Breast Cancer Phenotype Research Articles

Prognostic Significance of Combining Cytokeratin-19, E-Cadherin and Ki-67 Analysis in Triple-Negative Breast Cancer with Basal-Like and Non-Basal-Like Phenotype

Triple-negative breast cancer (TNBC) is known to have the worst outcome compared to the other forms of breast cancer. Moreover, molecular markers identified basal-like breast cancer (BLBC) phenotypes to be also related to a worse prognosis. In this study, we evaluated by immunohistochemistry (IHC) the prognostic significance of combining Cytokeratin-19 (CK19), E-cadherin, and Ki-67 tissue expression in triple-negative breast cancer (TNBC) cases presenting a basal-like (BLBC) or a non-basal-like (n-BLBC) phenotype to improve the selection and the monitoring of BC patients with a more aggressive outcome. Herein, when compared to n-BLBC, patients with BLBC showed a positive correlation with lymph node metastasis occurrence and lower survival rates. Immunohistochemistry analysis revealed significantly lower E-cadherin prevalence and higher prevalence of both CK19 and Ki-67 in BLBC when compared to n-BLBC. Spearman correlation showed that E-cadherin is negatively and significantly correlated to CK19 and Ki-67 expressions. Moreover, in BLBC, expressing both CK19 and Ki-67 combined with E-cadherin loss was associated with the worst relapse-free and overall survival. In conclusion, TNBC/BLBC phenotypes simultaneously losing E-cadherin and overexpressing CK19 and Ki-67 markers are the most aggressive forms. This combined analysis could be a predictive marker of poor prognosis.

Induction of Invasive Basal Phenotype in Triple-Negative Breast Cancers by Long Noncoding RNA BORG.

Long noncoding RNAs (lncRNAs) are known to play key roles in breast cancers; however, detailed mechanistic studies of lncRNA function have not been conducted in large cohorts of breast cancer tumors, nor has inter-donor and inter-subtype variability been taken into consideration for these analyses. Here we provide the first identification and annotation of the human BORG lncRNA gene. Using multiple tumor cohorts of human breast cancers, we show that while BORG expression is strongly induced in breast tumors as compared to normal breast tissues, the extent of BORG induction varies widely between breast cancer subtypes and even between different tumors within the same subtype. Elevated levels of BORG in breast tumors are associated with the acquisition of core cancer aggression pathways, including those associated with basal tumor and pluripotency phenotypes and with epithelial-mesenchymal transition (EMT) programs. While a subset of BORG-associated pathways was present in high BORG-expressing tumors across all breast cancer subtypes, many were specific to tumors categorized as triple-negative breast cancers. Finally, we show that genes induced by heterologous expression of BORG in murine models of TNBC both in vitro and in vivo strongly overlap with those associated with high BORG expression levels in human TNBC tumors. Our findings implicate human BORG as a novel driver of the highly aggressive basal TNBC tumor phenotype.

A Novel HER2 Protein Identification Methodology in Breast Cancer Cells Using Raman Spectroscopy and Raman Imaging: An Analytical Validation Study.

Conventional assays such as immunohistochemistry (IHC) and in situ hybridization (ISH) used in clinical procedures for quantification of the human epidermal growth factor receptor-2 (HER2) status in breast cancer have many limitations. In the current study, we have used HER2 expression in a broad range of breast cancer phenotypes to explore the potential utility of a novel immunodetection technique using Raman spectroscopy and Raman imaging combined with artificial intelligence models. The correlations between the Raman method and conventional HER2 testing methodologies (IHC and ISH) have been tested. Raman measurements showed a strong linear correlation (p = 0.05, R2=0,9816) with IHC analysis in the studied breast cell lines: MCF-10A, MCF-7, MDA-MB-231, HTB-30 (SK-BR-3), and AU-565 represent normal, nontumorigenic epithelial cells, triple-positive breast carcinoma, and triple-negative breast cancer cell lines. Analytic testing of Raman spectroscopy and Raman imaging demonstrated that this method may offer advantages over the currently used diagnostic methodologies.

