Breast cancer stem cells as novel biomarkers.

Breast cancer is the most common cancer and the leading cause of mortality worldwide. Despite advancements in detection and treatment, it remains a major cause of cancer-related deaths in women. Breast cancer stem cells (BCSCs) are a crucial group of cells responsible for carcinogenesis, metastasis, medication resistance, and tumor recurrence. Identifying and understanding their molecular pathways is essential for developing effective breast cancer therapy. BCSCs are responsible for tumor genesis, development, metastasis, treatment resistance, and recurrence. Biomarkers are essential tools for identifying high-risk patients, improving diagnostic accuracy, developing follow-up programs, assessing treatment susceptibility, and predicting prognostic outcomes. Stem cell intervention therapy can provide specialized tools for precision therapy. Biomarker analysis in cancer patients is crucial to identify cells associated with disease progression and post-therapeutic relapse. However, negative post-therapeutic impacts can enhance cancer stemness by boosting BCSCs plasticity phenotypes, activating stemness pathways in non-BCSCs, and promoting senescence escape, leading to tumor relapse and metastasis. Despite the advancements in precision medicine, challenges persist in identifying stem cell markers, limiting the number of eligible patients for treatment. The diversity of biomedical research hinders the development of individualization-based preventative, monitoring, and treatment strategies, especially in oncology. Integrating and interpreting clinical and scientific data remains challenging. The development of stem cell-related indicators could significantly improve disease precision, enabling stem cell-targeted therapy and personalized treatment plans, although BCSCs are promising for breast cancer treatment optimization, serving as biomarkers for current therapy modalities. This summary discusses recent advancements in breast cancer stem cell research, including biomarkers, identification methods, molecular mechanisms, and tools for studying their biological origin and lineage development for precision medicine.

Ginsenoside Rg3 decreases breast cancer stem-like phenotypes through impairing MYC mRNA stability.

Breast cancer stem cells (BCSCs) are responsible for breast cancer metastasis, recurrence and treatment resistance, all of which make BCSCs potential drivers of breast cancer aggression. Ginsenoside Rg3, a traditional Chinese herbal medicine, was reported to have multiple antitumor functions. Here, we revealed a novel effect of Rg3 on BCSCs. Rg3 inhibits breast cancer cell viability in a dose- and time-dependent manner. Importantly, Rg3 suppressed mammosphere formation, reduced the expression of stemness-related transcription factors, including c-Myc, Oct4, Sox2 and Lin28, and diminished ALDH(+) populations. Moreover, tumor-bearing mice treated with Rg3 exhibited robust delay of tumor growth and a decrease in tumor-initiating frequency. In addition, we found that Rg3 suppressed breast cancer stem-like properties mainly through inhibiting MYC expression. Mechanistically, Rg3 accelerated the degradation of MYC mRNA by enhancing the expression of the let-7 family, which was demonstrated to bind to the MYC 3' untranslated region (UTR). In conclusion, our findings reveal the remarkable suppressive effect of Rg3 on BCSCs, suggesting that Rg3 is a promising therapeutic treatment for breast cancer.

Targeting triple negative breast cancer stem cells using nanocarriers.

Breast cancer is a complex and heterogeneous disease, encompassing various subtypes characterized by distinct molecular features, clinical behaviors, and treatment responses. Categorization of subtypes is based on the presence or absence of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2), leading to subtypes such as luminal A, luminal B, HER2-positive, and triple-negative breast cancer (TNBC). TNBC, comprising around 20% of all breast cancers, lacks expression of ER, PR, and HER2 receptors, rendering it unresponsive to targeted therapies and presenting significant challenges in treatment. TNBC is associated with aggressive behavior, high rates of recurrence, and resistance to chemotherapy. Tumor initiation, progression, and treatment resistance in TNBC are attributed to breast cancer stem cells (BCSCs), which possess self-renewal, differentiation, and tumorigenic potential. Surface markers, self-renewal pathways (Notch, Wnt, Hedgehog signaling), apoptotic protein (Bcl-2), angiogenesis inhibition (VEGF inhibitors), and immune modulation (cytokines, immune checkpoint inhibitors) are among the key targets discussed in this review. However, targeting the BCSC subpopulation in TNBC presents challenges, including off-target effects, low solubility, and bioavailability of anti-BCSC agents. Nanoparticle-based therapies offer a promising approach to target various molecular pathways and cellular processes implicated in survival of BSCS in TNBC. In this review, we explore various nanocarrier-based approaches for targeting BCSCs in TNBC, aiming to overcome these challenges and improve treatment outcomes for TNBC patients. These nanoparticle-based therapeutic strategies hold promise for addressing the therapeutic gap in TNBC treatment by delivering targeted therapies to BCSCs while minimizing systemic toxicity and enhancing treatment efficacy.

