Levels In Breast Cancer Tissues Research Articles

PFDN5 plays a dual role in breast cancer and regulates tumor immune microenvironment: Insights from integrated bioinformatics analysis and experimental validation

BackgroundAlthough the prognosis for patients with breast cancer has improved, breast cancer remains the leading cause of death for women worldwide. Prefoldin 5 (PFDN5), as a subunit of the prefoldin complex, plays a vital role in aiding the correct folding of newly synthesized proteins. However, the exact impact of PFDN5 on breast cancer development and its prognostic implications remain unclear. MethodsWe conducted bioinformatics analysis to investigate the correlation between PFDN5 and patient survival, as well as various clinicopathological characteristics in breast cancer. Additionally, various assays were employed to validate the biological functions of PFDN5 in breast cancer. Finally, RNA sequencing (RNA-seq) was utilized to investigate the molecular mechanisms associated with PFDN5. ResultsCompared to normal tissues, PFDN5 exhibited lower expression levels in breast cancer tissues, and lower expression of PFDN5 is associated with poorer prognosis. PFDN5 led to G2/M phase arrest in the cell cycle and reduced proliferative potential in breast cancer cells. However, PFDN5 also promoted migration and invasion of breast cancer cells. Also, RNA-seq analysis revealed an involvement of PFDN5 in the cell cycle and TGF-β signaling pathway. Furthermore, PFDN5 had a significant impact on tumor immune microenvironment by promoting macrophage polarization towards the M1 phenotype and exhibited a positive correlation with CD8+ T cell infiltration levels. ConclusionsPFDN5 plays a dual role in breast cancer and serves as a key factor in tumor immune microenvironment. Therefore, PFDN5 holds promise as a valuable biomarker for predicting both metastatic and prognosis in breast cancer.

Prognostic value of PLEKHA4 and its correlation with tumor-infiltrating immune cells in breast cancer: a comprehensive study based on bioinformatics and clinical analysis validation.

Pleckstrin homology containing family A, number 4 (PLEKHA4) plays a role in a number of biological processes in human cells, including cell polarization, growth, and proliferation. However, the relationship between PLEKHA4 expression and survival in breast cancer (BC) remains unclear. The aim of this study is to investigate the potential of PLEKHA4 as a prognostic indicator in BC. We obtained gene expression profiles of BC and normal tissues from the Tumor Immune Estimation Resource (TIMER), UALCAN web. Immunohistochemistry (IHC) staining was performed to investigate the protein expression and prognostic value of PLEKHA4 in BC patients. The prognostic value was analyzed using Kaplan-Meier curve analysis and Cox regression analysis in R software after downloading The Cancer Genome Atlas (TCGA) databases. The correlations between PLEKHA4 and tumor immune infiltrates were investigated via gene set variation analysis (GSVA). Signaling pathways related to PLEKHA4 expression were identified by the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. Both bioinformatics and IHC results showed that PLEKHA4 was expressed at low levels in BC tissues compared with the adjacent tissues. Furthermore, the expression of PLEKHA4 was negatively correlated with ages (χ2=6.394, P=0.01), molecular subtype (χ2=15.606, P=0.001), lymph node metastasis (χ2=13.753, P=0.004), tumor-node-metastasis (TNM) stage (χ2=22.616, P<0.001). Kaplan-Meier curves implicated low expression of PLEKHA4 was associated with worse survival of BC patients [hazard ratio (HR) =0.46, P=0.01]. Cox regression models showed that low PLEKHA4 expression could be an independent risk factor for BC (HR =0.911, P=0.006). The results of gene set enrichment analysis (GSEA) showed that cell cycle, Notch signaling pathway, nuclear factor erythroid 2-related factor 2 (NRF2) signaling pathway, and Rho GTPases were highly enriched in the low PLEKHA4 expression group, as identified by GO and KEGG. Additionally, in BC, PLEKHA4 expression displayed a positive correlation with the infiltration of natural killer (NK) cells (P<0.001), CD8+ T cells (P<0.001), B cells (P<0.001), neutrophils (P<0.001), and dendritic cells (DCs) (P<0.001). The findings indicate that PLEKHA4 is an independent prognostic biomarker associated with key signaling pathways and immune infiltration in BC. Targeting PLEKHA4 may contribute to improving immunotherapy for BC.