The effects of perfluorooctanoic acid on breast cancer metastasis depend on the phenotypes of the cancer cells: An in vivo study with zebrafish xenograft model

Per- and polyfluorinated substances (PFAS) have been associated with numerous human diseases. Recent in vitro studies have implicated the association of PFAS with an increased risk of breast cancer in humans. This study aimed to assess the toxic effects of PFAS during the development of human breast cancer using a zebrafish xenograft model. Perfluorooctanoic acid (PFOA) was used as a PFAS chemical of interest for this study. Two common breast cancer cell lines, MCF-7 and MDA-MB-231, were used to represent the diversity of breast cancer phenotypes. Human preadipocytes were co-implanted with the breast cancer cells into the zebrafish embryos to optimize the microenvironment for tumor cells in vivo. With this modified model, we evaluated the potential effects of the PFOA on the metastatic potential of the two types of breast cancer cells. The presence of human preadipocytes resulted in an enhancement to the metastasis progress of the two types of cells, including the promotion of cell in vivo migration and proliferation, and the increased expression levels of metastatic biomarkers. The enhancement of MCF-7 proliferation by preadipocytes was observed after 2 days post injection (dpi) while the increase of MDA-MB-231 proliferation was seen after 6 dpi. The breast cancer metastatic biomarkers, cadherin 1 (cdh1), and small breast epithelial mucin (sbem) genes demonstrated significant down- and upregulations respectively, by the co-injection of preadipocytes. In the optimized xenograft model, the PFOA consistently promoted cell proliferation and migration and altered the metastatic biomarker expression in MCF-7, which suggested a metastatic effect of PFOA on MCF-7. However, those effects were not consistently observed in MDA-MB-231. The presence of the preadipocytes in the xenograft model may provide a necessary microenvironment for the progress of tumor cells in zebrafish embryos. The finding suggested that the impacts of PFOA exposure on different phenotypes of breast cancers may differ.

Is Oncoplastic Surgery Safe in High-Risk Breast Cancer Phenotypes?

Oncoplastic surgery (OPS) has increased in popularity over the recent years. It is a form of breast conservation surgery allowing for larger partial mastectomy (PM) resections followed by either volume displacement or volume replacement reconstruction techniques. However, there is a lack of evidence on the effectiveness and safety of OPS with radiotherapy (OPS + RT) in high-risk breast cancer phenotypes, such as triple negative breast cancer (TNBC) and HER2 positive (HER2+) patients. Our aim was to compare the breast cancer-specific survival (BCSS) and postoperative surgical complications in OPS + RT compared to PM alone with radiation (PM + RT) and total mastectomy (MTX) without radiotherapy (MTX-RT). Patient data were analyzed from the Surveillance, Epidemiology, and End Results (SEER) cancer registries from January 1, 2012 to December 31, 2020. Patients were stratified according to the type of surgery. Cox regression analysis was performed to assess prognostic factors of BCSS. A total of 24 621 patients with high-risk breast cancer phenotypes were identified, 180 underwent OPS + RT; 13 402, PM + RT; and 11 039 MTX-RT. OPS + RT was more frequently performed in younger (mean age of 65.53 years, SD: 9.29, p < 0.001), non-Hispanic White (90.5% vs. 77.7% vs. 76.3%) and single women (17.9% vs. 12.1% vs. 13.3%). MTX-RT was usually performed in patients with high histological grade, TNBC, and higher stages. Overall complication rates were higher in the MTX-RT, compared to OPS + RT and PM + RT, 2%, 1.1%, and 0.7%, respectively, p < 0.001. Rates of hematoma and surgical site infections were higher in the MTX-RT group. With a median follow-up of 46 months, OPS + RT had better BCSS rates at 5 years compared to PM + RT and MTX-RT (97.1% vs. 94.7% vs. 89.8%, p < 0.001). MTX-RT was found to be an independent prognostic factor of worse BCSS compared to OPS + RT (hazard ratio [HR] = 2.584; 95% confidence interval [CI]: 1.005-7.171), while PM + RT had no difference compared to OPS + RT (HR = 1.670, 95% CI: 0.624-4.469). OPS is a safe breast surgical option in patients with HER2+ and TNBC. Patients with high-risk phenotypes who underwent OPS + RT and have similar BCSS and complication rates compared to standard breast surgical options. As such, OPS should be considered as an option whenever breast conservation surgery is being discussed.

PFAS Levels, Early Life Factors, and Mammographic Breast Density in Premenopausal Women.

