Inhibited Breast Tumor Growth Research Articles

CircNFIB decreases synthesis of arachidonic acid and inhibits breast tumor growth and metastasis

We identified circNFIB (hsa_circ_0086376) as a down-regulated circRNA in breast cancer but its effect is unclear. We aimed to explore the roles of circNFIB in breast cancer. The expression levels of circNFIB in breast cancer tissues and cells were detected. Both in vitro and in vivo experiments were used to assess the effects and mechanisms of circNFIB. circNFIB was down-regulated in 29 breast cancer tissues compared to adjacent normal tissues. circNFIB is a highly conserved circRNA and mainly located in cytoplasm of breast cancer cells. In vitro experiments showed that overexpression of circNFIB inhibited proliferation and invasion of breast cancer cells, whereas knockdown of circNFIB induced proliferation and invasion. Animal experiments indicated that circNFIB inhibited tumor growth and metastasis in vivo. Bioinformatics analysis showed that circNFIB contained an open reading frame (ORF) spanning its spliced junction, an internal ribosome entry site (IRES) and a N6-methyladenosine (m6A) site, suggesting circNFIB had the potential to encode a 56 amino acid (aa) protein, which was then confirmed by experiments. Metabonomics analysis results indicated that circNFIB may inhibit synthesis of arachidonic acid (AA) by regulating phospholipase. EIF4A3 and U2AF65 may regulate circNFIB expression by binding to the flanking sequence of circNFIB. In conclusion, circNFIB is a down-regulated circRNA in breast cancer tissues and encodes a 56 aa protein. circNFIB down-regulates AA in breast cancer cells, thus decreasing AA metabolites. Based on reported evidences of AA metabolites on cancer, we speculated that circNFIB may inhibit breast tumor growth and metastasis partly by inhibiting AA.

Intelligent Nanoplatform Integrating Macrophage and Cancer Cell Membrane for Synergistic Chemodynamic/Immunotherapy/Photothermal Therapy of Breast Cancer.

Cell membrane-coated nanoplatforms for drug delivery have garnered significant attention due to their inherent cellular properties, such as immune evasion and homing abilities, making them a subject of widespread interest. The coating of mixed membranes from different cell types onto the surface of nanoparticles offers a way to harness natural cell functions, enhancing biocompatibility and improving therapeutic efficacy. In this study, we merged membranes from murine-derived 4T1 breast cancer cells with RAW264.7 (RAW) membranes, creating a hybrid biomimetic coating referred to as TRM. Subsequently, we fabricated hybrid TRM-coated Fe3O4 nanoparticles loaded with indocyanine green (ICG) and imiquimod (R837) for combination therapy in breast cancer. Comprehensive characterization of the RIFe@TRM nanoplatform revealed the inherent properties of both cell types. Compared to bare Fe3O4 nanoparticles, RIFe@TRM nanoparticles exhibited remarkable cell-specific self-recognition for 4T1 cells in vitro, leading to significantly prolonged circulation life span and enhanced in vivo targeting capabilities. Furthermore, the biomimetic RIFe@TRM nanoplatform induced tumor necrosis through the Fenton reaction and photothermal effects, while R837 facilitated enhanced uptake of tumor-associated antigens, further activating CD8+ cytotoxic T cells to strengthen antitumor immunotherapy. Hence, RIFe@TRM nanoplatform demonstrated outstanding synergy in chemodynamic/immunotherapy/photothermal therapies, displaying significant inhibition of breast tumor growth. In summary, this study presents a promising biomimetic nanoplatform for effective treatment of breast cancer.

Prox1 Suppresses the Proliferation of Breast Cancer Cells via Direct Inhibition of c-Myc Gene Expression.

Breast cancer is one of the most lethal malignancies in women worldwide and is characterized by rapid growth and low survival rates, despite advances in tumor biology and therapies. Novel therapeutic approaches require new insights into the molecular mechanisms of malignant transformation and progression. To this end, here, we identified Prox1 as a negative regulator of proliferation and tumor-related metabolism in breast cancer. In particular, we showed that breast tumors from human patients exhibited reduced levels of Prox1 expression, while high expression levels of Prox1 were associated with a favorable prognosis in breast cancer patients. Moreover, we experimentally demonstrated that Prox1 was sufficient to strongly suppress proliferation, migration, and the Warburg effect in human breast cancer cells without inducing apoptosis. Most importantly, over-expression of Prox1 inhibited breast tumor growth in vivo in both heterotopic and orthotopic xenograft mouse models. The anti-tumorigenic effect of Prox1 was mediated by the direct repression of c-Myc transcription and its downstream target genes. Consistently, c-Myc over-expression from an artificial promoter that was not targeted by Prox1 reversed Prox1's anti-tumor effects. These findings suggest that Prox1 has a tumor suppressive role via direct transcriptional regulation of c-Myc, making it a promising therapeutic gene for breast cancer.

