Unveiling the Mechanism of the ChaiShao Shugan Formula Against Triple-Negative Breast Cancer.

Background: The ChaiShao Shugan Formula (CSSGF) is a traditional Chinese medicine formula with recently identified therapeutic value in triple-negative breast cancer (TNBC). This study aimed to elucidate the underlying mechanism of CSSGF in TNBC treatment. Methods: TNBC targets were analyzed using R and data were from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. The major ingredients and related protein targets of CSSGF were explored via the Traditional Chinese Medicine Systems Pharmacology database, and an ingredient-target network was constructed via Cytoscape to identify hub genes. The STRING database was used to construct the PPI network. GO and KEGG enrichment analyses were performed via R to obtain the main targets. The online tool Kaplan‒Meier plotter was used to identify the prognostic genes. Molecular docking was applied to the core target genes and active ingredients. MDA-MB-231 and MCF-7 cell lines were used to verify the efficacy of the various drugs. Results: A total of 4562 genes were screened as TNBC target genes. The PPI network consisted of 89 nodes and 845 edges. Our study indicated that quercetin, beta-sitosterol, luteolin and catechin might be the core ingredients of CSSGF, and EGFR and c-Myc might be the latent therapeutic targets of CSSGF in the treatment of TNBC. GO and KEGG analyses indicated that the anticancer effect of CSSGF on TNBC was mainly associated with DNA binding, transcription factor binding, and other biological processes. The related signaling pathways mainly involved the TNF-a, IL-17, and apoptosis pathways. The molecular docking data indicated that quercetin, beta-sitosterol, luteolin, and catechin had high affinity for EGFR, JUN, Caspase-3 and ESR1, respectively. In vitro, we found that CSSGF could suppress the expression of c-Myc or promote the expression of EGFR. In addition, we found that quercetin downregulates c-Myc expression in two BC cell lines. Conclusion: This study revealed the effective ingredients and latent molecular mechanism of action of CSSGF against TNBC and confirmed that quercetin could target c-Myc to induce anti-BC effects.

GSH-activable heterotrimeric nano-prodrug for precise synergistic therapy of TNBC.

Combination chemotherapy is an effective approach for triple-negative breast cancer (TNBC) therapy, especially when drugs are administered at specific optimal ratios. However, at present, strategies involving precise and controllable ratios based on effective loading and release of drugs are unavailable. Herein, we designed and synthesized a glutathione (GSH)--responsive heterotrimeric prodrug and formulated it with an amphiphilic polymer to obtain nanoparticles (DSSC2 NPs) for precise synergistic chemotherapy of TNBC. The heterotrimeric prodrug was prepared using docetaxel (DTX) and curcumin (CUR) at the optimal synergistic ratio of 1: 2. DTX and CUR were covalently conjugated by disulfide linkers. Compared with control NPs, DSSC2 NPs had quantitative/ratiometric drug loading, high drug co-loading capacity, better colloidal stability, and less premature drug leakage. After systemic administration, DSSC2 NPs selectively accumulated in tumor tissues and released the encapsulated drugs triggered by high levels of GSH in cancer cells. In vitro and in vivo experiments validated that DSSC2 NPs released DTX and CUR at the predefined ratio and had a highly synergistic therapeutic effect on tumor suppression in TNBC, which can be attributed to ratiometric drug delivery and synchronous drug activation. Altogether, the heterotrimeric prodrug delivery system developed in this study represents an effective and novel approach for combination chemotherapy.

Nitric oxide nano-reactor DNMF/PLGA enables tumor vascular microenvironment and chemo-hyperthermia synergetic therapy.

