Aggressive Breast Cancer Cells Research Articles

Lung-resident alveolar macrophages regulate the timing of breast cancer metastasis.

Breast disseminated cancer cells (DCCs) can remain dormant in the lungs for extended periods, but the mechanisms limiting their expansion are not well understood. Research indicates that tissue-resident alveolar macrophages suppress breast cancer metastasis in lung alveoli by inducing dormancy. Through ligand-receptor mapping and intravital imaging, it was found that alveolar macrophages express transforming growth factor (TGF)-β2. This expression, along with persistent macrophage-cancer cell interactions via the TGF-βRIII receptor, maintains cancer cells in a dormant state. Depleting alveolar macrophages or losing the TGF-β2 receptor in cancer cells triggers metastatic awakening. Aggressive breast cancer cells are either suppressed by alveolar macrophages or evade this suppression by avoiding interaction and downregulating the TGF-β2 receptor. Restoring TGF-βRIII in aggressive cells reinstates TGF-β2-mediated macrophage growth suppression. Thus, alveolar macrophages act as a metastasis immune barrier, and downregulation of TGF-β2 signaling allows cancer cells to overcome macrophage-mediated growth suppression.

Antiproliferative Activity of an Organometallic Sn(IV) Coordination Compound Based on 1-Methylbenzotriazole against Human Cancer Cell Lines

A motivating class of compounds with interest in the research field of biological active metallopharmaceuticals for cancer treatment is based on organometallic complexes of Sn(IV), exhibiting advantages such as improved cellular uptake and body excretion, lower toxicity, and fewer side effects compared to platinum-based drugs. In this study, the mononuclear organotin coordination complex [(CH3)2SnCl2(mebta)2] was synthesized and characterized using vibrational spectroscopy (IR, Raman), 1H NMR, 13C{1H} NMR, and X-ray crystallography. Its antiproliferative properties were thoroughly assessed across an aggressive triple-negative human breast cancer cell line. Notably, comparative studies with precursor materials verified that the observed biological activity is intrinsic to the complex itself. This study highlights the compound’s ability to induce cell fate by disrupting essential cellular functions, such as proliferation. By exploring the antiproliferative effects of organotin(IV) derivatives, we introduce a novel class of Sn complexes with 1-methylbenzotriazole (mebta), demonstrating significant potential as promising antitumor agents in the field of organotin compounds.

The effects of an aggressive breast tumor on thrombosis after antithrombin downregulation in a hypercoagulable mouse model

BackgroundDespite improvements in therapy, breast cancer still contributes to high mortality rates. Survival of these patients becomes progressively worse upon diagnosis with cancer-associated thrombosis (CAT). Unfortunately, the mechanism causing CAT has remained unclear. ObjectiveSet up an acute and non-invasive hypercoagulable mouse model with an aggressive breast cancer and study the mechanism of cancer-associated thrombosis. MethodsMice were grafted with the aggressive breast cancer cell line MDA-MB-231 or sham-treated. Subsequently, an acute imbalance in coagulation was introduced by injecting a synthetic small interfering (si) RNA targeting hepatic Serpinc1 to knockdown antithrombin – a condition known to predispose to cause a hypercoagulant state in vivo. ResultsSilencing Serpinc1 with siRNA decreased plasma antithrombin levels. siRNA treatment had no short-term effects on tumor characteristics, but increased distant metastasis within the timeframe of this study. The systemic pro-inflammatory status, with elevated platelet counts and fibrinogen levels in tumor-bearing mice, was also not affected by antithrombin silencing. While elevated fibrin deposition in the liver upon Serpinc1 targeting was not significantly affected by the presence of breast cancer, knockdown of antithrombin did significantly increase intratumoral fibrin deposition and inflammation. Surprisingly, in the presence of an aggressive tumor, a protective outcome with less clinical features coinciding with venous thrombosis were observed in mice with antithrombin knockdown. ConclusionWe conclude that the presence of a breast tumor protects hypercoagulant mice from severe consumption of coagulation factors after lowering hepatic antithrombin levels, possibly due to elevated platelet counts. However, the consequences on cancer-associated thrombosis remained inconclusive.