The novel circFKBP8/miR-432-5p/E2F7 cascade functions as a regulatory network in breast cancer

BackgroundCircular RNAs (circRNAs) are capable of affecting breast cancer (BC) development. However, the role and underneath mechanism of circFKBP8 (also known as hsa_circ_0000915) in BC remain largely unknown.MethodsExpression analyses were performed using quantitative real-time polymerase chain reaction (qRT-PCR), western blot, and immunohistochemistry (IHC) assays. Effects on cell functional phenotypes were determined by assessing cell proliferation, migratory capacity, invasion, and stemness in vitro. The relationship between microRNA (miR)-432-5p and circFKBP8 or E2F transcription factor 7 (E2F7) was examined by RNA pull-down, dual-luciferase reporter, and RNA immunoprecipitation (RIP) assays. Xenograft assays were used to identify the function of circFKBP8 in vivo.ResultsCircFKBP8 was presented at high levels in BC tissues and cells. High circFKBP8 expression was associated with worse overall survival in BC patients. CircFKBP8 suppression inhibited BC cell proliferation, migratory capacity, invasion and stemness in vitro. CircFKBP8 suppression blocked xenograft tumor growth in vivo. Mechanistically, circFKBP8 functioned as a miR-432-5p sponge to modulate E2F7 expression. CircFKBP8 modulated BC cell malignant behaviors by miR-432-5p, and miR-432-5p affected these cell phenotypes through E2F7.ConclusionOur observations prove that circFKBP8 promotes BC malignant phenotypes through the miR-432-5p/E2F7 cascade, offering a promising therapeutic and prognostic target for BC.

Zinc transporter ZnT5 is associated with epithelial mesenchymal transition via SMAD1 in breast cancer.

Zinc levels in breast cancer tissues have been reported to be higher than those in normal tissues. In addition, the expression levels of zinc transporters, including ZnT5 and ZnT6, are reportedly higher in breast cancer than in normal breast tissues. ZnT5 and ZnT6 also contribute to heterodimer formation and are involved in several biological functions. However, the functions of ZnT5 and ZnT6 heterodimers in breast cancer remain unknown. Therefore, we first investigated the immunolocalization of ZnT5 and ZnT6 in pathological breast cancer specimens and in MCF-7 and T-47D breast cancer cells. Next, we used small interfering RNA to assess cell viability and migration in ZnT5 knockdown MCF-7 and T-47D cells. Immunohistochemical analysis showed that the number of ZnT5-positive breast cancer cells was inversely correlated with the pathologic N factor status. ZnT5 knockdown had no effect on cell viability in the presence of 100 μM ZnCl2 in MCF-7 and T-47D cells. In a wound healing assay, 100 μM ZnCl2 treatment inhibited cell migration of MCF-7 and T-47D cells, whereas ZnT5 knockdown promoted cell migration, decreased E-cadherin expression and increased vimentin, slug and matrix metalloproteinase 9 expression. Antibody arrays showed that ZnT5 knockdown increased the expression of SMAD1, and that dorsomorphin treatment inhibited the promotion of migratory ability induced by ZnT5 knockdown. The results of this study revealed that both ZnT5 may be involved in less aggressive breast cancer subtypes, possibly through inhibition of cell migration.

SPACA6P-AS: a trailblazer in breast cancer pathobiology and therapeutics

ObjectiveThe primary objective of this investigation is to delve into the involvement of the long noncoding RNA (lncRNA) SPACA6P-AS in breast cancer (BC) development, focusing on its expression pattern, association with clinical-pathological features, impact on prognosis, as well as its molecular and immunological implications.MethodsBioinformatics analysis was conducted utilizing RNA sequencing data of 1083 BC patients from the TCGA database. Functional exploration of SPACA6P-AS was carried out through the construction of survival curves, GO and KEGG enrichment analysis, and single-sample gene set enrichment analysis (ssGSEA). Furthermore, its functionality was validated through in vitro cell experiments and in vivo nude mouse model experiments.ResultsSPACA6P-AS showed a remarkable increase in expression levels in BC tissues (p < 0.001) and demonstrated a close relationship to poor prognosis (overall survival HR = 1.616, progression-free interval HR = 1.40, disease-specific survival HR = 1.54). Enrichment analysis revealed that SPACA6P-AS could impact biological functions such as protease regulation, endopeptidase inhibitor activity, taste receptor activity, taste transduction, and maturity-onset diabetes of the young pathway. ssGSEA analysis indicated a negative correlation between SPACA6P-AS expression and immune cell infiltration like dendritic cells and neutrophils, while a positive correlation was observed with central memory T cells and T helper 2 cells. Results from in vitro and in vivo experiments illustrated that silencing SPACA6P-AS significantly inhibited the proliferation, migration, and invasion capabilities of BC cells. In vitro experiments also highlighted that dendritic cells with silenced SPACA6P-AS exhibited enhanced capabilities in promoting the proliferation of autologous CD3 + T cells and cytokine secretion. These discoveries elucidate the potential multifaceted roles of SPACA6P-AS in BC, including its potential involvement in modulating immune cell infiltration in the tumor microenvironment.ConclusionThe high expression of lncRNA SPACA6P-AS in BC is closely linked to poor prognosis and may facilitate tumor progression by influencing specific biological processes, signaling pathways, and the immune microenvironment. The regulatory role of SPACA6P-AS positions it as a prospective biomarker and target for therapeutic approaches for BC diagnosis and intervention.