BACKGROUND: Breast cancer ranks first among malignant tumors, of which triple-negative breast cancer (TNBC) is characterized by its highly invasive behavior and the worst prognosis. Timely diagnosis and precise treatment of TNBC are substantially challenging. Abnormal tumor vessels play a crucial role in TNBC progression and treatment. Nitric oxide (NO) regulates angiogenesis and maintains vascular homeostasis, while effective NO delivery can normalize the tumor vasculature. Accordingly, we have proposed here a tumor vascular microenvironment remodeling strategy based on NO-induced vessel normalization and extracellular matrix collagen degradation with multimodality imaging-guided nanoparticles against TNBC called DNMF/PLGA. RESULTS: Nanoparticles were synthesized using a chemotherapeutic agent doxorubicin (DOX), a NO donor L-arginine (L-Arg), ultrasmall spinel ferrites (MnFe2O4), and a poly (lactic-co-glycolic acid) (PLGA) shell. Nanoparticle distribution in the tumor was accurately monitored in real-time through highly enhanced magnetic resonance imaging and photoacoustic imaging. Near-infrared irradiation of tumor cells revealed that MnFe2O4 catalyzes the production of a large amount of reactive oxygen species (ROS) from H2O2, resulting in a cascade catalysis of L-Arg to trigger NO production in the presence of ROS. In addition, DOX activates niacinamide adenine dinucleotide phosphate oxidase to generate and supply H2O2. The generated NO improves the vascular endothelial cell integrity and pericellular contractility to promote vessel normalization and induces the activation of endogenous matrix metalloproteinases (mainly MMP-1 and MMP-2) so as to promote extravascular collagen degradation, thereby providing an auxiliary mechanism for efficient nanoparticle delivery and DOX penetration. Moreover, the chemotherapeutic effect of DOX and the photothermal effect of MnFe2O4 served as a chemo-hyperthermia synergistic therapy against TNBC. CONCLUSION: The two therapeutic mechanisms, along with an auxiliary mechanism, were perfectly combined to enhance the therapeutic effects. Briefly, multimodality image-guided nanoparticles provide a reliable strategy for the potential application in the fight against TNBC.

Injectable Alginate Complex Hydrogel Loaded with Dual-Drug Nanovectors Offers Effective Photochemotherapy against Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC), accounting for approximately 20% of breast cancer cases, is a particular subtype that lacks tumor-specific targets and is difficult to treat due to its high aggressiveness and poor prognosis. Chemotherapy remains the major systemic treatment for TNBC. However, its applicability and efficacy in the clinic are usually concerning due to a lack of targeting, adverse side effects, and occurrence of multidrug resistance, suggesting that the development of effective therapeutics is still highly demanded nowadays. In this study, an injectable alginate complex hydrogel loaded with indocyanine green (ICG)-entrapped perfluorocarbon nanoemulsions (IPNEs) and camptothecin (CPT)-doped chitosan nanoparticles (CCNPs), named IPECCNAHG, was developed for photochemotherapy against TNBC. IPNEs with perfluorocarbon can induce hyperthermia and generate more singlet oxygen than an equal dose of free ICG upon near-infrared (NIR) irradiation to achieve photothermal and photodynamic therapy. CCNPs with positive charge may facilitate cellular internalization and provide sustained release of CPT to carry out chemotherapy. Both nanovectors can stabilize agents in the same hydrogel system without interactions. IPECCNAHG integrating IPNEs and CCNPs enables stage-wise combinational therapeutics that may overcome the issues described above. With 60 s of NIR irradiation, IPECCNAHG significantly inhibited the growth of MDA-MB-231 tumors in the mice without systemic toxicity within the 21 day treatment. We speculate that such anticancer efficacy was accomplished by phototherapy followed by chemotherapy, where cancer cells were first destroyed by IPNE-derived hyperthermia and singlet oxygen, followed by sustained damage with CPT after internalization of CCNPs; a two-stage tumoricidal process. Taken together, the developed IPECCNAHG is anticipated to be a feasible tool for TNBC treatment in the clinic.

Isobavachalcone, a natural sirtuin 2 inhibitor, exhibits anti-triple-negative breast cancer efficacy in vitro and in vivo.

Triple-negative breast cancer (TNBC) is the most aggressive and lethal clinical subtype and lacks effective targeted therapies at present. Isobavachalcone (IBC), the main active component of Psoralea corylifolia L., has potential anticancer effects. Herein, we identified IBC as a natural sirtuin 2 (SIRT2) inhibitor and characterized the potential mechanisms underlying the inhibition of TNBC. Molecular dynamics analysis, enzyme activity assay, and cellular thermal shift assay were performed to evaluate the combination of IBC and SIRT2. The therapeutic effects, mechanism, and safety of IBC were analyzed in vitro and in vivo using cellular and xenograft models. IBC effectively inhibited SIRT2 enzyme activity with an IC50 value of 0.84 ± 0.22 µM by forming hydrogen bonds with VAL233 and ALA135 within its catalytic domain. In the cellular environment, IBC bound to and stabilized SIRT2, consequently inhibiting cellular proliferation and migration, and inducing apoptosis and cell cycle arrest by disrupting the SIRT2/α-tubulin interaction and inhibiting the downstream Snail/MMP and STAT3/c-Myc pathways. In the in vivo model, 30 mg/kg IBC markedly inhibited tumor growth by targeting the SIRT2/α-tubulin interaction. Furthermore, IBC exerted its effects by inducing apoptosis in tumor tissues and was well-tolerated. IBC alleviated TNBC by targeting SIRT2 and triggering the reactive oxygen species ROS/ß-catenin/CDK2 axis. It is a promising natural lead compound for future development of SIRT2-targeting drugs.

