Clinical Significance In Breast Cancer Research Articles

Upregulation of vesicle-associated membrane protein 7 in breast cancer tissues.

Vesicle-associated membrane protein 7 (VAMP7) plays oncogenic roles in cancers. However, its clinical significance in breast cancer (BC) tissues remains unknown. To elucidate the clinical implications of VAMP7, as well as its involvement in the tumor microenvironment and molecular pathways of breast cancer. BC (n=100) and non-cancerous breast tissues (n= 100) were collected for an immunohistochemical experiment (1:200). The protein expression level of VAMP7 was determined by using a semi-quantitative scoring method. High-throughput RNA-sequencing data of BC tissues were analyzed to confirm the mRNA expression trend of VAMP7. Additionally, the largest BC prognosis cohort data were collected to mine the potential impact VAMP7 has on BC progression. The association between VAMP7 and the microenvironment of BC was evaluated by using a CIBERSORT algorithm. Moreover, we explored the co-expressed molecular mechanisms of VAMP7 in BC by calculating Pearson correlation coefficients and overexpressed genes. Finally, the biological mechanism underlying the relationship between VAMP7 and the key pathways was also explored using gene set enrichment analysis (GSEA). Potential therapeutic strategies were predicted targeting VAMP7. VAMP7 protein was significantly over-expressed in BC tissue than that in controls (p< 0.001). Compared with 459 normal breast tissues and 113 non-cancerous breast tissues, the expression level of VAMP7 mRNA was significantly increased in 1111 BC tissues. CD4+T cells, macrophages, and naïve B cells had a higher infiltration rate in BC tissues with high VAMP7 expression, while regulatory T cells and CD8+T cells had a lower infiltration rate. Over-expressed VAMP7 was associated with macrophages activation and transition from M1 to M2 polarization. Upregulated VAMP7 could predicted poorer OS, DMFS, PPS, and RFS outcomes. Upregulated VAMP7 co-expressed genes were significantly enriched in the cell cycle checkpoints. GSEA confirmed that over-expressed VAMP7 are markedly associated with functional enrichment in cell cycle related categories, including mitotic spindle, G2M checkpoint, and E2F targets. KU-55933 was predicted as a putative therapeutic drug for BC targeting VAMP7. VAMP7 was upregulated in BC tissue and correlated with poor prognosis of BC patients. VAMP7 may promote BC progression by targeting the cell cycle pathway.

Abstract C018: Prognostic worth of Nrf2/BACH1/HO-1 signals in the development of breast cancer

Abstract Nrf2/BACH1/HO-1 signals have been implicated in the development and progression of tumors. However, there has been no comprehensive analysis of their expressions in breast cancer. This study evaluated Nrf2/BACH1/HO-1 signals expression and its clinical significance in breast cancer. 114 female breast cancer of different pathologic parameters and non-cancerous tissues were evaluated for Nrf2/BACH1/HO-1 signals expression using immunohistochemistry and western blot. The relationship connecting Nrf2/BACH1/HO-1 expression and clinicopathologic factors were assessed using the chi-square test. Nrf2 protein in cancerous tissues (74%) was significantly higher than in the non-cancerous tissues (43% p &amp;lt; 0.002), BACH1 expression in cancerous tissues (61%) was significantly lower than that of non-cancerous tissues (20% p &amp;lt; 0.031) and HO-1 protein expression in cancerous tissues (67%) was highly significant than in the non-cancerous tissues (17% p &amp;lt; 0.001). The expressions of Nrf2 and HO-1 significantly correlated with tumor grade whilst BACH1 was significantly associated with tumor stage (p &amp;lt; 0.05). Nrf2, BACH1 and HO-1 play key roles in the growth and development of breast cancer and can serve as probable biological markers for breast cancer diagnosis, prognosis as well as therapeutic targets. Citation Format: Precious Barnes, Roland Osei Saahene. Prognostic worth of Nrf2/BACH1/HO-1 signals in the development of breast cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C018.

PATHOLOGICAL SIGNIFICANCE OF CDH1/E-CADHERIN GERMLINE SEQUENCE VARIANTS IN BREAST CANCER PATIENTS.

