Inhibited Triple-negative Breast Cancer Cell Research Articles

Integrating network pharmacology, molecular docking and experimental verification to explore the therapeutic effect and potential mechanism of nomilin against triple-negative breast cancer

BackgroundNomilin is a limonoid compound known for its multiple biological activities, but its role in triple negative breast cancer (TNBC) remains unclear. This study aims to uncover the potential therapeutic effect of nomilin on TNBC and elucidate the specific mechanism of its action.MethodsWe employed weighted gene co-expression network analysis (WGCNA), differential expression analysis, and the GeneCards database to identify potential targets for TNBC. Simultaneously, we utilized the Swiss Target Prediction, ChEMBL, and STITCH databases to identify potential targets of nomilin. The core targets and mechanisms of nomilin against TNBC were predicted through protein-protein interaction (PPI) network analysis, molecular docking, and enrichment analysis. The results of the network pharmacology were corroborated by conducting experiments.ResultsA total of 17,204 TNBC targets were screened, and 301 potential targets of nomilin were identified. Through the PPI network, eight core targets of nomilin against TNBC were pinpointed, namely BCL2, Caspase3, CyclinD1, EGFR, HSP90AA1, KRAS, PARP1, and TNF. Molecular docking, molecular dynamics simulation and proteome microarray revealed that nomilin exhibits strong binding activity to these core proteins. Enrichment analysis results indicated that the anti-TNBC effect of nomilin is associated with PI3K/Akt pathway. In vitro and in vivo experiments have demonstrated that nomilin inhibits TNBC cell proliferation and migration while promoting cell apoptosis through the PI3K/Akt pathway.ConclusionFor the first time, the research effectively discovered the objectives and mechanisms of nomilin in combating TNBC using network pharmacology, molecular docking, molecular dynamics simulation, proteome microarray and experimental confirmation, presenting a hopeful approach for treating TNBC.Graphical

Combined knockdown of CD151 and MMP9 may inhibit the malignant biological behaviours of triple-negative breast cancer through the GSK-3β/β-catenin-related pathway

Triple-negative breast cancer (TNBC) represents a significant health concern for women worldwide, and the overproduction of MMP9 and CD151 is associated with various cancers, influencing tumour growth and progression. This study aimed to investigate how CD151 and MMP9 affect TNBC cell migration, apoptosis, proliferation, and invasion. Immunohistochemical experiments revealed that CD151 and MMP9 were positively expressed in triple-negative breast cancer, and lymph node metastasis, the histological grade, and CD151 and MMP9 expression were found to be independent prognostic factors for the survival of patients with triple-negative breast cancer. Cytological experiments indicated that the knockdown of CD151 or MMP9 slowed triple-negative breast cancer cell growth, migration, and invasion and increased the apoptosis rate. Compared with CD151 knockdown, double MMP9 and CD151 knockdown further promoted cell death and inhibited TNBC cell proliferation, migration, and invasion. Moreover, β-catenin and p-GSK-3β were significantly downregulated. In summary, simultaneously silencing CD151 and MMP9 further suppressed the proliferation, migration and invasion of TNBC cells and promoted their apoptosis. One possible strategy for inducing this effect is to block the GSK-3β/β-catenin pathway.

Puerarin Decreases the Expression of FUS-Dependent MAPK4 to Inhibit the Development of Triple-Negative Breast Cancer.

Puerarin has been reported to have anticancer properties; however, its mechanism in regulating triple-negative breast cancer (TNBC) remains unclear. Cell function was assessed using a cell counting kit-8 assay, 5-ethynyl-2'-deoxyuridine assay, flow cytometry, and transwell assay. Additionally, the glucose assay kit, lactate assay kit, and ADP/ATP ratio assay kit were used to analyze glucose metabolism. mRNA and protein expression levels were analyzed using qRT-PCR and western blotting assays, respectively. The relationship between FUS RNA binding protein (FUS) and mitogen-activated protein kinase 4 (MAPK4) was determined using an RNA immunoprecipitation assay. TNBC cell malignancy invitro was validated using a xenograft mouse model assay. Puerarin treatment or MAPK4 knockdown effectively inhibited TNBC cell proliferation, invasion, and glucose metabolism, and induced cell apoptosis. Additionally, puerarin treatment downregulated MAPK4 and FUS expression. Conversely, MAPK4 overexpression attenuated the effects of puerarin in TNBC cells. FUS stabilized MAPK4 mRNA expression in TNBC cells. Furthermore, puerarin decreased MAPK4 expression by downregulating FUS in TNBC cells. Finally, puerarin inhibited tumor formation invivo. Puerarin inhibited TNBC development by decreasing the expression of FUS-dependent MAPK4, indicating that puerarin may serve as a promising therapeutic agent to hind TNBC.