Teadenol B as a Component of Microorganism-Fermented Tea Extract Inhibited Breast Cancers by Promoting Autophagy.

Breast cancer is a significant threat to life and health, which needs more safe and effective drugs to be explored. Teadenol B is a characteristic chemical component of microbial fermented tea. This study discovered that teadenol B could exhibit obvious inhibitory effects on all four different clinical subtype characteristics of breast cancer cells. Proteomic studies show that deoxycytidine triphosphate deaminase (DCTD), which could block DNA synthesis and repair DNA damage, had the most significant and consistent reduction in all four types of breast cancer cells with the treatment of teadenol B. Considering MDA-MB-231 cells exhibit poor clinical prognosis and displayed substantial statistical differences in KEGG pathway enrichment analysis results, we investigated its impact on the size and growth of MDA-MB-231 triple-negative breast tumors transplanted into nude mice and demonstrated that teadenol B significantly suppressed tumor growth without affecting body weight significantly. Finally, we found that the conversion of LC3-I to LC3-II in MDA-MB-231 increased significantly with teadenol B treatment. This proved that teadenol B could be a strong autophagy promotor, which explained the down-regulation of DCTD to some extent and may be the potential mechanism underlying teadenol B's anti-breast cancer effects. This finding provides new evidence for drinking fermented tea to prevent breast cancer and highlights the potential of teadenol B as a novel therapeutic option for breast cancer prevention and treatment, necessitating further investigations to clarify its exact target and the details involved.

Adjuvant platinum versus capecitabine for residual, invasive, triple-negative breast cancer: Patient-reported outcomes in ECOG-ACRIN EA1131.

BACKGROUND: Patient-reported outcomes (PROs) are a better tool for evaluating the experiences of patients who have symptomatic, treatment-associated adverse events (AEs) compared with clinician-rated AEs. The authors present PROs assessing health-related quality of life (HRQoL) and treatment-related neurotoxicity for adjuvant capecitabine versus platinum on the Eastern Cooperative Oncology Group-American College of Radiology Imaging Network (ECOG-ACRIN) EA1131 trial (ClinicalTrials.gov identifier NCT02445391). METHODS: Participants completed the National Comprehensive Cancer Network Functional Assessment of Cancer Therapy-Breast Cancer Symptom Index (NFBSI-16) and the Functional Assessment of Cancer Therapy-Gynecologic Oncology Group neurotoxicity subscale (platinum arm only) at baseline, cycle 3 day 1 (C3D1), 6 months, and 15 months. Because of early termination, power was insufficient to test the hypothesis that HRQoL, as assessed by the NFBSI-16 treatment side-effect (TSE) subscale, would be better at 6 and 15 months in the capecitabine arm; all analyses were exploratory. Means were compared by using t-tests or the Wilcoxon rank-sum test, and proportions were compared by using the χ2 test. RESULTS: Two hundred ninety-six of 330 eligible patients provided PROs. The mean NFBSI-16 TSE subscale score was lower for the platinum arm at baseline (p = .02; absolute difference, 0.6 points) and for the capecitabine arm at C3D1 (p = .04; absolute difference, 0.5 points), but it did not differ at other times. The mean change in TSE subscale scores differed between the arms from baseline to C3D1 (platinum arm, 0.15; capecitabine arm, -0.72; p = .03), but not from baseline to later time points. The mean decline in Functional Assessment of Cancer Therapy-Gynecologic Oncology Group neurotoxicity subscale scores exceeded the minimal meaningful change (1.38 points) from baseline to each subsequent time point (all p < .05). CONCLUSIONS: Despite the similar frequency of clinician-rated AEs, PROs identified greater on-treatment symptom burden with capecitabine and complemented clinician-rated AEs by characterizing patients' experiences during chemotherapy.

Photoresponsive 'chemo-free' phytotherapy: formulation development for the treatment of triple-negative breast cancer.

