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Aims/Background The clinical presentation of non-lactational mastitis (NLM) shares similarities with some symptoms and examination results of breast cancer (BC), which can lead to misdiagnosis or delayed treatment. Current studies on breast lesions mostly focus on the diagnostic performance of a single imaging technique. This study aims to construct a discrimination diagnostic model for NLM and BC based on such imaging features as ultrasound and magnetic resonance imaging (MRI) and to validate the application value of the model, assisting clinicians in improving disease diagnosis and refining medical decisions. Methods This study is a retrospective analysis. Clinical data of 108 patients suspected of NLM based on imaging diagnosis, admitted to The First Affiliated Hospital with Nanjing Medical University between May 2018 and August 2023, were collected. Among them, 94 cases were pathologically confirmed as NLM and 14 cases as BC. Univariate and multivariate logistic regression analyses were performed on the patients' clinical data, ultrasound features, and MRI features to select the risk factors for discriminating NLM and BC, and construct a discrimination model. The discrimination performance of the model was analyzed with the receiver operating characteristic (ROC) curve, decision curve analysis (DCA), and calibration curve. Results In the NLM group, there were 24 cases of granulomatous lobular mastitis (25.53%) and 70 cases of plasma cell mastitis (74.47%). In the BC group, there were 2 cases of infiltrating ductal carcinoma, 2 cases of atypical hyperplasia, 3 cases of papillary carcinoma, and 7 cases of ductal carcinoma in situ . Age, internal blood flow, calcification, edge, enhancement characteristics, apparent diffusion coefficient (ADC) values, and time-intensity curve (TIC) type were independent factors for differentiating NLM and BC ( p < 0.05). The ROC analysis showed that the area under the curve of the model for discriminating NLM and BC was 0.920. The DCA results showed that the model had high net benefits for discriminating NLM and BC. The calibration curve analysis showed that the model had good consistency with the actual diagnosis of NLM and BC, with a chi-square value of 4.545 and a p -value of 0.155 according to the Hosmer-Lemeshow test. Conclusion Age, internal blood flow, calcification, edge, enhancement characteristics, ADC, and TIC curve types are important factors in distinguishing NLM and BC, and the model based on the above characteristics to distinguish NLM and BC has a high net benefit in distinguishing the two.

Minimally invasive thermal therapy is a successful alternative treatment to surgery in solid tumors with high complete ablation rates, however, tumor recurrence remains a concern. Central memory CD8 + T cells (T CM ) play important roles in protection from chronic infection and cancer. Here we find, by single-cell RNA analysis of human breast cancer samples, that although the memory phenotype of peripheral CD8 + T cells increases slightly after microwave ablation (MWA), the metabolism of peripheral CD8 + T cells remains unfavorable for memory phenotype. In mouse models, glycolysis inhibition by 2-deoxy-D-glucose (2DG) in combination with MWA results in long-term anti-tumor effect via enhancing differentiation of tumor-specific CD44 hi CD62L + CD8 + T CM cells. Enhancement of CD8 + T CM cell differentiation determined by Stat-1, is dependent on the tumor-draining lymph nodes (TDLN) but takes place in peripheral blood, with metabolic remodeling of CD8 + T cells lasting the entire course of the the combination therapy. Importantly, in-vitro glycolysis inhibition in peripheral CD8 + T cells of patients with breast or liver tumors having been treated with MWA thrice leads to their differentiation into CD8 + T CM cells. Our work thus offers a potential strategy to avoid tumor recurrence following MWA therapy and lays down the proof-of-principle for future clinical trials., (© 2024. The Author(s).)

Background: Immune checkpoint blockade has shown low response rates for advanced breast cancer, and combination strategies are needed. Microwave ablation (MWA) may be a trigger of antitumor immunity. This window-of-opportunity trial (ClinicalTrials.gov: NCT04805736) was conducted to determine the safety and feasibility of preoperative camrelizumab (an anti-PD-1 antibody) combined with MWA in the treatment of early-stage breast cancer., Methods: Sixty participants were randomized to preoperatively receive single-dose camrelizumab alone (n = 20), MWA alone (n = 20), or camrelizumab+MWA (n = 20). A random number table was used to allocate interventions. The primary outcome was the safety and feasibility of MWA combined with camrelizumab., Findings: Camrelizumab and MWA were well tolerated alone and in combination without delays in prescheduled surgery. No treatment-related grade III/IV adverse events were observed. Different from in the single-dose camrelizumab or MWA group, participants showed stable counts of blood cells after combination therapy. After combination therapy, peripheral CD8 + T cells showed enhanced cytotoxic and effect-memory functions. Clonal expansional CD8 + T cells showed higher cytotoxic activity and effector memory- and tumor-specific signatures than emergent clones after combination therapy. Enhanced interactions between clonal expansional CD8 + T cells and monocytes were observed, suggesting that monocytes contributed to the enhanced functions of clonal expansional CD8 + T cells. Major histocompatibility complex (MHC) class I-related pathways and interferon signaling pathways were activated in monocytes by combination therapy., Conclusions: Camrelizumab combined with MWA was feasible for early-stage breast cancer. Peripheral CD8 + T cells were activated after combination therapy, dependent on monocytes with activated MHC class I pathways., Funding: This study was supported by the Natural Science Foundation of Jiangsu Province (BK20230017)., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Purpose: This study aimed to explore a novel position of mammography named axilla view in axillary lymph node (ALN) evaluation in breast cancer., Patients and Methods: Patients were prospectively enrolled and scheduled for mammography before surgery. Investigated imaging patterns included mediolateral oblique (2D-MLO) and axilla view (2D-axilla) of mammography, and axilla view of digital breast tomosynthesis (3D-axilla). The correlation of ALN numbers between imaging and pathology was analyzed. Diagnostic performance was analyzed via AUC., Results: 75 patients were included. A larger and clearer axillary region was displayed in axilla view. The total number of ALNs detected under 2D/3D-axilla view was significantly higher than that under 2D-MLO view (4.6 vs. 2.5, P < .001; 5.6 vs. 4.6, P = .034). Correlations between number of positive ALNs detected under 2D/3D-axilla view and pathologically confirmed metastatic ALNs were stronger than 2D-MLO view (Pearson correlation coefficients: 0.7084,0.7044 and 0.4744). The proportion of cases with ≥5 positive ALNs detected under 3D-axilla view was significantly higher than that under 2D-MLO (38.2% vs. 14.7%, P = .028). The overweight and obese group showed a higher AUC value than the underweight and lean group in ALN evaluation, although not significantly (2D-MLO: 0.7643 vs. 0.6458, P = .2656; 2D-axilla: 0.8083 vs. 0.6586, P = .1522; 3D-axilla: 0.8045 vs. 0.6615, P = .1874). This difference was more pronounced in axilla view., Conclusion: Axilla view exhibited advantages over conventional MLO view in the extent of axilla displayed by mammography in breast cancer. Further studies with larger sample sizes are needed., Competing Interests: Disclosure The authors have stated that they have no conflicts of interest., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Introduction: Locoregional recurrent breast cancers have a poor prognosis. Little is known about the prognostic impact of immune microenvironment, and tertiary lymphoid structures (TLSs) in particular have not been reported. Thus, we aimed to characterize the immune microenvironment in locoregional recurrent breast tumors and to investigate its relationship with prognosis., Methods: We retrospectively included 112 patients with locoregional recurrent breast cancer, and hematoxylin-eosin staining and immunohistochemical staining (CD3, CD4, CD8, CD19, CD38, and CD68) were performed on locoregional recurrent tumor samples. The association of immune cells and TLSs with progression-free survival (PFS) were analyzed by survival analysis., Results: We found more immune cells in the peritumor than stroma. After grouping according to estrogen receptor (ER) status, a low level of peritumoral CD3+ cells in ER+ subgroup (p = 0.015) and a low level of stromal CD68+ cells in ER- subgroup (p = 0.047) were both associated with longer PFS. TLSs were present in 68% of recurrent tumors, and CD68+ cells within TLSs were significantly associated with PFS as an independent prognostic factor (p = 0.035). TLSs and immune cells (CD3, CD38, and CD68) within TLSs were associated with longer PFS in ER- recurrent tumors (p = 0.044, p = 0.012, p = 0.050, p < 0.001, respectively), whereas CD38+ cells within TLSs were associated with shorter PFS in ER+ recurrent tumors (p = 0.037)., Conclusion: Our study proposes potential predictors for the clinical prognosis of patients with locoregional recurrent breast cancer, emphasizing the prognostic value of immune cells within TLSs, especially CD68+ cells., (© 2023 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Neoadjuvant chemotherapy (NCT) is the standard treatment for patients with locally advanced breast cancer (LABC). The predictive value of heterogeneous circulating tumor cells (CTCs) in NCT response has not been determined. All patients were staged as LABC, and blood samples were collected at the time of biopsy, and after the first and eighth NCT courses. Patients were divided into High responders (High-R) and Low responders (Low-R) according to Miller-Payne system and changes in Ki-67 levels after NCT treatment. A novel SE-i·FISH strategy was applied to detect CTCs. Heterogeneities were successfully analyzed in patients undergoing NCT. Total CTCs increased continuously and were higher in Low-R group, while in High-R group, CTCs increased slightly during NCT before returning to baseline levels. Triploid and tetraploid chromosome 8 increased in Low-R but not High-R group. The number of small CTCs in Low-R group increased significantly until the last sample, however, remained constant in High-R group. The patients with more CTCs had shorter PFS and OS than those with less CTCs after the eighth course of NCT. Total CTCs following NCT could predict patients' responses. More detailed characterizations of CTC blood profiles may improve predictive capacity and treatments of LABC., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Objective: We aimed to investigate the impact of human epidermal growth factor receptor 2 status (human epidermal growth factor receptor 2-low versus human epidermal growth factor receptor 2-zero) on pathological response to neoadjuvant chemotherapy and survival outcomes in early-stage breast cancer., Methods: Patients with primary invasive breast cancer received neoadjuvant chemotherapy between July 2018 and July 2021 were identified from six hospitals. The primary efficacy end-point was total pathological complete response. The second short-term efficacy end-points include breast pathological complete response, axillary lymph nodes pathological complete response and the score of Miller-Payne grade. Long-term efficacy end-point was disease-free survival., Results: 429 patients with human epidermal growth factor receptor 2 negative invasive tumors were included, 267 (62.24%) had human epidermal growth factor receptor 2-low tumors. Hormone receptor-positive patients had a higher percentage of human epidermal growth factor receptor 2-low tumors compared to hormone receptor-negative patients (71.97% versus 42.14%). The pathological response rate was significantly lower in human epidermal growth factor receptor 2-low tumors than in human epidermal growth factor receptor 2-zero tumors for total patients in univariate analysis, including the rates of total pathological complete response (5.2% versus 14.2%), breast pathological complete response (6.4% versus 17.3%), nodes pathological complete response (26.3% versus 37.7%) and MP4-5 (21.2% versus 33.8%). Subgroup analysis showed that the rates of total pathological complete response, breast pathological complete response and MP4-5 were also significantly lower in human epidermal growth factor receptor 2-low tumors versus human epidermal growth factor receptor 2-zero tumors in both univariate and multivariate analysis in hormone receptor-negative subgroup. With the median follow-up of 24 months, disease-free survival was comparable between these two subgroups (P = 0.816)., Conclusions: Our results demonstrate that human epidermal growth factor receptor 2-low tumors achieved a significantly lower pathological complete response rate with conventional chemotherapy than those with human epidermal growth factor receptor 2-zero tumors, especially for hormone receptor-negative group. Large, randomized, prospective studies are needed to confirm our data and further evaluate the prognostic value of human epidermal growth factor receptor 2-low expression., (© The Author(s) 2023. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Chemoresistance is a major obstacle in the neoadjuvant chemotherapy (NCT) of locally advanced breast cancer (LABC). Identification of miRNAs as prognostic biomarkers may help overcome chemoresistance of breast cancer (BC). This study aimed to evaluate the expression level of miR-1275 in plasma samples and its biological functions in the chemoresistance of BC. The expression levels of miR-1275 in plasma samples and cells were measured by RT-qPCR. CRISPR/Cas9-mediated gene editing was used to construct miR-1275 knock-out cells in MCF-7. We found that miR-1275 was significantly downregulated in plasma from patients resistant to chemotherapy and in chemoresistant BC cell lines, while patients with low levels of miR-1275 showed poor overall survival. miR-1275 knock-out promoted chemoresistance in BC cells by increasing the properties of cancer stem cells (CSCs). Mechanistically, we identified that MDK was determined to be direct downstream protein of miR-1275 which initiated PI3K/Akt signaling in breast cancer cells. We demonstrated that the high expression level of miR-1275 in plasma predicted better response to NCT. The reduction of miR-1275 promoted BC cells chemoresistance by increasing CSCs properties via targeting MDK/AKT axis. The potential of miR-1275 as a new prognostic biomarker and therapeutic target of BC patients was identified., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Objective: This nonrandomized prospective clinical trial aimed to assess the efficacy, safety and follow-up outcomes of ultrasound-guided high-intensity focused ultrasound (USgHIFU) surgery in patients with breast fibroadenoma., Methods: With the approval of the institutional ethics committee and written informed consent, a total of 113 patients diagnosed with breast fibroadenoma by core-needle biopsy in our hospital were recruited. USgHIFU surgery was performed under local anesthesia. Contrast-enhanced ultrasound (CEUS) or contrast-enhanced MRI (CEMRI) was performed to evaluate the nonperfused volume (NPV). The patients were followed up with physical examination and ultrasound imaging., Results: The clinical outcome of 85 patients with 147 fibroadenomas with a follow-up time of more than 3 months was analyzed in this study. Fifty-two patients had one lesion, twenty-one patients had two lesions and twelve patients had more than two lesions. During USgHIFU, the median localization time for all fibroadenomas was 3 (interquartile range: 1, 5) min, and the median treatment time was 9 (interquartile range: 5, 15) min. Under local anesthesia, all the patients tolerated the treatment well. No serious epidermal burns were observed in any of the patients. Based on CEUS or CEMRI imaging evaluation, the median NPV ratio was 100% (interquartile range: 79.2%, 116.8%). The VRR were 26.77 ± 50.05%, 50.22 ± 42.01% and 72.74 ± 35.39% at 3-6 months, 6-12 months and >12 months, respectively, which showed significant statistical difference ( p  < .001)., Conclusion: Ultrasound-guided HIFU surgery is an effective and safe noninvasive alternative technique for the treatment of breast fibroadenoma.