Mammographic breast density (MBD) is a strong risk factor and an intermediate phenotype for breast cancer, yet there are limited studies on how environmental pollutants are associated with MBD. We investigated associations of perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), and perfluorohexane sulfonate (PFHxS) levels with measures of MBD and evaluated if early life factors modified any associations. Metabolon performed metabolomics analysis using ultrahigh-performance liquid chromatography/tandem accurate mass spectrometry in fasting blood from 705 premenopausal women completing their annual screening mammogram in St. Louis, Missouri. We calculated least square means (LSM) of mammographic volumetric percent density (VPD), dense volume (DV), and nondense volume (NDV) by quartiles (Q) of PFOS, PFOA, and PFHxS from multivariable linear regression modeling overall and stratified by recruitment period, race, age at menarche, and body shape at age 10. Models were adjusted for age, age at menarche, body fat percentage, race, family history of breast cancer, oral contraceptive use, alcohol consumption, parity/age at first birth, and body shape at age 10. PFOS, PFOA, and PFHxS were not significantly associated with VPD or NDV. PFHxS was significantly positively associated with DV (, , , ; ). PFOS was positively associated with DV (, , , ; ) with DV being 8.1%, 12%, and 12.3% higher in Q2, Q3, and Q4 compared to Q1. Among women who were underweight/normal weight at age 10, PFOS was positively associated with VPD (, , , ; ) while there was an inverse association among women who were overweight/obese at age 10 (, , , ; ) (). We report novel associations of PFHxS and PFOS with DV in premenopausal women. PFOS, PFOA, and PFHxS were not associated with VPD and NDV. In addition, body shape at age 10 may modify the associations of PFOS with MBD. Further studies are needed to validate our findings and to evaluate the associations of other per- and polyfluoroalkyl substances (PFAS), as well as mixtures of PFAS, with MBD. https://doi.org/10.1289/EHP14065.

Direct inhibition of tumor hypoxia response with synthetic transcriptional repressors.

Many oncogenic transcription factors (TFs) are considered to be undruggable because of their reliance on large protein-protein and protein-DNA interfaces. TFs such as hypoxia-inducible factors (HIFs) and X-box-binding protein 1 (XBP1) are induced by hypoxia and other stressors in solid tumors and bind to unfolded protein response element (UPRE) and hypoxia-induced response element (HRE) motifs to control oncogenic gene programs. Here, we report a strategy to create synthetic transcriptional repressors (STRs) that mimic the basic leucine zipper domain of XBP1 and recognize UPRE and HRE motifs. A lead molecule, STR22, binds UPRE and HRE DNA sequences with high fidelity and competes with both TFs in cells. Under hypoxia, STR22 globally suppresses HIF1α binding to HRE-containing promoters and enhancers, inhibits hypoxia-induced gene expression and blocks protumorigenic phenotypes in triple-negative breast cancer (TNBC) cells. In vivo, intratumoral and systemic STR22 treatment inhibited hypoxia-dependent gene expression, primary tumor growth and metastasis of TNBC tumors. These data validate a novel strategy to target the tumor hypoxia response through coordinated inhibition of TF-DNA binding.

CAF-Associated Genes in Breast Cancer for Novel Therapeutic Strategies.

Breast cancer (BC) is the most common cancer in women, and therapeutic strategies for it are based on the molecular subtypes of luminal BC, HER2 BC, and triple-negative BC (TNBC) because each subtype harbors different unique genetic aberrations. Recently, features of the tumor microenvironment (TME), especially cancer-associated fibroblasts (CAFs), have been demonstrated to play a critical role in BC progression, and we would like to understand the molecular features of BC CAFs for novel therapeutic strategies. In a recent study, 115 CAF-associated genes (CAFGs) were identified in a public database of microdissection and microarray data (GSE35602) from 13 colorectal cancer (CRC) tumors. Using a public database (GSE10797) of 28 BC tumors, a similar analysis was performed. In BC, 59 genes from the 115 CAFGs identified in CRC (CRC CAFGs) were also closely associated with a CAFs marker, SPARC (R = 0.6 or beyond), and POSTN was of particular interest as one of the BC CAFGs with the highest expression levels and a close association with SPARC expression (R = 0.94) in the cancer stroma of BC tumors. In BC stroma, POSTN was followed in expression levels by DKK3, MMP2, PDPN, and ACTA2. Unexpectedly, FAP and VIM were not as highly associated with SPARC expression in the cancer stroma of BC tumors and exhibited low expression. These findings suggested that ACTA2 might be the most relevant conventional CAFs marker in BC, and ACTA2 was actually correlated in expression with many CRC CAFGs, such as SPARC. Surprisingly, the SE ratio values of the BC CAFGs were much lower (average SE = 3.8) than those of the CRC CAFGs (SE = 10 or beyond). We summarized the current understanding of BC CAFs from the literature. Finally, in triple-negative BC (TNBC) (n = 5), SPARC expression uniquely showed a close association with COL11A1 and TAGLN expression, representing a myofibroblast (myCAFs) marker in the cancer stroma of the BC tumors, suggesting that myCAFs may be molecularly characterized by TNBC in contrast to other BC phenotypes. In summary, CAFs could have unique molecular characteristics in BC, and such TME uniqueness could be therapeutically targeted in BC.