AGEs induce MMP-9 promoter demethylation through the GADD45α-mediated BER pathway to promote breast cancer metastasis in patients with diabetes.

Scientific evidence has linked diabetes to a higher incidence and increased aggressiveness of breast cancer; however, mechanistic studies of the numerous regulators involved in this process are insufficiently thorough. Advanced glycation end products (AGEs) play an important role in the chronic complications of diabetes, but the mechanisms of AGEs in breast cancer are largely unexplored. In this study, we first demonstrate that high AGE levels in breast cancer tissues are associated with the diabetic state and poor patient outcomes. Furthermore, AGEs interact with the receptor for AGEs (RAGE) to promote breast cancer cell migration and invasion. Mechanistically, based on RNA sequencing (RNA-seq) analysis, we reveal that growth arrest and DNA damage gene 45α (GADD45α) is a vital protein upregulated by AGEs through a P53-dependent pathway. Next, GADD45α recruits thymine DNA glycosylase for base excision repair to form the demethylation complex at the promoter region of MMP-9 and enhance MMP-9 transactivation through DNA demethylation. Overall, our results indicate a critical regulatory role of AGEs in patients with breast cancer and diabetes and reveal a novel mechanism of epigenetic modification in promoting breast cancer metastasis.

EXPRESSION OF SPP1 AND SPARC GENES IN TUMOR TISSUE OF PATIENTS WITH BREAST CANCER.

Breast cancer (BCa) is one of the most common oncological diseases in women in Ukraine and worldwide, which determines the need to search for new diagnostic and prognostic markers. In this aspect, the study of multicellular proteins, in particular osteopontin (OPN) and osteonectin (ON), in BCа tissue is relevant. The aim of the work was to investigate the expression of SPP1 and SPARC at the mRNA and protein levels in BCa tissue and to assess their relationship with the main clinicopathological BCa characteristics and the survival rates of patients. The work was based on the analysis of the results of the examination and treatment of 60 patients with stage II-III BCa and 15 patients with breast fibroadenomas. SPP1 and SPARC mRNA levels were determined by real-time PCR. The study of the expression of protein products of the SPP1 and SPARC genes was carried out by the immunohistochemical method. We have established that the BCa tissue was characterized by 3.5 (p < 0.05) and 7.4 (p < 0.05) lower levels of SPP1 and SPARC mRNA, respectively, compared to the tissue of benign neoplasms, while OPN and ON expression levels were 1.6 (p < 0.05) and 5.6 (p < 0.05) times higher, respectively, compared to fibroadenoma tissue. The analysis of the relationship between the expression of SPP1 and SPARC at the protein and mRNA levels in BCa tissue and the main clinicopathological BCa characteristics revealed its dependence on the presence of metastases in regional lymph nodes, differentiation grade, and the molecular BCa subtype. Also, high expression levels of SPP1 and OPN were associated with worse patient survival rates. The obtained results indicate the perspective of using SPP1 and SPARC expression indices in BCa tissue to assess the aggressiveness of the cancer course and optimize the tactics of treating patients.

Systematic proteomics analysis revealed different expression of laminin interaction proteins in breast cancer: lower in luminal subtype and higher in claudin-low subtype.

Breast cancer is a major public health concern. Proteomics enables identification of proteins with aberrant properties. Here, we identified proteins with abnormal expression levels in breast cancer tissues and systematically analyzed and validated the data to locate potential diagnostic and therapeutic targets. Protein expression level in breast cancer tissues and para-carcinoma tissues were detected by Isobaric Tags for Relative and Absolute Quantification (iTRAQ) technology and further screened through Gene Expression Profiling Interactive Analysis (GEPIA) database. Cellular components, protein domain and Reactome pathway analysis were performed to screen functional targets. Abnormal expression levels of functional targets were validated by Oncomine database, quantitative real time polymerase chain reaction (qRT-PCR) and proteomics detection. Protein correlation analysis was performed to explain the abnormal expression levels of potential targets in breast cancer. Overall, 207 and 207 proteins were up- and down-regulated, respectively, in breast cancer tissues, and approximately 50% were also detected in the GEPIA database. The overlapping proteins were mainly extracellular proteins containing epidermal growth factor-like domain in leukocyte adhesion molecule (EGF-Lam) domain and enriched in laminin interaction pathway. Moreover, the downregulated laminin interaction proteins could be functional targets, which were also validated through Oncomine-Richardson and Oncomine-Curtis database. However, the lower expression level of laminin interaction proteins only fit for luminal breast cancer cells with no or low metastasis ability because the proteins achieved higher expression level in more invasive claudin-low breast cancer cells. In addition, when compared with corresponding in situ carcinoma tissues, above-mentioned proteins also showed higher expression levels in invasive carcinoma tissues. Finally, we have revealed the negative correlation between the laminin interaction proteins and the claudins. The laminin interaction protein, especially for laminins with β1 and γ1 subunits and their integrin receptors with α1 and α6 subunits, showed lower expression levels in luminal breast cancer with no or lower metastatic ability, but showed higher expression levels in claudin-low breast cancer with higher metastatic ability; and their higher expression could be related to the low claudin expression.