Breast cancer is a top dangerous killer for women. An accurate early diagnosis of breast cancer is the primary step for treatment. A novel breast cancer detection model called SAFNet is proposed based on ultrasound images and deep learning. We employ a pre-trained ResNet-18 embedded with the spatial attention mechanism as the backbone model. Three randomized network models are trained for prediction in the SAFNet, which are fused by majority voting to produce more accurate results. A public ultrasound image dataset is utilized to evaluate the generalization ability of our SAFNet using 5-fold cross-validation. The simulation experiments reveal that the SAFNet can produce higher classification results compared with four existing breast cancer classification methods. Therefore, our SAFNet is an accurate tool to detect breast cancer that can be applied in clinical diagnosis., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Backgrounds: Breast cancer (BC) is the most frequent cancer diagnosed in women throughout the world. The purpose of this study was to explore new biomarkers for breast cancer diagnosis. CyclinB1 (CCNB1) is found in abundance in a wide range of human malignancies., Material and Methods: We evaluated the transcriptional, survival data and expression levels in tissue data of CCNB1 in patients with breast cancer from the Gene Expression Omnibus (GEO), The Cancer Genome Atlas (TCGA), The Human Protein Atlas (THAP) and Genome Tissue Expression (GTEx) database. A series of in silico analyses were used to investigate at noncoding RNAs (ncRNAs), gene ontology (GO) annotation analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG), and the Protein-Protein Interaction (PPI) network. A quantitative real-time polymerase chain reaction (qRT-PCR) was used to validate CCNB1 in BC cell lines., Results: CCNB1 expression was higher in BC tissues than in normal breast tissues. It was significantly related to survival time, tumor mutation burden (TMB), methylated, immune cell infiltration, and the expressed in estrogen receptor (ER) (-), lymphnode (+), and p53 (+) groups in BC. Moreover, The AC026401.3/CCNB1-miR-139-5p axis was discovered as the most promising upstream ncRNA-related pathway of CCNB1 in BC., Conclusion: CCNB1 can be used as an independent predictive factor for BC, indicating that this would be a target for highly precise therapy and a biomarker for the disease.

Minimally invasive thermal therapies have been attempted in the treatment of breast cancer, and the immune response induced by these therapies has not been fully reported. A clinical trial is performed to determine the effect of microwave ablation (MWA) in the treatment of early-stage breast cancer. The authors perform single-cell RNA sequencing on peripheral blood mononuclear cells (PBMCs) from six patients before and after ablation. NK and CD8 + T cells are activated by MWA of breast cancer, with the increased inhibitory signature of CD8 + T cells but not dysfunctional. Enhanced co-stimulatory signature of CD4 + T cells is observed and increased frequency of ICOS + CD4 + T cells after MWA is confirmed by flow cytometric analysis. After ablation, T-cell clones expand with increased T-cell receptor diversities. Activated antigen receptor-mediated signaling pathways are found in B cells. Enhanced interactions between B cells and CD4 + T cells are found, indicating that B cells are important antigen-presenting cells that initiate CD4 + T cells in MWA-induced immune response. Blockade of CTLA-4 or PD-1 of post-MWA PBMCs show higher T-cell activity than that of pre-MWA PBMCs. This study provide global characteristics of MWA-induced systemic immune response and pave a way for the identification of potential targets to improve the immune response., (© 2022 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Background: Circulating tumor cells (CTCs) have been shown to be associated with the response to neoadjuvant chemotherapy (NCT) and the prognosis of locally advanced breast cancer (LABC) patients. Our study aimed to investigate whether the change of CTC status during NCT could serve as a supplement to the Response Evaluation Criteria in Solid Tumors (RECIST) in the treatment and evaluation of LABC patients., Methods: 6 ml of blood samples were collected before NCT, after the first cycle of NCT and after the completion of NCT, respectively. According to the change of CTC number during NCT, the patients were divided into "CTC low-response (low-R)" group and "CTC high-response (high-R)" group. Survival data of each group of patients were obtained through long-term follow-up., Results: A total of 35 patients diagnosed with LABC were enrolled. The median follow-up for distant metastasis was 27 months (range 7-36 months). There was no significant difference in distant metastasis-free survival (DMFS) between PR/CR group and PD/SD group (P = 0.0914), while CTC low-R group had a worse DMFS than CTC high-R group (P = 0.0199). In PR/CR subgroup, patients with CTC low-R showed a lower DMFS compared with those with CTC high-R (P = 0.0159). However, in PD/SD subgroup, there was no significant difference in DMFS between CTC low-R and CTC high-R group (P = 0.7521). In terms of assessing response to NCT, CTC change or RECIST classification alone had an AUC of 0.533 (95% CI 0.277-0.790) and 0.700 (95% CI 0.611-0.789), respectively. When combining the two, the AUC slightly increased to 0.713 (95% CI 0.532-0.895)., Conclusion: The change of CTC number during NCT has a potential to serve as a supplement to RECIST in the assessment of NCT efficacy and the prognosis of LABC patients., (© 2022. The Author(s) under exclusive licence to Japan Society of Clinical Oncology.)

