Tumor-derived small extracellular vesicles promote breast cancer progression by upregulating PD-L1 expression in macrophages.

BACKGROUND: The metastasis of breast cancer (BC) is a complex multi-step pathological process, strictly dependent on the intrinsic characteristics of BC cells and promoted by a predisposing microenvironment. Although immunotherapy has made important progress in metastasis BC, the heterogeneity of PD-L1 in tumor associated macrophages (TAMs) in BC and the underlying mechanisms in the metastasis development of BC are still not completely elucidated. Small extracellular vesicles (sEVs) represent essential interaction mediators between BC cells and TAMs. It is worth noting to explore the underlying mechanisms typical of sEVs and their role in the metastasis development of BC. METHODS: The structure of sEVs was identified by TEM, while the particle size and amounts of sEVs were detected by BCA and NTA analysis. The specific PD-L1 + CD163 + TAM subpopulation in metastasis BC was identified by scRNA-seq data of GEO datasets and verified by IHC and IF. The function of TAMs and sEVs in metastasis BC was explored by RT-qPCR, WB, IF, flow cytometry and in vivo experiment. The expression profiles of plasma sEVs-miRNA in relation to BC metastasis was analyzed using next-generation sequencing. Further detailed mechanisms of sEVs in the metastasis development of BC were explored by bioinformatics analysis, RT-qPCR, WB and luciferase reporter assay. RESULTS: In this study, we identified that the immunosuppressive molecule PD-L1 was more abundant in TAMs than in BC cells, and a specific PD-L1 + CD163 + TAM subpopulation was found to be associated with metastasis BC. Additionally, we found that BC cells-derived sEVs can upregulate the PD-L1 expression and induce the M2 polarization, enhancing the metastasis development both in vitro and in vivo. Also, Clinical data showed that sEV-miR-106b-5p and sEV-miR-18a-5p was in relation to BC metastasis development and poor prognosis of BC patients. Further mechanistic experiments revealed that BC-derived sEV-miR-106b-5p and sEV-miR-18a-5p could synergistically promoted the PD-L1 expression in M2 TAMs by modulating the PTEN/AKT and PIAS3/STAT3 pathways, resulting in the enhancement of the BC cells invasion and metastasis. CONCLUSIONS: Our study demonstrated that BC-derived sEVs can induce metastasis in BC through miR-106b-5p/PTEN/AKT/PD-L1 and miR-18a-5p/PIAS3/STAT3/PD-L1 pathways in TAMs. Therefore, the inhibition of these specific interactions of signaling pathways would represent a promising target for future therapeutic strategies for treatment of BC.

CBX3 promotes breast cancer progression and high level of CBX3 predicts poor prognosis in patients.

Breast cancer is one of the leading cancer deaths around the world. Targeted drugs have greatly increased the survival rate of breast cancer patients in recent years. But in some patients, the current regimen is still ineffective. Therefore, more therapeutic targets for treating breast cancer are demanding. The core heterochromatin-related genes of breast cancer were identified by utilizing prognostic survival analysis and multivariate Cox hazard proportional regression analysis. Both breast cancer and adjacent normal tissue were collected and analyzed with western blot and immunohistochemistry. Colony formation assay, CCK-8 assay, and EdU assay were used to measure the effect of CBX3 on breast cancer cell growth, wound-healing assay and Transwell assay were used to analyze the effect of CBX3 on breast cancer cell migration and invasion. Flow cytometry assay and western blot were used to study the molecular mechanism of CBX3 in breast cancer. High expression of heterochromatin-related proteins CBX3, H2AFY, and SULF1 showed a poor prognosis in patients in both TCGA dataset and GEO datasets. Western blot demonstrated that the expression level of CBX3 was significantly higher in breast cancer than that in adjacent normal tissues. Colony formation assay, CCK-8 assay, and EdU assay showed that the knockdown of CBX3 could significantly inhibit breast cancer cell growth, and the overexpression of CBX3 could promote the growth of breast cancer cells. Transwell assay and wound healing assay showed that knockdown of CBX3 inhibited breast cancer cell migration and invasion, and the overexpression of CBX3 promoted breast cancer cell migration and invasion. Western blot showed that CBX3 might promote breast cancer cell proliferation, invasion, and migration in breast cancer by modulating the ERK1/2 signaling pathway and epithelial-mesenchymal transition (EMT)-related genes. CBX3 was a biomarker of poor prognosis in breast cancer patients. CBX3 promoted the proliferation of breast cancer cells through the ERK signaling pathway, and migration and invasion of breast cancer cells through EMT-related genes. The CBX3/p-ERK1/2 signaling axis might provide a new therapeutic method against breast cancer.

