Potential Biomarker For Breast Cancer Research Articles

Involvement of N4BP2L1, PLEKHA4, and BEGAIN genes in breast cancer and muscle cell development

Patients with breast cancer show altered expression of genes within the pectoralis major skeletal muscle cells of the breast. Through analyses of The Cancer Genome Atlas (TCGA)-breast cancer (BRCA), we identified three previously uncharacterized putative novel tumor suppressor genes expressed in normal muscle cells, whose expression was downregulated in breast tumors. We found that NEDD4 binding protein 2-like 1 (N4BP2L1), pleckstrin homology domain-containing family A member 4 (PLEKHA4), and brain-enriched guanylate kinase-associated protein (BEGAIN) that are normally highly expressed in breast myoepithelial cells and smooth muscle cells were significantly downregulated in breast tumor tissues of a cohort of 50 patients with this cancer. Our data revealed that the low expression of PLEKHA4 in patients with menopause below 50 years correlated with a higher risk of breast cancer. Moreover, we identified N4BP2L1 and BEGAIN as potential biomarkers of HER2-positive breast cancer. Furthermore, low BEGAIN expression in breast cancer patients with blood fat, heart problems, and diabetes correlated with a higher risk of this cancer. In addition, protein and RNA expression analysis of TCGA-BRCA revealed N4BP2L1 as a promising diagnostic protein biomarker in breast cancer. In addition, the in silico data of scRNA-seq showed high expression of these genes in several cell types of normal breast tissue, including breast myoepithelial cells and smooth muscle cells. Thus, our results suggest their possible tumor-suppressive function in breast cancer and muscle development.

Abstract PO1-24-07: LYSOSOMAL-ASSOCIATED TRANSMENBRANE 4B (LAPTM4B) AS POTENTIAL BIOMARKER IN BREAST CANCER

Abstract LAPTM4B is a transmembrane that promotes autophagy and renders tumor cells resistant to metabolic and genotoxic stress. Increased expression of LAPTM4B has been found in breast, liver, lung, ovarian, uterine and gastric cancers. Overexpression of LAPTM4B contributes to chemotherapy resistance, proliferation and metastases. We analyze the expression of LAPTM4B in serum by ELISA method in a series of breast cancer patients in different clinical situations to assess its possible potential as a biomarker. A total of 135 patients were analyzed, 95 patients were in early breast cancer and 50 in metastatic disease. Of the 95 patients with early disease, 52 had a luminal A phenotype, 19 luminal B, 15 HER2 positive and 9 triple negative. And of the other 50 they were luminal in 34, HER2 positive in 11 and triple negative in 5. The attached table specifies the values of the LAPTM4B expression in the different clinical situations. GLOBAL LUMINAL A LUMNAL B HER2 TRIPLE NEGATIVE SITUATIONS Laptm4b n Laptm4b n Laptm4b n Laptm4b n Laptm4b n EARLY 9,6 ng/ml 95 7,7 ng/ml 52 12,8 ng/ml 19 13,4 ng/ml 15 7,1 ng/ml 9 METASTATIC 16,8 ng/ml 50 13,7 ng/ml 34 7,2 ng/ml 11 58,9 ng/ml 5 We found a statistic difference in the expression of LAPTM4B in patients with early and metastatic disease (ANOVA test p:0,028 ). LAPTM4B was higher in the luminal B and HER2 subtypes in early breast cancer while in metastatic disease was overexpressed in triple negative. An interesting finding was that in metastatic disease, LAPTM4B was overexpressed in patients with progression compared to patients in response. 30 ng/ml vs 10 ng/ml (ANOVA test p:0,011) LAPTM4B expression may be a potential biomarker in breast cancer since its expression is statistically different in early and metastatic disease (9,6 vs 16,8 ng/ml).In addition, patients with metastatic breast cancer who re in response also have lower levels of LAPTM4B compared with patients in progression. Monitoring the levels of LAPTM4B in serum can help us to predict the treatment response. Citation Format: Serafin Morales Murillo, Ariadna Gasol Cudós, Noemí Tuset Der-Abrain, Izaskun Urdanibia, Ana Velasco Sánchez. LYSOSOMAL-ASSOCIATED TRANSMENBRANE 4B (LAPTM4B) AS POTENTIAL BIOMARKER IN BREAST CANCER [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-24-07.