CHMP4A stimulates CD8+ T-lymphocyte infiltration and inhibits breast tumor growth via the LSD1/IFNβ axis.

CD8+ T lymphocyte-mediated immunity strategies have represented attractive weapons against breast cancer (BC) recently. However, the mechanisms underlying CD8+ T-lymphocyte infiltration still remain obscure. Here, using bioinformatics analysis, we identified four hub prognostic genes related to CD8+ T-lymphocyte infiltration (CHMP4A, CXCL9, GRHL2, and RPS29), among which CHMP4A was the most significant gene. High CHMP4A mRNA expression was significantly associated with longer overall survival (OS) in BC patients. Functional experiments showed that CHMP4A had the ability to promote CD8+ T-lymphocyte recruitment and infiltration and suppressed BC growth in vitro and in vivo. Mechanistically, CHMP4A stimulates CD8+ T-lymphocyte infiltration by downregulating LSD1 expression, leading to HERV dsRNA accumulation, and promoting IFNβ and its downstream chemokine production. Collectively, CHMP4A is not only a novel positive predictor for prognosis in BC but also a stimulator of CD8+ T-lymphocyte infiltration regulated by the LSD1/IFNβ pathway. This study suggests that CHMP4A may be a novel target for improving the effectiveness of immunotherapy in patients with BC.

Breast Cancer Immunity: It is TIME for the Next Chapter.

Our ability to interrogate the tumor immune microenvironment (TIME) at an ever-increasing granularity has uncovered critical determinants of disease progression. Not only do we now have a better understanding of the immune response in breast cancer, but it is becoming possible to leverage key mechanisms to effectively combat this disease. Almost every component of the immune system plays a role in enabling or inhibiting breast tumor growth. Building on early seminal work showing the involvement of T cells and macrophages in controlling breast cancer progression and metastasis, single-cell genomics and spatial proteomics approaches have recently expanded our view of the TIME. In this article, we provide a detailed description of the immune response against breast cancer and examine its heterogeneity in disease subtypes. We discuss preclinical models that enable dissecting the mechanisms responsible for tumor clearance or immune evasion and draw parallels and distinctions between human disease and murine counterparts. Last, as the cancer immunology field is moving toward the analysis of the TIME at the cellular and spatial levels, we highlight key studies that revealed previously unappreciated complexity in breast cancer using these technologies. Taken together, this article summarizes what is known in breast cancer immunology through the lens of translational research and identifies future directions to improve clinical outcomes.

Natural product preferentially targets redox and metabolic adaptations and aberrantly active STAT3 to inhibit breast tumor growth in vivo

Dysregulated gene expression programs and redox and metabolic adaptations allow cancer cells to survive under high oxidative burden. These mechanisms also represent therapeutic vulnerabilities. Using triple-negative breast cancer (TNBC) as a model, we show that compared to normal human breast epithelial cells, the TNBC cells, MDA-MB-231 and MDA-MB-468 that harbor constitutively active STAT3 also express higher glucose-6-phosphate dehydrogenase (G6PD), thioredoxin reductase (TrxR)1, NADPH, and GSH levels for survival. Present studies discover that the natural product, R001, targets these adaptation mechanisms. Treatment of TNBC cells with R001 inhibited constitutively active STAT3, STAT3-regulated gene expression, and the functions of G6PD and TrxR1. Consequently, in the TNBC, but not normal cells, R001 suppressed GSH levels, but raised NADPH levels, reflective of a loss of mitochondrial respiration and which led to reactive oxygen species (ROS) induction, all of which led to loss of viable cells and inhibition of anchorage-dependent and independent growth. R001 treatment further led to early pyroptosis and late DNA damage, cell cycle arrest, and apoptosis only in the TNBC cells. Oral administration of 5 mg/kg R001 inhibited MDA-MB-468 xenografts growth in mice, with reduced pY705-STAT3, G6PD, TrxR1, and GSH levels. R001 serves as a therapeutic entity that targets the vulnerabilities of TNBC cells to inhibit tumor growth in vivo.