Aim: The present investigation aimed to develop a chemo-free, nanophytosomal system to treat triple-negative breast cancer (TNBC) via a phyto-photo dual treatment strategy. Method: Size, shape, surface analysis, photoprovoked release profile, photothermal stability, (3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide assay, apoptotic assay, DNA fragmentation, in vitro cellular uptake evaluation, mitochondrial membrane potential and caspase-3 assay, and photodynamic evaluation. Results: Biological experiments using MDA-MB-231 cells displayed dose-dependent synergistic anti-TNBC activity of PhytoS/Houttuynia cordata extract (HCE)/IR780 as compared with Phyto/HCE, PhytoS/IR780 and even more promising under laser treatment. Apoptotic assay and DNA fragmentation analysis also showed enhanced anti-TNBC effects. Investigation found that HCE acts via suppression of mitochondrial membrane potential and inducing caspase-3 activity in cells. Conclusion: Our findings suggested that photo-empowered phytotherapy can be employed effectively and safely against TNBC.

Brucea javanica derived exosome-like nanovesicles deliver miRNAs for cancer therapy.

Triple-negative breast cancer (TNBC) is characterized by complex heterogeneity, high recurrence and metastasis rates, and short overall survival, owing to the lack of endocrine and targeted receptors, which necessitates chemotherapy as the major treatment regimen. Exosome-like nanovesicles derived from medicinal plants have shown great potential as novel biotherapeutics for cancer therapy by delivering their incorporated nucleic acids, especially microRNAs (miRNAs), to mammalian cells. In this study, we isolated exosome-like nanovesicles derived from B. javanica (BF-Exos) and investigated their influence and underlying molecular mechanisms in TNBC. We found that BF-Exos delivered 10 functional miRNAs to 4T1 cells, significantly retarding the growth and metastasis of 4T1 cells by regulating the PI3K/Akt/mTOR signaling pathway and promoting ROS/caspase-mediated apoptosis. Moreover, BF-Exos were shown to inhibit the secretion of vascular endothelial growth factor, contributing to anti-angiogenesis in the tumor microenvironment. In vivo, BF-Exos inhibited tumor growth, metastasis, and angiogenesis in breast tumor mouse models, while maintaining high biosafety. Overall, BF-Exos are considered promising nanoplatforms for the delivery of medicinal plant-derived nucleic acids, with great potential to be developed into novel biotherapeutics for the treatment of TNBC.

In-vitro Antioxidant Activity and Flow Cytometric Analysis of Simarouba glauca DC Bark Extract Induced Apoptosis in Triple Negative Breast Cancer Cells.

OBJECTIVE: Ethnomedicinally Simarouba glauca DC is an important plant containing major class of phenols and terpenoids as bioactive compounds. The present study focuses on the evaluation of the anticancer effects of S. glauca bark UAE-EA (Ultrasonicator Assisted Extraction - Ethyl Acetate) fraction (SG-Fraction) against MDA-MB-231 triple negative breast cancer cell lines. METHODS: UAE-EA technique was used for the extraction of phytochemicals from S. glauca bark. Fractionation method was carried out to obtain Ethyl acetate fraction and PPS, TPC, and DPPH assays were performed to characterize the extract. MTT assay was then applied to analyse the viability of cells and MMP assay to confirm the initiation of drug induced apoptosis. Apoptotic morphology and quantification were assessed by DAPI and Annexin V/propidium iodide (PI) staining. RESULTS: UAE yielded 53g of crude extract in methanol. 16g Ethyl acetate fraction was obtained from fractionation. Phytoconstituents such as alkaloids, phenols, flavonoids, and triterpenoids were detected. The TPC was 278.65 mg GAE/100ml. The SG-Fraction showed maximum 66.38% RSA at 200 µg/ml and IC50 value was 101.72 µg/ml. MMP confirmed the induction of apoptosis. DAPI showed the reduction of nuclei with bright chromatin condensation, blebbing, nuclear fragmentation and apoptotic bodies. Annexin-V FITC/PI study showed 59.48% apoptosis induction. This fraction showed a similar trend of antioxidant effect as compared to ascorbic acid but, prominently lower cell viability than Camptothecin (P<0.005). In line with higher TPC in the SG-fraction, free radical scavenging activity was increased (r = 0.098**, p=0.002) and cell viability was reduced significantly (r = -0.097*** p<0.01). CONCLUSION: These results indicate that UAE-EA fraction of S. glauca bark inhibits the growth of MDA-MB-231 cells and can be considered for further neo-adjuvant chemotherapy drug research.

