Marker In Breast Cancer Research Articles

Abstract PO4-03-01: Enhancing Breast Cancer Subtyping: APIS Breast Cancer Subtyping Kit Proliferative Signature's Potential for Improved Proliferation Assessment and Recurrence Prediction

Abstract Background: Ki67, a widely recognized biomarker for measuring tumor cell proliferation in breast cancer (BC), plays a crucial role in classifying BC into luminal A and luminal B subtypes, subsequently aiding treatment decisions. However, the inconsistency in Ki67 assessment and the lack of standardization in methods and interpretation of immunohistochemistry (IHC) staining has hindered its recommendation for routine diagnostic use. The standardization of measurement techniques, especially for Ki67, holds immense potential for improving patient treatment outcomes. The APIS Breast Cancer Subtyping Kit (APIS BCS Kit) addresses these challenges by utilizing RT-qPCR to accurately determine the mRNA expression of BC markers, including Ki67, along with a novel four-gene proliferative signature, comprising four markers (MKI67, CCNA2, KIF23, and PCNA) associated with proliferation and expressed throughout all cell cycle stages, thereby providing a more accurate measure of tumor proliferation. In this study, we present the preliminary findings of our ongoing evaluation of the APIS BCS Kit proliferative signature's ability to distinguish between Luminal A and Luminal B subtypes and to determine the risk of recurrence in hormone receptor-positive (HR+) patients. Method: A total of N=153 retrospectively collected HR+/HER2 negative samples were provided by University of Basel. For each sample the following clinical annotations were included: relapse free survival, treatment regimen (Chemotherapy (ChT) or Hormone Therapy (HT)) as well as OncotypeDx (ODx) risk score (RS). N=40 samples underwent additional testing using PAM50 (Prosigna®) test. Only samples collected since 2018 were included, resulting in 10 observed relapse events (N=7 in HT cohort and N=3 in ChT cohort). RNA isolation from tissue sections and gene expression analysis were performed as per APIS BCS Kit instructions for use. The agreement of subtype calls between IHC, APIS BCS Kit, and PAM50 was assessed. The correlation between RS obtained from ODx, PAM50 and the APIS BCS Kit proliferation signature was also explored. Results: The agreement between IHC and the APIS BCS Kit subtype call, utilizing Ki67 alone as a measure of proliferation, was 67.3% (103/153). Replacing Ki67 with the APIS BCS Kit proliferation signature increased the concordance to 69.9% (107/153). The concordance between IHC and PAM50 subtype calls was comparable at 62.5% (25/40). The agreement between the APIS BCS Kit and PAM50 subtype call was notably higher (80%; 32/40). Significant association was observed between the APIS BCS Kit proliferation signature and ODx RS (P=.0001), and PAM50 RS (P=.002). In the HT cohort, both ODx and the APIS BCS Kit proliferation signature accurately predicted recurrence (by assigning high RS) in n=2 cases. In some cases both the APIS BCS Kit and ODx erroneously assigned low risk and subsequently missed n=4 recurrence events. Notably, APIS BCS Kit proliferation signature correctly identified one recurrence event (high RS), which was missed by ODx (low RS). Conclusion: The APIS Breast Cancer Subtyping Kit exhibits improved agreement with the PAM50 molecular subtype in contrast to the IHC-derived subtype, indicating that molecular testing may be a more suitable method for determining proliferation and distinguishing between luminal A and B subtypes. Furthermore, there is a significant correlation between the RS generated by APIS Breast Cancer Subtyping Kit proliferation signature and both PAM50 and Oncotype RS. These initial findings indicate that the APIS Breast Cancer Subtyping Kit proliferation signature has the potential to accurately identify patients at risk of recurrence, although additional studies including a larger proportion of events are needed to validate these results. Citation Format: Marcus Vetter, Anna Gasior, Elena-Diana Chiru, Joanna Gorniak, Leanne Gough, Mathew Harrison, Sara Rollinson, Simone Muenst, Christian Kurzeder. Enhancing Breast Cancer Subtyping: APIS Breast Cancer Subtyping Kit Proliferative Signature's Potential for Improved Proliferation Assessment and Recurrence Prediction [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 PO4-03-01.

Vascular Endothelial Growth Factor (VEGF) and Its Role in the Progression and Development of Breast Tumor

Breast cancer is the most common type of cancer in women and is widely known. Angiogenesis is the process through which new blood vessels develop from the body's current vascular system. It is essential for tumor growth, invasiveness, and metastasis, and so plays a pivotal role in the development of carcinoma. Proteolytic and proangiogenic catalyst activators and inhibitors regulate angiogenesis in a hierarchical fashion. The angiogenic process is largely under the direction of VEGF. In a select number of malignant tumors, the VEGF gene is overexpressed. The function of VEGF in angiogenesis has been the subject of intensive study in recent years. In breast cancer patients, VEGF plasma levels are highly predictive of tumor growth and survival. Several VEGF gene polymorphisms, have been identified to affect gene expression level in prior investigations. Epidemiological studies have linked polymorphisms in the VEGF gene to altered cancer risk, tumor growth, and metastasis. Previous research on VEGF polymorphism to evaluate the association between genes and breast cancer susceptibility was scant. The current review discusses the role of VEGF in the progression of breast cancer in addition to its promising usage as a predictive marker for breast cancer.