Gene expression-phenotype association study reveals the dual role of TNF-α/TNFR1 signaling axis in confined breast cancer cell migration

AimsWhile enhanced tumor cell migration is a key process in the tumor dissemination, mechanistic insights into causal relationships between tumor cells and mechanical confinement are still limited. Here we combine the use of microfluidic platforms to characterize confined cell migration with genomic tools to systematically unravel the global signaling landscape associated with the migratory phenotype of breast cancer (BC) cells. Meterials and methodsThe spontaneous migration capacity of seven BC cell lines was evaluated in 3D microfluidic devices and their migration capacity was correlated with publicly available molecular signatures. The role of identified signaling pathways on regulating BC migration capacity was determined by receptor stimulation through ligand binding or inhibition through siRNA silencing. Downstream effects on cell migration were evaluated in microfluidic devices, while the molecular changes were monitored by RT-qPCR. Key findingsExpression of 715 genes was correlated with BC cells migratory phenotype, revealing TNF-α as one of the top upstream regulators. Signal transduction experiments revealed that TNF-α stimulates the confined migration of triple negative, mesenchymal-like BC cells that are also characterized by high TNFR1 expression, but inhibits the migration of epithelial-like cells with low TNFR1 expression. TNFR1 was strongly associated with the migration capacity and triple-negative, mesenchymal phenotype. Downstream of TNF/TNFR1 signaling, transcriptional regulation of NFKB seems to be important in driving cell migration in confined spaces. SignificanceTNF-α/TNFR1 signaling axis reveals as a key player in driving BC cells confined migration, emerging as a promising therapeutic strategy in targeting dissemination and metastasis of triple negative, mesenchymal BC cells.

Tamoxifen metabolites treatment promotes ERα+ transition to triple negative phenotype in vitro, effects of LDL in chemoresistance.

Estrogen receptor-positive (ER+) breast cancer represents about 80% of cases, tamoxifen is the election neoadjuvant chemotherapy. However, a large percentage of patients develop chemoresistance, compromising recovery. Clinical evidence suggests that high plasmatic levels of low-density lipoproteins (LDL) could promote cancer progression. The present study analyzed the effect of LDL on the primary plasmatic active Tamoxifen's metabolites resistance acquisition, 4-hydroxytamoxifen (4OH-Tam) and 4-hydroxy-N-desmethyl-tamoxifen (endoxifen), in breast cancer ERα + cells (MCF-7). Two resistant cellular variants, MCF-7Var-H and MCF-7Var-I, were generated by a novel strategy and their phenotype features were evaluated. Phenotypic assessment was performed by MTT assays, cytometry, immunofluorescence microscopy, zymography and protein expression analysis. MCF-7Var-H, generated only with tamoxifen metabolites, showed a critical down-regulation in hormone receptors, augmented migration capacity, metalloprotease 9 extracellular medium excretion, and a mesenchymal morphology in contrast with native MCF-7, suggesting the transition towards Triple-negative breast cancer (TNBC) phenotype. In contrast, MCF-7Var-I which was generated in a high LDL media, showed only a slight upregulation in ER and other less noticeable metabolic adaptations. Results suggest a potential role of transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) in phenotypic differences observed among variants. LDL high or low concentrations during Tamoxifen´s metabolites chemoresistance acquisition leads to different cellular mechanisms related to chemoresistance. A novel adaptative cellular response associated with Nrf2 activity could be implicated.

Whole genome joint analysis reveals ATM:C.1564_1565del variant segregating with Ataxia-Telangiectasia and breast cancer

ATM gene is implicated in the development of breast cancer in the heterozygous state, and Ataxia-telangiectasia (A-T) in a homozygous or compound heterozygous state. Ataxia-telangiectasia (A-T) is a rare cerebellar ataxia syndrome presenting with progressive neurologic impairment, telangiectasia, and an increased risk of leukemia and lymphoma.Although the role of ATM, separately, in association with A-T and breast cancer is well documented, there is a limited number of studies investigating ATM variants when segregating with both phenotypes in the same family. Here, using joint analysis and whole genome sequencing, we investigated ATM c.1564_1565del in a family with one homozygous member presenting with A-T (OMIM # 208900) and three heterozygous members, of whom one had breast cancer (OMIM #114480). To our knowledge, this is the first study of ATM c.1564_1565del segregation with both A-T and breast cancer phenotypes within the same kindred. This study highlights the need for a comprehensive genomic approach in the appropriate cancer risk management of heterozygote carriers of ATM in families with A-T.

