Apoptosis In Breast Cancer Cells Research Articles

Anticancer Potential of Postbiotic Derived from Lactobacillus brevis and Lactobacillus casei: In vitro Analysis of Breast Cancer Cell Line.

Breast cancer has emerged as the most widespread and dangerous type of malignancy among women worldwide. Postbiotics have recently emerged as a promising novel adjunct in breast cancer therapy, due to their immunomodulatory effects and the potential to mitigate the adverse effects of conventional treatments. This study aims to investigate the therapeutic effects of postbiotics derived from Lactobacillus brevis (CSF2) and Lactobacillus casei (CFS5), specifically examining their ability to inhibit cell proliferation and induce apoptosis in MCF-7 breast cancer cells. In the current study, the anticancer activity of the cell-free supernatant of L. brevis and L. casei was investigated against MCF-7 cells using MTT assay, flow cytometry, and qRT-PCR technique. Both bacteria showed a high potential for the induction of cell death in MCF-7 cells. However, CFS2 cytotoxicity was significantly higher than CFS5. Flow cytometry results showed significant induction of early apoptosis in cells treated with both CFS2 and CFS5 within 48h. The induction was notably higher in cells treated with CFS2 compared to CFS5. Overall, CFS2 therapy resulted in a greater increase in BAX and CASP9 gene expression, as well as an elevated BAX/BCL2 ratio within 48h. These findings indicate that the CFS2 treatment showed a higher level of apoptotic activity than the CFS5 treatment. High biocompatibility was demonstrated following treatment with CFS2 and CFS5. These CFSs may serve as adjunctive medications for suppressing the proliferation of cancer cells. The results of the current study highlight the potential of postbiotics in cancer treatment and suggest that supernatants may serve as effective agents for suppressing cancer cell growth and viability.

MLLT6/ATF2 axis restrains breast cancer progression by driving DDIT3/4 expression.

Epigenetic deregulation is strongly associated with tumour progression. The identification of natural tumour suppressors to overcome cancer metastasis is urgent for cancer therapy. We investigate whether myeloid/lymphoid or mixed-lineage leukaemia translocated (MLLT) family members contribute to breast cancer progression and found that high MLLT6 expression predicted a better prognosis and that gradually decreased MLLT6 expression was accompanied by breast cancer malignancy. MLLT6 was downregulated by hypoxia-induced enrichment of DNMT1 at the MLLT6 promoter. The results of in vitro functional experiments indicated that MLLT6 depletion promoted colony formation and cell migration, probably by hampering apoptosis. RNA profiling revealed that the apoptotic pathway was downregulated following stable knockdown of MLLT6. DNA damage-inducible transcript 3/4 (DDIT3/4) were among the top 10 downregulated genes and may have expression patterns similar to that of MLLT6. Restoring DDIT3/4 expression in cells with MLLT6 depletion blocked colony formation and cell migration and attenuated the successful colonization of breast cancer cells in vivo. We also determined that the transcription factor activating transcription factor 2 (ATF2) is a binding partner of MLLT6 and participates in the MLLT6/ATF2 axis, which was reinforced by inhibition of AKT signalling, in turn inducing DDIT3/4 expression by establishing an active chromatin structure at the DDIT3/4 gene promoters. Because MLLT6 promotes breast cancer cell apoptosis by inducing DDIT3/4 expression during metastasis, it could be a novel tumour suppressor. Implications: Control of MLLT6 expression via inhibition of PI3K/AKT kinase activity is a potential therapeutic approach for the management of metastatic breast cancer.

CRISPR-Cas13d screens identify KILR, a breast cancer risk-associated lncRNA that regulates DNA replication and repair

BackgroundLong noncoding RNAs (lncRNAs) have surpassed the number of protein-coding genes, yet the majority have no known function. We previously discovered 844 lncRNAs that were genetically linked to breast cancer through genome-wide association studies (GWAS). Here, we show that a subset of these lncRNAs alter breast cancer risk by modulating cell proliferation, and provide evidence that a reduced expression on one lncRNA increases breast cancer risk through aberrant DNA replication and repair.MethodsWe performed pooled CRISPR-Cas13d-based knockdown screens in breast cells to identify which of the 844 breast cancer-associated lncRNAs alter cell proliferation. We selected one of the lncRNAs that increased cell proliferation, KILR, for follow-up functional studies. KILR pull-down followed by mass spectrometry was used to identify binding proteins. Knockdown and overexpression studies were performed to assess the mechanism by which KILR regulates proliferation.ResultsWe show that KILR functions as a tumor suppressor, safeguarding breast cells against uncontrolled proliferation. The half-life of KILR is significantly reduced by the risk haplotype, revealing an alternative mechanism by which variants alter cancer risk. Mechanistically, KILR sequesters RPA1, a subunit of the RPA complex required for DNA replication and repair. Reduced KILR expression promotes breast cancer cell proliferation by increasing the available pool of RPA1 and speed of DNA replication. Conversely, KILR overexpression promotes apoptosis in breast cancer cells, but not normal breast cells.ConclusionsOur results confirm lncRNAs as mediators of breast cancer risk, emphasize the need to annotate noncoding transcripts in relevant cell types when investigating GWAS variants and provide a scalable platform for mapping phenotypes associated with lncRNAs.