Potential values of circulating tumor cell for detection of recurrence in patients of thyroid cancer: a diagnostic meta-analysis.

BACKGROUND: Several studies have reported that circulating tumor cells (CTCs) are a promising marker for the diagnosis of thyroid cancer (TC) with recurrence or distant metastasis (DMs). However, some studies emerged with conflicting results. Therefore, we provide a meta-analysis to evaluate the diagnostic performance of CTC for detection of recurrence in patients of TC. METHODS: We searched PubMed, Web of Science, Cochrane library with the keywords "thyroid cancer" and "circulating tumor cells". Data extraction and risk of bias assessment were performed independently by two reviewers. The summary receiver operating characteristic curve (SROC) and other parameters were adopted to summarize the overall test performance. The sensitivity of CTCs in the detection of recurrent TC was reviewed. All analyses were performed by STATA 12.0 and Meta-disc software. RESULTS: For CTCs expressing epithelial cell adhesion molecule (EpCAM), seven studies were included in our meta-analysis. Pooled sensitivity, specificity, and diagnostic odds ratio were 0.71 (95% CI: 0.63-0.78), 0.89 (95% CI: 0.84-0.94), and 26.75 (95% CI: 9.11-78.53); 0.78 (95% CI: 0.65-0.89), 0.88 (95% CI: 0.76-0.96), and 40.01 (95% CI: 10.49-152.63) for CTCs expressing thyroid stimulating hormone receptor (TSHR). The area under the SROC for EpCAM and TSHR were both 0.91. CONCLUSION: CTC was a reliable marker for the diagnosis of TC patients with recurrence and DMs, and the sensitivity of CTCs expressing TSHR was higher than that of EpCAM. Additional research is warranted in order to establish uniformity in international guidelines, make up the drawbacks of conventional diagnostic methods and to prevent futile surgery.

Endoplasmic reticulum stress targeted therapy for breast cancer.

Recurrence, metastasis, and drug resistance are still big challenges in breast cancer therapy. Internal and external stresses have been proven to substantially facilitate breast cancer progression through molecular and systemic mechanisms. For example, endoplasmic reticulum stress (ERS) results in activation of the unfolded protein response (UPR), which are considered an important cellular stress response. More and more reports indicate its key role in protein homeostasis and other diverse functions involved in the process of breast cancer progression. Therefore, therapies targeting the activation of ERS and its downstream signaling pathways are potentially helpful and novel tools to counteract and fight breast cancer. However, recent advances in our understanding of ERS are focused on characterizing and modulating ERS between healthy and disease states, and so little attention has been paid to studying the role and clinical application of targeting ERS in a certain cancer. In this review, we summarize the function and main mechanisms of ERS in different molecular types of breast cancer, and focus on the development of agents targeting ERS to provide new treatment strategies for breast cancer. Video Abstract.

Hyperthermia promotes exosome secretion by regulating Rab7b while increasing drug sensitivity in adriamycin-resistant breast cancer.

PURPOSE: To investigate the mechanism through which hyperthermia promotes exosome secretion and drug sensitivity in adriamycin-resistant breast cancer. MATERIALS AND METHODS: We first evaluated the effect of hyperthermia on adriamycin-resistant breast cancer viability and used transmission electron microscopy, nanoparticle tracking analysis, and a bicinchoninic acid kit to validate the effect of hyperthermia on exosome secretion. The effective targeting molecules and pathways changed by hyperthermia were explored by RNA microarray and verified in vitro. The adriamycin-resistant MCF-7/ADR cells co-incubated with the exosomes produced by MCF-7/ADR cells after hyperthermia were assessed. The uptake of exosomes by MCF-7/ADR cells after hyperthermia treatment was evaluated by confocal microscopy. Finally, the mechanism through which hyperthermia promotes exosome secretion by hyperthermia was determined. RESULTS: Hyperthermia significantly suppressed the growth of adriamycin-resistant breast cancer cells and increased drug sensitivity by upregulating FOS and CREB5, genes related to longer overall survival in breast cancer patients. Moreover, hyperthermia promoted exosome secretion through Rab7b, a small GTPase that controls endosome transport. The upregulated FOS and CREB5 antioncogenes can be transferred to MCF-7/ADR cells by hyperthermia-treated MCF-7/ADR cell-secreted exosomes. CONCLUSIONS: Our results demonstrated a novel function of hyperthermia in promoting exosome secretion in adriamycin-resistant breast cancer cells and revealed the effects of hyperthermia on tumor cell biology. These hyperthermia-triggered exosomes can carry antitumor genes to the residual tumor and tumor microenvironment, which may be more beneficial to the effects of hyperthermia. These results represent an exploration of the relationship between therapeutic strategies and exosome biology.

