Role of CD44 as cancer stem cell marker in triple-negative breast cancer and its association with histological grade and angiogenesis

Introduction: Cancer stem cells (CSC) within the tumors play a central role in tumorigenesis. It is, thus, of utmost importance to identify these cells to develop effective cancer therapy. Triple-Negative Breast Cancer (TNBC) is an aggressive molecular subtype of breast cancer associated with poor patient outcomes. The role of CD44 immunohistochemistry (IHC) as a putative CSC in breast carcinomas, particularly of the TNBC-subtype is ambiguous, with equivocal results. Aims and Objectives: The present study aims to assess the role of CSC in breast carcinoma by immunohistochemical analysis of CD44 expression in TNBC. The association of TNBC expressing CSC with histological grade as well as with angiogenesis (using CD34 IHC) has been studied. Materials and Methods: Biopsy samples from 58 patients with infiltrating ductal carcinoma, NST were studied. The histology of the tumor was sub-classified into grades 1–3. Based on immunohistochemical analysis (ER, PR, HER2/Neu), the cases were divided into TNBC and NTNBC groups. The tissue sections were also subjected to analysis for CD44 to identify the CSC-phenotype and CD34 to evaluate angiogenesis, to determine the microvascular density (MVD). Results: Out of the 58 cases in the study, 28 were TNBC and 30 were NTNBC. CSC phenotype (CD44 positive) was expressed significantly higher in the TNBC (78%) versus the NTNBC (53%) (p-value 0.043). The MVD estimated using CD34 IHC was lower in the TNBC group in our study, though the difference was not statistically significant. A larger proportion of cases in TNBC showed a higher histological grade (35%) in comparison to NTNBC (27%). However, statistically, it was not significant. Conclusion: Our study demonstrated that CD44 as a CSC marker is seen significantly more in the TNBC category of invasive ductal carcinomas. Further large-scale studies, to confirm these findings, will be of potential therapeutic and prognostic value.

Expression of long non-coding RNA FRAPT in triple-negative breast cancer and its biological significance / 中国生物制品学杂志

@#Objective To investigate the expression of long non-coding RNA FRAPT(lncRNA FRAPT) in triple-negative breast cancer(TNBC) cells and analyze its effects on proliferation,cell cycle and drug resistance of TNBC cells.Methods Using scRNASeqDB and TCGA on-line bioinformatics database,the expression of lncRNA FRAPT in TNBC and its correlation with prognosis were detected;TNBC cell line MDA-MB-231 was transfected with si-lncRNA FRAPT and si-Ctrl(negative control) by using Lipofectamine~(TM) 2000 respectively,of which the proliferation and cell cycle changes after transfection were detected by SRB test and flow cytometry,and the drug resistance to chemotherapy drug paclitaxel(PTX)was detected.Results The expression of lncRNA FRAPT was up-regulated in TNBC tissues and cells,and its high expression was positively correlated with the poor prognosis of TNBC patients(Hazard Ratio=1.66,P=0.047).Silencing lncRNA FRAPT significantly inhibited the proliferation of TNBC cells,with the cell cycle arrested in G1 phase,while increased the sensitivity of TNBC cells to PTX.Conclusion LncRNA FRAPT was highly expressed in TNBC and related to tumor prognosis-Silencing lncRNA FRAPT inhibited proliferation and cell cycle of TNBC,and increased the sensitivity to chemotherapy drug PTX.

Roles of lncRNA in the diagnosis and prognosis of triple-negative breast cancer / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Breast cancer is a malignant tumor that seriously endangers women's lives. The prognosis of breast cancer patients differs among molecular types. Compared with other subtypes, triple-negative breast cancer (TNBC) has been a research hotspot in recent years because of its high degree of malignancy, strong invasiveness, rapid progression, easy of recurrence, distant metastasis, poor prognosis, and high mortality. Many studies have found that long non-coding RNA (lncRNA) plays an important role in the occurrence, proliferation, migration, recurrence, chemotherapy resistance, and other characteristics of TNBC. Some lncRNAs are expected to become biomarkers in the diagnosis and prognosis of TNBC, and even new targets for its treatment. Based on a PubMed literature search, this review summarizes the progress in research on lncRNAs in TNBC and discusses their roles in TNBC diagnosis, prognosis, and chemotherapy with the hope of providing help for future research.