Discovering the potential of biomaterial-based mesoporous and magneto-luminescent nanohybrid (Nd-doped Hydroxyapatite/Fe3O4) for cancer theragnosis

Multimodality nanoplatforms play a crucial role in advancing medical interventions by integrating multiple functionalities into a single system. However, issues like intricate production processes and biocompatibility persist. Herein, a facile synthesis of a biomaterial-based mesoporous nanocarrier, HAp:Nd+SPIONs@mSiO2 is reported. The nanohybrid with ∼100 nm average size, comprised of Nd-doped hydroxyapatite (HAp:Nd) nanophosphor, Fe3O4 superparamagnetic iron oxide nanoparticles (SPIONs), and mesoporous silica, exhibiting magneto-luminescent properties. The nanohybrid showed NIR to NIR photoluminescence properties important for deep tissue imaging. The mesoporous nanohybrid was loaded with Indocyanine green (ICG), a photosensitizer and photothermal dye, as a model drug (∼6 μg/mg of nanoparticles) with a high absorption stability retaining >75 % drug until 24 h incubation in pH 6 and 7.4, respectively. Nanoparticles demonstrated dual functionality by generating heat through magnetic and photonic stimulation, as well as producing reactive oxygen species (ROS) upon excitation with 808 nm light. In vitro assays on aggressive triple-negative breast cancer cells (MDA-MB-231) showed the high biocompatibility of nanohybrid with and without ICG, while a significant toxicity was seen after irradiation of NIR light due to ROS production. Noticeably, the nanohybrids also exhibit the ability to monitor temperature changes via Nd3+ associated NIR luminescence. The nanoplatform integrates clinically relevant components like hydroxyapatite, SPIONs, mesoporous silica and ICG, highlighting its potential for translational applications. The developed nanohybrids, with combined NIR-mediated photothermal and photodynamic effects, magnetic photothermal capabilities, and NIR/MR imaging, offer promise in addressing cancer heterogeneity and improving conventional treatments with reduced side effects.

Melatonin increases Olaparib sensitivity and suppresses cancer-associated fibroblast infiltration via suppressing the LAMB3-CXCL2 axis in TNBC

Triple-negative breast cancer (TNBC) is the most malignant breast cancer subtype, characterized with high aggressiveness and a high recurrence rate. Olaparib is the first US Food and Drug Administration-approved poly(ADP ribose) polymerase (PARP) inhibitor (PARPi) to treat breast cancer patients with a germline BRCA1 or BRCA2 mutation. However, resistance to Olaparib treatment restricts the therapeutic effects, and thus novel therapeutics are urgently required. In the present study, we identified that the combination of melatonin and Olaparib synergistically enhanced the sensitivity of TNBC cells. Moreover, melatonin exerted promising antitumor activities in Olaparib-resistant cells, implying the potential for its clinical application. An RNA-sequencing analysis revealed that melatonin treatment downregulated laminin subunit beta 3 (LAMB3) expression. Genetic ablation of LAMB3 significantly increased Olaparib sensitivity, and subsequently suppressed proliferation, epithelial-to-mesenchymal transition (EMT)-related gene expressions, and aggressiveness of breast cancer cells. Accordingly, LAMB3 expression was positively correlated with C-X-C motif chemokine ligand 2 (CXCL2), and they collaboratively promoted cancer-associated fibroblast (CAF) infiltration. An in vivo study demonstrated that combined treatment with melatonin and Olaparib showed enhanced inhibitory efficacy against tumor growth, LAMB3 expression, CXCL2 levels, and CAF infiltration compared to single treatment groups, and combined treatment with melatonin and Olaparib significantly ameliorated the immunosuppressive tumor microenvironment. These findings illustrate a promising therapeutic strategy using melatonin to overcome Olaparib resistance and activate antitumor immunity via attenuating the LAMB3-CXCL2 axis in breast cancer patients.

Synthetic Benzylic Diselenides and Disulfides: Potential Anticancer Activities via Modulation of the ROS-dependent Akt/β-Catenin Signaling Pathway.