Abstract PO2-03-08: Uncovering Novel Potential Prognostic Biomarkers in Basal-Like Breast Cancer using Transcriptomic Data of 1,899 Patients

Abstract Introduction: Patients with basal-like breast cancer (BLBC) predominantly represented by triple-negative breast cancer have shown a high recurrence rate and are characterized by poor prognosis. There is an urgent need to undercover reliable prognostic biomarkers that can help in the clinical management of such patients and identify additional therapeutic targets. The objective of this study was to create a comprehensive transcriptomic database on a large scale and leverage it to identify and prioritize cancer-related genes associated with BLBC patients’ outcomes. Methods: We identified breast cancer cohorts from public repositories that contained gene expression data at the transcriptome level, along with clinical follow-up information. BLBC were identified using the PAM50 signature. All samples were standardized using a standard array normalization coupled with scaling to have a mean expression across all genes of 1000 in each sample and incorporated into a unified database. Redundant samples were removed. For each gene, Cox univariate survival analysis was conducted, to account for multiple hypothesis testing, the false discovery rate was computed, and a significant cutoff of 1% was employed to determine the highest statistical significance. Association with RFS and OS was performed. Multivariate analysis was performed for selected genes involving clinical and pathological variables. To uncover higher-level functions related to altered RFS, gene ontology analysis was performed using the enrichGO function in the TNM plotter (http://www.tnmplot.com). Results: The complete integrated database comprises 1,899 samples from 52 breast cancer datasets. Altogether, 2,342 genes were correlated with relapse-free survival (RFS), and 1,149 genes were correlated with overall survival (OS). 619 genes were statistically significant for both RFS and OS. The most significant genes were ANGPTL4 (p=4.25E-08, HR=2.02), NHP2 (p=5.98E-10, HR=1.93), STK3 (p=4.86E-10, HR=1.93), GBE1 (p=2.77E-09, HR=1.86), and PMVK (p=3.65E-09, HR=1.85) for RFS and PINK1 (p=1.64E-05 , HR=3.31), CAMK2N1 (p=1.06E-07 , HR=2.93), CACFD1 (p=4.79E-04 , HR=2.61), SCAP (p=3.29E-04 , HR=2.6), SDC1 (p=2.81E-04 , HR=2.57), for OS. The most significant gene ontology biological processes upregulated in tumors with a worse prognosis include GO:0000184, nuclear-transcribed mRNA catabolic process, nonsense-mediated decay (p=6.64E-18); GO:0045047 , protein targeting to ER (p=6.64E-18); GO:0006614, SRP-dependent cotranslational protein targeting to membrane (p=9.08E-18); GO:0072599, establishment of protein localization to endoplasmic reticulum (p=1.30E-17); and GO:0006613, cotranslational protein targeting to membrane (p=1.90E-17). Conclusions: Our results help to prioritize genes and to neglect those which are most likely to fail in studies aiming to establish new clinically useful biomarkers and therapeutic targets in BLBC. Citation Format: Balazs Gyorffy, Libero Santarpia. Uncovering Novel Potential Prognostic Biomarkers in Basal-Like Breast Cancer using Transcriptomic Data of 1,899 Patients [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-03-08.

The Significance of Thyroid Hormone Receptors in Breast Cancer: A Hypothesis-Generating Narrative Review.

Breast cancer (BC) is frequently diagnosed among Canadian women. While targeted therapies are available for most BC patients; treatment resistance is common and novel therapeutic targets are of interest. Thyroid hormones (TH) bound to thyroid hormone receptors (THR) influence cell proliferation and differentiation; they are also involved in the growth and development of normal breast tissue. Evidence suggests that THRβ is a tumor suppressor in various solid tumors. This narrative review discusses retrospective studies regarding the clinical relevance of THRβ as a potential prognostic biomarker and therapeutic target in BC. We consulted with an information specialist to develop a search strategy to find all literature related to THRα expression as a potential prognostic and therapeutic biomarker in breast cancer. The primary search was developed for Medline and translated to Embase. The searches were conducted on the Ovid platform on 18 August 2023. Across seven retrospective studies identified, several have shown an association between higher THRβ1 expression with a lower risk of BC recurrence and with longer overall survival. Some evidence suggests that THRβ expression is associated with a lower risk of BC recurrence and death. Validation of THRβ as an independent prognostic biomarker and possible predictive biomarker of response to endocrine therapy and/or chemotherapy is of interest. Given that THRβ is upstream of the AKT/PI3K pathway, its potential as a predictive biomarker of response to AKT inhibitors and/or PI3K inhibitors may also be of value. Finally, the potential re-purposing of THRβ agonists as anti-cancer agents warrants investigation.

Development and validation of a model and nomogram for breast cancer diagnosis based on quantitative analysis of serum disease-specific haptoglobin N-glycosylation