Encapsulated Rose Bengal Enhances the Photodynamic Treatment of Triple-Negative Breast Cancer Cells.

Among breast cancer subtypes, triple-negative breast cancer stands out as the most aggressive, with patients facing a 40% mortality rate within the initial five years. The limited treatment options and unfavourable prognosis for triple-negative patients necessitate the development of novel therapeutic strategies. Photodynamic therapy (PDT) is an alternative treatment that can effectively target triple-negative neoplastic cells such as MDA-MB-231. In this in vitro study, we conducted a comparative analysis of the PDT killing rate of unbound Rose Bengal (RB) in solution versus RB-encapsulated chitosan nanoparticles to determine the most effective approach for inducing cytotoxicity at low laser powers (90 mW, 50 mW, 25 mW and 10 mW) and RB concentrations (50 µg/mL, 25 µg/mL, 10 µg/mL and 5 µg/mL). Intracellular singlet oxygen production and cell uptake were also determined for both treatment modalities. Dark toxicity was also assessed for normal breast cells. Despite the low laser power and concentration of nanoparticles (10 mW and 5 µg/mL), MDA-MB-231 cells experienced a substantial reduction in viability (8 ± 1%) compared to those treated with RB solution (38 ± 10%). RB nanoparticles demonstrated higher singlet oxygen production and greater uptake by cancer cells than RB solutions. Moreover, RB nanoparticles display strong cytocompatibility with normal breast cells (MCF-10A). The low activation threshold may be a crucial advantage for specifically targeting malignant cells in deep tissues.

Sulfated polysaccharides of Laetiporus sulphureus fruiting bodies exhibit anti-breast cancer activity through cell cycle arrest, apoptosis induction, and inhibiting cell migration.

ETHNOPHARMACOLOGICAL RELEVANCE: Laetiporus sulphureus has long been used as an edible and medicinal mushroom in Asia, America, and Europe. Its fruiting bodies are widely used in folk medicine for treating cancer, gastric diseases, cough, and rheumatism. Polysaccharides are an important bioactive component of mushrooms. In nature, sulfated polysaccharides have never been reported in mushrooms. Furthermore, there is no information on differences in physicochemical properties and anti-breast cancer activities between polysaccharides (PS) and sulfated polysaccharides (SPS) of L. sulphureus. AIM OF THE STUDY: This study aimed to investigate the physicochemical properties of PS and SPS isolated from fruiting bodies of L. sulphureus and examine their anti-proliferative effects and mechanism(s) of action on MDA-MB-231 breast cancer cells. METHODS: Polysaccharides (PS) were isolated using hot water and ethanol precipitation methods. Sulfated polysaccharides (SPS) were isolated by the papain-assisted hydrolysis method. Physicochemical properties comprising sugar, protein, uronic acid, and sulfate contents, and molecular weight, monosaccharide composition, and structural conformation were analyzed on PS and SPS. In the anti-cancer study, a triple-negative breast cancer cell line (MDA-MB-231) and a normal human mammary epithelial cell line (H184B5F5/M10) were used to evaluate the anti-proliferative activity of PS and SPS, and their mechanism(s) of action. RESULTS: The results showed that SPS, which had higher sulfate and protein contents and diversified monosaccharide composition, exhibited more potent anti-proliferative activity against MDA-MB-231 cells than PS. Furthermore, it had a selective cytotoxic effect on breast cancer cells but not the normal cells. SPS induced cell cycle arrest at G0/G1 phase via down-regulating CDK4 and cyclin D1 and up-regulating p21 protein expression. Breast cancer cell apoptosis was not observed until 72 h after SPS treatment. In addition, SPS also markedly inhibited breast cancer cell migration. CONCLUSION: This study demonstrates that SPS exhibited selective cytotoxicity and was more potent than PS in inhibiting MDA-MB-231 cell proliferation. The contents of sulfate and protein, and monosaccharide composition could be the main factors affecting the anti-breast cancer activity of L. sulphureus SPS.

Danggui Buxue Tang improves therapeutic efficacy of doxorubicin in triple negative breast cancer via ferroptosis.

