Suppressed Triple-negative Breast Cancer Cell Research Articles

Exploring the regulatory role of FBXL19-AS1 in triple-negative breast cancer through the miR-378a-3p/OTUB2 axis.

The regulatory potential of long noncoding RNA (lncRNA) FBXL19-AS1 has been highlighted in various cancers, but its effect on triple-negative breast cancer (TNBC) remains unclear. Here, we aimed to elucidate the role of FBXL19-AS1 in TNBC and its underlying mechanism. RT-qPCR was employed to detect the expressions of FBXL19-AS1 and miR-378a-3p in tissues and cells. Immunohistochemical staining and western blot were utilized to detect the expression levels of proteins. Cell activities were detected using flow cytometry, CCK-8, and transwell assay. Dual-luciferase reporter and RNA immunoprecipitation (RIP) assays were deployed to investigate interactions of different molecules. Protein-protein interaction (PPI) network, gene ontology (GO), and Kyoto encyclopedia of genes and genomes (KEGG) pathways were used to analyze the downstream pathway. In vivo xenograft model was conducted to detect the effect of FBXL19-AS1 on tumor growth. FBXL19-AS1 was overexpressed in TNBC tissues and cell lines compared with counterparts. FBXL19-AS1 knockdown suppressed TNBC cell activities, whereas its overexpression exhibited the opposite effect. Mechanistically, FBXL19-AS1 was found to interact with miR-378a-3p. Further analysis revealed that miR-378a-3p exerted tumor-suppressive effects in TNBC cells. Additionally, miR-378a-3p targeted and downregulated the expression of ubiquitin aldehyde binding 2 (OTUB2), a deubiquitinase associated with TNBC progression. In vivo experiments substantiated the inhibitory effects of FBXL19-AS1 knockdown on TNBC tumorigenesis, and a miR-378a-3p inhibitor partially rescued these effects. The downstream pathway of the miR-378a-3p/OTUB2 axis was explored, revealing connections with proteins involved in modifying other proteins, removing ubiquitin molecules, and influencing signaling pathways, including the Hippo signaling pathway. Western blot analysis confirmed changes in YAP and TAZ expression levels, indicating a potential regulatory network. In summary, FBXL19-AS1 promotes exacerbation in TNBC by suppressing miR-378a-3p, leading to increased OTUB2 expression. The downstream mechanism may be related to the Hippo signaling pathway. These findings propose potential therapeutic targets for TNBC treatment.

PYCR3 modulates mtDNA copy number to drive proliferation and doxorubicin resistance in triple-negative breast cancer

Triple-negative breast cancer (TNBC) poses significant challenges in treatment due to its aggressive nature and limited therapeutic targets. Understanding the underlying molecular mechanisms driving TNBC progression and chemotherapy resistance is imperative for developing effective therapeutic strategies. Thus, in this study, we aimed to elucidate the role of pyrroline-5-carboxylate reductase 3 (PYCR3) in TNBC pathogenesis and therapeutic response. We observed that PYCR3 is significantly upregulated in TNBC specimens compared to normal breast tissues, correlating with a poorer prognosis in TNBC patients. Knockdown of PYCR3 not only suppresses TNBC cell proliferation but also reverses acquired resistance of TNBC cells to doxorubicin, a commonly used chemotherapeutic agent. Mechanistically, we identified the mitochondrial localization of PYCR3 in TNBC cells and demonstrated its impact on TNBC cell proliferation and sensitivity to doxorubicin through the regulation of mtDNA copy number and mitochondrial respiration. Importantly, Selective reduction of mtDNA copy number using the mtDNA replication inhibitor 2′, 3′-dideoxycytidine effectively recapitulates the phenotypic effects observed in PYCR3 knockout, resulting in decreased TNBC cell proliferation and the reversal of doxorubicin resistance through apoptosis induction. Thus, our study underscores the clinical relevance of PYCR3 and highlight its potential as a therapeutic target in TNBC management. By elucidating the functional significance of PYCR3 in TNBC, our findings contribute to a deeper understanding of TNBC biology and provide a foundation for developing novel therapeutic strategies aimed at improving patient outcomes.