The natural and synthetic organodiselenides have garnered much research attention due to their chemotherapeutic and chemopreventive activities. Herein, we describe the synthesis of a series of benzylic diselenides, which were synthesized by coupling the in situ generated disodium diselenide with the corresponding benzylic halides. The diselenides were evaluated for their anticancer activities in the highly aggressive triple-negative breast cancer cells. Preliminary anti-proliferative activities indicated 4-cyano-substituted diselenide 7 to be most potent with an IC50 value of 1.9 ± 0.3 μM. Detailed mechanistic investigations showed that diselenide 7 induces apoptosis and causes G1 phase arrest of the cell cycle. It exhibits anticancer activity by suppressing the Akt/β-catenin signaling pathway. Further control experiments with LiCl (inhibitor of GSK-3β) revealed that down-regulation of β-catenin was promoted by GSK-3β-induced phosphorylation of β-catenin and its subsequent proteasomal degradation. Moreover, the intracellular ROS was found to act as an upstream mediator for the inactivation of the Akt/β-catenin signaling pathway. The present study describing the efficient anticancer activity of a synthetic benzylic diselenide towards triple-negative breast cancer cells through the modulation of ROS-dependent Akt/β-catenin signaling pathway would certainly be helpful in the future towards the development of small-molecule organoselenium compounds for the treatment of cancer.

Antioxidant Properties of Lippia alba Essential Oil: A Potential Treatment for Oxidative Stress-Related Conditions in Plants and Cancer Cells.

Lippia alba (Mill.) N.E.Br. ex Britton and P. Wilson is used in folk medicine of Central and South America for its biological activities: i.e., antifungal, antibacterial, antiviral, and anti-inflammatory. Based on ethnopharmacological information and the increasing interest in this species, this work aimed to test a possible wide use of its essential oil (EO) in pharmaceutical and horticultural applications. Therefore, we focused the attention on the antioxidant activity of the oil as a possible tool to overcome the oxidative stress in both applications. For this purpose, we have chosen three aggressive breast cancer cell lines and two horticultural species (Solanum lycopersicum L. and Phaseolus acutifolius L.) that are very sensitive to salt stress. We determined the antioxidant activity of L. alba EO through the quantification of phenols and flavonoids. Regarding tomato and bean plants under salt stress, L. alba EO was used for the first time as a seed priming agent to enhance plant salt tolerance. In this case, the seed treatment enhanced the content of phenolic compounds, reduced power and scavenger activity, and decreased membrane lipid peroxidation, thus mitigating the oxidative stress induced by salt. While in breast cancer cells the EO treatment showed different responses according to the cell lines, i.e., in SUM149 and MDA-MB-231 the EO decreased proliferation and increased antioxidant activity and lipid peroxidation, showing high cytotoxic effects associated with the release of lactate dehydrogenase, vice versa no effect was observed in MDA-MB-468. Such antioxidant activity opens a new perspective about this essential oil as a possible tool to counteract proliferation in some cancer cell lines and in horticulture as a seed priming agent to protect from oxidative damage in crops sensitive to salinity.

DMF-DM-seq: Digital-Microfluidics Enabled Dual-Modality Sequencing of Single-Cell mRNA and microRNA with High Integration, Sensitivity, and Automation.

Multimodal measurement of single cells provides deep insights into the intricate relationships between individual molecular layers and the regulatory mechanisms underlying intercellular variations. Here, we reported DMF-DM-seq, a highly integrated, sensitive, and automated platform for single-cell mRNA and microRNA (miRNA) co-sequencing based on digital microfluidics. This platform first integrates the processes of single-cell isolation, lysis, component separation, and simultaneous sequencing library preparation of mRNA and miRNA within a single DMF device. Compared with the current half-cell measuring strategy, DMF-DM-seq enables complete separation of single-cell mRNA and miRNA via a magnetic field application, resulting in a higher miRNA detection ability. DMF-DM-seq revealed differential expression patterns of single cells of noncancerous breast cells and noninvasive and aggressive breast cancer cells at both mRNA and miRNA levels. The results demonstrated the anticorrelated relationship between miRNA and their mRNA targets. Further, we unravel the tumor growth and metastasis-associated biological processes enriched by miRNA-targeted genes, along with important miRNA-interaction networks involved in significant signaling pathways. We also deconstruct the miRNA regulatory mechanisms underlying different signaling pathways across different breast cell types. In summary, DMF-DM-seq offers a powerful tool for a comprehensive study of the expression heterogeneity of single-cell mRNA and miRNA, which will be widely applied in basic and clinical research.