ETHNOPHARMACOLOGICAL RELEVANCE: Danggui Buxue Tang (DBT) has been used for over 800 years to enhance Qi and nourish Blood, and it is particularly beneficial for cancer patients. Recent research has shown that combining DBT with chemotherapy agents leads to superior anti-cancer effects, thereby enhancing therapeutic efficacy. AIM OF THE STUDY: The aim of this study was to evaluate the effectiveness of a combination therapy involving doxorubicin (DOX) and Danggui Buxue Tang (DBT) in the treatment of triple-negative breast cancer (TNBC) and to elucidate the underlying mechanisms of action. MATERIALS AND METHODS: In vitro experiments were performed using MDA-MB-231 and 4T1 cells, while in vivo experiments were carried out using MDA-MB-231 xenograft mice. The therapeutic effects of the combination therapy were evaluated using various techniques, including MTT assay, colony formation assay, flow cytometry, transwell assay, immunofluorescence, transmission electron microscopy (TEM), histological analysis, western blotting, and bioluminescence assay. RESULTS: DBT was found to enhance DOX's anti-TNBC activity in vitro by promoting ferroptosis, as evidenced by the observed mitochondrial morphological changes using TEM. The combination therapy was also found to reduce the expression of Nrf2, HO-1, and GPX4, which are all targets for ferroptosis induction, while simultaneously increasing ROS production. Additionally, the combination therapy reduced nuclear accumulation and constitutive activation of Nrf2, which is a significant cause of chemotherapy resistance and promotes cancer growth. In vivo experiments using an MDA-MB-231 xenograft animal model revealed that the combination therapy significantly reduced tumor cell proliferation and accelerated TNBC deaths by modulating the Nrf2/HO-1/GPX4 axis, with no evidence of tissue abnormalities. Moreover, the combination therapy exhibited a liver protective effect, and administration of Fer-1 was able to reduce the ROS formation produced by the DBT + DOX combination therapy. CONCLUSION: This study provides evidence that the combination therapy of DOX and DBT has the potential to treat TNBC by promoting ferroptosis through the Nrf2/HO-1/GPX4 axis.

Therapeutic effect and metabolic fingerprinting of triple-negative breast cancer cells following exposure to a novel pH-responsive, gambogic acid-loaded micelle.

Triple-negative breast cancer (TNBC) is a subtype of breast cancer with a poor prognosis and lacks effective therapeutic targets. The use of gambogic acid (GA), a class of active ingredients in traditional Chinese medicine with anti-tumour potential, is limited in tumour therapy owing to its drawbacks and unclear organ toxicity. In this study, we used the pH-responsive amphiphilic block copolymer, PEOz-PCL, to create nanodrugs for GA delivery to MDA-MB-231 cells. The pH-responsive GA-loaded micelles were prepared through nanoprecipitation with a more homogeneous size. The average particle size was 42.29 ± 1.74 nm, and the zeta potential value was 9.88 ± 0.17 mV. The encapsulation rate was 85.06%, and the drug loading rate was 10.63%. The process was reproducible, and sustained release reached 80% in 96 h at acid pH 5.0. Furthermore, cellular tests using CCK-8, TUNEL, and flow cytometry revealed that pH-responsive GA-loaded micelles killed MDA-MB-231 cells more effectively and had much higher activity and targeting compared with free drugs. Metabolomic analysis of the changes in differential metabolites revealed that pH-responsive GA-loaded micelles may inhibit TNBC cells by causing amino acid anabolism, nucleotide metabolism, and glucose metabolism, as well as by affecting their energy sources. The study outcomes will help understand the mechanism of action and the therapeutic efficacy of pH-responsive GA-loaded micellesin vivo.

Multiomics of HER2-low triple-negative breast cancer identifies a receptor tyrosine kinase-relevant subgroup with therapeutic prospects.

To provide complementary information and reveal the molecular characteristics and therapeutic insights of HER2-low breast cancer, we performed this multiomics study of hormone receptor-negative (HR-) and HER2-low breast cancer, also known as HER2-low triple-negative breast cancer (TNBC), and identified 3 subgroups: basal-like, receptor tyrosine kinase-relevant (TKR), and mesenchymal stem-like. These 3 subgroups had distinct features and potential therapeutic targets and were validated in external data sets. Interestingly, the TKR subgroup (which exists in both HR+ and HR- breast cancer) had activated HER2 and downstream MAPK signaling. In vitro and in vivo patient-derived xenograft experiments revealed that pretreatment of the TKR subgroup with a tyrosine kinase inhibitor (lapatinib or tucatinib) could inhibit HER2 signaling and induce accumulated expression of nonfunctional HER2, resulting in increased sensitivity to the sequential HER2-targeting, Ab-drug conjugate DS-8201. Our findings identify clinically relevant subgroups and provide potential therapeutic strategies for HER2-low TNBC subtypes.