Transcriptional regulation of DLGAP5 by AR suppresses p53 signaling and inhibits CD8+T cell infiltration in triple-negative breast cancer

Triple-negative breast cancer (TNBC) is a challenging subtype with unclear biological mechanisms. Recently, the transcription factor androgen receptor (AR) and its regulation of the DLGAP5 gene have gained attention in TNBC pathogenesis. In this study, we found a positive correlation between high AR expression and TNBC cell proliferation and growth. Furthermore, we confirmed DLGAP5 as a critical downstream regulator of AR with high expression in TNBC tissues. Knockdown of DLGAP5 significantly inhibited TNBC cell proliferation, migration, and invasion. AR was observed to directly bind to the DLGAP5 promoter, enhancing its transcriptional activity and suppressing the activation of the p53 signaling pathway. In vivo experiments further validated that downregulation of AR or DLGAP5 inhibited tumor growth and enhanced CD8+T cell infiltration. This study highlights the crucial roles of AR and DLGAP5 in TNBC growth and immune cell infiltration. Taken together, AR inhibits the p53 signaling pathway by promoting DLGAP5 expression, thereby impacting CD8+T cell infiltration in TNBC.

Screening of a kinase inhibitor library identified novel targetable kinase pathways in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is a highly invasive breast cancer subtype that is challenging to treat due to inherent heterogeneity and absence of estrogen, progesterone, and human epidermal growth factor 2 receptors. Kinase signaling networks drive cancer growth and development, and kinase inhibitors are promising anti-cancer strategies in diverse cancer subtypes. Kinase inhibitor screens are an efficient, valuable means of identifying compounds that suppress cancer cell growth in vitro, facilitating the identification of kinase vulnerabilities to target therapeutically. The Kinase Chemogenomic Set is a well-annotated library of 187 kinase inhibitor compounds that indexes 215 kinases of the 518 in the known human kinome representing various kinase networks and signaling pathways, several of which are understudied. Our screen revealed 14 kinase inhibitor compounds effectively inhibited TNBC cell growth and proliferation. Upon further testing, three compounds, THZ531, THZ1, and PFE-PKIS 29, had the most significant and consistent effects across a range of TNBC cell lines. These cyclin-dependent kinase (CDK)12/CDK13, CDK7, and phosphoinositide 3-kinase inhibitors, respectively, decreased metabolic activity in TNBC cell lines and promote a gene expression profile consistent with the reversal of the epithelial-to-mesenchymal transition, indicating these kinase networks potentially mediate metastatic behavior. These data identified novel kinase targets and kinase signaling pathways that drive metastasis in TNBC.

Hippophae Rhamnoides-derived Phytomedicine Nano-System Modulates Bax/Fas Pathways to Reduce Proliferation in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is the most common primary tumor of the breast with limited effectual drug availability. Therefore, the aim of the study is to develop an innovative phyto-nanomedicine (PNM) to cure TNBC with the least genotoxicity. Hereinafter, the sea buckthorn' extracted polyphenols (SBP), combine with metformin (MET), are synthesized as a novel PNM to evaluate its anti-tumor properties, effectiveness, and mechanism of action in TNBC in vitro and in vivo models. The SBP exhibits 16 new kinds of polyphenols that are been reported earlier which regulated cell development, proliferation, and programmed cell death (PCD) effectively. SBP-MET PNM inhibits MDA-MB-231 (47%), MDA-MB-436 (46%), and 4T1 (46%) cell proliferation but does not affect L929 normal murine cell development and successfully induce PCD (73.19%) in MDA-MB-231 cells. Mechanistically, in vivo SBP-MET proteome expression profiling reveals upregulation of proapoptotic Bax protein and activation of Fas signaling pathways convince downstream Daxx and FADD proteins, which further triggers Caspase-3 that prompts apoptosis in human TNBC cells by cleaving PARP-1 protein. Current findings establish innovative highly biocompatible phyto-nanomedicine that has significant potential to inhibit TNBC cell growth and induce regulated cell death (RCD) in vivo model, thereby opening a new arena for TNBC therapy.