Germline alterations of the CDH1 (E-cadherin) tumor suppressor gene have been reported in several epithelial malignancies like hereditary diffuse gastric cancer and lobular breast cancer. E-cadherin plays a central role in proliferation, maintenance of cell-to-cell adhesion, polarity, and epithelial-mesenchymal transition of tissue cells. It is necessary to analyze the impact of the CDH1 germline sequence variants on protein and predict its clinical significance in breast cancer (BC) progression. The aim of the current study was to evaluate the impact and association of CDH1 gene potentially pathogenic variants/likely pathogenic variants (PVs/LPVs) with the initiation and progression of BC. In this study, the clinical data of 200 BC patients have been analyzed based on the type of BC, age, grade, stage, hormonal status, and risk factors. Blood samples from 50 healthy donors were used as a control. Furthermore, CDH1 gene molecular analysis, along with in silico analysis, was provided to assess the invasiveness and progression of BC caused by the E-cadherin protein. Four variants were identified by genetic screening within the CDH1 gene that included variations in exons 7, 8, 10, 11, and 13. Exon 10 had splice site mutation at position c.1337C>A, affecting the protein structure. In exon 11, there was an insertion of T base at position 1669, resulting in truncated protein compared to a normal one that can lead to the disease-causing non- sense-mediated decay and exon 13 variant c.2076T>C has already known polymorphism. In silico analysis of CDH1 showed the presence of the different variants that indicated the overall disruption of protein structure and function. The further functional analysis of these variants and their association with BC can be ensured by increasing the sample size and in vivo studies using mouse models.

Comprehensive Analysis of SLC35A2 in Pan-Cancer and Validation of Its Role in Breast Cancer.

Elucidation of the oncogenic role of SLC35A2 in human tumors and the potential function and clinical significance in breast cancer. Pan-cancer analysis was performed via various bioinformatics tools to explain the pathogenic role of SLC35A2. A prognostic nomogram was also developed based on the SLC35A2 expression and clinicopathological characteristics in breast cancer patients. In addition, the role of SLC35A2 was validated in breast cancer by in vivo and in vitro experiments. SLC35A2 expression is increased in 27 tumor types, and its high expression is substantially correlated with poor prognosis in patients with a variety of cancers. Receiver operating characteristic (ROC) curves showed that SLC35A2 expression levels could accurately distinguish most tumor tissues from normal tissues. High SLC35A2 expression was linked to increased immune infiltration in myeloid-derived suppressor cells (MDSC), as well as immune checkpoints, ferroptosis-related genes, tumor mutational burden (TMB), and microsatellite instability (MSI). SLC35A2 may be involved in tumorigenesis by regulating the glycosylation process. Furthermore, multivariate Cox analysis showed that SLC35A2 was an independent prognostic factor for breast cancer. And the nomogram model had good predictive accuracy for the prognosis of breast cancer patients. Meanwhile, cellular experiments demonstrated that knockdown of SLC35A2 could significantly inhibit the proliferation, migration and invasion of breast cancer cells, while increasing the protein level of E-cadherin and decreasing N-cadherin. A nude mouse xenograft model showed that inhibition of SLA35A2 expression could significantly inhibit tumor growth. SLC35A2 has good diagnostic and prognostic values in multiple cancers and is closely related to tumor immune infiltration. In addition, SLA35A2 as an oncogene in breast cancer may be involved in the progression of epithelial mesenchymal transition (EMT).

Evaluating the role of MEN1 gene expression and its clinical significance in breast cancer patients.

Breast cancer is a multifactorial disease which involves number of molecular factors that are critically involved in proliferation of breast cancer cells. MEN1 gene that is traditionally known for its germline mutations in neuroendocrine tumors is associated with high risk of developing breast cancer in females with MEN1 syndrome. However, the paradoxical role of MEN1 is reported in sporadic breast cancer cases. The previous studies indicate the functional significance of MEN1 in regulating breast cells proliferation but its relevance in development and progression of breast cancer is still not known. Our study targets to find the role of MEN1 gene aberration and its clinical significance in breast cancer. Breast tumor and adjacent normal tissue of 142 sporadic breast cancer patients were collected at the time of surgery. The expression analysis of MEN1 mRNA and protein was done through RT-PCR, immunohistochemistry and western blotting. Further to find the genetic and epigenetic alterations, automated sequencing and MS-PCR was performed respectively. Correlation between our findings and clinical parameters was determined using appropriate statistical tests. MEN1 expression was found to be significantly increased in the breast tumor tissue with its predominant nuclear localization. The elevated expression of MEN1 mRNA (63.38% cases) and protein (60.56% cases) exhibited a significant association with ER status of the patients. Most of the cases had unmethylated (53.52%) MEN1 promoter region, which can be a key factor responsible for dysregulated expression of MEN1 in breast cancer cases. Our findings also revealed the significant association of MEN1 mRNA overexpression with Age and lymph node status of the patients. Our results indicate upregulated expression of MEN1 in sporadic breast cancer patients and it could be critically associated with development and advancement of the disease.