Surface-enhanced Raman scattering-fluorescence dual-mode probes for target imaging of tumors, organoids and cancerous cells

A dual-mode nanoprobe (DMNP) overcomes the limitations of single probes and offers superior performance by combining their advantages. Herein, we report a DMNP design that features a gold nanosphere core and two layers of silica wrapping. The gold core surface is coated with 4-mercaptobenzoic acid(4-MBA), which serves as a Raman reporter. After the initial silica encapsulation, fluorophores in the form of conjugated polymer nanoparticles (CPNs) are applied to the surface. Finally, the encapsulation process is completed by adding the second layer of silica. The probe has dual-mode capabilities, including surface-enhanced Raman scattering (SERS) with a Raman enhancement factor of 134.1, and metal-enhanced fluorescence (MEF) with a fluorescence enhancement factor of 1.42. When modified with glycoprotein-3 (GPC3) antibody, the DMNPs probe shows excellent performance in fluorescence and SERS imaging for HepG2 liver cancer cells. When DMNP is conjugated with the breast tumor-specific antibody CA153, the DMNPs-AbCA153 probe enables imaging and induces apoptosis of breast cancer cells. These probes can serve as a platform for visualizing the distribution of cancer cell surface markers by modifying antibodies to recognize the cell surface. They have also shown significant potential as probes for oncology diagnosis and treatment monitoring.

Mechanistic exploration and experimental validation of the Xiaochaihu decoction for the treatment of breast cancer by network pharmacology.

Xiaochaihu (XCH) decoction is a traditional Chinese prescription that has been recorded in the pharmacopeia of the People's Republic of China. In China, the XCH decoction is used clinically to treat a variety of tumors, including breast cancer. However, its potential mechanism of action is still undefined. The chemical compounds in the XCH decoction were identified via Q Exactive Orbitrap LC-MS/MS. Then, we screened the active ingredients and targets in the XCH decoction from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP). Next, Cytoscape and Metascape were used to construct an active ingredient-target-disease network, which included a protein-protein interaction (PPI) network, GO enrichment analysis, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. Finally, we used molecular docking and in vitro experiments to verify the results of network pharmacology analysis. More than 70 major compounds were identified by Q Exactive Orbitrap LC-MS/MS analysis from the XCH decoction. A total of 162 active ingredients and 153 targets related to the XCH decoction and breast cancer were identified, and a compound-target-disease network was constructed. GO and KEGG analyses revealed that the XCH decoction regulated the drug response, apoptosis process, cancer pathway, and PI3K/Akt signaling pathway. Molecular docking and experimental validation indicated that the XCH decoction suppressed proliferation and induced apoptosis in breast cancer cells by regulating the expression of apoptosis-related proteins and inhibiting the PI3K/Akt pathway. This study suggested that the XCH decoction can be used to treat breast cancer by inhibiting cell proliferation, inducing apoptosis and downregulating the PI3K/Akt signaling pathway.

Molecular interactions between metformin and D-limonene inhibit proliferation and promote apoptosis in breast and liver cancer cells

BackgroundCancer is a fatal disease that severely affects humans. Designing new anticancer strategies and understanding the mechanism of action of anticancer agents is imperative.Hypothesis/PurposeIn this study, we evaluated the utility of metformin and D-limonene, alone or in combination, as potential anticancer therapeutics using the human liver and breast cancer cell lines HepG2 and MCF-7.Study designAn integrated systems pharmacology approach is presented for illustrating the molecular interactions between metformin and D-limonene.MethodsWe applied a systems-based analysis to introduce a drug–target–pathway network that clarifies different mechanisms of treatment. The combination treatment of metformin and D-limonene induced apoptosis in both cell lines compared with single drug treatments, as indicated by flow cytometric and gene expression analysis.ResultsThe mRNA expression of Bax and P53 genes were significantly upregulated while Bcl-2, iNOS, and Cox-2 were significantly downregulated in all treatment groups compared with normal cells. The percentages of late apoptotic HepG2 and MCF-7 cells were higher in all treatment groups, particularly in the combination treatment group. Calculations for the combination index (CI) revealed a synergistic effect between both drugs for HepG2 cells (CI = 0.14) and MCF-7 cells (CI = 0.22).ConclusionOur data show that metformin, D-limonene, and their combinations exerted significant antitumor effects on the cancer cell lines by inducing apoptosis and modulating the expression of apoptotic genes.

Marine seaweed endophytic fungi-derived active metabolites promote reactive oxygen species-induced cell cycle arrest and apoptosis in human breast cancer cells.