Spectroscopic insight into breast cancer: profiling small extracellular vesicles lipids via infrared spectroscopy for diagnostic precision

Breast cancer, a leading cause of female mortality due to delayed detection owing to asymptomatic nature and limited early diagnostic tools, was investigated using a multi-modal approach. Plasma-derived small EVs from breast cancer patients (BrCa, n = 74) and healthy controls (HC, n = 30) were analyzed. Small EVs (n = 104), isolated through chemical precipitation, underwent characterization via transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA). Validation involved antibody-based tests (TSG101, CD9, CD81, CD63). Infrared spectra of small EVs were obtained, revealing significant differences in lipid acyl chains, particularly in the C–H stretching of CH3. The study focused on the lipid region (3050–2900 cm−1), identifying peaks (3015 cm−1, 2960 cm−1, 2929 cm−1) as distinctive lipid characteristics. Spectroscopic lipid-to-lipid ratios [(I3015/I2929), (I2960/I2929)] emerged as prominent breast cancer markers. Exploration of protein, nucleic acid, and carbohydrate ratios indicated variations in alpha helices, asymmetric C–H stretching vibrations, and C–O stretching at 1033 cm−1. Principal component analysis (PCA) successfully differentiated BrCa and HC small EVs, and heatmap analysis and receiver operating characteristic (ROC) curve evaluations underscored the discriminatory power of lipid ratios. Notably, (I2960/I2929) exhibited 100% sensitivity and specificity, highlighting its potential as a robust BrCa sEV marker for breast cancer detection.

SLC35A2 expression is associated with HER2 expression in breast cancer

The role of SLC35A2 in breast cancer remains poorly understood, with limited available information on its significance. This study aimed to investigate the expression of SLC35A2 and clinicopathological variables in breast cancer patients. Immunohistochemical analysis of SLC35A2 protein was conductedon 40 adjacent non-neoplastic tissues and 320 breast cancer tissues. The study also assesed the association between SLC35A2 expression and breast cancer clinicopathological features of breast cancer, as well as its impact on overall survival. In comparison to adjacent non-neoplastic tissues, a significantly higher expression of SLC35A2 was observed in breast cancer tissues (P = 0.020), and this expression was found to be independently correlated with HER2 positivity (P = 0.001). Survival analysis indicated that patients with low SLC35A2 expression had a more favorable prognosis in HER2-positive subtype breast cancer (P = 0.017). These results suggest that SLC35A2 is overexpressed in breast cancer tissues compared to adjacent non-neoplastic tissues and may serve as a potential prognostic marker for HER2-positive subtype breast cancer. Furthermore, breast cancer patients with the HER2 positive subtype who exhibited decreased levels of SLC35A2 expression demonstrated improved long-term prognostic outcomes.

The Utility of [18]F-Fluorocholine Positron Emission Computed Tomography and [18]F-Fluorodeoxyglucose Positron Emission Tomography-Computed Tomography in Evaluating Breast Cancer Phenotypes: A Pilot Study.

The utility of the [18]F fluorodeoxyglucose positron emission tomography-computed tomography ([18]F FDG PET-CT) marker for breast cancer is well established. Given its limitations in localizing FDG-negative malignant tumors, the expression of [18]F-fluorocholine ([18]-FCH) may potentially be helpful to improve the overall accuracy in evaluating breast cancer. This study determined the potential of [18]- FCH PET CT as a potential marker in assessing breast cancer phenotypes. We recruited consecutive patients with biopsy-proven breast carcinoma who underwent [18] F-FCH PET-CT following the [18]F-FDG PET-CT imaging. The subjects were dichotomized into human epidermal growth factor receptor 2 (HER2)-negative and HER2-positive genotypes. The maximum standardized uptake value (SUVmax; g/dL) was used to predict the two groups of variables. Global health status (GHS) score based on the EORTC quality of life questionnaire (QLQ) was used to evaluate the outcome of the cohort subjects at 6, 12, and 24 months. There were 21 females with a mean age of 54.48 ± 12.17 years. Eighteen patients had invasive ductal carcinoma (18/21;85.8%) on histology, with 11 (52.4%) were HER2-negative genotype. There was higher sensitivity and specificity of [18]-FCH-PET/CT in breast lesions at 40% and 68.8% compared to [18]FDGPET/CT with 33.3% and 66.7%, respectively. There were significant differences between [18]F-FCH SUVmax (g/dL) of the HER-negative as compared to the HER2- positive group (1.99 g/dL vs. 0.2 g/dL; P < .05). High SUVmax (g/dL) of [18]F-FCH had predicted the HER-negative genotype at the cutoff value of 0.75 (P < .05). High [18]F-FCH showed significantly poor scoring of GHS parameters compared to low FCH at 6 months (mean SUVmax 8.06 vs. 5.40 respectively; P < .05). [18]F-FCH PET-CT is a potential marker in localizing and predicting aggressive breast carcinoma phenotypes.

