Prognostic Marker In Breast Cancer Research Articles

A cell-to-patient machine learning transfer approach uncovers novel basal-like breast cancer prognostic markers amongst alternative splice variants

BackgroundBreast cancer is amongst the 10 first causes of death in women worldwide. Around 20% of patients are misdiagnosed leading to early metastasis, resistance to treatment and relapse. Many clinical and gene expression profiles have been successfully used to classify breast tumours into 5 major types with different prognosis and sensitivity to specific treatments. Unfortunately, these profiles have failed to subclassify breast tumours into more subtypes to improve diagnostics and survival rate. Alternative splicing is emerging as a new source of highly specific biomarkers to classify tumours in different grades. Taking advantage of extensive public transcriptomics datasets in breast cancer cell lines (CCLE) and breast cancer tumours (TCGA), we have addressed the capacity of alternative splice variants to subclassify highly aggressive breast cancers.ResultsTranscriptomics analysis of alternative splicing events between luminal, basal A and basal B breast cancer cell lines identified a unique splicing signature for a subtype of tumours, the basal B, whose classification is not in use in the clinic yet. Basal B cell lines, in contrast with luminal and basal A, are highly metastatic and express epithelial-to-mesenchymal (EMT) markers, which are hallmarks of cell invasion and resistance to drugs. By developing a semi-supervised machine learning approach, we transferred the molecular knowledge gained from these cell lines into patients to subclassify basal-like triple negative tumours into basal A- and basal B-like categories. Changes in splicing of 25 alternative exons, intimately related to EMT and cell invasion such as ENAH, CD44 and CTNND1, were sufficient to identify the basal-like patients with the worst prognosis. Moreover, patients expressing this basal B-specific splicing signature also expressed newly identified biomarkers of metastasis-initiating cells, like CD36, supporting a more invasive phenotype for this basal B-like breast cancer subtype.ConclusionsUsing a novel machine learning approach, we have identified an EMT-related splicing signature capable of subclassifying the most aggressive type of breast cancer, which are basal-like triple negative tumours. This proof-of-concept demonstrates that the biological knowledge acquired from cell lines can be transferred to patients data for further clinical investigation. More studies, particularly in 3D culture and organoids, will increase the accuracy of this transfer of knowledge, which will open new perspectives into the development of novel therapeutic strategies and the further identification of specific biomarkers for drug resistance and cancer relapse.

Downregulated ZNF132 predicts unfavorable outcomes in breast Cancer via Hypermethylation modification

BackgroundAn important mechanism that promoter methylation-mediated gene silencing for gene inactivation is identified in human tumorigenesis. Methylated genes have been found in breast cancer (BC) and beneficial biomarkers for early diagnosis. Prognostic assessment of breast cancer remain little known. Zinc finger protein 132 (ZNF132) is downregulated by promoter methylation in prostate cancer and esophageal squamous cell carcinoma. However, no study provides information on the status of ZNF132, analyzes diagnosis and prognostic significance of ZNF132 in BC.MethodsIn the present study, the expression of ZNF132 mRNA and protein level was determined based on the Cancer Genome Atlas (TCGA) RNA-Seq database and clinical samples analysis and multiple cancer cell lines verification. P rognostic significance of ZNF132 in BC was assessed using the Kaplan-Meier plotter. Molecular mechanisms exploration of ZNF132 in BC was performed using the multiple bioinformatic tools. Hypermethylated status of ZNF132 in BC cell lines was confirmed via Methylation specific polymerase chain reaction (MSP) analysis.ResultsThe expression of ZNF132 both the mRNA and protein levels was downregulated in BC tissues. These results were obtained based on TCGA database and clinical sample analysis. Survival analysis from the Kaplan-Meier plotter revealed that the lower level of ZNF132 was associated with a shorter Relapse Free Survival (RFS) time. Receiver operating characteristic curve (ROC) of 0.887 confirmed ZNF132 had powerful sensitivity and specificity to distinguish between BC and adjacent normal tissues. Bioinformatic analysis showed that 6% ((58/960)) alterations of ZNF132 were identified from cBioPortal. ZNF132 participated in multiple biological pathways based on the Gene Set Enrichment Analysis (GSEA) database including the regulation of cell cycle and glycolysis. Finally, MSP analysis demonstrated that ZNF132 was hypermethylated in a panel of breast cancer cell lines and 5-aza-2′-deoxycytidine (5-Aza-dC) treatment restored ZNF132 expression in partial cell lines.ConclusionsResults revealed that hypermethylation of ZNF132 contributed to its downregulated expression and could be identified as a new diagnostic and prognostic marker in BC.