Endophytic fungi have an abundant sources rich source of rich bioactive molecules with pivotal pharmacological properties. Several studies have found that endophytic fungi-derived bioactive secondary metabolites have antiproliferative, anti-oxidant, and anti-inflammatory properties, but the molecular mechanism by which they induce cell cycle arrest and apoptosis pathways is unknown. This study aimed to determine the molecular mechanism underlying the anticancer property of the endophytic fungi derived active secondary metabolites on human breast cancer cells. In this study, we identified four endophytic fungi from marine seaweeds and partially screened its phytochemical properties by Chromatography-Mass Spectrometry (GC-MS) analysis. Moreover, the molecular mechanism underlying the anticancer property of these active secondary metabolites (FA, FB, FC and FE) on human breast cancer cells were examined on MCF-7 cells by TT assay, Apoptotic assay by Acridine orang/Ethidium Bromide (Dual Staining), DNA Fragmentation by DAPI Staining, reactive oxygen species (ROS) determination by DCFH-DA assay, Cell cycle analysis was conducted Flow cytometry and the apoptotic signalling pathway was evaluated by westernblot analysis. Doxorubicin was used as a positive control drug for this experiment. The GC-MS analysis of ethyl acetate extract of endophytic fungi from the marine macro-algae revealed the different functional groups and bioactive secondary metabolites. From the library, we observed the FC (76%), FB (75%), FA (73%) and FE (71%) have high level of antioxidant activity which was assessed by DPPH scavenging assay. Further, we evaluated the cytotoxic potentials of these secondary metabolites on human breast cancer MCF-7 cells for 24h and the IC50 value were calculated (FA:28.62 ± 0.3µg/ml, FB:49.81 ± 2.5µg/ml, FC:139.42 ± µg/ml and FE:22.47 ± 0.5µg/ul) along with positive control Doxorubicin 15.64 ± 0.8µg/ml respectively by MTT assay. The molecular mechanism by which the four active compound induced apoptosis via reactive oxygen species (ROS) and cell cycle arrest in MCF-7 cells was determined H2DCFDA staining, DAPI staining, Acridine orange and ethidium bromide (AO/EtBr) dual staining, flowcytometry analysis with PI staining and apoptotic key regulatory proteins expression levels measured by westernblot analysis. Our findings, revealed the anticancer potential of endophytic fungi from marine seaweed as a valuable source of bioactive compounds with anticancer properties and underscore the significance of exploring marine-derived endophytic fungi as a promising avenue for the development of novel anticancer agents. Further investigations are necessary to isolate and characterize specific bioactive compounds responsible for these effects and to validate their therapeutic potential in preclinical and clinical settings.

Abstract PO3-27-02: A novel PTK6 PROTAC degrader induces apoptosis of drug resistant breast cancer cells

Abstract Background. PTK6/Brk, a non-receptor tyrosine kinase, is an oncogenic driver for several tumor types, including breast, prostate, liver, and pancreatic cancers. PTK6 is expressed in all breast cancer subtypes and higher levels of expression are associated with worse patient outcomes. The optimal approach to targeting PTK6 for therapeutic benefit has not been established. Although several inhibitors of PTK6 kinase activity have been developed, none have thus far been successfully translated to clinical trials. This may be in part due to kinase activity-independent functions of PTK6, for which there is growing evidence, that are not effectively targeted by traditional small molecule kinase inhibitors. Downregulation of PTK6 using shRNA inhibits growth, invasion/migration, survival and metastasis of breast cancer cells, including those that are resistant to standard therapies. PTK6 kinase inhibitor treatment phenocopies effects of PTK6 downregulation with respect to some, but not all PTK6-driven oncogenic activities. Moreover, growth of PTK6 CRISPR-targeted cells can be rescued either with wild type or catalytically inactive PTK6, supporting kinase activity-independent activities of PTK6 in regulating growth and viability of breast cancer cells. Therefore, PROTAC degraders may be a better way to chemically target and downregulate PTK6 expression in cancer, thereby phenocopying the effects of PTK6 shRNA. Methods. We developed a first-in-class PTK6 degrader using PROTAC (proteolysis-targeting chimeric) technology to target PTK6 expression in breast cancer cells of diverse subtypes. Degrader controls were also synthesized to assess specificity of MS105’s effects. Results. PTK6 degrader, but not degrader controls, potently downregulates PTK6 expression in multiple breast cancer cell types. Proteomic and kinome profiling confirm the specificity of our PTK6 degrader. PTK6 PROTAC degrader inhibits viability and induces apoptosis of PTK6-expressing breast cancer cells, whereas treatment with parent kinase inhibitor or degrader controls that do not engage PTK6 or E3 ligase have minimal effect. Treatment with PTK6 degrader, but not kinase inhibitor or degrader controls, induces expression of pro-apoptotic Bim, phenocopying the effects observed with PTK6 shRNA. In contrast, PTK6 degrader and kinase inhibitor are comparably effective in inhibiting cell migration, supporting the differential kinase dependency of PTK6-dependent oncogenic functions. Therefore, our PTK6 PROTAC degrader effectively inhibits both kinase-dependent and independent oncogenic activities of PTK6. Our studies support the development of PTK6 PROTAC degraders as a preferred approach to clinically targeting PTK6 in cancer. Citation Format: Hanna Irie, Criseyda Martinez, Yan Xiong, Alison Bartkowski, Jessica Byerly, Jian Jin. A novel PTK6 PROTAC degrader induces apoptosis of drug resistant breast cancer cells [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 PO3-27-02.