Metal-phenolic networks with ferroptosis to deliver NIR-responsive CO for synergistic therapy.

As an endogenous gasotransmitter, CO has achieved tremendous advances in cancer treatment through selectively killing cancer cells. However, the application of CO in tumor immunotherapy has not been reported and the tumor targeting delivery is still a tremendous challenge. Herein, thermosensitive boronic acid group-containing CO prodrug was synthesized and fabricated with tannic acid (TA) and iron (Fe) to form metal-phenolic networks, and then loaded with near-infrared (NIR) photothermal agent IR820 to form FeCO-IR820@FeIIITA for combinational therapy of CO and photothermal therapy. Ferroptosis can also be enhanced due to the Fe3+ incorporation. After TA reduced Fe3+ into Fe2+, Fe2+ might lead to intracellular Fenton reaction. Furthermore, in combination with CTLA-4 blockade immunotherapy, FeCO-IR820@FeIIITA remarkably inhibited breast tumor growth, suppressed the lung metastasis and improved the antitumor immune response. To summarize, FeCO-IR820@FeIIITA provides a potential novel option for CO/photothermal/immune synergistic therapy with enhanced ferroptosis through simple compositions and facile synthesis process.

Sabizabulin, a Potent Orally Bioavailable Colchicine Binding Site Agent, Suppresses HER2+ Breast Cancer and Metastasis.

HER2+ breast cancer accounts for 15% of all breast cancer cases. Current frontline therapy for HER2+ metastatic breast cancer relies on targeted antibodies, trastuzumab and pertuzumab, combined with microtubule inhibitors in the taxane class (paclitaxel or docetaxel). It is well known that the clinical efficacy of taxanes is limited by the development of chemoresistance and hematological and neurotoxicities. The colchicine-binding site inhibitors (CBSIs) are a class of promising alternative agents to taxane therapy. Sabizabulin (formerly known as VERU-111) is a potent CBSI that overcomes P-gp-mediated taxane resistance, is orally bioavailable, and inhibits tumor growth and distant metastasis in triple negative breast cancer (TNBC). Herein, we demonstrate the efficacy of sabizabulin in HER2+ breast cancer. In vitro, sabizabulin inhibits the proliferation of HER2+ breast cancer cell lines with low nanomolar IC50 values, inhibits clonogenicity, and induces apoptosis in a concentration-dependent manner. In vivo, sabizabulin inhibits breast tumor growth in the BT474 (ER+/PR+/HER2+) xenograft model and a HER2+ (ER-/PR-) metastatic patient-derived xenograft (PDX) model, HCI-12. We demonstrate that sabizabulin is a promising alternative agent to target tubulin in HER2+ breast cancer with similar anti-metastatic efficacy to paclitaxel, but with the advantage of oral bioavailability and lower toxicity than taxanes.

Novel bio-inspired lipid nanoparticles for improving the anti-tumoral efficacy of fisetin against breast cancer

• The anticancer activity of fisetin against breast cancer has been successfully emphasized through encapsulation into lipoprotein bioinspired lipid nanoparticles (LPINs). • The pharmaceutical and therapeutic activity of developed LPINs have been appraised in comparison with CD-44 targeting polysaccharide coated LPINs. • In-vitro cell line studies against breast cancer cells (MDA-MB-231) have shown significant four times increase in cytotoxicity of all developed nanosystems relative to free fisetin. • The preclinical studies revealed superior inhibition of breast tumor growth by fisetin loaded- LPINs in comparison with fisetin solution.

A Facile Total Synthesis of Kilogram-Scale Production of SKLB1039: A Novel and Selective Hexahydroisoquinolin-Containing EZH2 Inhibitor.