BackgroundA better diagnostic marker is in need to distinguish breast cancer from suspicious breast lesions. The abnormal glycosylation of haptoglobin has been documented to assist cancer diagnosis. This study aims to evaluate disease-specific haptoglobin (DSHp)-β N-glycosylation as a potential biomarker for breast cancer diagnosis.MethodsDSHp-β chains of 497 patients with suspicious breast lesions who underwent breast surgery were separated from serum immunoinflammatory-related protein complexes. DSHp-β N-glycosylation was quantified by mass spectrometric analysis. After missing data imputation and propensity score matching, patients were randomly assigned to the training set (n = 269) and validation set (n = 113). Logistic regression analysis was employed in model and nomogram construction. The diagnostic performance was analyzed with receiver operating characteristic and calibration curves.Results95 N-glycopeptides at glycosylation sites N207/N211, N241, and N184 were identified in 235 patients with benign breast diseases and 262 patients with breast cancer. DSHp-β N-tetrafucosyl and hexafucosyl were significantly increased in breast cancer compared with benign diseases (p < 0.001 and p = 0.001, respectively). The new diagnostic model and nomogram included GN2F2, G6N3F6, GN2FS at N184, G-N&G2S2, G2&G3NFS, G2N3F, GN3 at N207/N211, CEA, CA153, and could reliably distinguish breast cancer from benign diseases. For the training set, validation set, and training and validation sets, the area under the curves (AUCs) were 0.80 (95% CI: 0.75–0.86, specificity: 87%, sensitivity: 62%), 0.77 (95% CI:0.69–0.86, specificity: 75%, sensitivity: 69%), and 0.80 (95% CI:0.76–0.84, specificity: 77%, sensitivity: 68%), respectively. CEA, CA153, and their combination yielded AUCs of 0.62 (95% CI: 0.56–0.67, specificity: 29%, sensitivity: 90%), 0.65 (95% CI: 0.60–0.71, specificity: 74%, sensitivity: 51%), and 0.67 (95% CI: 0.62–0.73, specificity: 60%, sensitivity: 68%), respectively.ConclusionsThe combination of DSHp-β N-glycopeptides, CEA, and CA153 might be a better serologic marker to differentiate between breast cancer and benign breast diseases. The dysregulated N-glycosylation of serum DSHp-β could provide insights into breast tumorigenesis.

UTRN as a potential biomarker in breast cancer: a comprehensive bioinformatics and in vitro study

Utrophin (UTRN), known as a tumor suppressor, potentially regulates tumor development and the immune microenvironment. However, its impact on breast cancer’s development and treatment remains unstudied. We conducted a thorough examination of UTRN using both bioinformatic and in vitro experiments in this study. We discovered UTRN expression decreased in breast cancer compared to standard samples. High UTRN expression correlated with better prognosis. Drug sensitivity tests and RT-qPCR assays revealed UTRN’s pivotal role in tamoxifen resistance. Furthermore, the Kruskal–Wallis rank test indicated UTRN’s potential as a valuable diagnostic biomarker for breast cancer and its utility in detecting T stage of breast cancer. Additionally, our results demonstrated UTRN’s close association with immune cells, inhibitors, stimulators, receptors, and chemokines in breast cancer (BRCA). This research provides a novel perspective on UTRN’s role in breast cancer’s prognostic and therapeutic value. Low UTRN expression may contribute to tamoxifen resistance and a poor prognosis. Specifically, UTRN can improve clinical decision-making and raise the diagnosis accuracy of breast cancer.

Abstract 6990: RHBDF1 is a potential modulator of EGFR activation and oncogenic biomarker in triple-negative breast cancer

Abstract Recently, our proteomics study on breast cancer (BC) reported a possible association of RHBDF1 with triple-negative breast cancer (TNBC). The overall survival of TNBC patients on METABRIC data set implicated that the RHBDF1 high-expression group has a shorter life expectancy than the RHBDF1 low-expression group. Although many TNBC patients have high expression levels of RHBDF1 and poor prognosis, the mechanism of RHBDF1 in the pathogenesis of TNBC remained unclear. Rhomboid 5 homolog 1 (RHBDF1) is a catalytically inactive pseudoprotease on the Golgi and endoplasmic reticulum membrane. Despite its inactive status of peptidase, it regulates the activity of ADAM17 protease that cleaves the extracellular portion of EGFR ligands such as TGF alpha and EGF. Eventually, RHBDF1 and ADAM17 complex activate the EGFR signaling pathway, thereby increasing cell proliferation, migration, and tumorigenesis. First of all, we recruited the clinical data of the overall survival (OS) and disease-free survival (DFS) in the hospital of Seoul National University (SNUH cohort, n= 203) and compared IHC score 0 or 1 with survival rate. To investigate the underlying biological mechanism of RHBDF1 in TNBC, the effect of knockdown or overexpression of RHBDF1 was observed in TNBC cell lines (MDA-MB-468, HCC1143 and BT20) using siRNA (small interfering RNA) and lentiviral systems. Cell proliferation assay was performed to clarify whether RHBDF1 affects cell growth. Then, we estimated the ability of migration, invasion, and sphere formation to confirm the characteristics of cancer cells including cancer stemness and metastasis. SNUH cohort with TNBC patients showed that the RHBDF1 high-expression group has a poorer disease-free survival (DFS) than the low-expression group (p&amp;lt;0.05). In vitro assay, RHBDF1 knockdown inhibited proliferation in MDA-MB-468 and HCC1143. RHBDF1 downregulation also decreased the ability of migration and invasion. We found that RHBDF1 activates specific cell growth signaling pathways and increases cell mobility in TNBC. The sphere formation assay was performed to measure cancer stemness characteristics. As a result, RHBDF1 knockdown TNBC cell lines couldn’t form spheres relative to RHBDF1 overexpressed TNBC cell lines. In conclusion, we demonstrated that RHBDF1, a potential triple-negative breast cancer biomarker, is highly amplified in TNBC patients and TNBC cell lines. Its functions of accelerating cell proliferation, metastasis and cancer stemness were unveiled. But we need to study ultimately how RHBDF1 regulates ADAM17 activity at Golgi and endoplasmic reticulum membrane and which transcription factor activates RHBDF1 expression. Citation Format: Hayeon Kim, Soo Young Park, Da Sol Kim, Cheng Hyun Lee, Min Ji Song, Han Suk Ryu. RHBDF1 is a potential modulator of EGFR activation and oncogenic biomarker in triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6990.