Abstract PO5-23-11: Elevated linoleic acid levels in red blood cells membrane predicts response to neoadjuvant chemotherapy in breast cancer patients

Abstract Introduction: Fatty acids (FA) consumed in the diet have been implicated in increasing the risk of breast cancer (BC). Humans have a wide variety of FAs, including saturated (SFA), monounsaturated (MUFA), and polyunsaturated (PUFA) FAs. PUFAs can be further classified into omega-3 (n-3) and omega-6 (n-6) PUFAs. While n-3 has been associated with reducing inflammation and promoting apoptosis in BC cells, n-6 has been linked to increased inflammation, oxidative stress, and the promotion of BC cell proliferation and treatment resistance. In healthy women, the n-3 content in mammary tissue correlates with n-3 content in red blood cells (RBCs). The FA profile of the RBC membrane provides an unbiased option to assess lipid intake and composition, reflecting long-term FA intake and providing an objective and sensitive measure of lipid intake and body fat deposits. Notably, BC patients have been found to display a high n-6:n-3 ratio in RBCs, which is associated with inflammatory and oxidative stress biomarkers. The neoadjuvant chemotherapy treatment (NACT) model presents a unique opportunity to study resistance mechanisms in non-metastatic BC patients. In this approach, the absence of tumor (pathologic complete response: pCR, or Residual Cancer Burden (RCB) = 0) after NACT indicates treatment sensitivity and better overall survival (OS). Objective: The objective of this study is to determine whether differences in FA profiles determined in the RBC membrane and plasma are associated with the response to NACT in non-metastatic BC patients. Methods: A prospective study was conducted, including non-metastatic BC patients scheduled to receive NACT. Blood samples were collected before the first cycle of NACT to measure plasma and RBC FA derivatives, SFAs, MUFAs, and PUFAs. The pathological complete response (pCR) and residual cancer burden (RCB) were assessed. Results: A total of twenty-eight BC patients were included in the study. When comparing pCR vs. no-pCR using total SFA, MUFA, and PUFA plasma samples, no significant differences were observed. However, in RBC, an increase in PUFA was associated with a higher rate of no-pCR (**p< 0.01). Furthermore, comparing omega-6 and omega-3 (both PUFA) levels, an increase in omega-6 was associated with no-pCR (**p< 0.01), while no significant differences were observed with omega-3. No differences were obtained in plasma samples for either omega-6 and omega-3. To determine which specific omega-6 PUFAs were responsible for the association with no-pCR, elevated levels of linoleic acid (LA) in RBC were found to be associated with no-pCR (**p< 0.01). Other omega-6 PUFAs, including gamma-linolenic acid (GLA), eicosadienoic acid (EDA), dihomo-gamma linolenic acid (DGLA), and arachidonic acid (ARA), showed no significant associations. Multivariate and regression analysis confirmed LA (C18:2 n-6, LA) as the most relevant FA when assessing RCB (0-1 vs. 2-3) in RBC membranes, but not in plasma. The receiver operating characteristic (ROC) curve analyzing LA levels in RBC showed an area under the curve (AUC) of 0.855 for RCB (sensitivity: 76.9%, specificity: 92.3%). Conclusions: To the best of our knowledge, this is the first study demonstrating a clear connection between RBC membrane FA composition, specifically LA concentration, and the pathological response in BC patients after NACT. The measurements taken in plasma did not show significant associations, suggesting that RBC membrane FA profile of could serve as a relevant prognostic biomarker for studying resistance mechanisms to NACT in early BC patients Table. Measured FAs in RBC membrane and plasma by pathologic complete response *Indicates p<0.05; abbreviations: pCR: pathological complete response; SD: standard deviation; yr: years; BMI: body mass index; HR: hormone receptor; HER2: human epidermal growth factor receptor type-2; NLR: neutrophil to lymphocyte ratio; HG: histological grade; RCB: residual cancer burden; NA: not applicable Citation Format: Benjamin Walbaum, César Sánchez, Francisco Acevedo. Elevated linoleic acid levels in red blood cells membrane predicts response to neoadjuvant chemotherapy in breast cancer patients [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 PO5-23-11.

