breast cancer is mainly caused by the metastatic ability of cancer cells, resistance to chemotherapy and radiotherapy, and tumor regression capacity. In recent years, it has been shown that the presence of breast cancer stem cells is closely associated with the migration and metastatic ability of cancer cells, as well as with their resistance to chemotherapy and radiotherapy. The tumor microenvironment is one of the main molecular factors involved in cancer and metastatic processes development, in this sense it is interesting to study the role of platelets, one of the main communicator cells in the human body which are activated by the signals they receive from the microenvironment and can generate more than one response. Platelets can ingest and release RNA, proteins, cytokines and growth factors. After the platelets interact with the tumor microenvironment, they are called "tumor-educated platelets." Tumor-educated platelets transport material from the tumor microenvironment to sites adjacent to the tumor, thus helping to create microenvironments conducive for the development of primary and metastatic tumors. It has been observed that the clone capable of carrying out the metastatic process is a cancer cell with stem cell characteristics. Cancer stem cells go through a series of processes, including epithelial-mesenchymal transition, intravasation into blood vessels, movement through blood vessels, extravasation at the site of the establishment of a metastatic focus, and site colonization. Tumor-educated platelets support all these processes.