Scutellarin, a flavonoid compound from Scutellaria barbata, suppresses growth of breast cancer stem cells in vitro and in tumor-bearing mice.

BACKGROUND: Scutellaria barbata D. Don (SB), commonly known as Ban Zhi Lian and firstly documented by Shigong Chen, is a dried whole plant that has been studied for its therapeutic effects on breast cancer, colon cancer, and prostate cancer. Among its various compounds, scutellarin (SCU) has been demonstrated with anti-tumor effects. PURPOSE: This study aimed to evaluate the effects of SB water extract (SBW) and scutellarin on breast cancer stem cells (BCSCs), and to investigate their potential therapeutic effects on breast tumors in mice. METHODS: BCSCs were enriched from human breast cancer cells (MDA-MB-231 and MDA-MB-361) and their characteristics were analyzed. The effects of varying concentrations of SBW and scutellarin on cell viability, proliferation, self-renewal, and migration abilities were studied, along with the underlying mechanisms. The in vivo anti-tumor effects of scutellarin were further evaluated in SCID/NOD mice. Firstly, mice were inoculated with naïve BCSCs and subjected to treatment with scutellarin or vehicle. Secondly, BCSCs were pre-treated with scutellarin or vehicle prior to inoculation into mice. RESULTS: The derived BCSCs expressed CD44, CD133 and ALDH1, but not CD24, indicating that BCSCs have been successfully induced from both MDA-MB-231 and MDA-MB-361 cells. Both SBW and scutellarin reduced the viability, proliferation, sphere and colony formation, and migration of BCSCs. In mice with tumors derived from naïve BCSCs, scutellarin significantly reduced tumor growth, expression of proliferative (Ki67) and stem cell markers (CD44), and lung metastasis. In addition, pre-treatment with scutellarin also slowed tumor growth. Western blot results suggested the involvement of Wnt/ß-catenin, NF-κB, and PTEN/Akt/mTOR signaling pathways underlying the inhibitory effects of scutellarin. CONCLUSION: Our study demonstrated for the first time that both SB water extract and scutellarin could reduce the proliferation and migration of BCSCs in vitro. Scutellarin was shown to possess novel inhibitory activities in BCSCs progression. These findings suggest that Scutellaria barbata water extract, in particular, scutellarin, may have potential to be further developed as an adjuvant therapy for reducing breast cancer recurrence.

Transcriptomics analysis reveals distinct mechanism of breast cancer stem cells regulation in mammospheres from MCF-7 and T47D cells.