Isotoosendanin exerts inhibition on triple-negative breast cancer through abrogating TGF-β-induced epithelial-mesenchymal transition via directly targeting TGFβR1

As the most aggressive breast cancer, triple-negative breast cancer (TNBC) is still incurable and very prone to metastasis. The transform growth factor β (TGF-β)-induced epithelial-mesenchymal transition (EMT) is crucially involved in the growth and metastasis of TNBC. This study reported that a natural compound isotoosendanin (ITSN) reduced TNBC metastasis by inhibiting TGF-β-induced EMT and the formation of invadopodia. ITSN can directly interact with TGF-β receptor type-1 (TGFβR1) and abrogated the kinase activity of TGFβR1, thereby blocking the TGF-β-initiated downstream signaling pathway. Moreover, the ITSN-provided inhibition on metastasis obviously disappeared in TGFβR1-overexpressed TNBC cells in vitro as well as in mice bearing TNBC cells overexpressed TGFβR1. Furthermore, Lys232 and Asp351 residues in the kinase domain of TGFβR1 were found to be crucial for the interaction of ITSN with TGFβR1. Additionally, ITSN also improved the inhibitory efficacy of programmed cell death 1 ligand 1 (PD-L1) antibody for TNBC in vivo via inhibiting the TGF-β-mediated EMT in the tumor microenvironment. Our findings not only highlight the key role of TGFβR1 in TNBC metastasis, but also provide a leading compound targeting TGFβR1 for the treatment of TNBC metastasis. Moreover, this study also points out a potential strategy for TNBC treatment by using the combined application of anti-PD-L1 with a TGFβR1 inhibitor.

Effect of Chaihu Guizhitang on Triple-negative Breast Cancer Cells： Based on HIF-1α/VEGFA Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the effect of Chaihu Guizhitang on triple-negative breast cancer （TNBC） cells based on hypoxia-inducible factor-1α （HIF-1α）/vascular endothelial growth factor A （VEGFA） signaling pathway. MethodTNBC xenograft model was established and the cells were randomized into model group， capecitabine group （0.2 mg·kg-1）， Chaihu Guizhitang low-dose group， medium-dose group， and high-dose group （10.62， 21.23， 42.46 g·kg-1）， with 10 mice in each group. After 21 days of medication， the content of tumor necrosis factor-α （TNF-α） in serum was detected by enzyme-linked immunosorbent assay （ELISA）. The expression of HIF-1α mRNA was detected by real-time fluorogenic quantitative polymerase chain reaction （real-time PCR）. Immunohistochemistry （IHC） was employed to detect the expression of HIF-1α， TNF-α， and VEGFA in tumor tissues， and CD34 staining to examine the angiogenesis in tumor tissues. Microvessel density （MVD） was calculated， and the protein expression of HIF-1α， VEGFA， and epidermal growth factor receptor （EGFR） in tumor tissues was measured by Western blot. ResultCompared with the model group， the rest four groups showed low levels of TNF-α （P<0.01）， HIF-1α mRNA （P<0.01）， expression of HIF-1α， TNF-α， VEGFA， and CD34 in cells， and MVD （P<0.05， P<0.01）， and low protein levels of HIF-1α， VEGFA， and EGFR （P<0.01）. Compared with capecitabine group， medium-dose and high-dose Chaihu Guizhitang decreased the level of TNF-α （P<0.01）， HIF-1α mRNA （P<0.01）， expression of HIF-1α， TNF-α， and VEGFA in cells （P<0.01）， CD34 expression， MVD， and protein levels of HIF-1α， VEGFA， and EGFR （P<0.01）. ConclusionChaihu Guizhitang may inhibit the angiogenesis in TNBC cells by regulating the expression of HIF-1α/VEGFA signaling pathway， thus exerting anti-tumor effect.

Management of human epidermal growth factor receptor 2杗egative metastatic breast cancer: Role of poly adenosine diphosphate (ADP?ribose) polymerase inhibitors

Human epidermal growth factor receptor 2 (HER2)?negative subset is the most heterogeneous group of metastatic breast cancers (MBCs) as it includes both hormone receptor (HR)?positive and HR?negative breast cancer (or TNBC), which have different therapies and treatment challenges. Though endocrine therapy (ET) remains the treatment backbone in HR?positive HER2?negative cases, about 40% of the patients show intrinsic or acquired resistance to ET due to multiple mechanisms. Combining different therapies such as ET and other targeted therapies with or without chemotherapy fails to give continued benefit, unlike cyclin?dependent kinase (CDK) 4/6 inhibitors that have shown a great benefit. TNBC has conventionally been treated ineffectively with systemic chemotherapy. Recently, poly (ADP?ribose) polymerase inhibitors (PARPi) have emerged for HER2?negative breast cancer (BC) patients, including TNBC. Olaparib and talazoparib have recently been approved in germline BRCA?mutated (gBRCAm) HER2?negative MBC. Additionally, ongoing trials of PARPi in combination with various therapies are expected to provide more and better treatment options for gBRCAm HER2?negative breast cancer.