Abstract PO4-28-04: Curcumin inhibits breast cancer cell proliferation by regulating ciRS-7/miR-7-5p/CKS2 axis

Abstract Background Breast cancer (BC) is the most common cancer in women worldwide. Curcumin is a polyphenolic turmeric-derived compound that has anti-proliferative and anti-tumor properties in different cancer types by acting on multiple molecules. Circular RNAs (circRNAs) are non-coding, single-stranded, covalently closed RNA molecules that act as regulators of the microRNA (miRNA) activity. Recent studies show that circRNAs are potential contributors to the onset and progression of many cancer types. CircRNA ciRS-7 acts as an oncogene and accelerates tumor progression by competitively suppressing miR-7-5p in BC. However, whether curcumin can regulate ciRS-7 to inhibit BC progression is still unclear. Method Breast Cancer cell lines (MCF-7 and T47D) and normal epithelial cell line (MCF-10A) were cultivated and treated with Curcumin (5μM, 10 μM, 20 μM) or DMSO as a control. Also, the cell lines were transfected with ciRS-7 siRNA, miR-7-5p mimic, and their non-targeted controls. The cell viability was detected by Cell Viability Detection Kit-8 (CVDK-8) using an ELISA plate reader. To detect cell migration, scratch assay was performed after 24hs of transfection and a phase contrast microscope was used to acquire images. The effect on apoptosis was detected by Annexin V-FITC Apoptosis Detection Kit using flow cytometry. Potential target genes of miR-7-5p were identified by searching for the overlapping genes in miRNet and miRTarBase v8 with the overexpressed genes in BC patient tissue samples in the TCGA database. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression of ciRS-7, miR-7-5p, and the selected genes. qRT-PCR experiments were performed in duplicate and the 2−ΔΔCt method was used for relative quantitation analysis. Results Curcumin treatment, depending on the increased doses, decreased cell proliferation through inducing apoptosis in MCF-7 and T47D cancer cells. Curcumin's anti-proliferative effect was shown to be quite restricted on normal MCF-10A cells, as compared to MCF-7 and T47D cancer cells. In MCF-7 and T47D cells treated with curcumin, cell migration was dramatically inhibited. Curcumin has very little influence on the migration of MCF-10A cells. In both cancer cell group that was transfected with ciRS-7 siRNA and miR-7-5p mimic, it was observed that apoptosis was increased, proliferation was suppressed, and migration was decreased compared to its control groups. Moreover, the expression of ciRS-7 was found to be significantly decreased in the curcumin-treated group, while miR-7-5p was shown to be significantly higher. CKS2 gene one of the possible target genes of miR-7-5p that was identified by using in silico approaches, was also downregulated in curcumin-treated BC cell lines compared to its control group. ciRS-7 and CKS2 expression levels were found to be downregulated, whereas miR-7-5p expression level was found to be elevated in MCF-7 and T47D cells that were transfected with ciRS-7 siRNA. Additionally, CKS2 gene expression was found to be downregulated in miR-7-5p mimic transfected cells. Conclusion Curcumin is derived from the turmeric plant (Curcuma longa) and has been used since 3000 B.C. as a food additive. Nowadays, curcumin is one of the most essential anti-cancer substances that was examined. However, investigations on the influence of curcumin on the circRNA-miRNA-mRNA axis are scarce. Curcumin has been demonstrated to inhibit proliferation and induce apoptosis of BC cells via the ciRS-7/miR-7-5p/CKS2 axis in the present study. Keywords Breast Cancer, Curcumin, ciRS-7, miR-7-5p, CKS2 Citation Format: Asmaa Abuaisha, Murat Kaya, Ilknur Suer, Selman Emiroglu, Fahrunnisa Abanoz, Mustafa Tukenmez, Neslihan Cabıoğlu, Mahmut Muslumanoglu, Kivanc Cefle, Sukru Palanduz, Sukru Ozturk. Curcumin inhibits breast cancer cell proliferation by regulating ciRS-7/miR-7-5p/CKS2 axis [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 PO4-28-04.

Synergistic Effect of Layered Double Hydroxides Nanodosage Form to Induce Apoptosis and Ferroptosis in Breast Cancer.

Breast cancer is the most common cancer in women and one of the leading causes of cancer death worldwide. Ferroptosis, a promising mechanism of killing cancer cells, has become a research hotspot in cancer therapy. Simvastatin (SIM), as a potential new anti-breast cancer drug, has been shown to cause ferroptosis of cancer cells and inhibit breast cancer metastasis and recurrence. The purpose of this study is to develop a novel strategy boosting ferroptotic cascade for synergistic cancer therapy. In this paper, iron base form of layered double hydroxide supported simvastatin (LDHs-SIM) was synthesized by hydrothermal co-precipitation method. The characterization of LDHs-SIM were assessed by various analytical techniques, including ultraviolet-visible (UV-vis) spectroscopy, X-ray diffraction (XRD), Fourier transform infrared (FTIR) spectroscopy, and transmission electron microscopy (TEM). Biological activity, ferroptosis mechanism and biocompatibility were analyzed through in vivo and in vitro analysis, so as to evaluate its therapeutic effect on breast cancer. The constructed LDHs-SIM nanosystem can not only release SIM through mevalonate (MVA) pathway, inhibit the expression of glutathione peroxidase 4 (GPX4), inhibit the expression of SLC7A11 and reduce the synthesis efficiency of GSH, but also promote the accumulation of Fe2+ in cells through the release of Fe3+, and increase the intracellular ROS content. In addition, LDHs-SIM nanosystem can induce apoptosis of breast cancer cells to a certain extent, and achieve the synergistic effect of apoptosis and ferroptosis. In the present study, we demonstrated that nanoparticles of layered double hydroxides (LDHs) loaded with simvastatin were more effective than a free drug at inhibiting breast cancer cell growth, In addition, superior anticancer therapeutic effects were achieved with little systemic toxicity, indicating that LDHs-SIM could serve as a safe and high-performance platform for ferroptosis-apoptosis combined anticancer therapy.