Prognostic significance of heat shock protein 90AA1 (HSP90α) in invasive breast cancer

AimsThe mechanisms that drive breast cancer (BC) progression and poor outcome are not fully understood. The human heat shock protein 90 alpha family class A member 1 (HSP90α) encoded by...

Targeting of FK506 binding protein 5 by miR-203 affects the progression of breast cancer via regulating the fatty acid degradation pathway and potential drug-repurposing.

Increasing number of studies have suggested that microRNA (miR)-203 is a potential prognostic marker for breast cancer. However, the specific molecular mechanism underlying the effects of miR-203 remains unknown. The present study aimed to explore the molecular target and underlying mechanisms of action of miR-203 in breast cancer via bioinformatics analysis and cellular assays, such as wound healing assay and western blotting. In the present study, 17 candidate target genes of miR-203 were identified in the downregulated differentially expressed genes from Affymetrix microarray and TargetScan 7.2 database. Subsequently, FK506 binding protein 5 (FKBP5) was considered as the miR-203 target by 3 different hub gene analysis methods (EcCentricity, Betweenness and Stress). FKBP5 protein expression was significantly downregulated in SUM159 cells transfected with miR-203 mimics compared with SUM159 cells transfected with miR-203 negative control (NC) in western blot analysis. High expression of FKBP5 was associated with poor prognosis in breast cancer based on the results obtained from the Kaplan-Meier Plotter database. In addition, the wound healing assay indicated that the inhibition of migration due to miR-203 overexpression in SUM159 cells was reversed by FKBP5 overexpression. These results suggested that miR-203 may directly target FKBP5. In addition, Gene Set Enrichment Analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis revealed that miR-203 might play a role in breast cancer via the ‘fatty acid degradation’ KEGG pathway. Notably, the levels of fatty acids were significantly reduced in SUM159 cells transfected with miR-203 mimics compared with SUM159 cells transfected with miR-203 NC when assessed by the fatty acid content assay. Finally, virtual screening analysis revealed that ZINC000003944422 may be a potential inhibitor of FKBP5. In summary, the present study demonstrated that miR-203 may directly target FKBP5 in breast cancer via fatty acid degradation and potential drugs, hence providing a novel treatment approach for breast cancer.

Hippo/TEAD4 signaling pathway as a potential target for the treatment of breast cancer.

Breast cancer is the most common type of cancer among women worldwide. The Hippo signaling pathway is strongly associated with cell proliferation, migration, invasion, metastasis and resistance to breast cancer treatment. The upstream factors involved in the Hippo signaling pathway, including mammalian Ste20 kinases 1/2, large tumor suppressor kinases 1/2 and transcription coactivator Yes-associated protein (YAP)/transcriptional coactivator with PDZ-binding motif (TAZ), have been extensively studied as they are considered therapeutic targets for breast cancer. Recently, it has been suggested that the transcriptional enhancer factor domain (TEAD) family of transcription factors, particularly TEAD4, plays an important role in breast cancer. TEADs interact with YAP/TAZ to act as transcription factors. Notably, recent studies have demonstrated that TEAD4 may also function in a YAP/TAZ-independent manner and serve as a prognostic marker for breast cancer. The present review summarizes the current research on the effect of the aberrant activation of the Hippo signaling pathway on breast cancer progression. Furthermore, the latest advances on the role of the TEAD family in breast cancer are highlighted, and the role of TEAD4 as a potential target for therapeutic intervention in breast cancer is discussed.

Abstract PS6-58: The impact of baseline modified glasgow prognostic score (mGPS) on survival outcomes in in indigenous and non-indigenous patients with advanced breast cancer patients of Western Australia