Blood-based microRNA profiling unveils complex molecular dynamics in breast cancer.

Breast cancer, a genetically intricate disease with diverse subtypes, exhibits heightened incidence globally. In this study, we aimed to investigate blood-based microRNAs (miRNAs) as potential biomarkers for breast cancer. The primary objectives were to explore the role of miRNAs in cancer-related processes, assess their differential expression between breast cancer patients and healthy individuals, and contribute to a deeper understanding of the molecular underpinnings of breast cancer. MiRNA extraction was performed on 40 breast cancer patients and adjacent normal tissues using a commercial RNA isolation kit. Total RNA quantification and quality assessment were conducted with advanced technologies. MiRNA profiling involved reverse transcription, labeling, and hybridization on Agilent human miRNA arrays (V2). Bioinformatics analysis utilized the DIANA system for target gene prediction and the DIANA-mirPath tool for pathway enrichment analysis. Selected miRNAs underwent validation through quantitative real-time PCR. Principal component analysis revealed overlapping miRNA expression patterns in primary and malignant breast tumors, underscoring the genetic complexity involved. Statistical analysis identified 54 downregulated miRNAs in malignant tumors and 38 in primary tumors compared to controls. Bioinformatics analysis implicated several pathways, including Wnt, TGF-b, ErbB, and MAPK signaling. Validation through qRT-PCR confirmed altered expression of hsa-miR-130a, hsa-miR-21, hsa-miR-223, and hsa-let-7c key miRNAs, highlighting their significance in breast cancer. The results from microarray were further validated by qPCR and the expression of which are downregulated in breast cancer was detected. This study provides significant insights into distinct miRNA expression patterns in normal and malignant breast tissues. The overlapping miRNA profiles in primary and malignant tumors underscore the complexity of genetic regulation in breast cancer. The identification of deregulated miRNAs and affected pathways contributes to our understanding of breast cancer pathogenesis. The validated miRNAs hold potential as diagnostic and prognostic markers, offering avenues for further clinical exploration in breast cancer research.

Targeted Sequencing of Human Satellite 2 Repeat Sequences in Plasma cfDNA Reveals Potential Breast Cancer Biomarkers.

Liquid biopsies are revolutionizing the detection and management of malignant diseases. While repetitive DNA sequences, such as LINE-1 and ALU are established in cell-free DNA (cfDNA) research, their clinical applications remain limited. In this study, we explore human satellite 2 (HSATII), a prevalent repeat DNA sequence in plasma that exhibits increased levels in cancer patients, thereby positioning it as a potential pan-cancer biomarker. We employed targeted sequencing and copy number variation (CNV) analysis using two primer pairs to assess the differential abundance of HSATII sequences in the plasma of breast cancer patients compared to healthy individuals. PCR amplicons of HSATII from 10 patients and 10 control subjects were sequenced, generating 151 bp paired-end reads. By constructing a pooled reference dataset, HSATII copy ratios were estimated in the patients. Our analysis revealed several significant CNVs in HSATII, with certain sequences displaying notable gains and losses across all breast cancer patients, suggesting their potential as biomarkers. However, we observed pronounced fragmentation of cfDNA in cancer, leading to the loss of longer PCR amplicons (>180 bp). While not all observed losses can be attributed to fragmentation artifacts, this phenomenon does introduce complexity in interpreting CNV data. Notably, this research marks the first instance of targeted HSATII sequencing in a liquid biopsy context. Our findings lay the groundwork for developing sequencing-based assays to detect differentially represented HSATII sequences, potentially advancing the field of minimally-invasive cancer screening.

Impact of NDUFAF6 on breast cancer prognosis: linking mitochondrial regulation to immune response and PD-L1 expression