The up-regulation of PD-L1 during boningmycin-induced senescence in human cancer cells depends on the activation of the JAK/STAT signaling pathway mediated by SASP.

Therapy-induced senescence can regulate both the innate and adaptive immune systems, thereby affecting therapeutic efficacy. Bleomycin is a major component of combined chemotherapy regimens, utilized for the treatment of multiple tumors, whereas pulmonary toxicity severely restricts its clinical benefits. As a member of the bleomycin family, boningmycin (BON) exhibits potent anticancer activity with minimal pulmonary toxicity, making it a potential alternative to bleomycin. Low concentrations of BON can induce senescence, but the impact of BON-induced senescence on anticancer immunity remains unclear. This study investigates the effects of BON-induced senescence on PD-L1 expression and the underlying mechanisms in human cancer cells. Firstly, the elevation of PD-L1 protein during BON-induced senescence was confirmed by a senescence β-galactosidase staining assay, detection of the senescence-associated secretory phenotype (SASP), western blot and flow cytometry in human lung cancer NCI-H460 cells and breast cancer MDA-MB-231 cells. Subsequently, it was shown that the increase in PD-L1 protein is mediated by SASP, as evidenced by the use of conditional media, knockdown of cyclic GMP-AMP synthase and inhibition of stimulator of interferon genes. Ultimately, it was demonstrated that SASP-mediated PD-L1 up-regulation is dependent on the activation of the JAK/STAT pathway through the use of specific inhibitors and siRNAs. These findings clarify the impact of BON-induced senescence on PD-L1 expression and may contribute to the optimization of the therapeutic efficacy of bleomycin-related compounds and the clinical transformation of BON.

Androgen receptor-mediated pharmacogenomic expression quantitative trait loci: implications for breast cancer response to AR-targeting therapy

BackgroundEndocrine therapy is the most important treatment modality of breast cancer patients whose tumors express the estrogen receptor α (ERα). The androgen receptor (AR) is also expressed in the vast majority (80–90%) of ERα-positive tumors. AR-targeting drugs are not used in clinical practice, but have been evaluated in multiple trials and preclinical studies.MethodsWe performed a genome-wide study to identify hormone/drug-induced single nucleotide polymorphism (SNP) genotype - dependent gene-expression, known as PGx-eQTL, mediated by either an AR agonist (dihydrotestosterone) or a partial antagonist (enzalutamide), utilizing a previously well characterized lymphoblastic cell line panel. The association of the identified SNPs-gene pairs with breast cancer phenotypes were then examined using three genome-wide association (GWAS) studies that we have published and other studies from the GWAS catalog.ResultsWe identified 13 DHT-mediated PGx-eQTL loci and 23 Enz-mediated PGx-eQTL loci that were associated with breast cancer outcomes post ER antagonist or aromatase inhibitors (AI) treatment, or with pharmacodynamic (PD) effects of AIs. An additional 30 loci were found to be associated with cancer risk and sex-hormone binding globulin levels. The top loci involved the genes IDH2 and TMEM9, the expression of which were suppressed by DHT in a PGx-eQTL SNP genotype-dependent manner. Both of these genes were overexpressed in breast cancer and were associated with a poorer prognosis. Therefore, suppression of these genes by AR agonists may benefit patients with minor allele genotypes for these SNPs.ConclusionsWe identified AR-related PGx-eQTL SNP-gene pairs that were associated with risks, outcomes and PD effects of endocrine therapy that may provide potential biomarkers for individualized treatment of breast cancer.