The visual inspection of histopathological samples is the benchmark for detecting breast cancer, but a strenuous and complicated process takes a long time of the pathologist practice. Deep learning models have shown excellent outcomes in clinical diagnosis and image processing and advances in various fields, including drug development, frequency simulation, and optimization techniques. However, the resemblance of histopathologic images of breast cancer and the inclusion of stable and infected tissues in different areas make detecting and classifying tumors on entire slide images more difficult. In breast cancer, a correct diagnosis is needed for complete care in a limited amount of time. An effective detection can relieve the pathologist's workload and mitigate diagnostic subjectivity. Therefore, this research work investigates improved the pre-trained xception and deeplabv3+ design semantic model. The model has been trained on input images with ground masks on the tuned parameters that significantly improve the segmentation of ultrasound breast images into respective classes, that is, benign/malignant. The segmentation model delivered an accuracy of greater than 99% to prove the model's effectiveness. The segmented images and histopathological breast images are transferred to the 4-qubit-quantum circuit with six-layered architecture to detect breast malignancy. The proposed framework achieved remarkable performance as contrasted to currently published methodologies. HIGHLIGHTS: This research proposed hybrid semantic model using pre-trained xception and deeplabv3 for breast microscopic cancer classification in to benign and malignant classes at accuracy of 95% accuracy, 99% accuracy for detection of breast malignancy., (© 2022 Wiley Periodicals LLC.)

Breast cancer is an aggressive malignancy with high morbidity in females worldwide. Extensive studies reveal that long noncoding RNAs (lncRNAs) are abnormally expressed and act as key regulators in various cancers, including breast cancer. In this work, we investigated the role and regulatory mechanism of lncRNA prostate cancer-associated transcript 6 (PCAT6) in breast cancer progression. Our findings revealed that PCAT6 was overexpressed in breast cancer tissues and cell lines. Furthermore, elevation of PCAT6 reflected an adverse prognosis of patients. Functional experiments indicated that PCAT6 knockdown hampered cell proliferation, facilitated apoptosis and cell cycle arrest in vitro, and inhibited tumor growth in vivo. We also found that the transcription factor SP1 could bind to the PCAT6 promoter and promoted its expression. Subsequently, it was verified that PCAT6 was a molecular sponge for microRNA-326 (miR-326), and the leucine-rich repeat containing the eight family member E (LRRC8E) was a direct target of miR-326. Rescue assays revealed that LRRC8E overexpression attenuated the suppressive effect of PCAT6 knockdown on cellular progression of breast cancer. In summary, this study demonstrated that SP1-activated PCAT6 promoted the malignant behaviors of breast cancer cells by regulating the miR-326/LRRC8E axis., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Surgery is essential for controlling the symptoms and complications of stage IV breast cancer. However, locoregional treatment of primary tumors often results in distant progression, including lung metastasis, the most common type of visceral metastasis. As a minimally invasive thermal therapy, microwave ablation (MWA) has been attempted in the treatment of breast cancer, but the innate immune response after MWA has not yet been reported. Using two murine models of stage IV breast cancer, we found that MWA of primary breast cancer inhibited the progression of lung metastasis and improved survival. NK cells were activated after MWA of the primary tumor and exhibited enhanced cytotoxic functions, and the cytotoxic pathways of NK cells were activated. Depletion experiments showed that NK cells but not CD4+ or CD8+ T cells played a pivotal role in prolonging survival. Then, we found that compared with surgery or control treatment, MWA of the primary tumor induced completely different NK-cell-related cytokine profiles. Macrophages were activated after MWA of the primary tumor and produced IL-15 that activated NK cells to inhibit the progression of metastasis. In addition, MWA of human breast cancer stimulated an autologous NK-cell response. These results demonstrate that MWA of the primary tumor in metastatic breast cancer inhibits metastatic progression via the macrophage/IL-15/NK-cell axis. MWA of the primary tumor may be a promising treatment strategy for de novo stage IV breast cancer, although further substantiation is essential for clinical testing., (© 2020. CSI and USTC.)

Background: Despite great advances in the treatment of breast cancer, innovative approaches are still needed to reduce metastasis. As a minimally invasive local therapy (not standard therapy for breast cancer), microwave ablation (MWA) has been attempted to treat breast cancer, but the local effect and immune response induced by MWA have seldom been reported., Methods: The clinical study was performed to determine the complete ablation rate of MWA for early-stage breast cancer. Secondary endpoints included safety and antitumor immune response. 35 subjects from this clinical study were enrolled in the current report, and the local effect was determined by pathological examinations or follow-up. To investigate MWA-induced immune response, patients treated with surgery (n=13) were enrolled as control, and blood samples were collected before and after MWA or surgery. The immune cell populations, serum cytokines, secretory immune checkpoint molecules, and T-cell receptor sequencing were analyzed., Results: Of 35 enrolled patients, 32 (91.4%) showed complete ablation. Compared with surgery, MWA induced significantly increased levels of inducible co-stimulator (ICOS)+ activated CD4+ T cells and serum interferon gamma, indicating a shift in the Th1/Th2 balance toward Th1. The activated ICOS pathway was involved in the MWA-induced adaptive immune response. T-cell receptor sequencing revealed MWA of primary tumor activated T lymphocytes expansion and recognized some cancer-specific antigens. Moreover, CD4+ effector memory T-cell response was induced by MWA, and the immune response still existed after surgical resection of the ablated tumor., Conclusions: MWA may not only be a promising local therapy but also a trigger of antitumor immunity for breast cancer, opening new avenues for the treatment of breast cancer. Combinatorial strategy using additional agents which boost MWA-induced immune response could be considered as potential treatment for clinical study for early breast cancer therapy., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Importance: There is a lack of studies exploring whether the survival of patients with distant lymph node metastases (DLNM) is different from that of patients with ipsilateral supraclavicular lymph node metastases (ISLM) and other stage IV breast cancer., Objective: To assess the survival of patients with DLNM from breast cancer vs ISLM and other stage IV breast cancer., Design, Setting, and Participants: This cohort study included 2033 patients diagnosed with breast cancer between January 1, 2010, and December 31, 2014, from the Surveillance, Epidemiology and End Results registries database. Three groups of patients were included: (1) patients with ISLM without any distant metastasis, (2) patients with DLNM, and (3) patients with distant metastases (DLNM excluded). Patients younger than 18 years or older than 100 years were excluded. The data were analyzed in February 2020., Exposures: Surgery for primary tumor, surgery for distant lymph nodes, and radiotherapy., Main Outcomes and Measures: Overall survival (OS) and breast cancer-specific survival (BCSS)., Results: Of the 2033 women (mean [SD] age, 62.03 [14.62] years [range, 23.00-99.00 years]; 1510 White participants [74.3%]) with breast cancer included in the study, 346 patients (17.0%) had DLNM, 212 (10.4%) had ISLM, and 1475 (72.6%) had distant metastases (DLNM excluded). The 3-year BCSS rates were 63.24% for ISLM, 64.54% for DLNM, and 41.20% for distant metastases. The 3-year OS rates were 53.46% for ISLM, 62.67% for DLNM, and 38.21% for distant metastases. Compared with patients with ISLM, patients with DLNM showed similar BCSS (hazard ratio [HR], 0.81; 95% CI, 0.52-1.25; P = .34) and OS (HR, 0.73; 95% CI, 0.51-1.05; P = .09), whereas patients with distant metastases showed significantly poorer BCSS (HR, 1.99; 95% CI, 1.43-2.78; P < .001) and OS (HR, 1.79; 95% CI, 1.35-2.38; P < .001). Of the 346 patients with DLNM, primary surgery (HR, 0.21; 95% CI, 0.12-0.39; P < .001) and radiotherapy (HR, 0.46; 95% CI, 0.25-0.87; P = .02) were significantly associated with improved OS., Conclusions and Relevance: The results of this cohort study suggest that DLNM of breast cancer, with similar survival to N3c disease (indicating metastases to the ipsilateral supraclavicular lymph nodes), might be a regional disease, and reassessment of the role of lymph node metastases in breast cancer may be necessary. Given these findings, aggressive locoregional therapies for this disease are recommended, although future studies are still needed to confirm these results.