Exosomal circRHCG promotes breast cancer metastasis via facilitating M2 polarization through TFEB ubiquitination and degradation

Triple-negative breast cancer (TNBC) is a highly aggressive cancer with distant metastasis. Accumulated evidence has demonstrated that exosomes are involved in TNBC metastasis. Elucidating the mechanism underlying TNBC metastasis has important clinical significance. In the present study, exosomes were isolated from clinical specimens and TNBC cell lines. Colony formation, EdU incorporation, wound healing, and transwell assays were performed to examine TNBC cell proliferation, migration, and metastasis. Macrophage polarization was evaluated by flow cytometry and RT-qPCR analysis of polarization markers. A mouse model of subcutaneous tumor was established for assessment of tumor growth and metastasis. RNA pull-down, RIP and Co-IP assays were used for analyzing molecular interactions. Here, we proved that high abundance of circRHCG was observed in exosomes derived from TNBC patients, and increased exosomal circRHCG indicated poor prognosis. Silencing of circRHCG suppressed TNBC cell proliferation, migration, and metastasis. TNBC cell-derived exosomes promoted M2 polarization via delivering circRHCG. Exosomal circRHCG stabilized BTRC mRNA via binding FUS and naturally enhanced BTRC expression, thus promoting the ubiquitination and degradation of TFEB in THP-1 cells. In addition, knockdown of BTRC or overexpression of TFEB counteracted exosomal circRHCG-mediated facilitation of M2 polarization. Furthermore, exosomal circRHCG promoted TNBC cell proliferation and metastasis by facilitating M2 polarization. Knockdown of circRHCG reduced tumor growth, metastasis, and M2 polarization through the BTRC/TFEB axis in vivo. In summary, exosomal circRHCG promotes M2 polarization by stabilizing BTRC and promoting TFEB degradation, thereby accelerating TNBC metastasis and growth. Our study provides promising therapeutic strategies against TNBC.

Fbxw7 suppresses carcinogenesis and stemness in triple-negative breast cancer through CHD4 degradation and Wnt/β-catenin pathway inhibition

BackgroundCancer stem cells (CSCs) are a small population of cells in tumor tissues that can drive tumor initiation and promote tumor progression. A small number of previous studies indirectly mentioned the role of F-box and WD repeat domain-containing 7 (FBXW7) as a tumor suppressor in Triple-negative breast cancer (TNBC). However, few studies have focused on the function of FBXW7 in cancer stemness in TNBC and the related mechanism.MethodsWe detected FBXW7 by immunohistochemistry (IHC) in 80 TNBC patients. FBXW7 knockdown and overexpression in MD-MBA-231 and HCC1937 cell models were constructed. The effect of FBXW7 on malignant phenotype and stemness was assessed by colony assays, flow cytometry, transwell assays, western blot, and sphere formation assays. Immunoprecipitation-Mass Spectrometry (IP-MS) and ubiquitination experiments were used to find and verify potential downstream substrate proteins of FBXW7. Animal experiments were constructed to examine the effect of FBXW7 on tumorigenic potential and cancer stemness of TNBC cells in vivo.ResultsThe results showed that FBXW7 was expressed at low levels in TNBC tissues and positively correlated with prognosis of TNBC patients. In vitro, FBXW7 significantly inhibited colony formation, cell cycle progression, cell migration, EMT process, cancer stemness and promotes apoptosis. Further experiments confirmed that chromodomain-helicase-DNA-binding protein 4 (CHD4) is a novel downstream target of FBXW7 and is downregulated by FBXW7 via proteasomal degradation. Moreover, CHD4 could promote the nuclear translocation of β-catenin and reverse the inhibitory effect of FBXW7 on β-catenin, and ultimately activate the Wnt/β-catenin pathway. Rescue experiments confirmed that the FBXW7-CHD4-Wnt/β-catenin axis was involved in regulating the maintenance of CSC in TNBC cells. In animal experiments, FBXW7 reduced CSC marker expression and suppressed TNBC cell tumorigenesis in vivo.ConclusionsTaken together, these results highlight that FBXW7 degrades CHD4 protein through ubiquitination, thereby blocking the activation of the Wnt/β-catenin pathway to inhibit the stemness of TNBC cells. Thus, targeting FBXW7 may be a promising strategy for therapeutic intervention against TNBC.

