METRN modulates immune escape via CD8+ T cells in breast cancer progression through IL-6 signaling pathway

The aberrant expression of stromal gene signatures in breast cancer has been widely studied. However, the association of stromal gene signatures with tumor immunity, progression, and clinical outcomes remains lacking. Based on eight breast tumor stroma (BTS) transcriptomics datasets, we identified differentially expressed genes (DEGs) between BTS and normal breast stroma. Based on the DEGs, we identified dysregulated pathways and prognostic hub genes, hub oncogenes, hub protein kinases, and other key marker genes associated with breast cancer. Moreover, we compared the enrichment levels of stromal and immune signatures between breast cancer patients with bad and good clinical outcomes. We also investigated the association between tumor stroma-related genes and breast cancer progression. The DEGs included 782 upregulated and 276 downregulated genes in BTS versus normal breast stroma. The pathways significantly associated with the DEGs included cytokine-cytokine receptor interaction, chemokine signaling, T cell receptor signaling, cell adhesion molecules, focal adhesion, and extracellular matrix-receptor interaction. Protein-protein interaction network analysis identified the stromal hub genes with prognostic value in breast cancer, including two oncogenes (COL1A1 and IL21R), two protein kinases encoding genes (PRKACA and CSK), and a growth factor encoding gene (PLAU). Moreover, we observed that the patients with bad clinical outcomes were less enriched in stromal and antitumor immune signatures (CD8 + T cells and tumor-infiltrating lymphocytes) but more enriched in tumor cells and immunosuppressive signatures (MDSCs and CD4 + regulatory T cells) compared with the patients with good clinical outcomes. The ratios of CD8 + /CD4 + regulatory T cells were lower in the patients with bad clinical outcomes. Furthermore, we identified the tumor stroma-related genes, including MCM4, SPECC1, IMPA2, and AGO2, which were gradually upregulated through grade I, II, and III breast cancers. In contrast, COL14A1, ESR1, SLIT2, IGF1, CH25H, PRR5L, ABCA6, CEP126, IGDCC4, LHFP, MFAP3, PCSK5, RAB37, RBMS3, SETBP1, and TSPAN11 were gradually downregulated through grade I, II, and III breast cancers. It suggests that the expression of these stromal genes has an association with the progression of breast cancers. These progression-associated genes also displayed an expression association with recurrence-free survival in breast cancer patients. This study identified tumor stroma-associated biomarkers correlated with deregulated pathways, tumor immunity, tumor progression, and clinical outcomes in breast cancer. Our findings provide new insights into the pathogenesis of breast cancer.

Improvement of understanding of the safety profile and biological significance of antidiabetic agents in breast cancer (BC) progression may shed new light on minimizing the unexpected side effect of antidiabetic reagents in diabetic patients with BC. Our recent finding showed that Saxagliptin (Sax) and Sitagliptin (Sit), two common antidiabetic dipeptidyl peptidase-4 inhibitors (DPP-4i) compounds, promoted murine BC 4T1 metastasis via a ROS–NRF2–HO-1 axis in nonobese diabetic–severe combined immunodeficiency (NOD-SCID) mice. However, the potential role of DPP-4i in BC progression under immune-competent status remains largely unknown. Herein, we extended our investigation and revealed that Sax and Sit also accelerated murine BC 4T1 metastasis in orthotopic, syngeneic, and immune-competent BALB/c mice. Mechanically, we found that DPP-4i not only activated ROS–NRF2–HO-1 axis but also triggered reactive oxygen species (ROS)-dependent nuclear factor kappa B (NF-κB) activation and its downstream metastasis-associated gene levels in vitro and in vivo, while NF-кB inhibition significantly abrogated DPP-4i-driven BC metastasis in vitro. Meanwhile, inhibition of NRF2–HO-1 activation attenuated DPP-4i-driven NF-кB activation, while NRF2 activator ALA enhanced NF-кB activation, indicating an essential role of ROS–NRF2–HO-1 axis in DPP-4i-driven NF-кB activation. Furthermore, we also found that DPP-4i increased tumor-infiltrating CD45, MPO, F4/80, CD4, and Foxp3-positive cells and myeloid-derived suppressor cells (MDSCs), and decreased CD8-positive lymphocytes in metastatic sites, but did not significantly alter cell viability, apoptosis, differentiation, and suppressive activation of 4T1-induced splenic MDSCs. Moreover, we revealed that DPP-4i triggered ROS-NF-κB-dependent NLRP3 inflammasome activation in BC cells, leading to increase in inflammation cytokines such as interleukin (IL)-6, tumor necrosis factor alpha (TNF-α), vascular endothelial growth factor (VEGF), intercellular cell adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), IL-1β and IL-33, and MDSCs inductors granulocyte-macrophage colony-stimulating factor (GM-CSF), G-CSF, and M-CSF, which play a crucial role in the remodeling of tumor immune-suppressive microenvironment. Thus, our findings suggest that antidiabetic DPP-4i reprograms tumor microenvironment that facilitates murine BC metastasis by interaction with BC cells via a ROS–NRF2–HO-1–NF-κB–NLRP3 axis. This finding not only provides a mechanistic insight into the oncogenic ROS–NRF2–HO-1 in DPP-4i-driven BC progression but also offers novel insights relevant for the improvement of tumor microenvironment to alleviate DPP-4i-induced BC metastasis.