Abstract Introduction: The modified Glasgow prognostic score (mGPS) is known to be useful in prognostication of multiple cancers. The mGPS, which integrates albumin and C-reactive protein, could assist as a possible prognostic marker in breast cancer. Indigenous women have inferior breast cancer survival to non-Indigenous women and may also have a differing inflammatory environment. Here, we examine the utility and impact of baseline mGPS on survival outcomes in women with metastatic breast cancer by Indigenous status. Methods: We retrospectively collected data from the Western Australian Cancer Registry and electronic records for patients diagnosed with breast cancer between 2001 and 2016 with confirmed metastatic disease. Overall survival (OS) were measured from the date of diagnosis until death. mGPS comprised scores of one point given for CRP > 10 mg/L and/or albumin < 3.5 g/dL, therefore having a value of 0 to 2. Results: Of 152 patients with metastatic breast cancer, 89 patients had all relevant data available and were included with a median follow up of 120 months. The median age was 55.3 years. Baseline mGPS was 0 in 46.6 %, 1 in 34.8 % and 2 in 18.6 %. Median OS across Indigenous and non-Indigenous patients combined was significantly worse moving from the mGPS-0 group through mGPS-1 to mGPS 2, 50.0 v 30.0 v 8.0 months respectively (p<0.0001). Looking at the groups separately, both cohorts separately demonstrated inferior median OS in mGPS-2 compared to mGPS-1 patients, 9 v 32 months (p=0.02) for Indigenous patients, and 2.0 vs 25.0 months for non_Indigenous patients (p=0.001). The correlation between mGPS and the neutrophil-to-lymphocyte ratio (NLR) was weak with a Pearson correlation R-value of 0.184 (p=0.085) Conclusion: The study shows that the mGPS is an independent prognostic factor in advanced-stage disease. A higher baseline mGPS score was associated with worse survival in Indigenous and non-Indigenous patients. A larger prospective study is needed to validate the results, inclusive of assessing links between mGPS and OS in different breast cancer sub-types. Citation Format: Azim Khan, Hilary Martin, Lisa Spalding, Andrew Redfern. The impact of baseline modified glasgow prognostic score (mGPS) on survival outcomes in in indigenous and non-indigenous patients with advanced breast cancer patients of Western Australia [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS6-58.

Abstract PS6-42: Prognostic value of neutrophil to lymphocyte ratio in patients with breast cancer in a Mexican population

Abstract Background: Inflammatory serum markers (IBM) have emerged as prognostic factors in solid tumors. In breast cancer (BC) in particular, an elevated neutrophil to lymphocyte ratio (NLR) has been linked to lower overall survival (OS) and disease-free survival (DFS), although a standard cutoff value has not been established. In Mexico there is no information regarding NLR as a prognostic marker in BC. The aim of the present study was to assess NLR as prognostic factor in BC patients in a Mexican population. Methods: This single-center retrospective and descriptive study included patients, >18 years old with histological diagnosis of BC who were treated at Medica Sur oncological center in Mexico City between January 2008 and December 2019. The patients were divided into two groups according to their NLR. NLR was calculated using the following formula: absolute neutrophil count / absolute lymphocyte count (mm3). NLR was considered as elevated (>2), and as low (≤2) based on data from previous studies. The primary endpoint was OS. Statistical analysis was performed with SPSS v25. The associations between PNI and clinicopathologic characteristics were analyzed using Pearson's χ2 test. Kaplan-Meier and log-rank test methods were used for survival analysis. The prognostic value of the pre-treatment NLR was assessed by univariate and multivariate analysis. P<0.05 was considered to indicate a statistically significant difference. This study was approved by our scientific and bioethical committee. Results: A total of 110 patients were included in the final analysis. All patients had infiltrating ductal carcinoma; 18.2% were triple negative, 23.6% over-expressed HER 2 and around 51% had hormone receptor expression. 15.5% had metastatic disease. Median follow-up was 65 months. Mean NLR at diagnosis was 2.82 (SD 2.59). Mean NLR in patients with triple negative BC (TNBC) was significantly higher than in patients with non TNBC (p= 0.055), no other statistically significant associations were found between mean NLR and clinical characteristics. Median OS was not reached in two groups, but the 100 months OS was higher in NLR ≤2 group vs NLR > 2 group (90 vs 78%) (p= 0.079). In univariate and multivariate analysis, triple negative histological subtype and elevated NLR were independent prognostic indicators of poor survival. Conclusion: An elevated pretreatment NLR was an independent prognostic factor, associated with lower OS in patients with BC with local and advanced disease, although statistically significant association was not found. NLR is an accessible and minimally invasive marker with clinical value for evaluation of the prognosis of BC patients, especially in resource-limited settings. Citation Format: Andrés Mauricio Arroyave-Ramírez, Edgar Varela-Santoyo, Pamela Denisse Soberanis-Piña, Hector Hugo Buerba-Vieregge, Daniel Motola-Kuba. Prognostic value of neutrophil to lymphocyte ratio in patients with breast cancer in a Mexican population [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS6-42.