Inhibition of ABI2 ubiquitination-dependent degradation suppresses TNBC cell growth via down-regulating PI3K/Akt signaling pathway

Triple negative breast cancer (TNBC) is a type of cancer that lacks receptor expression and has complex molecular mechanisms. Recent evidence shows that the ubiquitin-protease system is closely related to TNBC. In this study, we obtain a key ubiquitination regulatory substrate-ABI2 protein by bioinformatics methods, which is also closely related to the survival and prognosis of TNBC. Further, through a series of experiments, we demonstrated that ABI2 expressed at a low level in TNBC tumors, and it has the ability to control cell cycle and inhibit TNBC cell migration, invasion and proliferation. Molecular mechanism studies proved E3 ligase CBLC could increase the ubiquitination degradation of ABI2 protein. Meanwhile, RNA-seq and IP experiments indicated that ABI2, acting as a crucial factor of tumor suppression, can significantly inhibit PI3K/Akt signaling pathway via the interaction with Rho GTPase RAC1. Finally, based on TNBC drug target ABI2, we screened and found that FDA-approved drug Colistimethate sodium(CS) has significant potential in suppressing the proliferation of TNBC cells and inducing cell apoptosis, making it a promising candidate for impeding the progression of TNBC.

Exploration of the sensitization effect of Chaihu Shugan powder on chemotherapy for triple-negative breast cancer and its active ingredients.

Chemotherapy plays a crucial role in the clinical treatment of triple-negative breast cancer (TNBC), but drug resistance limits its clinical application. The active ingredients of Chaihu Shugan Powder (CSP; Bupleurum Liver-Coursing Powder), quercetin and luteolin, both belong to flavonoid compounds and have significant anti-tumor potential, which can promote chemotherapy sensitivity. However, the correlation between the two and TNBC paclitaxel (PTX) chemotherapy sensitivity is unknown. We collected herbal components of CSP from the TCMSP database, and screened effective molecules and corresponding targets. STRING database was utilized to construct a protein-protein interaction network combining effective molecules and target genes. The top 50 nodes ranked by affinity were chosen for subsequent functional analysis, and the drug-active ingredient-gene interaction network was established using Cytoscape software. Molecular docking was used to determine the small molecules that target TNBC PTX resistance. The "clusterProfiler" package was utilized for GO and KEGG enrichment analyses on the top 50 genes to determine the pathways affected by CSP. Cell counting and colony formation assays evaluated cell viability, IC50 values, and proliferation capacity. Flow cytometry tested PTX intracellular accumulation. Western blot assayed the expression of TNF pathway-related proteins. Active ingredients of CSP, quercetin and luteolin, could inhibit TNBC cell proliferation and promote PTX chemotherapy sensitization. Quercetin and luteolin repressed the TNF signaling pathway and promoted PTX chemotherapy sensitization. Quercetin and luteolin could inhibit TNBC cell proliferation and promote PTX chemotherapy sensitization through the TNF signaling pathway. Therefore, the use of quercetin and luteolin plus PTX treatment provides a prospective strategy for TNBC treatment.

Erianin inhibits the progression of triple-negative breast cancer by suppressing SRC-mediated cholesterol metabolism

Triple-negative breast cancer (TNBC) is highly malignant and lacks effective biotherapeutic targets. The development of efficient anticancer drugs with low toxicity and few side effects is a hotspot in TNBC treatment research. Although erianin is known to have potent antitumor activity, its regulatory mechanism and target in TNBC have not been fully elucidated, hampering further drug development. This study showed that erianin can significantly inhibit TNBC cell proliferation and migration, promote cell apoptosis, and inhibit the growth of transplanted tumors in mice. Mechanistically, through network pharmacology analysis, molecular docking and cellular thermal shift assays, we preliminarily identified SRC as the cellular target of erianin. Erianin potently inhibited the expression of SRC, which mediated the anticancer effect of erianin in TNBC. Moreover, erianin can downregulate the expression of genes related to cholesterol synthesis and uptake by targeting SRC, interfering with cholesterol levels in TNBC, thereby inhibiting the progression of TNBC in vivo and in vitro. Taken together, our results suggest that erianin may inhibit the progression of TNBC by suppressing SRC-mediated cholesterol metabolism, and erianin has the great potential to be an effective treatment for TNBC patients.