The Protective Mechanism of Tao Hong Si Wu Decoction Against Breast Cancer Through Regulation of EGFR/ERK1/2 Signaling

Ethnopharmacological relevanceTao Hong Si Wu Decoction (THSWD), a traditional Chinese herbal medicine, is widely utilized in clinical settings, either alone or in combination with other medications, for the treatment of breast cancer.Aim of the study: The specific targeting molecule(s) of THSWD and its associated molecular mechanisms remain unclear. This research aims to elucidate the underlying molecular mechanisms of THSWD in the treatment of breast cancer. Materials and MethodsThe pharmacological properties of THSWD were investigated in breast cancer cells and tumor tissues using a range of methods including Acridine Orange/Ethidium Bromide (AO/EB) staining, Transwell assay, flow cytometry, immunofluorescence assay, and breast cancer mice models. ResultsOur findings demonstrate that THSWD induces necrosis and/or apoptosis in breast cancer cells, while significantly inhibiting cell migration. Target proteins of THSWD in anticancer activity include EGFR, RAS, and others. THSWD treatment for breast cancer is associated with the EGFR/ERK1/2 signaling pathway. ConclusionOur findings offer initial insights into the primary mechanism of action of THSWD in breast cancer treatment, indicating its potential as a complementary therapy deserving further investigation.

Marine-derived EGFR inhibitors: novel compounds targeting breast cancer growth and drug resistance.

Breast cancer (BC) continues to be a major health challenge globally, ranking as the fifth leading cause of cancer mortality among women, despite advancements in cancer detection and treatment. In this study, we identified four novel compounds from marine organisms that effectively target and inhibit the Epidermal Growth Factor Receptor (EGFR), crucial for BC cell growth and proliferation. These compounds not only induced early apoptosis through Caspase-3 activation but also showed significant inhibitory effects on EGFR mutations associated with drug resistance (T790M, L858R, and L858R/T790M), demonstrating high EGFR kinase selectivity. Cell Thermal Shift Assay (CETSA) experiments indicated that Tandyukisin stabilizes EGFR in a concentration-dependent manner. Furthermore, binding competition assays using surface plasmon resonance technology revealed that Tandyukisin and Trichoharzin bound to distinct sites on EGFR and that their combined use enhanced apoptosis in BC cells. This discovery may pave the way for developing new marine-derived EGFR inhibitors, offering a promising avenue for innovative cancer treatment strategies and addressing EGFR-mediated drug resistance.

LINC02535 + miR-30a-5p combination enhances proliferation and inhibits apoptosis in metastatic breast Cancer cells

Current clinical therapies for metastatic breast cancer (MBC) have limited therapeutic efficacy and induce significant systemic side effects, leading to poor patient compliance. To address this challenge, this investigation focuses on the design of LINC02535 + miR-30a-5p for treating breast cancer. In vitro cytotoxicity studies confirmed that LINC02535 + miR-30a-5p was more effective in 4 T1 cells, with reduced toxicity in NIH3T3 cells. Further verification of cellular morphology was achieved through various biochemical staining methods. Additionally, the antimetastatic attributes of LINC02535 + miR-30a-5p have been evaluated using both migration scratch and Transwell migration assays, which have collectively revealed excellent antimetastatic potential. The DNA fragmentation of the 4 T1 cells was assessed using a comet assay. LINC02535 + miR-30a-5p improved ROS levels and caused mitochondrial membrane potential alterations and DNA damage, which resulted in apoptosis. Therefore, we propose that LINC02535 + miR-30a-5p could be an alternative therapeutic strategy for breast cancer therapy.

Inhibitory effects of medium-chain fatty acids on the proliferation of human breast cancer cells via suppression of Akt/mTOR pathway and modulating the Bcl-2 family protein.

Medium-chain fatty acids (MCFAs) have 6-12 carbon atoms and are instantly absorbed into the bloodstream before traveling to the portal vein and the liver, where they are immediately used for energy and may have antitumor effects. Its role in breast cancer is poorly understood. To investigate the apoptosis-inducing effect of MCFAs in breast cancer cells, cell viability assay, colony formation assay, cell migration assay, cell invasion assay, nuclear morphology, cell cycle assay, intracellular reactive oxygen species (ROS), matrix metalloproteinase (MMP), apoptosis, RT-qPCR analysis, and Western blot analysis were performed. In the present study, MCFA treatments reduced proliferative capability, increased ROS level, increased the depletion of MMP, induced G0/G1 and S phase cell cycle arrest, and late apoptosis of breast cancer cells in an effective concentration. Besides, MCFA treatment contributed to the upregulation of proapoptotic protein (BAK) and caspase-3, and the downregulation of antiapoptotic protein (Bcl-2). Mechanistically, phosphorylation levels of EGFR, Akt, and mTOR were significantly reduced in breast cancer cells treated with MCFAs. However, no significant changes in apoptosis and signaling-related proteins were observed in lauric acid-treated ER-positive cancer cells. Our findings suggested that MCFAs suppressed breast cancer cell proliferation by modulating the PI3K/Akt/mTOR signaling pathway. MCFAs may be a promising therapeutic drug for treating breast cancer.