The influence of the chlorine atom on the biological activity of 2′-hydroxychalcone in relation to the lipid phase of biological membranes - Anticancer and antimicrobial activity

The study investigates the effect of the presence of a chlorine atom in the 2′-hydroxychalcone molecule on its interaction with model lipid membranes, in order to discern its potential pharmacological activity. Five chlorine derivatives of 2′-hydroxychalcone were synthesized and evaluated against liposomes composed of POPC and enriched with cationic (DOTAP) or anionic (POPG) lipids. The physicochemical properties of the compounds were initially simulated using SwissAdame software, revealing high lipophilicity (ilogP values: 2.79–2.90). The dynamic light scattering analysis of liposomes showed that chloro chalcones induce minor changes in the diameter of liposomes of different surface charges. Fluorescence quenching assays with a TMA-DPH probe demonstrated the strong ability of the compounds to interact with the lipid bilayer, with varying quenching capacities based on chlorine atom position. FTIR studies indicated alterations in carbonyl, phosphate, and choline groups, suggesting a transition area localization rather than deep penetration into the hydrocarbon chains. Additionally, dipole potential reduction was observed in POPC and POPC-POPG membranes, particularly pronounced by derivatives with a chlorine atom in the B ring. Antibacterial and antibiofilm assays revealed enhanced activity of derivatives with a chlorine atom compared to 2′-hydroxychalcone, especially against Gram-positive bacteria. The MIC and MBIC50 values showed increased efficacy in the presence of chlorine with 3′-5′-dichloro-2′-hydroxychalcone demonstrating optimal antimicrobial and antibiofilm activity. Furthermore, antiproliferative assays against breast cancer cell lines indicated higher activity of B-ring chlorine derivatives, particularly against MDA-MB-231 cells. In general, the presence of a chlorine atom in 2′-hydroxychalcone improves its pharmacological potential, with derivatives showing improved antimicrobial, antibiofilm, and antiproliferative activities, especially against aggressive breast cancer cell lines. These findings underscore the importance of molecular structure in modulating biological activity and highlight chalcones with a chlorine as promising candidates for further drug development studies.

Mitigation of Breast Cancer Cells' Invasiveness via Down Regulation of ETV7, Hippo, and PI3K/mTOR Pathways by Vitamin D3 Gold-Nanoparticles.

Metastasis in breast cancer is the major cause of death in females (about 30%). Based on our earlier observation that Vitamin D3 downregulates mTOR, we hypothesized that Vitamin D3 conjugated to gold nanoparticles (VD3-GNPs) reduces breast cancer aggressiveness by downregulating the key cancer controller PI3K/AKT/mTOR. Western blots, migration/invasion assays, and other cell-based, biophysical, and bioinformatics studies are used to study breast cancer cell aggressiveness and nanoparticle characterization. Our VD3-GNP treatment of breast cancer cells (MCF-7 and MDA-MB-231) significantly reduces the aggressiveness (cancer cell migration and invasion rates > 45%) via the simultaneous downregulation of ETV7 and the Hippo pathway. Consistent with our hypothesis, we, indeed, found a downregulation of the PI3K/AKT/mTOR pathway. It is surprising that the extremely low dose of VD3 in the nano formulation (three orders of magnitude lower than in earlier studies) is quite effective in the alteration of cancer invasiveness and cell signaling pathways. Clearly, VD3-GNPs are a viable candidate for non-toxic, low-cost treatment for reducing breast cancer aggressiveness.

MCT4 knockdown by tumor microenvironment-responsive nanoparticles remodels the cytokine profile and eradicates aggressive breast cancer cells

Breast cancer is a wide-spread threat to the women’s health. The drawbacks of conventional treatments necessitate the development of alternative strategies, where gene therapy has regained hope in achieving an efficient eradication of aggressive tumors. Monocarboxylate transporter 4 (MCT4) plays pivotal roles in the growth and survival of various tumors, which offers a promising target for treatment. In the present study, pH-responsive lipid nanoparticles (LNPs) based on the ionizable lipid,1,2-dioleoyl-3-dimethylammonium propane (DODAP), were designed for the delivery of siRNA targeting MCT4 gene to the breast cancer cells. Following multiple steps of characterization and optimization, the anticancer activities of the LNPs were assessed against an aggressive breast cancer cell line, 4T1, in comparison with a normal cell line, LX-2. The selection of the helper phospholipid to be incorporated into the LNPs had a dramatic impact on their gene delivery performance. The optimized LNPs enabled a powerful MCT4 silencing by ∼90 % at low siRNA concentrations, with a subsequent ∼80 % cytotoxicity to 4T1 cells. Meanwhile, the LNPs demonstrated a 5-fold higher affinity to the breast cancer cells versus the normal cells, in which they had a minimum effect. Moreover, the MCT4 knockdown by the treatment remodeled the cytokine profile in 4T1 cells, as evidenced by 90 % and ∼64 % reduction in the levels of TNF-α and IL-6; respectively. The findings of this study are promising for potential clinical applications. Furthermore, the simple and scalable delivery vector developed herein can serve as a breast cancer-targeting platform for the delivery of other RNA therapeutics.