TRPS1 is a Highly Sensitive Marker for Breast Cancer: A Tissue Microarray Study Evaluating More Than 19,000 Tumors From 152 Different Tumor Entities.

Trichorhinophalangeal syndrome 1 (TRPS1) is a nuclear protein highly expressed in breast epithelial cells. TRPS1 immunohistochemistry (IHC) has been suggested as a breast cancer marker. To determine the diagnostic and prognostic utility of TRPS1 IHC, tissue microarrays containing 19,201 samples from 152 different tumor types and subtypes were analyzed. GATA3 IHC was performed in a previous study. TRPS1 staining was seen in 86 of 152 tumor categories with 36 containing at least one strongly positive case. TRPS1 staining predominated in various types of breast carcinomas (51%-100%), soft tissue tumors (up to 100%), salivary gland tumors (up to 46%), squamous cell carcinomas (up to 35%), and gynecological cancers (up to 40%). TRPS1 positivity occurred in 1.8% of 1083 urothelial neoplasms. In invasive breast carcinoma of no special type, low TRPS1 expression was linked to high grade ( P = 0.0547), high pT ( P < 0.0001), nodal metastasis ( P = 0.0571), loss of estrogen receptor and progesterone receptor expression ( P < 0.0001 each), and triple-negative status ( P < 0.0001) but was unrelated to patient survival ( P = 0.8016). In squamous cell carcinomas from 11 different sites, low TRPS1 expression was unrelated to tumor phenotype. Positivity for both TRPS1 and GATA3 occurred in 47.4% to 100% of breast cancers, up to 30% of salivary gland tumors, and 29 (0.3%) of 9835 tumors from 134 other cancer entities. TRPS1 IHC has high utility for the identification of cancers of breast (or salivary gland) origin, especially in combination with GATA3. The virtual absence of TRPS1 positivity in urothelial neoplasms is useful for the distinction of GATA3-positive urothelial carcinoma from breast cancer.

Exosomal miRNA-92a derived from cancer-associated fibroblasts promote invasion and metastasis in breast cancer by regulating G3BP2

Cancer-associated Fibroblasts (CAFs) exert a tumor-promoting effect in various cancers, including breast cancer. CAFs secrete exosomes containing miRNA and proteins, influencing the tumor microenvironment. In this study, we identified CAF-derived exosomes that transport functional miR-92a from CAFs to tumor cells, thereby intensifying the aggressiveness of breast cancer. CAFs downregulate the expression of G3BP2 in breast cancer cells, and a significant elevation in miR-92a levels in CAF-derived exosomes was observed. Both in vitro and in vivo experiments demonstrate that miR-92a enhances breast cancer cell migration and invasion by directly targeting G3BP2, functioning as a tumor-promoting miRNA. We validated that the RNA-binding proteins SNRPA facilitate the transfer of CAF-derived exosomal miR-92a to breast cancer cells. The reduction of G3BP2 protein by CAF-derived exosomes releases TWIST1 into the nucleus, promoting epithelial-mesenchymal transition (EMT) and further exacerbating breast cancer progression. Moreover, CAF-derived exosomal miR-92a induces tumor invasion and metastasis in mice. Overall, our study reveals that CAF-derived exosomal miR-92a serves as a promoter in the migration and invasion of breast cancer cells by reducing G3BP2 and may represent a potential novel tumor marker for breast cancer.

Involvement of FAM170B-AS1, hsa-miR-1202, and hsa-miR-146a-5p in breast cancer.