Signaling Pathways for the Targeted Therapy of Triple-Negative Breast Cancer and Their Clinical Applications / 中国生物化学与分子生物学报

Triple negative breast cancer (TNBC) accounts for 15%-20 % of all breast cancer cases, with the negative expressions of estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2. TNBC is a highly aggressive and malignant subtype, which also has a high risk of recurrence and poor prognosis. Due to its high heterogeneity and complicated clinical features, chemotherapy, radiotherapy, and surgical resection are the dominant methods for TNBC treatment currently. However, severe side effects, high risk of recurrence, and damages to health are not neglectable. With the progress of basic research on TNBC, more and more signaling pathways suitable for the targeted therapy of TNBC have been revealed, and some of them have successfully entered into clinical trials, suggesting promising molecular targets in TNBC treatment. Moreover, some of these theraputic targets play important roles in the classification and precise treatment of TNBC. This paper reviewed the research progress and clinical trials of the classic signaling pathways in the targeted therapy of TNBC, including synthetic lethality pathway, PI3K/AKT/mTOR pathway, PD-1/PD-L1 immune pathway, et al. Meanwhile, we also introduced potential signaling pathways revealed in recent years, including tumor angiogenesis pathway, polyamine synthesis and metabolism pathway, SLC3A2/LAT1 transportation pathway, and IGF-1/IGF-1R/FAK/YAP signal transduction pathway,et al.

Reprogramming of Glucose Metabolism Pathways in Triple-negative Breast Cancer Cells under Hypoxia / 中国生物化学与分子生物学报

Metabolic changes are recognized as one of the hallmarks of cancer cells. Previous studies have shown that hypoxia can change the glucose metabolism of cancer cells. However, the mechanisms still need to be studied in detail. In this study, by using RNA-Sequencing (RNA-seq) and bioinformatics analysis, we found that there is a significant change in the expression of 334 genes in BT549 cells and 215 genes in MDA-MB-231 cells induced by hypoxia at the mRNA level. Most of these genes were associated with glucose metabolism. RNA-seq data, Western blot, enzyme activity assays, and metabolite quantification experiments showed that the glucose uptake increased by elevating the expression of glucose transporter protein 1 (GLUT1) in BT549 cells and GLUT1 and GLUT3 in MDA-MB-231 cells induced by hypoxia. Hypoxia promotes glycolysis by increasing the expression of at least one isozyme or enzyme protein subunit of the enzymes that catalyze the each reaction in the glycolysis pathway, as well as the regulatory enzymes 6-phosphofructo-2-kinase/fructose-2, 6-biphosphatase 3 (PFKFB3) and 4 (PFKFB4) isozymes. Hypoxia increases the expression of pyruvate dehydrogenase kinase 1 (PDK1) and PDK3 and reduces the expression of isocitrate dehydrogenase 3(IDH3), succinate dehydrogenase subunit B(SDHB) and D(SDHD) to reduce the aerobic oxidation pathway. The expression of glucose-6-phosphate dehydrogenase (G6PD) and glycogen synthase was significantly increased to promote the pentose phosphate pathway and glycogen synthesis pathway. The mRNA levels of enzyme genes had no significant difference in gluconeogenesis and glycogenolysis. The breast cancer database suggests that the mRNA levels of the glucose metabolism enzymes were consistent in vivo and in vitro. Hypoxia regulates the reprogramming of glucose metabolism by altering the gene expression of isozymes or subunits of these enzymes. The study comprehensively analyzed the effects of hypoxia on the expression of all enzyme genes and major regulatory enzymes in six glucose metabolism pathways and provides an understanding of glucose metabolism in cancer cells under a hypoxia environment.

Advances in Treatment of Triple Negative Breast Cancer / 肿瘤防治研究

TNBC is a special type of breast cancer with strong aggressiveness and poor prognosis. Chemotherapy is still the main treatment for TNBC, due to poor efficacy of endocrine therapy and targeted therapy. However, TNBC is a kind of heterogeneous disease, so it is urgent to study the precise molecular types and explore new precision treatment. This paper will summarize the results of clinical trials and analyze treatment strategies for TNBC, including surgical treatment, radiotherapy, chemotherapy, targeted therapy and immunotherapy, in order to provide evidence for clinical management.