LIFU/MMP-2 dual-responsive release of repurposed drug disulfiram from nanodroplets for inhibiting vasculogenic mimicry and lung metastasis in triple-negative breast cancer

BackgroundVasculogenic mimicry (VM), when microvascular channels are formed by cancer cells independent of endothelial cells, often occurs in deep hypoxic areas of tumors and contributes to the aggressiveness and metastasis of triple-negative breast cancer (TNBC) cells. However, well-developed VM inhibitors exhibit inadequate efficacy due to their low drug utilization rate and limited deep penetration. Thus, a cost-effective VM inhibition strategy needs to be designed for TNBC treatment.ResultsHerein, we designed a low-intensity focused ultrasound (LIFU) and matrix metalloproteinase-2 (MMP-2) dual-responsive nanoplatform termed PFP@PDM-PEG for the cost-effective and efficient utilization of the drug disulfiram (DSF) as a VM inhibitor. The PFP@PDM-PEG nanodroplets effectively penetrated tumors and exhibited substantial accumulation facilitated by PEG deshielding in a LIFU-mediated and MMP-2-sensitive manner. Furthermore, upon exposure to LIFU irradiation, DSF was released controllably under ultrasound imaging guidance. This secure and controllable dual-response DSF delivery platform reduced VM formation by inhibiting COL1/pro-MMP-2 activity, thereby significantly inhibiting tumor progression and metastasis.ConclusionsConsidering the safety of the raw materials, controlled treatment process, and reliable repurposing of DSF, this dual-responsive nanoplatform represents a novel and effective VM-based therapeutic strategy for TNBC in clinical settings.Graphical

UPR-Activated IRE1 Signaling Pathway Promotes Malignant Growth of Breast Cancer

Initially, it was believed that the unfolded protein response (UPR) was a network signal that synchronized apoptotic and adaptive responses to the buildup of unfolded proteins in the endoplasmic reticulum (ER). The endoplasmic reticulum stress sensor IRE1 is a major player in the UPR, and overactivation of IRE1 can promote a variety of pathological phenomena, such as diabetes, neurodegenerative diseases, and cancer. Under endoplasmic reticulum stress, IRE1 is activated through autophosphorylation and oligomerization, leading to conformational changes in the RNase domain, thereby promoting two signaling pathways: 1) unconventional splicing of XBP1 and 2) regulated IRE1 dependence Decay (RIDD). Research has revealed that aggressive B-type breast cancer cells typically have an overexpressed and amplified IRE1 gene. Through the RIDD mechanism, IRE1 processes and regulates the breakdown of a fraction of tumor-suppressing micro-RNA, which in breast cancer cells results in the Ras oncogene GTPase. RAB3B is elevated. This article reveals the mechanism of action of the IRE1 signaling pathway and the RIE1-RIDD-miRNA pathway in promoting the malignant development of luminal breast cancer.

Abstract 3385: Modeling evolutionary fitness in resistant cancers based on a common adaptability trait