Initial Identification of UDP-Glucose Dehydrogenase as a Prognostic Marker in Breast Cancer Patients, Which Facilitates Epirubicin Resistance and Regulates Hyaluronan Synthesis in MDA-MB-231 Cells.

UDP-glucose-dehydrogenase (UGDH) synthesizes UDP-glucuronic acid. It is involved in epirubicin detoxification and hyaluronan synthesis. This work aimed to evaluate the effect of UGDH knockdown on epirubicin response and hyaluronan metabolism in MDA-MB-231 breast cancer cells. Additionally, the aim was to determine UGDH as a possible prognosis marker in breast cancer. We studied UGDH expression in tumors and adjacent tissue from breast cancer patients. The prognostic value of UGDH was studied using a public Kaplan–Meier plotter. MDA-MB-231 cells were knocked-down for UGDH and treated with epirubicin. Epirubicin-accumulation and apoptosis were analyzed by flow cytometry. Hyaluronan-coated matrix and metabolism were determined. Autophagic-LC3-II was studied by Western blot and confocal microscopy. Epirubicin accumulation increased and apoptosis decreased during UGDH knockdown. Hyaluronan-coated matrix increased and a positive modulation of autophagy was detected. Higher levels of UGDH were correlated with worse prognosis in triple-negative breast cancer patients that received chemotherapy. High expression of UGDH was found in tumoral tissue from HER2--patients. However, UGDH knockdown contributes to epirubicin resistance, which might be associated with increases in the expression, deposition and catabolism of hyaluronan. The results obtained allowed us to propose UGDH as a new prognostic marker in breast cancer, positively associated with development of epirubicin resistance and modulation of extracellular matrix.

Low Expression of the Polycomb Protein RING1 Predicts Poor Prognosis in Human Breast Cancer.

BackgroundTo date, breast cancer remains the most common malignant tumor in women. In recent years, a growing number of studies on polycomb proteins have been conducted. The Ring finger protein1 (RING1), an essential component of the polycomb family of proteins, plays vital roles in the tumorigenesis of various cancer types. However, further research is required in determining RING1 expression and prognostic value in breast cancer.MethodRING1 expression level in multiple cancer types was evaluated using the XENA and UALCAN databases. Real-time quantitative PCR (real-time qPCR) and immunohistochemistry (IHC) were used to confirm this expression. The prognostic value was analyzed using our follow-up data and the Kaplan–Meier plotter website. RING1 co-expressed genes and its promoter methylation level were calculated using the cBioPortal and UALCAN online tools. The gene ontology (GO) and the Kyoto encyclopedia of Genes and Genomes (KEGG) pathway enrichment were analyzed using the DAVID online analysis tool.ResultRING1 expression was upregulated in CHOL (Bile Duct Cancer), ESCA (Esophageal Cancer), LIHC (Liver Cancer), and PCPG (Pheochromocytoma & Paraganglioma). However, its expression level was decreased in COAD (Colon Cancer), KICH (Kidney Chromophobe), KIRP (kidney papillary cell carcinoma), THCA (Thyroid Cancer), and BRCA (Breast carcinoma). RING1 low expression is an unfavorable prognostic factor in many cancer patients, especially in breast cancer patients. For breast cancer, the IHC result showed that RING1 protein expression significantly and negatively correlates with tumor size (P = 0.029), LNM (P = 0.017), TNM stage (P = 0.016), ER (P = 0.005), Ki67 (P = 0.015), and p53 status (P = 0.034). Moreover, the multivariate Cox regression model indicated that RING1 (P = 0.038) and ER (P = 0.029) expressions were independent prognostic markers for breast cancer. RING1 co-expressed genes were selected and included HDAC10, PIN1, CDK3, BAX, and BAD. GO analysis and KEGG pathway analyses revealed that RING1 related genes, were mainly enriched in “regulation of transcription”, “apoptotic process”, “protein transport”, “protein binding”, “Notch signaling pathway”, and “Homologous recombination”.ConclusionRING1 expression was downregulated in breast cancer, and its low expression was associated with worse disease outcomes. RING1 may act as a new prognostic biomarker for breast cancer.