The Role of Melatonin and Matrix Metalloproteinases in Breast Cancer Pathogenesis: A Comprehensive Review

Background: Breast cancer is a heterogeneous disease with molecular subtypes including Triple Negative Breast Cancer (TNBC), HER2/neu-positive, Luminal Type A, and Luminal Type B, each with unique clinical behaviors and therapeutic responses. This study investigates the role of melatonin, matrix metalloproteinases (MMPs), pro-inflammatory cytokines (IL6 and TNF-alpha), and lactate dehydrogenase (LDH) isoenzymes in breast cancer pathogenesis. Methods: Over three years, 171 breast cancer patients were studied. Blood and urine samples were collected to measure melatonin, MMPs, IL-6, TNF-alpha, LDH-1, and LDH-5 levels. Circulating tumor cells (CTCs) were isolated using density gradient centrifugation and immunomagnetic separation. Melatonin receptor expression on CTCs was analyzed by flow cytometry and immunocytochemistry. Plasma concentrations of melatonin were measured using high-performance liquid chromatography (HPLC), and MMPs, IL-6, TNF-alpha, LDH isoenzymes were quantified using enzyme-linked immunosorbent assays (ELISAs) and electrophoretic methods. Results: The expression of melatonin receptors was lowest in TNBC patients, higher in HER2/neu-positive and ER/PRnegative patients, and highest in Luminal Type A and B patients. TNBC patients exhibited elevated LDH-1 and inflammatory cytokines (IL-6, TNF-alpha), with significantly decreased melatonin levels. HER2/neu-positive patients showed elevated LDH5 and moderate decreases in melatonin. Luminal Type B and A patients had varying degrees of biomarker elevation and melatonin receptor expression. Discussion: The findings suggest a correlation between reduced melatonin signaling and aggressive breast cancer phenotypes, particularly TNBC. Elevated MMPs and pro-inflammatory cytokines underscore the role of inflammation and ECM degradation in cancer progression. Melatonin’s potential inhibitory effects on these pathways highlight its therapeutic promise. The combined assessment of melatonin, MMPs, cytokines, and LDH could guide personalized treatment strategies. Conclusion: This study underscores melatonin’s multifaceted role in breast cancer pathogenesis. Melatonin receptor expression, along with MMP, IL-6, TNF-alpha, and LDH levels, provides a detailed molecular landscape of breast cancer subtypes. Integrating melatonin supplementation into therapeutic regimens holds promise for improving outcomes, particularly inaggressive subtypes like TNBC. Further research and clinical trials are essential to validate these findings and develop effective melatonin-based therapies.

Stimulator of Interferon Genes Protein (STING) Expression in Cancer Cells: A Tissue Microarray Study Evaluating More than 18,000 Tumors from 139 Different Tumor Entities.

Stimulator of interferon genes protein (STING) activates the immune response in inflammatory cells. STING expression in cancer cells is less well characterized, but STING agonists are currently being evaluated as anticancer drugs. A tissue microarray containing 18,001 samples from 139 different tumor types was analyzed for STING by immunohistochemistry. STING-positive tumor cells were found in 130 (93.5%) of 139 tumor entities. The highest STING positivity rates occurred in squamous cell carcinomas (up to 96%); malignant mesothelioma (88.5%-95.7%); adenocarcinoma of the pancreas (94.9%), lung (90.3%), cervix (90.0%), colorectum (75.2%), and gallbladder (68.8%); and serous high-grade ovarian cancer (86.0%). High STING expression was linked to adverse phenotypes in breast cancer, clear cell renal cell carcinoma, colorectal adenocarcinoma, hepatocellular carcinoma, and papillary carcinoma of the thyroid (p < 0.05). In pTa urothelial carcinomas, STING expression was associated with low-grade carcinoma (p = 0.0002). Across all tumors, STING expression paralleled PD-L1 positivity of tumor and inflammatory cells (p < 0.0001 each) but was unrelated to the density of CD8+ lymphocytes. STING expression is variable across tumor types and may be related to aggressive tumor phenotype and PD-L1 positivity. The lack of relationship with tumor-infiltrating CD8+ lymphocytes argues against a significant IFN production by STING positive tumor cells.

FOXCUT regulates the malignant phenotype of triple-negative breast Cancer via the miR-337-3p/ANP32E Axis