Association of ADP‑ribosylation factor family genes with prognosis and immune infiltration of breast cancer.

Breast cancer (BC) is the most common type of cancer found in women. ADP-ribosylation factors (ARFs) are a group of small proteins that bind to GTP and are involved in controlling different cellular functions. The function and evolution of multiple ARFs in BC have remained to be fully elucidated, despite existing studies on this protein family in Homo sapiens and other species. In the present study, a systematic analysis of ARF expression levels in BC tissues compared to normal breast tissues was performed using data from The Cancer Genome Atlas database. The analysis revealed significantly higher expression of ARFs in BC tissues. In addition, the prognostic significance of ARF1 and ARF3-6 expression levels was assessed in patients with BC. Of note, elevated ARF1 expression was associated with reduced rates of distant metastasis-free survival (DMFS), overall survival (OS) and recurrence-free survival (RFS) in affected individuals. Similarly, patients with high expression levels of ARF3 had lower post-progression survival (PPS) rates. In addition, patients with higher ARF4 expression had worse PPS and patients with high ARF5 expression exhibited lower DMFS. Patients with high ARF6 expression had worse DMFS, OS, RFS and predictive power score values. Furthermore, the expression of ARF was found to be strongly linked to the infiltration of various immune cell types, namely dendritic cells, macrophages, neutrophils, CD8+ T cells and B cells. These significant associations offer a solid foundation for the potential utilization of new therapeutic targets and predictive markers for the treatment of BC.

Clinical value of KiSS-1 and MMP-2 expression levels in breast cancer tissue in evaluating prognosis of elderly breast cancer patients after modified radical mastectomy.

We attempted to clarify clinical value of KiSS-1 and MMP-2 levels in breast cancer (BC) tissue in evaluating prognosis of elderly BC patients after modified radical mastectomy (MCM). The data of 192 elderly female BC patients receiving MCM in our hospital from January 2018 to December 2022 were collected. According to prognosis, patients received division into poor prognosis group (n = 43) and good prognosis group (n = 149). The serum CEA level and KiSS-1 and MMP-2 levels in BC tissue received measurement in both groups. The predictive value of KiSS-1 and MMP-2 alone and jointly in adverse prognosis of elderly BC patients after MCM received assessment. Results showed that No statistical significance was exhibited between both groups in general data (P > 0.05). The serum CEA level and MMP-2 expression in BC tissue in poor prognosis group exhibited elevation relative to those in good prognosis group, and KiSS-1 expression in BC tissue in poor prognosis group exhibited depletion relative to that in good prognosis group, indicating statistical significance (P < 0.05). The high-level KiSS-1 might be a protective element for adverse prognosis of elderly BC patients after MCM, and high-level CEA and MMP-2 might be an independent risk element for adverse prognosis of elderly BC patients after MCM (P < 0.05). KiSS-1 and MMP-2 alone and jointly predicted AUC of adverse prognosis in elderly BC patients after MCM were 0.93, 0.802 and 0.958, with certain predictive values; when cutoff values of KiSS-1 and MMP-2 were 6.15 and 2.26, the predictive value was the best. In conclusion, KiSS-1 and MMP-2 levels in BC tissue possess relation to adverse prognosis of MCM. KiSS-1 and MMP-2 levels in elderly BC patients before surgery may be detected in the future to assist in prognosis evaluation of elderly BC patients after MCM.

TFAP2C Activates CST1 Transcription to Facilitate Breast Cancer Progression and Suppress Ferroptosis.

Cystatin SN (CST1) appears to have pro-tumor effects in breast cancer (BC) and is involved in ferroptosis; however, there is no report on the regulation of ferroptosis by CST1 for BC development. The purpose of this study is to investigate the functions and mechanisms operated by CST1 in BC development and ferroptosis. Transcription Factor Activator Protein 2γ (TFAP2C) and CST1 levels in BC tissues and estrogen receptor (ER)+ cells were quantified by RT-qPCR and western blotting. After knocking down TFAP2C and CST1 expression in MCF7 and T47D cells, the proliferation, colony formation ability, apoptosis, and cell cycle were assessed. Ferroptosis was verified by detecting glutathione peroxidase 4 (GPX4) and 4-hydroxy-2-nonenal (4HNE) levels. The kits were used to test Fe2+, reactive oxygen species, malondialdehyde, and glutathione levels, and ultrastructure of mitochondria was observed through transmission electron microscope. Dual-luciferase reporter assay and chromatin immunoprecipitation test were carried out to investigate the interaction of TFAP2C and CST1. A transplanted tumor model was established to explore the function of TFAP2C in tumorigenesis by quantifying TFAP2C, CST1, Ki67, and GPX4 levels through western blotting and immunochemistry after silencing TFAP2C. TFAP2C and CST1 were predominantly expressed in BC cells. Silencing of TFAP2C or CST1 expression suppressed ER+ BC cell proliferation, promoted apoptosis and ferroptosis, and blocked cell cycle transition from G1 phase to S phase. TFAP2C knockdown in transplanted tumors inhibited tumor growth and GPX4 level. Upregulating CST1 nullified the anti-tumor effects of TFAP2C knockdown and TFAP2C promoted CST1 expression through transcription activation. TFAP2C activates CST1 transcription to facilitate BC development and block ferroptosis.