Abstract PO3-25-10: Selective and Effective Targeting of Triple-Negative Breast Cancer by CMPD1: Inhibition of MAP kinase-activated protein kinase 2 and Microtubule Dynamics

Abstract Background: Microtubule-targeting agents (MTAs) have been widely utilized in cancer treatment, but the development of novel MTAs has faced limitations. CMPD1, a highly selective inhibitor of MAP kinase-activated protein kinase 2 (MK2) with dual functions as a microtubule inhibitor, shows promise as a targeted therapy for triple-negative breast cancer (TNBC). This study aimed to investigate the effects of CMPD1, targeting MK2 and acting as a microtubule inhibitor, on cell cycle progression, microtubule dynamics, and TNBC-related processes. Methods: CMPD1 was evaluated in non-transformed retinal pigment epithelial cells (RPE1) and TNBC cell lines, including MDA-MB-231 and MDA-MB-468 cells. Time-lapse microscopy and immunofluorescence staining were employed to observe the effects of CMPD1 on mitotic progression, cellular morphology, activation of the spindle assembly checkpoint (SAC), and microtubule dynamics. Anchorage-independent growth assays and in vivo studies using a TNBC xenograft model were performed to assess the impact of CMPD1 on colony formation, tumor growth, and metastasis. Results: CMPD1 treatment induced prometaphase arrest and mitotic slippage in both non-transformed and TNBC cells. This arrest was associated with SAC activation, evidenced by increased levels of Mad1 at kinetochores. CMPD1 exhibited robust microtubule depolymerization, primarily targeting microtubule plus-ends and inhibiting microtubule polymerization in vitro. Furthermore, CMPD1 disrupted the organization of the microtubule network in interphase cells and perturbed the mitotic spindle during mitosis. Additionally, CMPD1 treatment resulted in the formation of cytoplasmic protrusions, membrane deformations, and dynamic movement of clustered mitotic chromosomes. These morphological alterations were indicative of the impact of CMPD1 on cellular structures and processes during mitosis. Notably, CMPD1 demonstrated significant efficacy in inhibiting anchorage-independent growth in TNBC cells, highlighting its potential as an anti-TNBC agent. In the TNBC xenograft model, CMPD1 exhibited superior anti-tumor activity compared to the standard chemotherapeutic agent Taxol. Tumor growth was markedly suppressed, and the formation of metastatic lesions was significantly reduced upon CMPD1 treatment. Furthermore, CMPD1 treatment effectively inhibited TNBC cell migration and invasion, likely through its modulation of microtubule dynamics. The disruption of microtubule dynamics by CMPD1 impaired the ability of TNBC cells to migrate and invade surrounding tissues, which are crucial processes in cancer metastasis. Importantly, CMPD1 showed high selectivity for TNBC cells, exerting minimal cytotoxic effects on normal cells, and could be efficiently washed out. This selectivity and washout ability enhance its potential as a targeted therapeutic agent for TNBC treatment. Conclusions: CMPD1, a highly selective MK2 inhibitor and microtubule inhibitor, exerts profound effects on cell cycle progression, microtubule dynamics, and TNBC-related processes. It induces prometaphase arrest, mitotic slippage, and microtubule depolymerization, resulting in altered cellular morphology. CMPD1 displays high selectivity for TNBC cells, while sparing normal cells and allowing efficient washout, making it a promising therapeutic agent for TNBC treatment. Its inhibition of anchorage-independent growth, suppression of tumor growth in vivo, and ability to impair TNBC cell migration and invasion further underscore its potential as a targeted anti-TNBC therapy. Citation Format: Mamoru Takada, Yu-Chia Chen, Yu Muhan, Hideyuki Yamada, Junta Sakakibara, Hiroshi Fujimoto, Takeshi Nagashima, Masayuki Otsuka, Motoki Takaku, Aussie Suzuki. Selective and Effective Targeting of Triple-Negative Breast Cancer by CMPD1: Inhibition of MAP kinase-activated protein kinase 2 and Microtubule Dynamics [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-25-10.