PP2A is a major tumor suppressor whose inactivation is frequently found in a wide spectrum of human tumors. In particular, deletion or epigenetic silencing of genes encoding the B55 family of PP2A regulatory subunits is a common feature of breast cancer cells. A key player in the regulation of PP2A/B55 phosphatase complexes is the cell cycle kinase MASTL (also known as Greatwall). During cell division, inhibition of PP2A-B55 by MASTL is required to maintain the mitotic state, whereas inactivation of MASTL and PP2A reactivation is required for mitotic exit. Despite its critical role in cell cycle progression in multiple organisms, its relevance as a therapeutic target in human cancer and its dependence of PP2A activity is mostly unknown. Here we show that MASTL overexpression predicts poor survival and shows prognostic value in breast cancer patients. MASTL knockdown or knockout using RNA interference or CRISPR/Cas9 systems impairs proliferation of a subset of breast cancer cells. The proliferative function of MASTL in these tumor cells requires its kinase activity and the presence of PP2A-B55 complexes. By using a new inducible CRISPR/Cas9 system in breast cancer cells, we show that genetic ablation of MASTL displays a significant therapeutic effect in vivo. All together, these data suggest that the PP2A inhibitory kinase MASTL may have both prognostic and therapeutic value in human breast cancer.

Breast cancer (BC) is a diverse disease with heterogeneous subgroups in pathophysiology and behavior. Recent advances in biomarker-based therapy may help improve treatment options for BC. Thus, this study aimed to assess the role of interleukin (IL)-31 and IL-33 as biomarkers associated with BC risk. Serum concentrations of IL-31 and IL-33 were quantified in women with BC (n = 77), women with benign breast lesion (BBL; n = 51), and healthy women (n = 75), using enzyme-linked immunosorbent assay kits. Compared with BBL or healthy women, IL-33 concentrations were significantly greater in BC patients and were associated with a 1.33-fold increased risk of disease and had excellent prognostic value in BC (area under the curve = 0.89). In addition, IL-33 expression was linked to triple-negative BC, distant metastasis, Tumor-Node-Metastasis (TNM) stage, status of estrogen and progesterone receptors, and human epidermal growth factor receptor 2. IL-31 concentrations showed no significant differences between BC, BBL, and healthy women, but were greater in patients with distant metastasis and those who were negative for estrogen and progesterone receptors. In conclusion, IL-33 and to a lesser extent IL-31 can be used as biomarkers associated with BC risk and to identify patients with a poor prognosis especially those with triple-negative BC. Furthermore, understanding the connection between IL-33 and BC progression may aid in the development of safe and effective therapy.