MicroRNA‑606 inhibits the growth and metastasis of triple‑negative breast cancer by targeting Stanniocalcin1.

Triple‑negative breast cancer (TNBC) is associated with a poor prognosis; however, treatments for TNBC are limited, with poor outcomes. MicroRNAs (miRNAs/miRs) are small non‑coding RNA molecules that are able to regulate gene expression. The present study aimed to identify differentially expressed miRNAs inpatients with breast cancer, and to investigate the functional role of the identified miRNA targets and their effects invitro and invivo. Transfection with miR‑606 suppressed TNBC cell proliferation, migration, invasion and tumor sphere‑forming ability, as determined using trypan blue, Transwell and sphere formation assays. Moreover, miR‑606 induced the apoptosis of TNBC cells, as determined by flow cytometric analysis. Furthermore, intratumoral injections of miR‑606 mimics suppressed tumor growth in MDA‑MB‑231 xenografts. In addition, MDA‑MB‑231 cells transfected with miR‑606 mimics exhibited decreased lung metastatic nodules in a mouse tail vein injection model. Notably, miR‑606 and STC1 expression had opposing effects on the overall survival ofpatients with TNBC. The results of the present study suggested a novel tumor suppressor function for miR‑606 in TNBC, thus indicating its potential application in the development of anticancer miRNA therapeutics.

WTAP Mediates NUPR1 Regulation of LCN2 Through m6A Modification to Influence Ferroptosis, Thereby Promoting Breast Cancer Proliferation, Migration and Invasion.

Ferroptosis is involved in various pathophysiological diseases, including triple-negative breast cancer (TNBC). Targeting ferroptosis is considered as a novel anti-TNBC strategy. Nevertheless, the regulatory mechanism of ferroptosis during TNBC progression is unclear. Here, the role of WTAP in ferroptosis during TNBC progression was investigated.The clinicopathological significance of WTAP, NUPR1 and LCN2 was analyzed by Kaplan-Meier method. Cell viability was assessed using MTT assay. Transwell assay was employed to analyze cell migration and invasion. GSH/GSSG and Fe2+ levels in TNBC cells were analyzed using kits. m6A level was examined using m6A dot blot assay. NUPR1 mRNA stability was analyzed using RNA degradation assay. RIP was performed to analyze the interaction between eIF3a and NURP1.Herein, our results revealed that WTAP, NUPR1 and LCN2 expressions were significantly elevated in TNBC. NUPR1 silencing inhibited TNBC cell proliferation, migration and invasion by inducing ferroptosis. NUPR1 positively regulated LCN2 expression in TNBC cells, and LCN2 knockdown induced ferroptosis to suppress TNBC cell malignant behaviors. Our molecular study further revealed that WTAP promoted NUPR1 expression in an m6A-EIF3A mediated manner. And, as expected, WTAP knockdown promoted ferroptosis to suppress TNBC cell malignant behaviors, which were abrogated by NUPR1 overexpression.WTAP upregulated LCN2 by regulation of NUPR1 m6A modification, thereby suppressing ferroptosis to contribute to accelerate TNBC progression. Our study revealed the cancer-promoting effect of WTAP, NUPR1 and LCN2 in TNBC and clarified the relevant mechanism, providing a theoretical basis for developing novel diagnostic and therapeutic strategies for TNBC.

Natural compound So-2 suppresses triple-negative breast cancer through inducing ferroptosis via downregulating transcription factor E2F7