Abstract Cancer is composed of a major subpopulation of proliferative cells and a minor subpopulation of highly adaptable cells. Although it is evident that cancer is an evolution like process driven by rare adaptable cancer cells that persist under various selection pressures, most of our knowledge about cancer comes from studying proliferative cancer cells. To overcome the hurdle in modeling adaptable cancer cell state that defeats all currently offered therapies, we are exploiting a linkage that exists between different adaptability substates such as metabolic, regulatory, and structural substates. Our studies, thus far carried out with aggressive triple-negative breast cancer and melanoma cell lines, suggest that it is feasible to apply severe metabolic challenges as realistic selection pressures/bottlenecks for modeling highly abnormal and highly adaptable cancer cells that drive cancer evolution and therapy resistance. The most significant aspect of our approach is an ability to model rare resistant cells (approximately 0.01% of cells) that survive in reversible quiescence under selection pressures. Monitoring of cells under microscope for several weeks under a severe and prolonged metabolic challenge, for example, a lack of glutamine in culture medium, revealed perhaps most interesting and cancer-relevant cells that survive in quiescence for weeks and then advance to generate therapy-resistant cells. Such microscopic monitoring revealed a significant heterogeneity among surviving cells. As examples, 1) surviving cells differ in the depth of quiescence, some cells trying to proliferate sooner than others; 2) as cells progress from quiescence to proliferation, their fate is far from certain: cells may progress to yield few cells, tiny colony, or large colony and then stop proliferating and in most instances die (abortive attempts at evolution). Of significance, the cell lines derived inflammatory breast cancer (IBC), an aggressive subtype of breast cancer, possessed a higher evolutionary fitness than non-IBC cell lines. This approach also eliminates 99.99% of cells that proliferate in cell culture but would not survive selection pressures in the body. The adaptable cells that survived the bottleneck and then proliferated indefinitely were highly resistant and metastatic. Molecular analyses of adaptable cells, including gene expression microarrays, CGH microarrays, and whole genome sequencing revealed mechanisms for their plasticity (exemplified by markers of high EMT), and for generating cellular diversity by altering epigenome (exemplified by low TET2) and by structural modifications in transcriptome (exemplified by high FTO). We conclude that the phenotype-based approach described here is good at modeling deep intrinsic resistance and may be useful in developing strategies for improving outcomes in resistant cancers. Supported by a State of Texas Grant for Rare and Aggressive Cancers. Citation Format: Balraj Singh, Vanessa N. Sarli, Anthony Lucci. Modeling evolutionary fitness in resistant cancers based on a common adaptability trait [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3385.

Abstract 6772: A 3D model of breast cancer-bone metastasis to mimic dysregulation of bone remodeling and drug response

Abstract Introduction: Bone metastases are common among advanced stage breast cancer patients and induce dysregulation of bone remodeling. Crosstalk between cancer cells and the bone niche drives the vicious cycle of bone loss and breast cancer growth. To accelerate discovery of targeted therapies, there is a critical need for experimental models that can mimic breast cancer-induced dysregulation of bone remodeling in patients. Mouse models are the current gold standard, but are low throughput, costly, and cannot fully recapitulate human-human cell interactions. To address these limitations, our goal is to engineer a fully human 3D in vitro model of breast cancer-bone metastasis that can mimic dysregulation of bone remodeling and drug responses in patients, and enable screening of potential new drug candidates. Methods: Two human breast cancer cell lines (MDA-MB-231, MCF-7) were chosen given their well characterized clinical features and tendency to invade bone. Our 3D model comprises an outer ring of tissue engineered (TE) bone and a center of breast cancer cells to mimic invasion through the bone marrow/long bone interface. Human mesenchymal stem cells were encapsulated in 3D gelatin microribbon scaffolds and cultured in osteogenic media for 28 days to form TE bone. Human monocyte-derived osteoclasts were seeded onto the bone, and the center was replaced with GFP-labeled breast cancer cells. Denosumab and zoledronic acid were selected as they are standard treatments for reducing bone loss in patients, and were used to treat the model at 30 µg/mL and 2 µg/mL, respectively. Intermittent parathyroid hormone (iPTH) treatment was selected as it showed efficacy in animal models, and was used to treat the model at 10 µg/mL for 6 hours daily. All groups were treated with drugs for 14 days. Bone volume was assessed using microCT. Cancer cell proliferation and invasion into bone was monitored with fluorescence confocal microscopy. Results: MicroCT imaging showed both breast cancer cell lines induced bone loss in the triculture model, compared to the control with no breast cancer cells in the center. Consistent with known clinical features, the more aggressive cell line (MDA-MB-231) induced a greater amount of bone loss than the less aggressive breast cancer cell line (MCF-7). Consistent with clinical findings, both denosumab and zoledronic acid reduced bone loss in our 3D model, and zoledronic acid further reduced breast cancer cell invasion and proliferation. Finally, the 3D model recapitulated the in vivo drug response of iPTH treatment, reducing bone resorption and breast cancer invasion. Conclusions: Here we report a fully human triculture model of breast cancer-bone metastasis that can mimic key clinical features of breast cancer-induced bone resorption and drug response in patients. Such a 3D model could enable screening of drug candidates with reduced time and cost and enable identification of novel druggable targets. Citation Format: Michelle Tai, Eva C. González Díaz, Callan E. Monette, Joy Wu, Fan Yang. A 3D model of breast cancer-bone metastasis to mimic dysregulation of bone remodeling and drug response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6772.

