Target In Breast Cancer Research Articles

Targeting breast cancer resistance protein (BCRP/ABCG2): Functional inhibitors and expression modulators

The primary source of failure of cancer therapies is multidrug resistance (MDR), which can be caused by different mechanisms, including the overexpression of ABC transporters in cancer cells. Among the 48 human ABC proteins, the breast cancer resistance protein (BCRP/ABCG2) has been described as a pivotal player in cancer resistance. The use of functional inhibitors and expression modulators is a promising strategy to overcome the MDR caused by ABCG2. Despite the lack of clinical trials using ABCG2 inhibitors, many compounds have already been discovered. This review presents an overview about various ABCG2 inhibitors that have been identified, discussing some chemical aspects and the main experimental methods used to identify and characterize the mechanisms of new inhibitors. In addition, some biological requirements to pursue preclinical tests are described. Finally, we discuss the potential use of ABCG2 inhibitors in cancer stem cells (CSC) for improving the objective response rate and the mechanism of ABCG2 modulators at transcriptional and protein expression levels.

Mitigating off-target distribution and enhancing cytotoxicity in breast cancer cells with alpha-ketoglutaric acid-modified Fe/Mg-CA nanoparticles

Abstract Purpose In this work, pH-sensitive alpha-ketoglutaric acid-modified Fe/Mg-carbonate apatite (α-KAM-Fe/Mg-CA) NPs were introduced and found to be capable of promoting the selective delivery of cancer-killing drug doxorubicin (DOX) in breast cancer cells, while simultaneously mitigating DOX toxicity on normal cells. Methods As part of the characterization and evaluation of α-KAM-Fe/Mg-CA NPs to target breast cancer cells, a series of assessments were performed, which included size measurements, morphological analysis, FTIR, cytotoxicity assessment, hemolysis, drug binding, cellular uptake, and pH-responsive drug release tests. Liquid chromatography-mass spectrometry was used to conduct the protein corona analysis of α-KAM-Fe/Mg-CA using 10% FBS (fetal bovine serum) and mice plasma. Furthermore, to investigate the distribution of DOX-loaded α-KAM-Fe/Mg-CA NPs in major tissues and the tumor, a biodistribution investigation was conducted in mammary tumor-induced Balb/c mouse models 24 h after the intravenous administration of DOX-loaded α-KAM-Fe/Mg-CA NPs. Results The in vitro pH-dependent release of DOX over time demonstrated that α-KAM-Fe/Mg-CA NPs were pH-responsive and degraded rapidly at acidic pH levels. When compared to free DOX, the DOX-loaded α-KAM-Fe/Mg-CA NPs demonstrated a potent antiproliferative effect on breast cancer cells. Confocal microscopy confirmed the effective internalization of DOX-loaded α-KAM-Fe/Mg-CA NPs in breast cancer cells. The protein corona analysis revealed an affinity for dysopsonins (serum albumin, apolipoproteins) and transport proteins that may assist in extending their blood circulation period. Furthermore, biodistribution data of DOX-loaded α-KAM-Fe/Mg-CA NPs in the mammary tumor-induced Balb/c mouse model indicated extended circulation in the bloodstream, reduced non-target distribution in major tissues, and increased drug accumulation in the tumor. Conclusion The results obtained suggest that α-KAM-Fe/Mg-CA NPs may emerge as a prospective candidate for delivering therapeutic cargos to treat malignant mammary tumors.

Pyrimidine-5-carbonitrile based potential anticancer agents as apoptosis inducers through PI3K/AKT axis inhibition in leukaemia K562