SKLB1039 is a potent, highly selective, and orally bioavailable EZH2 inhibitor, which significantly inhibited breast tumor growth and metastasis in pre-clinical studies. In a previously reported synthesis of SKLB1039, the yields of several steps were low, which led to an overall yield of less than 10%. In addition, flash chromatography was required for the purification of several intermediates using this route. To optimize the synthesis and establish an efficient commercial-scale method for the production of SKLB1039. The reaction time, solvent, reactant ratio, temperature, and mode of addition of reactants in the reductive amination, hydrolysis, hexahydroisoquinoline formation, hydrogenolysis, condensation and Suzuki crosscoupling reactions were optimized. A chromatography-free seven-step process starting from a commercially available material was developed that afforded SKLB1039 in 36% overall yield with > 99% purity. A cost-effective, high-yielding, and convergent kilo-scale synthesis for the EZH2 inhibitor SKLB1039 was developed. The operation was simple, and the pure product was easily obtained without column chromatography. This method will be economical and convenient for the subsequent industrial scale-up production of SKLB1039, which will be conducive for this promising EZH2 inhibitor to enter clinical studies of its antitumor effects.

Abstract 5476: Novel azetidine-based STAT3 inhibitors induce misfolded protein response, endoplasmic reticulum stress and mitophagy, and inhibit breast tumor growth in vivo

Abstract Anticancer drugs that directly target Signal Transducer and Activator of Transcription (Stat)3 have remained elusive. We present azetidine-based small molecules as a new class of potent Stat3 inhibitors. Compounds H172 and H182 have potency (IC50) of 0.38-0.98 µM, H105 and H120 have IC50 of 1.75-2.07 µM against Stat3 DNA-binding activity in vitro, compared to the IC50 of >17 µM against Stat1 or Stat5 activity. Treatment of the triple-negative breast cancer (TNBC), MDA-MB-231 and MDA-MB-468 cells with the new compounds inhibited constitutive Stat3 activation, promoted Stat3 ubiquitination, blocked anchorage-dependent and independent growth, with potency, EC50 of 1.0-1.9 μM, and induced apoptosis of these cells. By contrast, pYStat1, pYStat5, pS727Stat3, pSrc, pJanus kinase, or the induction of other signaling events are not inhibited in compound-treated cells, and normal or tumor cells that do not harbor aberrantly-active Stat3 are only weakly inhibited. In vivo delivery of H120 or H182 as a single agent inhibited growth of human TNBC xenografts, and of H278 (HCl salt of H182) combined with radiation abrogated tumor growth and improved survival in mouse TNBC syngeneic models. These studies provide novel mechanism of disruption of aberrantly-active Stat3 and its dynamics in showing that the azetidine-based Stat3 inhibitors trigger Stat3 ubiquitination, and tumor cell death. H182 and H172 are new entities for further therapeutic development against TNBC and other cancers that harbor aberrantly-active Stat3. Citation Format: Peibin Yue, Francisco Lopez-Tapia, Yinsong Zhu, Nagendra Verma, Felix Alonso-Valenteen, Simoun Mikhael, Lali Medina-Kauwe, Marcus Tius, James Turkson. Novel azetidine-based STAT3 inhibitors induce misfolded protein response, endoplasmic reticulum stress and mitophagy, and inhibit breast tumor growth in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5476.

Mitophagy Induction and Aryl Hydrocarbon Receptor-Mediated Redox Signaling Contribute to the Suppression of Breast Cancer Cell Growth by Taloxifene via Regulation of Inflammasomes Activation.

Aims: Raloxifene, a selective estrogen receptor (ER) modulator, has been reported to exert the tumor-suppressive effects in both ER-positive and ER-negative cancer cells; however, the mechanisms underlying its ER-independent anti-cancer effects are poorly understood. The NLRP3 inflammasome, a critical component of the innate immune system, has recently received growing attention owing to its multifaceted roles in various aspects of cancer development. The present study aimed at examining the involvement of NLRP3 inflammasomes in the anti-breast cancer effects of raloxifene and its underlying mechanisms. Results: Raloxifene significantly inhibited the activation of NLRP3 inflammasomes in various breast cancer cell lines. Importantly, forced expression of a gain-of-function variant of NLRP3 rescued breast cancer cells from growth arrest by raloxifene, suggesting that the suppression of NLRP3 inflammasomes activation mediates the raloxifene-induced inhibition of breast cancer growth. Mechanistically, raloxifene suppressed NLRP3 inflammasomes activation by lowering the cellular levels of reactive oxygen species (ROS) through the modulation of redox signaling mediated via aryl hydrocarbon receptor (AhR)-nuclear factor erythroid 2-related factor 2 (Nrf2)-heme oxygenase-1 (HO-1) axis or the impaired generation of mitochondrial ROS in a mitophagy-dependent manner. Further, the blockage of AhR signaling or inhibition of mitophagy abolished the tumor-suppressive effect of raloxifene in a human breast tumor xenograft model. Innovation: We elucidate a novel molecular mechanism underlying the breast tumor suppressing effect of raloxifene. Conclusion: The results observed in this study suggest that the modulation of NLRP3 inflammasomes activation is a critical event in the inhibition of breast tumor growth by raloxifene. Antioxid. Redox Signal. 37, 1030-1050.