Expression and Gene Regulation Network of Metabolic Enzyme Phosphoglycerate Mutase Enzyme 1 in Breast Cancer Based on Data Mining

The metabolic enzyme phosphoglycerate mutase enzyme 1 (PGAM1) is a key enzyme in the glycolysis pathway, and glycolysis is closely related to cancer progression, suggesting that PGAM1 may have important functions in breast cancer. We used sequencing data from the Oncomine database and UALCAN database to analyze the expression of PGAM1 and its influence on the clinicopathological characteristics of breast cancer. LinkedOmics was used to identify genes related to PGAM1 expression, kinases, miRNAs, and transcription factors that were significantly related to PGAM1 through GSEA. cBioPortal was used to identify the alternation frequency and form of PGAM1 in breast cancer. The expression level of PGAM1 in breast cancer was significantly higher than that in normal tissues. Moreover, the expression level of PGAM1 is closely related to the molecular subtype and TP53 mutation status. The expression level of PGAM1 in HER2-positive and triple-negative tumors was significantly higher than that of luminal type. The expression level of PGAM1 in TP53-mutant tumors was higher than that in non-TP53-mutant tumors. In addition, the overall survival of patients with high PGAM1 expression was significantly worse than that of patients with low expression ( P = 0.0077 ). Through GSEA analysis, we found multiple kinases, miRNAs, and transcription factors significantly related to PFKFB4. cBioPortal analysis showed that the mutation rate of PGAM1 in breast cancer was relatively low (4%), and the main form of mutation was high mRNA expression. This study suggests that PGAM1 is a potential diagnostic and prognostic marker in breast cancer. Through data mining, we revealed the potential regulatory network information of PGAM1, laying a foundation for further research on the role of PGAM1 in breast cancer.

COX-2 “Cyclooxygenase 2“as a Prognostic marker in Breast Cancer

Background: Breast cancer is the most commonly diagnosed cancer and the leading cause of cancer death among women in both developed and developing countries. Globally, the incidence rate of breast cancer has been rising rapidly over the past few decades.Cycloxygenase-2 (COX-2) regulates tumor growth, invasion and metastasis in breast cancer. Objective: The objective of our study is to evaluate COX-2 expression in breast cancer as a poor prognostic factor. Methods: Formalin-fixed and paraffin-embedded tissue blocks were studied for COX-2 expression by immunohistochemistry in 100 patients diagnosed as breast carcinoma. The relationship between COX-2 expression and various clinico-pathological parameters was studied. Results: The results of our study suggest an association of the expression of COX-2 to the poor prognostic factors in breast cancer, such as larger tumor size, positive lymph node status, higher T stage and N stage as studying the association between Cox-2 protein expression and different clinco-pathologic features revealed that larger tumor size (> 5) and lymph node metastasis showed statistical significant association with Cox2 protein expression (p = 0.014 and p = 0.031, respectively). While rest of clinico-pathologic features such as age, stage, hormonal receptor status, HER2/NEU and histopathological features showed no statistical significant association. Conclusion: Our study established the role of COX-2 in carcinogenesis and its association with adverse prognostic factors.

GATA3 - IS IT A PROGNOSTIC MARKER IN BREAST CARCINOMA?

GATA binding proteins are among the new prognostic factors being explored. My area of interest is GATA3 to correlate it with tumor size, nodal status ,tumor grade, Estrogen receptor, Progesterone receptor and HER2neu receptors in breast carcinoma patients This was a quantitative correlational study Patient were enrolled using consecutive non probability sampling method. GATA3 was inversely related with tumor size ,nodal status and grade. Estrogen positivity was associated with higher GATA3. However Progesterone and HER2Neu receptors statuses did not show any significant association with GATA3 levels. This study concludes that GATA3 expression can be used as a prognostic marker of breast cancer.

Increased Expression of DNA2 Was Linked to Poor Prognosis in Breast Cancer.

DNA double-strand break (DSB) repaired by homologous recombination (HR) is an essential process for breast cancer cells to survive. DNA2 nuclease acts parallel to homologous recombination (HR). Here, we investigated the detailed clinical attribute of DNA2 in breast cancer and the role of DNA2 in breast cancer cells' growth. We found that elevated expression of DNA2 was obviously linked to poor prognosis in breast cancer. Further, DNA2 expression was increased in the ER-negative group, PR-negative group, HER2-positive group, and high-grade group via analyzing 2,509 breast cancers in “cBioportal” and 3,063 breast cancer data in “bc-GenExMiner.” Besides, the immunohistochemical staining in 26 breast cancer tissues also showed that elevated expression of DNA2 was correlated with ER-/PR-/HER+. To further detect the role of DNA2 in breast cancer cells, we took GESA, GO, and KEGG analyses and found that DNA2 was enriched in cell cycle and DNA replication pathways. Furthermore, silencing of DNA2 inhibited cell growth in T47D and MD-MB-231 breast cancer cells and suppressed tumor growth in vivo, indicating DNA2 functioned importantly in breast cancer progression and maybe a potential prognostic marker in breast cancer. Our research reveals that DNA2 is a biomarker for diagnosis and prognosis in breast cancer from multiple perspectives and gives a new clue for further preclinical and clinical investigation.