Prognostic and Immunological Roles of CES2 in Breast Cancer and Potential Application of CES2-Targeted Fluorescent Probe DDAB in Breast Surgery.

The expression and function of CES2 in breast cancer (BRCA) has not been fully elucidated. The purpose of this study was to investigate its clinical significance in BRCA. Bioinformatics analysis tools and databases, including The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO) databases, SURVIVAL packages, STRING database, Gene Ontology (GO) enrichment, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, Gene set variation analysis (GSVA), and Tumor Immunity Estimation Resource (TIMER), were utilized to measure the expression level and clarify the clinical significance of CES2 in BRCA. In addition, we verified the expression level of CES2 in BRCA at the cellular and tissue levels by Western blot, immunohistochemistry (IHC) and real-time fluorescence quantitative PCR assays. Furthermore, DDAB is the first reported near-infrared fluorescent probe that can be used to monitor CES2 in vivo. We applied the CES2-targeted fluorescent probe DDAB in BRCA for the first time and verified its physicochemical properties and labeling sorting ability by CCK-8, cytofluorimetric imaging, flow cytometry fluorescence detection, and isolated human tumor tissue imaging assays. The expression of CES2 was higher in normal tissues than that in BRCA tissues. Patients with lower CES2 expression in the BRCA T4 stage had a poorer prognosis. Finally, we applied the CES2-targeted fluorescent probe DDAB in BRCA for the first time, which was demonstrated to have good cellular imaging performance with low biological toxicity in BRCA cells and ex vivo human breast tumor tissue models. CES2 can be considered a potential biomarker to predict the prognosis of breast cancer at stage T4 and might contribute to the development of immunological treatment strategies. Meanwhile, CES2 is able to distinguish between breast normal and tumor tissues, the CES2-targeting NIR fluorescent probe DDAB may have potential for surgical applications in BRCA.

Molecular and Immunohistochemical Alterations in Breast Cancer Patients in Upper Egypt.

Breast cancer (BC) plays a major public health in Egyptian woman. In Upper Egypt, there is an increase in incidence of BC compared to other Egyptian areas. Triple-negative BC, estrogen receptor (ER)-negative, progesterone receptor (PR)-negative, and HER2-neu-negative, is a high-risk BC that lacks the benefit of specific therapy that targets these proteins. Accurate determination of Caveolin-1(Cav-1), Caveolin-2 (Cav-2) and HER-2/neu status have become of major clinical significance in BC by focusing about its role as a tumor marker for response to different therapies. The present study was performed on 73 female BC patients in the South Egypt Cancer Institute. Blood samples were used for Cav-1, Cav-2, and HER-2/neu genes amplification and expression. In addition, immunohistological analysis of mammaglobin, GATA3, ER, PR, and HER-2/neu was done. There was a statistically significant association between Cav-1, 2 and HER-2/neu genes expression and the age of patients (P< 0.001). There are increase in the level of Cav-1, 2 and increase in HER-2/neu mRNA expression in groups treated with chemotherapy and group treated with both chemotherapy and radiotherapy compared to each group baseline level of genes mRNA expression before treatment. On the contrary, the group treated with chemotherapy, radiotherapy and hormonal therapy revealed increase on the level of Cav-1, 2 and HER-2/neu mRNA expression when compared with their baseline for the same patients before treatment. Noninvasive molecular biomarkers such as Cav-1 and Cav-2 have been proposed for use in the diagnosis and prognosis for women with BC.

Investigation of miR-133a, miR-637 and miR-944 genes expression and their relationship with PI3K/AKT signaling in women with breast cancer.