The PRMT5-LSD1 axis confers Slug dual transcriptional activities and promotes breast cancer progression

BackgroundDownregulation of epithelial markers and upregulation of mesenchymal markers are the characteristics of the epithelial to mesenchymal transition (EMT) program, which provides the metastatic advantage of breast cancer. However, the mechanism underlying the switch of EMT markers remains poorly understood.MethodsIn this study, we used the affinity purification and mass spectrometry coupled approach to identify the interactome of Slug. CoIP, GST-pulldown, ChIP, Re-ChIP, qPCR and Immunoblot were used to investigate the underlying mechanism of Slug-PRMT5-LSD1 complex. The role of PRMT5 and LSD1 in breast cancer progression was evaluated both in vivo and in vitro.ResultsHere we found that the transcription factor Slug associates with PRMT5 and LSD1 in a complex and facilitates the breast cancer invasion in vitro. Mechanistically, PRMT5 and LSD1 work with Slug to exert dual transcriptional activities to inhibit E-cadherin expression by PRMT5-catalyzed H4R3me2s and LSD1-mediated demethylation of H3K4me2 on the E-cadherin (CDH1) promoter, and activate vimentin (VIM) expression via PRMT5-driven H3R2me2s and LSD1-mediated removal of H3K9me2. Importantly, PRMT5 and LSD1 are coordinately expressed in breast cancer patients and pharmacologic perturbation of both PRMT5 and LSD1 shows a synergetic effect on the inhibition of breast tumor growth and metastasis in vivo.ConclusionsOur study suggests that PRMT5 and LSD1 function as a dual epigenetic modifier to promote Slug induced EMT program, suggesting that the inhibition of PRMT5 and LSD1 presents a potential therapeutic strategy against cancer metastasis.

Targeted Drug Delivery Biopolymers Effectively Inhibit Breast Tumor Growth and Prevent Doxorubicin-Induced Cardiotoxicity.

The anticancer agent doxorubicin(dox) has been widely used in the treatment of a variety of hematological malignancies and solid tumors. Despite doxorubicin’s efficiency in killing tumor cells, severe damage to healthy tissues, along with cardiotoxicity, limits its clinical use. To overcome these adverse side effects, improve patient safety, and enhance therapeutic efficacy, we have designed a thermally responsive biopolymer doxorubicin carrier that can be specifically targeted to tumor tissue by locally applying mild hyperthermia (41 °C). The developed drug vehicle is composed of the following: a cell penetrating peptide (SynB1) to promote tumor and cellular uptake; thermally responsive Elastin-like polypeptide (ELP); and the (6-maleimidocaproyl) hydrazone derivative of doxorubicin (DOXO-EMCH) containing a pH-sensitive hydrazone linker that releases doxorubicin in the acidic tumor environment. We used the in vivo imaging system, IVIS, to determine biodistribution of doxorubicin-delivered ELP in MDA-MB-231 xenografts in nude mice. Tumor bearing mice were treated with a single IV injection of 10 mg/kg doxorubicin equivalent dose with free doxorubicin, thermally responsive SynB1 ELP 1-DOXO, and a thermally nonresponsive control biopolymer, SynB1 ELP 2-DOXO. Following a 2 h treatment with hyperthermia, tumors showed a 2-fold higher uptake when treated with SynB1 ELP 1-DOXO compared to free doxorubicin. Accumulation of the thermally non-responsive control SynB1 ELP2 –DOXO was comparable to free doxorubicin, indicating that an increase in dox accumulation with ELP is due to aggregation in response to thermal targeting. Higher levels of SynB1 ELP1–DOXO and SynB1 ELP2 –DOXO with respect to free doxorubicin were observed in kidneys. Fluorescence intensity from hearts of animals treated with SynB1 ELP1–DOXO show a 5-fold decrease in accumulation of doxorubicin than the same dose of free doxorubicin. SynB1-ELP1-DOXO biopolymers demonstrated a 6-fold increase in tumor/heart ratio in comparison to free doxorubicin, indicating preferential accumulation of the drug in tumors. These results demonstrate that thermally targeted polymers are a promising therapy to enhance tumor targeting and uptake of anticancer drugs and to minimize free drug toxicity in healthy tissues, representing a great potential for clinical application.