RecQ-mediated genome instability 2 (RMI2) maintains genome stability by promoting DNA damage repair. It has been reported to accelerate the progression of several tumors. However, the functional mechanism of RMI2 in breast cancer remains unclear. Gene expression profiles were obtained from TCGA, GTEx, and GEO databases. The expression of RMI2 and its prognostic value in breast cancer was explored. In addition, we calculated pooled standardized mean deviation (SMD) and performed a summary receiver operating characteristic (sROC) curve analysis to further determine RMI2 expression status and diagnostic significance. The functions and related signaling pathways were investigated based on GO and KEGG analyses. The PPI network was constructed by combining the STRING database and Cytoscape software. Subsequently, in vitro assays were conducted to detect the effect of RMI2 on the proliferation and migration of breast cancer cells. The expression of RMI2 was markedly upregulated in breast cancer tissues relative to that in normal tissues. Moreover, pooled SMD further confirmed the overexpression of RMI2 in breast cancer (SMD=1.29, 95% confidence interval (CI): 1.18-1.41, p=0.000). The sROC curve analysis result suggested that RMI2 had a relatively high diagnostic ability in breast cancer (AUC=0.87, 95% CI: 0.84-0.90). High RMI2 expression was associated with poor prognosis. GO and KEGG analyses revealed that RMI2 was closely related to cell adhesion, various enzyme activities, and PI3K/AKT signaling pathway. PPI analysis showed that RMI2 had interactions with proteins involved in DNA damage repair. knockdown of RMI2 remarkably inhibited the proliferation and migration of breast cancer cells, while overexpression of RMI2 exerted the opposite effects. Furthermore, we identified that RMI2 accelerates the proliferation and migration of breast cancer cells via activation of the PI3K/AKT pathway. The results suggest that RMI2 is a potential diagnostic and prognostic biomarker associated with cell proliferation and migration, and may be used as a novel therapeutic target for breast cancer in the future.

Background Circulating microRNAs (miRNAs) as regulators of gene expression have recntly been promising suitable potential biomarkers due to their stability and ease of detection in blood. In this study, we aimed to determine the plasma expression levels of 372 different miRNAs in patients with invasive ductal breast cancer (IDC). Methods The expression levels of 372 circulating miRNAs in plasma samples of 20 patients with operable stage 1-III IDC and 10 healthy controls were determined using RT-PCR arrays. Mean ages of patients and healthy controls were 45.9+/-8.8) and 45.4+/- 5) respectively. Of 20 breast tumors, 12 were luminal breast cancer, whereas 8 were non-luminal as pure HER2-neu or triple negative breast cancer. RNA was isolated using miRNeasy Serum/Plasma Kit (Qiagen, Hilden, Germany, Cat. No: 217004). cDNA synthesis was performed according to manufacturer's instructions with miScript II RT Kit (Qiagen,Hilden, Germany, Cat. No: 218161). Serum/Plasma 384 HC PCR arrays with miScript SYBR Green PCR Kit (Qiagen, Hilden, Germany, Cat. No:218076) were used RT-PCR analysis. These assays included 372 miRNAs in additon to housekeeping genes and reaction controls. All reactions were performed in triplicates. Ct values were analyzed via an online software developed by Sabiosciences. P values were calculated using Student's t-test and p values lower than 0.05 were considered significant. Results Among 372 miRNAs, 19 were found to be deregulated in plasma samples of patients with IDC. 8 miRNAs were upregulated, and the other 11 were downregulated. Upregulated and Downregulated Circulating miRNAs in Patients with Invasive Ductal CarcinomaUpregulated miRNAsFold regulationp valueDownregulated miRNAsFold Regulationp valuemiR-29a-3p2.0240.032miR-19b-1-5p-2.3750.048miR-101-3p2.4000.040miR-4732-5p-2.0910.015miR-542-3p2.2910.042miR-4687-5p-4.6230.005miR-199b-3p2.0190.020miR-3135b-2.7920.0005miR-98-5p2.4830.003miR-4770-2.4150.0103miR-424-5p3.0550.034miR-4301-2.6680.007miR-374c-5p2.1100.049miR-1247-5p-2.8130.0003miR-19b-3p3.7590.048miR-1287-5p-2.2020.0007 miR-197-3p-4.090.023 miR-126-5p-2.0520.029 miR-671-3p-2.5350.014Table 1 miR-19b-3p was the most upregulated miRNA with a fold change of 3.759 (95%CI:1.83- 5.68, p=.048) while miR-4687-5p was the most downregulated with a fold change of 0.216 (95%CI:0.00001-0.43, p=.005). Conclusion Our findings indicate that these 19 deregulated circulating miRNAs might be promising biomarker candidates for detection of IDC. The two most deregulated miRNAs were miR-19b-3p and miR-4687-5p. miR19b-3p belongs to a cluster of miRNAs which has been shown to function as oncogenes resulting in the downregulation of tissue factor expression in breast cancer cells. Further validation studies are ongoing in order to determine their clinicopathological prognostic value in breast cancer. Citation Format: Tiryakioglu NO, Cabioglu N, Coskunpinar E, Tukenmez M, Ozturk D, Ozkurt E, Igci A, Pence S, Muslumanoglu M. miR-19b-3p and miR-4687-5p as novel circulating miRNAs as potential prognostic biomarkers in breast cancer [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P6-07-24.