Background: Core needle biopsy (CNB) plays an important role in the preoperative axillary lymph node (ALN) assessment in breast cancer (BC) patients with the development of treatment, but little is known about the axillary lymph flow after CNB of ALNs. This study aimed to investigate the changes of lymphatic flow after CNB of sentinel lymph node (SLN) in a rabbit model., Methods: The axillary SLN was biopsied in a rabbit model, and the changes of sentinel lymph flow were observed by methylthioninium chloride imaging at 1 and 12 days after the biopsy. Furthermore, the afferent lymphatic vessel was ligated and imaged once every 3 days to assess the changes of lymphatic flow., Results: The SLN biopsied was characterized by disorganized medullary sinus containing erythrocytes, whereas clean medullary sinus containing a normal population of circulating lymphoid cells was observed in the contralateral normal SLN. At 1 day after biopsy, the sentinel lymphatic drainage was blocked. At 12 days after biopsy, the sentinel lymphatic flow was reconstructed or repaired. Ligation of afferent lymphatic vessel further confirmed the reconstruction of lymphatic flow., Conclusions: The sentinel lymphatic flow changes after CNB in a rabbit model, but it can be reconstructed or repaired.

Objective: To evaluate the diagnostic performance of contrast-enhanced ultrasound (CEUS) combined with conventional ultrasound of axillary lymph nodes (ALNs) in predicting metastatic ALNs in patients with breast cancer., Methods: This retrospective study included 259 patients with breast cancer who underwent conventional ultrasound and CEUS. The parameters and patterns evaluated on conventional ultrasound included short axis diameter (S), long axis/short axis (L/S) ratio, cortical thickness, resistive index (RI), lymph node (LN) morphology of greyscale ultrasound, hilum and vascular pattern. Meanwhile, enhancement pattern, wash-in time, time to peak (TP), maximum signal intensity, and duration of contrast enhancement were evaluated on CEUS. Univariate and multiple logistic regression analyses were performed to identify independent factors of ALN status. Three models (conventional ultrasound, CEUS, and combined parameters) were established. Receiver operating characteristic (ROC) curves were applied to evaluate the accuracy of the three predictive models., Results: On conventional axillary ultrasound, LN morphology and vascular pattern were independent factors in predicting metastatic ALNs. On CEUS, maximum signal intensity, duration of contrast enhancement, and TP were independent factors in predicting metastatic ALNs. When combining conventional ultrasound and CEUS features, five independent factors obtained from the conventional ultrasound and CEUS were associated with ALN status. ROC curve analysis showed that the use of CEUS markers combined with conventional ultrasound features (AUC = 0.965) was superior to the use of CEUS markers (AUC = 0.936) and conventional ultrasound features alone (AUC = 0.851)., Conclusion: Combining conventional ultrasound and CEUS features can enable discrimination of ALN status better than the use of CEUS and conventional ultrasound features alone., Advances in Knowledge: The axillary lymph node status in breast cancer patients impacts the treatment decision. Our ultrasonic data demonstrated that CEUS features of ALNs in breast cancer patients could be image markers for predicting ALN status. Combining conventional ultrasound and CEUS features of ALNs can improve specificity discrimination of ALN status better than the use of CEUS and the conventional ultrasound features alone, which will help the treatment planning optimization.

Palbociclib, a CDK4/6 inhibitor, has been granted accelerated approval by US FDA for hormone receptor-positive HER2-negative metastatic breast cancer. To determine potential biomarkers of palbociclib sensitivity to assist in patient selection and clinical development, we investigated the effects of palbociclib in a panel of molecularly characterized breast cancer cell lines. We quantified palbociclib sensitivity and c-myc expression in 11 breast cancer cell lines, 124 breast cancer samples, and The Cancer Genome Atlas database. We found non-TNBC subtypes were more sensitive to palbociclib than TNBC. Activation of c-myc led to differential palbociclib sensitivities, and further inhibition of c-myc enhanced palbociclib sensitivity. Moreover, we identified for the first time a c-myc/miR-29b-3p/CDK6 axis in breast cancer that could be responsible for c-myc-induced palbociclib insensitivity, in which c-myc activation resulted in downregulation of miR-29b-3p, further activated CDK6 and inhibited cell-cycle arrest at G 1 phase. Moreover, downregulated (inactived) c-myc-induced oncogenic addiction could increase palbociclib efficacy, using both Xenograft model and patient-derived tumor xenograft (PDTX) model. Our finding extends the concept of combined blockade of the CDK4/6 and c-myc signaling pathways to increase palbociclib sensitivity, making c-myc a promising biomarker for palbociclib sensitivity in breast cancer.

Breast cancer (BC) is the most common malignancy among women worldwide. However, identifying effective biomarkers for the diagnosis and treatment of BC is challenging. Based on our previously developed 'humanized' mouse model of BC, microarray expression analysis was performed and multiple differentially expressed genes, including ribosomal protein (RP) L32, were screened. Recent reports have revealed that RPs are relevant to the development and progression of cancer. However, the expression and function of RPL32 in BC remains unknown. Therefore, in the present study, the role of RPL32 in the development of BC was explored. Immunohistochemical staining and reverse transcription‑quantitative PCR were used, and it was found that RPL32 was upregulated in human BC tissues and cells. Cell Counting Kit‑8, cell invasion and migration assays were performed, which demonstrated that RPL32 knockdown using lentivirus‑delivered small interfering RNA inhibited the migration and invasion of BC cells in vitro and in vivo (nude mouse model). Moreover, western blotting showed that RPL32 knockdown decreased the expression levels of matrix metalloproteinase (MMP)‑2 and MMP‑9. Thus, the present findings indicated a potential oncogenic role of RPL32, suggesting that it may be a novel target for molecular targeted therapy in patients with BC.

Tamoxifen resistance is a major roadblock in the treatment of patients with breast cancer. Ribonucleotide reductase M2 (RRM2) was found to be involved in acquired resistance of breast cancer cells (BCCs) to tamoxifen. Here, we used GW8510, which has been identified as a potential RRM2 inhibitor, to evaluate the effect of RRM2 inhibition on reversing resistance of BCCs to tamoxifen and investigate its mechanisms. We showed that RRM2 overexpression played a key role in the development of acquired tamoxifen resistance in BCCs through downregulation of autophagy level. Combination treatment with tamoxifen and GW8510 significantly inhibited survival of the tamoxifen-resistant BCCs through induction of autophagic cell death compared to either of the two drugs. Furthermore, combination of tamoxifen and GW8510 resulted in marked growth inhibition of tamoxifen-resistant BBC xenograft tumor in vivo compared to tamoxifen or GW8510 alone. In conclusion, tamoxifen in combination with GW8510 can overcome acquired tamoxifen resistance in BCCs and may be a rational therapeutic approach against breast cancer with high RRM2 expression., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

This study aimed to retrospectively evaluate the circulating free DNA (cfDNA) level in patients with locally advanced breast cancer (LABC) having different neoadjuvant chemotherapy (NCT) responses and to investigate whether dynamic changes in cfDNA level could predict the effectiveness of NCT in patients with LABC. Data on 61 patients with LABC were included. NCT responses were evaluated using the response evaluation criteria. Blood samples were collected for cfDNA detection before treatment and after the first and eighth courses of chemotherapy. The Alu 111-bp and 260-bp fragment levels were evaluated by polymerase chain reaction, and the predictive value of the cfDNA level in the NCT response was determined. In vitro, the MCF-7 and MCF-7/ADR cell lines were applied to simulate the phenomenon of drug resistance and explain the underlying mechanism. The Alu 111-bp level increased after the first NCT course (P = .014) and then remained high after NCT in the high-R group (P = .047), but it remained steady in the low-R group during NCT. A similar tendency in the Alu 260-bp level was revealed in different groups. The ∆∆Ct value of Alu 260-bp had good diagnostic efficiency in assessing predictive ability. The area under the curve for the ∆∆Ct1 and ∆∆Ct2 of Alu 260-bp was 0.697 and 0.647, respectively. The cfDNA level was closely related to epirubicin-induced apoptosis and changes in the Ki-67 index in vitro. The elevation of cfDNA after one chemotherapy cycle was mediated by the apoptosis of tumor cells and related to the improved chemotherapy response., (© 2020 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Drug resistance, including epirubicin-based therapeutic resistance, is one of the major reasons responsible for the unfavorable prognosis of patients diagnosed with breast cancer (BC). Acquired chemoresistance and metastatic properties have been identified to be closely associated with the process of epithelial-mesenchymal transition (EMT). Recently, dysregulation of long non-coding RNAs (lncRNAs) have been increasingly reported to perform promotive or suppressive functions in chemoresistance and EMT process in multiple cancers. However, relevant novel lncRNA participating in epirubicin resistance and EMT and its underlying molecular mechanisms remain unknown in BC. Herein, we established the epirubicin-resistant breast cancer cell subline (MCF-7/ADR), which presented mesenchymal phenotype and increased metastatic potential. A panel of differentially expressed lncRNAs, including 268 upregulated and 49 downregulated lncRNAs, were identified by high-flux microarray investigation in MCF-7 and MCF-7/ADR cells. The novel lncRNA NONHSAT101069 was significantly overexpressed in BC specimens, BC cell lines, and epirubicin-resistant cell sublines. The knockdown of NONHSAT101069 significantly repressed, whereas overexpression of NONHSAT101069 promoted the epirubicin resistance, migration, invasion and EMT process of BC cells both in vitro and in vivo. Further mechanism-related researches uncovered that NONHSAT101069 functioned as a ceRNA (competing endogenous RNA) via sponging miR-129-5p. Twist1 was a direct downstream protein of NONHSAT101069/miR-129-5p axis in BC cells. To conclude, NONHSAT101069 was upregulated in BC tissues and promoted epirubicin resistance, migration and invasion of BC cells via regulation of NONHSAT101069/miR-129-5p/Twist1 axis, highlighting its potential as an oncogene and a therapeutic biomarker for BC.