CENPA knockdown restrains cell progression and tumor growth in breast cancer by reducing PLA2R1 promoter methylation and modulating PLA2R1/HHEX axis.

Breast cancer is a lethal malignancy affecting females worldwide. It has been reported that upregulated centromere protein A (CENPA) expression might indicate unfortunate prognosis and can function as a prognostic biomarker in breast cancer. This study aimed to investigate the accurate roles and downstream mechanisms of CENPA in breast cancer progression. CENPA protein levels in breast cancer tissues and cell lines were analyzed by Western blot and immunohistochemistry assays. We used gain/loss-of-function experiments to determine the potential effects of CENPA and phospholipase A2 receptor (PLA2R1) on breast cancer cell proliferation, migration, and apoptosis. Co-IP assay was employed to validate the possible interaction between CENPA and DNA methyltransferase 1 (DNMT1), as well as PLA2R1 and hematopoietically expressed homeobox (HHEX). PLA2R1 promoter methylation was determined using methylation-specific PCR assay. The biological capabilities of CENPA/PLA2R1/HHEX axis in breast cancer cells was determined by rescue experiments. In addition, CENPA-silenced MCF-7 cells were injected into mice, followed by measurement of tumor growth. CENPA level was prominently elevated in breast cancer tissues and cell lines. Interestingly, CENPA knockdown and PLA2R1 overexpression both restrained breast cancer cell proliferation and migration, and enhanced apoptosis. On the contrary, CENPA overexpression displayed the opposite results. Moreover, CENPA reduced PLA2R1 expression through promoting DNMT1-mediated PLA2R1 promoter methylation. PLA2R1 overexpression could effectively abrogate CENPA overexpression-mediated augment of breast cancer cell progression. Furthermore, PLA2R1 interacted with HHEX and promoted HHEX expression. PLA2R1 knockdown increased the rate of breast cancer cell proliferation and migration but restrained apoptosis, which was abrogated by HHEX overexpression. In addition, CENPA silencing suppressed tumor growth in vivo. CENPA knockdown restrained breast cancer cell proliferation and migration and attenuated tumor growth in vivo through reducing PLA2R1 promoter methylation and increasing PLA2R1 and HHEX expression. We may provide a promising prognostic biomarker and novel therapeutic target for breast cancer.

FEATURES OF COL1A1 EXPRESSION IN BREAST CANCER TISSUE OF YOUNG PATIENTS.

In the last decades, the incidence of breast cancer (BCa) in young women has been increasing steadily. The quantitative indicators of expression of collagen, which play important role in stromal microenvironment, and their association with the age and survival rates of BCa patients have not been yet definitively clarified. To investigate the relationship between the COL1A1 gene expression at the mRNA and protein levels in BCa tissue and the clinicopatological features and survival rates of BCa patients of different age groups. The study was conducted on the clinical material of 50 patients with stage I-III BCa. COL1A1 gene expression at the mRNA and protein levels in BCa tissue were studied using the real-time PCR and immunohistochemical methods, as well as the bioinformatic analysis (UALCAN and Kaplan - Meier Plotter databases). The bioinformatic analysis showed that BCa tissue is characterized by 6.0 times (p < 0.05) higher level of COL1A1 mRNA compared to normal breast tissue. The correlation of COL1A1 expression at the mRNA and protein levels with the molecular subtype of neoplasms was demonstrated. According to Kaplan - Meier Plotter database, a low level of expression of COL1A1 protein level in BCa tissue is associated with lower rates of relapse-free survival of patients. The ex vivo study of the clinical material revealed a decrease in COL1A1 protein expression in tumor tissue of young patients with BCa of T3 category (p < 0.0374), low differentiation grade (p < 0.0163) and basal molecular subtype (p < 0.0001). A correlation between the expression of COL1A1 at the mRNA and protein levels and the expression status of estrogen receptors (p < 0.0001) and progesterone receptors (p < 0.0040) was established. The relapse-free 3-year survival rate of young BCa patients is significantly lower in the presence of a low COL1A1 optical density index in the tumor tissue. The identified relationship between COL1A1 expression and such indicators of BCa malignancy as tumor size, differentiation grade, molecular subtype, receptor status, and the recurrencefree survival of patients indicates the prospects of its use to predict the aggressiveness of the BCa course in young patients.