Abstract P3-12-01: Regular aspirin use, breast tumor characteristics and long-term breast cancer survival

Abstract Background: Experimental evidence suggests that aspirin may inhibit breast tumor growth and reduce invasiveness of breast cancer cells. In large population-based studies (including our own), post-diagnostic aspirin use was associated with reduced breast cancer-specific and all-cause mortality. Here, we have extended our prior study with an additional 10 years of follow-up (3,785 more breast cancer cases) and investigation of the molecular underpinnings of the role of aspirin in breast cancer prognosis. Methods: Our study included 7,949 women diagnosed with stage I, II or III breast cancer from the Nurses’ Health Study (NHS) and NHSII. Cox proportional hazards regression was used to compute the multivariate hazard ratio (HR) for death, adjusting for tumor characteristics, treatment information and lifestyle factors. Post-diagnostic aspirin use was obtained at least 12 months after diagnosis and updated at every 2-year follow-up interval. We also evaluated the association between post-diagnostic aspirin use and survival according to tumor characteristics. Estrogen receptor (ER), insulin receptor (IR), PTEN, and Ki67 protein expression was evaluated by immunohistochemistry. PIK3CA mutation status was determined via polymerase chain reaction and pyrosequencing. In an exploratory analysis to identify functional enrichment of biologic pathways associated with long-term pre-diagnostic aspirin use (ever use of aspirin for ≥10 years with >2d/wk), we used a competitive gene-set testing procedure in a subgroup of cases with gene expression data (N=453). In this subset, we also computed the abundance of immune cell infiltration (B cell, CD4+ T cell, CD8+T cell, macrophage, neutrophil and dendritic cell) using Tumor IMmune Estimation Resource (TIMER). Results: During a median follow-up of 12 years after breast cancer diagnosis, we documented 2,502 deaths, including 1,373 from breast cancer. Compared with nonusers, women who regularly used aspirin after diagnosis had lower breast cancer-specific mortality: HR for 1, 2-5, 6-7 days of aspirin use per week were 0.65 (95% CI: 0.49, 0.86), 0.41 (95% CI: 0.29, 0.57) and 0.61 (95% CI: 0.47, 0.78) for, respectively (p-trend<0.0001). The association did not differ statistically by PTEN loss, PIK3CA mutation, or expression of IR (PI3K—AKT—mTOR), ER or Ki67 expression (proliferation pathways), or immune cell infiltration of the primary tumors (p-heterogeneity>0.05). Long-term regular aspirin use before diagnosis was associated with the downregulation of pathways related to inflammation (INF-α, INF-γ, and TNF-α), PI3K—AKT—mTOR signaling (mTOR and PI3K—AKT), and other proliferation (E2F and myc targets) in primary breast tumors and/or normal-adjacent tissues (FDR≤0.05). Long-term regular aspirin use prior to diagnosis was also associated with higher CD8+T cell (mean±SD=0.20±0.02 for non-users and 0.21±0.02 for users, p=0.01) and macrophage (mean±SD=0.05±0.03 for non-users and 0.06±0.02 for users, p=0.0002) relative abundance in primary tumors. Conclusion: Regular aspirin use after breast cancer diagnosis was associated with lower risk of breast cancer-specific and total mortality. The association between aspirin and mortality did not differ by molecular characteristics of primary tumors. Long-term aspirin use prior to breast cancer diagnosis was associated with downregulation of inflammatory and proliferation pathways and higher immune cell infiltration of CD8+ T cells and macrophages in breast tumors. Taken together, our large-scale population-based analysis with long-term of follow-up highlight the potential benefit of aspirin as a secondary prevention strategy across different tumor molecular characteristics. Citation Format: Cheng Peng, Michelle D. Holmes, Tengteng Wang, Alexandra Harris, Wendy Chen, Kristen Brantley, Yujing J Heng, Vanessa C. Bret-Mounet, Gabrielle M Baker, Bernard Rosner, Walter Willett, Rulla Tamimi, A. Heather Eliassen. Regular aspirin use, breast tumor characteristics and long-term breast cancer survival [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P3-12-01.