Caracterização funcional do gene codificador de proteína CD14 e do gene não codificador de proteína LINC01133 em linhagens celulares humanas de câncer de mama triplo negativo / Functional characterization of the CD14 protein-coding gene and the non-protein-coding LINC01133 gene in triple negative human breast cancer cell lines

Dentre os subtipos de câncer de mama, o triplo negativo (TNBC) é o que apresenta as maiores taxas de mortalidade, sendo, portanto, considerado um enorme desafio para a clínica. O uso de moléculas como marcadores tumorais vem auxiliando o clínico no diagnóstico, no prognóstico e, até mesmo, no tratamento do TNBC, sendo essenciais na redução de suas altas taxa de mortalidade. No entanto, um pequeno grupo de marcadores tumorais são validados na prática clínica, estimulando à busca por novos alvos, e sua caracterização funcional, como forma de se entender a Biologia desta doença. Assim, o objetivo deste trabalho é caracterizar funcionalmente o gene codificador de proteína CD14 e o gene não codificador de proteína LINC01133 em linhagens celulares humanas de TNBC, no intuito de descobrir o papel destas moléculas na progressão tumoral. Na primeira parte deste trabalho, analisou-se a expressão do CD14 frente à um painel de linhagens celulares que representam os diferentes subtipos dos tumores mamários. O CD14 exibiu elevados níveis de expressão nas linhagens nãotumorigênicas MCF10A e MCF12A e baixos níveis na linhagem triplo negativa Hs578T. A partir destes resultados, o CD14 foi superexpresso na linhagem Hs578T. Ensaios de caracterização funcional mostraram que a superexpressão do CD14 reduziu a capacidade migratória e invasiva das células, efeito que foi hipoteticamente relacionado ao aumento da expressão da E-caderina. No entanto, observou-se aumento no potencial tumorigênico, levando-nos a sugerir seu envolvimento num possível mecanismo utilizado pelas células para compensar a significativa redução do potencial migratório e invasivo. Os resultados obtidos indicam que o nível basal de expressão do CD14 observado na linhagem Hs578T é importante, podendo contribuir para a desenvolvimento primário do tumor, atuando como um oncogene. Na segunda parte deste trabalho, analisou-se a expressão de 10 RNAs longos não codificadores (lncRNAs), frente ao mesmo painel de linhagens descritoanteriormente. Dentre estes, o lncRNA LINC01133 exibiu baixos níveis de expressão nas linhagens não-tumorigênicas MCF10A e MCF12A e elevados níveis na linhagem triplo negativa Hs578T, sendo, então, escolhido como alvo de estudo. A partir destes resultados, decidimos superexpressar, de forma indutível, o LINC01133 na linhagem MCF10A e nocautear este gene, via sistema CRISPR/Cas9, na linhagem Hs578T. Ensaios de caracterização funcional mostraram que a superexpressão do LINC01133 na linhagem MCF10A reduziu a proliferação celular e inibiu o crescimento de colônias dependente de ancoragem, mas, em contrapartida, aumentou o crescimento de colônias independente de ancoragem e a capacidade migratória e invasiva destas células. No entanto, sugerimos que isto não seja suficiente para tornar estas células tumorigênicas e metastáticas. Por outro lado, o nocauteamento do LINC01133 na linhagem triplo negativa Hs578T aumentou de forma considerável todos os parâmetros de malignidade analisados. Baseado nos dados obtidos, sugerimos que o elevado nível de expressão do LINC01133 na linhagem Hs578T é importante na regulação negativa de processos relacionados com a progressão tumoral, atuando com um supressor tumoral. Os dados obtidos em nosso estudo contribuem para o enriquecimento de informações relacionadas à Biologia do TNBC, auxiliando, desta forma, no desenvolvimento de potenciais protocolos clínicos e terapêuticos utilizandos estes biomarcadores