Estimation of gene expression level of CDH1 as a predisposing factor for metastasis in localized and locally advanced and metastatic breast cancer Iraqi patients

Breast cancer (BC) is a prevalent malignancy among women, ranking as the second most commonly diagnosed cancer globally. Notably, a substantial proportion of breast cancer-related fatalities, up to 90 percent, are attributed to the development of distant organ metastases. While Cadherin1 (CDH1) has conventionally been considered a tumor-suppressor gene in cancer research, recent investigations have unequivocally revealed that both CDH1 and its encoded E-cadherin exhibit oncogenic characteristics. The primary focus of this case-control study is to ascertain the involvement of the CDH1 gene in a specific subset of Iraqi female patients. A total of ninety patients sought diagnosis and treatment at the Oncology Teaching Hospital in Medical City and the Oncology Unit at Al-Yarmouk Teaching Hospital in Baghdad. In addition, we included 30 apparently healthy individuals as blood control subjects and another 30 women with benign breast tumors as tissue control subjects for the study. In the initial phase of the study, we conducted a serological analysis using enzyme-linked immunosorbent assay (ELISA) to detect the concentration of E-cadherin in serum samples from two groups of BC patients. The group with locally advanced and metastatic BC exhibited a significantly higher E-cadherin concentration (963.4 ± 89.8 pg/mL) compared to the group with localized BC. In the second part of the research, qRT-PCR was performed to analyze the expression of the CDH1 gene across all sample types. CDH1 was shown to have the greatest fold expression (2.550 ± 0.164) in cases with locally progressed and metastatic BC. Compared to the seemingly healthy control group, the fold expression for localized BC was 1.456 ± 0.055, and for malignant tissue it was 1.886 ± 0.08621. In conclusion, this study provides compelling evidence supporting CDH1 as an oncogene in BC. The significance of CDH1 in BC tumorigenesis underscores its potential for the development of novel detection biomarkers and targeted therapeutic approaches for BC treatment. Notably, CDH1 exhibited elevated expression levels in BC tissues and demonstrated an association with an unfavorable distant metastasis-free survival outcome.

Prognostic value of lncRNA AFAP1-AS1 in breast cancer: a meta-analysis and validated study in Chinese population.

Long non encoding RNA (lncRNA) plays a crucial role in breast cancer. However, the prognostic role of AFAP1-AS1 in breast cancer remains unclear. To investigate the relationship between the expression of long non-coding RNA actin filament-associated protein1 antisense RNA1 (AFAP1-AS1) and prognosis of breast cancer. Meta-analysis was performed to explore the correlation between AFAP1-AS1 and breast cancer. The AFAP1-AS1expression in patients with breast cancer tissue and adjacent normal tissue from 153 patients was determined by qRT-PCR. Bioinformatics and Cox proportional-hazards risk model were used to explore the relationship between expression of AFAP1-AS1 and prognosis. The combined analysis revealed a significant correlation between AFAP1-AS1 expression and both overall survival (hazard ratios, HR = 2.33, 95%Cl: 1.94-2.81, p < 0.001) as well as disease-free survival/progression-free survival (HR = 2.94, 95%CI: 2.35-3.67, p < 0.001). The relation between expression of AFAP1-AS1 and breast cancer was determined in 153 breast cancer and adjacent normal tissues. The findings revealed a significantly higher AFAP1-AS1expression levels in breast cancer tissues compared to adjacent normal tissues (p < 0.001). Additionally, patients exhibiting heightened levels of AFAP1-AS1 expression were correlated with an unfavorable prognosis (HR = 2.35, 95%CI: 1.47-3.74, p < 0.001), which aligns consistently with the findings of the pooled analysis. The subgroup analysis of clinical characteristics revealed a significant association between high expression of AFAP1-AS1 and TNM stage (HR = 1.72, 95%CI: 1.11-2.65, p = 0.015). This study demonstrated that AFAP1-AS1 acts as an oncogene and may serve as a novel prognostic marker for breast cancer, particularly in the Chinese population.

UBE3B promotes breast cancer progression by antagonizing HIF-2α degradation.

Mutations in E3 ubiquitin ligase UBE3B have been linked to Kaufman Oculocerebrofacial Syndrome (KOS). Accumulating evidence indicates that UBE3B may play an important role in cancer. However, the precise role of UBE3B in cancer and the underlying mechanism remain largely uncharted. Here, we reported that UBE3B is an E3 ligase for hypoxia-inducible factor 2α (HIF-2α). Mechanically, UBE3B physically interacts with HIF-2α and promotes its lysine 63 (K63)-linked polyubiquitination, thereby inhibiting the Von Hippel-Lindau (VHL) E3 ligase complex-mediated HIF-2α degradation. UBE3B depletion inhibits breast cancer cell proliferation, colony formation, migration, and invasion in vitro and suppresses breast tumor growth and lung metastasis in vivo. We further identified K394, K497, and K503 of HIF-2α as key ubiquitination sites for UBE3B. K394/497/503R mutation of HIF-2α dramatically abolishes UBE3B-mediated breast cancer growth and lung metastasis. Intriguingly, the protein levels of UBE3B are upregulated and positively correlated with HIF-2α protein levels in breast cancer tissues. These findings uncover a critical mechanism underlying the role of UBE3B in HIF-2α regulation and breast cancer progression.