Abstract P3-12-03: Pre-diagnostic plasma 25-hydroxyvitamin D and mortality among women with breast cancer in the nurses’ health study cohorts

Abstract Background: Sufficient levels of vitamin D are critical for optimal health. 25-hydroxyvitamin D (25(OH)D) is an essential steroid hormone, derived from sun exposure, diet, and supplement use, that is hypothesized to influence breast cancer prognosis by inhibiting breast tumor growth. While this has been observed in laboratory-based studies, findings from observational studies are inconclusive. We examined the association between pre-diagnostic 25(OH)D concentrations and both breast cancer-specific and overall mortality to explore the potential effects of vitamin D, a modifiable exposure. Methods: We evaluated the association between pre-diagnostic plasma 25(OH)D concentrations and breast cancer-specific and total mortality among 1,300 women diagnosed with invasive, non-metastatic breast cancer enrolled in the Nurses’ Health Study 1 (NHS1) and Nurses’ Health Study 2 (NHS2) cohorts. In this analysis, women provided a blood sample from 1989-1990 and a second blood sample in 2000-2001 (NHS1) or a blood sample from 1996-1999 (NHS2). Women were diagnosed with breast cancer after blood collection up through 2016 (NHS1) and 2017 (NHS2) and censored at end of study (2016 NHS1; 2017 NHS2) or death, whichever came first. Cox proportional hazards models were used to estimate hazard ratios (HR) and 95% confidence intervals (95%CI) for mortality after adjustment for tumor characteristics, type of cancer treatment, and lifestyle factors. For women with two blood samples, most recent exposure status was used in the analyses. Results: A total of 396 total deaths and 145 breast cancer-specific deaths occurred during follow-up through 2017. We observed a non-statistically significant 25% lower breast cancer-specific mortality rate among those with the highest, versus lowest, quartile of pre-diagnostic 25(OH)D levels (HRQ4vsQ1 0.75, 95%CI 0.45-1.25; p for trend=0.32), that did not vary significantly by estrogen receptor status of the tumor (p for interaction=0.61). Women in the highest vs. lowest quartile of plasma 25(OH)D experienced a 31% lower overall mortality rate (HRQ4vsQ1 0.69, 95%CI 0.51-0.93; p for trend=0.03). In cause-specific mortality analyses among non-breast cancer deaths, non-statistically significant decreases were observed for other cancer deaths and cardiovascular disease deaths. Conclusion: Pre-diagnostic plasma 25(OH)D levels were not significantly associated with lower breast cancer-specific mortality among women with breast cancer. However, significant inverse associations were observed for total mortality, which supports a potential benefit of vitamin D among these women. Citation Format: Etienne Holder. Pre-diagnostic plasma 25-hydroxyvitamin D and mortality among women with breast cancer in the nurses’ health study cohorts [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P3-12-03.

Dietary Piperine Suppresses Obesity-Associated Breast Cancer Growth and Metastasis by Regulating the miR-181c-3p/PPARα Axis.

Adipocyte-derived leptin activates multiple oncogenic signaling, leading to breast cancer cell progression and metastasis. Hence, finding effective strategies to inhibit the oncogenic effects of leptin would provide a novel approach for disrupting obesity-associated breast cancer. In the current study, we explored the role of piperine, a major plant alkaloid from Piper nigrum (black pepper), against leptin-induced breast cancer. Piperine treatment significantly inhibited leptin-induced breast cancer cell proliferation, colony formation, migration, and invasion. We found that piperine downregulated the expression of PPARα, a predicted target of miR-181c-3p. Mechanistically, piperine potentiates miR-181c-3p-mediated anticancer potential in leptin-induced breast cancer cells. Interestingly, the knockdown of PPARα reduced the proliferative potential of leptin-induced breast cancer cells. Further, oral administration of piperine inhibited breast tumor growth in diet-induced obese mice, accompanied by the upregulation of miR-181c-3p and downregulation of PPARα expression. Together, piperine represents a potential candidate for further development as an anticancer agent for treating obesity-associated breast cancer.