ObjectiveWe used bioinformatics analysis to identify potential biomarker genes and their relationship with breast cancer (BC).Materials and MethodsWe used a weighted gene co-expression network analysis (WGCNA) to create a co-expression network based on the top 25% genes in the GSE24124, GSE33926, and GSE86166 datasets obtained from the Gene Expression Omnibus. We used the DAVID online platform to perform GO and KEGG pathway enrichment analyses and the Cytoscape CytoHubba plug-in to screen the potential genes. Then, we related the genes to prognostic values in BC using the Oncomine, GEPIA, and Kaplan–Meier Plotter databases. Findings were validated by immunohistochemical (IHC) staining in the Human Protein Atlas and the TCGA-BRCA cohort. LinkedOmics identified the interactive expressions of hub genes. We used UALCAN to evaluate the methylation levels of these hub genes. MethSurv and SurvivalMeth were used to assess the multilevel prognostic value. Finally, we assessed hub gene association with immune cell infiltration using TIMER.ResultsThe mRNA levels of MKI67, UBE2C, GTSE1, CCNA2, and MND1 were significantly upregulated in BC, whereas ESR1, THSD4, TFF1, AGR2, and FOXA1 were significantly downregulated. The DNA methylation signature analysis showed a better prognosis in the low-risk group. Further subgroup analyses revealed that MND1 might serve as an independent risk factor for unfavorable BC prognosis. Additionally, MND1 expression levels positively correlate with the immune infiltration statuses of CD4+ T cells, CD8+ T cells, B cells, neutrophils, dendritic cells, and macrophages.ConclusionOur results indicate that the ten hub genes may be involved in BC’s carcinogenesis, development, or metastasis, and MND1 may be a potential biomarker and therapeutic target for BC.