Triple-negative breast cancer (TNBC), accounting for about 15∼18% of all breast cancers, is notorious for its poor prognosis, high rate of relapse and short overall survival. Because of lacking effective therapeutic targets or drugs, treatment of TNBC in clinical encounters great obstacle. Siegesbeckiaorientalis L. have been used as a traditional Chinese medicine “Xi-Xian-Cao” for centuries with multiple medicinal benefits including cancerous treatment. We have reported the isolation of twenty-seven germacranolides including So-2 from the aerial parts of S. orientalis with potent cytotoxicity against breast cancer cells. The studyaims to verified the anti-TNBC function of the natural compound So-2 both in vitro and vivo and uncover the underlying mechanism. The results showed that So-2 caused cell cycle arrest and suppress TNBC cell proliferation and migration. Also, So-2 was first identified to be a bona fide ferroptosis inducer in TNBC cells. So-2 effectively suppressed tumor growth of TNBC by using an orthotopic transplantation tumor model. We also characterized the oncogenic role of the transcription factor E2F7 in TNBC. E2F7 was demonstrated to be involved in the ferroptosis-inducing and tumor suppression effect of So-2. Altogether, So-2 exhibits inhibitory effect on TNBC both in vitro and vivo by inducing TNBC ferroptosis via downregulating the expression of E2F7. These findings provide valuable insight into the pathogenesis of TNBC. The natural compound So-2, isolated from Chinese traditional medicine, might be a prospective drug candidate in TNBC therapy.

Identification of lactate-related subgroups and prognostic model in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is a highly aggressive subtype of breast cancer that exhibits elevated glycolytic capacity. Lactate, as a byproduct of glycolysis, is considered a major oncometabolite that plays an important role in oncogenesis and remodeling of the tumor microenvironment. However, the potential roles of lactate in TNBC are not yet fully understood. In this study, our goal was to identify prognosis-related lactate genes (PLGs) and construct a lactate-related prognostic model (LRPM) for TNBC. First, we applied lactate-related genes to classify TNBC samples using a hierarchical clustering algorithm. Then, we performed the log-rank analysis and the least absolute shrinkage and selection operator analysis to screen PLGs and construct the LRPM. The biological functions of the identified PLGs in TNBC were investigated using CCK8 assay and clone formation assay. Finally, we constructed a nomogram based on the lactate-risk score and tumor clinical stage. We used the operating characteristic curve and decision curve analysis to evaluate the predictive capability of the nomogram. Our results showed that the TNBC samples could be classified into two subgroups with different survival probabilities. Three genes (NDUFAF3, CARS2 and FH), which can suppress TNBC cell proliferation, were identified as PLGs. Moreover, the LRPM and nomogram exhibited excellent predictive performance for TNBC patient prognosis. We have developed a novel LRPM that enables risk stratification and identification of poor molecular subtypes in TNBC patients, showing great potential in clinical practice.

CircKIF4A combines EIF4A3 to stabilize SDC1 expression to activate c-src/FAK and promotes TNBC progression

Triple-negative breast cancer (TNBC) is recognized for its poor prognosis and limited options for treatment. Circular RNA KIF4A (circKIF4A) was documented to be abnormally overexpressed in TNBC and was correlated with a poor survival rate. The objective of this study is to further examine the functional role of circKIF4A and its underlying mechanism. CircKIF4A was significantly upregulated in TNBC and the knockdown of circKIF4A suppressed TNBC cell proliferation, migration, and invasion. CircKIF4A was directly bound to EIF4A3, which interacted with SDC1. Knockdown of circKIF4A reduced interaction between EIF4A3 and SDC1 as well as SDC1 mRNA stability. SDC1 activated the c-src/FAK signaling pathways and finally promoted TNBC progression. circKIF4A induced TNBC progress in the in vivo mouse model via SDC1. CircKIF4A interacts with EIF4A3 to stabilize SDC1 mRNA, which activates the c-src/FAK signaling pathways and promotes TNBC progression. This may provide a potential therapy for TNBC treatment.

S1P1 Threonine 236 Phosphorylation Mediates the Invasiveness of Triple-Negative Breast Cancer and Sensitivity to FTY720.