BackgroundBreast cancer is a major global health concern, and there is a continuous search for novel biomarkers to predict its prognosis. The mitochondrial protein NDUFAF6, previously studied in liver cancer, is now being investigated for its role in breast cancer. This study aims to explore the expression and functional significance of NDUFAF6 in breast cancer using various databases and experimental models.MethodsWe analyzed breast cancer samples from The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO), and Human Protein Atlas (HPA) databases, supplemented with immunohistochemistry (IHC) staining to assess NDUFAF6 expression. A breast cancer cell xenograft mouse model was used to evaluate tumor growth, apoptosis, and NDUFAF6 expression. Survival probabilities were estimated through Kaplan–Meier plots and Cox regression analysis. A Protein–Protein Interaction (PPI) network was constructed, and differentially expressed genes related to NDUFAF6 were analyzed using GO, KEGG, and GSEA. The relationship between NDUFAF6 expression, immune checkpoints, and immune infiltration was also evaluated.ResultsNDUFAF6 was found to be overexpressed in breast cancer patients and in the xenograft mouse model. Its expression correlated with worse clinical features and prognosis. NDUFAF6 expression was an independent predictor of breast cancer outcomes in both univariate and multivariate analyses. Functionally, NDUFAF6 is implicated in several immune-related pathways. Crucially, NDUFAF6 expression correlated with various immune infiltrating cells and checkpoints, particularly promoting PD-L1 expression by inhibiting the NRF2 signaling pathway.ConclusionThe study establishes NDUFAF6 as a potential prognostic biomarker in breast cancer. Its mechanism of action, involving the inhibition of NRF2 to upregulate PD-L1, highlights its significance in the disease's progression and potential as a target for immunotherapy.

Plasma Thioredoxin Reductase as a Potential Diagnostic Biomarker for Breast Cancer

BackgroundEarly diagnosis of breast cancer is critical to the treatment and prognosis of breast cancer patients. Our aim is to explore more practical and effective diagnostic methods to facilitate early treatment and improve prognosis for breast cancer patients. Materials and MethodsThe Mann–Whitney U test, receiver operating characteristic curve, Youden index, Chi-square test, and Fisher's exact test were used to determine whether plasma thioredoxin reductase (TrxR) could be used for the clinical diagnosis of breast cancer. The Wilcoxon signed-rank test was used to validate the prognostic potential of plasma TrxR activity assessment. ResultsA total of 761 patients were included, including 537 cases of breast cancer and 224 cases of benign breast diseases. Plasma TrxR activity in the breast cancer group [8.0 (6.0, 9.45) U/mL] was significantly higher than that in the benign group [3.05 (1.20, 6.275) U/mL]. The diagnostic efficiency of TrxR for breast cancer was higher than that of other conventional breast cancer biomarkers, with an area under the curve of 0.821 (95% CI = 0.791-0.852). In addition, TrxR can be used in combination with conventional tumor markers to further improve the diagnostic efficiency. The optimal TrxR threshold for identifying benign and malignant diseases is 7.45 U/mL. We detected plasma TrxR activity and serum tumor markers before and after antitumor therapies in 333 breast cancer patients and found that their trends were basically the same, with a significant decrease in plasma TrxR activity after treatment. ConclusionPlasma TrxR activity can be used as a suitable biomarker for breast cancer diagnosis and efficacy assessment.

Multi-targeted therapeutic potential of stigmasterol from the Euphorbia ammak plant in treating lung and breast cancer

Cancer is the most prevalent disease globally, which presents a significant challenge to the healthcare industry, with breast and lung cancer being predominant malignancies. This study used RNA-seq data from the TCGA database to identify potential biomarkers for lung and breast cancer. Tumor Necrosis Factor (TNFAIP8) and Sulfite Oxidase (SUOX) showed significant expression variation and were selected for further study using structure-based drug discovery (SBDD). Compounds derived from the Euphorbia ammak plant were selected for in-silico study with both TNFAIP8 and SUOX. Stigmasterol had the greatest binding scores (normalized scores of −8.53 kcal/mol and −9.69 kcal/mol) with both proteins, indicating strong stability in their binding pockets throughout the molecular dynamics’ simulation. Although Stigmasterol first changed its initial conformation (RMSD = 0.5 nm with the starting conformation) in SUOX, it eventually reached a stable conformation (RMSD of 1.5 nm). The compound on TNFAIP8 showed a persistent shape (RMSD of 0.35 nm), indicating strong protein stability. The binding free energy of the complex was calculated using the MM/GBSA technique; TNFAIP8 had a ΔGTOTAL of −24.98 kcal/mol, with TYR160 being the most significant residue, contributing −2.52 kcal/mol. On the other hand, the SUOX complex had a binding free energy of −16.87 kcal/mol, with LEU151 being the primary contributor (-1.17 kcal/mol). Analysis of the complexes' free energy landscape unveiled several states with minimum free energy, indicating robust interactions between the protein and ligand. In its conclusion, this work emphasises the favourable ability of Stigmasterol to bind with prospective targets for lung and breast cancer, indicating the need for more experimental study.

The presence of autoantibodies as a potential prognostic biomarker for breast cancer.