Abstract PO1-24-08: Targeting of HER3 potentiates the antitumor activity of paclitaxel against triple negative breast cancer

Abstract Background: Triple negative breast cancer (TNBC) is characterized by the absence of expression of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) overexpression. This aggressive form of breast cancer carries a higher risk of recurrence compared to other subtypes, presenting a significant clinical challenge. Although chemotherapy is commonly used as the primary treatment for a substantial number of TNBC patients, it is often accompanied by drug resistance and frequent tumor recurrence. Urgent efforts are needed to identify novel molecular targets specific to TNBC and develop effective therapies to combat this aggressive disease. The ultimate objective is to discover treatments capable of overcoming drug resistance and improving the overall survival of patients with TNBC. Methods: Immunohistochemistry was performed to examine the expression of HER3 in TNBC samples. Western blots were used to assess protein expression and activation. Cell proliferation and viability were determined by cell growth (MTS) assays. TCGA databases were analyzed to correlate HER3 mRNA expression with the clinical outcomes of TNBC patients. Specific shRNA was used to knockdown HER3 expression. LIVE/DEAD Cell Imaging was to detect live and dead cells. Orthotopic tumor models were established in nude mice to determine the capability of TNBC cells forming tumors and to test if our newly developed anti-HER3 monoclonal antibody (mAb) 4A7 could potentiate the antitumor activity of paclitaxel in vivo. Results: The chemo-resistant subline HCC1806-TR was obtained by subjecting TNBC HCC1806 cells to prolonged treatment with paclitaxel. In comparison to the parental cell line HCC1806-P, HCC1806-TR exhibited enhanced HER3 expression and activation. It is noteworthy that approximately half of the tested TNBC specimens and cell lines displayed elevated expression of HER3. Analyzing TCGA databases revealed that TNBC patients with high levels of HER3 mRNA expression in their tumors had significantly poorer overall survival (OS) and relapse-free survival (RFS) compared to those with low HER3 expression. Specifically knockdown of HER3 markedly inhibited TNBC cell proliferation and mammosphere formation in vitro and tumor growth in vivo. Blockade of HER3 signaling with our mAb (4A7) in combination with paclitaxel exerted significant antitumor activity against TNBC in vitro and in vivo. Conclusions: Our data demonstrate that increased HER3 is an effective therapeutic target for TNBC and our anti-HER3 mAb (4A7) may enhance the efficacy of paclitaxel in TNBC. Keywords: HER3, Anbibody, Chemotherapy, Triple Negative Breast Cancer Citation Format: Hui Lyu, Sanbao Ruan, Congcong Tan, Ann Thor, Bolin Liu. Targeting of HER3 potentiates the antitumor activity of paclitaxel against triple negative breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-24-08.

High MCM6 expression promotes proliferation and correlates with poor prognosis in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is an aggressive subtype with a poor prognosis. Minichromosome maintenance genes (MCM2-7) crucial for DNA replication are significant biomarkers for various tumor types; however, their roles in TNBC remain underexplored. We utilized four TNBC-related GEO databases to examine MCM2-7 gene expression and predict its prognosis in TNBC, performing single-cell analysis and GSEA to discover MCM6's potential function. The Cancer Dependency Map gene effect scores and CCK8 assay were used to assess MCM6's impact on TNBC cell proliferation. The correlations between MCM6 expression, immune infiltrates, and immune cells were also analyzed. WGCNA and LASSO Cox regression built a risk score model predicting TNBC patient survival based on MCM6-related gene expression. MCM2-7 gene expression was higher in TNBC tissues compared to adjacent normal tissues. High MCM6 expression correlated with shorter TNBC patient survival time. GSEA and single-cell analysis revealed a relationship between elevated MCM6 expression and the cell cycle pathway. MCM6 knockdown inhibited TNBC cell proliferation. A risk model featuring MCM6, CDC23, and CCNB1 effectively predicts TNBC patient survival. MCM6 overexpression in TNBC links to a worse prognosis and reduced cell proliferation upon MCM6 knockdown. We developed a risk score model based on MCM6-related genes predicting TNBC patient prognosis, potentially assisting future treatment strategies.

YAP1-induced RBM24 promotes the tumorigenesis of triple-negative breast cancer through the β-catenin pathway.