A novel series of 4-(4-Methoxyphenyl)-2-(methylthio)pyrimidine-5-carbonitrile was developed linked to an aromatic moiety via N-containing bridge and then evaluated for their cytotoxic activity against MCF-7 and K562 cell lines. Seven compounds exhibited the highest activity against both cell lines where compounds 4d and 7f were the most active against K562 cell line. Exploring their molecular mechanisms by enzyme inhibition assay on PI3Kδ/γ and AKT-1 showed that compound 7f was promising more than 4d with IC50 = 6.99 ± 0.36, 4.01 ± 0.55, and 3.36 ± 0.17 uM, respectively. Also, flowcytometric analysis revealed that 7f caused cell cycle arrest at S-phase followed by caspase 3 dependent apoptosis induction. Mechanistically, compound 7f proved to modulate the expression of PI3K, p-PI3K, AKT, p-AKT, Cyclin D1, and NFΚβ. Furthermore, in-vivo toxicity study indicated good safety profile for 7f. These findings suggest that the trimethoxy derivative 7f has strong potential as a multi-acting inhibitor on PI3K/AKT axis targeting breast cancer and leukaemia.

Biomarkers to target and silence stemness of breast cancer stem cell model: silencing MDR1 by siRNA

Abstract Objectives Aim of the study was to reveal new biomarker genes to target breast cancer stem-like cells (BCSC-like) and then sensitize BCSC-like cells to chemotherapy by silencing MDR1 gene found to be the most suitable target. Methods Drug resistance associated genes were screened by cDNA microarray to unveil biomarker genes in drug resistance breast cancer model cells. Drug resistance was then reversed by silencing MDR1 gene in BCSC-like cells. The effect of silencing was monitored by real-time cell proliferation analysis. Differential expressions of MDR1, ALDH1A3, EGFR and BAG4 genes were identified by real-time PCR. P-glycoprotein (P-gp) expression level and its activity were investigated by Western blot and flow cytometry measurements, respectively. Results 16 new biomarker genes were identified upon gene expression analysis by cDNA microarray. MDR1 gene was selected as the most potent target gene and silencing of it caused down-regulation of MDR1, ALDH1A3, EGFR, BAG4 expression and P-glycoprotein activity and expression in BCSC-like cells. At the end, silenced BCSC-like cells were found to be more responsive to paclitaxel therapy. Conclusions In conclusion, siMDR1 silencing is an effective way to reverse multidrug resistance and malignancy. New biomarker genes revealed in this study require to be investigated to target stemness of BC.

A universal strategy of glyconanoparticle preparation using a bifunctional linker for lectin sensing and cell imaging.

Glyconanoparticles (G-NPs), biofunctional nanomaterials that can fully combine the unique properties of nanoparticles (NPs) with the bioactivities of carbohydrates, have become an appealing nanoplatform in analytical chemistry and biomedical research. However, there is currently a lack of an efficient and universal method for facile immobilization of reducing carbohydrates on NPs while maintaining their structure integrity, greatly limiting the preparation and application of G-NPs. Herein, a new and universal strategy for preparing carbohydrate-functionalized gold nanoclusters (Au NCs) was developed by using S-(3-(methoxyamino)propyl) thioacetate (MPTA) as a new bifunctional linker. MPTA with an N-methoxyamine group (-NHOMe) and a thioacetyl group (-SAc) was synthesized by a two-step strategy and then grafted onto Au NCs by an efficient click reaction. Subsequently, reducing carbohydrates could be readily immobilized onto MPTA-functionalized Au NCs (MPTA-Au NCs) by a reducing end ring-closure reaction under mild conditions. The obtained G-NPs showed average size of 1.9 ± 0.42 nm and strong fluorescence at 610 nm. Carbohydrates grafted on G-NPs still retained their structure integrity and specific recognition ability toward their receptor proteins. Notably, the affinity between G-NPs and proteins was increased by 1300 times compared with free carbohydrates with an association constant of (1.47 ± 0.356) × 106 M−1. The prepared fluorescent G-NPs were also successfully applied to lectin sensing and targeted breast cancer cell imaging with good performance. These results indicated that the intact immobilization of reducing carbohydrates (whether naturally or chemically accessed) on NPs could be easily achieved using MPTA, providing a simple, efficient, and universal strategy for G-NP preparation.Graphical abstract Supplementary InformationThe online version contains supplementary material available at 10.1007/s00604-022-05220-w.