Calycosin inhibits triple-negative breast cancer progression through down-regulation of the novel estrogen receptor-α splice variant ER-α30-mediated PI3K/AKT signaling pathway

BackgroundTriple-negative breast cancer (TNBC) is a heterogeneous carcinoma characterized by the most aggressive phenotype among all breast cancer subtypes. However, therapeutic options for TNBC patients have limited clinical efficacy due to lack of specific target and efficient targeted therapeutics. AimTo investigate the biological characteristics of a novel estrogen receptor (ER)-α splice variant ER-α30 in breast cancer cells, and its possible role in the anticancer effects of calycosin, a typical phytoestrogen derived from the herbal plant Astragalus membranaceus, against TNBC. This may also provide a better understanding of the inhibitory activity of calycosin on TNBC progression. MethodsBreast cancer tissues and para-cancer tissues were collected and analyzed for the expression levels of ER-α30 using immunohistochemistry (IHC), and its expression in two TNBC cell lines (MDA-MB-231 and BT-549) was detected by western blot and qRT-PCR assays. Then the alteration of cell viability, apoptosis, migration, invasion and epithelial-mesenchymal transition (EMT) in response to overexpression or knockdown of ER-α30 was separately determined by CCK-8, Hoechst 33258, wound healing, transwell and western blot assays in two TNBC cell lines. Next, the anticancer effects of calycosin on MDA-MB-231 cells were evaluated through CCK-8, colony formation, flow cytometry, Hoechst 33258 and western blot assays, along with the role of ER-α30 in these effects and the possible downstream targets of ER-α30. In addition, the in vivo experiments were carried out using MDA-MB-231 xenograft model intraperitoneally treated with calycosin. The volume and weight of xenograft tumor were measured to evaluate the in vivo anticancer activities of calycosin, while the corresponding changes of ER-α30 expression in tumor tissues were detected by IHC. ResultsIt was demonstrated that the novel ER-α splice variant ER-α30 was primarily distributed in the nucleus of TNBC cells. Compared with normal breast tissues, ER-α30 expression was found in significantly higher levels in breast cancer tissues of ER- and progesterone receptor (PR)-negative subtype, so did in TNBC cell lines (MDA-MB-231 and BT-549) when compared to normal breast cell line MCF10A. Moreover, ER-α30 overexpression strikingly enhanced cell viability, migration, invasion and EMT progression and reduced apoptosis in TNBC cells, whereas shRNA-mediated knockdown of ER-α30 revealed the opposite results. Notably, calycosin suppressed the expression of ER-α30 in a dose-dependent manner, accompanied with the inhibition of TNBC growth and metastasis. A similar finding was observed for the xenografts generated from MDA-MB-231 cells. The treatment with calycosin suppressed the tumor growth and decreased ER-α30 expression in tumor tissues. Furthermore, this inhibition by calycosin was more pronounced in ER-α30 knockdown cells. Meanwhile, we found a positive relationship between ER-α30 and the activity of PI3K and AKT, which could also be inactivated by calycosin treatment. ConclusionFor the first time, it is demonstrated that the novel estrogen receptor-α splice variant ER-α30 could function as pro-tumorigenic factor in the context of TNBC by participating in cell proliferation, apoptosis, invasion and metastasis, thus it may serve as a potential therapeutic target for TNBC therapy. Calycosin could reduce the activation of ER-α30-mediated PI3K/AKT pathway, thereby inhibited TNBC development and progression, suggesting that calycosin may be a potential therapeutic option for TNBC.

Clinical Significance of ABCG2/BCRP Quantified by Fluorescent Nanoparticles in Breast Cancer Patients Undergoing Neoadjuvant Chemotherapy

Breast cancer resistance protein (BCRP), also known as ATP-binding cassette transporter G2 (ABCG2), is associated with chemotherapy resistance. BCRP is also implicated in breast cancer stem cells, and is reported as a poor prognostic factor. However, the relationship of BCRP levels in breast cancer tissues with chemotherapy resistance and prognosis has not been clarified. We aimed to evaluate the correlation between BCRP expression and prognosis in breast cancer using immunohistochemistry with fluorescent phosphor-integrated dots (IHC-PIDs). A total of 37 breast cancer patients with residual cancer in the primary tumor and axillary lymph nodes were evaluated. BCRP levels in breast cancer tissue and metastatic lymph nodes were quantitatively detected after neoadjuvant chemotherapy (NAC). Among these 37 patients, 24 had corresponding core needle biopsies obtained before NAC. Biomarker assay with IHC-PIDs showed high accuracy for the quantitative assessment of BCRP with low expression. High BCRP expression in the primary tumor and metastatic lymph nodes after preoperative chemotherapy was associated with worse overall survival. In conclusion, high BCRP levels may be associated with poor prognosis in patients with breast cancer, having residual tumors within the primary tumor and lymph nodes after preoperative chemotherapy. These findings provide a basis for further appropriate adjuvant therapy in these patients.