Aun (n = 1-16) clusters on the ZrO2(111) surface: a DFT+U investigation.

The growth patterns and electronic structures of Aun clusters (n = 1-16) supported on the monoclinic ZrO2(111) surface were investigated using a DFT+U approach. We found that the supported Aun clusters prefer quasi-planar structures and lay flat on the ZrO2 surface. This result agrees well with the experimental results. Both orbital overlap and dispersion interactions contribute to the interaction between the Aun cluster and the ZrO2 surface. Electrons were transferred from the ZrO2 surface to the Aun cluster. Small energy gaps between unoccupied states in the Aun clusters and occupied states in the ZrO2 surface were found, especially for the supported Aun clusters with odd n, which may indicate that more electrons are excited from the ZrO2 surface to the Aun cluster even under visible-light irradiation. In other words, the ZrO2 support may be involved in the photocatalytic process when Aun/ZrO2 is used as a photocatalyst.

Pitavastatin induces autophagy-dependent ferroptosis in MDA-MB-231 cells via the mevalonate pathway.

Triple-negative breast cancer (TNBC) is more prone to recurrence and metastasis relative to other subtypes of breast cancer, leading to an extremely poor prognosis. The increasing potential chemoresistance of TNBC patients is mainly due to that tumor cells escape from apoptosis. In recent years, statins have demonstrated extensive anti-tumor effects. It is worth noting that statins have more effective anti-tumor effects on TNBC cells and drug-resistant breast cancer cells. Therefore, this study examines the superior cytotoxic effects of statins on TNBC cell lines and further explores their potential therapeutic mechanisms. We detected different cell phenotypes and found that statins significantly reduced the cell viability of TNBC cells. Specifically, pitavastatin showed an obvious induction in cell death, cell cycle arrest and oxidative stress in TNBC MDA-MB-231 cells. The reversal effect of iron chelator desferrioxamine (DFO) on the morphological and molecular biological changes induced by pitavastatin has revealed a new mode of cell death induced by pitavastatin: ferroptosis. This ferroptotic effect was strengthened by the decreased expression of glutathione peroxidase 4 (GPx4) as well as newly discovered ferroptosis suppressor protein 1 (FSP1). The data showed that ferroptotic death of MDA-MB-231 cells is autophagy-dependent and mediated by the mevalonate pathway. Finally, we found that therapeutic oral doses of statins can inhibit the growth of transplanted tumors, which establishes statins as a potential treatment for TNBC patients. In conclusion, we found pitavastatin could induce autophagy-dependent ferroptosis in TNBC cells via the mevalonate pathway which may become a potential adjuvant treatment option for TNBC patients.

Vaspin accelerates the proliferation, invasion and metastasis of Triple-Negative breast cancer through MiR-33a-5p/ABHD2.

OBJECTIVE: To explore the influence and the underlying mechanism of vaspin (visceral adipose tissue-derived serpin) on the development of triple-negative breast malignancy. METHODS: First, we analyzed medical records and screened out 22 breast cancer patients with different BMI according to inclusion and exclusion criterion, and measured serum vaspin of those patients. Then we studied the effects of vaspin on TNBC cell lines by using EdU assay, colony formation, transwell and wound-healing assay. Later, we used bioinformatics analysis to identify downstream effectors and verify with qRT-PCR, luciferase assay, western blot, etc. RESULTS: We found the vaspin level was positively correlated with BMI in breast malignant patients and vaspin could significantly enhance the proliferation, infiltration and transferring of triple-negative breast cancer cells by restraining the expression of miR-33a-5p. By using bioinformatic analysis and luciferase assay, we identified miR-33a-5p directly regulating ABHD2. CONCLUSION: Vaspin, as a cancer-promoting cytokine, may inhibit miR-33a-5p thus increasing the level of ABHD2 to promote the development of the triple-negative breast cancer.