Glycolysis inhibition for synergistic phototherapy of triple-negative breast cancer.

Phototherapy is a local and precise therapeutic technique for tumor treatment. However, the therapeutic effects of photothermal and photodynamic therapies are inevitably encountered by hypoxia of the tumor microenvironment and heat shock protein induced by hyperthermia, respectively. Herein, we found that mannose, a glucose analog, could reverse tumor hypoxia by inhibiting glycolysis of cancer cells and suppressing the expression of heat shock protein through inhibiting cellular adenosine triphosphate (ATP) generation. Next, we used lipid nanoparticles simultaneously loaded with indocyanine green (ICG) and mannose molecules, named imLipo, for tumor therapy. Both in vitro and in vivo experiments evidenced that the imLipo nanoplatform has significant therapeutic efficacy through synergistic phototherapy under single near-infrared laser irradiation. This work shows that glycolysis inhibition can overcome the challenges of phototherapy. In addition, all three parts (mannose, ICG, and lipid) of imLipo are clinically approved and our designed nanoplatforms have great potential for future tumor treatment.

Inhibition of glycolysis-driven immunosuppression with a nano-assembly enhances response to immune checkpoint blockade therapy in triple negative breast cancer.

Immune-checkpoint inhibitors (ICI) are promising modalities for treating triple negative breast cancer (TNBC). However, hyperglycolysis, a hallmark of TNBC cells, may drive tumor-intrinsic PD-L1 glycosylation and boost regulatory T cell function to impair ICI efficacy. Herein, we report a tumor microenvironment-activatable nanoassembly based on self-assembled aptamer-polymer conjugates for the targeted delivery of glucose transporter 1 inhibitor BAY-876 (DNA-PAE@BAY-876), which remodels the immunosuppressive TME to enhance ICI response. Poly ß-amino ester (PAE)-modified PD-L1 and CTLA-4-antagonizing aptamers (aptPD-L1 and aptCTLA-4) are synthesized and co-assembled into supramolecular nanoassemblies for carrying BAY-876. The acidic tumor microenvironment causes PAE protonation and triggers nanoassembly dissociation to initiate BAY-876 and aptamer release. BAY-876 selectively inhibits TNBC glycolysis to deprive uridine diphosphate N-acetylglucosamine and downregulate PD-L1 N-linked glycosylation, thus facilitating PD-L1 recognition of aptPD-L1 to boost anti-PD-L1 therapy. Meanwhile, BAY-876 treatment also elevates glucose supply to tumor-residing regulatory T cells (Tregs) for metabolically rewiring them into an immunostimulatory state, thus cooperating with aptCTLA-4-mediated immune-checkpoint inhibition to abolish Treg-mediated immunosuppression. DNA-PAE@BAY-876 effectively reprograms the immunosuppressive microenvironment in preclinical models of TNBC in female mice and provides a distinct approach for TNBC immunotherapy in the clinics.

[Hydnocarpin inhibits malignant progression of triple negative breast cancer via CNOT4-mediated ubiquitination and degradation of YAP].

This study aims to investigate the effect and mechanism of hydnocarpin(HC) in treating triple negative breast cancer(TNBC). Cell counting kit-8(CCK-8), xCELLigence real-time cellular analysis(RTCA), and colony formation assay were employed to determine the effects of HC on the proliferation of two TNBC cell lines: MDA-MB-231 and MDA-MB-436. The effects of HC on the migration and invasion of TNBC cells were detected by high-content analysis, wound-healing assay, and Transwell assay. The changes in the epithelial-mesenchymal transition(EMT) and the expression of invasion-and migration-associated proteins [E-cadherin, vimentin, Snail, matrix metalloproteinase-2(MMP-2), and MMP-9] were detected by Western blot. Western blot and RT-qPCR were employed to determine the protein and mRNA levels of Yes-associated protein(YAP) and downstream targets(CTGF and Cyr61). TNBC cells were transfected with Flag-YAP for the overexpression of YAP, and the role of YAP as a key target for HC to inhibit TNBC malignant progression was examined by CCK-8 assay, Transwell assay, and wound-healing assay. The pathway of HC-induced YAP degradation was detected by the co-treatment of proteasome inhibitor with HC and ubiquitination assay. The binding of HC to YAP and the E3 ubiquitin ligase Ccr4-not transcription complex subunit 4(CNOT4) was detected by microscale thermophoresis(MST) assay and drug affinity responsive target stability(DARTS) assay. The results showed that HC significantly inhibited the proliferation, colony formation, invasion, and EMT of TNBC cells. HC down-regulated the protein and mRNA levels of CTGF and Cyr61. HC down-regulated the total protein level of YAP, while it had no effect on the mRNA level of YAP. The overexpression of YAP antagonized the inhibitory effects of HC on the proliferation, migration, and invasion of TNBC cells. HC promoted the degradation of YAP through the proteasome pathway and up-regulated the ubiquitination level of YAP. The results of MST and DARTS demonstrated direct binding between HC, YAP, and CNOT4. The above results indicated that HC inhibited the malignant progression of TNBC via CNOT4-mediated degradation and ubiquitination of YAP.