Breast cancer (BC) is one of the most common cancers in females. Since early detection can bring prognosis benefit, discovery of novel noninvasive biomarkers for BC diagnosis is in urgent need. In this four-phase study, we profiled miRNA expression in plasma samples from a total of 257 BC patients and 257 normal controls (NCs). Exiqon miRNA qPCR panel was used to select candidate miRNAs in the screening phase which were further analyzed using qRT-PCR in the following training, testing and external validation phases. Finally, we identified five plasma miRNAs (let-7b-5p, miR-122-5p, miR-146b-5p, miR-210-3p and miR-215-5p) whose expression levels were significantly different between BC patients and NCs. A 5-miRNA panel in plasma with high sensitivity and specificity was then constructed to detect BC. The areas under the receiver-operating characteristic curves (AUCs) of the panel were 0.683, 0.966, 0.978 for the training, testing and external validation sets, respectively. Expression of the identified miRNAs was further analyzed among 32 pairs of BC tissue and the adjacent normal tissue samples as well as plasma-derived exosome samples from 32 BC patients vs 32 NCs. Let-7b-5p was contrarily down-regulated in BC tissue. In exosomes samples, only miR-122-5p was significantly up-regulated as in plasma for BC patients. In conclusion, we identified a 5-miRNA plasma panel (let-7b-5p, miR-122-5p, miR-146b-5p, miR-210-3p and miR-215-5p) that could serve as a promising biomarker for BC detection., (© 2019 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Most N6-methyladenosine (m 6 A) associated regulatory proteins (i.e., m 6 A writer, eraser, and reader proteins) are involved in the pathogenesis of various cancers, mostly in m 6 A-dependent manners. As a component in the m 6 A 'writers', KIAA1429 is reported to be an RNA-binding protein and involved in the m 6 A modification, mRNA splicing and processing. Till now, the functions of KIAA1429 in tumorigenesis and related mechanism have not been reported. In the present study, we found KIAA1429 was highly expressed in breast cancer tissues, but frequently down-regulated in non-cancerous breast tissues. The overall survival of breast cancer patients with high-expression KIAA1429 was significantly shorter than those with low-expression KIAA1429. Then, we demonstrated that KIAA1429 was associated with breast cancer proliferation and metastasis in vivo and in vitro. The potential targeting genes of KIAA1429 in breast cancer were identified by RNA immunoprecipitation sequencing. One of these genes is cyclin-dependent kinase 1 (CDK1), which plays an oncogenic role in cancers. Furthermore, we confirmed that KIAA1429 played its oncogenic role in breast cancer by regulating CDK1 by an m 6 A-independent manner. 5'-fluorouracil was found to be very effective in reducing the expression of KIAA1429 and CDK1 in breast cancer. These findings indicated that KIAA1429 could promote breast cancer progression and was correlated with pathogenesis. It may represent a promising therapeutic strategy on breast cancer, especially in combination with CDK1 treatment.

Aims: To determine the factors influencing ultrasound breast tumor size assessment accuracy., Material and Methods:  Five factors (tumor type, molecular subtype, histological size, histological grade, and breast density) were used to assess the measurement accuracy of breast ultrasound in tumor size. Size underestimation was defined as ultrasound index lesion diameter < histological size by at least 5 mm., Results: Breast ultrasound underestimated tumor size significantly, especially in cases with intraductal components (p=0.002). There was a tendency for higher size underestimation in breast cancer tumors with high-histological grade (p=0.03), human epidermal growth factor receptor type 2 (HER2)-overexpressing breast cancer tumors (p=0.02) and hormone receptor (HR)-/HER2+ breast cancer tumors (p=0.008). Furthermore, core biopsy revealedhigher probability of size underestimation with intraductal components (p=0.002). Size underestimation was more frequent with larger histological size (p<0.001). Masses in non-dense breasts were significantly underestimated (p=0.036) compared to dense breasts., Conclusions: The size underestimation was influenced by pathological type, molecular subtype, and histological size. The pathological results of core biopsy were conducive for predicting tumor size pre-surgery in precise breast cancer diagnosis.

Activation of oncogenes by promoter hypomethylation plays an important role in tumorigenesis. Zinc finger protein 57 (ZFP57), a member of KRAB-ZFPs, could maintain DNA methylation in embryonic stem cells (ESCs), although its role and underlying mechanisms in breast cancer are not well understood. In this study, we found that ZFP57 had low expression in breast cancer, and overexpression of ZFP57 could inhibit the proliferation of breast cancer cells by inhibiting the Wnt/β-catenin pathway. MEST was validated as the direct target gene of ZFP57 and MEST may be down-regulated by ZFP57 through conserving DNA methylation. Furthermore, overexpression of MEST could restore the tumour-suppressed and the Wnt/β-catenin pathway inactivated effects of ZFP57. ZFP57-MEST and the Wnt/β-catenin pathway axis are involved in breast tumorigenesis, which may represent a potential diagnostic biomarker, and provide a new insight into a novel therapeutic strategy for breast cancer patients.

Background: Many elderly breast cancer patients might just receive palliative local surgery, especially those with locally advanced breast cancer (LABC). However, palliative tumor removal might lead to perioperative residual tumor growth. In this study, we aimed to determine the survival effect of palliative local surgery without definitive axillary surgery for LABC in elderly patients., Patients and Methods: Patients age 70 years or older diagnosed with T3/4M0 breast cancer, who received no surgery, mastectomy, or lumpectomy without axillary surgery, were identified in the Surveillance, Epidemiology, and End Results cancer database from 1973 to 2014. The overall survival effect of palliative local surgery was determined by using multivariable Cox regression, and propensity score matching was applied to confirm the results., Results: A total of 2616 female breast cancer patients age 70 years or older diagnosed with T3/4M0 (without inflammatory breast cancer) were identified; 1374 received no cancer-directed surgery, 583 received lumpectomy without axillary surgery, and 659 received mastectomy without axillary surgery. Adjusted for potential confounders, both types of palliative local surgeries (lumpectomy: hazard ratio [HR], 0.95; 95% confidence interval [CI], 0.71-1.27; P = .719; mastectomy: HR, 0.88; 95% CI, 0.65-1.17; P = .371) were not associated with overall survival compared with no surgery within hormone receptor-positive patients. However, mastectomy strongly improved survival within hormone receptor-negative patients. Palliative local surgery did not change the patterns of mortality., Conclusion: For elderly patients diagnosed with LABC, not candidates for standard therapies, mastectomy should be recommended as palliative therapy for hormone receptor-negative, but not for hormone receptor-positive patients., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Introduction: Adjuvant docetaxel-based chemotherapy is frequently used in the treatment of operable early breast cancer (EBC). This study investigated patterns of docetaxel use in patients with EBC in real-world clinical practice in China., Methods: This was a multicenter, prospective, observational study of Chinese women with operable breast cancer in tier 1 non-oncology-specific hospitals and other city hospitals in China. Adult Chinese female patients (≥ 18 years) with newly diagnosed breast cancer who underwent surgery and received docetaxel-containing adjuvant chemotherapy were eligible for inclusion. The primary end point of the study was to assess patterns of docetaxel use for adjuvant therapy in Chinese clinical practice. Secondary end points included factors associated with treatment selection, patterns of G-CSF use and safety., Results: In total, 502 patients were enrolled, 475 of whom were included in the analysis. The results showed that 31.6% of patients received docetaxel and anthracyclines in combination, 47.2% received docetaxel and anthracyclines sequentially, 18.5% received docetaxel and other medication either in combination or sequentially and 2.7% received 'other' chemotherapy regimens. Factors influencing selection of docetaxel/anthracycline in combination versus sequentially included age, breast cancer subgroup, PR status, and preoperative neoadjuvant chemotherapy. A total of 258 (54.3%), 35 (7.4%), and 108 (22.7%) patients received G-CSF as primary prevention, secondary prevention and treatment, respectively. Treatment-emergent adverse events (TEAEs) occurred in 54.5% (259) of patients, and the most frequently reported TEAEs (≥ 5% of patients) included bone marrow failure (10.1%), granulocytopenia (9.9%), nausea (8.8%) and vomiting (6.9%). Neutropenia and febrile neutropenia were reported by 10 and 12 patients, respectively., Conclusion: Docetaxel for adjuvant chemotherapy of operable EBC in China was most commonly given in combination or sequentially with anthracyclines. The study also showed that in China G-CSF is most frequently used as primary prophylactic, and no unexpected safety events were observed during docetaxel treatment., Funding: Sanofi (China).