Hyperthermia promotes M1 polarization of macrophages via exosome-mediated HSPB8 transfer in triple negative breast cancer.

PURPOSE: To investigate the mechanism underlying the modulation of M1 macrophage polarization by exosomes released from hyperthermia-treated triple-negative breast cancer (TNBC) cells. MATERIALS AND METHODS: In this study, the effects of hyperthermia on TNBC cells were examined using cell counting kit-8, apoptosis, and cell cycle assays. Transmission electron microscopy was used to identify the structure of exosomes, while bicinchoninic acid and nanoparticle tracking analysis were used to detect particle size and amounts of exosomes released after hyperthermia. The polarization of macrophages incubated with exosomes derived by hyperthermia-pretreated TNBC cells were assessed by RT-qPCR and flow cytometry analysis. Next, RNA sequencing was performed to determine the targeting molecules changed in hyperthermia-treated TNBC cells in vitro. Finally, the mechanism underlying the modulation of macrophage polarization by exosomes derived from hyperthermia-treated TNBC cells was examined by using RT-qPCR, immunofluorescence and flow cytometry analysis. RESULTS: Hyperthermia markedly reduced cell viability in TNBC cells and promoted the secretion of TNBC cell-derived exosomes. The hub genes of hyperthermia-treated TNBC cells were significantly correlated with macrophage infiltration. Additionally, hyperthermia-treated TNBC cell-derived exosomes promoted M1 macrophage polarization. Furthermore, the expression levels of heat shock proteins, including HSPA1A, HSPA1B, HSPA6, and HSPB8, were significantly upregulated upon hyperthermia treatment, with HSPB8 exhibiting the highest upregulation. Moreover, hyperthermia can induce M1 macrophage polarization by promoting exosome-mediated HSPB8 transfer. CONCLUSION: This study demonstrated a novel mechanism that hyperthermia can induce M1 polarization of macrophages via exosome-mediated HSPB8 transfer. These results will help with future development of an optimized hyperthermia treatment regime for clinical application, especially for combination treatment with immunotherapy.

Development and evaluation of an adenosine-to-inosine RNA editing-based prognostic model for survival prediction of bladder cancer patients.

Adenosine-to-inosine RNA editing (ATIRE) is a common form of ribonucleic acid (RNA) editing, which has highlighted the importance of ATIRE in tumors. However, its role in bladder cancer (BLCA) remains poorly understood. To study ATIRE impact on BLCA patient prognosis, we obtained ATIRE, gene expression, and clinical data from the Cancer Genome Atlas (TCGA) database for 251 patients, randomly dividing them into training and testing groups. Univariate proportional hazards model (COX) regression identified prognosis-associated ATIRE loci, while the least absolute shrinkage and selection operator (LASSO) selected final loci to construct prognostic models and generate ATIRE scores. We developed a nomogram to predict BLCA patients' overall survival (OS) and analyzed the effect of ATIRE editing levels on host gene expression. We also compared immune cell infiltration and drug treatment between patients with high and low ATIRE scores. The ATIRE prognostic prediction model was constructed using ten ATIRE loci that are closely associated with BLCA survival. Patients with high ATIRE scores showed significantly worse OS than those with low ATIRE scores. Furthermore, the nomogram, which incorporates the ATIRE score, can better predict the prognosis of patients. Multiple functional and pathway changes associated with immune responses, as well as significant differences in immune cell infiltration levels and response to drug therapy were observed between patients with high and low ATIRE scores. This study represented the first comprehensive analysis of the role of ATIRE events in BLCA patient prognosis and provided new insights into potential prognostic markers for BLCA research.

Construction of an m6A-related lncRNA model for predicting prognosis and immunotherapy in patients with lung adenocarcinoma.