FAM170B-AS1 is usually expressed low in all organs except for testicular tissues. No study was performed to explore its role in breast cancer (BC). Contradictory results were reported about hsa-miR-1202 and hsa-miR-146a-5p in BC. The present study aimed to explore the involvement of FAM170B-AS1 in BC using bioinformatics predictive tools, followed by a practical validation besides exploring the impact of hsa-miR-1202 and hsa-miR-146a-5p in BC. This study enrolled 96 female patients with BC, 30 patients with benign breast diseases (BBD), and 25 control subjects. The expressions of circulating FAM170B-AS1, hsa-miR-1202, and hsa-miR-146a-5p were quantified using qRT-PCR. These ncRNAs' associations, predictive, and diagnostic roles in BC were statistically tested. The underlying miRNA/mRNA targets of FAM170B-AS1 in BC were bioinformatically predicted followed by confirmation based on the GEPIA and TCGA databases. The expression of FAM170B-AS1 was upregulated in sera of BC patients and hsa-miR-1202 was upregulated in sera of BBD and BC patients while that of hsa-miR-146a-5p was downregulated in BC. These FAM170B-AS1 was significantly associated with BC when compared to BBD. FAM170B-AS1 and hsa-miR-1202 were statistically associated with the BC's stage, grade, and LN metastasis. FAM170B-AS1 and hsa-miR-146a-5p gave the highest specificity and sensitivity for BC. KRAS and EGFR were predicted to be targeted by FAM170B-AS1 through interaction with hsa-miR-143-3p and hsa-miR-7-5p, respectively. Based on the TCGA database, cancer patients having mutations in FAM170B show good overall survival. The present study reported that for the first time, FAM170B-AS1 may be a potential risk factor, predictive, and diagnostic marker for BC. In addition, FAM170B-AS1 might be involved in BC by interacting with hsa-miR-143-3p/KRAS and hsa-miR-7-5p/EGFR through enhancement or repression that may present a new therapeutic option for BC.

Exploring research progress in studying serum exosomal miRNA-21 as a molecular diagnostic marker for breast cancer.

Breast cancer is one of the most prevalent malignancies affecting women globally and poses a significant public health challenge. Early clinical detection plays a pivotal role in providing optimal treatment opportunities and favorable prognoses, crucial for reducing breast cancer mortality and enhancing patients' quality of life. Therefore, the timely identification and diagnosis of breast cancer are imperative. Conventional tumor markers, such as carcinoembryonic antigen (CEA) and carbohydrate antigen 15-3 (CA15-3), serve as reliable methods for actively monitoring disease progression and have become a routine auxiliary diagnostic approach in clinical settings. However, these biomarkers exhibit limitations in sensitivity and specificity, particularly in the early screening and diagnosis of tumors, often yielding results inconsistent with clinical manifestations. In recent years, research has increasingly focused on exosomes released by tumor cells as potential new biomarkers for early stage breast cancer screening. Exosomes carry various components, including tumor-derived proteins, nucleic acids, and lipids. This paper delves into the specific utilization of serum exosomal microRNA-21 (miR-21) as a biomarker for early detection and diagnosis of breast cancer, evaluating its efficacy within this framework.

Expression of Cathepsin D in early-stage breast cancer and its prognostic and predictive value

PurposeCathepsin D is a proteolytic enzyme that is normally localized in the lysosomes and is involved in the malignant progression of breast cancer. There are conflicting results regarding Cathepsin D significance as prognostic and predictor marker in breast cancer. This study aimed to evaluate the expression and prognostic significance of Cathepsin D in early-stage breast cancer.MethodsExpression of Cathepsin D was assessed by immunohistochemical staining of tissue microarrays, in a large well-characterized series of early-stage operable breast cancer (n = 954) from Nottingham Primary Breast Carcinoma Series between the period of 1988 and 1998 who underwent primary surgery. Correlation of Cathepsin D expression with clinicopathological parameters and prognosis was evaluated.ResultsCathepsin D expression was positive in 71.2% (679/954) of breast cancer tumours. Positive expression of Cathepsin D was significantly associated with high histological grade (p = 0.007), pleomorphism (p = 0.002), poor Nottingham Prognostic Index (NPI) score (p < 0.002), recurrence (p = 0.005) and distant metastasis (p < 0.0001). Kaplan–Meier analysis showed that Cathepsin D expression was significantly associated with shorter breast cancer-specific survival (p = 0.001), higher risk of recurrence (p = 0.001) and distant metastasis (p < 0.0001). ER-positive tumours expressing Cathepsin D and treated with tamoxifen demonstrated a significantly higher risk of distant metastasis.ConclusionCathepsin D expression significantly predicts poor prognosis in breast cancer and is associated with variables of poor prognosis and shorter outcome. The strong association of Cathepsin D with aggressive tumour characteristics and poor outcomes warrants further research of its potential as a therapeutic target The results also suggest a possible interaction between Cathepsin D and tamoxifen therapy in ER-positive breast cancer which needs further investigation to elucidate the underlying mechanisms.