Objectives of the Study: Platelet-derived growth factor (PDGF) mediated signaling is pro-tumorigenic in many types of cancer, including breast cancer. However, the requirement of individual PDGF ligands in mediating breast cancer progression remains unclear. Our evaluation of publicly available breast cancer patient datasets shows that high primary tumor expression of PDGFB correlates with reduced overall survival (OS) and reduced metastasis free survival (MFS). This is in contrast to the lack of prognostic power of PDGFA and PDGFC and the association with improved OS and MFS for PDGFD. Based on our analysis of patient datasets, we set out to test the requirement of PDGFB in mammary tumor growth. Methods Used: Gain-of-function and loss-of-function experiments were performed wherein PDGFB was either overexpressed or knocked-down in mammary tumor cells. Manipulated tumor cells were injected directly into the mammary fat pads of adult female mice, and tumor growth was monitored over time. PDGFB is produced by tumor cells whereas the corresponding receptor, PDGFRβ, is expressed by mesenchymal cells in the stroma. As another way to mimic oncogenic PDGFB-to-PDGFRβ signaling in the breast, we developed a mouse model of stroma-specific PDGFRβ activation using the Fsp-cre transgene. We evaluated changes mammary gland development upon mesenchymal specific PDGFRβ activation, and performed orthotopic injections with mammary tumor cells in these animals to test the functional role of receptor activation in mammary tumor growth. Results and Conclusions: We evaluated expression of PDGF ligands in FVB/N murine mammary tumor cell lines and found that the PDGFB is dramatically higher in DB7 tumor cells compared to other syngeneic cell lines. In the high-PDGFB expressing DB7 cells, we used shRNA technology to knockdown the ligand. At the same time, we overexpressed the ligand in an isogenic cell line that expresses low levels of PDGFB. These cells were injected orthotopically into the mammary fat pads of adult female mice, and in both cases, expression of PDGFB dictated tumor growth. There was a significant reduction in tumor growth with shRNA-mediated knockdown of PDGFB whereas overexpression of the ligand accelerated tumor growth. In our mouse model of mesenchymal-specific PDGFRβ activation, we reveal that activation of the receptor exerts pro-proliferative signals on adjacent mammary epithelial cells and accelerates orthotopic tumor growth of PDGFB-expressing cells. These findings indicate that PDGFB-to-PDGFRβ signaling is a viable therapeutic target for breast cancer. In fact, treatment with the PDGFR inhibitor, imatinib, impedes tumor cell proliferation when mouse mammary fibroblasts are co-injected orthotopically with DB7 cells. Significance: The requirement of other PDGF ligands in breast cancer remains to be evaluated, but our data support a pro-tumorigenic role for PDGFB in the breast. Importantly, PDGFR inhibitors are being used in clinical trials for several cancer types. Our data advocate for (1) the potential utility of PDGFB as a prognostic biomarker and (2) the pre-clinical evaluation of PDGFR inhibitors in breast cancer models. Citation Format: Katie A Thies, Anisha M Hammer, Sarah A Steck, Manjusri Das, Raleigh D Kladney, Steven T Sizemore, Michael C Ostrowski, Gina M Sizemore. Platelet derived growth factor-b (PDGFB) promotes breast cancer progression [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P6-06-06.

BackgroundMetastasis is the leading cause of mortality in patients with breast cancer (BC). Studies demonstrate that circular RNAs (circRNAs) were involved in BC progression, while the molecular mechanisms remain largely unclear.MethodsThe microArray circRNA profiles were used to explore the differential expression circRNAs in BC and paracancerous normal tissues, and the quantitative reverse transcription-polymerase chain reaction was used to validate their expression level in clinical samples and cell lines. Nuclear/cytosolic fractionation and fluorescence in situ hybridization (FISH) assays were performed to examine circRRM2 (hsa_circ_0052582) subcellular location. The scratch wound healing and transwell assays were conducted to evaluate the impact of circRRM2 on BC cell migration and invasion. We predicted miRNAs that might bind with cricRRM2 and the downstream target genes using bioinformatics analysis and explored their expression levels and prognostic value in BC. FISH, RNA immunoprecipitation, Co-immunoprecipitation, Western blot, and rescue experiments were implemented to figure out circRRM2 function and underlying mechanisms in BC.ResultsThe present study revealed several aberrant circRNAs in BC tissues and observed that circRRM2 was upregulated in tumor tissues of 40 patients with BC. High circRRM2 was significantly associated with advanced N stage in patients with BC. Gain- and loss- of function experiments revealed that circRRM2 promoted the migration and invasion of cells and functioned as an oncogene in BC. Mechanism studies showed that circRRM2 competed with miR-31-5p/miR-27b-3p to upregulate the IGF2BP1 expression. Furthermore, IGF2BP1 upregulated the circRRM2 level via interacting with MYC, which functioned as the transcriptional factor of circRRM2. Thus, the positive feedback loop that was composed of circRRM2/IGF2BP1/MYC was identified.ConclusionThis study confirms that upregulated circRRM2 functions an oncogenic role in BC metastasis. The positive feedback loop of circRRM2/IGF2BP1/MYC enforces the circRRM2 expression, which might offer a potential target for BC treatment.