MicroRNAs (miRNAs) are regulatory molecules capable of positively or negatively regulating signaling pathways, and are involved in tumorigenesis as well as various aspects of cancer. The purpose of this study was to investigate the expression levels of miR-133a, miR-637, and miR-944 in serum and tumor tissues as well as their relationship with the expression level of phosphatidylinositol-3-kinase (PI3K) and protein kinase-B (AKT) genes and proteins along with their clinical significance in breast cancer. The expressions of miR-133a, miR-637, miR-944, PI3K, and AKT genes were examined in the tumor and tumor margin tissues of 40 patients with breast cancer, as well as the serum levels of miR-133a, miR-637, and miR-944 in these patients and 40 healthy groups by quantitative real-time PCR (qRT-PCR). PI3K and AKT proteins expression in tumor and tumor margin tissues were detected using immunohistochemistry (IHC). The expression levels of miR-133a and miR-637 in the tumor tissue and serum of patients were lower than those in the tumor margin tissue and serum of the healthy group, respectively. In addition, the expression level of miR-944 in the tumor tissue was lower than that in the tumor margin tissue, but its expression increased in the serum of cancer patients compared to that in the healthy group. The expression of miR-637 was correlated with tumor location and Her2 receptors, and the expression of miR-944 was correlated with tumor location and family history. PI3K and AKT mRNA and protein levels were higher in the tumor tissues than in the tumor margin tissues (p < 0.05). The results of our study revealed that miR-637 has a better diagnostic value in breast cancer than miR-133a and miR-944.

Striatins and STRIPAK complex partners in clinical outcomes of patients with breast cancer and responses to drug treatment.

Striatins (STRNs) family, which contains three multi-domain scaffolding proteins, are cornerstones of the striatins interacting phosphatase and kinase (STRIPAK) complex. Although the role of the STRIPAK complex in cancer has become recognized in recent years, its clinical significance in breast cancer has not been fully established. Using a freshly frozen breast cancer tissue cohort containing both cancerous and adjacent normal mammary tissues, we quantitatively evaluated the transcript-level expression of all members within the STRIPAK complex along with some key interacting and regulatory proteins of STRNs. The expression profile of each molecule and the integrated pattern of the complex members were assessed against the clinical-pathological factors of the patients. The Cancer Genome Atlas (TCGA) dataset was used to evaluate the breast cancer patients' response to chemotherapies. Four human breast cancer cell lines, MDA-MB-231, MDA-MB-361, MCF-7, and SK-BR-3, were subsequently adopted for in vitro work. Here we found that high-level expressions of STRIP2, calmodulin, CCM3, MINK1 and SLMAP were respectively associated with shorter overall survival (OS) of patients. Although the similar pattern observed for STRN3, STRN4 and a contrary pattern observed for PPP2CA, PPP2CB and PPPR1A were not significant, the integrated expression profile of STRNs group and PPP2 group members constitutes a highly significant prognostic indicator for OS [P<0.001, hazard ratio (HR)=2.04, 95% confidence interval (95% CI), 1.36-3.07] and disease-free survival (DFS) (P=0.003, HR=1.40, 95% CI, 1.12-1.75). Reduced expression of STRN3 has an influence on the biological functions including adhesiveness and migration. In line with our clinical findings, the breast cancer cells responded to STRN3 knockdown with changes in their chemo-sensitivity, of which the response is also breast cancer subtype dependent. Our results suggest a possible role of the STRIPAK complex in breast cancer development and prognosis. Among the members, the expression profile of STRN3 presents a valuable factor for assessing patients' responses to drug treatment.

LINC00536 Promotes Breast Cancer Progression by Regulating ROCK1 via Sponging of miR-214-5p.

Accumulating evidence has shown that long noncoding RNAs (lncRNAs) play a significant role in regulating gene expression and participating in the progression of various malignancies. In our study, by analyzing data from The Cancer Genome Atlas (TCGA), LINC00536 was found to be highly expressed in breast cancer (BC) tissues, but its function and clinical significance in BC are still unknown. Therefore, we aimed to explore the role and molecular mechanism of LINC00536 in BC. We collected human BC tissue specimens and validated that LINC00536 was overexpressed in BC tissues. Increased LINC00536 expression was associated with advanced TNM stage, larger tumor diameter, lymph node metastasis and poor prognosis in patients with BC. Univariate and multivariate Cox regression analyses showed that high LINC00536 expression was an independent prognostic risk factor for overall survival in BC patients. Furthermore, quantitative reverse transcription PCR (qRT-PCR) showed that LINC00536 was upregulated in BC cell lines. Then, we confirmed that LINC00536 silencing-inhibited BC cell proliferation, migration, and invasion and led to cell cycle arrest in vitro. Animal experiments showed that knockdown of LINC00536 expression suppressed tumorigenesis in vivo. Mechanistically, LINC00536 serves as a ceRNA for miR-214-5p, increasing the expression of ROCK1, which acts as a tumor promoter in BC. Rescue assays revealed that miR-214-5p inhibition or ROCK1 overexpression could neutralize the suppressive effects of LINC00536 knockdown on cell proliferation, migration and invasion. Our data indicated that LINC00536 accelerates BC progression by regulating the miR-214-5p/ROCK1 pathway, which might provide a new perspective to investigate the development process of BC.