Purpose: To determine the characteristics of ultrasound (US) imaging of completely ablated cases and the effects of duration and clinical experience on accurate microwave ablation (MWA) for the treatment of benign breast tumours., Methods: With written informed consent and approval of the institutional ethics committee, patients with symptomatic or palpable benign breast tumours (longest diameter, 7-32 mm), to whom MWA (2450 MHz) was performed, were enrolled in this prospective nonrandomised study. US and contrast-enhanced US (CEUS) images were applied for follow-up and analysed., Results: Forty-seven consecutive patients with 52 completely ablated tumours were enrolled. Of these 52 tumour ablations in US, 16 ablations were defined as concentric type, and 36 were defined as nonconcentric type. Of these 52 ablations, 7 cases were defined as nonaccurate ablation with the largest margin ≥10 mm in US. The nonaccurate ablation rate in the training group (the first consecutive 30 cases, 7/30) was significant higher than that (the last 22 cases, 0/22) in the practiced group (p = 0.016). Of 38 completely ablated cases (9 mm < the longest diameter <20 mm), the average largest margin in >70 s group was significant larger than that in <70 s group (p = 0.019)., Conclusions: Experience was important for accurate MWA in the treatment of benign breast tumour, and at least 30 cases training was recommended. Nevertheless, clinical trials are still required to validate our findings in the future.

Background: The wide adoption of electronic health record systems (EHRs) in hospitals in China has made large amounts of data available for clinical research including breast cancer. Unfortunately, much of detailed clinical information is embedded in clinical narratives e.g., breast radiology reports. The American College of Radiology (ACR) has developed a Breast Imaging Reporting and Data System (BI-RADS) to standardize the clinical findings from breast radiology reports., Objectives: This study aims to develop natural language processing (NLP) methods to extract BI-RADS findings from breast ultrasound reports in Chinese, thus to support clinical operation and breast cancer research in China., Methods: We developed and compared three different types of NLP approaches, including a rule-based method, a traditional machine learning-based method using the Conditional Random Fields (CRF) algorithm, and deep learning-based approaches, to extract all BI-RADS finding categories from breast ultrasound reports in Chinese., Results: Using a manually annotated dataset containing 540 reports, our evaluation shows that the deep learning-based method achieved the best F1-score of 0.904, when compared with rule-based and CRF-based approaches (0.848 and 0.881 respectively)., Conclusions: This is the first study that applies deep learning technologies to BI-RADS findings extraction in Chinese breast ultrasound reports, demonstrating its potential on enabling international collaborations on breast cancer research., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Purpose: Novel noninvasive biomarkers with high sensitivity and specificity for the diagnosis of breast cancer (BC) are urgently needed in clinics. The aim of this study was to explore whether miRNAs from the miR-106a-363 cluster can be detected in the circulation of BC patients and whether these miRNAs can serve as potential diagnostic biomarkers., Methods: The expression of 12 miRNAs from the miR-106a-363 cluster was evaluated using qRT-PCR in 400 plasma samples (from 200 BC patients and 200 healthy controls (HCs)) and 406 serum samples (from 204 BC patients and 202 HCs) via a three-phase study. The identified miRNAs were further examined in tissues (32 paired breast tissues), plasma exosomes (from 32 BC patients and 32 HCs), and serum exosomes (from 32 BC patients and 32 HCs)., Results: Upregulated levels of four plasma miRNAs (miR-106a-3p, miR-106a-5p, miR-20b-5p, and miR-92a-2-5p) and four serum miRNAs (miR-106a-5p, miR-19b-3p, miR-20b-5p, and miR-92a-3p) were identified and validated in BC. A plasma 4-miRNA panel and a serum 4-miRNA panel were constructed to discriminate BC patients from HCs. The areas under the receiver-operating characteristic curves of the plasma panel were 0.880, 0.902, and 0.858, and those of the serum panel were 0.910, 0.974, and 0.949 for the training, testing, and external validation phases, respectively. Two overlapping miRNAs (miR-106a-5p and miR-20b-5p) were consistently upregulated in BC tissues. Except for the expression of the plasma-derived exosomal miR-20b-5p, the expression patterns of exosomal miRNAs were concordant between plasma and serum, indicating the potential use of exosomal miRNAs as biomarkers., Conclusion: We identified four plasma miRNAs and four serum miRNAs from the miR-106a-363 cluster as promising novel biomarkers for the diagnosis of BC.

Breast cancer is the most commonly diagnosed cancer and the leading cause of cancer death in females worldwide. Chemoresistance has been a major reason for the drug therapy failure. The present study performed a microarray analysis between MCF‑7 and MCF‑7/adriamycin (ADR) cells, and intended to identify long non‑coding (lnc)RNA expression character in drug resistant breast cancer cells. MCF‑7/ADR cells were induced from MCF‑7 cells via pulse‑selection with doxorubicin for 4 weeks, and the resistance to doxorubicin of ADR cells was confirmed by MTT assay. Microarray analysis was performed between MCF‑7 and MCF‑7/ADR cells. Total RNA was extracted from the two cell lines respectively and was transcribed into cDNA. The results of the microarray were verified by reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR). Gene Ontology (GO) and pathways analysis were conducted to enrich the dysregulated lncRNAs presented in the microarray results. Compared to the MCF‑7 cells, 8,892 lncRNAs were differentially expressed in MCF/ADR cells (absolute fold‑change >2.0). A total of 32 lncRNAs were selected for RT‑qPCR by fold‑change filtering, standard Student's t‑test, and multiple hypothesis testing. Among the dysregulated lncRNAs, AX747207 was prominent because its associated gene RUNX3 was previously reported to be relative to malignant tumor chemoresistance. GO analysis results also indicated some biological processes and molecular functions linked to chemoresistance. The pathway enrichment results provided some potential pathways associated with chemoresistance. In the present study, the authors intended to identify lncRNA expression character in drug resistant cell line MCF‑7/ADR, corresponding to the parental MCF‑7 cell line. In addition, the study identified the lncRNA AX747207, and its potential targeted gene RUNX3, may be related to chemoresistance in breast cancer. These results may new insights into exploring the mechanisms of chemoresistance in breast cancer.

Breast cancer is the second leading cause of cancer‑associated mortality, and metastatic breast cancer is responsible for 90% of patient mortalities. Given that JWA represses the proliferation, invasion and metastasis of a number of other human tumor cells, including melanoma, esophageal, hepatocellular and gastric carcinomas, via mitogen‑activated protein kinase or integrin signaling, the present study investigated the expression and function of JWA in human breast cancers. The results showed that the expression level of JWA was significantly reduced in human primary breast cancers when compared with the paired adjacent tissues. Downregulating JWA enhanced, while overexpressing JWA suppressed, the migration and invasion abilities of the two breast cancer cell lines, MDA‑MB‑468 and MDA‑MB‑231, without affecting their proliferations in vitro. In addition, JWA negatively regulated the surface expression of CXCR4 in the two cell lines via proteasome degradation, though not via transcriptional inhibition. Functionally, normalizing the disturbed expressions of CXCR4 largely reversed the inhibitory effect of JWA on cell invasion. These data demonstrated that JWA suppressed the migration/invasion of breast carcinoma cells by downregulating the expression of CXCR4, and suggested that JWA may harbor prognostic and therapeutic potential in patients with breast cancer.