The presence of autoantibodies is closely associated with the occurrence and development of cancer. Autoantibodies can be used as biomarkers for early breast cancer diagnosis. However, the relationship between autoantibodies and the prognosis of breast cancer patients remains elusive. This retrospective study aimed to investigate the correlation between the presence of autoantibodies and outcomes in breast cancer patients. This study included a total of 155 patients from People's Hospital of Henan University (Zhengzhou, China) who were diagnosed with breast cancer from January 2017 to December 2021. The enrolled patients' clinicopathological features were collected, and 88 patients were ultimately involved in the analysis. Univariate and multivariate Cox regression analyses were performed to search for the risk factors related to the poor prognosis of breast cancer patients. The association between the presence of autoantibodies and patients' survival was analysed using Kaplan-Meier curves. After screening, there were 38 autoantibody-positive cases and 50 autoantibody-negative cases. Breast cancer patients with autoantibody-positive had a 57% lower risk of death compared with autoantibody-negative patients. Multivariate Cox regression analysis indicated that the presence of autoantibody could be a potential prognostic predictor for breast cancer, independent of age, histological grade, pathological classification, clinical stage, and the expression levels of hormonal receptors. In addition, autoantibody-positive breast cancer patients had longer progression-free survival and overall survival compared with autoantibody-negative cases. Positive autoantibody was found as an independent biomarker of better prognosis in breast cancer patients, providing a new strategy for the prognostic assessment of breast cancer patients.

MiRNA-449c-5p regulates the JAK-STAT pathway in inhibiting cell proliferation and invasion in human breast cancer cells by targeting ERBB2.

Breast cancer is a highly prevalent disease worldwide, and early diagnosis and treatment could reduce the mortality rate of breast cancer patients. microRNAs (miRNA) have been shown to regulate the occurrences and progression of many types of cancers. Thus, it is crucial to find novel biomarkers in breast cancer. miR-449c-5p acted as a biomarker in non-small cell lung cancer, gastric carcinoma, and so forth. ERBB2 is an ideal target for breast cancer therapy. However, the molecular mechanisms between miR-449c-5p and ERBB2 in breast cancer remain poorly understood. Our study focused on the regulatory role of miR-449c-5p in breast cancer and its targeting relationship with ERBB2. The miR-449c-5p expression in breast cancer tissue and normal tissue was searched from the online database (Starbase). The clinical prognosis of miR-449c-5p and ERBB2 was predicted by using the Kaplan-Meier analysis method. The expression of miR-449c-5p mimics and inhibitors was measured by qRT-PCR. T47D cells were transfected with miR-449c-5p mimics and miR-449c-5p inhibitors. After that, CCK-8, colony formation assays and Transwell assays were used to evaluate the cell proliferation ability, migration and invasion. Whether ERBB2 was the target gene of the miR-449c-5p was predicted by Starbase and verified by dual-luciferase activity assay. In addition, protein levels and the relationship between signalling pathways were measured and validated using western blotting analysis. We confirmed that miR-449c-5p was highly expressed in breast cancer tissue, and its downregulation was linked with poor prognosis. Overexpression of miR-449c-5p inhibited the proliferation, migration and invasion of breast cancer cells. ERBB2 was a target of miR-449c-5p. The invasion, migration, and proliferation of breast cancer cells were inhibited by miR-449c-5p/ERBB2 through JAK-STAT. This study demonstrated that miR-449c-5p inhibits breast cancer cell proliferation, migration and invasion by targeting ERBB2 via JAK/STAT, which means miR-449c-5p, is a potential biomarker for breast cancer and provides a novel insight for diagnosis.

Abstract A006: Spatial localization of Caveolin-1 protein in triple negative breast cancer is related to different molecular features

Abstract Background: Triple negative breast cancer (TNBC) is an aggressive and heterogeneous form of breast cancer with few targeted therapies and clinically used biomarkers. Caveolin-1 (CAV1) is a marker of stromal activation and vascularization. Molecular features of the tumor microenvironment (TME), important for tumor metastasis, may improve precision oncology in TNBC. CAV1 protein levels have previously been reported as a potential biomarker in breast cancer depending on their localization. Therefore, we aimed to investigate CAV1 protein levels in malignant and stromal cells of TNBC and their relation to CAV1 gene expression, clinicopathological factors, and molecular features. Methods: Tumor-specific CAV1 protein levels were evaluated with immunohistochemistry in malignant and stromal cells in tumor tissue microarrays from a cohort of 242 TNBC patients (inclusion 2010-2015, Sweden) from SCAN-B (NCT03758976). The protein levels of CAV1 in malignant and stromal cells were dichotomized as ‘strong’ (1) vs ‘negative to moderate’ (0) The CAV1 categories in malignant and stromal cells, respectively, were combined to create a joint CAV1 status with four categories: malignant/stromal cells, 0/0, 0/1, 1/0, and 1/1. Gene expression profiles from massive parallel paired-end sequencing of mRNA (RNA-seq) were available for all the tumors and from which PAM50 category and TNBC subtype were assigned. CAV1 status in malignant and stromal cells in relation to clinicopathological factors was analyzed using Chi-Square test. CAV1 mRNA expression in relation to CAV1 status in malignant and stromal cells was evaluated using Kruskal-Wallis test. Results: CAV1 status could be evaluated in 231/242 tumors. Strong CAV1 staining in malignant cells was associated with higher histological grade but no axillary lymph node involvement (both P &amp;lt;0.007). In contrast, strong CAV1 staining in stromal cells was associated with lower histological grade but axillary lymph node involvement (both P &amp;lt;0.001). Strong CAV1 staining in stromal cells was also associated with higher age at diagnosis (P =0.025). With regards to PAM50 subtypes, CAV1 in malignant cells was positively associated with the basal subtype while CAV1 in malignant cells was positively associated with the HER2 enriched subtype (P &amp;lt;0.001). Depending on localization, CAV1 expression was associated with different TNBCsubtypes. CAV1 in malignant cells was positively associated with the mesenchymal and negatively associated with the immunomodulatory subtype (P &amp;lt;0.001). CAV1 in stromal cells was positively associated with the luminal androgen (LAR) subtype (P &amp;lt;0.001). Neither CAV1 protein levels in malignant cells nor in stromal cells were correlated with CAV1 gene expression in the tumors, and the combined CAV1 status was not associated with CAV1 gene expression. Conclusion: CAV1 protein levels in malignant cells were related to different molecular features of TNBC compared to CAV1 protein levels in stromal cells. The results suggest that depending on localization, CAV1 protein levels function as two distinct biomarkers. Citation Format: Christopher Godina, Somayeh Khazaei, Mattias Belting, Johan Vallon-Christersson, Björn Nodin, Karin Jirström, Karolin Isaksson, Ana Bosch, Helena Jernström. Spatial localization of Caveolin-1 protein in triple negative breast cancer is related to different molecular features [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr A006.