Abstract LB392: Transformative pathomics in oncology: Harnessing FFPE tissue slide imagery for clinically relevant gene expression prediction in invasive breast carcinoma

Abstract Purpose: Pathologists routinely analyze H&amp;E-stained FFPE tissue slides microscopically to arrive at diagnoses for patients. Pathomics, the study of quantitative imaging from such samples, aims to elevate diagnostic accuracy by revealing intricate tissue and cellular details. Our research evaluates how pathomic features from tissue imagery can predict gene expression, as determined by RNA sequencing, offering advancements in the molecular profiling and targeted therapy for invasive breast carcinoma (IBC). Method: We conducted an analysis of 90 regions of interest (ROIs) on FFPE tissue slide images from the TCGA registry for patients with IBC. Using HistomicsTK software, we extracted nearly 300 pathomic features depicting cell morphometry, intensity, and gradient from these regions of interest (ROIs). We then assessed the intra-correlation of the pathomic features within the ROIs of the same tissue slide to identify the most heterogeneous and highly correlating features (n=20) across categories—morphometry, intensity, and gradient—with a Pearsonʼs Correlation coefficient (r) greater than 0.9 and a FDR adjusted p-value less than 0.05. Subsequently, gene expression data in the form of FPKM were obtained for the corresponding tissues from TCGA. We trained a Multitask Elastic Net (MTEN) model on these pathomic features using an 80:20 split for the training and testing sets, with a three-fold cross-validation applied to the training set utilizing ImaGene software. The model's performance was evaluated by measuring the AUC and R² value for the predicted gene expression on the testing set. Result and Biological significance: The testing of the MTEN model using a testing set predicted the expression of three genes identified in the literature as prognostic markers in breast cancer, namely ALDH1L2, MFAP5, and MXRA8, with an AUC greater than 0.8 and an R² above 0.5 at a p-value of less than 0.002. According to the literature, expression of genes from the ALDH1 superfamily in breast cancer correlates with the stage of the disease, triple-negative status, and response to neoadjuvant therapy. The upregulation of MFAP5 in invasive breast carcinomas has been associated with high risk prognostic features, such as higher tumor grade and stage and increased angiogenesis, and poorer outcomes, such as lymph node metastasis. Furthermore, MXRA8 is involved in modulating the progression of human triple-negative breast cancer, likely through its influence on the interactions of tumor cells with their microenvironment. Conclusion: The present study predicts the expression of clinically relevant genes in IBC using a heterogeneous set of pathomic features extracted from FFPE tissue slide imagery. By linking cell and tissue-based morphometric, intensity, and gradient features with gene expression, researchers can gain insights into the molecular mechanisms underlying disease progression. Digital pathology and pathomics may reduce the need for additional genetic testing in critical patient-cases by providing predictive information from routinely acquired pathology slides. Bridging the gap between phenotypic tissue data and molecular data, the predictive capabilities of pathomic features represent a significant advancement in the field of precision medicine. Citation Format: Shrey S. Sukhadia, Digvijay Yadav, Kristen E. Muller. Transformative pathomics in oncology: Harnessing FFPE tissue slide imagery for clinically relevant gene expression prediction in invasive breast carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB392.

Clinical and Analytical Validation of Two Methods for Ki-67 Scoring in Formalin Fixed and Paraffin Embedded Tissue Sections of Early Breast Cancer.

Proliferation determined by Ki-67 immunohistochemistry has been proposed as a useful prognostic and predictive marker in breast cancer. However, the clinical validity of Ki-67 is questionable. In this study, Ki-67 was retrospectively evaluated by three pathologists using two methods: a visual assessment of the entire slide and a quantitative assessment of the tumour margin in 411 early-stage breast cancer patients with a median follow-up of 26.8 years. We found excellent agreement between the three pathologists for both methods. The risk of recurrence for Ki-67 was time-dependent, as the high proliferation group (Ki-67 ≥ 30%) had a higher risk of recurrence initially, but after 4.5 years the risk was higher in the low proliferation group. In estrogen receptor (ER)-positive patients, the intermediate Ki-67 group initially followed the high Ki-67 group, but eventually followed the low Ki-67 group. ER-positive pN0-1 patients with intermediate Ki-67 treated with endocrine therapy alone had a similar outcome to patients treated with chemotherapy. A cut-off value of 20% appeared to be most appropriate for distinguishing between the high and low Ki-67 groups. To summarize, a simple visual whole slide Ki-67 assessment turned out to be a reliable method for clinical decision-making in early breast cancer patients. We confirmed Ki-67 as an important prognostic and predictive biomarker.

Long-Noncoding-RNA HOTAIR Upregulation is Associated with Poor Breast Cancer Outcome: A Systematic Review and Meta Analysis.