An update on nanoparticle usage in breast cancer imaging

AbstractIntroductionThe objective of this literature review is to highlight both the recent promises of Nanoparticles (NPs) for breast imaging and potential solutions to the challenges that arise with using different types of NPs.MethodsWe searched for key terms on PubMed including NPs, MRI, CT, and contrast agent (CA). We selected 57 articles published from 2012 to 2020 and discussed NP such as nanocarriers, titanium oxide tubes, and carbon nanotubes, as well as gold, gold‐silver alloy, silver, magnetic, gadolinium dicarbide, allochroic, and calcium phosphate.ResultsWhile early studies on NPs investigated their ability to target breast cancer stem cells, as well as enable the visualization of metastasis and deliver therapeutic agents with minimal toxicity, newer particles demonstrate a greater multifunctionality of NPs than previously thought. In this review, we use existing research to demonstrate the surprisingly broad range of applications of NPs with regards to breast cancer and imaging, which includes activating immune cells, identifying protein signatures for early‐stage breast cancer, and ablating tumors through a laser irradiation‐triggered photothermal effect.ConclusionWith the multifunctionality of NPs in mind and the challenges associated with imaging breast tissue, specifically mammographically dense breast tissue, the current knowledge of NPs should be reviewed for potential use in mammograms.

Exploration of novel VEGFR2 tyrosine kinase inhibitors via design and synthesis of new alkylated indolyl-triazole Schiff bases for targeting breast cancer

According to the World Health Organization (WHO) statistics: In 2020, there were 2.3 million women diagnosed with breast cancer and 685,000 deaths globally. Therefore, searching for new leads for fighting this type of cancer is necessary. VEGFR-2 kinase plays a crucial role in the proliferation, migration, and survival of breast cancer cells so, identifying novel inhibitors for VEGFR-2 could be effective in breast cancer treatment. Accordingly, novel heterocyclic compounds containing indole, 1,2,4-triazole, and glycosyl or allyl moieties were synthesized. The synthesized compounds were evaluated for their cytotoxic and apoptotic activities towards breast cancer cell lines (MCF7). In this regard, compounds 6, 17, and 18 exhibited promising cytotoxic activity against MCF-7 cells with IC50 values of 3.06, 1.18, and 3.02 μM compared to Sorafenib (IC50 = 2.13 μM). Interestingly, among the identified lead molecules, compound 17 displayed remarkable VEGFR2 inhibition activity with IC50 value of 19.8 nM compared to Sunitinib (IC50 = 75 nM) and Sorafenib (IC50 = 30 nM). Moreover, it is significantly stimulated apoptotic breast cancer cell death; it induced apoptosis by 17.4 %, arresting the cell cycle phases at G1 and S-phases. Additionally, in vivo (Xenograft model) study validated the anticancer activity of the hit compound 17, which showed a tumor inhibition ratio of 54.2 % compared to 5-FU (49.5%) with an improvement of hematological and biochemical parameters. The results disclosed that the identified hit compound 17 is validated for impeding cell proliferation and migration through apoptosis activation and VEGFR2 inhibition.

“Double hit” strategy: Removal of sialic acid from the dendritic cell surface and loading with CD44+/CD24-/low cell lysate inhibits tumor growth and metastasis by targeting breast cancer stem cells

Cancer stem cells (CSCs), which represent the root cause of resistance to conventional treatments, recurrence, and metastasis, constitute the critical point of failure in cancer treatments. Targeting CSCs with dendritic cell (DC)-based vaccines have been an effective strategy, but sialic acids on the surface of DCs limit the interaction with loaded antigens. We hypothesized that removal of sialic acid moieties on immature DCs (iDCs) could significantly affect DC-CSC-antigen loading, thereby leading to DC maturation and improving immune recognition and activity. The lysate of CD44+/CD24-/low breast CSCs (BCSCs) was pulsed with sialidase-treated DCs to obtain mature dendritic cells (mDCs). The roles of cytoskeletal elements in antigen uptake and dendritic cell maturation were determined by immunofluorescence staining, flow cytometry, and cytokine measurement, respectively. To test the efficacy of the vaccine in vivo, CSCs tumor-bearing mice were immunized with iDC or mDC. Pulsing DCs with antigen increased the expression levels of actin, gelsolin, talin, WASp, and Arp2, especially in podosome-like regions. Compared with iDCs, mDCs expressed high levels of CD40, CD80, CD86 costimulatory molecules and increased IL-12 production. Vaccination with mDC: i) increased CD8+ and CD4 + T-cell numbers, ii) prevented tumor growth with anti-mitotic activity and apoptotic induction, iii) suppressed metastasis by decreasing Snail, Slug, and Twist expressions. This study reveals for the first time that sialic acid removal and loading with CSC antigens induces significant molecular, morphological, and functional changes in DCs and that this new DC identity may be considered for future combined immunotherapy strategies against breast tumors.