Luminal A breast cancer, constituting 70 % of breast cancer cases, presents a challenge due to the development of resistance and recurrence caused by breast cancer stem cells (BCSC). Luminal breast tumors are characterized by TP53 expression, a tumor suppressor gene involved in maintaining stem cell attributes in cancer. Although a previous study successfully developed mammospheres (MS) from MCF-7 (with wild-type TP53) and T47D (with mutant TP53) luminal breast cancer cells for BCSC enrichment, their transcriptomic profiles remain unclear. We aimed to elucidate the transcriptomic disparities between MS of MCF-7 and T47D cells using bioinformatics analyses of differentially expressed genes (DEGs), including the KEGG pathway, Gene Ontology (GO), drug-gene association, disease-gene association, Gene Set Enrichment Analysis (GSEA), DNA methylation analysis, correlation analysis of DEGs with immune cell infiltration, and association analysis of genes and small-molecule compounds via the Connectivity Map (CMap). Upregulated DEGs were enriched in metabolism-related KEGG pathways, whereas downregulated DEGs were enriched in the MAPK signaling pathway. Drug-gene association analysis revealed that both upregulated and downregulated DEGs were associated with fostamatinib. The KEGG pathway GSEA results indicated that the DEGs were enriched for oxidative phosphorylation, whereas the downregulated DEGs were negatively enriched for the p53 signaling pathway. Examination of DNA methylation revealed a noticeable disparity in the expression patterns of the PKM2, ERO1L, SLC6A6, EPAS1, APLP2, RPL10L, and NEDD4 genes when comparing cohorts with low- and high-risk breast cancer. Furthermore, a significant positive correlation was identified between SLC6A6 expression and macrophage presence, as well as MSN, and AKR1B1 expression and neutrophil and dentritic cell infiltration. CMap analysis unveiled SA-83851 as a potential candidate to counteract the effects of DEGs, specifically in cells harbouring mutant TP53. Further research, including in vitro and in vivo validations, is warranted to develop drugs targeting BCSCs.

Construction and in vitro evaluation of pH-sensitive nanoparticles to reverse drug resistance of breast cancer stem cells.

Breast cancer is a major threat to safety and health of women. The breast cancer stem cells (BCSCs) have multi-drug resistance to chemotherapy drugs, which leads to chemotherapy failure. We proposed a strategy of delivery of tumor-killing drugs and a resistance reversal agent, to enhance inhibition of BCSCs. Here, schisandrin B (SchB)/AP NPs are constructed using acid-grafted-poly (ß-amino ester) (ATRA-g-PBAE, AP) grafted polymer nanoparticle encapsulated SchB, with pH-sensitive release function. This drug delivery system has good pharmacological properties and can increase the SchB release with the decrease of pH. The NPs showed cytotoxic effects in reversing ATRA resistance to BCSCs. Lysosomal escape was achieved when the nanoparticles were taken up by BCSCs. In addition, we found that NPs may reverse MDR by inhibiting the expression of P-glycoprotein (P-gp) and affecting the energy supply of drug efflux. This study provides a nanodelivery therapy strategy that reverses BCSCs multidrug resistance (MDR) and demonstrates that it did so by interfering with cancer cell energy metabolism. Therefore, the co-delivery strategy of ATRA and SchB provides a new option for the treatment of breast cancer.

Impact of De Novo Cholesterol Biosynthesis on the Initiation and Progression of Breast Cancer.

Cholesterol (CHOL) is a multifaceted lipid molecule. It is an essential structural component of cell membranes, where it cooperates in regulating the intracellular trafficking and signaling pathways. Additionally, it serves as a precursor for vital biomolecules, including steroid hormones, isoprenoids, vitamin D, and bile acids. Although CHOL is normally uptaken from the bloodstream, cells can synthesize it de novo in response to an increased requirement due to physiological tissue remodeling or abnormal proliferation, such as in cancer. Cumulating evidence indicated that increased CHOL biosynthesis is a common feature of breast cancer and is associated with the neoplastic transformation of normal mammary epithelial cells. After an overview of the multiple biological activities of CHOL and its derivatives, this review will address the impact of de novo CHOL production on the promotion of breast cancer with a focus on mammary stem cells. The review will also discuss the effect of de novo CHOL production on in situ and invasive carcinoma and its impact on the response to adjuvant treatment. Finally, the review will discuss the present and future therapeutic strategies to normalize CHOL biosynthesis.

Liposome-enabled bufalin and doxorubicin combination therapy for trastuzumab-resistant breast cancer with a focus on cancer stem cells.