Among the breast cancer subtypes, the triple negative (TNBC) displays the highest mortality rates, being, therefore, considered a major challenge for the clinic. The use of molecules as tumor markers has helped clinicians in the diagnosis, prognosis and even in treatment of TNBC, being essential in reducing its high mortality rate. However, a small group of tumor markers is validated in clinical practice, stimulating the search for new targets, and their functional characterization, as a way to understand the biology of this disease. Thus, the aim of this work is to functionally characterize the CD14 protein-coding gene and the non-protein-coding LINC01133 gene in human TNBC cell lines, in order to probe into the role of these molecules in tumor progression. In the first part of this work, the expression of CD14 was analyzed in a panel of cell lines that represent the different subtypes of breast tumors. High expression levels of CD14 were observed in the non-tumorigenic MCF10A and MCF12A lineages and low levels in the triple negative Hs578T lineage. Based on these results, CD14 was overexpressed in the Hs578T lineage. Functional characterization assays showed that CD14 overexpression reduced the migratory and invasive capacity of cells, an effect that was hypothetically related to increased E-cadherin expression. However, increased in the tumorigenic potential was observed, leading us to suggest its involvement in a possible mechanism used by cells to compensate for the significant reduction in the migratory and invasive potential. The results obtained indicate that CD14 expression basal level observed in the Hs578T lineage may be important to contribute to the primary development of tumor, thus acting as an oncogene. In the second part of this work, the expression of 10 long non-coding RNAs (lncRNAs) was analyzed against the same lineage panel described above. Among these, the LINC01133 lncRNA exhibited low expression levels in the non-tumorigenic MCF10A and MCF12A lineages and high levels in the triple negative Hs578T lineage, being, then, chosen as a target for this study. Based on these results, we decided toinducibly overexpress LINC01133 in the MCF10A lineage and knockout this gene, via the CRISPR/Cas9 system, in the Hs578T lineage. Functional characterization assays showed that overexpression of LINC01133 in the MCF10A lineage reduced cell proliferation and inhibited anchorage-dependent colony growth, but, on the other hand, increased anchorage-independent colony growth and the migratory and invasive capacity of these cells. However, we suggest that this is not sufficient to render these cells tumorigenic and metastatic. On the other hand, the knockout of LINC01133 in the triple negative Hs578T lineage considerably increased all the analyzed malignancy parameters. Based on the results obtained, we suggest that the high expression level of LINC01133 in the Hs578T lineage is important for down-regulation of processes related to tumor progression, acting as a tumor suppressor. The data obtained in our study contribute to the enrichment of information related to TNBC Biology, thus assisting in the development of potential clinical and therapeutic protocols using these biomarkers

Immunohistochemical Survey of Invasive Ductal Carcinoma of the Breast, using ER, PR, HER2 and KI-67 biomarkers, in Uyo, Nigeria

Background: Breast's Invasive Ductal Carcinoma (IDC), which is the commonest type of malignancy in females worldwide, can be characterized using immunohistochemistry in view of personalized cancer therapy. In this study, we aimed to determine the pattern of immunohistochemical profiles of IDC using oestrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor 2 receptor (HER2) and proliferative index (Ki-67) biomarkers in our tertiary healthcare facility in Uyo, Akwa Ibom State, Nigeria given the dearth of its data in our environment. Materials and methods: We carried out a retrospective hospital-based immunohistochemical study of archival IDC tissue blocks over a four- and half-year period. Using systematic random sampling method, 64 formalin fixed paraffin embedded (FFPE) IDC tissue blocks were selected for this study. We carried out immunohistochemical evaluation using ER, PR, HER2 and Ki-67 biomarkers. Subsequently, we presented the results and classification schemes as text, tables, graphs, and photomicrographs. Results: We found that the proportion of expressions were ER-negative (88.7%), PR-negative (87.3%), HER2-negative (68.3%) and Ki-67 (<20%) being 83.6% respectively. The immunohistochemical-based classification which was done using combined immunohistochemical profiles of ER/PR/HER2 and ER/PR/HER2/Ki-67 biomarkers respectively, revealed five immunohistochemical-based subtypes. These subtypes were ER-positive luminal A (ER+/±PR+/HER2-) [5.56%], ER-positive luminal B (ER+/±PR+/HER2+) [5.56%], HER2-overexpression (ER-/±PR+/HER2+) [16.66%], Triple negative (ER-/PR-/HER2-) [66.67%] and Unclassified subtypes (ER-/PR+/HER2-) [5.56%]. Furthermore, these five subtypes were further subcategorized into low (Ki-67 <20%) and high (Ki-67 ≥20%) proliferation subtypes accordingly. Conclusion: The commonest pattern of immunohistochemical profile expression of IDC in Uyo was found to be the Triple negative subtype.

NELFE promotes the migration and invasion of triple negative breast cancer cells by inhibiting NDRG2 expression / 西安交通大学学报(医学版)

【Objective】 To explore the effect of negative elongation factor E(NELFE) on the invasion and migration of triple-negative breast cancer(TNBC) cells and its mechanisms. 【Methods】 qRT-PCR was used to detect the expression of negative elongation factor E(NELFE) in triple-negative breast cancer(TNBC) tissues and adjacent tissues. NELFE expression was upregulated in MDA-MB-231 cell lines via lentivirus transfection. Transwell assay was used to observe the effect of elevated NELFE expression on the migration and invasion of MDA-MB-231 cells. The RNA stability assay was used to detect the influence of NELFE on the stability of the mRNA of N-myc downstream regulated gene2(NDRG2). Western blotting analysis was used to detect the regulatory function of NELFE on NDRG2 protein expression. 【Results】 NELFE expression was increased in TNBC tissues. After the overexpression of NELFE, the migration and invasion of MDA-MB-231 cells were significantly enhanced, NDRG2 mRNA stability was decreased, and NDRG2 protein expression level was downregulated. NDRG2 expression was upregulated in MDA-MB-231 cell lines with overexpressed NELFE; cell migration and invasion abilities were decreased. 【Conclusion】 NELFE inhibited NDRG2 expression by decreasing the stability of NDRG2 mRNA and promoted the migration and invasion of TNBC cells.