Identification of CISD1 as a Prognostic Biomarker for Breast Cancer.

Although CISD1 (CDGSH iron sulfur domain 1) is upregulated in many cancer types, the potential role of CISD1 in breast cancer is still unclear. The purpose of this study was to investigate its clinical significance in breast cancer. We obtained 1109 breast cancer samples and 113 normal samples from The Cancer Genome Atlas (TCGA) and GTEx databases to demonstrate the relationship between CISD1 expression and pancancer characteristics. We analysed the relationship between CISD1 and breast cancer using the t-test and the chi-square test to evaluate the expression level of CISD1 and its clinical significance in breast cancer. The prognostic value of CISD1 in breast cancer was determined by Kaplan‒Meier and Cox regression analyses. The biological pathways were screened by gene set analysis and Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and single-sample gene set enrichment analysis (ssGSEA), of which the correlation between the level of immune infiltration and the expression of CISD1 in breast cancer was then analysed. Finally, we verified the conclusion by qPCR, immunohistochemistry, and CCK8. CISD1 is highly expressed in breast cancer patients. In addition, we verified a higher expression of CISD1 expressed in the BRCA (breast cancer) cell line, whereas CISD1 has a high diagnostic value, with an AUC of 0.718. Kaplan‒Meier survival and Cox regression analyses showed that high expression of CISD1 was independently associated with adverse clinical outcomes. In turn, GO and KEGG analyses showed that most genes were related to rRNA metabolic process, rRNA processing. Moreover, PCR and immunohistochemistry showed that CISD1 in breast cancer tissues was upregulated significantly, with CCK8 results showing that the proliferation of breast cancer cells decreased after CISD1 knockout. A high level of CISD1 is associated with poor prognosis and immune infiltration in breast cancer.

Clinical relevance of CERK and SPHK1 in breast cancer and their association with metastasis and drug resistance

Despite numerous reports on the altered sphingolipids metabolism in human cancers, their clinical significance in breast cancer remains obscure. Previously, we identified the high levels of sphingolipids, ceramide phosphates and sphingosine phosphates, and the genes involved in their synthesis, CERK and SPHK1, in breast cancer patients. The present study aimed to determine the correlations of CERK and SPHK1 with clinical outcomes as well as metastasis and drug resistance markers. Both local and TCGA cohorts were analysed. High-confidence regulatory interaction network was constructed to find association of target genes with metastasis and drug resistance. Furthermore, correlations of CERK and SPHK1 with selected metastasis and drug resistance markers were validated in both cohorts. Overexpression of CERK and SPHK1 was associated with nodal metastasis, late tumor stage and high proliferation potency. In addition, increased CERK expression was also indicative of poor patient survival. Computational network analysis revealed the association of CERK and SPHK1 with known metastasis markers MMP-2 and MMP-9 and drug resistance markers ABCC1 and ABCG2. Correlation analysis confirmed the associations of target genes with these markers in both local as well as TCGA cohort. The above findings suggest clinical utility of CERK and SPHK1 as potential biomarkers in breast cancer patients and thus could provide novel leads in the development of therapeutics.

The expression and clinical significance of STAMBP in breast cancer.