Surface Proteome of Extracellular Vesicles and Correlation Analysis Reveal Breast Cancer Biomarkers

Breast cancer (BC) is the second most frequently diagnosed cancer and accounts for approximately 25% of new cancer cases in Canadian women. Using biomarkers as a less-invasive BC diagnostic method is currently under investigation but is not ready for practical application in clinical settings. During the last decade, extracellular vesicles (EVs) have emerged as a promising source of biomarkers because they contain cancer-derived proteins, RNAs, and metabolites. In this study, EV proteins from small EVs (sEVs) and medium EVs (mEVs) were isolated from BC MDA-MB-231 and MCF7 and non-cancerous breast epithelial MCF10A cell lines and then analyzed by two approaches: global proteomic analysis and enrichment of EV surface proteins by Sulfo-NHS-SS-Biotin labeling. From the first approach, proteomic profiling identified 2459 proteins, which were subjected to comparative analysis and correlation network analysis. Twelve potential biomarker proteins were identified based on cell line-specific expression and filtered by their predicted co-localization with known EV marker proteins, CD63, CD9, and CD81. This approach resulted in the identification of 11 proteins, four of which were further investigated by Western blot analysis. The presence of transmembrane serine protease matriptase (ST14), claudin-3 (CLDN3), and integrin alpha-7 (ITGA7) in each cell line was validated by Western blot, revealing that ST14 and CLDN3 may be further explored as potential EV biomarkers for BC. The surface labeling approach enriched proteins that were not identified using the first approach. Ten potential BC biomarkers (Glutathione S-transferase P1 (GSTP1), Elongation factor 2 (EEF2), DEAD/H box RNA helicase (DDX10), progesterone receptor (PGR), Ras-related C3 botulinum toxin substrate 2 (RAC2), Disintegrin and metalloproteinase domain-containing protein 10 (ADAM10), Aconitase 2 (ACO2), UTP20 small subunit processome component (UTP20), NEDD4 binding protein 2 (N4BP2), Programmed cell death 6 (PDCD6)) were selected from surface proteins commonly identified from MDA-MB-231 and MCF7, but not identified in MCF10A EVs. In total, 846 surface proteins were identified from the second approach, of which 11 were already known as BC markers. This study supports the proposition that Evs are a rich source of known and novel biomarkers that may be used for non-invasive detection of BC. Furthermore, the presented datasets could be further explored for the identification of potential biomarkers in BC.

Circulating tumor DNA as prognostic markers of relapsed breast cancer: a systematic review and meta-analysis