Objective: This study aims to investigate the correlations between rapidly accelerated fibrosarcoma/mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase signaling pathway and clinicopathological features and prognosis for patients with breast cancer having axillary lymph node metastasis., Methods: A total of 118 breast cancer tissues with axillary lymph node metastasis (axillary lymph node metastasis group), 150 breast cancer tissues with non-axillary lymph node metastasis (non-axillary lymph node metastasis group), and 216 normal breast tissues (normal group) were enrolled in this study. The messenger RNA and protein expressions of rapidly accelerated fibrosarcoma, MEK, extracellular signal-regulated kinase, and their phosphorylated (p-) proteins were examined by reverse transcriptase quantitative polymerase chain reaction and immunohistochemistry, respectively. All patients received a 1-year follow-up, and the clinical follow-up data were collected. The multiple factors on the prognosis of patients with breast cancer having axillary lymph node metastasis were tested by Cox regression analysis., Results: The messenger RNA expressions of rapidly accelerated fibrosarcoma, MEK, and extracellular signal-regulated kinase and positive rates of rapidly accelerated fibrosarcoma, MEK, phosphorylated MEK, extracellular signal-regulated kinase, and p-extracellular signal-regulated kinase in the axillary lymph node metastasis group were higher than in the non-axillary lymph node metastasis and normal groups (all P < .05). The protein expressions of rapidly accelerated fibrosarcoma, MEK, phosphorylated MEK, extracellular signal-regulated kinase, and p-extracellular signal-regulated kinase were associated with tumor size, clinical stage, and axillary lymph node metastasis number (all P < .05). Rapidly accelerated fibrosarcoma, MEK, and extracellular signal-regulated kinase expressions were significantly correlated with the prognosis of patients with breast cancer (all P < .05). Patients with BC having positive rapidly accelerated fibrosarcoma, MEK, phosphorylated MEK, extracellular signal-regulated kinase, and phosphorylated ERK expressions had a higher survival rate than patients with BC having the negative ones (all P < .05). Rapidly accelerated fibrosarcoma and extracellular signal-regulated kinase protein expressions, clinical stage, pathological grade, and axillary lymph node metastasis number were independent prognostic factors in patients with breast cancer having axillary lymph node metastasis (all P < .05)., Conclusion: Our study proved that rapidly accelerated fibrosarcoma/MEK/extracellular signal-regulated kinase signaling pathway is significantly correlated with the clinicopathological features and prognosis for patients with BC having axillary lymph node metastasis. Rapidly accelerated fibrosarcoma and extracellular signal-regulated kinase protein expressions are independent prognostic factors for patients with breast cancer having axillary lymph node metastasis.

Neoadjuvant chemotherapy (NCT) is the standard treatment for locally advanced breast cancer (LABC). Pathological complete response (pCR) is commonly used as a valid predictor of NCT long-term outcomes. Blood-based tumor biomarkers have the potential to predict response to NCT at early stage non-invasively. We believed plasma CCL5 could be a potential marker to predict NCT of LABC. Its efficiency and possible mechanism was studied in this work. Human Cytokine Antibody Microarray was applied to screen different cytokine concentration in plasma between low histological regression (Low-R) and high histological regression (High-R) patients. LABC patients were divided into two groups according to pathological reactivity. The concentration of plasma CCL5 in different groups was determined by ELISA analysis. CCK8 assay was performed to analyze epirubicin susceptibility of breast cancer cells. Transwell assay was performed to determine the effect of CCL5 on breast cancer cells' migration and invasion. qRT-PCR and western blot were used to verify the EMT (epithelial-mesenchymal transition) markers in CCL5-treated and epirubicin-treated breast cancer cells. The concentration of plasma CCL5 of Low-R group was higher than High-R group before NCT. The plasma levels of CCL5 were significantly reduced after NCT in the group of high histological regression (High-R). Epirubicin susceptibility decreased in the breast cancer cells treated by recombinant CCL5. Migration and invasion were significantly enhanced in breast cancer cells treated by recombinant CCL5. E-cadherin expression was decreased whereas vimentin increased significantly in CCL5-treated breast cancer cells. The phosphorylation of ezrin in Y-567 and its downstream protein cortactin increased significantly in CCL5-treated breast cancer cells. Plasma CCL5 level could be a promised candidate to predict chemotherapy response of breast cancer. Plasma CCL5 plays an important role in EMT process of breast cancer.

In breast cancer, cell-free DNA (cfDNA) has been proven to be a diagnostic and prognostic biomarker. However, there have been few studies on the origin and biological significance of cfDNA. In this study, we assessed the release pattern of cfDNA from breast cancer cell lines under different culture conditions and investigated the biological significance of cfDNA. The cfDNA concentration increased rapidly (6 h) after passage, decreased gradually, and was then maintained at a relatively stable level after 24 h. In addition, the cfDNA concentration did not correlate with the amount of apoptotic and necrotic cells. Interestingly, if more cells were in the G1 phase, more cfDNA was detected (p < 0.01) and the cfDNA concentration correlated positively with the percent of cells in the G1 phase (p < 0.05). We observed that cells could release cfDNA actively, but not exclusively, via exosomes. Furthermore, we showed that cfDNA could stimulate hormone receptor-positive breast cancer cell proliferation by activating the TLR9-NF-κB-cyclin D1 pathway. In conclusion, cfDNA is released from breast cancer mainly by active secretion, and cfDNA could stimulate proliferation of breast cancer cells.

BACKGROUND Epithelial to mesenchymal transition (EMT) contributes to metastases in various types of tumors, and is also the key step in the breast cancer metastatic cascade. In our previous study, a mouse model containing human-derived normal breast tissue was established and allowed EMT/MET process of human breast cancer cells to be mimicked in a humanized mammary microenvironment. MATERIAL AND METHODS Two-dimensional electrophoresis (2-DE) and mass spectrometry were used to detect different proteins between parental MDA-MB-231 and its variant sub-line obtained from tumors grown in transplanted normal human breast tissue (MDA-MB-231br). We knocked down the ezrin in 2 cell lines (MDA-MB-231 and SUM1315). The migration and invasion ability was assessed. EMT markers were examined by real-time reverse transcription PCR analysis and Western blot analysis. The relationship of ezrin with cortactin was tested by tissue microarray and co-immunoprecipitation. RESULTS Proteomic analysis revealed 81 differentially expressed proteins between parental MDA-MB-231 and MDA-MB-231br. Among these proteins, the expression of ezrin and cortactin and the phosphorylation of ezrin were significantly correlated, accompanied with a group of classic EMT makers. Knockdown of ezrin reversed the expression of EMT markers and downregulated cortactin and EMT transcription factors. Ezrin silencing inhibited tumor cell migration and invasion. Breast cancer tissue microarray and immunohistochemistry showed a significant positive association between ezrin and cortactin. CONCLUSIONS These findings indicate that ezrin is correlated with cortactin in facilitating EMT in breast cancer. The interaction between ezrin and cortactin is a novel mechanism contributing to the EMT process in cancer metastases.

Vasohibin 2 (VASH2) is identified as an angiogenic factor, and has been implicated in tumor angiogenesis, proliferation and epithelial-mesenchymal transition (EMT). To investigate the EMT role of VASH2 in breast cancer, we overexpressed or knocked down expression of VASH2 in human breast cancer cell lines. We observed that VASH2 induced EMT in vitro and in vivo. The transforming growth factor β1 (TGFβ1) pathway was activated by VASH2, and expression of a dominant negative TGFβ type II receptor could block VASH2-mediated EMT. In clinical breast cancer tissues VASH2 positively correlated with TGFβ1 expression, but negatively correlated with E-cadherin (a marker of EMT) expression. Under hypoxic conditions in vitro or in vivo, we found that down-regulation of estrogen receptor 1 (ESR1) in VASH2 overexpressing ESR1 positive cells suppressed E-cadherin. Correlation coefficient analysis indicated that VASH2 and ESR1 expression were negatively correlated in clinical human breast cancer tissues. Further study revealed that a transcription factor of ESR1, GATA-binding factor 3 (GATA3), was down-regulated by VASH2 under hypoxia or in vivo. These findings suggest that VASH2 drives breast cancer cells to undergo EMT by activation of the TGFβ1 pathway and hypoxia dependent repression GATA3-ESR1 pathway, leading to cancer metastasis., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Vasohibin 2 (VASH2) is an angiogenic factor and cancer-related protein that acts via paracrine mechanisms. Here, we investigated the angiogenic function and mechanism of action of VASH2 in 200 human breast cancer tissues by performing immunohistochemical staining, western blot, indirect sandwich enzyme-linked immunosorbent assay (ELISA), and a semi-quantitative sandwich-based antibody array. Breast cancer cells stably overexpressing VASH2 or with knocked-down VASH2 were established and used for in vivo and in vitro models. In human luminal tissue, but not in HER2-positive or basal-like breast cancer tissues, VASH2 was positively correlated with CD31-positive microvascular density, induced angiogenesis in xenograft tumors, and promoted human umbilical vein endothelial cell tube formation in vitro. VASH2 expression was absent in the concentrated conditioned medium collected from knocked-down VASH2 and VASH2-overexpressing luminal breast cancer cells. Further, VASH2 regulated the expression of fibroblast growth factor 2 (FGF2) in human luminal breast cancer cells, and the pro-angiogenic effect induced by VASH2 overexpression was blocked by FGF2 neutralization in vitro. Additionally, dual luciferase reporter assay and Chromatin immunoprecipitation analysis results showed that FGF2 promoter was transcriptionally activated by VASH2 via histone modifications. In conclusion, VASH2 expression is positively correlated with FGF2 expression and promotes angiogenesis in human luminal breast cancer by transcriptional activation of fibroblast growth factor 2 through non-paracrine mechanisms., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Background: Sentinel lymph node biopsy (SLNB) is well accepted to be a standard procedure in breast cancer surgery with clinically negative lymph nodes. Isosulfan blue is the first dye approved by the USA Food and Drug Administration for the localization of the lymphatic system. Few alternative tracers have been investigated. In this study, we aimed to compare the differences between 10 % fluorescein sodium and 1 % isosulfan blue in breast sentinel lymph node biopsy and to investigate the feasibility of using 10 % fluorescein sodium as a new dye for breast sentinel lymph node biopsy., Methods: A total of 30 New Zealand rabbits were randomly divided into the fluorescein sodium group and the isosulfan blue group (15 rabbits per group). Fluorescein sodium or isosulfan blue was injected subcutaneously into the second pair of mammary areolas., Results: The average fading time of the second lymph nodes in the isosulfan blue group was significantly shorter than that in the fluorescein sodium group. Moreover, the detection rates of SLNs were higher in the fluorescein sodium group than in the isosulfan blue group. No significant differences between the fluorescein sodium group and isosulfan blue group were observed regarding the distances between the detected sentinel lymph nodes and second pair of mammary areolas, the distances between the second lymph nodes and second pair of mammary areolas, the number of detected sentinel lymph nodes and second lymph nodes, the average dyeing time of the sentinel and the second lymph nodes, and the average fading time of the second lymph nodes., Conclusions: In summary, we first reported that fluorescein sodium is a potential new tracer for breast sentinel lymph node biopsy.