Introduction: Minichromosome maintenance complex component 7 (MCM7) plays an essential role in proliferation and DNA replication of cancer cells. However, the expression and prognostic significance of MCM7 in breast cancer (BC) remain to be defined. In this study, we aimed to evaluate the role of MCM7 in BC. Methods: We conducted immunohistochemistry staining of MCM7 in 1,156 operable early-stage BC samples and assessed MCM7 at the transcriptomic levels using publicly available cohorts (n = 13,430). MCM7 expression was evaluated and correlated with clinicopathological parameters including Ki67 labelling index and patient outcome. Results: At the transcriptomic level, there was a significant association between high MCM7 mRNA levels and shorter patient survival in the whole cohort and in luminal BC class but not in the basal-like molecular subtype. High MCM7 protein expression was detected in 43% of patients and was significantly associated with parameters characteristic of aggressive tumour behaviour. MCM7 was independently associated with shorter survival, particularly in oestrogen receptor-positive (luminal) BC. MCM7 stratified luminal tumours with aggressive clinicopathological features into distinct prognostic groups. In endocrine therapy-treated BC patients, high MCM7 was associated with poor outcome, but such association disappeared with administration of adjuvant chemotherapy. Patients with high expression of Ki67 and MCM7 showed worst survival, while patients with double low expression BC showed the best outcome compared with single expression groups. Conclusion: The current findings indicate that MCM7 expression has a prognostic value in BC and can be used to identify luminal BC patients who can benefit from adjuvant chemotherapy. Introduction: Minichromosome maintenance complex component 7 (MCM7) plays an essential role in proliferation and DNA replication of cancer cells. However, the expression and prognostic significance of MCM7 in breast cancer (BC) remain to be defined. In this study, we aimed to evaluate the role of MCM7 in BC. Methods: We conducted immunohistochemistry staining of MCM7 in 1,156 operable early-stage BC samples and assessed MCM7 at the transcriptomic levels using publicly available cohorts (n = 13,430). MCM7 expression was evaluated and correlated with clinicopathological parameters including Ki67 labelling index and patient outcome. Results: At the transcriptomic level, there was a significant association between high MCM7 mRNA levels and shorter patient survival in the whole cohort and in luminal BC class but not in the basal-like molecular subtype. High MCM7 protein expression was detected in 43% of patients and was significantly associated with parameters characteristic of aggressive tumour behaviour. MCM7 was independently associated with shorter survival, particularly in oestrogen receptor-positive (luminal) BC. MCM7 stratified luminal tumours with aggressive clinicopathological features into distinct prognostic groups. In endocrine therapy-treated BC patients, high MCM7 was associated with poor outcome, but such association disappeared with administration of adjuvant chemotherapy. Patients with high expression of Ki67 and MCM7 showed worst survival, while patients with double low expression BC showed the best outcome compared with single expression groups. Conclusion: The current findings indicate that MCM7 expression has a prognostic value in BC and can be used to identify luminal BC patients who can benefit from adjuvant chemotherapy.