Moringin, an isothiocyanate modulates multiple cellular signalling molecules in breast cancer cells

Prohibitin (PHB) is a pleiotropic molecule with a variety of known functions and subcellular locations. PHB's function in breast cancer is poorly understood. Herein, we report that PHB is expressed in cancer types of diverse origin including breast cancer. The cancer patients with changes in PHB were reported to have significantly reduced ‘overall survival’ in comparison to the cases without alterations in PHB. The expression of PHB was increased by H2O2 and also by Moringin (MG), which is an isothiocyanate derived from the seeds of Moringa oleifera. MG interacted with PHB, DRP1, and SLP2 and inhibited the growth of MCF-7 and MDAMB-231 cells. The isothiocyanate triggered apoptosis in breast cancer cells as revealed by AO/PI assay, phosphatidylserine externalization, cell cycle analysis and DAPI staining. MG induced proapoptotic proteins expression such as cytochrome c, p53, and cleaved caspase-7. Further, cell survival proteins such as survivin, Bcl-2, and Bcl-xL were suppressed. A depolarization of membrane potential suggested that the apoptosis was triggered through mitochondria. The isothiocyanate suppressed the cancer cell migration and interacted with NF-κB subunits. MG suppressed p65 nuclear translocation induced by TNF-α. The reactive oxygen species generation was also induced by the isothiocyanate in breast cancer cells. MG also modulated the expression of lncRNAs. Collectively, the functions of PHB in breast cancer growth is evident from this study. The activities of MG against breast cancer might result from its ability to modulate multiple cancer-related targets.

Daurisoline inhibits proliferation, induces apoptosis, and enhances TRAIL sensitivity of breast cancer cells by upregulating DR5.

Daurisoline (DS) is an isoquinoline alkaloid that exerts anticancer activities in various cancer cells. However, the underlying mechanisms through which DS affects the survival of breast cancer cells remain poorly understood. Therefore, the present study was undertaken to investigate the potential anticancer effect of DS on breast cancer cells and reveal the mechanism underlying the enhancedtumor necrosis factor-related apoptosis-inducing ligand(TRAIL)-mediated apoptosis by DS. Cell counting kit-8(CCK-8) and 5-ethynyl-2-deoxyuridine (EdU) assay were used to evaluate the ability of cell proliferation. Flow cytometry was selected to examine the cell cycle distribution. TUNELassay was used to detect the cell apoptosis. The protein expression was measured by Western blot analysis. DS was found to reduce the cell viability and suppress the proliferation of MCF-7 and MDA-MB-231 cells by causing G1 phase cell cycle arrest. DS could trigger apoptosis by promoting the cleavage of caspase-8 and PARP. The phosphorylation of ERK,JNK,and p38MAPK was upregulated clearly following DS treatment. Notably, SP600125 (JNK inhibitor) pretreatment significantly abrogated DS-induced PARP cleavage. DS inactivatedAkt/mTOR and Wnt/β-catenin signaling pathwayand upregulated the expression of ER stress-related proteins. Additionally, DS amplified TRAIL-caused viability reduction and apoptosis in breast cancer cells. Mechanismly, DS upregulated the protein level of DR4 and DR5, and knockdown of DR5 attenuated the cotreatment-induced cleavage of PARP. Inhibition of JNK could block DS-induced upregulation of DR5. This study provides valuable insights into the mechanisms of DS inhibiting cell proliferation, triggering apoptosis, and enhancing TRAIL sensitivity of breast cancer cells.

Apoptosis as a therapeutic strategy for breast cancer: the role of Thymax, a gross thymic extract, in modulating cell death pathways