Postoperative Survival of Triple Negative Breast Cancer Patients After High Exposure to Traditional Chinese Medicine： A Cohort Study / 中国实验方剂学杂志

Objective:To evaluate the 3-year survival outcomes of postoperative patients after high exposure to traditional Chinese medicine （TCM） for triple negative breast cancer （TNBC）. Method:The complete 3-year follow-up data of 150 postoperative patients with stage I–III TNBC were retrospectively analyzed. All the patients received routine western medical treatments （surgery， chemotherapy， and/or radiotherapy） according to the National Comprehensive Cancer Network （NCCN） clinical practice guidelines in oncology as well as TCM. According to the degree of exposure to TCM， they were divided into the high- and low-exposure cohorts， with the oral administration of Chaihu Longmu Decoction with or without anti-cancer Chinese patent medicine for at least six months annually， or 18 months or more in the three years as the inclusion criterion for the former cohort. The metastatic sites of recurrent TNBC and the recurrent metastasis/death time were observed in both cohorts to compare the disease-free survival （DFS） and overall survival （OS）. The influences of onset age， pathological type， histopathological grade， vascular invasion， clinical stage， and exposure to TCM on survival were subjected to statistical analysis， followed by the observation of adverse effects. Result:There was no significant difference in the metastatic sites between the two cohorts （<italic>P</italic>>0.05）. The high-exposure cohort had a longer 3-year DFS than the low-exposure cohort， and the 3-year DFS rate in the high-exposure cohort was increased by 16.9% （χ²=6.995， <italic>P</italic>=0.008） as compared with that in the low-exposure cohort， exhibiting a significant difference （<italic>P</italic><0.05）. As revealed by the Cox proportional-hazards model， patients in the low-exposure cohort had a 3.724-fold as high risk of recurrent metastasis as that in the high-exposure cohort （95%CI 1.399~9.915）. There was no significant difference in the 3-year OS between the two cohorts （<italic>P</italic>>0.05）. The overall incidence of adverse effects in both groups was 7.3%， mainly manifested as gastrointestinal discomfort. Conclusion:High exposure to TCM contributes to reducing postoperative recurrence and metastasis and prolonging DFS.

Regulatory Effect of Huadu Sanyinfang on PI3K/Akt/NF-κB Pathway in Triple-negative Breast Cancer Patients with Qi-deficiency Constitution： An Exploration Based on Differential Expression of miRNA / 中国实验方剂学杂志

Objective:To explore the regulatory effect of Huadu Sanyinfang on phosphatidylinositol 3 kinase（PI3K）/protein kinase B（Akt）/transcription factor nuclear factor-<italic>κ</italic>B （NF-<italic>κ</italic>B） in triple-negative breast cancer （TNBC） patients with qi-deficiency constitution based on the differential expression of miRNA. Method:Based on previous research results， this study conducted the bioinformatics analysis to predict the target genes responsible for regulating the differential expression of miRNA between patients with qi-deficiency constitution and those with moderate constitution， which were intersected with TNBC target genes. The resulting intersection targets were then subjected to Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis and protein-protein interaction （PPI） network analysis to obtain the key pathways and target genes for differentially expressed miRNA in regulating TNBC. TNBC patients with Qi-deficiency constitution were treated with Huadu Sanyinfang for three years after they completed the standard Western medical treatment. The peripheral blood of the patients was sampled before and after medication for detecting gene expression in the key pathways. Result:The comparison between patients with Qi-deficiency constitution and those with moderate constitution revealed 49 differentially expressed miRNAs （16 up-regulated and 33 down-regulated）， which regulated 1 445 TNBC target genes. As demonstrated by PPI and KEGG pathway enrichment analysis， the key genes were mainly tumor protein p53 （TP53）， Akt1， epidermal growth factor receptor （EGFR）， mitogen-activated protein kinase 3 （MAPK3）， vascular endothelial growth factor A （VEGRA）， and tumor necrosis factor （TNF）. The key pathways included PI3K/Akt， MAPK， and RAS signaling pathways. A total of 11 TNBC patients with qi-deficiency constitution were enrolled. Compared with the situations before treatment， the expression levels of p105 subunit of NF-κB （NF-<italic>κ</italic>B1） and Akt1 in the PI3K/Akt signaling pathway were down-regulated after medication， while the levels of catalytic subunit alpha of PI3K （PIK3CA） and B-cell lymphoma-xL （Bcl-xL） were up-regulated. The differences in NF-<italic>κ</italic>B1 and Akt1 expression were statistically significant. Conclusion:Huadu Sanyinfang is able to affect the gene expression of PI3K/Akt/NF-<italic>κ</italic>B signaling pathway in TNBC patients with Qi-deficiency constitution. Specifically， it down-regulates NF-<italic>κ</italic>B1 and Akt1 expression and up-regulates PIK3CA and Bcl-xL.