Liquid-liquid phase separation (LLPS) impact immune signaling in cancer and related genes have shown prognostic value in breast cancer (BRCA). However, the crosstalk between LLPS and immune infiltration in BRCA remain unclear. Therefore, we aimed to develop a novel prognostic model of BRCA related to LLPS and immune infiltration. BRCA-related, liquid-liquid phase separation (LLPS)-related genes, and differentially expressed genes (DEGs) were identified using public databases. Mutation and drug sensitivity analyses were performed using Gene Set Cancer Analysis database. Univariate cox regression and LASSO Cox regression were used for the construction and verification of prognostic model. Kaplan-Meier analysis was performed to evaluate overall survival (OS). Gene set variation analysis was conducted to analyze key pathways. CIBERSORT was used to assess immune infiltration and its correlation with prognostic genes was determined through Pearson analysis. A total of 6056 BRCA-associated genes, 3775 LLPS-associated genes, and 4049 DEGs, resulting in 314 overlapping genes. Twenty-eight prognostic genes were screened, and some of them were mutational and related to drug sensitivity Subsequently, a prognostic model comprising L1CAM, EVL, FABP7, and CST1 was built. Patients in high-risk group had shorter OS than those in low-risk group. The infiltrating levels of CD8+ T cells, macrophages M0, macrophages M2, dendritic cells activated, and mast cells resting was altered in high-risk group of breast cancer patients compared to low-risk group. L1CAM, EVL, FABP7, and CST1 were related to these infiltrating immune cells. L1CAM, EVL, FABP7, and CST1 were potential diagnostic biomarkers and therapeutic targets for BRCA.

Breast cancer is the most common malignancies worldwide. LncRNA HOX transcript antisense intergenic RNA (HOTAIR) has been shown to promote progression and metastasis of various cancers, including breast cancer. This reasearch aimed to investigate the downstream regulatory pathways of HOTAIR in breast cancer. The levels of HOTAIR and miR-129-5p were examined in breast cancer tissues and SKBR3 and MCF7 cells by quantitative real-time PCR (qRT-PCR). Cell proliferation was examined by Cell Counting Kit-8 (CCK-8) assay. Cell migration and invasion were estimated by transwell assay. Epithelial-to-mesenchymal transition (EMT)-related markers (E-cadherin, N-cadherin and Vimentin) were measured by Western blot assay. The expression of Frizzled 7 (FZD7) was detected using qRT-PCR or Western blot assay. Bioinformatics analysis, luciferase reporter assay or RNA Immunoprecipitation (RIP) assay was performed to explore the molecular mechanism of HOTAIR in breast cancer. Xenograft analysis was utilized to evaluate the tumor growth in vivo. HOTAIR and FZD7 were upregulated, while miR-129-5p was down-regulated in breast cancer tissues and cells. Knockdown of miR-129-5p reversed the effect of HOTAIR knockdown on cell proliferation, migration, invasion and EMT. FZD7 restored the inhibition of miR-129-5p on breast cancer progression. Furthermore, HOTAIR was a sponge of miR-129-5p and FZD7 was a target of miR-129-5p. Knockdown of HOTAIR inhibited the tumor growth in vivo. HOTAIR facilitated breast cancer progression by regulating the miR-129-5p/FZD7 axis, indicating that HOTAIR may be a potential biomarker and therapeutic target for breast cancer.

BackgroundLong noncoding RNAs (lncRNAs) play crucial roles in tumor progression and are aberrantly expressed in various cancers. However, the functional roles of lncRNAs in breast cancer remain largely unknown.MethodsBased on public databases and integrating bioinformatics analyses, the overexpression of lncRNA BCRT1 in breast cancer tissues was detected and further validated in a cohort of breast cancer tissues. The effects of lncRNA BCRT1 on proliferation, migration, invasion and macrophage polarization were determined by in vitro and in vivo experiments. Luciferase reporter assay and RNA immunoprecipitation (RIP) were carried out to reveal the interaction between lncRNA BCRT1, miR-1303, and PTBP3. Chromatin immunoprecipitation (ChIP) and RT-PCR were used to evaluate the regulatory effect of hypoxia-inducible factor-1α (HIF-1α) on lncRNA BCRT1.ResultsLncRNA BCRT1 was significantly upregulated in breast cancer tissues, which was correlated with poor prognosis in breast cancer patients. LncRNA BCRT1 knockdown remarkably suppressed tumor growth and metastasis in vitro and in vivo. Mechanistically, lncRNA BCRT1 could competitively bind with miR-1303 to prevent the degradation of its target gene PTBP3, which acts as a tumor-promoter in breast cancer. LncRNA BCRT1 overexpression could promote M2 polarization of macrophages, mediated by exosomes, which further accelerated breast cancer progression. Furthermore, lncRNA BCRT1 was upregulated in response to hypoxia, which was attributed to the binding of HIF-1α to HREs in the lncRNA BCRT1 promoter.ConclusionsCollectively, these results reveal a novel HIF-1α/lncRNA BCRT1/miR-1303/PTBP3 pathway for breast cancer progression and suggest that lncRNA BCRT1 might be a potential biomarker and therapeutic target for breast cancer.