Breast cancer is the most frequent cancer among women worldwide with significant disproportionate mortality rates in developing countries. Although clinical management of breast cancer has been immensely improved, refinement in the prognostic and recurrent markers is still needed. Long non-coding RNAs (lncRNA) HOTAIR has recently been associated with poor outcome and is potentially used as a prognostic marker in breast cancer. We comprehensively reviewed studies evaluating lncRNA HOTAIR in association with overall and disease-free survivals in breast cancers. Systematic searches were performed in Pubmed, ProQuest, ScienceDirect, Scopus, Google Scholar, Semantic Scholar, Springer, Nature, Sage Journals, and Wiley databases using combination keywords "long non-coding RNA," "lncRNA," "HOX transcript antisense RNA," "HOTAIR," "breast can-cer," "carcinoma mammae," "prognosis," and "survival." Risk of bias score was used to assess quality of studies, I2 test was conducted to assess heterogeneity. Meta-analysis was performed to compare HOTAIR expression with breast cancer survival rates using STATA v.17 software. Of the total 1,504 screened studies, seven studies were included in the meta-analysis involving 533 patients. High expression of HOTAIR was associated with poor survival rates (pooled HR: 1.69; 95%CI: 1.11-2.59; p=0.015), shorter overall survival (OS) (pooled HR: 1.33; 95%CI: 0.78-2.26; p=0.455), poor disease-free survival (DFS) (pooled HR: 2.40; 95%CI: 1.63-3.53; p<0.001), poor distant metastatic-free survival (MFS) (HR: 1.75; 95%CI: 1.13-2.71; p=0.012). In addition, overexpression of HOTAIR was associated with positive lymph node infiltration (pooled OR: 2.38; 95%CI: 0.53-10.69; p=0.26) and ductal type cancer (pooled OR: 3.27; 95%CI: 1.15-9.30; p=0.03). Upregulation of lncRNA HOTAIR is associated with worse DFS aand MFS that can potentially be used as a prognostic marker in breast cancer patients.

Expression and Significance of LINC02418 in Breast Cancer.

The complicated pathogenesis and poor prognosis of breast cancer have become a major difficulty in medical research. This study aims to explore new lncRNA as prognostic markers for breast cancer and explore their roles and molecular mechanisms to lay a foundation for the treatment of cancer patients. The expression of LINC02418 and miR-766-5p in breast cancer tissues and cells was first identified using polymerase chain reaction, and Pearson was used to examine the correlation between the two. The cancer cells activities under different transfection conditions were detected using the Transwell assay and CCK8 assay. The correlation between LINC02418 and patient prognosis was analyzed using multifactor Cox regression and Kaplan-Meier. It was shown that LINC02418 expression was upregulated in breast cancer tissues and cells. There are significant differences in lymph node metastasis and TNM stage between high and low LINC02418 expression groups. The higher the expression of LINC02418, the higher the mortality rate of breast cancer patients. miR-766-5p expression was downregulated and negatively correlated with LINC02418. There are binding sites between LINC02418 and miR-766-5p; Transfection with miR-766-5p inhibitor boosted LINC02418 luciferase activity, but transfection with miR-766-5p mimic decreased it. Knockdown of LINC02418 promoted miR-766-5p expression and inhibited cancer progression, which was alleviated to some extent by transfection with miR-766-5p inhibitors. LINC02418 has the potential to serve as a poor prognostic marker for breast cancer and plays a pro-oncogenic role by targeting miR-766-5p.

The Influence of Tumor-Specific Markers in Breast Cancer on Other Blood Parameters.

Breast cancer is the most frequently diagnosed cancer among women, responsible for the highest number of cancer-related deaths worldwide. There is limited data available related to serum tumor markers in breast cancer and other blood parameters or other glandular laboratory parameters. This study aims to evaluate the correlation of tumor-specific markers for breast cancer with other blood parameters and how these correlations could impact clinical management. This retrospective study represents a data analysis from 1 January 2020 to 31 May 2023, in the County Hospital of Timisoara, Romania. We reviewed all the cases where, in the laboratory analyses, the serum tumor specific biomarkers for breast cancer were analyzed. A statistical analysis was performed in order to identify a possible relationship between CA 15-3 and the various biomarkers and blood parameters included in the present study. Values were classified according to reference ranges. The tests revealed no statistically significant associations between CA 15-3 values and the levels of CA125 (χ2(1) = 1.852, p = 0.174), CEA (χ2(1) = 1.139, p = 0.286), AFP (Fisher's exact test, p = 0.341), fT4 (Fisher's exact test, p = 0.310), TSH (Fisher's exact test, p = 0.177), or PTH (Fisher's exact test, p = 0.650). The findings indicate a lack of strong correlation between CA 15-3 and CA125, CEA, AFP, thyroid function markers, or PTH within this cohort.

Is fibroblast growth factor 11 (FGF11) a predictive marker for breast cancer?