New Advances in Targeted Therapy of HER2-Negative Breast Cancer.

Breast cancer has an extremely high incidence in women, and its morbidity and mortality rank first among female tumors. With the increasing development of molecular biology and genomics, molecular targeted therapy has become one of the most active areas in breast cancer treatment research and has also achieved remarkable achievements. However, molecular targeted therapy is mainly aimed at HER2-positive breast cancer and has not yet achieved satisfactory curative effect on HER2-negative breast cancer. This article describes the potential targets that may be used for breast cancer treatment from the aspects of PI3K/AKT signaling pathway, DDR, angiogenesis, the cell cycle, breast cancer stem cells, etc., and explores possible inhibitors for the treatment of HER2-negative breast cancer, such as PI3K inhibitors, AKT inhibitors and m-TOR inhibitors that inhibit the PI3K/AKT signaling pathway, small molecule tyrosine kinase inhibitors that restrain angiogenesis, CDK inhibitors, aurora kinase inhibitors and HDAC inhibitors that block cell cycle, as well as the drugs targeting breast cancer stem cells which have been a hit, aiming to provide a new idea and strategy for the treatment of HER2-negative breast cancer.

Metabolic syndrome and breast cancer survivors: a follow-up analysis after completion of chemotherapy

BackgroundWe previously reported that (neo)adjuvant chemotherapy adversely altered metabolic syndrome (MetS) components, body composition, and related biomarkers after a 12 to 18-week chemotherapy treatment course in women. Here, we sought to determine whether these measures worsened within 4–5 years post-chemotherapy among the same sample of early stage breast cancer survivors.MethodsTwenty-eight breast cancer survivors were reassessed within 4–5 years post-chemotherapy. Participants were tested for MetS, lipid profile (total cholesterol; TC, low-density lipoprotein cholesterol; LDL-C), glucose metabolism (insulin, homeostatic model- insulin resistance; HOMA-IR, glycosylated hemoglobin; HbA1c), inflammation (C-reactive protein; CRP) and body composition (body weight; BW, percent body fat; BF, fat mass; FM) during follow-up physical exams. A comparison of measurements between post-chemotherapy and follow-up periods was performed using repeated measures analysis of covariance.ResultsMost study patients were Caucasian (44%) or Hispanic (30%) with a mean age of 48.2 years. Average time from completion of chemotherapy was 4.75 years. At follow-up, MetS components significantly increased (p < 0.01) compared with the post chemotherapy assessment. Additionally, BF, FM, lipids (TC, LDL), glucose metabolism (HOMA-IR, insulin, HbA1c), and inflammation (CRP) significantly increased (p < 0.01). Notably BW significantly increased; mean weight gain after chemotherapy was 6.1 kg and increased an additional 8.2% at follow-up (p < 0.01).ConclusionMetS components, body composition, and biomarkers continued to worsen within 4–5 years post-chemotherapy in breast cancer survivors. Energy balance interventions should target breast cancer patients to reduce the exacerbation of MetS.