Thymoquinone, a Novel Multi-Strike Inhibitor of Pro-Tumorigenic Breast Cancer (BC) Markers: CALR, NLRP3 Pathway and sPD-L1 in PBMCs of HR+ and TNBC Patients.

Breast cancer (BC) is not only a mass of malignant cells but also a systemic inflammatory disease. BC pro-tumorigenic inflammation has been shown to promote immune evasion and provoke BC progression. The NOD-like receptor (NLR) family pyrin domain-containing protein 3 (NLRP3) inflammasome is activated when pattern recognition receptors (PRRs) sense danger signals such as calreticulin (CALR) from damaged/dying cells, leading to the secretion of interleukin-1ß (IL-1ß). CALR is a novel BC biological marker, and its high levels are associated with advanced tumors. NLRP3 expression is strongly correlated with an elevated proliferative index Ki67, BC progression, metastasis, and recurrence in patients with hormone receptor-positive (HR+) and triple-negative BC (TNBC). Tumor-associated macrophages (TAMs) secrete high levels of IL-1ß promoting endocrine resistance in HR+ BC. Recently, an immunosuppressive soluble form of programmed death ligand 1 (sPD-L1) has been identified as a novel prognostic biomarker in triple-negative breast cancer (TNBC) patients. Interestingly, IL-1ß induces sPD-L1 release. BC Patients with elevated IL-1ß and sPD-L1 levels show significantly short progression-free survival. For the first time, this study aims to investigate the inhibitory impact of thymoquinone (TQ) on CALR, the NLRP3 pathway and sPD-L1 in HR+ and TNBC. Blood samples were collected from 45 patients with BC. The effect of differing TQ concentrations for different durations on the expression of CALR, NLRP3 complex components and IL-1ß as well as the protein levels of sPD-L1 and IL-1ß were investigated in the peripheral blood mononuclear cells (PBMCs) and TAMs of TNBC and HR+ BC patients, respectively. The findings showed that TQ significantly downregulated the expression of CALR, NLRP3 components and IL-1ß together with the protein levels of secreted IL-1ß and sPD-L1. The current findings demonstrated novel immunomodulatory effects of TQ, highlighting its potential role not only as an excellent adjuvant but also as a possible immunotherapeutic agent in HR+ and TNBC patients.

Targeting Breast Cancer Signaling via Phytomedicine and Nanomedicine.

BACKGROUND: The development of breast cancer (BC) and how it responds to treatment have both been linked to the involvement of inflammation. Chronic inflammation is critical in carcinogenesis, leading to elevated DNA damage, impaired DNA repair machinery, cell growth, apoptosis, angiogenesis, and invasion. Studies have found several targets that selectively modulate inflammation in cancer, limit BC's growth, and boost treatment effectiveness. Drug resistance and the absence of efficient therapeutics for metastatic and triple-negative BC contribute to the poor outlook of BC patients. SUMMARY: To treat BC, small-molecule inhibitors, phytomedicines, and nanoparticles are conjugated to attenuate BC signaling pathways. Due to their numerous target mechanisms and strong safety records, phytomedicines and nanomedicines have received much attention in studies examining their prospects as anti-BC agents by such unfulfilled demands. KEY MESSAGES: The processes involved in the affiliation across the progression of tumors and the spread of inflammation are highlighted in this review. Furthermore, we included many drugs now undergoing clinical trials that target cancer-mediated inflammatory pathways, cutting-edge nanotechnology-derived delivery systems, and a variety of phytomedicines that presently address BC.