BackgroundThe lack of specific molecular targets and the rapid spread lead to a worse prognosis of triple-negative breast cancer (TNBC). Therefore, identifying new therapeutic and prognostic biomarkers helps to develop effective treatment strategies for TNBC. MethodsThrough preliminary bioinformatics analysis, FOXCUT was found to be significantly overexpressed in breast cancer, especially in TNBC. Tissue samples were collected from 15 TNBC patients, and qRT-PCR was employed to validate the expression of FOXCUT in both TNBC patient tissues and TNBC cell lines. We also carried out the GSEA analysis and KEGG enrichment analysis of FOXCUT. Additionally, the effects of FOXCUT knockdown on TNBC cell malignant behaviors, and aerobic glycolysis were assessed by methods including CCK-8, Transwell, western blot, and Seahorse XF 96 analyses. Moreover, utilizing databases predicting interactions between ceRNAs, corresponding lncRNA-miRNA binding relationships, and miRNA-mRNA interactions were predicted. These predictions were subsequently validated through RNA immunoprecipitation and dual-luciferase reporter assays. ResultsFOXCUT exhibited high expression in both TNBC tissues and cell lines, fostering cell malignant behaviors and glycolysis. FOXCUT was found to sponge miR-337-3p, while miR-337-3p negatively regulated the expression of ANP32E. Consequently, FOXCUT ultimately facilitated the malignant phenotype of TNBC by upregulating ANP32E expression. ConclusionThis study elucidated the role of FOXCUT in elevating aerobic glycolysis levels in TNBC and driving malignant cancer cell development via the miR-337-3p/ANP32E regulatory axis.

Germline rare variants in HER2-positive breast cancer predisposition: a systematic review and meta-analysis.

Approximately 10% of breast cancer (BC) cases result from hereditary causes. Genetic testing has been widely implemented in BC care to determine hereditary cancer syndromes and personalized medicine. Thus, identification of individuals carrying germline pathogenic variants could be useful to provide appropriate prophylactic or screening measures for each BC subtype, however, there are few formal recommendations for genetic testing in this sense so far. In this study, we assessed rare germline variants in a specific group of genes in order to determine the association with human epidermal growth factor 2 enriched (HER2+) BC phenotype through a systematic review and meta-analysis comparing subtypes overexpressing HER2 with other clinically recognized subtypes of BC. This review was registered with PROSPERO (ID: CRD42023447571). We conducted an online literature search in PubMed (MEDLINE), Scopus, and EMBASE databases. We included original studies that investigated germline variants in HER2+ BC patients and selected the studies that reported only rare and/or pathogenic germline variants. We assessed the risk of bias and quality of the studies using the Joanna Briggs Institute Critical Appraisal checklists and the Modified Newcastle-Ottawa Scale for Genetic Studies, respectively. Considering hormone receptor and HER2 expression status, we compared gene-based risks initially in HR-HER2-, HR+HER2-, HR+HER2+, and HR-HER2+ groups, conducting separate meta-analyses using the random effects model for each comparison, and within them for each gene. Of the total 36 studies describing germline variants, 11 studies provided information on the prevalence of variants in the different clinically relevant BC subtypes and allowed comparisons. Germline variants within eight genes showed significant differences when meta-analyzed between the BC groups: BRCA1, BRCA2, TP53, ATM, CHEK2, PALB2, RAD51C, and BARD1. Notably, TP53, ATM, and CHEK2 germline variants were identified as predisposing factors for HER2+ subtypes, whereas BRCA1, BRCA2, PALB2, RAD51C, and BARD1 germline variants were associated with a predisposition to low HER2 expression. Main concerns about bias and quality assessment were the lack of confounding factors control; and comparability or outcome assessment, respectively. Our findings underscore the connection between germline variants and differential expression of the HER2 protein and BC subtypes. https://www.crd.york.ac.uk/PROSPERO, identifier CRD42023447571.

TFAP2A downregulation mediates tumor-suppressive effect of miR-8072 in triple-negative breast cancer via inhibiting SNAI1 transcription

BackgroundTriple-negative breast cancer (TNBC) represents a highly aggressive subset of breast malignancies characterized by its challenging clinical management and unfavorable prognosis. While TFAP2A, a member of the AP-2 transcription factor family, has been implicated in maintaining the basal phenotype of breast cancer, its precise regulatory role in TNBC remains undefined.MethodsIn vitro assessments of TNBC cell growth and migratory potential were conducted using MTS, colony formation, and EdU assays. Quantitative PCR was employed to analyze mRNA expression levels, while Western blot was utilized to evaluate protein expression and phosphorylation status of AKT and ERK. The post-transcriptional regulation of TFAP2A by miR-8072 and the transcriptional activation of SNAI1 by TFAP2A were investigated through luciferase reporter assays. A xenograft mouse model was employed to assess the in vivo growth capacity of TNBC cells.ResultsSelective silencing of TFAP2A significantly impeded the proliferation and migration of TNBC cells, with elevated TFAP2A expression observed in breast cancer tissues. Notably, TNBC patients exhibiting heightened TFAP2A levels experienced abbreviated overall survival. Mechanistically, TFAP2A was identified as a transcriptional activator of SNAI1, a crucial regulator of epithelial-mesenchymal transition (EMT) and cellular proliferation, thereby augmenting the oncogenic properties of TFAP2A in TNBC. Moreover, miR-8072 was unveiled as a negative regulator of TFAP2A, exerting potent inhibitory effects on TNBC cell growth and migration. Importantly, the tumor-suppressive actions mediated by the miR-8072/TFAP2A axis were intricately associated with the attenuation of AKT/ERK signaling cascades and the blockade of EMT processes.ConclusionsOur findings unravel the role and underlying molecular mechanism of TFAP2A in driving tumorigenesis of TNBC. Targeting the TFAP2A/SNAI1 pathway and utilizing miR-8072 as a suppressor represent promising therapeutic strategies for treating TNBC.