To investigate the value of Anoctamin 6 (ANO6) in breast cancer (BC) by analyzing its expression, prognostic impact, biological function, and its association with immune characteristics. We initially performed the expression and survival analyses, followed by adopting restricted cubic spline to analyze the nonlinear relationship between ANO6 and overall survival (OS). Stratified and interaction analyses were conducted to further evaluate its prognostic value in BC. Next, we performed enrichment analyses to explore the possible pathways regulated by ANO6. Finally, the correlations between ANO6 and immune characteristics were analyzed to reveal its role in immunotherapy. Lower ANO6 expression was observed in BC than that in the normal breast group, but its overexpression independently predicted poor OS among BC patients (P < .05). Restricted cubic spline analysis revealed a linear relationship between ANO6 and OS (P-Nonlinear > 0.05). Interestingly, menopause status was an interactive factor in the correlation between ANO6 and OS (P for interaction = 0.016). Additionally, ANO6 was involved in stroma-associated pathways, and its elevation was significantly linked to high stroma scores and macrophage polarization (P < .05). Moreover, ANO6 was notably correlated with immune checkpoint expression levels, and scores of tumor mutation burden and microsatellite instability (all P < .05). ANO6 was an independent prognostic factor for BC, and might be a potential target for the BC treatment. Besides, ANO6 might affect BC progression via the regulation of stroma-related pathways and macrophage polarization.

Editorial on the Research Topic Breast cancer (BC) is the most common type of cancer and the fourth cause of cancer death among women worldwide. Steroid (estrogens, progestogens, androgens and glucocorticoid), thyroid, TSH, prolactin and growth/insulin receptors signaling pathways activated by their hormones have a major role in the mammary gland development and in the etiology, and progression of BC. Although classification of BC is defined by hormone receptor expression, (estrogen receptor-ER and/or progesterone receptor-PR), there is evidence indicating that estradiol and progesterone are not the unique hormones driving BC progression. During the last years, the role of different hormones has been unravelled in biological aspects to regulate tumor progression, metastasis, and treatment response, as well as their receptors crosstalks. The study of hormonal receptors expression as well as the molecular mechanism induced could have a prognostic value in BC. The present Research Topic aims to provide a platform for novel research in BC, including original research articles that would help in the development of better diagnostic, prognostic and/or therapeutic strategies for this disease. ER remains the most important biomarker in breast oncology. ER low tumors represent a relatively small subgroup of BC patients, being similar to ER neg disease in their molecular landscape, clinicopathological characteristics, and response to therapy. Nevertheless, a proportion may retain some ER signalling dependency, and so the possibility of responding to some degree to endocrine therapy and should be considered as a heterogeneous number of tumors to be fully characterized from the molecular point of view. Reinert et al. reviewed the most important considerations regarding the definition of ER positivity, pathology assessment, prognosis, and therapeutic implication of ER low BC from the medical oncology perspective. Whether to offer endocrine therapy as part of the overall strategy under the possibility of some remaining endocrine sensitivity should remain an individual discussion. Wright et al. analysed the signalling network of PR in a BC model cell line, T47D by phospho-proteomics. Similar to ER, PR signals mostly through the genomic pathway, but membrane bound forms have also been described. The T47D cell line is a model often applied for PR research; however, the phospho-proteome determination combined with pathway enrichment analysis provides new insights into the molecular function of this hormone. Especially, these data suggest progesterone mediated changes in the nuclear Frontiers in Endocrinology frontiersin.org 01