ALDH1A1 Activity in Tumor-Initiating Cells Remodels Myeloid-Derived Suppressor Cells to Promote Breast Cancer Progression.

Tumor-initiating cells (TIC) are associated with tumor initiation, growth, metastasis, and recurrence. Aldehyde dehydrogenase 1A1 (ALDH1A1) is a TIC marker in many cancers, including breast cancer. However, the molecular mechanisms underlying ALDH1A1 functions in solid tumors remain largely unknown. Here we demonstrate that ALDH1A1 enzymatic activity facilitates breast tumor growth. Mechanistically, ALDH1A1 decreased the intracellular pH in breast cancer cells to promote phosphorylation of TAK1, activate NFκB signaling, and increase the secretion of GM-CSF, which led to myeloid-derived suppressor cell expansion and immunosuppression. Furthermore, the ALDH1A1 inhibitor disulfiram and chemotherapeutic agent gemcitabine cooperatively inhibited breast tumor growth and tumorigenesis by purging ALDH+ TICs and activating T-cell immunity. These findings elucidate how active ALDH1A1 modulates the immune system to promote tumor development, highlighting new therapeutic strategies for malignant breast cancer. SIGNIFICANCE: ALDH1A1 enzyme activity induces MDSC expansion and triggers a procancer immune microenvironment to facilitate breast cancer progression, providing a novel therapeutic vulnerability in this disease.

Nuclear receptor 4A1 (NR4A1) antagonists target paraspeckle component 1 (PSPC1) in cancer cells.

Paraspeckles compound 1 (PSPC1) is a multifunctional protein that plays an important role in cancer cells, where PSPC1 is a master regulator of pro-oncogenic responses that includes activation of TGFβ (TGFβ1),TGFβ-dependent EMT, and metastasis. The pro-oncogenic activities of PSPC1 closely resembled those observed for the orphan nuclear receptor 4A1 (NR4A1, Nur77) and knockdown of NR4A1 decreased expression of PSPC1 in MDA-MB-231 breast, H1299 lung, and SNU449 liver cancer cells. Similar results were observed in these same cell lines after treatment with bisindole-derived (CDIMs) NR4A1 antagonists. Moreover, PSPC1-dependent regulation of TGFβ, genes associated with cancer stem cells and epithelial to mesenchymal transition (EMT) were also downregulated after NR4A1 silencing or treatment of breast, lung, and liver cancer cells with CDIM/NR4A1 antagonists. Results of chromatin immunoprecipitation (ChIP) assays suggest that NR4A1 regulates PSPC1 through interaction with an NBRE sequence in the PSPC1 gene promoter. These results coupled with in vivo studies showing that NR4A1 antagonists inhibit breast tumor growth and downregulate PSPC1 in tumors indicate that the pro-oncogenic nuclear PSPC1 factor can be targeted by CDIM/NR4A1 antagonists.

Transcriptome study of oleanolic acid in the inhibition of breast tumor growth based on high-throughput sequencing.

The function of oleanolic acid (OA) in various types of cancer has been reported frequently, especially for breast cancer. However, the regulation of breast tumor growth in response to OA treatment has not been studied in depth. Here, we first explored the effect of OA treatment on breast tumors in vitro and in vivo and then used RNA-seq technology to study the effect and molecular mechanism of OA treatment of MCF-7 cells, particularly at the level of functional genomics. The results showed that 40 μM OA treatment could significantly inhibit the proliferation and induce the apoptosis of MCF-7 cells. Through analysis of RNA sequencing data quality and differentially expressed genes (DEGs), 67 significantly downregulated genes and 260 significantly upregulated genes were identified to be involved in OA treatment of MCF-7 cells. Among these genes, 43 unique DEGs were enriched in several signaling pathways and Gene Ontology terms, such as p53 signaling pathway, TNF signaling pathway and mTOR signaling pathway. Six downregulated genes, including THBS1, EDN1, CACNG4, CCN2, AXIN2 and BMP4, as well as six upregulated genes, including ATF4, SERPINE1, SESN2, PPARGC1A, EGR1 and JAG1, were selected as target genes in response to OA treatment. The inhibitory effect of OA on breast cancer was also found in the following mouse experiments. Our study provides evidence and molecular support for the treatment of breast cancer with OA.