We investigated whether great tumour burden in the primary draining lymph node would lead to obstructed lymphatic flow in breast cancer patients. Breast cancer patients with false-negative sentinel lymph nodes (SLNs) were enrolled from January 2001 to March 2011, retrospectively. A further 45 breast cancer patients were recruited prospectively from December 2013 to November 2014. Carbon nanoparticles, a lymphatic tracer, were injected into the subareolar area 24 h before surgery, followed by axillary lymph node dissection. In the SLN cohort, among the 28 false-negative cases, >50 % showed great tumour burden in the axilla. In the carbon nanoparticles cohort, we found that cases with <3 nodes involved in the pathology had more lymph nodes stained by carbon nanoparticles than the subgroup with ≥3 involved nodes (P = 0.003). Nodes stained with carbon nanoparticles showed smaller tumour burdens compared with unstained nodes (P < 0.05). Furthermore, five cases showed metastatic nodes that were not stained with carbon nanoparticles, and all the lymph nodes that were free of metastasis were stained with carbon nanoparticles. Great tumour burden in the axilla might lead to lymphatic flow obstructions in clinical practice. Nevertheless, clinical trials are still needed to validate our findings.

In clinical practice, it is necessary to define an optimal choice from many different therapeutic regimens. This study aimed to assess the efficacy and safety of neoadjuvant endocrine therapy (NET) for breast cancer patients. Randomized clinical trials were included. Nine studies comprising 2133 patients were included in the final analysis. Network meta-analysis showed that everolimus plus letrozole was more easily accepted by patients than exemestane (≥20wks) (odds ratio (OR): 856697.02, 95% confidence intervals (95%CI): 1.88 to 87242934...); exemestane (≥20wks) had worse acceptability than letrozole (OR: 0.00, 95%CI: 0.00 to 0.98). Letrozole produced a better clinical objective response (COR) than tamoxifen (OR: 1.99, 95%CI: 1.04 to 3.80). The incidence of fatigue between the anastrozole plus gefitinib group and the everolimus plus letrozole group was significantly different (OR: 0.08, 95%CI: 0.01 to 0.83). The exemestane (<20wks) plus celecoxib group had fewer hot flushes than others. Ranking showed the everolimus plus letrozole was most likely rank first in comparisons of COR and acceptability, and had a 64% possibility to rank first after stochastic multi-criteria acceptability analysis. In conclusion, our study showed that letrozole plus everolimus is the most effective treatment for postmenopausal, hormone receptor-positive breast cancer in the neoadjuvant setting.

Given the greatly elevated risks of contralateral breast cancer (CBC) observed in breast cancer patients who carry mutation in BRCA1 or BRCA2, the present meta-analysis was addressed to determine whether adjuvant tamoxifen treatment for breast cancer is associated with reduced CBC risk for these patients. We searched the database of PubMed for eligible studies about the effectiveness of tamoxifen on CBC risk among BRCA1/2 mutation carriers, and we calculated the pooled relative risks (RRs) for CBC risk along with 95 % confidence intervals (CIs). Fixed- or random-effects model was conducted according to study heterogeneity. Four non-overlapping studies met the inclusion criteria for the meta-analysis. Tamoxifen was found to be significantly associated with reduced risk of CBC among BRCA1/2 mutation carriers (summary RR = 0.56, 95 % CI = 0.41-0.76). Similar findings were observed in BRCA1 mutation carriers (summary RR = 0.47, 95 % CI = 0.37-0.60) and BRCA2 mutation carriers (summary RR = 0.39, 95 % CI = 0.28-0.54), respectively. In conclusion, tamoxifen significantly reduces the incidence of CBC among BRCA1/2 mutation carriers with primary unilateral breast cancer. It can reasonably be offered to breast cancer patients with BRCA1 or BRCA2 mutation for the prevention of CBC. Further studies are required to get a more precise estimation of the benefits and harms of tamoxifen in these patients.

Breast cancer remains a world-wide challenge, and additional anti-cancer therapies are still urgently needed. Emerging evidence has demonstrated the potent anti-tumor effect of biguanides, among which phenformin was reported to potentially be a more active anti-cancer agent than metformin. However, little attention has been given to the role of phenformin in breast cancer. In this study, we reveal the role of phenformin in cell death of the MCF7, ZR-75-1, MDA-MB-231 and SUM1315 breast cancer cell lines. The respective IC50 values of phenformin in MCF7, ZR-75-1, MDA-MB-231 and SUM1315 cells were 1.184±0.045 mM, 0.665±0.007 mM, 2.347±0.010 mM and 1.885±0.015 mM (mean± standard error). Phenformin induced cell cycle change and apoptosis in breast cancer cells via the AMPK/mTOR/p70s6k and MAPK/ERK pathways. Interestingly, phenformin induced MET (mesenchymal-epithelial transition) and decreased the migration rate in breast cancer cell lines. Furthermore, our results suggest that phenformin inhibits breast cancer cell metastasis after intracardiac injection into nude mice. Taken together, our study further confirms the potential benefit of phenformin in breast cancer treatment and provides novel mechanistic insight into its anti-cancer activity in breast cancer.

Background: Aldehyde dehydrogenase 1 (ALDH1) is an important marker of tumor-initiating cells. We aimed to investigate ALDH1 expression in benign breast disease and human breast cancer of different histologic stages., Methods: Immunohistochemical staining of ALDH1 was applied to 21 cases with benign breast diseases, 47 ductal carcinoma in situ (DCIS) cases, 62 cases diagnosed with invasive cancer with extensive intraductal component (EIC), and 58 cases diagnosed with invasive cancer without EIC., Results: ALDH1 was expressed in tumor cells in 61.0% of 164 breast cancer cases, which was higher than that in benign breast disease (3/21) (P < 0.001). Of these 167 breast cancer cases, a significantly higher rate (54/58) of intratumoral ALDH1 expression was observed in invasive cancer without EIC cases than that in DCIS cases (19/46, one case not available) and invasive cancer with EIC cases (27/60, two cases not available) (P < 0.001). Interestingly, a significantly higher rate (22/48) of intratumoral ALDH1 expression in invasive component was observed than that in in situ component (7/48) in the same tumor (P = 0.001). In 47 DCIS cases, no significant association was observed between ALDH1 positivity and any clinicopathological parameter (all P > 0.05). However, ALDH1 positive invasive breast cancers were significantly more likely to be with large tumor size (P = 0.001), high grade (P < 0.001), and high Ki67 expression (P = 0.009)., Conclusions: ALDH1 may play an important role in the invasion of breast cancer, and may be associated with aggressive phenotypes of breast cancer., Virtual Slides: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1608671725154947 .

Background: Tubular adenoma of the breast is a rare benign epithelial tumor and only a few literatures have been reported; so far, no cases of tubular adenoma occurred in the accessory breast have been reported in the English literature. Clinical presentation and management of our patient are discussed along with a review of the literature on accessory mammary and tubular adenoma., Case Presentation: We present a case of 26-year-old woman (gravid 4, para 1) at 37 weeks of pregnancy with rapid enlargement in left anterior chest wall during pregnancy. Physical examination showed the left accessory breast was obviously bigger than the right one that only had a light areola around a small nipple. An elastic, mobile well-circumscribed mass measuring approximately 15 cm × 15 cm was palpated. Moreover, it was edematous and congestive with an increase in local temperature. The breast ultrasound further demonstrated the mass was a relatively homogeneous solid with short stripe blood flow signal. A single live fetus of 37 weeks gestation was observed by abdominal ultrasound scan. After a 2850 g male neonate was delivered, the right accessory breast and the mass in left accessory breast were removed. The resected specimen appeared as a solid white elastic mass with a smooth surface and the cut surface was red-grayish. Microscopically, the lesion consisted of tightly packed homogenous glandular structures which are supported by a single layer of myoepithelial cells with sparse intervening stroma., Conclusions: We describe a very rare case of giant tubular adenoma arising within an accessory breast in the anterior chest wall in a late pregnancy woman. The high concentrations of estrogen, progesterone and prolactin might account for the significant tumor enlargement during pregnancy. To our knowledge, this is the first case of giant tubular adenoma occurred within the accessory breast in the anterior chest wall., Virtual Slides: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/6210811191552106 .