The prognostic role of fibroblast growth factor 11 (FGF11) has only been reported in cancers such as nasopharyngeal carcinoma and prostate cancer. The role of FGF11 in breast cancer is not fully known. It was aimed to compare FGF11 expression levels in de novo metastatic hormone receptor-positive, human epidermal reseptor-2-negative breast tumor tissue and healthy breast tissue and investigate the effect of the FGF11 expression on survival in breast cancer patients. To determine the FGF11 expression rate, breast tumor tissue of breast cancer patients diagnosed by breast biopsy and healthy breast tissue of healthy individuals who underwent breast biopsy due to benign lesions were used. The study population included 38 breast cancer patients and 24 healthy controls. The number of patients with a FGF11 expression level score of 1 (15.8% vs 12.5%), score of 2 (18.4% vs 12.5%), and score of 3 (31.6% vs 0%) was significantly higher in the patient group compared to the healthy control group. The median overall survival and progression-free survival were numerically better in the group with a FGF11 expression score of 0 to 1 than the group with a FGF11 expression score of 2 and 3, but this difference was not statistically significant. FGF11 may be a predictive marker for breast cancer formation. Additionally, with new FGF11-targeted treatment agents to be developed, endocrine resistance may be reduced, and better survival results may be achieved in hormone receptor-positive, human epidermal reseptor-2-negative breast cancer.

Identification of miR-125a and miR-106b signature as a potential diagnostic biomarker in breast cancer tissues

MicroRNAs (miRNAs) have essential roles in the etiology of breast cancer and are regarded as possible markers in this malignancy. In order to find new markers for breast cancer, the current study has measured expression level of four miRNAs, namely miR-125a, miR-106b, miR-96 and miR-92a-3p in the paired breast samples. Expression levels of miR-125a and miR-106b were higher in tumoral tissues compared with control tissues (Expression ratios (95% CI) = 4.01 (1.96–8.19) and 3.9 (1.95–7.81); P values = 0.0005 and 0.0003, respectively). miR-106b and miR-125a differentiated between malignant and non-malignant tissues with AUC values of 0.7 and 0.67, respectively. We detected association between expression of miR-106b and clinical stage (P = 0.03), in a way that its expression was the lowest in the advanced stages. Finally, significant relationships were found between miR-96 and miR-125a in both tumoral and non-tumoral specimens (ρ = 0.76 and 0.69, respectively). This nonparametric measure of rank correlation also showed relationship between miR-106b and miR-96 in both sets of samples (ρ = 0.63 and 0.61, respectively). Cumulatively, the assessed miRNAs, particularly miR-125a and miR-106b are putative targets for further expression and functional assays in breast cancer.

Abstract 3486: Triple hormone receptor signatures as novel prognostic markers in breast cancer

Abstract Background: Tumor phenotype across various cancers, including breast cancer, is predominantly shaped by a synergy between genomic alterations and the cell-of-origin from which the tumor originates. These elements collectively influence crucial aspects like tumor aggressiveness, treatment response, and patient prognosis. While existing research highlights that the role of cell-of-origin in shaping the molecular architecture of breast cancer, leveraging this information for actionable clinical insights has proven challenging. Here, we introduce two robust models leveraging cellular ancestry signatures of breast luminal cells to improve patient stratification and outcome prediction. Methods: In this study, we developed a unique gene signature anchored in the expression levels of triple hormone receptors (THR)—namely androgen (AR), estrogen (ER), and vitamin D (VDR)—in breast cancer cells. Building on this, we formulated two distinct mRNA markers, THR-50 and THR-70, aimed at categorizing breast tumors based on their THR expression profiles. These markers were rigorously validated across 65 independent breast cancer studies, involving a total of 6,679 patients, utilizing Kaplan-Meier survival curves, Cox proportional hazards models, and unsupervised clustering analyses. Results: Our findings indicate that both THR-50 and THR-70 are robustly associated with overall survival and recurrence-free survival in breast cancer patients across all datasets evaluated. Importantly, these THR signatures demonstrate broad applicability across various breast cancer subtypes, grades, and treatment phases—unlike existing prognostic markers that tend to be subtype-specific. Additionally, the THR signatures have revealed four unique patient clusters with divergent survival outcomes, three of which are ER-positive and one is ER-negative. Additionally, we developed an immune-based signature (i20) which can further categorize patients within the ER-negative cluster into two subgroups: one with a highly favorable prognosis, comparable to the ER-positive subtypes, and another that is characterized by exceedingly poor survival outcomes. Conclusion: The THR-based cellular ancestry signatures open a new avenue in understanding the intricacies of breast cancer biology. These signatures offer a robust and stable framework for developing other prognostic markers, thus providing an enhanced stratification of existing breast cancer categories as well as creating new classifications with more distinct survival estimates. Citation Format: Mohamed Omar, J. Chuck Harrell, Rulla M. Tamimi, Luigi Marchionni, Tan Ince. Triple hormone receptor signatures as novel prognostic markers in 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 3486.