Microenvironment-driven sequential ferroptosis, photodynamic therapy, and chemotherapy for targeted breast cancer therapy by a cancer-cell-membrane-coated nanoscale metal-organic framework

Designing and developing nanomedicine based on the tumor microenvironment (TME) for effective cancer treatment is highly desirable. In this work, polyvinyl pyrrolidone (PVP) dispersed nanoscale metal-organic framework (NMOF) of Fe-TCPP (TCPP = tetrakis (4-carboxyphenyl) porphyrin) loaded with hypoxia-activable prodrug tirapazamine (TPZ) and coated by the cancer cell membrane (CM) is constructed (the formed nanocomposite denoted as PFTT@CM). Due to the functionalization with the homologous cancer cell membrane, PFTT@CM is camouflaged to evade the immune clearance and preferentially accumulates at the tumor site. Once internalized by cancer cells, PFTT@CM is activated by the TME through redox reaction and Fenton reaction between Fe3+ in nano-platform and endogenous glutathione (GSH) and hydrogen peroxide (H2O2) to promote GSH exhausting as well as •OH and O2 production, which triggers ferroptosis and dramatically enhances photodynamic therapy (PDT) efficacy. Subsequently, the PDT process mediated by TCPP and light would consume oxygen and aggravate tumor hypoxia to further activate the prodrug TPZ for cancer chemotherapy. As a consequence, the TME-driven PFTT@CM nano-platform not only demonstrated its TME modulation ability but also showed a sequential synergistic therapy, which eventually inhibited the cancer cell proliferation. This multimodal nano-platform is expected to shed light on the design of TME-activatable reaction to reinforce the synergistic therapeutic outcome and facilitate the development of effective cancer nanomedicine.

Untargeted LC-MS/MS Metabolomics Study on the MCF-7 Cell Line in the Presence of Valproic Acid.

To target breast cancer (BC), epigenetic modulation could be a promising therapy strategy due to its role in the genesis, growth, and metastases of BC. Valproic acid (VPA) is a well-known histone deacetylase inhibitor (HDACi), which due to its epigenetic focus needs to be studied in depth to understand the effects it might elicit in BC cells. The aim of this work is to contribute to exploring the complete pharmacological mechanism of VPA in killing cancer cells using MCF-7. LC-MS/MS metabolomics studies were applied to MCF-7 treated with VPA. The results show that VPA promote cell death by altering metabolic pathways principally pentose phosphate pathway (PPP) and 2′deoxy-α-D-ribose-1-phosphate degradation related with metabolites that decrease cell proliferation and cell growth, interfere with energy sources and enhance reactive oxygen species (ROS) levels. We even suggest that mechanisms such as ferropoptosis could be involved due to deregulation of L-cysteine. These results suggest that VPA has different pharmacological mechanisms in killing cancer cells including apoptotic and nonapoptotic mechanisms, and due to the broad impact that HDACis have in cells, metabolomic approaches are a great source of information to generate new insights for this type of molecule.

N-acetyl-d-glucosamine decorated nano-lipid-based carriers as theranostics module for targeted anti-cancer drug delivery

The present investigation was carried out to engineer N-acetyl-d-glucosamine (NADG) coupled quercetin-loaded (Q) nano-lipid-based carriers (NADG-Q-NLBCs) to target breast cancer. The constructed nano-bioconjugate was characterized by various techniques, and the in vitro drug release profiles were examined using zero and first-order kinetic models. The characterization data confirmed the morphology, size, charge distribution, crystallinity, and chemical interactions among the various moieties of the nano-bioconjugate. Further, the synthesized NADG-Q-NLBCs were applied to target the human breast cancer cells (MCF-7), which interestingly showed a more cytotoxic effect compared to the lone NLBCs and free Quercetin. The flow cytometry study confirmed NADG-Q-NLBCs induced apoptosis in MCF-7 cells in a targeted manner. The percentage of early apoptotic cells was found to be 25% in the case of NADG-Q-NLBCs, which is almost 2.5 times higher than the Q-NLBCs. However, the number of viable cells reached the maximum when treated with NLBCs. The present investigation suggests that the constructed nano-bioconjugate could be a capable carrier of drugs with sustained pharmacokinetics and improved physicochemical properties.

Design, synthesis, and molecular modeling of coumarin derivatives as MDM2 inhibitors targeting breast cancer.