Breast cancer stem cells (BCSCs) play a key role in therapeutic resistance in breast cancer treatments and disease recurrence. This study aimed to develop a combination therapy loaded with pH-sensitive liposomes to kill both BCSCs and the okbulk cancer cells using trastuzumab-sensitive and resistant human epidermal growth factor receptor 2 positive (HER2+) breast cancer cell models. The anti-BCSCs effect and cytotoxicity of all-trans retinoic acid, salinomycin, and bufalin alone or in combination with doxorubicin were compared in HER2+ cell line BT-474 and a validated trastuzumab-resistant cell line, BT-474R. The most potent anti-BCSC agent was selected and loaded into a pH-sensitive liposome system. The effects of the liposomal combination on BCSCs and bulk cancer cells were assessed. Compared with BT-474, the aldehyde dehydrogenase positive BCSC population was elevated in BT-474R (3.9 vs. 23.1%). Bufalin was the most potent agent and suppressed tumorigenesis of BCSCs by ∼50%, and showed strong synergism with doxorubicin in both BT-474 and BT-474R cell lines. The liposomal combination of bufalin and doxorubicin significantly reduced the BCSC population size by 85%, and inhibited both tumorigenesis and self-renewal, although it had little effect on the migration and invasiveness. The cytotoxicity against the bulk cancer cells was also enhanced by the liposomal combination than either formulation alone in both cell lines (p < 0.001). The liposomal bufalin and doxorubicin combination therapy may effectively target both BCSCs and bulk cancer cells for a better outcome in trastuzumab-resistant HER2+ breast cancer.

Targeting the Key Signaling Pathways in Breast Cancer: From Molecular Mechanism to Therapeutic Interventions.

Breast cancer is a public health issue in developing and developed countries. Nowadays, the concept of BCSC (breast cancer stem cell) is gaining popularity among oncology researchers. The breast cancer stem cell is a tiny cell fraction inside the tumor mass that shows features that look like stem cells that are implicated in the genesis, recurrence, and metastasis of breast cancer tumors. Extracellular cues, mutations, and epigenetic control all contribute to the intricacy of gene expression control in Breast cancer stem cells. Thus, signaling pathways identified in breast cancer are Hedgehog and NOTCH, signal transducer and transcription 3, wingless-type MMTV integration site family (Wnt)/-catenin, and nuclear factor-kappa B, particularly connected with a phenotype of stem cell. Furthermore, the tumor microenvironment, such as hypoxic regions, can impact these BCSCs. Various approved signaling pathway targeted molecules have been patented, which show protective effects against breast cancer and have been used in clinical uses. PARP inhibitors are found to be very useful in the treatment of breast cancer. Promoting studies on the molecular pathways underlying the development of cancer in breast cancer patients was one of the main objectives of this study topic. The objective of this review Topic was to discover new intrinsic and extrinsic molecular pathways. Research focusing on novel signaling pathways that may lead to novel treatments or identifying patients at-risk of not responding to standard therapy approaches were the areas of focus we highlighted. The paper covers the linkage between breast cancer stem cells and cellular signaling, the tumor microenvironment in BC, and the relevance of signaling pathways and their therapeutic interventions. The review also covered patent applications associated with these signaling pathways and their prospects.

Cancer stem cell-immune cell crosstalk in breast tumor microenvironment: a determinant of therapeutic facet.

Breast cancer (BC) is globally one of the leading killers among women. Within a breast tumor, a minor population of transformed cells accountable for drug resistance, survival, and metastasis is known as breast cancer stem cells (BCSCs). Several experimental lines of evidence have indicated that BCSCs influence the functionality of immune cells. They evade immune surveillance by altering the characteristics of immune cells and modulate the tumor landscape to an immune-suppressive type. They are proficient in switching from a quiescent phase (slowly cycling) to an actively proliferating phenotype with a high degree of plasticity. This review confers the relevance and impact of crosstalk between immune cells and BCSCs as a fate determinant for BC prognosis. It also focuses on current strategies for targeting these aberrant BCSCs that could open avenues for the treatment of breast carcinoma.

Unraveling the Potential of miRNAs from CSCs as an Emerging Clinical Tool for Breast Cancer Diagnosis and Prognosis.