Triple-negative breast cancer (TNBC) is the most aggressive breast cancer subtype and refractory to current treatments. RBM24 is an RNA-binding protein and shows the ability to regulate tumor progression in multiple cancer types. However, its role in TNBC is still unclear. In this study, we analyzed publicly available profiling data from TNBC tissues and cells. Loss- and gain-of-function experiments were performed to determine the function of RBM24 in TNBC cells. The mechanism for RBM24 action in TNBC was investigated. RBM24 was deregulated in TNBC tissues and TNBC cells with depletion of SIPA1, YAP1, or ARID1A, three key regulators of TNBC. Compared to MCF10A breast epithelial cells, TNBC cells had higher levels of RBM24. Knockdown of RBM24 inhibited TNBC cell proliferation, colony formation, and tumorigenesis, while overexpression of RBM24 promoted aggressive phenotype in TNBC cells. YAP1 overexpression induced the expression of RBM24 and the RBM24 promoter-driven luciferase reporter. YAP1 was enriched at the promoter region of RBM24. Overexpression of RBM24 increased β-catenin-dependent transcriptional activity. Most importantly, knockdown of CTNNB1 rescued RBM24 aggressive phenotype in TNBC cells. Collectively, the YAP1/RBM24/β-catenin axis plays a critical role in driving TNBC progression. RBM24 may represent a novel therapeutic target for TNBC treatment.

The deubiquitinase BRCC3 increases the stability of ZEB1 and promotes the proliferation and metastasis of triple-negative breast cancer cells.

Triple negative breast cancer (TNBC) has a high recurrence rate, metastasis rate and mortality rate. The aim of this study is to identify new targets for the treatment of TNBC. Clinical samples are used for screening deubiquitinating enzymes (DUBs). MDA-MB-231 cells and a TNBC mouse model are used for in vitro and in vivo experiments, respectively. Western blot analysis is used to detect the protein expressions of DUBs, zinc finger E-box binding homeobox 1 (ZEB1), and epithelial-mesenchymal transition (EMT)-related markers. Colony formation and transwell assays are used to detect the proliferation, migration and invasion of TNBC cells. Wound healing assay is used to detect the mobility of TNBC cells. Immunoprecipitation assay is used to detect the interaction between breast cancer susceptibility gene 1/2-containing complex subunit 3 (BRCC3) and ZEB1. ZEB1 ubiquitination levels, protein stability, and protein degradation are also examined. Pathological changes in the lung tissues are detected via HE staining. Our results show a significant positive correlation between the expressions of BRCC3 and ZEB1 in clinical TNBC tissues. Interference with BRCC3 inhibits TNBC cell proliferation, migration, invasion and EMT. BRCC3 interacts with ZEB1 and interferes with BRCC3 to inhibit ZEB1 expression by increasing ZEB1 ubiquitination. Interference with BRCC3 inhibits TNBC cell tumorigenesis and lung metastasis in vivo. In all, this study demonstrates that BRCC3 can increase the stability of ZEB1, upregulate ZEB1 expression, and promote the proliferation, migration, invasion, EMT, and metastasis of TNBC cells, providing a new direction for cancer therapy.

Mechanism of Erianin anti-triple negative breast cancer based on transcriptomics methods and network pharmacology.

Triple negative breast cancer (TNBC) is a highly aggressive illness that lacks effective targeted treatments. Although Erianin has shown potential antitumor properties, its precise mechanism of action and target in TNBC remain unclear, hampering the development of drugs. The present study investigated the underlying mechanism of action of Erianin in treating TNBC by using transcriptomics and network pharmacology approaches. We evaluated Erianin's bioactivity in TNBC cell lines and xenograft tumor models. The results showed that Erianin significantly inhibited TNBC cell proliferation and impeded tumor growth. A subsequent analysis of transcriptomic and network pharmacological data identified 51 mutual targets. Analysis of protein-protein interactions identified eight hub targets. Furthermore, molecular docking indicated that the PPARA binding energy was the lowest for Erianin among the hub targets, followed by ROCK2, PDGFRB, CCND1, MUC1, and CDK1. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes functional enrichment analysis showed that the common targets were associated with multiple cancer-related signaling pathways, including focal adhesion, PI3K-Akt signaling pathway, Rap1 signaling pathway, microRNAs in cancer, and human papillomavirus infection. The results of the Western blot and immunohistochemistry experiment further showed that Erianin could suppress PI3K/Akt signaling pathway activation. After co-incubation with SC79, the cell inhibition rate of Erianin was decreased, which further confirmed that Erianin inhibits TNBC progression via the PI3K-AKT signaling pathway. In conclusion, our results indicated that Erianin has the potential to inhibit the proliferation of TNBC by downregulating the PI3K/AKT signaling pathway by transcriptomics and network pharmacology. Therefore, Erianin appears to be a promising compound for the effective treatment of TNBC.