Hyperactive sphingosine 1-phosphate (S1P) signaling is associated with a poor prognosis of triple-negative breast cancer (TNBC). Despite recent evidence that links the S1P receptor 1 (S1P1) to TNBC cell survival, its role in TNBC invasion and the underlying mechanisms remain elusive. Combining analyses of human TNBC cells with zebrafish xenografts, we found that phosphorylation of S1P receptor 1 (S1P1) at threonine 236 (T236) is critical for TNBC dissemination. Compared to luminal breast cancer cells, TNBC cells exhibit a significant increase of phospho-S1P1 T236 but not the total S1P1 levels. Misexpression of phosphorylation-defective S1P1 T236A (alanine) decreases TNBC cell migration in vitro and disease invasion in zebrafish xenografts. Pharmacologic disruption of S1P1 T236 phosphorylation, using either a pan-AKT inhibitor (MK2206) or an S1P1 functional antagonist (FTY720, an FDA-approved drug for treating multiple sclerosis), suppresses TNBC cell migration in vitro and tumor invasion in vivo. Finally, we show that human TNBC cells with AKT activation and elevated phospho-S1P1 T236 are sensitive to FTY720-induced cytotoxic effects. These findings indicate that the AKT-enhanced phosphorylation of S1P1 T236 mediates much of the TNBC invasiveness, providing a potential biomarker to select TNBC patients for the clinical application of FTY720.

Icariin Induces Triple-Negative Breast Cancer Cell Apoptosis and Suppresses Invasion by Inhibiting the JNK/c-Jun Signaling Pathway.

Breast cancer is a common cancer worldwide. Triple-negative breast cancer (TNBC) is an aggressive form of breast cancer characterized by a poor prognosis. Icariin (ICA) is a flavonoid glycoside purified from the natural product Epimedium, which is reported to exert an inhibitory effect on a variety of cancers. However, molecular mechanisms behind ICA suppressed TNBC remain elusive. The curative effects of ICA on TNBC cells and potential targets were predicted by network pharmacology and molecular biology methods screening, and the mechanism of inhibition was explained through in vitro experiments such as cell function determination, Western blot analysis, molecular docking verification, etc. This study showed that ICA inhibits TNBC cell functions such as proliferation, migration, and invasion in a dose-dependent manner. ICA could induce redox-induced apoptosis in TNBC cell, as shown by ROS upregulation. As a result of network pharmacology, ICA was predicted to be able to inhibit the MAPK signaling pathway. ICA treatment inhibited the expression of JNK and c-Jun and downregulated the antiapoptotic gene cIAP-2. Our results suggested that ICA could induce apoptosis by inducing an excessive accumulation of ROS in cells and suppress TNBC cell invasion via the JNK/c-Jun signaling pathway. We demonstrated that ICA can effectively inhibit cell proliferation and induced apoptosis of TNBC cells. In addition, ICA could inhibit TNBC cell invasion through the JNK/c-Jun signaling pathway. The above suggests that ICA may become a potential drug for TNBC.

Disruption of estrogen receptor beta's DNA binding domain impairs its tumor suppressive effects in triple negative breast cancer.

Triple negative breast cancer (TNBC) is an aggressive sub-type of the disease which accounts for a disproportionately high percentage of breast cancer morbidities and mortalities. For these reasons, a better understanding of TNBC biology is required and the development of novel therapeutic approaches are critically needed. Estrogen receptor beta (ERβ) is a reported tumor suppressor that is expressed in approximately 20% of primary TNBC tumors, where it is associated with favorable prognostic features and patient outcomes. Previous studies have shown that ERβ mediates the assembly of co-repressor complexes on DNA to inhibit the expression of multiple growth promoting genes and to suppress the ability of oncogenic transcription factors to drive cancer progression. To further elucidate the molecular mechanisms by which ERβ elicits its anti-cancer effects, we developed MDA-MB-231 cells that inducibly express a mutant form of ERβ incapable of directly binding DNA. We demonstrate that disruption of ERβ's direct interaction with DNA abolishes its ability to regulate the expression of well characterized immediate response genes and renders it unable to suppress TNBC cell proliferation. Loss of DNA binding also diminishes the ability of ERβ to suppress oncogenic NFκB signaling even though it still physically associates with NFκB and other critical co-factors. These findings enhance our understanding of how ERβ functions in this disease and provide a model system that can be utilized to further investigate the mechanistic processes by which ERβ elicits its anti-cancer effects.

Sanyin Formula Enhances the Therapeutic Efficacy of Paclitaxel in Triple-Negative Breast Cancer Metastases through the JAK/STAT3 Pathway in Mice.