Breast cancer (BC) is the most diagnosed cancer in women and the second most common cancer globally. Significant advances in BC research have led to improved early detection and effective therapies. One of the key challenges in BC is the presence of BC stem cells (BCSCs). This small subpopulation within the tumor possesses unique characteristics, including tumor-initiating capabilities, contributes to treatment resistance, and plays a role in cancer recurrence and metastasis. In recent years, microRNAs (miRNAs) have emerged as potential regulators of BCSCs, which can modulate gene expression and influence cellular processes like BCSCs' self-renewal, differentiation, and tumor-promoting pathways. Understanding the miRNA signatures of BCSCs holds great promise for improving BC diagnosis and prognosis. By targeting BCSCs and their associated miRNAs, researchers aim to develop more effective and personalized treatment strategies that may offer better outcomes for BC patients, minimizing tumor recurrence and metastasis. In conclusion, the investigation of miRNAs as regulators of BCSCs opens new directions for advancing BC research through the use of bioinformatics and the development of innovative therapeutic approaches. This review summarizes the most recent and innovative studies and clinical trials on the role of BCSCs miRNAs as potential tools for early diagnosis, prognosis, and resistance.

SGCE promotes breast cancer stemness by promoting the transcription of FGF-BP1 by Sp1.

Breast cancer stem cells are mainly responsible for poor prognosis, especially in triple-negative breast cancer (TNBC). In a previous study, we demonstrated that ε-Sarcoglycan (SGCE), a type Ⅰ single-transmembrane protein, is a potential oncogene that promotes TNBC stemness by stabilizing EGFR. Here, we further found that SGCE depletion reduces breast cancer stem cells, partially through inhibiting the transcription of FGF-BP1, a secreted oncoprotein. Mechanistically, we demonstrate that SGCE could interact with the specific protein 1 transcription factor and translocate into the nucleus, which leads to an increase in the transcription of FGF-BP1, and the secreted FBF-BP1 activates FGF-FGFR signaling to promote cancer cell stemness. The novel SGCE-Sp1-FGF-BP1 axis provides novel potential candidate diagnostic markers and therapeutic targets for TNBC.

Mechanisms of Breast Cancer Stem Cell Specification and Self-Renewal Mediated by Hypoxia-Inducible Factor 1.

Many advanced human cancers contain regions of intratumoral hypoxia, with O2 gradients extending to anoxia. Hypoxia-inducible factors (HIFs) are activated in hypoxic cancer cells and drive metabolic reprogramming, vascularization, invasion, and metastasis. Hypoxia induces breast cancer stem cell (BCSC) specification by inducing the expression and/or activity of the pluripotency factors KLF4, NANOG, OCT4, and SOX2. Recent studies have identified HIF-1-dependent expression of PLXNB3, NARF, and TERT in hypoxic breast cancer cells. PLXNB3 binds to and activates the MET receptor tyrosine kinase, leading to activation of the SRC non-receptor tyrosine kinase and subsequently focal adhesion kinase, which promotes cancer cell migration and invasion. PLXNB3-MET-SRC signaling also activates STAT3, a transcription factor that mediates increased NANOG gene expression. Hypoxia-induced NARF binds to OCT4 and serves as a coactivator by stabilizing OCT4 binding to the KLF4, NANOG, and SOX2 genes and by stabilizing the interaction of OCT4 with KDM6A, a histone demethylase that erases repressive trimethylation of histone H3 at lysine 27, thereby increasing KLF4, NANOG, and SOX2 gene expression. In addition to increasing pluripotency factor expression by these mechanisms, HIF-1 directly activates expression of the TERT gene encoding telomerase, the enzyme required for maintenance of telomeres, which is required for the unlimited self-renewal of BCSCs. HIF-1 binds to the TERT gene and recruits NANOG, which serves as a coactivator by promoting the subsequent recruitment of USP9X, a deubiquitinase that inhibits HIF-1α degradation, and p300, a histone acetyltransferase that mediates acetylation of H3K27, which is required for transcriptional activation.

Estrogen-Related Receptor Alpha (ERRα) Promotes Cancer Stem Cell-Like Characteristics in Breast Cancer.