The coumarin ring was used as a central scaffold that was substituted with a variety of bioactive functional groups, for designing and synthesizing novel MDM2 inhibitors targeting breast cancer. The synthesized derivatives, 3c, 3d, 3g, 7b, 7c and 8 with IC50s ranging from 9.4 to 9.9µM were evaluated for their safety on MCF10a normal breast cell line. The compounds showed selectivity indices of 2.15, 3.85, 2.75, 1.38, 3.72 and 5.20 respectively. 7c was selected for further investigation, the compound was capable of down-regulating MDM2 and the anti-apoptosis proteins Bcl-2 and Bcl-xL, up-regulating the level of p53 and the pro-apoptosis protein BAX, causing cell cycle arrest at G2/M phase and activating Caspase-9 to induce apoptosis. Molecular docking study revealed the capability of derivative 7c to interact with the key amino acids in p53 binding pocket of MDM2 protein. Moreover, the physicochemical properties of compound 7c were studied in silico.

Co-axial electrospraying of injectable multi-cancer drugs nanocapsules with polymer shells for targeting aggressive breast cancers

BackgroundThere is growing potential for nanocarrier-based drug delivery in cancer. However, an incomplete understanding of nano–bio interactions and the challenges regarding processing and fabrication in scale-up engineering techniques, controls over drug release, efficacy, and cytotoxicity to the human cell are the major challenges for its clinical success. The purpose of the study was to develop an electrospraying processing of injectable nanonized encapsulated chemotherapeutics to target primary and metastatic breast cancer tumor microenvironment for precise and controlled delivery.ResultsA novel coaxial electrospraying of multiple cancer drugs (paclitaxel and GW2580) as core and polycaprolactam (PCL) as the shell has been developed to produce multi-cancer drug nanocapsules. Using electrospraying process, we have successfully made nanocapsules containing paclitaxel to target breast cancer cells and GW2580, a colony-stimulating factor 1 receptor (CSF1R) inhibitor to target CSF1R+ myeloid cells in the tumor microenvironments (TME). The UV–vis drug release test for 14 days shows a prolonged and sustained release pattern of both the drugs. In vitro and in vivo results showed the effects of nanocapsules containing multiple drugs in controlling the growth of tumor cells and increased survival of the animal bearing breast cancers.ConclusionNanonized multi-cancer drugs were encapsulated in a PCL shell. The drug doses ratio and the polymer-to-drug ratio were controlled by engineered process parameters. The studies showed the importance of making nanocapsules containing nanocrystals of multiple drugs, which will pave the way of making multiple drug combinations in a controlled manner and capsules can be designed for sustained release of the drugs after accumulation into the TME. TME-directed therapy can be a norm in future cancer treatment strategies. These injectable nanocapsules will allow cancer site-specific precision and controlled delivery to cure primary and metastatic breast cancer and to overcome the chemotherapy resistance.

Smart Lipid-Polysaccharide Nanoparticles for Targeted Delivery of Doxorubicin to Breast Cancer Cells.

In this study, actively-targeted (CD44-receptors) and dual stimuli (pH/redox)-responsive lipid–polymer nanoparticles were proposed as a delivery vehicle of doxorubicin hydrochloride in triple negative breast cancer cell lines. A phosphatidylcholine lipid film was hydrated with a solution of oxidized hyaluronic acid and doxorubicin, chosen as model drug, followed by a crosslinking reaction with cystamine hydrochloride. The obtained spherical nanoparticles (mean diameter of 30 nm) were found to be efficiently internalized in cancer cells by a receptor-mediated endocytosis process, and to modulate the drug release depending on the pH and redox potential of the surrounding medium. In vitro cytotoxicity assays demonstrated the safety and efficacy of the nanoparticles in enhancing the cytotoxic effect of the free anticancer drug, with the IC50 values being reduced by two and three times in MDA-MB-468 and MDA-MB-231, respectively. The combination of self-assembled phospholipid molecules with a polysaccharide counterpart acting as receptor ligand, and stimuli-responsive chemical moieties, was carried out on smart multifunctional nanoparticles able to actively target breast cancer cells and improve the in vitro anticancer activity of doxorubicin.

New and Emerging Targeted Therapies for Advanced Breast Cancer.