METTL14/miR-29c-3p axis drives aerobic glycolysis to promote triple-negative breast cancer progression though TRIM9-mediated PKM2 ubiquitination.

The energy metabolic rearrangement of triple-negative breast cancer (TNBC) from oxidative phosphorylation to aerobic glycolysis is a significant biological feature and can promote the malignant progression. However, there is little knowledge about the functional mechanisms of methyltransferase-like protein 14 (METTL14) mediated contributes to TNBC malignant progression. Our study found that METTL14 expression was significantly upregulated in TNBC tissues and cell lines. Silencing METTL14 significantly inhibited TNBC cell growth and invasion invitro, as well as suppressed tumour growth. Mechanically, METTL14 was first found to activate miR-29c-3p through m6A and regulate tripartite motif containing 9 (TRIM9) to promote ubiquitination of pyruvate kinase isoform M2 (PKM2) and lead to its transition from tetramer to dimer, resulting in glucose metabolic reprogramming from oxidative phosphorylation to aerobic glycolysis to promote the progress of TNBC. Taken together, these findings reveal important roles of METTL14 in TNBC tumorigenesis and energy metabolism, which might represent a novel potential therapeutic target for TNBC.

Decanoylcarnitine Inhibits Triple-Negative Breast Cancer Progression via Mmp9 in an Intermittent Fasting Obesity Mouse.

Purpose: Treatment of triple-negative breast cancer (TNBC) remains challenging. Intermittent fasting (IF) has emerged as a promising approach to improve metabolic health of various metabolic disorders. Clinical studies indicate IF is essential for TNBC progression. However, the molecular mechanisms underlying metabolic remodeling in regulating IF and TNBC progression are still unclear. Methods: In this study, we utilized a robust mouse model of TNBC and exposed subjects to a high-fat diet (HFD) with IF to explore its impact on the metabolic reprogramming linked to cancer progression. To identify crucial serum metabolites and signaling events, we utilized targeted metabolomics and RNA sequencing (RNA-seq). Furthermore, we conducted immunoblotting, real-time quantitative polymerase chain reaction (RT-qPCR), cell migration assays, lentivirus-mediated Mmp9 overexpression, and Mmp9 inhibitor experiments to elucidate the role of decanoylcarnitine/Mmp9 in TNBC cell migration. Results: Our observations indicate that IF exerts notable inhibitory effects on both the proliferation and cancer metastasis. Utilizing targeted metabolomics and RNA-seq, we initially identified pivotal serum metabolites and signaling events in the progression of TNBC. Among the 349 serum metabolites identified, decanoylcarnitine was picked out to inhibit TNBC cell proliferation and migration. RNA-seq analysis of TNBC cells treated with decanoylcarnitine revealed its suppressive effects on extracellular matrix-related protein components, with a notable reduction observed in Mmp9. Further investigations confirmed that decanoylcarnitine could inhibit Mmp9 expression in TNBC cells, primary tumors, lung, and liver metastasis tissues. Mmp9 overexpression abolished the inhibitory effect of decanoylcarnitine on cell migration. Conclusion: This study pioneers the exploration of IF intervention and the role of decanoylcarnitine/Mmp9 in the progression of TNBC in obese mice, enhancing our comprehension of the potential roles of various dietary patterns in the process of cancer treatment.