High B3GALT5 expression confers poor clinical outcome and contributes to tumor progression and metastasis in breast cancer

BackgroundExistence of breast cancer stem cells (BCSCs) is implicated in disease relapse, metastasis, and resistance of treatment. β1,3-Galactosyltransferase 5 (B3GALT5) has been shown to be a pro-survival marker for BCSCs. However, little is known about the prognostic significance of B3GALT5 in breast cancer.MethodsPaired tissues (tumor part and adjacent non-tumor part) from a cohort of 202 women with breast cancer were used to determine the expression levels of B3GALT5 mRNA by qRT-PCR. Kaplan–Meier and multivariable Cox proportional hazard models were used to assess survival differences in terms of relapse-free survival (RFS) and overall survival (OS). Both breast cancer cells and cancer stem cells (BCSCs) were used to see the in vitro effects of knockdown or overexpression of B3GALT5 on cell migration, invasion, and epithelial-to-mesenchymal transition (EMT). A patient-derived xenograft (PDX) model was used to see the in vivo effects of knockdown of B3GALT5 in BCSCs on tumor growth and metastasis.ResultsHigher expression of B3GALT5 in 202 breast cancer tissues, especially in adjacent non-tumor tissue, correlated with poor clinical outcomes including shorter OS and RFS in all patients, especially those with early stage breast cancer. In vitro studies showed B3GALT5 could enhance cell migration, invasion, mammosphere formation, and EMT. Of note, B3GALT5 upregulated the expression of β-catenin and EMT activator zinc finger E-box binding homeobox 1 (ZEB1) pathway in BCSCs. In vivo studies showed B3GALT5 expression in BCSCs is critical for not only tumor growth but also lymph node and lung metastasis in PDX mice.ConclusionOur results demonstrated the value of B3GALT5 as a prognostic marker of breast cancer, especially among the early stage patients, and its crucial roles in regulating EMT, cell migration, and stemness thereby promoting breast cancer progression.

Circulating Tumor Cells and Bevacizumab Pharmacokinetics during Neoadjuvant Treatment Combining Chemotherapy and Bevacizumab for Early Breast Cancer: Ancillary Analysis of the AVASTEM Trial.

Simple SummaryWe recently published the results of the AVASTEM study, in which we explored the impact of the addition of an angiogenesis inhibitor (bevacizumab) to standard pre-operative chemotherapy for breast cancer. In this work, we aimed to identify biological parameters correlated to prognosis and treatment efficacy. To do so we explored if circulating tumor cells (CTCs), that are cells released by the tumor and that can be detected in the bloodstream of a patient; can predict the outcome of the women treated in this study. We also analyzed if bevacizumab concentration in the blood during treatment was associated with outcome, i.e., response to treatment and survival. We observed that CTCs detection before treatment initiation is associated with survival but was not correlated to local response to treatment. We moreover found that CTC status after three weeks of chemotherapy was not correlated to outcome as well as bevacizumab levels before and during treatment. However, bevacizumab concentration tended to be associated with an increase of hematological toxicities during the study. We thus show in this work that CTCs detection at baseline is prognostic for patients with breast cancer receiving a pre-operative chemotherapy-bevacizumab combination, and that this prognostic value was independent of tumor response.The phase II AVASTEM trial explored the impact of chemotherapy-bevacizumab combination on breast cancer stem cells in the neoadjuvant setting. We aimed to identify biological features associated with preoperative chemotherapy efficacy and prognosis by analyses of circulating tumor cells (CTCs) and bevacizumab pharmacokinetics (PK). The main objective was to assess the prognostic (relapse-free survival and overall survival) and predictive (pathological complete response, pCR) values of CTCs (CellSearch technology) and bevacizumab PK (ELISA). Seventy-five patients were included. Out of them 50 received bevacizumab-chemotherapy and 25 received chemotherapy alone. CTC results were available for 60 patients and PK data for 29 patients in the experimental arm. The absence of CTC at inclusion was correlated to better outcome. Five-years overall survival (OS) was 91% for CTC-negative patients vs. 54% for CTC-positive cases (HR = 6.21; 95%CI (1.75–22.06), p = 0.001, log-rank test). Similar results were observed for RFS with 5 y-RFS of 78% vs. 44% (HR = 3.51; 95%CI (1.17–10.52), p = 0.017, log-rank test). However, CTC status at baseline was not predictive of pCR (p = 0.74). CTC status after one cycle was not a significant prognostic factor (HR = 1.56; 95%CI (0.19–12.67); p = 0.68 for OS and HR = 2.76; 95%CI (0.60–12.61); p = 0.17 for RFS, log-rank test). Bevacizumab serum levels could not predict pCR and survival. PK values were not associated with treatment-related toxicities. In conclusion, CTCs detection at baseline is a prognostic marker for breast cancer receiving a neoadjuvant chemotherapy-bevacizumab combination independently of tumor response.