Cancer stem cells drive tumor initiation, invasion, metastasis and recurrence. In the present study, we have evaluated the role of ERRα in the maintenance of breast cancer stem cells (BCSCs) using breast cancer cell lines. The inhibition of ERRα with the inverse agonist, XCT-790, or the knockdown of ERRα in breast cancer cells significantly reduced the mammosphere formation efficiency and mammosphere size along with a significant reduction in the CD44+/CD24- BCSCs. Treatment with XCT-790 significantly downregulated expression of the transcription factors involved in stem cell maintenance such as Oct4, Klf4, Sox2, Nanog and c-Myc in the mammosphere forming stem cells of MCF7 and MDA-MB-231. In addition, XCT-790 induced cell cycle arrest and apoptosis in the mammosphere-forming cells. The knockdown or inhibition of ERRα downregulated the expression of Notch1 and ß-catenin, whereas the overexpression of ERRα in MCF7 cells upregulated the expression of these proteins. Moreover, the inhibition of ERRα synergistically enhanced the efficacy of paclitaxel in inhibiting the BCSCs. These results show that ERRα is crucial for the maintenance of BCSCs and suggest that ERRα could be a potential target for breast cancer treatment.

Cancer Cachexia and breast cancer stem cell signalling - A crosstalk of signalling molecules.

Cancer Cachexia is a condition characterized by the involuntary loss of lean body mass, a negative protein and energy balance, and systemic inflammation. This syndrome profoundly impacts the patient's quality of life and is linked to poor chemotherapy response and reduced survival. Despite multiple mechanisms being implicated in its development, and various cytokines believed to contribute to the persistent catabolic state, cachexia is still not fully recognized and is often left untreated. Cachexia is caused by altered metabolic adaptation and lack of anticactic therapy due to systemic cytokines promoting and fuelling cancer growth. The exact molecular mechanisms and clinical endpoints remain poorly defined. It has an occurrence rate of 30%-80%, accounting for 20% of total cancer mortality. Tumor cells remodel the microenvironment suitable for their proliferation, wherein they communicate with fibroblast cells to modulate their expression and induce tumor progressive cytokines. Several studies have reported its strong correlation with systemic cytokines that initiate and aggravate the condition. Plenty of studies show the prominent role of cancer-induced cachexia in pancreatic cancer, colon cancer, and lung cancer. However, limited data are available for breast cancer-induced cachexia, highlighting the need for studying it. Breast cancer stem cells (BCSCs) are a prominently explored area in breast cancer research. They are characterized by CD44+/CD24-/ALDH+ expression and are a focus of cancer research. They are a source of renewal and differentiation within the tumor environment and are responsible for progression, and chemotherapeutic resistance. The tumor microenvironment and its cytokines are responsible for maintaining and inducing their differentiation. Cytokines significantly impact BCSC development and self-renewal, stimulating or inhibiting proliferation depending on cytokine and environment. Pro-inflammatory mediators like IL-6, TNF-α, and IL-8 increase proliferation, promoting tumor growth. Experimental models and clinical studies have shown a direct relationship between cytokines and BCSC proliferation. Several of them seem to be interconnected as they initiate signalling down different pathways but converge at BCSC increase and tumor proliferation. This review highlights the common pathways between cachexia and BCSC signalling, to identify potential therapeutic targets that can aid both conditions.

GNPNAT1 promotes the stemness of breast cancer and serves as a potential prognostic biomarker.

Glucosamine­phosphate N­acetyltransferase 1 (GNPNAT1) is a member of the acetyltransferase superfamily, related to general control non­depressible 5 (GCN5). It has been documented that GNPNAT1 expression is increased in lung cancer, whereas its involvement in breast cancer (BC) remains to be further investigated. The present study aimed to evaluate the expression levels of GNPNAT1 in BC and its effect on BC stem cells (BCSCs). The Cancer Genome Atlas (TCGA) database was used for the analysis of the expression of GNPNAT1 and its clinical significance. Cox regression and logistic regression analyses were used to evaluate prognosis­related factors. The GNPNAT1­binding protein network was constructed using the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) application. The biological signaling pathways implicated in GNPNAT1 were investigated through function enrichment analysis including Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, and gene set enrichment analysis. The single­sample GSEA method was used to investigate the connection between the level of immune infiltration and GNPNAT1 expression in BC. GNPNAT1 expression was upregulated in patients with BC and was significantly associated with a poor prognosis. GNPNAT1 and its co­expressed genes were mostly enriched in nuclear transport, Golgi vesicle transport, ubiquitin­like protein transferase activity and ribonucleoprotein complex binding, as determined using functional enrichment analysis. GNPNAT1 expression was positively associated with Th2 cells and T­helper cells, and negatively associated with plasmacytoid dendritic cells, CD8+ T­cells and cytotoxic cells. Additionally, the GNPNAT1 expression levels were considerably increased in BCSCs. GNPNAT1 knockdown markedly decreased the stemness ability of SKBR3 and Hs578T cells, including the production of CSC markers and mammosphere or clone formation, while GNPNAT1 overexpression increased the stemness level. Hence, the findings of the present study demonstrate that GNPNAT1 may be exploited as a novel prognostic biomarker and therapeutic target for BC.