In the United States, breast cancer is among the most frequently diagnosed cancers in women. Breast cancer is classified into four major subtypes: human epidermal growth factor receptor 2 (HER2), Luminal-A, Luminal-B, and Basal-like or triple-negative, based on histopathological criteria including the expression of hormone receptors (estrogen receptor and/or progesterone receptor) and/or HER2. Primary breast cancer treatments can include surgery, radiation therapy, systemic chemotherapy, endocrine therapy, and/or targeted therapy. Endocrine therapy has been shown to be effective in hormone receptor-positive breast cancers and is a common choice for adjuvant therapy. However, due to the aggressive nature of triple-negative breast cancer, targeted therapy is becoming a noteworthy area of research in the search for non-endocrine-targets in breast cancer. In addition to HER2-targeted therapy, other emerging therapies include immunotherapy and targeted therapy against critical checkpoints and/or pathways in cell growth. This review summarizes novel targeted breast cancer treatments and explores the possible implications of combination therapy.

Screening and Identification of Human Endogenous Retrovirus-K mRNAs for Breast Cancer Through Integrative Analysis of Multiple Datasets.

ObjectiveHuman endogenous retroviruses (HERVs) make up 8% of the human genome. HERVs are biologically active elements related to multiple diseases. HERV-K, a subfamily of HERVs, has been associated with certain types of cancer and suggested as an immunologic target in some tumors. The expression levels of HERV-K in breast cancer (BCa) have been studied as biomarkers and immunologic therapeutic targets. However, HERV-K has multiple copies in the human genome, and few studies determined the transcriptional profile of HERV-K copies across the human genome for BCa.MethodsNinety-one HERV-K indexes with entire proviral sequences were used as the reference database. Nine raw sequencing datasets with 243 BCa and 137 control samples were mapped to this database by Salmon software. The differential proviral expression across several groups was analyzed by DESeq2 software.ResultsFirst, the clustering of each dataset demonstrated that these 91 HERV-K proviruses could well cluster the BCa and control samples when the normal controls were normal cells or healthy donor tissues. Second, several common HERV-K proviruses that are closely related with BCa risk were significantly differentially expressed (padj < 0.05 and absolute log2FC > 1.5) in the tissues and cell lines. Additionally, almost all the HERV-K proviruses had higher expression in BCa tissue than in healthy donor tissue. Notably, we first found the expression of 17p13.1 provirus that located with TP53 should regulate TP53 expression in ER+ and HER2+ BCa.ConclusionThe expression profiling of these 91 HERV-K proviruses can be used as biomarkers to distinguish individuals with BCa and healthy controls. Some proviruses, especially 17p13.1, were strongly associated with BCa risk. The results suggest that HERV-K expression profiles may be appropriate biomarkers and targets for BCa.

Abstract P1-05-08: Targeting tumor heterogeneity and breast cancer metastasis through the metastatic microenvironment mediated epigenetic reprogramming

Abstract Targeting tumor heterogeneity and breast cancer metastasis through the metastaticmicroenvironment mediated epigenetic reprogrammingLi Yang and Jae Young So, Nicolas Skrypek, Anand Merchant, George Nelson, Howard H.Yang, Maxwell P. LeeNational Cancer Institute, National Institutes of Health, Bethesda, MD 20892Current cancer treatments are largely based on the genetic characterization of primarytumors and are ineffective for metastatic disease. There is lack of mechanistic understanding ashow cancer cells are selected and evolved to establish distant metastatic colonies. In addition,tumor heterogeneity and biomarker identification remain to be some of the most difficultchallenges in cancer treatment. In the current study, we discovered an increased DNAmethyltransferase 3B (DNMT3B) in metastatic nodules and in tumor cells with epithelia tomesenchymal (EMT) phenotype. Of great interest, high levels of DNMT3B were correlated withpoor clinical outcomes in multiple human breast cancer datasets. Mechanistically, DNMT3Balters multiple pathways including STAT3, NFkappaB, PI3K/Akt, beta-catenin, Notch signalingas well as EMT, which are critical for cancer cell survival, apoptosis, proliferation, invasion, andcolonization. We further identified PGE2 and IL6 as critical inflammatory mediators inDNMT3B induction. Importantly, targeting IL6 or PGE2 production reduced DNMT3Bexpression and improved chemo treatment or PD1 immune therapy. Furthermore, perioperative(surgical removal of primary tumors) targeting DNMT3B in combination with chemotherapymarkedly suppressed tumor recurrence and metastasis in preclinical mouse models for triplenegative breast cancer. Our studies identify DNMT3B as an important mechanism fortranscription regulation in tumor heterogeneity that is critical for tumor invasion and metastasis,thus suggesting a validated target for treating metastatic disease. Citation Format: Li Yang, Jae Young So, Nicolas Skrypek, Anand Merchant, George Nelson, Howard Yang, Maxwell Lee. Targeting tumor heterogeneity and breast cancer metastasis through the metastatic microenvironment mediated epigenetic reprogramming [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 P1-05-08.