KCTD12 is a prognostic marker of breast cancer and correlates with tumor immune cell infiltration.

BackgroundAnnually, breast cancer (BC) is the most common newly diagnosed cancer in females. The relatively crude measures of the molecular phenotypes of BC have not provided a comprehensive understanding of its molecular architecture. To a certain extent, this has resulted in many patients being over- or undertreated. Therefore, novel biomarkers that help to improve patients’ outcomes are required. The potassium channel tetramerization domain containing 12 (KCTD12) is one such candidate.MethodsRibonucleic acid-sequencing (RNA-Seq) filings along with corresponding clinical information of BC samples were obtained from The Cancer Genome Atlas (TCGA) program databases to evaluate the associations between KCTD12 expression levels and clinical features. The prognostic value of KCTD12 in patients was examined by Kaplan-Meier survival analysis and PrognoScan database analysis. To identify the main functions of KCTD12 in BC, we performed gene set enrichment analysis (GSEA) in BC samples and cell lines. The correlations between KCTD12 expression and tumor-infiltrating lymphocyte quantities was confirmed using two online tools: Tumor Immune Estimation Resource and the Gene Expression Profiling Interaction Analysis 2.ResultsKCTD12 expression was significantly decreased in cancer samples compared to normal samples, and was lowly expressed in aggressive disease relative to initial disease. Patients with lower KCTD12 expression levels showed a shorter overall survival and a shorter recurrence-free survival, indicating a worse prognosis. We found that genes of BC in the high-KCTD12 expression group were enriched in immune response pathways. Finally, the positive correlations between the expression of tumor-infiltrating lymphocytes, programmed cell-death ligand 1 (PD-L1), and programmed cell-death protein 1 (PD-1), and KCTD12 expression were confirmed.ConclusionsKCTD12 can be considered a biomarker to predict the prognosis of BC patients. KCTD12 may also help to predict patient response to PD-L1 or PD-1 inhibitor treatment.

The impact of G protein-coupled oestrogen receptor 1 on male breast cancer: a retrospective analysis.

IntroductionThe G protein-coupled oestrogen receptor 1 (GPER-1) is a potential prognostic marker in breast cancer. However, its role in male breast cancer (MBC) is still unknown. This study evaluates the expression of GPER-1 in MBC samples and correlates these data with clinical and pathological parameters including patients’ survival.Material and methodsFor this retrospective analysis of a prospectively maintained cohort of patients with MBC, we examined 161 specimens for GPER-1 expression using immunohistochemistry. An immunoreactive score (IRS) was calculated based on staining intensity and the percentage of positive tumour cells. Then, we correlated GPER-1 IRS with clinical and pathological parameters, and overall and relapse-free survival.ResultsAbout 40% of MBC samples were positive for GPER-1 expression (IRS ≥ 4). There was no significant correlation with clinicopathological parameters, such as hormone receptor status or grading. However, a statistical trend was observed for tumour size (≥ 2 cm, p = 0.093). Kaplan-Meier survival analysis revealed no significant correlation with relapse-free survival. However, there was a significant correlation with overall survival, but when we adjusted the log-rank p-value to compensate for the cut-off point optimization method, it rose above 0.1. Additionally, GPER-1-positive patients were older at diagnosis. When adjusted for age by multivariable Cox regression analysis, the significance of GPER-1 status for survival was further reduced.ConclusionsWe found no significant prognostic value of GPER-1 in this MBC cohort as anticipated from studies on female BC. Future studies with higher sample size are needed to further verify a potential sex-specific role of GPER-1.