AURKAIP1 actuates tumor progression through stabilizing DDX5 in triple negative breast cancer

Aurora-A kinase interacting protein 1 (AURKAIP1) has been proved to take an intermediary role in cancer by functioning as a negative regulator of Aurora-A kinase. However, it remains unclear whether and how AURKAIP1 itself would directly engage in regulating malignancies. The expression levels of AURKAIP1 were detected in triple negative breast cancer (TNBC) by immunohistochemistry and western blots. The CCK8, colony formation assays and nude mouse model were conducted to determine cell proliferation whereas transwell and wound healing assays were performed to observe cell migration. The interaction of AURKAIP1 and DEAD-box helicase 5 (DDX5) were verified through co-immunoprecipitation and successively western blots. From the results, we found that AURKAIP1 was explicitly upregulated in TNBC, which was positively associated with tumor size, lymph node metastases, pathological stage and unfavorable prognosis. AURKAIP1 silencing markedly inhibited TNBC cell proliferation and migration in vitro and in vivo. AURKAIP1 directly interacted with and stabilized DDX5 protein by preventing ubiquitination and degradation, and DDX5 overexpression successfully reversed proliferation inhibition induced by knockdown of AURKAIP1. Consequently, AURKAIP1 silencing suppressed the activity of Wnt/β-catenin signaling in a DDX5-dependent manner. Our study may primarily disclose the molecular mechanism by which AURKAIP1/DDX5/β-catenin axis modulated TNBC progression, indicating that AURKAIP1 might serve as a therapeutic target as well as a TNBC-specific biomarker for prognosis.

Ezetimibe inhibits triple-negative breast cancer proliferation and promotes cell cycle arrest by targeting the PDGFR/AKT pathway

Cholesterol levels were strongly associated with tumor progression and metastasis. Targeted cholesterol metabolism has broad prospects in tumor treatment. Ezetimibe, the only FDA-approved inhibitor of cholesterol absorption, has been reported to be able to inhibit angiogenesis in liver cancer. However, the efficacy and specific mechanisms of Ezetimibe in the treatment of Triple-Negative Breast Cancer (TNBC)have not been reported. Our research shows Ezetimibe inhibits TNBC cell proliferation and blocks the cell cycle in the G1 phase. Mechanistically, Ezetimibe inhibits the activation of PDGFRβ/AKT pathway, thereby promoting cell cycle arrest and inhibiting cell proliferation. By overexpressing PDGFRβ in TNBC cells, we found that PDGFRβ significantly reduced the inhibitory effect of Ezetimibe on TNBC cell proliferation and the cell cycle. Similarly, SC79, an AKT agonist, can reduce the proliferation inhibitory and cycle-blocking effects of Ezetimibe on TNBC cells. Furthermore, the AKT inhibitor MK2206 enhanced the inhibitory effect of Ezetimibe on the cell cycle and proliferation ability of TNBC cells overexpressing PDGFRβ. In xenograft tumor models, we also found that Ezetimibe inhibited TNBC growth, an effect that can be blocked by overexpression of PDGFR or activation of AKT. In summary, we have demonstrated that EZ inhibits the PDGFR/AKT pathway, thereby halting TNBC cycle progression and tumor growth.

SILAC proteomics based on 3D cell spheroids unveils the role of RAC2 in regulating the crosstalk between triple-negative breast cancer cells and tumor-associated macrophages

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer, and is characterized by a high infiltration of tumor-associated macrophages (TAMs). TAMs contribute significantly to tumor progression by intricately interacting with tumor cells. Deeply investigating the interaction between TNBC cells and TAMs is of great importance for finding potential biomarkers and developing novel therapeutic strategies to further improve the clinical outcomes of TNBC patients. In this study, we confirmed the interplay using both 3D and 2D co-culture models. The stable-isotype labeling by amino acids in cell culture (SILAC)-based quantitative proteomics was conducted on 3D cell spheroids containing TNBC cells and macrophages to identify the potential candidate in regulating the crosstalk between TNBC and TAMs. Ras-related C3 botulinum toxin substrate 2 (RAC2) was identified as a potential molecule for further exploration, given its high expression in TNBC and positive correlation with M2 macrophage infiltration. The suppression of RAC2 inhibited TNBC cell proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) in vitro. Meanwhile, knocking down RAC2 in TNBC cells impaired macrophage recruitment and M2 polarization. Mechanistically, RAC2 exerted its roles in TNBC cells and TAMs by regulating the activation of P65 NF-κB and P38 MAPK, while TAMs further elevated RAC2 expression and P65 NF-κB activation by secreting soluble mediators including IL-10. These findings highlight the significance of RAC2 as a crucial molecule in the crosstalk between TNBC and TAMs, suggesting it could be a promising therapeutic target in TNBC.