Functional variant rs10175368 which affects the expression of CYP1B1 plays a protective role against breast cancer in a Chinese Han population.

Cytochrome P450 1B1 ( CYP1B1 ) genetic variants are relevant in the pathogenesis of breast cancer. Exploring the relationships between CYP1B1 functional variants and breast cancer could improve our understanding of breast cancer molecular pathophysiology. This is a two-stage hospital-based case-control study of a Chinese Han population. Genotyping was performed to identify candidate gene variants. 3DSNP, ANNOVAR, and RegulomeDB were used to determine functional single nucleotide polymorphisms (SNPs). The relationship between candidate variants and breast cancer risk was evaluated through unconditional logistic regression analysis. The PancanQTL platform was used to perform cis and trans expression quantitative trait loci (eQTL) analysis of positive SNPs. The GSCA platform was then used to compare the gene expression levels of potential target genes between breast cancer tissue and normal tissue adjacent to the cancer. rs10175368-T acted as a protective factor against breast cancer based on an additive model [odds ratio (OR) = 0.722, 95% confidence interval (CI) = 0.613-0.850; P < 0.001], and was identified as a protective factor in the postmenopausal population (OR = 0.601; 95% CI, 0.474-0.764; P < 0.001). eQTL analysis and analysis of differential expression in carcinoma and paracancerous tissues revealed that the expression level of CYP1B1 - AS1 was associated with rs10175368 and that CYP1B1-AS1 had significantly higher expression levels in breast cancer tissues than in paracancerous tissues. We show, for the first time in a Chinese Han population, that the functional variant rs10175368 plays a protective role against breast cancer, especially in the postmenopausal population.

Abstract P3-05-15: Clinical significance of breast cancer resistance protein (BCRP) quantified by fluorescence nanoparticle in breast cancer patients undergoing neoadjuvant chemotherapy

Abstract Breast cancer resistance protein (BCRP), also known as ATP-binding cassette transporter G2 (ABCG2), is associated with chemotherapy resistance. BCRP is also implicated in breast cancer stem cells and has been reported as a poor prognostic factor. However, the relationship of BCRP levels in breast cancer tissues with chemotherapy resistance and prognosis has not been clarified to date. Our laboratory recently developed nanoparticles suitable for the quantitative immunohistochemical (IHC) method called phosphor-integrated dots (PIDs). Using PID-based IHC (IHC-PIDs), many biomarker proteins can be visualized and quantitatively analyzed at the single-particle level in paraffine-embedded, formalin-fixed tumor samples. We aimed to evaluate the correlation between quantitatively analyzed BCRP expression and prognosis in breast cancer tissue samples using immunohistochemistry with IHC-PIDs. Thirty-seven Japanese patients diagnosed with invasive ductal carcinoma of the breast who underwent mastectomy and lymph node dissection at Tohoku University Hospital (Sendai, Japan) between 2004 and 2010 were included. All patients had undergone NAC and were pathologically diagnosed with non-pathological complete response or progressive disease and lymph node metastasis postoperatively. The patients were followed up within a median of 10.1 years (range, 1.1–18.2 years). Among the thirty-seven patients, twenty-eight patients had core needle biopsy specimens taken before NAC. BCRP levels in breast cancer tissue and metastatic lymph nodes were quantitatively detected after preoperative chemotherapy and core needle biopsy. Biomarker assay with IHC-PIDs showed high accuracy for quantitative assessment of BCRP with low expression. The optimal PT+LN BCRP cut-off score for predicting survival was 27.4, with a sensitivity of 60.6% and a specificity of 77.3%. BCRP expression in PT+LN samples was dichotomized into high and low groups at this cutoff value, and OS was compared between these group. The results showed that OS was significantly worse in the high BCRP-expression group (log-rank p=0.0089). In conclusion, high BCRP levels are associated with poor prognosis in patients with breast cancer with residual tumors within the primary tumor and lymph nodes after preoperative chemotherapy. These findings provide a basis for further appropriate adjuvant therapy in these patients. Citation Format: Hiroshi Tada, Kohsuke Gonda, Narufumi Kitamura, Minoru Miyashita, Narumi Harada-Shoji, Yohei Hamanaka, Akiko Ebata, Takanori Ishida. Clinical significance of breast cancer resistance protein (BCRP) quantified by fluorescence nanoparticle in breast cancer patients undergoing neoadjuvant chemotherapy [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P3-05-15.