Breast cancer is the leading cause of death from cancer in women worldwide. STAMBP functions as a JAMM family deubiquitinating enzyme that modulates the stability of substrate proteins in cells by cleaving ubiquitin moieties. The expression of STAMBP and its clinical significance in breast cancer remain unclear. The level of the STAMBP protein in noncancerous and tumor tissues of breast cancer patients was examined by immunohistochemical staining. The expression of STAMBP mRNA in tissues based on healthy individual and breast cancer patient data in the TCGA database was evaluated. The association between the expression of STAMBP mRNA and clinical features and prognosis was evaluated using TCGA database. Cell growth was assessed by Cell Counting Kit-8 (CCK-8) assay, and cell migration and invasion were assessed by wound healing and Transwell assays. Activation of the ERK signaling was detected by Western blotting. The expression of STAMBP was markedly upregulated in the cytoplasm of tumor cells from breast cancer patients. The level of STAMBP was closely associated with the tumor subtype and size and the TNM stage of the breast cancer patients. Importantly, high expression of STAMBP predicted poor overall survival (OS) for breast cancer patients. Furthermore, knockdown of STAMBP expression reduced cell mobility and invasion of breast cancer cells. Notably, the phosphorylation of EGFR and ERK was markedly reduced in STAMBP-knockdown cells. STAMBP plays a critical role in the progression of breast cancer and may serve as a biomarker to monitor the progression of the disease.

Expression characteristics of the yes-associated protein in breast cancer: A meta-analysis.

Background:The yes-associated protein (YAP) gene plays an important role in many malignant tumors, but its clinical significance in breast cancer remains unclear. This study aimed to explore the significance of YAP expression in breast cancer using meta-analysis.Methods:Seven databases will be searched to collect the case–control studies published on the association between YAP expression and clinical pathogenic features in breast cancer until December 2021: PubMed, EMBASE, Web of Science, China National Knowledge Infrastructure, Chinese Scientific Journal Database, Wan Fang Database, and the Chinese Biomedical Literature Database. To perform meta-analysis, STATA 14.0 and RevMan5 software were used with odds ratio (OR) and 95% confidence interval (95% CI) as the effect index, and publication bias and sensitivity analysis were subsequently tested.Results:Form a total of 10 articles used in this study, 8 studies consisted of nontriple negative breast cancer (non-TNBC) and the other 2 of TNBC. Meta-analysis indicated a positive expression rate of YAP in non-TNBC tissues that was lower than in normal breast tissue (OR = 0.15, 95% CI = 0.10–0.21, P < .001). In contrast, the positive rate of YAP expression in TNBC was significantly higher than that in normal breast tissue (OR = 18.23, 95% CI = 8.20–40.52, P < .001). Furthermore, the positive expression rate was higher in the patients with lymph node metastasis, higher tumor node metastasis stage and histologic grade, and larger diameter in TNBC. However, there was no statistical difference in the positive expression rate of YAP between non-TNBC patients and lymph node metastasis, tumor node metastasis stage, histologic grade, and tumor size.Conclusions:YAP may participate in the occurrence and development of non-TNBC as a tumor suppressor gene; however, it may also be a carcinogenic factor in TNBC and may be a potential therapeutic target for TNBC.

CK+/CD45+ (dual-positive) circulating cells are associated with prognosis in patients with advanced breast cancer.

1093 Background: Circulating tumor cells (CTCs) expressing epithelial markers (EPCAM, cytokeratin (CK)) and lacking CD45 (a leukocyte marker) have been associated with poor outcome in many cancer types. Nonetheless, the presence of cells expressing both CK and CD45 (CK+/CD45+), circulating in the blood of cancer patients (pts) have also been reported, but not widely investigated. Early evidence indicates that circulating dual-positive cells (DPcells) are hybrids deriving from the fusion of tumor cells and macrophages. We previously reported that it is possible to detect DPcells in the blood of pts with metastatic breast cancer (BC) and that they are associated with shorter progression-free survival (PFS), in pts with &lt;5 CK+/CD45- CTCs. Here, we investigated the impact of DPcells on overall survival (OS) in pts with advanced BC (aBC). Methods: Blood samples (7.5 ml) were collected from aBC pts before starting a new therapy and processed with the FDA-approved CellSearch platform for CTCs and DPcells enumeration. The prognostic role of CTCs and DPcells was assessed through the Kaplan-Meier method using the log-rank test. Single DPcells were isolated using the DEPArray platform and underwent whole genome amplification and lowpass whole genome sequencing (Ampli1 WGA and Ampli1 Lowpass kits). Results: Blood samples from 341 pts with luminal (n=168), HER2+ (n=76) and triple negative (n=88) BC were analyzed. Of these, 131 samples (38.4%) contained ≥5 CTCs (CTCpos), whereas DPcell were detected in 152 samples (44.6%, range 0-53), of which 66 (43.4%) were CTCpos and 86 (56.6%) CTCneg. Overall, DPcells were associated with a shorter OS: median OS 24.5 vs 35.0 months, p=0.046. However, when analyzing CTCpos and CTCneg separately, only the latter group showed a difference in OS according to DPcells presence. In particular, among CTCneg pts, those with ≥4 DPcells showed a 2.3-fold shorter OS (26.7 vs 60.6 months, p=0.025). Moreover, pts with ≥4 DPcells were less likely to experience a 6-months PFS clinical benefit (p=0.015). Interestingly, in the analysis by BC subtype, DPcells were confirmed to be associated with worse OS only in pts with triple negative BC (median OS 11.5 vs 16.9, p=0.048). To explore the exiology of DPcells, 2 out of 3 cells analyzed after single-cell isolation from 1 patient were confirmed to have copy number alterations (CNA) consistent with malignant cells. CNA and mutational profiling of additional single DPcells and CTCs are ongoing. Conclusions: DPcells are associated with worse OS in aBC pts, with the prognostic impact primarily in pts with &lt;5 CTCs and triple negative BC. This suggests that DPcells might be an alternative way of tumor dissemination in specific pts, in which CK+/CD45- CTCs are less prevalent. More studies are needed to better elucidate DPcell clinical significance in BC, and to confirm their fusion-hybrid origin.