Economic evaluation of the SP142 versus 22C3 PD-L1 assays in the treatment of atezolizumab plus nab-paclitaxel for patients with advanced triple negative breast cancer in the Brazilian private healthcare systemp / Avaliação econômica dos testes de PD-L1 SP142 versus 22C3 no tratamento com atezolizumabe mais nab-paclitaxel em pacientes com câncer de mama triplo-negativo avançado no sistema de saúde suplementar no Brasil

Objective: The aim of the study was to demonstrate the economic impact of two PD-L1 immunohistochemistry (IHC) assays, SP142 versus 22C3, in the treatment with atezolizumab plus nab-paclitaxel in patients with advanced triple negative breast cancer (aTNBC) in the Brazilian private healthcare system (BPHS). Methods: The study performed two analyses: one per patient and other of the potential population projected for the BPHS (budget impact analysis). Data of progressionfree survival and overall survival were extracted from a post hoc analysis of the IMpassion130 trial to develop a partitioned-survival model to simulate the economic impact of the treatment with atezolizumab plus nab-paclitaxel guided by the SP142 and 22C3 assays on patients with aTNBC. The analyses included only direct costs that were based on CBHPM (Classificação Brasileira Hierarquizada de Procedimentos Médicos) and CMED (Câmara de Regulação do Mercado de Medicamentos) PF18% tables. A univariate sensitivity analysis was performed with the parameters varying ± 20%. Results: The study has demonstrated that the SP142 assay has the potential to save ­BRL 179,730 with the treatment of atezolizumab plus nab-paclitaxel per patient with aTNBC in five years. Conclusion: The SP142 assay can optimize the use of atezolizumab plus nab-paclitaxel avoiding its prescription in patients who will not have a significant clinical improvement.

Objetivo: O objetivo do estudo foi demonstrar o impacto econômico de dois testes de imuno-histoquímica, SP142 versus 22C3, no tratamento com atezolizumabe + nab-paclitaxel em pacientes com câncer de mama triplo-negativo avançado (CMTNa) no sistema de saúde suplementar (SSS) no Brasil. Métodos: O estudo realizou duas análises: uma por paciente e outra na população potencial projetada para o SSS (análise de impacto no orçamento). Dados de sobrevida livre de progressão e de sobrevida global foram extraídos da análise post hoc do estudo IMpassion130 para o desenvolvimento de um modelo de sobrevida particionado que simulasse o impacto econômico do tratamento com atezolizumabe + nab-paclitaxel direcionado pelos testes SP142 e 22C3 em pacientes com CMTNa. A análise considerou somente os custos diretos baseados nas tabelas CBHPM (Classificação Brasileira Hierarquizada de Procedimentos Médicos) e CMED (Câmara de Regulação do Mercado de Medicamentos) PF18%. Uma análise de sensibilidade univariada foi realizada variando os parâmetros em ± 20%. Resultados: O estudo demonstrou que o teste SP142 apresenta um potencial de economia de -179.730 reais (BRL) no tratamento de atezolizumabe + nab-paclitaxel por paciente com CMTNa em cinco anos. Conclusão: O uso do teste SP142 possibilita otimizar o uso de atezolizumabe + nab-paclitaxel evitando a sua prescrição em pacientes que não irão se beneficiar de forma significativa.