Sanyin formula (SYF) is used as a complementary treatment for triple-negative breast cancer (TNBC). The purpose of this study was to identify the potential functional components and clarify the underlying molecular mechanisms of SYF in TNBC. High-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was used to identify the main components of SYF extracts. Network pharmacology and bioinformatic analyses were carried out to identify potential candidate targets of SYF in TNBC. Cell proliferation was determined with a Celigo imaging cytometer. Wound-healing and Transwell assays were adopted to evaluate cell migration. A Transwell cell-invasion assay was performed with Matrigel-coated membranes. In vivo bioluminescence imaging (BLI) and pathological analyses illustrated the effect of SYF on cancer cell metastasis in tumour-bearing mice. The inhibitory mechanism of SYF was investigated via quantitative PCR (qPCR) and Western blotting. We found that 3,4-dihydroxyphenyllactic acid, kaempferol, p-coumaric acid, and vanillic acid may be the active components of SYF. Molecular docking confirmed that kaempferol, p-coumaric acid, vanillic acid, and 3,4-dihydroxyphenyllactic acid bound stably to proteins such as AKR1C3, MMPs, and STAT3. SYF extract suppressed TNBC cell proliferation, migration, invasion, and metastasis by inhibiting JAK/STAT3 signalling and then regulating downstream genes, such as MMP-2/MMP-9. SYF regulates the expression of genes involved in cell proliferation, migration, and invasion by regulating the JAK/STAT3 signalling pathway and finally inhibits tumour cell metastasis in TNBC. The present study clarifies the mechanism by which SYF inhibits TNBC metastasis and lays an experimental foundation for the continued clinical development of SYF targeting the JAK/STAT3 pathway.

A novel Pim-1 kinase-targeted photosensitizer to combat triple-negative breast cancer with enhanced photodynamic efficacy and reduced metastasis

As one of the most malignant breast cancer types, triple-negative breast cancer (TNBC) treatment is a particularly great challenge in the current era of precision medicine. Due to the limited effectiveness and targeting of current treatment methods, it is urgently need to develop an effective precision strategy to combat TNBC. Herein, a novel TNBC targeted molecular drug based on zinc(II) phthalocyanine (BF-Pc) was rationally designed by conjugating a Pim-1 kinase inhibitor with high TNBC selectivity. In our in vitro investigation, BF-Pc not only exhibits extremely high targeting to TNBC cells overexpressed Pim-1 kinases, but also shows ultra-efficient photocytotoxicity toward TNBC cells far beyond the normal cells. Significantly, BF-Pc-induced PDT can obviously suppress TNBC cell migration and invasion. The proposed manner in our work offers a promising approach to combat TNBC effectively and precisely.

GRWD1 affects the proliferation, apoptosis, invasion and migration of triple negative breast cancer through the Notch signaling pathway.

Breast cancer is a highly heterogeneous tumor, among which triple negative breast cancer (TNBC) is the most invasive and prone to recurrence and metastasis. The present study aimed to investigate the regulatory mechanisms of glutamate-rich WD-repeat-containing protein 1 (GRWD1) in TNBC cells. The expression of GRWD1 in the normal human breast epithelial cells and human breast cancer cells was detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis. The transfection effects of small interfering RNA (siRNA)-GRWD1 and overexpression (Ov)-Notch1 were also confirmed by RT-qPCR and western blotting. The proliferation, apoptosis, invasion and migration of transfected cells were in turn analyzed by Cell Counting Kit-8, 5-Ethynyl-2'-deoxyuridine, Matrigel and wound healing assays. The expression of proteins related to proliferation, apoptosis, metastasis, epithelial-mesenchymal transition and the Notch signaling pathway was detected by western blotting. As a result, GRWD1 expression was upregulated in breast cancer cells and was revealed to be highest in MDA-MB-231 and HCC1937 cells. GRWD1 knockdown suppressed TNBC cell proliferation, invasion and migration and promoted TNBC cell apoptosis. Furthermore, the expression of Notch1 and Notch4 was inhibited by GRWD1 knockdown. The expression of downstream genes of the Notch signaling pathway Hes1, Hes5, Hey1, Hey2, p21, c-Myc, cyclin D1, human epidermal growth factor 2 receptor and NF-κB were all suppressed after siRNA-GRWD1 transfection. However, Notch1 overexpression reversed the effect of GRWD1 knockdown on biological behaviors of TNBC cells. In conclusion, GRWD1 knockdown could suppress the proliferation, invasion and migration and promoted apoptosis of TNBC cells through inhibiting the Notch signaling pathway.