Expression Patterns of Ezrin and AJAP1 and Clinical Significance in Breast Cancer.

Ezrin and adherens junction-associated protein 1 (AJAP1) are structural proteins which are involved in numerous human malignancies. However, little is known about the relationship between them in breast cancer. This study was set out to investigate the relationship between them and to further explore the mechanism of AJAP1-mediating cytoskeleton in breast cancer progression. Ezrin and AJAP1 expressions were detected in 377 samples of breast cancer by immunohistochemistry, and different expression patterns between AJAP1 and Ezrin with clinicopathological parameters were analyzed. Besides, univariate and multivariate Cox models were used to evaluate their prognostic potential. Enzyme-linked immunosorbent assay, Western blot, qRT-PCR, and phalloidin staining of F-actin were used to explore the relationship and the mechanism between AJAP1 and Ezrin in cytoskeleton arrangement. 377 cases of breast cancer results showed that AJAP1 expression was negatively related with histological grade and lymph node involvement and could be an independent prognosis marker of breast cancer. AJAP1 expression tended to be higher in the Ezrin-negative expression case. Patients with AJAP1negative and Ezrinpositive expression had a worse prognosis (p < 0.0001) and shorter DFS (p = 0.015). More importantly, AJAP1 depletion increased the cell ability of F-actin formation through promoting Ezrin expression. AJAP1 depletion might mediate breast cancer malignancy potential through promoting Ezrin expression and cytoskeleton formation.

Key Role of MCUR1 in Malignant Progression of Breast Cancer.

BackgroundMitochondrial calcium uniporter regulator 1 (MCUR1, also known as CCDC90A) is a protein-coding gene that plays a key role in mitochondrial calcium uptake. However, knowledge about its clinical significance in breast cancer is still limited.MethodsThe expression profile of MCUR1 in various cancers was analyzed via the ONCOMINE and Tumor Immune Estimation Resource databases. The correlation between MCUR1 expression and the clinical features of breast cancer was investigated using UALCAN and MEXPRESS. Immunohistochemical analysis was applied to verify the expression of MCUR1 in breast cancer. The prognostic significance of MCUR1 in breast cancer was evaluated using Kaplan–Meier plotter and the PrognoScan database. Gene Set Enrichment Analysis (GSEA) was performed to explore the possible biological functions of MCUR1. In addition, the function of MCUR1 was examined by gene silencing in vitro. Western blotting was applied to detect the expression of proteins.ResultsMCUR1 was overexpressed in breast cancer and significantly related to the clinical characteristics of breast cancer. Results from the public databases and IHC analysis indicated that MCUR1 expression was the highest in triple-negative breast cancer (TNBC). The high expression of MCUR1 was associated with poor overall survival, distant metastasis-free survival, and recurrence-free survival. GSEA showed that the hypoxia pathway, cellular reactive oxygen species (ROS) pathway, epithelial–mesenchymal transition pathway, and notch signaling pathway were differentially enriched in the high MCUR1 expression phenotype. In vitro experiments showed that MCUR1 knockdown in TNBC cell lines led to a decrease in cellular ROS and weakened cell migration and invasion abilities. Moreover, Western blotting showed that MCUR1 knockdown inhibited the epithelial–mesenchymal transition of TNBC cells via the ROS/Nrf2/Notch pathways.ConclusionOur study suggests that MCUR1 plays a pivotal role in the malignant progression of breast cancer.