N6-methyladenosine (m6A)-related lncRNAs could be involved in the development of multiple tumors with an unknown role in lung adenocarcinoma (LUAD). Hence, gene expression data and clinical data of LUAD patients were acquired from The Cancer Genome Atlas Database. The prognostic m6A-related lncRNAs were identified through differential lncRNA expression analysis and Spearman's correlation analysis. The least absolute shrinkage and selection operator regression was used to establish the prognostic risk model, so as to evaluate and validate the predictive performance with survival analysis and receiver operating characteristic curve analysis. The expression of immune checkpoints, immune cell infiltration and drug sensitivity of patients in different risk groups were analyzed separately. A total of 19 prognostic m6A-related lncRNAs were identified to set up the prognostic risk model. The patients were divided into high- and low-risk groups based on the median value of the risk scores. Compared with the patients in the low-risk group, the prognosis of the patients in the high-risk group was relatively worse. The receiver operating characteristic curves indicated that this model had excellent sensitivity and specificity. Multivariate Cox regression analysis demonstrated that the risk score could be supposed as an independent prognostic risk factor. We highlighted that the risk scores were correlated with immune cell infiltration and drug sensitivity for constructing a prognostic risk model in LUAD patients based on m6A-related lncRNAs.

Integration of bioinformatics and machine learning strategies identifies APM-related gene signatures to predict clinical outcomes and therapeutic responses for breast cancer patients.

BACKGROUND: Tumor antigenicity and efficiency of antigen presentation jointly influence tumor immunogenicity, which largely determines the effectiveness of immune checkpoint blockade (ICB). However, the role of altered antigen processing and presentation machinery (APM) in breast cancer (BRCA) has not been fully elucidated. METHODS: A series of bioinformatic analyses and machine learning strategies were performed to construct APM-related gene signatures to guide personalized treatment for BRCA patients. A single-sample gene set enrichment analysis (ssGSEA) algorithm and weighted gene co-expression network analysis (WGCNA) were combined to screen for BRCA-specific APM-related genes. The non-negative matrix factorization (NMF) algorithm was used to divide the cohort into different clusters and the fgsea algorithm was applied to investigate the altered signaling pathways. Random survival forest (RSF) and the least absolute shrinkage and selection operator (Lasso) Cox regression analysis were combined to construct an APM-related risk score (APMrs) signature to predict overall survival. Furthermore, a nomogram and decision tree were generated to improve predictive accuracy and risk stratification for individual patients. Based on Tumor Immune Dysfunction and Exclusion (TIDE) method, random forest (RF) and Lasso logistic regression model were combined to establish an APM-related immunotherapeutic response score (APMis). Finally, immune infiltration, immunomodulators, mutational patterns, and potentially applicable drugs were comprehensively analyzed in different APM-related risk groups. IHC staining was used to assess the expression of APM-related genes in clinical samples. RESULTS: In this study, APMrs and APMis showed favorable performances in risk stratification and therapeutic prediction for BRCA patients. APMrs exhibited more powerful prognostic capacity and accurate survival prediction compared to conventional clinicopathological features. APMrs was closely associated with distinct mutational patterns, immune cell infiltration and immunomodulators expression. Furthermore, the two APM-related gene signatures were independently validated in external cohorts with prognosis or immunotherapeutic responses. Potential applicable drugs and targets were mined in the APMrs-high group. APM-related genes were further validated in our in-house samples. CONCLUSION: The APM-related gene signatures established in our study could improve the personalized assessment of survival risk and guide ICB decision-making for BRCA patients.

Diagnostic accuracy of de-escalated surgical procedure in axilla for node-positive breast cancer patients treated with neoadjuvant systemic therapy: A systematic review and meta-analysis.