Background Breast cancer is a prevalent disease in women and a leading cause of cancer-related health issues. Thymax, a thymic extract, has shown potential for inducing breast cancer cell apoptosis in vitro. Objective This study aims to investigate how Thymax induces apoptosis and inhibits breast cancer growth and metastasis in vivo. Materials and methods Thymax treatment was divided into five groups: the first group (negative control) − normal rats without tumors. In the second group (positive control), rats were injected subcutaneously in the mammary gland with a single dose of 50 mg/kg b.w. of 7,12-Dimethylbenz(a)anthracene (in 2 ml of corn oil) and allowed to develop tumors for 120 days. Group 3: Thymax was orally administered 6 days a week to tumor-bearing rats (0.4 mg/rat) and continued for 5 weeks. Tumor-bearing rats in group 4 (Thymax injection) received 0.1 ml of Thymax solution through intraperitoneal injection twice weekly for 5 weeks. The last group was Thymax mix (oral and injection); tumor-bearing rats received Thymax solution by dual routes: orally with 0.4 ml six times per week and intraperitoneally with 0.1 ml twice weekly for 5 weeks. Thymax treatment, beginning after 120 days of tumor induction, continued for 5 weeks. Results and conclusion Thymax- induced apoptosis in breast cancer cells by increasing cytochrome c, tumor necrosis factor receptor type 1-associated death domain protein (TRADD), and Fas associated death domain (FADD) levels. It also activated the mitochondrial-dependent pathway with up-regulation of tumor protein gene (P53) expression and cysteine-dependent, aspartate-specific peptidase (caspase-8) activation. Thymax restored normal renal and hepatic cell function and enhanced the immune system by improving total antioxidant levels and inhibiting malondialdehyde levels in treated animals. Histopathological results showed a significant apoptotic effect in the group receiving Thymax injections, demonstrating its capability to induce apoptosis without tumors or atypia in mammary glands. Our findings indicate that Thymax has a significant effect on enhancing tumor cell death and inducing apoptosis in vivo. Thymax may also modulate proapoptotic and antiapoptotic protein expression and activity, regulate the penetrability of the mitochondrial membrane, and release cytochrome c. Furthermore, our findings show that the injection route of Thymax is the fastest and most efficient method to deliver the extract to the tumor site and exert its antitumor effects. These results suggest that Thymax has the potential to be a novel adjuvant in the treatment of breast cancer, as it can enhance the efficacy of conventional therapies and reduce the risk of recurrence and metastasis.

PDP1 promotes the progression of breast cancer through STAT3 pathway.

This study aimed to investigate the expression pattern and mechanisms of Pyruvate Dehydrogenase Phosphatase Catalytic Subunit 1 (PDP1) in the progression of breast cancer (BC). PDP1, known for its involvement in cell energy metabolism, was found to be overexpressed in BC tissues. Notably, low PDP1 expression aligns with improved overall survival (OS) in BC patients. In this study, we found that PDP1 was overexpressed among BC tissues and low PDP1 expression showed a better prognosis for the patients with BC. PDP1 knockdown suppressed cell amplification and migration and triggered cell apoptosis in BC cells. In vivo assessments through a xenograft model unveiled the pivotal role and underlying mechanisms of PDP1 knockdown. RNA sequencing and kyoto encyclopedia of genes and genomes analysis of RNAs from PDP1 knockdown and normal MCF7 cells revealed 1440 differentially expressed genes, spotlighting the involvement of the JAK/STAT3 signaling pathway in BC progression. Western blot results implied that PDP1 knockdown led to a loss of p-STAT3, whereas overexpression of PDP1 induced the p-STAT3 expression. Cell counting kit-8 assay showed that PDP1 overexpression significantly raised MDA-MB-231 and MCF7 cell viability while STAT3 inhibitor S3I-201 recovered the cell growth to normal level. To summarize, PDP1 promotes the progression of BC through STAT3 pathway by regulating p-STAT3. The findings contribute to understanding the molecular mechanisms underlying BC progression, and opening avenues for targeted therapeutic approaches.

Elevated level of neuroserpin is an indication for the resistance to gambogic acid-induced apoptosis and oxidative stress in triple-negative breast cancer cells.

The triple-negative breast cancer (TNBC) subtype, characterized by loss of HER2, estrogen, and progesterone receptors, displays aggressive phenotype and poor prognosis compared to other BC subtypes. Since the TNBC cells are devoid of receptors, endocrine therapy is an ineffective option for TNBC patients, necessitating canonical chemotherapy strategies to treat TNBC. It is crucial to use alternative and natural agents to support chemotherapy in TNBC. To clarify the molecular mechanism of the tumorigenic effects of gambogic acid (GA) on TNBC cells with different epithelial character since GA has a wide spectrum of anticancer activity for most cancer types. We determined the cytotoxic dose of GA incubation of TNBC cells (MDA-MB-231 and BT-20 cells) for 24 h. We performed the MTT test and toluidine blue (TB) staining protocol for TNBC cells. We analyzed E-cadherin, N-cadherin, Bax, and neuroserpin mRNAs in both cells by qPCR. We evaluated apoptosis using DAPI staining and assessed the ROS using the 2',7'-dichlorofluorescin diacetate (DCFH-DA) method. We determined the IC50 concentrations of GA in MDA-MB-231 and BT-20 cells to be 315.8 nM and 441.8 nM, respectively. TB staining showed that BT-20 cells survive at excessive cytotoxic doses of GA, while most of the MDA-MB-231 cells were killed. Also, we found that BT-20 cells are more resistant to GA-induced apoptosis and oxidative stress than the MDA-MB-231 cells. qPCR results showed that GA upregulated neuroserpin, an oxidative stress-relieving factor in the BT-20 cells, but not in the MDA-MB-231 cells. The elevated level of neuroserpin could be a predictive marker to determine the development of resistance to chemotherapeutic agents.