CTLA-4 Expression and Its Clinical Significance in Breast Cancer.

Breast cancer is the leading cause of women's death among all cancers. The main reason associated with this is the development of metastasis and therapy-resistant breast carcinoma (BC), which pose the main challenge of oncology nowadays. Evidence suggest that these tumors seem to have inhibitory mechanisms that may favor their progression and surveillance. Cancer cells can evade antitumor T cell responses by expressing some immune inhibitory molecules such as the cytotoxic T-lymphocyte antigen-4 (CTLA-4), whose clinical meaning has emerged in the last few years and is poorly understood in the BC context. This systematic literature review aims at identifying studies on CTLA-4 expression in BC, and address what is known about its clinical meaning. A literature search was performed in PubMed and LILACS databases, using the MESH terms "breast cancer"; "CTLA-4 Antigen/antagonists and inhibitors"; and "Lymphocytes, Tumor-Infiltrating/immunology", published in the last 10years. In total, 12 studies were included in this review. Systematic review used the Preferred Reporting Items for Systematic Reviews and Meta-Analyses. Despite the small number of eligible studies, the literature reports some associations between CTLA-4 expression in the tumor microenvironment and worse BC outcomes, regardless of its molecular subtype. CTLA-4 expression in BC is a putative marker of clinical significance and a rationale therapeutic target in the emerging field of immunotherapy.

The impact of CBP expression in estrogen receptor-positive breast cancer

BackgroundThe development of new biomarkers with diagnostic, prognostic and therapeutic prominence will greatly enhance the management of breast cancer (BC). Several reports suggest the involvement of the histone acetyltransferases CREB-binding protein (CBP) and general control non-depressible 5 (GCN5) in tumor formation; however, their clinical significance in BC remains poorly understood. This study aims to investigate the value of CBP and GCN5 as markers and/or targets for BC prognosis and therapy. Expression of CBP, GCN5, estrogen receptor α (ERα), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2) in BC was analyzed in cell lines by western blot and in patients’ tissues by immunohistochemistry. The gene amplification data were also analyzed for CBP and GCN5 using the publicly available data from BC patients.ResultsElevated expression of CBP and GCN5 was detected in BC tissues from patients and cell lines more than normal ones. In particular, CBP was more expressed in luminal A and B subtypes. Using chemical and biological inhibitors for CBP, ERα and HER2 showed a strong association between CBP and the expression of ERα and HER2. Moreover, analysis of the CREBBP (for CBP) and KAT2A (for GCN5) genes in a larger number of patients in publicly available databases showed amplification of both genes in BC patients. Amplification of CREBBP gene was observed in luminal A, luminal B and triple-negative but not in HER2 overexpressing subtypes. Furthermore, patients with high CREBBP or KAT2A gene expression had better 5-year disease-free survival than the low gene expression group (p = 0.0018 and p < 0.00001, respectively).ConclusionsWe conclude that the persistent amplification and overexpression of CBP in ERα- and PR-positive BC highlights the significance of CBP as a new diagnostic marker and therapeutic target in hormone-positive BC.

Correlation of miR-600 with WT1 expression and its potential clinical significance in breast cancer.

Aim: The study aimed to explore miR-600 and WT1 expression and its potential clinical significance in breast cancer. Materials & methods: The expression of miR-600 and WT1 in tumor and non-tumor adjacent tissues in 45 breast cancer patients as well as serum level of miR-600 in these patients and 45 healthy group were analyzed. Results: The expression level of miR-600 in tumor tissue and serum of patients was significantly lower than non-tumor adjacent tissues and serum of controls, respectively, while WT1 mRNA and protein levels were higher in tumor tissues compared with non-tumor adjacent tissues. The miR-600 expression was correlated with lymph node metastasis and clinical stage. Conclusion: The miR-600 acts as tumor suppressor and a diagnostic and prognostic biomarker in breast cancer patients.