Quinacrine inhibits HIF-1α/VEGF-A mediated angiogenesis by disrupting the interaction between cMET and ABCG2 in patient-derived breast cancer stem cells.

BACKGROUND: Breast cancer stem cells (BCSCs) have a critical role in progression of breast cancer by inducing angiogenesis. Several therapeutic strategies have been designed for the treatment of breast cancer by specifically preventing angiogenesis. But there is a dearth of study regarding the treatment procedure which can specifically target and kill the BCSCs and cause lesser harm to healthy cells of the body. A plant-based bioactive compound Quinacrine (QC) specifically kills cancer stem cells (CSCs) without harming healthy cells and also inhibits cancer angiogenesis but the detailed mechanistic study of its anti-CSCs and anti-angiogenic activity is yet to explore. HYPOTHESIS: Earlier report showed that both cMET and ABCG2 play an essential role in cancer angiogenesis. Both are present on the cell surface of CSCs and share an identical ATP-binding domain. Interestingly, QC a plant based and bioactive compound which was found to inhibit the function of CSCs marker cMET and ABCG2. These relevant evidence led us to hypothesize that cMET and ABCG2 may interact with each other and induce the production of angiogenic factors, resulting in activation of cancer angiogenesis and QC might disrupt the interaction between them to stop this phenomena. METHODS: Co-immunoprecipitation assay, immunofluorescence assay, and western blotting were performed by using ex vivo patient-derived breast cancer-stem-cells (PDBCSCs) and human umbilical vein endothelial cells (HUVECs). In silico study was carried out to check the interaction between cMET and ABCG2 in presence or absence of QC. Tube formation assay using HUVECs and in ovo Chorioallantoic membrane (CAM) assay using chick fertilized eggs were performed to monitor angiogenesis. In vivo patient-derived xenograft (PDX) mice model was used to validate in silico and ex vivo results. RESULTS: Data revealed that in a hypoxic tumor microenvironment (TME), cMET and ABCG2 interact with each other and upregulate HIF-1α/VEGF-A axis to induce breast cancer angiogenesis. In silico and ex vivo study showed that QC disrupted the interaction between cMET and ABCG2 to inhibit the angiogenic response in endothelial cells by reducing the secretion of VEGF-A from PDBCSCs within the TME. Knockdown of cMET, ABCG2 or both, significantly downregulated the expression of HIF-1α and reduced the secretion of pro-angiogenic factor VEGF-A in the TME of PDBCSCs. Additionally, when PDBCSCs were treated with QC, similar experimental results were obtained. CONCLUSION: In silico, in ovo, ex vivo and in vivo data confirmed that QC inhibited the HIF-1α/VEGF-A mediated angiogenesis in breast cancer by disrupting the interaction between cMET and ABCG2.

ZEB1 potentiates chemoresistance in breast cancer stem cells by evading apoptosis.