BRCA1 deficiency enhances the aggressiveness of breast cancer cells expressing HPV16 oncoproteins.

The precise etiology of breast cancer is not completely understood, although women with BRCA1 gene mutations have a significantly increased risk of developing the disease. In addition, sporadic breast cancer is frequently associated with decreased BRCA1 gene expression. Growing evidence of Human papillomaviruses (HPVs) infections in breast tumors has raised the possibility of the involvement of HPVs in the pathogenesis of breast cancer. We investigated whether the effects of HPV oncoproteins E6 and E7 were influenced by the expression levels of BRCA1. HPV16E6E7 (prototype or E6D25E/E7N29S Asian variant type) were stably expressed in MDA-MB231 breast cancer cells, wild type for BRCA1, or with BRCA1 knocked down. Expression of HPV16E6E7 oncogenes did not affect BRCA1 levels and the abundance of HPV16E6E7 was not altered by BRCA1 knockdown. BRCA1 levels did not alter HPV16E6E7-dependent degradation of G1-S cell cycle proteins p53 and pRb. However, we found that the expression of G2-M cell cycle protein cyclin B1 enhanced by HPV16E6E7 was impacted by BRCA1 levels. Especially, we found the correlation between BRCA1 and cyclin B1 expression and this was also confirmed in breast cancer samples from a Thai cohort. We further demonstrated that the combination of HPV oncoproteins and low levels of BRCA1 protein appears to enhance proliferation and invasion. Transactivation activities of HPV16E6E7 on genes regulating cell proliferation and invasion (TGF-β and vimentin) were significantly increased in BRCA1-deficient cells. Our results indicate that a deficiency of BRCA1 promotes the transactivation activity of HPV16E6E7 leading to increase of cell proliferation and invasion. HPV infection appears to have the potential to enhance the aggressiveness of breast cancers, especially those deficient in BRCA1.

Evaluation of mAb 2C5-modified dendrimer-based micelles for the co-delivery of siRNA and chemotherapeutic drug in xenograft mice model

Combination therapy with small interfering RNA (siRNA) and chemotherapeutic drug is proven to be effective in downregulating cancer resistance proteins, such as P-glycoprotein (P-gp). These proteins are involved in multidrug resistance (MDR) of tumors. A targeted formulation capable of delivering siRNA and chemotherapeutic drug will not only downregulate P-gp but also increase the concentration of the chemotherapeutic drug at the site of tumor thereby increasing the therapeutic effect and lowering the systemic exposure. In this study, monoclonal antibody 2C5-modified dendrimer-based micelles were used to co-deliver siRNA and doxorubicin (DOX) to the tumor site in both male and female xenograft mouse model. The nucleosome-specific 2C5 antibody recognizes the cancer cells via the cell-surface bound nucleosomes. The ability of ability of the 2C5-modified formulation to affect the metastasis of highly aggressive triple negative breast cancer cell migration in (MDA-MB-231) was assessed by a wound healing. Further, the therapeutic efficacy of the formulation was assessed by measuring the tumor volume progression in which the 2C5-modified nanoparticle group had a similar tumor volume to the free drug group at the end of the study, although a 50% increase in DOX concentrations in blood was observed after the last dose of nanoparticle. The free drug group on the other hand showed body weight reduction as well as the visible irritation around the injection spot. The treatment group with 2C5-modified micelles has shown to be safe at the current dose of DOX and siRNA. Furthermore, the siRNA mediated P-gp downregualtion was studied using western blotting assay. We observed a 29% reduction of P-gp levels in both males and females with respect to the control (BHG). We also conclude that the dose of DOX and siRNA should be further optimized to have a better efficacy in a metastatic tumor model, which will be the subject of our future studies.Graphical abstract

Effect of Pistacia Terebinthus (terebinth) resin extract on aggressive breast cancer cells.