MiR-181c-5p/DERL1 pathway controls breast cancer progression mediated by TRAF6-linked K63 ubiquitination of AKT

BackgroundAberrant Derlin-1 (DERL1) expression is associated with an overactivation of p-AKT, whose involvement in breast cancer (BRCA) development has been widely speculated. However, the precise mechanism that links DERL1 expression and AKT activation is less well-studied.MethodsBioinformatic analyses hold a promising approach by which to detect genes’ expression levels and their association with disease prognoses in patients. In the present work, a dual-luciferase assay was employed to investigate the relationship between DERL1 expression and the candidate miRNA by both in vitro and in vivo methods. Further in-depth studies involving immunoprecipitation-mass spectrum (IP-MS), co-immunoprecipitation (Co-IP), as well as Zdock prediction were performed.ResultsOverexpression of DERL1 was detected in all phenotypes of BRCA, and its knockdown showed an inhibitory effect on BRCA cells both in vitro and in vivo. The Cancer Genome Atlas (TCGA) database reported that DERL1 overexpression was correlated with poor overall survival in BRCA cases, and so the quantification of DERL1 expression could be a potential marker for the clinical diagnosis of BRCA. On the other hand, miR-181c-5p was downregulated in BRCA, suggesting that its overexpression could be a potent therapeutic route to improve the overall survival of BRCA cases. Prior bioinformatic analyses indicated a somewhat positive correlation between DERL1 and TRAF6 as well as between TRAF6 and AKT, but not between miR-181c-5p and DERL1. In retrospect, DERL1 overexpression promoted p-AKT activation through K63 ubiquitination. DERL1 was believed to directly interact with the E3 ligase TRAF6. As Tyr77Ala or Tyr77Ala/Gln81Ala/Arg85Ala/Val158Ala attempts to prevent the interaction between DERL1 and TRAF domain of TRAF6, resulted in a significant reduction in K63-ubiquitinated p-AKT production. However, mutations in Gln81Ala, Arg85Ala, or Val158Ala could possibly interrupt with these processes.ConclusionsOur data confirm that mediation of the miR-181c-5p/DERL1 pathway by TRAF6-linked AKT K63 ubiquitination holds one of the clues to set our focus on toward meeting the therapeutic goals of BRCA.

Assessing the impact of extracellular matrix fiber orientation on breast cancer cellular metabolism

The extracellular matrix (ECM) is a dynamic and complex microenvironment that modulates cell behavior and cell fate. Changes in ECM composition and architecture have been correlated with development, differentiation, and disease progression in various pathologies, including breast cancer [1]. Studies have shown that aligned fibers drive a pro-metastatic microenvironment, promoting the transformation of mammary epithelial cells into invasive ductal carcinoma via the epithelial-to-mesenchymal transition (EMT) [2]. The impact of ECM orientation on breast cancer metabolism, however, is largely unknown. Here, we employ two non-invasive imaging techniques, fluorescence-lifetime imaging microscopy (FLIM) and intensity-based multiphoton microscopy, to assess the metabolic states of cancer cells cultured on ECM-mimicking nanofibers in a random and aligned orientation. By tracking the changes in the intrinsic fluorescence of nicotinamide adenine dinucleotide and flavin adenine dinucleotide, as well as expression levels of metastatic markers, we reveal how ECM fiber orientation alters cancer metabolism and EMT progression. Our study indicates that aligned cellular microenvironments play a key role in promoting metastatic phenotypes of breast cancer as evidenced by a more glycolytic metabolic signature on nanofiber scaffolds of aligned orientation compared to scaffolds of random orientation. This finding is particularly relevant for subsets of breast cancer marked by high levels of collagen remodeling (e.g. pregnancy associated breast cancer), and may serve as a platform for predicting clinical outcomes within these subsets [3–6].