Analyzing The Role Of Phytochemicals In Targeting Drug Transporter Protein Abcc6 Using Molecular Docking And Molecular Dynamics Simulations

Triple negative breast cancer (TNBC) is an aggressive breast cancer subtype that lacks hormonal receptors. This reduces the therapeutic options for TNBC patients creating more focus on chemotherapy. Drug resistance has posed as a major hurdle in treating TNBC patients. Deregulation of drug transporter proteins is one of major factors that cause resistance to chemotherapeutic drugs. In this study, ABCC6 a drug transporter protein that is found dysregulated in several resistant cancer cells has been docked with natural compounds or phytochemicals with known anti-cancer activities. Subtrifloralactone G, a withanolide extracted from Deprea subtriflora is found to show highest binding energy with ABCC6 protein. Molecular dynamics simulations further prove the stability of the ABCC6 protein- Subtrifloralactone G ligand complex. ADMET analysis shows that phytochemical Subtrifloralactone G can be used as an anti-cancer therapeutic drug in treating resistant cancer cells. The study mainly focuses on the role of phytochemicals in treating resistant TNBC cells.

LncRNA XIST interacts with miR-454 to inhibit cells proliferation, epithelial mesenchymal transition and induces apoptosis in triple-negative breast cancer

Triple-negative breast cancer (TNBC) is a one of the subtypes of breast cancer which accounts for approximately 10–20% of all breast cancers. LncRNA XIST (XIST) is reported to be dysfunctional in numerous tumortypes and is involved in the key pathways of cancer initiation, progression and metastasis. Thus, in the presentstudy, we explored the detailed molecular mechanism of XIST in TNBC. XIST was down-regulated in TNBCtissues and cell lines. Overexpressed XIST inhibited cell proliferation, epithelial mesenchymal transition(EMT) and induced apoptosis in vitro as well as suppressed TNBC tumor growth in vivo. MicroRNA (miR)-454 was up-regulated in TNBC tissues and cell lines. Knockdown of miR-454 inhibited TNBC progression bysuppressing cell proliferation, EMT and inducing cell apoptosis. Moreover, miR-454 was predicted andconfirmed to be a target of XIST, and rescue assay indicated that overexpressed miR-454 could reverse XISTrestoration mediated-anti-tumor effects on TNBC cells. In conclusion, XIST interacts with miR-454 to inhibitcells proliferation, EMT and induce apoptosis in TNBC, indicating a promising treatment strategy for TNBCpatients.

Afzelin inhibits migration of MDA-MB-231 cells by suppressing FAK expression and Rac1 activation

Triple-negative breast cancer (TNBC) has the worst prognosis and the highest rate of metastasis among other types ofbreast cancer. These characteristics are supported by the dysregulation of focal adhesion kinase (FAK) and Rac1 whichare the key players of mesenchymal cell migration on TNBC. Afzelin is a secondary metabolite that is contained ina variety of plants. This study explored the anti-migration effect of afzelin and its interaction with FAK and Rac1 onthe highly invasive TNBC cell line, MDA-MB-231. Cell viability was assessed by 3-(4,5-dimethyl 2-thiazolyl)-2,5-diphenyltetrazolium bromide assay, and cell migration was evaluated using in vitro scratch assay. Rac1 activation wasanalyzed using the colorimetric assay, while vinculin and actin filaments were stained through immunofluorescence. Thequantity of total FAK and phosphorylated FAK tyr397 was detected by Western blotting. Afzelin decreased cell viabilityand inhibited two-dimensional cell migration in a dose-dependent manner. Under confocal laser scanning microscopy,vinculin localization at the cell edge demonstrated a reduction of focal adhesion formation by afzelin. Further explorationshowed that afzelin decreased FAK expression but did not affect FAK phosphorylation at tyr397. In addition, afzelindecreased Rac1-GTPase activation, which is a downstream effector of FAK. Taken together, these results suggest thatafzelin suppresses TNBC cell migration, through inhibition of FAK expression and Rac1-GTPase activation.

Roles of miRNA and lncRNA in triple-negative breast cancer / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Triple-negative breast cancer (TNBC) is currently the most malignant subtype of breast cancer without effective targeted therapies, which makes its pathogenesis an important target for research. A growing number of studies have shown that non-coding RNA (ncRNA), including microRNA (miRNA) and long non-coding RNA (lncRNA), plays a significant role in tumorigenesis. This review summarizes the roles of miRNA and lncRNA in the progression, diagnosis, and neoadjuvant chemotherapy of TNBC. Aberrantly expressed miRNA and lncRNA are listed according to their roles. Further, it describes the multiple mechanisms that lncRNA shows for regulating gene expression in the nucleus and cytoplasm, and more importantly, describes lncRNA-regulated TNBC progression through complete combining with miRNA at the post-transcriptional level. Focusing on miRNA and lncRNA associated with TNBC can provide new insights for early diagnosis and treatment-they can be targeted in the future as a novel anticancer target of TNBC.