Abstract 650: Pim3 kinase is a poor prognostic marker and novel molecular target for triple negative breast cancer

Abstract Triple-negative Breast cancer (TNBC) is characterized by the absence of ER, PR, and HER-2 receptors and is associated with poor prognosis, early relapse, and distant metastasis. In neoadjuvant setting, most patients (~50-70%) with TNBC do not achieve complete remission. Currently, there is no effective targeted therapy for TNBC patients. Therefore, identifying novel molecular targets and developing alternative therapeutic strategies are urgently needed. PIM3 is a serine/threonine kinase and is upregulated in leukemia and various solid cancers. Although PIM1 has been shown to play a critical role in cell proliferation in cMyc-enriched TNBC tumors, the role of PIM3 in TNBC is not known. To determine the clinical significance of PIM3 in TNBC patient prognosis and progression, we performed a Kaplan-Meier analysis in the TCGA TNBC patient database. We found that PIM3 expression is significantly associated with shorter patient survival in TNBC patients (n=77, p=0.0088). PIM3 protein is highly expressed in TNBC cells compared to normal breast epithelium and other ER+ and HER2+ breast cancer subtypes. Inhibition of PIM3 by siRNA significantly suppressed TNBC cell proliferation, migration, and invasion of TNBC cells including MDA-MB-231, MDA-MB-436, and BT20 cells. Lenti-based PIM3 overexpression induced TNBC cell proliferation, invasion, and migration. Non-biased reverse phase protein array (RPPA) assay and Western blot analysis showed that PIM3 inhibition markedly reduced the expression of EF2 Kinase protein, and we found it to be associated with shorter patient survival and poor prognosis. Immunoprecipitation of PIM3 indicated that it forms a heterodimer with EF2K whose inhibition by siRNA also suppressed TNBC cell proliferation, migration, invasion, and tumor growth. Furthermore, PIM3 inhibition by siRNA significantly enhanced the antiproliferative effect of Doxorubicin, which is one of the first-line chemotherapeutics used in patients. Moreover, in vivo therapeutic delivery of PIM3 siRNA loaded albumin nanoparticles (ALNPs-PIM3 siRNA) markedly delayed tumor growth of two different TNBC tumor xenograft models (MDA-MB-231 and MDA-MB-436) in mice. Analysis of tumors by IHC after 4 weeks of treatment demonstrated that ALNP-PIM3 siRNA treatment led to inhibition of intra-tumoral proliferation and induction of apoptosis. Currently, we are conducting an in vivo study in TNBC tumor models by combining ALNP-PIM3 siRNA therapy with doxorubicin. Overall, our studies suggest that PIM3 is a marker for poor patient survival that drives TNBC tumor growth and progression and a novel potential molecular target in TNBC. Citation Format: Pinar Atalay, Goknur Kara, Rumeysa Ozyurt, Nermin Kahraman, Bulent Ozpolat. Pim3 kinase is a poor prognostic marker and novel molecular target for 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 650.

Multi-omics analysis reveals the unique landscape of DLD in the breast cancer tumor microenvironment and its implications for immune-related prognosis

BackgroundCuproptosis, i.e., copper-induced programmed cell death, has potential implications in cancer therapy. However, the impact of the cuproptosis-related gene (CRG) dihydrolipoyl dehydrogenase (DLD) on breast cancer (BC) prognosis remains underexplored. MethodsWe employed real-time quantitative PCR and multiplexed immunostaining techniques to quantify DLD expression in both BC and the adjacent non-cancerous tissues. Immunofluorescence analysis was employed to assess the influence of DLD on immune cells and immunological checkpoints in the BC microenvironment. DLD knockdown experiments were conducted in BC cell lines MDA-MB-468 and SK-BR-3, with knockdown efficiency validated via western blot. Subsequently, we performed the cell counting kit-8 (CCK-8) assay, clone formation assay, Transwell migration assay, and invasion assay. To construct a prognostic model, we employed a Lasso-Cox regression analysis of immune-related genes associated with DLD. Additionally, we established a competing endogenous RNA network based on CRGs to evaluate potential regulatory pathways. ResultsCompared to the adjacent tissues, BC tissues exhibited markedly elevated DLD expression levels. In vitro experiments demonstrated that DLD knockdown effectively inhibited BC cell migration, invasion, and proliferation. DLD exhibited positive correlations with CD68+ macrophages and PD-L1 in the tumor, as well as with macrophages and CD4+ T cells in the stroma. Tumor regions with high DLD expression were enriched in PD-L1 and macrophages, while stromal regions with high DLD expression contained CD4+ T cells and macrophages. The AUC values for 1-, 3-, and 5-year overall survival in TCGA-BRCA training set were 0.67, 0.66, and 0.66, respectively. A nomogram with a C-index of 0.715 indicated that risk score, tumor stage, and age could serve as independent prognostic factors for BC. ConclusionOur findings underscore the significant predictive significance of DLD in BC and its influence on the tumor microenvironment. DLD represents a promising diagnostic and prognostic marker for BC, offering novel avenues for the identification of therapeutic targets and the enhancement of immunotherapy in BC.