Abstract PD3-08: Novel cancer stem cell inhibitor 108600 modulates tumor immunomicroenvironment of triple negative breast cancer (TNBC)

Abstract 108600, a novel CK2/DYRK1/TNIK inhibitor, targets and inhibits cancer stem cells (CSC) in triple negative breast cancer (TNBC), inhibiting tumor growth and metastases in patient-derived xenograft models. CSC’s have been shown to promote immune evasion of several types of cancer. Specifically, over-expression of CK2 has been shown to promote intratumoral recruitment of myeloid derived suppressor cells. We investigated the effects of 108600 treatment on the immune microenvironment of triple negative breast cancer since targeting CSC’s could promote favorable anti-tumor immune responses and mechanistically contribute to tumor growth inhibition. Methods:C57BL/6 mice bearing murine triple negative E0771 tumors were generated by orthotopic injection and treated either with vehicle control or 108600 for 5 days. Tumor growth was assessed by daily caliper measurement All tumors were recovered at endpoint, and processed for RNA sequencing, flow cytometry or Western blot analysis. Results:108600 treatment significantly inhibited growth of E0771 tumors in vivo, demonstrating for the first time efficacy of 108600 against TNBC in immunocompetent models. 108600 treatment decreased intratumoral phosphorylation and expression of 108600 targets AKT1 (Ser 129) and Cyclin D1, which are substrates of CK2α and Dyrk1, respectively. To further explore nature of transcriptional and signaling pathways affected by 108600 treatment in vivo, RNA sequencing was performed. DGE and subsequent annotation analysis showed that various immune (GO:0006955, GO) and inflammatory (GO:0006954, GO) signaling pathways associated with Stat1 (mmu04062, KEGG) and IFNγ (mmu04060, KEGG) were down-regulated in 108600-treated E0771 tumors. GSEA (Gene set enrichment analysis) indicated that the regulatory T cell (Treg) population (GSE42021, GSE40685) was suppressed by 108600 treatment. To validate these findings, tumor infiltrating leukocytes (TIL) were isolated from vehicle or 108600-treated tumors and analyzed by flow cytometry. CD4, CD25, and FOXP3 expression was used to gate the Treg population. The Treg population was significantly suppressed by 108600 treatment, confirming the RNA sequencing results. 108600 also likely regulates expression of immunomodulatory molecules in TNBC tumor cells since 108600 treatment increased PD-L1 surface expression on E0771 cells in vitro. Conclusions:Our study supports 108600, an inhibitor that targets breast cancer stem cells, as a modulator of the immune microenvironment of TNBC. 108600 suppresses the Treg population among the TIL population and increases tumoral expression of PD-L1. Tregs have been associated with disease progression and metastases of TNBC. Further studies are ongoing to clarify the functional consequences of these changes in the setting of tumor growth inhibition. Synergies between 108600 and checkpoint inhibition are also under investigation and could lead to novel combination therapies for TNBC. Citation Format: Katsutoshi Sato, Stacey J. Baker, E. P. Reddy, Hanna Y. Irie. Novel cancer stem cell inhibitor 108600 modulates tumor immunomicroenvironment of triple negative breast cancer (TNBC) [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 PD3-08.