Gypensapogenin H suppresses tumor growth and cell migration in triple-negative breast cancer by regulating PI3K/AKT/NF-κB/MMP-9 signaling pathway

Due to its aggressiveness and high metastasis rates, triple-negative breast cancer (TNBC) is a ubiquitous and deadly disease for the majority of women globally. Gypensapogenin H (GH) is a novel dammarane-type triterpene isolated from hydrolyzate of total saponins from Gynostemma pentaphyllum. Our previous work demonstrated that GH promoted apoptosis in TNBC. In the present study, xenograft TNBC models (xenotransplantation of MDA-MB-231 cells in nude mice) were used to evaluate the efficacy of GH in vivo. We preliminarily predicted the mechanism of GH inhibiting breast cancer tumors at the gene level through transcriptome screening. Through western blot analysis of tumor tissue, we found that GH could inhibit tumor proliferation and migration by regulating the PI3K/AKT/NF-κB/MMP-9 signaling pathway in vivo. We also analyzedthe results at the cell level in vitro, which were consistent with those in vivo. In summary, GH inhibited TNBC growth in vivo and suppressed TNBC cell migration in vitro. Our findings could help understand the mechanism of action of GH and suggest that GH would be a promising agent for TNBC therapy.

Circ_0041732 regulates tumor properties of triple-negative breast cancer cells by the miR-149-5p/FGF5 pathway.

Triple-negative breast cancer (TNBC) is a subtype of breast cancers with a high recurrence and mortality. The important factors promoting the TNBC process have not been fully identified. In this research, the role of a TNBC-related circular RNA (circRNA), circ_0041732, was revealed in TNBC cell tumor properties. The expression levels of circ_0041732, microRNA-149-5p (miR-149-5p) and fibroblast growth factor 5 (FGF5) were detected by quantitative real-time polymerase chain reaction. The protein expression was determined by Western blot analysis or immunohistochemistry assay. Cell proliferation was detected by cell counting kit-8 and cell colony formation assays. Cell apoptosis was analyzed by flow cytometry and caspase-3 activity assays. Cell migration and invasion were evaluated by wound-healing and transwell invasion assays. Cell angiogenic capacity was investigated by a tube formation assay. The targeting relationship between miR-149-5p and circ_0041732 or FGF5 was identified by dual-luciferase reporter and RNA immunoprecipitation assays. The impacts of circ_0041732 knockdown on tumor formation were determined by an in vivo assay. Circ_0041732 and FGF5 expression were significantly upregulated, whereas miR-149-5p was downregulated in TNBC tissues and cells compared with normal breast tissues and cells, respectively. Circ_0041732 silencing inhibited TNBC cell proliferation, migration, invasion, and tube formation, but induced apoptosis. Additionally, circ_0041732 regulated TNBC cell tumor properties by binding to miR-149-5p. MiR-149-5p also modulated TNBC cell tumor properties by targeting FGF5. Furthermore, circ_0041732 knockdown hindered tumor formation in vivo. Circ_0041732 silencing suppressed TNBC cell tumor properties by decreasing FGF5 expression through miR-149-5p. This finding demonstrated that circ_0041732 had the potential as a therapeutic target for TNBC.

PGM5P3-AS1 regulates MAP1LC3C to promote cell ferroptosis and thus inhibiting the malignant progression of triple-negative breast cancer.

Triple-negative breast cancer (TNBC) represents an aggressive subtype of breast cancer characteristic of high recurrence rate and poor prognosis. According to previous studies and bioinformatics prediction, PGM5P3-AS1 has been found to be significantly down-regulated in TNBC cells. In addition, cell ferroptosis has become a hotspot in breast cancer research and TNBC has been reported to be more sensitive to ferroptosis than receptor positive breast cancer. Hence, we aim at exploring the molecular mechanism of PGM5P3-AS1 in TNBC cells and further explore whether PGM5P3-AS1 can inhibit TNBC progression via promoting cell ferroptosis. The expression of genes in TNBC cells was verified by RT-qPCR assay. Functional assays were taken to evaluate the impact PGM5P3-AS1 may exert on TNBC progression. The regulatory pattern of PGM5P3-AS1 on cell ferroptosis in TNBC was validated through mechanism assays. PGM5P3-AS1 was proved to be down-regulated in TNBC cells and suppressed TNBC cell proliferation as well as migration. PGM5P3-AS1 promoted cell ferroptosis in TNBC by recruiting RNA-binding protein (RBP) NOP58 to stabilize MAP1LC3C mRNA, and thus inhibiting TNBC progression. PGM5P3-AS1 regulated MAP1LC3C to promote cell ferroptosis and thus inhibiting the malignant progression of TNBC.