BACKGROUND: More initial clinical node-positive breast cancer patients achieve axillary pathological complete response (ax-pCR) after neoadjuvant systemic therapy (NST). Restaging axillary status and performing de-escalated surgical procedures to replace routine axillary lymph nodes dissection (ALND) is urgently needed. Targeted axillary lymph node biopsy (TLNB) is a novel de-escalated surgical strategy marking metastatic axillary nodes before NST and targeted dissection and biopsy intraoperatively to tailor individual axillary management. METHODS: This study provided a systematic review and meta-analysis to evaluate the feasibility and diagnosis accuracy of TLNB. Prospective and retrospective clinical trials on TLNB were searched from Pubmed, Embase, and Cochrane. Identification rate (IFR), false-negative rate (FNR), negative predictive value (NPV), and rate of ax-pCR were the outcomes of this meta-analysis. RESULTS: One thousand nine hundred and twenty patients attempted TLNB, with an overall IFR of 93.5% (95% confidence interval [CI] 90.1%-96.2%). IFR of three nodal marking methods, namely iodine seeds, clips, and carbon dye, was 95.6% (95% CI 91.2%-98.7%), 91.7% (95% CI 87.3%-95.4%), and 97.1% (95% CI 89.1%-100.0%), respectively. Of them, 847 patients received ALND, with an overall FNR of 5.5% (95% CI 3.3%-8.0%), and NPV ranged from 90.1% to 96.1%. Regression analysis showed that the overlap of targeted and sentinel biopsied nodes might associate with IFRs and FNRs. CONCLUSION: TLNB is a novel, less invasive surgical approach to distinguish initial node-positive breast cancer that achieves negative axillary conversion after NST. It yields an excellent IFR with a low FNR and a high NPV. A combination of preoperative imaging, intraoperative TLNB with SLNB, and postoperative nodal radiotherapy might affect the future treatment paradigm of primary breast cancer with nodal metastases.

Extracellular vesicles as drug vectors for precise cancer treatment.

Extracellular vesicles (EVs) are nano-sized vesicle structures secreted from a variety of cells, which carry numerous biological macromolecules, participate in cell signal transduction and avoid immune system clearance. EVs have a plethora of specific signal recognition factors, and many studies have shown that they can play an important role in the precise treatment of tumors. This review aims to compile the applications of EVs as nanocarriers for antitumor drugs, gene drugs and other nanomaterials with anticancer capability. Additionally, we systematically summarize the preparation methodology and expound upon how to improve the drug loading and cancer-targeting capacity of EVs. We highlight that EV-based drug delivery has the potential to become the future of precise cancer treatment.

Variation of Long Non-Coding RNA And mRNA Profiles in Breast Cancer Cells With Influences of Adipocytes.

BACKGROUND: It is well known that obesity is one of the risks for incurrence and development in breast cancer patients. Long non-coding RNAs (lncRNAs) are reported to participate in the composition of tumor microenvironment and to regulate breast cancer cell metabolic activities. However, there was rare study focused on the lncRNAs in breast cancer with the influences of adipocytes. The study aimed to investigate lncRNAs expression profiles and discover potential biomarkers to predict the incidence and progression of adipocyte-associated-breast cancer. METHODS: We co-cultured adipocytes with breast cancer cells and profiled the expression of lncRNAs as well as mRNAs by using the RNA-sequencing method. Wound Healing, Migration assays and Invasion assays were applied to verify the invasion and metastasis of cancer cells. RESULTS: MDA-MB-231/Hpa-V and SK-BR-3/Hpa-V cells showed elevated migration and invasiveness compared to the control group. A sum of 371 mRNAs (181 upregulated and 190 downregulated) and 850 lncRNAs(414 upregulated and 436 downregulated) were differentially expressed in MDA-MB-231/Hpa-V comparing to MDA-MB-231(P < 0.05; |log2 (fold change)|>1.2). GO enrichment, KEGG pathway and interaction networks demonstrated that differentially expressed lncRNAs were involved in functional categories, such as material metabolism, which might lead to the progression of breast cancer. CONCLUSION: Our study detected a lncRNA profile in breast cancer cells affecting by adipocytes and provided a better understanding of the tumor microenvironment. LncRNAs may be helpful to predict the therapeutic responses and prognosis of obese breast cancer patients.

The anti-cancer properties of heparin and its derivatives: a review and prospect.

Heparin, including unfractionated heparin (UFH), low-molecular-weight heparin (LMWH) and heparin derivatives, are commonly used in venous thromboembolism treatment and reportedly have beneficial effects on cancer survival. Heparin can affect the proliferation, adhesion, angiogenesis, migration and invasion of cancer cells via multiple mechanisms. The main mechanisms involve inhibition of heparanase, P-/L-selectin, angiogenesis, and interference with the CXCL12-CXCR4 axis. Here we summarize the current experimental evidence regarding the anti-cancer role of heparin and its derivatives, and conclude that there is evidence to support heparin's role in inhibiting cancer progression, making it a promising anti-cancer agent.