Breast cancer is one of the most common types of cancer causing high mortality rates among women worldwide. This study was aimed to evaluate the effect of Pistacia terebinthus (terebinth) resin extract (TRE) on the MDA-MB-231 breast cancer cell line. In the study, the cytotoxic dose of the resin extract in MDA-MB-231 cells was evaluated by MTS analysis. The effect of TRE on apoptosis was examined by Hoechst staining. Caspase-3 and cleaved caspase-3 protein expressions were determined by western blot analysis. Based on the outcomes of our MTS analysis, the IC50 dose of TRE was calculated at 56.54μg/mL during a 24-h application period. With Hoechst staining analysis, an increase was observed in cells that underwent apoptotic change at 10 and 100μg/ml TRE concentrations compared to the control. At 25 and 50μg/mL TRE concentrations, no apoptotic change was found in comparison to the control; however, a significant drop in the number of viable cells was observed because 200, 300, and 500μg/mL TRE concentrations were above the toxic dose. The caspase-3 protein expression level was significantly higher in cells treated with 100μg/ml TRE compared to the control group, while there was no significant change in cleaved caspase-3 protein expression. It was thought that P. terebinthus resin might cause cell death in MDA-MB-231 cells via caspase-independent apoptosis pathway or other cell death pathways, and it was concluded that it could be a supportive treatment for breast cancer treatment.

NOX4 alleviates breast cancer cell aggressiveness by co-ordinating mitochondrial turnover through PGC1α/Drp1 axis

Triple Negative Breast Cancer (TNBC) is a highly aggressive form of breast cancer, with few treatment options. This study investigates the complex molecular mechanism by which NADPH oxidase 4 (NOX4), a major ROS producer in mitochondria, affects the aggressiveness of luminal and triple-negative breast cancer cells (TNBCs). We found that NOX4 expression was differentially regulated in luminal and TNBC cells, with a positive correlation to their epithelial characteristics. Time dependent analysis revealed that TNBCs exhibits higher steady-state ROS levels than luminal cells, but NOX4 silencing increased ROS levels in luminal breast cancer cells and enhanced their ability to migrate and invade. In contrast, NOX4 over expression in TNBCs had the opposite effect. The mouse tail-vein experiment showed that the group injected with NOX4 silenced luminal cells had a higher number of lung metastases compared to the control group. Mechanistically, NOX4 enhanced PGC1α dependent mitochondrial biogenesis and attenuated Drp1-mediated mitochondrial fission in luminal breast cancer cells, leading to an increased mitochondrial mass and elongated mitochondrial morphology. Interestingly, NOX4 silencing increased mitochondrial ROS (mtROS) levels without affecting mitochondrial (Δψm) and cellular integrity. Inhibition of Drp1-dependent fission with Mdivi1 reversed the effect of NOX4-dependent mitochondrial biogenesis, dynamics, and migration of breast cancer cells. Our findings suggest that NOX4 expression diminishes from luminal to a triple negative state, accompanied by elevated ROS levels, which may modulate mitochondrial turnover to attain an aggressive phenotype. The study provides potential insights for targeted therapies for TNBCs.

177Lu-Labeled Iron Oxide Nanoparticles Functionalized with Doxorubicin and Bevacizumab as Nanobrachytherapy Agents against Breast Cancer.

The use of conventional methods for the treatment of cancer, such as chemotherapy or radiotherapy, and approaches such as brachytherapy in conjunction with the unique properties of nanoparticles could enable the development of novel theranostic agents. The aim of our current study was to evaluate the potential of iron oxide nanoparticles, coated with alginic acid and polyethylene glycol, functionalized with the chemotherapeutic agent doxorubicin and the monoclonal antibody bevacizumab, to serve as a nanoradiopharmaceutical agent against breast cancer. Direct radiolabeling with the therapeutic isotope Lutetium-177 (177Lu) resulted in an additional therapeutic effect. Functionalization was accomplished at high percentages and radiolabeling was robust. The high cytotoxic effect of our radiolabeled and non-radiolabeled nanostructures was proven in vitro against five different breast cancer cell lines. The ex vivo biodistribution in tumor-bearing mice was investigated with three different ways of administration. The intratumoral administration of our functionalized radionanoconjugates showed high tumor accumulation and retention at the tumor site. Finally, our therapeutic efficacy study performed over a 50-day period against an aggressive triple-negative breast cancer cell line (4T1) demonstrated enhanced tumor growth retention, thus identifying the developed nanoparticles as a promising nanobrachytherapy agent against breast cancer.