Maackiain Modulates miR-374a/GADD45A Axis to Inhibit Triple-Negative Breast Cancer Initiation and Progression

Breast cancer ranks as the leading cause of death in lethal malignancies among women worldwide, with a sharp increase of incidence since 2008. Triple negative breast cancer (TNBC) gives rise to the largest proportion in breast cancer-related deaths because of its aggressive growth and rapid metastasis. Hence, searching for promising targets and innovative approaches is indispensable for the TNBC treatment. Maackiain (MA), a natural compound with multiple biological activities, could be isolated from different Chinese herbs, such as Spatholobus suberectus and Sophora flavescens. It was the first time to report the anti-cancer effect of MA in TNBC. MA could suppress TNBC cell proliferation, foci formation, migration, and invasion. MA also exerted a significant inhibitory effect on tumor growth of TNBC. Furthermore, MA could induce apoptosis with an increase of GADD45α and a decrease of miR-374a. In contrast, overexpressing miR-374a would result in at least partly affecting the proapoptotic effect of MA and suppressing GADD45α stimulated by MA. These results reveal the anti-TNBC effect of MA in vitro and in vivo, providing evidence for its potential as a drug candidate utilized in TNBC therapy.

RNA N6 -methyladenosine reader YTHDC1 is essential for TGF-beta-mediated metastasis of triple negative breast cancer.

RNA N6-methyladenosine (m6A) modification and its regulators fine tune gene expression and contribute to tumorigenesis. This study aims to uncover the essential role and the underlying molecular mechanism(s) of the m6A reader YTHDC1 in promoting triple negative breast cancer (TNBC) metastasis.Methods: In vitro and in vivo models were employed to determine the pathological function of YTHDC1 in TNBC metastasis. To identify bona fide YTHDC1 target RNAs, we conducted RNA-seq, m6A-seq, and RIP-seq, followed by integrative data analysis and validation assays.Results: By analyzing The Cancer Genome Atlas (TCGA) dataset, we found that elevated expression of YTHDC1 is positively correlated with poor prognosis in breast cancer patients. Using a mammary fat pad mouse model of TNBC, YTHDC1 significantly promoted lung metastasis of TNBC cells. Through multiple transcriptome-wide sequencing and integrative data analysis, we revealed dysregulation of metastasis-related pathways following YTHDC1 depletion and identified SMAD3 as a bona fide YTHDC1 target RNA. Depletion of YTHDC1 caused nuclear retention of SMAD3 mRNA, leading to lower SMAD3 protein levels. Loss of YTHDC1 led to impaired TGF-β-induced gene expression, leading to inhibition of epithelial-mesenchymal transition (EMT) and suppressed TNBC cell migration and invasion. SMAD3 overexpression was able to restore the response to TGF-β in YTHDC1 depleted TNBC cells. Furthermore, we demonstrated that the oncogenic role of YTHDC1 is mediated through its recognition of m6A as m6A-binding defective mutants of YTHDC1 were unable to rescue the impaired cell migration and invasion of YTHDC1 knockout TNBC cells.Conclusions: We show that YTHDC1 plays a critical oncogenic role in TNBC metastasis through promoting the nuclear export and expression of SMAD3 to augment the TGF-β signaling cascade. Overall, our study demonstrates that YTHDC1 is vital for TNBC progression by enhancing TNBC cell survival and TGF-β-mediated EMT via SMAD3 to enable the formation of distant metastasis and highlights the therapeutic potential of targeting the YTHDC1/m6A/SMAD3 axis for TNBC treatment.