Assessment of Ki67 in Breast Cancer: Updated Recommendations From the International Ki67 in Breast Cancer Working Group.

Ki67 immunohistochemistry (IHC), commonly used as a proliferation marker in breast cancer, has limited value for treatment decisions due to questionable analytical validity. The International Ki67 in Breast Cancer Working Group (IKWG) consensus meeting, held in October 2019, assessed the current evidence for Ki67 IHC analytical validity and clinical utility in breast cancer, including the series of scoring studies the IKWG conducted on centrally stained tissues. Consensus observations and recommendations are: 1) as for estrogen receptor and HER2 testing, preanalytical handling considerations are critical; 2) a standardized visual scoring method has been established and is recommended for adoption; 3) participation in and evaluation of quality assurance and quality control programs is recommended to maintain analytical validity; and 4) the IKWG accepted that Ki67 IHC as a prognostic marker in breast cancer has clinical validity but concluded that clinical utility is evident only for prognosis estimation in anatomically favorable estrogen receptor–positive and HER2-negative patients to identify those who do not need adjuvant chemotherapy. In this T1-2, N0-1 patient group, the IKWG consensus is that Ki67 5% or less, or 30% or more, can be used to estimate prognosis. In conclusion, analytical validity of Ki67 IHC can be reached with careful attention to preanalytical issues and calibrated standardized visual scoring. Currently, clinical utility of Ki67 IHC in breast cancer care remains limited to prognosis assessment in stage I or II breast cancer. Further development of automated scoring might help to overcome some current limitations.

Identification of breast cancer prognostic modules via differential module selection based on weighted gene Co-expression network analysis

Breast cancer is a complex cancer which includes many different subtypes. Identifying prognostic modules, i.e., functionally related gene networks that play crucial roles in cancer development is essential in breast cancer study. Different subtypes of breast cancer correspond to different treatment methods. The purpose of this study is to use a new method to divide breast cancer into different prognostic modules, so as to provide scientific basis for improving clinical management.The method is based on comparing similarities between modules detected from different weighted gene co-expression networks. The method was applied on genomic data of breast cancer from The Cancer Genome Atlas database and was applied to select differential modules between two groups of patients with significant differences in survival times. It was compared with a previously proposed module selection method. The result shows that our method outperforms the previously proposed one. Moreover, within the identified two differential modules, the first one is highly enriched with genes involved in hormone responds, the second one is highly related with biological process engaged in M-phase. The two modules were further validated by log-rank test in the validation dataset GSE3494. Both of the two modules show significantly different with p-values less than 0.02. The identified two modules confirmed previous findings including importance of biological networks in breast cancer involved in hormone response and M-phase. Out of the top twenty hub genes in the two modules, fifteen genes were previously shown to be prognostic markers for breast cancer.

Potential Diagnostic and Prognostic Utility of miR-141, miR-181b1, and miR-23b in Breast Cancer.

miRNAs, a group of short noncoding RNAs, are key regulators of fundamental cellular processes and signaling pathways. Dysregulation of miRNA expression with known oncogenic or tumor suppressor functions has been associated with neoplastic transformation. Numerous studies have reported dysregulation of miRNA-141, miR-181b1, and miR-23b in a wide range of malignancies, including breast cancer. To the best of our knowledge, no previous study had demonstrated the expression of miR-141-3p, miR-181b1-5p, and miR-23b-3p in different histological grades and molecular subtypes of breast cancer. Here, we identified differential expression of these three miRNAs in breast cancer tissues compared with benign breast fibroadenomas. In addition, high expression levels of miR-141-3p and miR-181b1-5p are strongly associated with aggressive breast carcinomas. We also confirmed the clinical potential of using the three miRNAs individually or combined as diagnostic and prognostic markers in breast cancer. Using bioinformatics analyses, we identified 23 hub genes of these three miRNAs which are involved in key signaling pathways in breast cancer. Furthermore, the KM plotter online database analysis demonstrates the association between elevated expression of miR-141 and miR-181b and shorter overall survival of breast cancer patients. Together, our data suggest an oncogenic role of the studied miRNAs and highlight their molecular roles and potential clinical applications in breast cancer.