Chemoresistance renders a challenge to the clinics to treat breast cancer patients. Current treatment strategies are effective in mitigating tumor growth but remain largely ineffective against cancer-initiating cells or breast Cancer Stem Cells (CSCs). Epithelial-to-mesenchymal-transition (EMT) regulates breast CSC physiology. Zinc finger E-box binding homeobox 1 (ZEB1) is a key EMT-transcription factor that regulates breast CSC - differentiation and metastasis. However, its potential role in modulating tumor chemoresistance has not yet been fully understood. In-silico analysis revealed a higher ZEB1 expression in breast cancer patients that leads to decreased overall and relapse-free survival. We generated sorted breast CSC with stable ZEB1 overexpression (CD24-/CD44+GFP-ZEB1) and/or silencing (CD24-/CD44+ZEB1 shRNA) as well as breast cancer cells with stable ZEB1 overexpression (CD24+GFP-ZEB1) and/or silencing (CD24+ZEB1 shRNA). An increased colony-forming efficiency and doxorubicin accumulation correlated with decreased promoter activity and expression profile of ABCC1 drug-efflux ABC transporter in CD24-/CD44+GFP-ZEB1. Additionally, CD24-/CD44+GFP-ZEB1 demonstrated doxorubicin-induced higher anti-apoptotic and lower pro-apoptotic protein expressions in the mitochondrial and cytosolic fractions. Chemoresistant CD24-/CD44+GFP-ZEB1 cells depicted 1000-fold higher IC-50 values of doxorubicin and decreased activation of JNK-p38 stress kinase molecular signaling-dependent mammosphere forming efficiency to evade apoptosis. Thus, ZEB1 and its downstream effectors are plausible therapeutic targets for the mitigation of breast cancer chemoresistance in patients.

Long non-coding RNAs in breast cancer stem cells.

Breast cancer, one of the most commonly diagnosed cancers worldwide, is a heterogeneous disease with high rates of recurrence and metastasis that contribute to its high mortality rate. Breast cancer stem cells (BCSCs) are a small but significant subset of heterogeneous breast cancer cells that possess stem cell characteristics such as self-renewal and differentiation abilities that may drive metastasis and recurrence. Long non-coding RNAs (lncRNAs) are a class of RNAs that are longer than 200 nucleotides in length and do not possess protein-coding properties. An increasing number of studies have shown that some lncRNAs are abnormally expressed in BCSCs, and have great biological significance in the occurrence, progression, invasion, and metastasis of various cancers. However, the importance of lncRNAs, as well as the molecular mechanisms that regulate and promote the stemness of BCSCs, are still poorly understood. In the current review, we aim to summarize recent studies that highlight the role of lncRNAs in tumor occurrence and progression through BCSCs. In addition, the utility of lncRNAs as biomarkers of breast cancer progression, and their potential use as therapeutic targets for treatment of breast cancer, will be discussed.

Growth and invasion inhibition of T47D ductal carcinoma cells by the association of docetaxel with a bioactive agent in neutral nanosuspension.

Introduction: The approach for drug delivery has impressively developed with the emergence of nanosuspension, particularly the targeted nanoemulsions (NEs). It can potentially improve the bioavailability of drugs, enhancing their therapeutic efficiency. This study aims to examine the potential role of NE as a delivery system for the combination of docetaxel (DTX), a microtubule-targeting agent, and thymoquinone (TQ) in the treatment of human ductal carcinoma cells T47D. Methods: NEs were synthesized by ultra-sonication and characterized physically by dynamic light scattering (DLS). A sulforhodamine B assay was performed to evaluate cytotoxicity, and a flow cytometry analysis for cell cycle, apoptosis, autophagy, and cancer stem cell evaluations. A quantitative polymerase chain reaction further assessed the epithelial-mesenchymal transition gene expirations of SNAIL-1, ZEB-1, and TWIST-1. Results: The optimal sizes of blank-NEs and NE-DTX+TQ were found at 117.3 ± 8 nm and 373 ± 6.8 nm, respectively. The synergistic effect of the NE-DTX+TQ formulation significantly inhibited the in vitro proliferation of T47D cells. It caused a significant increase in apoptosis, accompanied by the stimulation of autophagy. Moreover, this formulation arrested T47D cells at the G2/M phase, promoted the reduction of the breast cancer stem cell (BCSC) population, and repressed the expression of TWIST-1 and ZEB-1. Conclusion: Co-delivery of NE-DTX+TQ may probably inhibit the proliferation of T47D via the induction of apoptosis and autophagy pathways and impede the migration by reducing the BCSC population and downregulating TWIST-1 expression to decrease the epithelial-to-mesenchymal transition (EMT) of breast cancer cells. Therefore, the study suggests the NE-DTX+TQ formula as a potential approach to inhibit breast cancer growth and metastasis.