ObjectiveCirculating tumor DNA (ctDNA) is increasingly being used as a potential prognosis biomarker in patients of breast cancer. This review aims to assess the clinical value of ctDNA in outcome prediction in breast cancer patients throughout the whole treatment cycle. MethodsPubMed, Web of Science, Embase, Cochrane Library, Scopus, and clinical trials.gov were searched from January 2016 to May 2022. Conference abstracts published in last three years were also included. The following search terms were used: ctDNA OR circulating tumor DNA AND breast cancer OR breast carcinoma. Only studies written in English languages were included. The following pre-specified criteria should be met for inclusion: (1) observational studies (prospective or retrospective), randomized control trials, case-control studies and case series studies; (2) patients with breast cancer; (3) ctDNA measurement; (4) clinical outcome data such as objective response rate (ORR), pathological complete response (pCR), relapse-free survival (RFS), overall survival (OS), and so on. The random-effect model was preferred considering the potential heterogeneity across studies. The primary outcomes included postoperative short-term outcomes (ORR and pCR) and postoperative long-term outcomes (RFS, OS, and relapse). Secondary outcomes focused on ctDNA detection rate. ResultsA total of 30 studies, comprising of 19 cohort studies, 2 case-control studies and 9 case series studies were included. The baseline ctDNA was significantly negatively associated with ORR outcome (Relative Risk [RR] = 0.65, 95% confidence interval [CI]: 0.50–0.83), with lower ORR in the ctDNA-positive group than ctDNA-negative group. ctDNA during neoadjuvant therapy (NAT) treatment was significantly associated with pCR outcomes (Odds Ratio [OR] = 0.15, 95% CI: 0.04–0.54). The strong association between ctDNA and RFS or relapse outcome was significant across the whole treatment period, especially after the surgery (RFS: Hazard Ratio [HR] = 6.74, 95% CI: 3.73–12.17; relapse outcome: RR = 7.11, 95% CI: 3.05–16.53), although there was heterogeneity in these results. Pre-operative and post-operative ctDNA measurements were significantly associated with OS outcomes (pre-operative: HR = 2.03, 95% CI: 1.12–3.70; post-operative: HR = 6.03, 95% CI: 1.31–27.78). ConclusionsIn this review, ctDNA measurements at different timepoints are correlated with evaluation indexes at different periods after treatment. The ctDNA can be used as an early potential postoperative prognosis biomarker in breast cancer, and also as a reference index to evaluate the therapeutic effect at different stages.

High-contrast NIR fluorescent probes for selective detection of NQO1 in breast cancer

NAD(P)H:quinone oxidoreductase 1 (NQO1) is a potential biomarker for breast cancer (BC) diagnosis and prognosis. However, existing fluorescent probes for NQO1 detection have limitations such as short emission wavelength, weak fluorescence response, or large background interference. Here, we developed two novel near-infrared (NIR) fluorescent probes, DCl-Q and DCl2-Q, that selectively detect NQO1 activity in BC cells and tissues. They consist of a trimethyl-locked quinone as the recognition group and a donor-π-acceptor structure with halogen atoms as the reporter group. They exhibit strong fluorescence emission at around 660 nm upon binding to NQO1. We demonstrated that they can distinguish BC cells with different NQO1 expression levels and image endogenous NQO1 in tumor-bearing mice. Our probes provide a convenient and highly sensitive tool for BC diagnosis and prognosis based on NQO1 detection.

Comparative profiling by data-independent acquisition mass spectrometry reveals featured plasma proteins in breast cancer: a pilot study.

Breast cancer is known to be influenced by genetic and environmental factors, and several susceptibility genes have been discovered. Still, the majority of genetic contributors remain unknown. We aimed to analyze the plasma proteome of breast cancer patients in comparison to healthy individuals to identify differences in protein expression profiles and discover novel biomarkers. This pilot study was conducted using bioresources from Seoul National University Bundang Hospital's Human Bioresource Center. Serum samples from 10 breast cancer patients and 10 healthy controls were obtained. Liquid chromatography-mass spectrometry analysis was performed to identify differentially expressed proteins. We identified 891 proteins; 805 were expressed in the breast cancer group and 882 in the control group. Gene set enrichment and differential expression analysis identified 30 upregulated and 100 downregulated proteins in breast cancer. Among these, 10 proteins were selected as potential biomarkers. Three proteins were upregulated in breast cancer patients, including cluster of differentiation 44, eukaryotic translation initiation factor 2-α kinase 3, and fibronectin 1. Seven proteins downregulated in breast cancer patients were also selected: glyceraldehyde-3-phosphate dehydrogenase, α-enolase, heat shock protein member 8, integrin-linked kinase, tissue inhibitor of metalloproteinases-1, vasodilator-stimulated phosphoprotein, and 14-3-3 protein gamma. All proteins had been previously reported to be related to tumor development and progression. The findings suggest that plasma proteome profiling can reveal potential diagnostic biomarkers for breast cancer and may contribute to early detection and personalized treatment strategies. A further validation study with a larger sample cohort of breast cancer patients is planned.

CirculatingmicroRNAs as potential biomarkers in triple-negative breast cancer: a translational research study of the NACATRINE trial.

Liquid biopsy is increasingly vital in monitoring neoadjuvant breast cancer treatment. This study collected plasma samples at three time points from participants in the Neoadjuvant Carboplatin in Triple Negative Breast Cancer (NACATRINE), analyzing miRNA expression with NanoString's nCounter® Human v3 miRNA assay. In the carboplatin arm, four ct-miRNAs exhibited dynamic changes linked to pathologic complete response, with a combined area under the curve of 0.811. Similarly, the non-carboplatin arm featured four ct-miRNAs with an area under the curve of 0.843. These findings underscore the potential of ct-miRNAs as personalized tools in breast cancer treatment, assisting in predicting treatment response and assessing the risk of relapse. Integrating ct-miRNA analysis into clinical practice can optimize decisions and enhance patient outcomes.